42

STUDIES ON PRODUCTION OF PECTIN FROM FRUIT WASTES

AVAILABLE IN PAKISTAN. IT’S BIO- CHARACTERIZATION

AND UTILIZATION IN THE DEVELOPMENT OF

PHARMACEUTICAL AND FOOD PRODUCTS

BY

NAUSHEEN HAMEED SIDDIQUI

B-Pharm., M-Pharm.

Thesis submitted for the partial fulfillment of the degree of

DOCTOR OF PHILOSOPHY

DEPARTMENT OF PHARMACOGNOSY

FACULTY OF PHARMACY AND PHARMACEUTICAL SCIENCES

UNIVERSITY OF KARACHI

KARACHI 75250

PAKISTAN

2016

43

DEDICATION

Dedicated to my parents for their continuous support

To my husband for be my strength and

to my priceless loving children

Maryam, Mysha and Ayaan

who are source of inspiration for me.

44

TABLE OF CONTENTS

List of Tables vii

List of Figures xvi

List of Abbreviations xix

Acknowledgements xxii

Abstract xxv

Urdu Translation xix

PART - I

INTRODUCTION 1

REVIEW OF LITERATURE 14

History of pectin 14

Chemistry and type of pectin 15

Biosynthesis of pectin 16

Classification of Pectin 17

Extraction and purification of pectin 18

Traditional method of extraction 22

Microwave- assisted extraction 23

Enzymatic Extraction of pectin 25

Purification of Pectin 27

Uses and application of pectin 34

Large scale production of pectin 39

45

PART - II

MATERIALS AND METHODS 42

Equipment 42

List of chemicals and reagents used 45

List of apparatus 48

List of solutions used 49

Extraction of pectin 53

Processing and preparation of peel 53

Screening of seasonal fruits and their wastes for the presence of pectin

contents

53

Extraction of pectin from selected fruit wastes and their comparative

study

54

Mechanical, pH and heating factors used during the study 55

Effects of organic acid and inorganic acids and their strength on pectin

yield from sapodilla

58

Extraction with 0.1N HCl, using 5 mechanical procedures, 5 pH, two

boiling methods and varying time of boiling (10, 20, 40 and 60 min)

58

Extraction using 1N HCl, using 5 mechanical procedure, 5 pH, two

boiling methods and varying time of boiling (10, 20, 40 and 60 min)

58

Extraction using organic acids 59

Extraction with different strengths of inorganic acid 59

Physical and biochemical characterization of pectin from Sapodilla 59

46

Percent yield 59

Identification tests for pectin 59

Stiff gel test 60

Test with 95% ethanol 60

Test with potassium hydroxide (KOH) 60

Iodine test 60

Biochemical characterization 61

Preparation of sample 61

Qualitative test for ammonia 61

Detection of moisture, ash, methoxy content and equivalent weight 61

Moisture 61

Ash 61

Equivalent weight 62

Methoxyl content 62

Anhydrouronic Acid 62

Grading of pectin 63

Galacturonic acid content 64

Water holding capacity (WHC) 64

Water binding capacity (WBC) 65

Fat binding capacity (FBC) 66

Fourier Transform Infrared Spectroscopy of selected pectin 66

47

Dynamic Light Scattering studies of selected pectin 66

Optimization of the yield of pectin from sapodilla peel thorough

Response Surface Methodology (RSM)

67

Pharmaceutical preparation 68

Formulation of tablet using extracted pectin from sapodilla peel 68

Evaluation of granules 71

Angle of repose (α) 71

Bulk density (ρb) 71

Tapped density (ρt) 72

Compressibility Index 72

Hausner’s ratio 72

Loss on drying (LOD) 72

Tablet compression 73

Tablets testing 73

Weight variation 74

Tablet thickness and diameter 74

Tablet hardness 74

In-Vitro dissolution studies 74

Formulation of antidiarrheal preparation (suspension) from extracted

sapodilla pectin.

75

Evaluation of Suspension 76

48

Color, odor and taste 76

Viscosity 76

Sedimentation Volume 76

Redispersibility 77

Food preparation 79

Preparation of Sapodilla jam 79

Physico-chemical analysis of jam 80

Determination of pH 80

Determination of viscosity 80

Determination of moisture and ash contents 80

Determination of total titratable acidity (TTA) 80

Determination of total soluble sugar 80

Determination of total solids 80

Determination of vitamin C (Ascorbic Acid) 81

Sensory Evaluation 81

Preparation of pudding from extracted sapodilla pectin 83

PART - III

RESULTS 86

Extraction and purification of pectin from the selected fruit waste. 86

Physical and Biochemical characterization of the purified pectin 88

Optimization of the yield of pectin from sapodilla peel thorough 88

49

response surface methodology

Utilization of the purified pectin for the development of pharmaceutical

and food products

89

PART – IV

DISCUSSION AND CONCLUSION 169

DISSCUSSION 169

CONCLUSION 206

REFERENCES 212

ANNEXURES

PUBLICATIONS

50

LIST OF TABLES

Table 1 Showing the uses of different types of pectin in various types of

food products

19

Table 2 Showing some investigated sources of pectin with their applied

mode of extractions

20

Table 3 Showing different uses and properties of pectin. (Source: J.F.

Hydrocolloids)

37

Table 4 Composition of Paracetamol tablet with different concentrations of

pectin used

69

Table 5 Composition of Ibuprufen tablet with different concentrations of

pectin used

70

Table 6 Composition of antidiarrheal preparation using extracted sapodilla

pectin.

78

Table 7 Formulation of the sapodilla Jam (1KG) with different concentration

of pectin

82

Table 8 Showing different pectin concentration used for each selected

extracted fruit pectin

85

Table 9 List of fruits used in the screening study for the presence of pectin 91

Table 10 Showing percent yield of pectin from five selected fruits 92

Table 11 Analysis of variance (mean squares) of yield for five selected fruits 93

Table 12 Means comparison of yield for sapodilla fruit peel (Mechanical

procedure pH interaction mean±SE )

94

Table 13 Means comparison of yield for sapodilla fruit peel (Mechanical

procedure boiling method interaction mean±SE)

94

Table 14 Means comparison of yield for sapodilla fruit peel (Mechanical

procedure pH boiling method interaction mean±SE)

95

51

Table 15 Means comparison of yield for banana fruit peel (Mechanical

procedure pH interaction mean±SE)

96

Table 16 Means comparison of yield for banana fruit peel (Mechanical

procedure boiling method interaction mean±SE)

96

Table 17 Means comparison of yield for banana fruit peel (Mechanical

procedure pH boiling method interaction mean±SE)

97

Table 18 Means comparison of yield for muskmelon fruit peel (Mechanical

procedure pH interaction mean±SE)

98

Table 19 Means comparison of yield for muskmelon fruit peel (Mechanical

procedure boiling method interaction mean±SE)

98

Table 20 Means comparison of yield for muskmelon fruit peel (Mechanical

procedure pH boiling method interaction mean±SE)

99

Table 21 Means comparison of yield for apple fruit peel (Mechanical

procedure pH interaction mean±SE)

100

Table 22 Means comparison of yield for apple fruit peel (Mechanical

procedure boiling method interaction mean±SE)

100

Table 23 Means comparison of yield for apple fruit peel (Mechanical

procedure pH boiling method interaction mean±SE)

101

Table 24 Means comparison of yield for orange fruit peel (Mechanical

procedure pH interaction mean±SE)

102

Table 25 Means comparison of yield for orange fruit peel (Mechanical

procedure boiling method interaction mean±SE)

102

Table 26 Means comparison of yield for orange fruit peel (Mechanical

procedure pH boiling method interaction mean±SE)

103

Table 27 Percentage yield of pectin from sapodilla fruit peel after using

different physicomechanical procedures (pH, mechanical procedure,

104

52

boiling method and time of boling) with 0.1N HCl

Table 28 Analysis of variance (mean squares) of yield of pectin from

sapodilla fruit peel after using different physicomechanical

procedure different fruits with 0.1N HCl

105

Table 29 Means comparison of yield for sapodilla fruit peel after 10 min of

boiling using 0.1N HCl (Mechanical procedure pH interaction

mean±SE)

106

Table 30 Means comparison of yield for sapodilla fruit peel after 10 min of

boiling using 0.1N HCl (Mechanical procedure boiling method

interaction mean±SE)

106

Table 31 Means comparison of yield for sapodilla fruit peel after 10 min of

boiling using 0.1N HCl (Mechanical procedure pH boiling

method interaction mean±SE)

107

Table 32 Means comparison of yield for sapodilla fruit peel after 20 min of

boiling using 0.1N HCl (Mechanical procedure pH interaction

mean±SE)

108

Table 33 Means comparison of yield for sapodilla fruit peel after 20 min of

boiling using 0.1N HCl (Mechanical procedure boiling method

interaction mean±SE)

108

Table 34 Means comparison of yield for sapodilla fruit peel after 20 min of

boiling using 0.1N HCl ( Mechanical procedure pH boiling

method interaction mean±SE)

109

Table 35 Means comparison of yield for sapodilla fruit peel after 40 min of

boiling using 0.1N HCl (Mechanical procedure pH interaction

mean±SE)

110

Table 36 Means comparison of yield for sapodilla fruit peel after 40 min of

boiling using 0.1N HCl ( Mechanical procedure boiling method

interaction mean±SE)

110

53

Table 37 Means comparison of yield for sapodilla fruit peel after 40 min of

boiling using 0.1N HCl (Mechanical procedure pH boiling

method interaction mean±SE)

111

Table 38 Means comparison of yield for sapodilla fruit peel after 60 min of

boiling using 0.1N HCl Mechanical procedure pH interaction

mean±SE)

112

Table 39 Means comparison of yield for sapodilla fruit peel after 60 min of

boiling using 0.1N HCl (Mechanical procedure boiling method

interaction mean±SE)

112

Table 40 Means comparison of yield for sapodilla fruit peel after 60 min of

boiling using 0.1N HCl (Mechanical procedure pH boiling

method interaction mean±SE)

113

Table 41 Percentage yield of pectin from sapodilla fruit peel after using

different physicomechanical procedures (pH, mechanical procedure,

boiling method and time of boling) with IN HCl

114

Table 42 Analysis of variance (mean squares) of yield of pectin from

sapodilla fruit peel after using different physicomechanical

procedure different fruits with 1N HCl

115

Table 43 Means comparison of yield for sapodilla fruit peel after 10 min of

boiling using 1N HCl ( Mechanical procedure pH interaction

mean±SE)

116

Table 44 Means comparison of yield for sapodilla fruit peel after 10 min of

boiling using 1N HCl ( Mechanical procedure boiling method

interaction mean±SE)

116

Table 45 Means comparison of yield for sapodilla fruit peel after 10 min of

boiling using 1N HCl (Mechanical procedure pH boiling method

interaction mean±SE)

117

54

Table 46 Means comparison of yield for sapodilla fruit peel after 20 min of

boiling using 1N HCl (Mechanical procedure pH interaction

mean±SE)

118

Table 47 Means comparison of yield for sapodilla fruit peel after 20 min of

boiling using 1N HCl ( Mechanical procedure boiling method

interaction mean±SE)

119

Table 48 Means comparison of yield for sapodilla fruit peel after 20 min of

boiling using 1N HCl (Mechanical procedure pH boiling method

interaction mean±SE)

120

Table 49 Means comparison of yield for sapodilla fruit peel after 40 min of

boiling using 1N HCl (Mechanical procedure pH interaction

mean±SE)

121

Table 50 Means comparison of yield for sapodilla fruit peel after 40 min of

boiling using 1N HCl (Mechanical procedure boiling method

interaction mean±SE)

121

Table 51 Means comparison of yield for sapodilla fruit peel after 40 min of

boiling using 1N HCl ( Mechanical procedure pH boiling

method interaction mean±SE)

122

Table 52 Means comparison of yield for sapodilla fruit peel after 60 min of

boiling using 1N HCl (Mechanical procedure pH interaction

mean±SE)

123

Table 53 Means comparison of yield for sapodilla fruit peel after 60 min of

boiling using 1N HCl (Mechanical procedure boiling method

interaction mean±SE)

123

Table 54 Means comparison of yield for sapodilla fruit peel after 60 min of

boiling using 1N HCl ( Mechanical procedure pH boiling

method interaction mean±SE)

124

55

Table 55 Means comparison of yield for sapodilla fruit peel after using

different inorganic acid

125

Table 56 Analysis of variance (mean squares) of yield of pectin from

sapodilla fruit peel after using different inorganic acid

126

Table 57 Means comparison of yield for sapodilla fruit peel after using citric

acid (Boiling method x mechanical procedure interaction mean±SE)

126

Table 58 Means comparison of yield for sapodilla fruit peel after using citric

acid (Acid % x boiling method interaction mean±SE)

127

Table 59 Means comparison of yield for sapodilla fruit peel after using citric

acid (Boiling method x Acid% x mechanical procedure interaction

mean±SE)

127

Table 60 Means comparison of yield for sapodilla fruit peel after using oxalic

acid (Boiling method x mechanical procedure interaction mean±SE)

128

Table 61 Means comparison of yield for sapodilla fruit peel after using oxalic

acid (Acid% x boiling method interaction mean±SE

128

Table 62 Means comparison of yield for sapodilla fruit peel after using oxalic

acid (Boiling method x Acid% x mechanical procedure interaction

mean±SE)

129

Table 63 Means comparison of yield for sapodilla fruit peel after using

tartaric acid (Boiling method x mechanical procedure interaction

mean±SE)

130

Table 64 Means comparison of yield for sapodilla fruit peel after using

tartaric acid (Acid% x boiling method interaction mean±SE)

130

Table 65 Means comparison of yield for sapodilla fruit peel after using

tartaric acid( Boiling method x Acid% x mechanical procedure

interaction mean±SE)

131

Table 66 Percentage yield of pectin from sapodilla fruit peel using different 132

56

strengths of same inorganic acid.

Table 67 Analysis of variance (mean squares) of yield of pectin from

sapodilla fruit peel after using different strength of same inorganic

acid

133

Table 68 Means comparison for yield of pectin from sapodilla fruit peel after

using 0.1N HCl (Mechanical procedure pH interaction mean±SE )

134

Table 69 Means comparison for yield of pectin from sapodilla fruit peel after

using 0.1N HCl (Mechanical procedure boiling method interaction

mean±SE)

134

Table 70 Means comparison for yield of pectin from sapodilla fruit peel after

using 0.1N HCl (Mechanical procedure pH boiling method

interaction mean±SE)

135

Table 71 Means comparison for yield of pectin from sapodilla fruit peel after

using 0.5N HCl (Mechanical procedure pH interaction mean±SE)

136

Table 72 Means comparison for yield of pectin from sapodilla fruit peel after

using 0.5N HCl (Mechanical procedure boiling method interaction

mean±SE)

136

Table 73 Means comparison for yield of pectin from sapodilla fruit peel after

using 0.5N HCl (Mechanical procedure pH boiling method

interaction mean±SE)

137

Table 74 Means comparison for yield of pectin from sapodilla fruit peel after

using 1N HCl (Mechanical procedure pH interaction mean±SE)

138

Table 75 Means comparison for yield of pectin from sapodilla fruit peel after

using 1N HCl (Mechanical procedure boiling method interaction

mean±SE)

138

Table 76 Means comparison for yield of pectin from sapodilla fruit peel after

using 1N HCl (Mechanical procedure pH boiling method

139

57

interaction mean±SE)

Table 77 Identification tests for presence of pectin from three selected fruits 140

Table 78 Biochemical characterization of the purified pectin extracted from

sapodilla pectin at different pH

140

Table 79 Water holding, water binding and fat binding capacity of sapodilla

pectin

141

Table 80 Showing FTIR spectral values of sample and standard pectin along

with associated functional groups.

141

Table 81 Summary of the physical characterization of standard pectin and the

different pectin extracted from various source and at different pH by

DLS studies.

144

Table 82 Box-Bechen experimental design and levels of factors used for

optimization of pectin yield

150

Table 83 Box-Behnken experimental design and corresponding results for

responses

150

Table 84 Analysis of Variance for Yield (Box-Bechen experimental design,

response surface methodology)

151

Table 85 Estimated Regression Coefficients for Yield (Box-Bechen

experimental design, response surface methodology)

152

Table 86 Predicted values of yield. (Box-Bechen experimental design,

response surface methodology)

152

Table 87 Flow properties of granules made for paracetamol tablets 157

Table 88 Pharmaceutical characteristics of compressed formulation of

paracetamol tablet

158

Table 89 Dissolution studies of paracetamol tablet 159

Table 90 Flow properties of granules made for ibuprofen tablets 163

58

Table 91 Pharmaceutical characteristics of compressed formulation of

ibuprofen tablet

163

Table 92 Dissolution studies of ibuprufen tablets 164

Table 93 Basic evaluation test of antidiarrheal formulation prepared from

sapodilla pectin

166

Table 94 Effect of different concentration of sapodilla pectin on the chemical

properties of the jam samples

167

Table 95 Scores for sensory parameters of jam as judged by twenty (20)

panelists.

167

59

LIST OF FIGURES

Fig. 1 Structure of plant cell showing location of pectin in the cell wall 1

Fig. 2 Showing colour and texture of pectin 2

Fig. 3 (A) Showing formation of viscous gel and (B) showing increased bulk

volume of stool by Pectin

3

Fig. 4 Showing effect of retention of NFM by pectin on outer and inner

stratum corneum of the skin. The improved hydration of the

epidermis leads of enhanced mechanical stability of the stratum

granulosum, stratum spinosum and stratum basle

4

Fig. 5 Showing glycoside linkage 5

Fig. 6 α-4-linked galactosyluronic acid (GaIA) 6

Fig. 7 Schematic representations of the conventional (A) and alternative

(B) structures of pectin.

7

Fig. 8 Showing major sources of pectin used arround the world 10

Fig. 9 Showing The overall application and uses of pectin in

pharmaceutical, food, cosmetics and chemical industries

10

Fig. 10 Showing major pectin producers and their market share 11

Fig. 11 Showing the global pectin market 12

Fig. 12 Showing current and growing market size of hydrocolloids in the

different regions of the world

13

Fig. 13 Schematic diagram for biosynthesis of pectin 17

Fig. 14 Extraction of pectin from soy hull. 28

Fig. 15 Extraction of pectin from leaves of Nephrolepis biserrata 29

Fig. 16 Extraction of pectin from raw papaya peel 30

Fig. 17 Commercial extraction of pectin ( Ensymm) 31

60

Fig. 18 Commercial extraction of pectin (Cargillfood) 32

Fig. 19 Commercial extraction of pectin (GENU) 33

Fig. 20 Showing marketed of pectin in Middle east and Australia. 38

Fig. 21 Flow chart showing general procedure employed for the extraction

of pectin on commercial scale.

41

Fig. 22 Showing flowchart of extraction of pectin followed during the

current study

57

Fig. 23 FTIR spectra of food grade pectin 142

Fig. 24 FTIR spectra of sapodilla peel pectin extracted at pH 5 142

Fig. 25 FTIR spectra of sapodilla peel pectin extracted at pH 3 143

Fig. 26 FTIR spectra of sapodilla peel pectin extracted at pH 1 143

Fig. 27 Physical characterization of standard pectin by DLS studies 145

Fig. 28 Physical characterization of apple pectin extracted at pH5.0 by

DLS studies

146

Fig. 29 Physical characterization orange pectin extracted at pH5.0 by DLS

studies

147

Fig. 30 Physical characterization of sapodilla pectin extracted at pH3.0 by

DLS studies.

148

Fig. 31 Physical characterization of sapodilla pectin extracted at pH5.0 by

DLS studies

149

Fig. 32 Showing the optimal conditions for the extraction of pectin from

sapodilla fruit peel(Box-Bechen experimental design, response

surface methodology)

153

Fig. 33 Response surface graph and contour plot of effect of pH and

temperature on yield of pectin at constant time

154

Fig. 34 Response surface graph and contour plot of effect of pH and time 155

Boiling for 5 min

61

on yield of pectin at constant temperature

Fig. 35 Response surface graph and contour plot of effect of temperature

and time on yield of pectin at constant pH

156

Fig. 36 Formulated paracetamol tablest ( F1, F2, F3) 160

Fig. 37 Formulated paracetamol tablest ( F4, F5, F6) 161

Fig. 38 Formulated paracetamol tablest ( F7, F8, F9) 162

Fig. 39 Formulated ipubrufen tablets ( R1, R2, R3, R4) 165

Fig. 40 Formulated antidiarrheal preparation 166

Fig. 41 Formulation of Jam made from extracted pectin 168

Fig. 42 Formulation of pudding made from extracted pectin. 168

62

Abbreviation

Abbreviation Full Form

AGA Apigalacturonan

AOAC Association of Official Analytical Chemists

ADI Accepted Daily Intake

AUA Anhydrouronic acid

ANOVA Analysis of Variance

ACF Autocorrealtion function

API Active Pharmaceutical ingredients

Ca++ Calcium ion

CAS Chemical abstract service

CDTA Cyclohexanediaminetetraacetic acid

CFR Code of Federal Regulations

CeO Cerium oxide

CP Centipoise

Coef Coefficient

DLS Dynamic light scattering

DM Degree of methoxylation

DE Degree of esterification

DI Deionized water

EDTA Ethylenediaminetetraacetic Acid

FTIR Fourier transform infrared spectroscopy

63

FDA Food and Drug Administration

FAO Food and Agriculture Organization of the United Nations

FGP

g

Food grade pectin

Gram

GA Galacturonic acid

GI Gastrointestinal

GC-MS Gas chromatography–mass spectrometry

Gal A Galacturonic acid

GRAS Generally Recognized as Safe

HG Homogalacturonan

HGA Homogalacturonan

H2SO4 Sulphuric acid

HPLC High Performance Liquid Chromatography

HCL Hydrochloric acid

HM High methoxyl pectin

H2O Water

HIV Human immunodeficiency virus

IMS Industrial methylated spirit

JECFA The Joint FAO/WHO Expert Committee on Food Additives

KOH Potassium hydrochloride

Kg Kilogram

KP Kilopond

64

LM Low methoxyl pectin

MT Metric ton

MW Microwave

MAE Microwave assisted extraction

ml Millilitre

mg Milligram

mW Milliwatt

MOH Ministry of health

mM Millimolar

N Normal

NFM Natural moisturizing factor

NaOH Sodium hydroxide

nm Nanometer

NH4OH Ammonium hydroxide

NaN3 Sodium azide

NaCl Sodium chloride

%PD Perrcent polydispersity

PCSIR Pakistan Council of Scientific and Industrial Research

ppm Parts pur million

RG-I Rhamnogalacturonan I

RG-II Rhamnogalacturonan II

RSM Response surface methodology

65

RH Radius of hydration

Rpm

s

Revolutions per minute

second

SE Standard error

SD Standard deviation

TSS Total soluble solids

TTA Total titratable acidity

TS Total solids

USP 36/NF United States Pharmacopeia (USP) and the National Formulary

VIFS Variance influence factor

Vit C Vitamin C

W Watt

w/v Weight by volume

WBC water binding capacity

WHC Water holding capacity

WHO World health organization

XGA Xylogalacturonan

66

ACKNOWLEDGEMENT

All praises goes to Allah Almighty because without his countless blessing and love this

work would have been possible.

I would like to express my deep and sincere gratitude to my supervisor, Professor Dr

Iqbal Azhar, Dean Faculty of Pharmacy and Pharmaceutical Sciences, University of

Karachi for his kind support, advices, productive and useful opinions and suggestions

throughout my work. His constant encouragement and attitude has always been a moral

support for me. His continuous help and confidence in my abilities have been an integral

part for my understanding I will always be grateful.

I owe a lot of gratitude to my second supervisor Dr Zafar Alam Mahmood, Country

Manager, Colorcon Limited-England. It is my pride and privilege to express my sincere

thanks and deep sense of gratitude for his motivation, earnestness and immense

knowledge. His preeminent guidance supported me during the time of research and

writing of this thesis. His tutoring and keen interest made my research both enjoyable and

exciting. It is my great honor to study and do research under his talented supervision.

In addition, I would in particular like to thank Dr. M. Mohtasheemul Hasan (Chairman),

Prof Dr. Waseemuddin Ahmed, Professor Dr Mansoor Ahmed, Prof. Dr. Ghazala H.

Rizwan, (former Dean of Faculty of Pharmacy), Ms Farah Mazhar (Assistant prof), Mr.

Salman Ahmed and Mrs. Safia Abdi (Lecturer) Department of Pharmacognosy

University of Karachi and Prof. Dr. M. Shaiq Ali (Assistant professor International

Centre for Chemical & Biological Sciences, ICCBS, HEJ Research Institute of

Chemistry, University of Karachi) for their special supports. I also give thanks to my

67

colleagues and workers at Department of Pharmacognosy Faculty of Pharmacy for their

kind support. I am very grateful to Dr. Abid Ali (Assistant professor International Centre

for Chemical & Biological Sciences, ICCBS, HEJ Research Institute of Chemistry,

University of Karachi) for providing me support in the DLS studies and the important

extensive discussions and contributions concerning my work

I owe loving thanks to my husband, Tariq for his amazing love and support especially

through the hard times. His soothing words always made me feel better and urged me on.

I thank him for his sacrifices and supports throughout the course of my PhD.

I feel particularly indebted to my parents for their ever-lasting love, understanding and

encouragement. It was their unshakable faith in me that has always helped me to proceed

further. I cannot express my love and gratitude I have for them. Thank you for

encouraging me and supporting me in every step so that I can fulfill my dreams.

I wish to thank my brothers and sisters and their spouses who were always there for me in

my difficulties, without their encouragement, understanding, support and prayers it would

have been impossible for me to finish this work. Thanks to my in-laws specially my

Father in law and friends for their constant encouragement when I needed it. I feel short

of words to express my most heartfelt and cordial thanks to my children, who have

always stood by my side at the toughest times their unconditional love kept me going.

Finally, I wish to extend warm thanks to my aunt Mrs Shamsunisa Siddiqui, my sister

Ambreen, nieces Romina and Adina, cousins Marya and Nida and to everybody involved

directly or indirectly with my work.

Nausheen Hameed Siddiqui

68

ABSTRACT

With the rising awareness and use of plant based functional fibers there is a growing

market demand of pectin throughout the world. However, this is mainly linked with the

price and quality of the products availability in the market for different purposes. The

market price and current production of pectin has been influenced by many factors

among which the provision of raw material for the extraction of pectin is of main

concern. The purpose of this research investigations was not only to explore new source

of pectin which can aid production of pectin but also to extract it in a simpler way as to

minimize or reduce the cost of production. A systematic literature search was also carried

out to evaluate the present findings comprising the earlier results in term of its application

in developing food and pharmaceutical products.

This thesis is based on work to find out the effective extraction of pectin from various

fruit wastes available in Pakistan, its extraction in a most simple and effective way and

then after extracting its usage in both pharmaceutical and food product. For this purpose

numerous seasonal fruits available in Pakistan were studied in their respective time of

availability as a screening program.The present research was focused on the isolation,

physicochemical characterization and functional properties of pectin extracted from a

new source. The extraction process for effective extraction was developed after using

different solvents (organic and inorganic acids of different strengths), five extracting pH

(1,3 ,5 6 and 7), five mechanical procedure (homogenizing , grinding, hammering, cutting

and chopping) , two different boiling techniques, (Bunsen burner and microwave heating)

. Sapodilla fruits when selected finally was subjected to other affecting parameters for

better yield, like time of boiling (10, 20, 40 and 60 min respectively) with two different

69

strength of acid (0.1 and 1N HCl) strength of inorganic acid (0.1, 0.5 and 1N HCl

respectively), and effect of organic acid (critic, tartaric and acetic acid)

Initially five fruits, sapodilla, banana, muskmelon, apple and orange were selected for

studying the physic mechanical effects among which two fruits (Apple and orange) were

purely used for comparative study purpose. After this study three fruits (banana sapodilla

and muskmelon) were selected for further evaluation. . Out of three fruits, highest pectin

yield was recorded from banana (10.5%) followed by sapodilla (4.7%) and muskmelon

(4.4%) respectively. In comparison orange peel indicated 22.7% and apple peel 4.85%.

Identification and jelly grade tests were performed which supported sapodilla as most

appropriate against banana and muskmelon pectin for further investigation to find out a

new source of pectin which can be chemically stable, easily available, and

pharmaceutically useful in nature. Hence sapodilla peel was used to extract pectin after

using different phyico-mechanical processes. It was observed that the best extraction

parameter to obtain maximum yield of pectin was after 10 min of boiling of chopped

peels of sapodilla maintaining the pH with 1NHCl to be at pH 5.

Present research was also concerned with the bio characterization of extracted sapodilla

pectin. With the equivalent weight of 1700, degree of esterification 73.63% and jelly

grade 100, sapodilla pectin was evaluated as high methoxyl pectin. Water binding, water

holding and fat binding capacity was also assessed which also showed promising results.

FTIR spectroscopy performed found that the pectin at pH5 was of best quality as

compared to the other extracted pectin at different pH. Dynamic light scattering studies

(DLS) were also performed to determine the partial side and molecular weight of

extracted pectin and was compared with the commercial food grade pectin. The DLS

70

studies showed similarity in the particle size and molecular weight between standard and

extracted pectin which also proved that the extraction performed was effective.

Optimization of process of extraction of pectin was a studied after applying statistical

software Box Behnken design. According to response surface methodology the best yield

of pectin (3.7%) from sapodilla fruits can be achieved by keeping the pH 5 at 61.11 oC

for 90 min of heating time. The verification of predicted model also gave similar results,

3.5% pectin was extracted on the predicted pH, temperature and time of boiling.

The extracted pectin was also used in the formulation of two types of solid

pharmaceutical dosage form and an oral liquid preparation (suspension). The tablets were

first tested for its micromeritics properties of the granules. After the formulation of the

desired tablets the tablets were compressed and were tested for hardness, thickness, loss

on drying and dissolution properties. It was found that the concentration of added pectin

has influence on both the micrometric properties of granules as well as on the dissolution

profile of formulated tablets. Both the tablets showed increased in hardness and lowering

of dissolution rate with the addition of increased amount of pectin. However the best

formulation for paracetamol was F4 and F5 with 40 and 50 mg of pectin respectively

while for ibuprofen R1 with 50mg of pectin concentration was best. The antidiarrheal

preparation also exibited similar results in terms of its evaluation as suspension.

The extracted sapodilla pectin was then used in preparation of food and pharmaceutical

products. Two types of food preparation, jam and pudding were made using the extracted

sapodilla pectin. The jam was also evaluated for its chemical and sensory attributes and

found sapodilla pectin can be used in making of jam with a slightly higher concentration

(10mg) than the food grade pectin while pudding was evaluated for its textural properties

71

mainly and found that the addition of sapodilla pectin (10 and 15g) has no significant

impact on the textural properties of pudding and the pudding was equally acceptable as

pudding made from equal amount of food grade pectin

72

73

74

INTRODUCTION

Considerable focus, both on global and regional level has been given during last decade

towards the systematic research and application of the various findings or outcomes to

support the pharmaceutical, food and cosmetic industries in developing useful

formulations or products. This idea has created a great impact and geared up the research

activity in the field of both primary and secondary metabolites of different biochemical

groups, especially the high molecular weight compounds produced naturally and with

biodegradable property. Perhaps, it will not be exaggerated to interpret that the scientific

knowledge of phytochemicals or phytopharmaceuticals with the current and future

resources will have significant impact in the process development for the introduction of

varieties of industrial or commercial products from renewable plants, fruits and

vegetables. Among various biologically active groups, one of the leading groups of

natural product, which has been given especial attention, is the polysaccharide moiety.

Figure – 1: Structure of plant cell showing location of pectin in the cell wall.

(Source: Molecular expressions, 2016)

75

The polysaccharides are of tremendous importance with regard to their application in

pharmaceutical, nutraceuticals, cosmeceuticals and chemical industry and of best

example which can be quoted here is “Pectin” which is the most structurally complicated.

Polysaccharide in the plant cell wall (Figure – 1) with vast applications in these

commercial organizations with respect to health, food and personal care productsIn plant

cells, pectin is linked to cellulose and hemicellulose. The initial form is protopectin

which is a water insoluble pectic substance and are present as an important parts of

middle lamella among the cells as calcium pectate and magnesium pectate (Figure -1).

The cellulose to which pectin in linked in the cell, provides sufficient support to the

tissues in maintaining their rigidity while the pectic component makes the plant flexible

in nature. Pectin is present as protopectin in unripe fruits which is transformed when the

fruit ripens into pectin, which when mixed with water has a tendency of forming gel and

also among sources of water soluble fibres. However, the jellying capacity of pectin

diminishes when in over ripe fruits pectin is converted into pectic acid. Comes from the

Greek word meaning congealed or curdled, pectin was first isolated and reported in 1790

by Vauquelin, followed by its characterization 1825 Barconnot from the cell wall of fruits

and vegetables. At present, commercial pectin is mainly obtained from citrus fruits and is

Figure – 2: Showing colour and texture of pectin. (Source: Wikipedia, 2016)

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a white to light brown powder (Figure - 2). However, production methods (isolation,

extraction and purification) used by different manufactures are highly confidential and

has patent protection.

The role of pectin is quite diversified in pharmaceuticals. These include both therapeutic

as well as excipient. In view of its properties to increase the viscosity and volume of stool

it has its applications both in diarrhoea and constipation. Though, FDA has discontinued

pectin in 1992, it is still use in other countries. It is also extensively used in Alternative

System of Medicine (such as Ayurvedic and Unani system of medicine) alone or in

combination with certain herbs or minerals as a compound or poly herbal formulations.

Pectin has the property to absorb water and can swell more than 40 times in volume to

form a viscous gel that helps to lubricate the lining of the intestine for ease of defecation.

It also increases bulk volume.

Figure – 3 (A) Showing formation of viscous gel and (B) showing increased bulk

volume of stool by Pectin. (Source: Anonymous)

The viscous gel that is formed softens and adds bulk to the stool to accelerate and

regulate the movement of food through the intestine leading to overall bowel cleansing

and detoxification. Although a bactericidal action of pectin has been proposed to explain

A B

77

the effectiveness of pectin treating diarrhoea, most experimental results do not support

this theory. However, some evidence suggests that under certain in-vitro conditions,

pectin may have a light antimicrobial action toward Echerichia coli (Thakur et al., 1997).

Pectin promisingly affects cholesterol levels in blood and is reported to reduce blood

cholesterol in a wide range of subjects and experimental conditions (Ginter et al., 1979;

Miettinen and Tarpila, 1977; Sriamornsak, 2001; Sundar et al., 2012). As excipients, the

functionalities include, binding and gelling agent. In the medical device, application of

pectin has mostly been focused in wound healing or dressing and as an adhesive, such as

colostomy devices. Ostomy care products.

Figure – 4: Showing effect of retention of NFM by pectin on outer and inner stratum

corneum of the skin. The improved hydration of the epidermis leads of enhanced

Outer stratum

corneum

Inner stratum

corneum

Stratum

granulosum

Stratum

spinosum

Stratum basale

78

mechanical stability of the stratum granulosum, stratum spinosum and stratum basale.

(Source: Anonymous)

When used in personal care products, pectin forms a layer of moisture on the surface of

the skin (Figure - 4) which helps optimise the fluid balance of the epidermis. This not

only supports the skin to retain and maintain its own natural moisturising factor (NMF)

but skin becomes smoother, firmer and full of moisture.

The family of polysaccharides containing α-(1-4)-linked D-galactopyranosyl uronic acid

residue is commonly known as pectin (Yeh et al., 2011). The primary cell wall is mainly

comprised of polysaccharide apart of cellulose and hemicellulose pectin, is one of the

class of polysaccharide which constitutes around 34% portion of call wall. (Darvill et al.,

1980). Pectin is a complex polysaccharide, made up of D--galacturonic acid in which

few are naturally present free carboxyl group are methylated ester while rest can join

with calcium and magnesium ions (Jain et al., 2005). The basic structure of pectin

polysaccharide is D-galacturonic acid, joined by glycoside linkages ( Figure-

5) (Abbaszadeh., 2008).

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Figure - 5: Showing : Showing glycoside linkageglycoside linkage. (

Source : Study blue, 2016)

Structurally the pectic polysaccharides are partitioned into homogalacturonan (HG),

Xylogalacturonan(XGA), apiogalacturonan(AGA), rhamnogalacturonan II(RG-II), and

rhamnogalacturonan I(RG-I) (Caffall & Mohnen, 2009). While during extraction of

pectin two types of pectins are isolated. The difference in types depends upon the degree

of methoxylation (DM). The pectin obtained with a DM more than 50% require high

amount of sucrose and low pH values to form gels while pectins with lower DM values

than 50% need a divalent cation for ex Ca++ to form gels (Carr et al., 1995). One another

group which is also low-methoxYl pectin known as amidated pectin are extracted in the

presence of ammonia. In these types of pectin polymers, amide groups are formed with

ester and acids moieties (May, 1990).

A general structural pattern of pectin polysaccharides is described by many scientists

(Willats, 2001; Caffall and Mohnen, 2009; Ridley et al., 2001). Mainly the structure of

pectin consist of two regions the linear and ramified region. The linear region of

homogalacturonan composed of 1,4-linked D- galactopyranosyluronic acid residues.

These regions are linked by L-rhamnopyranose residues one or two in numbers,

which are involved in the linear chain by a 1,2-linkage. Most of the pectins has this very

structure as their backbone, the difference among them is the length of the chain. The

ramified region composed of three auxiliary units: RG-I, arabinogalactan, and

xylogalacturonan, which can be found in different ratios (Ovodov, 2009).

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Due to the effect of the isolation procedure, storage and processing of plant material. It is

difficult to describe the exact structure of pectin (Sriamornsak, 2002). However it is

presumed that RG-I and RG-II are covalently linked to HGA domains but the direct

linkage between RG-I and RG-II does not have a proper proof. HGA is a

linear homopolymer of (1→4)-αlinked- D-galacturonic acid and is thought to include

some100–200 Galacturonic acid (GalA) residues. HGA is the most common and

frequently found part of pectin which is synthesized in Golgi. RG-I composed of about

100 repeats of the disaccharide (1→2)-α-L-rhamnose-(1→4)-α-D-galacturonic acid is the

acidic pectin domain/part. Here the chains are mainly consisting of of arabinan

andgalactan. RG-II sounds similar to RG-I due to its name but it has no structural

relationship with RG-I. It is a part of pectin which has numerous branches and a major

part of HGA. RG-II consists of almost 9 GalA residues which are connected by a (1→4)-

α-linkage which is substituted by 4 heteropolymeric side chains of known and consistent

lengths. There are eleven types of sugars attached with the side chains such as, 2-keto-3-

deoxy-D manno-octulosonic acid, apiose and acetic acid. (Figure-6) (Abbaszadeh, 2008)

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Figure – 6: α-4-linked galactosyluronic acid (GaIA) (Ridley et al., 2001)

In foods, including confectionery, beverage and canning industries, the application is

mainly confined as thickening and gelling agent while in cosmetic formulations it is

incorporated for its gelling, thickening and stabilizing properties (May, 1990;

Sriamornsak, 2001 and 2002 and Srivastava and Malviya, 2011). The food additive code

number for pectin is 440 or E440. The applications of pectin in solid oral dosage form

Figure-7: Schematic representations of the conventional (A) and alternative (B)

structures of pectin. Willats et al. (2006).

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have been comprehensively represented in a review article (Sriamornsak, 2001). As a

therapeutic agent, it has two major applications, which include its application in

antidiarrheal formulations for infants and children as well as a dislipidemia, especially to

reduce elevated blood cholesterol (Sriamornsak, 2001). However, it requires a very high

intake of pectin, above 6 g/day which may reduce around 13% of serum cholesterol in

two week time (Ginter et al., 1979; Mietinnen and Tarplia, 1977). In addition, pectin has

been reported to be useful in elimination of lead and mercury from respiratory organs and

gastrointestinal tract (Kohn, 1982). Another application to treat disorders relating to

overeating has also been reported (Di Lorenzo et al., 1988). As an excipient, pectin act as

binder and compressibility enhancer (Slany et al., 1981a, 1981b; Kim et al., 1998) as well

as in the development of modified release dosage form, such as controlled release matrix

formulations (Krusteve et al., 1990; Naggar et al., 1992; Sungthongjeen et al., 1999),

pectin beads as well as insoluble hydrophilic coating for sustained release drug delivery

system (Aydin and Akbuga, 1996; Sriamornsak, 1996; Sriamornsak et al., 1997a and

1997b), and above all its application as a carrier material in colon-specific drug delivery

system due to its insolubility and resistant property against intestinal enzymes but

sensitivity against colonic pectinolytic enzymes (Ashford et al., 1993; Rubinstein et al.,

1993; Englyst et al., 1987; Sandberg et al., 1983 and Rubinstein et al., 1992)

Pectin provides a natural structure to various cosmetic preparations, such as pastes,

creams and ointments, while its thickening and stabilizing action is useful in applying

hair tonics, body lotions, shampoos and conditioners (Ref: Cargill). When used in skin

formulations, pectin forms a layer of moisture on the surface of the skin. The major

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source as reported by IMR international (2009) for the production of pectin is citrus fruits

(Lemon, lime or oranges), followed by apple and sugar beet (Figure-8). The industrial

applications of pectin include use in plasticizers, edible films, foams and paper substitute.

The overall application and uses of pectin in pharmaceutical, food, cosmetics and

chemical industries have been graphically presented in Figure –9

Figure-8: Showing major sources of pectin used arround the world. ( Source: IMR

International 2016)

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Figure-9: Showing the overall application and uses of pectin in pharmaceutical,

food, cosmetics and chemical industries. (Source: IMR International 2016)

The major pectin manufacturing companies include: Danisco, , CP Kelco, Herbstreith &

Fox (Germany), Cargill and Yantal. And according an estimate (IMR 2009) 42,000 MT

of pectin was produced worldwide with a positive growth between 3 to 6%. CP Kelco is

the largest pectin producer in the world (Figure-10).

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.

Figure-10: Showing major pectin producers and their market share. ( Source: IMR

International 2016)

In view the importance and diversified applications of pectin which categorises it as an

exciting and promising component for the pharmaceuticals, nutraceuticals and

cosmeceuticals, the search of new sources for its production demands a comprehensive

studies by applying both pure and applied knowledge. Based on this approach, present

study was taken to investigate the possibility of having a better quality of pectin from

seasonal fruits available in Pakistan and to explore and synchronize the physic-chemical

parameters for increased yield on lab scale basis. This was followed by the application of

the crude as well as refined pectin in some pharmaceutical oral and liquid dosage form

and to calculate the economics of overall process. According to the most recent survey

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available on internet (Vision, 2015) the size of the global pectin market would be of $1 B

by 2017 and the compound annual growth rate will be 8% during the year 2012-2017

(Figure-11)

Figure -11: Showing the global pectin market. (Source: Vision, 2015)

87

Figure-12: Showing current and growing market size of hydrocolloids in the

different regions of the world (Source: Markets and Markets, 2016)

Hydrocolloid market in general is on constant growth which can be seen in figure-12.

According to this market survey report hydrocolloid market is growing fastest in Asia-

Pacific while due to the changing lifestyle and growing understanding of healthy and

nutritional foods the demand of hydrocolloids, of which pectin is a part, also increasing in

south-east Asia (Markets and Markets).

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REVIEW OF LITERATURE

History of pectin

The pectic substances were first discovered by the French Chemist Louis Vauquelin in

1790, while its properties were first reported by Henri Braconnot another French chemist

in 1825 (Georgiev et al.,2012). Pectin has a key action in the maintenance of the

structural integrity of plant by providing firmness to the plant by composing a hydrated

cross linked three dimensional networks (Shi et al., 2002). Pectin not only has a role in

ripening and texture building of fresh fruits in plants but it is an important agent in

processing industry for its characteristics property of jelling and thickening agent in

canned and processed food products (Nurdjanah,2013).The traditional use of pectin was

due to its gelling property and it was used in making jam jellies and marmalades and

other bakery products (Jameson et al., 1923; Nanji et al 1934).Pectin was also used as

emulsifying agent in many food products containing oils and fats. It was presumed that

pectin, due to its gelling property will not only help in stabilizing a product under regular

climatic conditions but also helps in pasteurizing the product (Douglas and Loesch,

1927). Other food items were also investigated to improve the quality of food products in

which pectin was used. Breakfast foods and cereals were studied to improve the texture

and quality of this food product (Spalding and Conn, 1936).

Numerous application of pectin has been studies over the years which include its

traditional uses to modern usage in pharmaceutical application. Pectin was also studied

for its other medicinal properties like hemostasis (Kertesz, 1951), lipid lowering (Ershoff

and Wells, 1962), activity on enzymes and hormones (Morgan et al., 1979) and

cholesterol lowering agent (Ink and Hurt, 1987). Pectin also got under investigation in the

89

management of both types of diabetes mellitus and showed that it helps in lowering blood

glucose and insulin levels (Jenkins et al., 1976). Due to its ability to absorb water it is

also used in weight reduction (Holt et al., 1979). The addition of pectin in pharmaceutical

preparation due to its properties like delaying absorption of drug (Kertesz, 1951). It has

been used in combination with many other agents in treatment of diarrhea (Tompkins,

1938; Bennett, 1958). Pectin was also used in the prophylaxes and management of heavy

metal toxicity (Paskins- Hurlburt et al., 1977). Studies were also found for the use of

pectin in dentistry and skin care products (Lochridge, 1951).Other interesting uses

reported in literature were the use of pectin in therapeutic gels (Raudnitz, 1979).

Chemistry and Type of Pectin

In search for naturally occurring compounds plants are being studied with great interest.

Likewise pectin has also been studies because of its complex structures and vast number

of uses (Cosgrove and Jarvis, 2012).Although pectin is being considered as most complex

and heterogeneous naturally originated structure and its being studied upon for more than

180 years, it is astonishing to learn that due to different physiological changes occurring

in call wall the actual molecular structure of pectin are still studied so as to understand

the basic structure of pectin molecule (Schols et al., 2009; Cybulska et al., 2015).

Numerous techniques and apparatus are being used in order to understand the actual

chemical structure of pectin such as atomic force microscopy and force microscopy

(Morris et al., 2011).Studies on the structure of pectin are important as the properties and

functions of pectin depend upon its chemical configuration. (Funami et al., 2011).The

studies on structure of pectin further extend the possibility of its uses and application in a

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more scientific way both in food and pharmaceutical properties (Sila et al., 2009).The

structure of pectin is mainly consist of D-galacturonic acid (Gal A) units joined in chains

by means of alpha-(1-4) glycosidic linkage (Srivastavaand Malviya, 2011).The

proportion of occurrence among HG, XGA, RGI, and RGII is also variable; typically HG

is the most common polysaccharide, composing around 65% of the pectin, while RGI

makes 20% to 35%. XGA and RGII are other lessor segments, each constituting less than

10 % (Acton, 2013).

Biosynthesis of pectin

The parts of the plants involved in the biosynthesis of pectin are mainly Golgi vesicles

however it is believed that some preliminary steps occurs in the endoplasmic reticulum. A

large numbers of enzymes are also involved in synthesis of these complex polysaccharides

which are mostly located in the Golgi apparatus. There are almost 67 enzymes which have

been identified which are required during the process which include glycosyltransferases,

methyltransferases and acetyltransferases (Harholt et al., 2010). The initiation of the

formation of pectic polysaccharides is still not known and the involvement of lipids and

protein donors are also not established. The pectin biosynthesis investigation has been a

tough task for researchers with very passive growth. Figure -13 shows a diagrammatic

representation of biosynthesis of pectin. According to this predictive model the pectin

biosynthesis mainly rely on nucleotide-sugars which are formed by nucleotide-sugar

transformation reactions which takes place in cytosyolic side of Golgi. A specialized

nucleotide-sugar which is called nucleoside monophosphate antiporters are responsible for

the movement of nucleotide-sugar into the Golgi (Ridley et al., 2001)

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Figure- 13: Schematic diagram for biosynthesis of Pectin. (Ridley et al., 2001)

Classification of Pectin

Commercially pectin is divided into different types on the basis of its gel forming

property. The gel forming property of pectin depends upon the degree of esterification.

The backbone structure of pectin which is partially methyl-esterified is used for

classification of pectin. The degree of esterification (DE) less than 50% is low DE pectin

while DE more than 50% is high DE pectin (Thirawong et al., 2007). It was also reported

that lower the methoxyl content present in pectin reduces is its gelling property

(Srivastava and Malviya, 2011).Activities like cholesterol lowering properties, also

depends upon the natural chemical properties of pectin (Brouns et al.,2012).

The types of pectin also effect drug permeability when used as films (Hagesaether et al.,

2009). It also has an effect on food grade of pectin, when pectin used in jams it effects the

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texture and color of the jam (Kopjar et al., 2009). Other effects like mucoadhesive,

(Hagesaether and Sande, 2007) and the relationship between pectin and mucin by mucin-

particle method are also under investigation (Sriamornsak et al., 2010).Little influence

has been of the degree of esterification (DE) of the pectin molecules on chemical abstract

service (CAS) emulsion stability at different pH.(Surh et al., 2006).Pectin used in making

different food items and the type of pectin used are presented in Table – 1.

Extraction and purification of pectin

Kertesz in 1951 suggested that the extraction of pectin is a complex physico-chemical

process which may involve solubilization, extraction and DE polymerization of pectin

complex molecules from plant tissues. In lab-scale extraction, hot water extraction is used for

high esterified pectin. While the low-esterified pectin is not extracted efficiently when

extracted with the same procedure for them, chelating agents like Ethylenediaminetetraacetic

acid (EDTA) and Cyclohexylenedinitrilotetraacetic acid (CDTA) are used (QI et al., 2002).

While commercially, pectin is extracted through hot dilute mineral acid pH around 2 with

varying length of time. The time depends upon the type of pectin required and from one

manufacturer to another (May, 1990).Various factors can affect the extraction of pectin. The

extracting conditions may affect the probable structure of pectin. Studies on, the use of

different pH levels, reveals that it has a profound effect on both the extractability and in the

breakdown of pectin molecule (Kaya et al., 2014).). Some of the recent referenced natural

sources of pectin with respective method of extraction have been highlighted in the Table-2.

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Table - 1: Showing the uses of different types of pectin in various types of food

products

Food item Type of pectin Reference

Jams, marmalades, fruit

jellies and fruit spreads

HM Pectin Rababah et al., (2014)

Beverages HM pectin Walsh et al., (2014)

Fruit preparations for

yougurt

LM pectin

LM amidated pectin

Rinaldia et al., (2014)

Fruit preparations for

bakery products

LM pectin Wuestenberg, (2014)

Confectionery HM pectin Wuestenberg, (2014)

Glazes LM pectin

LM amidated pectin

Wuestenberg, (2014)

Deserts LM pectin

LM amidated pectin

Thakur., et al (1997)

Acidified dairy drinks HM pectin Bonnaillie., et al (2014).

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Table - 2: Showing some investigated sources of pectin with their applied mode of

extractions.

SOME SOURCES OF PECTIN

Source (Common Names) Mode of extraction Scientist/scientists

Gold kiwifruit Acid, water, enzymetic Yuliarti et al., (2015)

Water Melon Microwave Assisted Maran et al., (2014)

Grape fruit Ultrasound-assisted heating

extraction Xu et al., (2014)

Passion fruit peel Microwave-induced heating Seixas et al., (2014)

Grape Pomace Ultrasound-assisted

extraction

Minjares-Fuentes et al.,

(2014)

Dragon Fruit Citric Acid Muhammad et al., (2014)

Apple pomace and citrus

peel Subcritical water Wang et al., (2014)

Pumpkin Enzymatic extraction Cui and Chang., ( 2014)

Citrus limon Alcohol precipitation

method Kanmani et al., (2014)

Pomelo Microwave Tan et al., (2014)

Lemon pomace Different solvents Azad et al., (2014)

Mango Chelating agents Kermani et al., (2014)

Sisal waste Aqueous extraction Santos et al,. (2013)

lime peel High hydrostatic pressure

treatment Naghshineh et al., (2013)

Sugar beet Organic acids Ma et al., (2013)

Orange

Ultra-high pressure,

microwave or traditional heating

Guo et al., (2012)

Dragon Fruit Extraction using ammonium Ismail et al., (2012)

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SOME SOURCES OF PECTIN

Source (Common Names) Mode of extraction Scientist/scientists

oxalate/oxalic acid

Sugar Beet Microwave-assisted Li et al., (2012)

Sweet potato Disodium phosphate

solution Zhang and Mu., (2012)

Sugar Palm Microwave assisted Rungrodnimitchai., ( 2011)

Apple pomace Hydrochloric acid and citric

acid

Kumar and Chauhan., (

2010)

Red Dragon Fruit Extraction using citric acid Woo et al., ( 2010)

Mulberry Use of Hydrocholoric acid Liu et al., (2010)

Passion fruit Mixture of acids Kliemann et al., (2009)

Taiwan tangerine HCl extraction Tamaki et al., (2008)

Passion Fruit Peel Citric acid Pinheiro et al., (2008)

Pumpkin Microbial enzymes Ptichkina et al., (2008)

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Traditional method of extraction

Extraction with hot water is by far the easiest and old method of extraction of pectin and

its related compounds substances (Huang, 1973) but the study of literature also revealed

the use of organic and dilute inorganic acids such as sulfuric acid aids in hydrolysis of

pectin thus giving good yield of pectin in less time (Jameson et al., 1924; Nanji et al

1927). Traditionally for commercial purposes the pectin was extracted with the help of

different mineral acids like sulfuric, hydrochloric and nitric acids. Sometimes

combination of hydrochloric acid with ion exchange resins were also used in the

commercial production of pectin (Rouse and Crandall, 1978).The main emphasis during

extraction of pectin historically was to improve the grade of pectin as jellying was only

considered the important parameter of pectin later it was found that there are other factors

which are also important to study as they affect in the jellying property of pectin.

Techniques like HPLC were used to determine the degree of methoxylation,

anhydrogalauronic acid and degree of acetylation (Voragen et al., 1986).

Different mode of extraction was also studied in order to understand the factors affecting

the yield of pectin. It was learned that temperature influenced the production of pectin in

the most positive manner and factors like rate of extraction and diffusion coefficient

investigated further. Although commercially pectin was extracted with the help of hot

diluted mineral acids like hydrochloric acid and sulfuric acid however the factors

influencing the yield were dependent on the type of source used to extract pectin and also

on the production methods of company’s manufacturing pectin.(May , 1990). The

industrial extraction of pectin by the use of mineral acids aroused numerous problems

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faced during the production of pectin which lead to the investigation of new process to

overcome those challenges. Enzyme extraction was one of the methods used so that the

problems like pulp maceration, difficult residual filtration and corrosion of the used

equipment were few of them (Sakamoto et al., 1995).

Physicochemical properties of pectin are also very important to understand and

investigate. The physicochemical properties of pectin depend upon many factors like

source from which pectin is extracted and also the process used for extraction. There are

others factors also which influence the extraction and yield of pectin are pH, temperature,

extraction time, type of solvents selected and the involvement of chelating agents like

EDTA and CDTA (Yeoh et al., 2008).Different techniques were used to determine the

actual structure of pectin in which spectroscopic techniques are used effectively.

Microwave was also investigated for better extraction of pectin for both in terms of yield

and quality. Image study of microwave extracted pectin was carried out to understand the

difference in quality or quantity of pectin extracted by using microwave (Zhongdong et

al., 2006).The method for the extraction of pectin has to be carefully designed in order to

get maximum yield with good quality of pectin. For this, numerous methods have been

studied and applied like direct boiling and microwave heating.

Microwave- assisted extraction

The traditional or conventional method of extraction of pectin is time consuming, it is

important to investigate for other methods for the extraction of pectin with good yield in

less time. For this purpose microwave heating is used presently (Koh et al.,

2014).Extraction with the help of microwave heating has given a desirable and positive

98

effect in getting good quantity of pectin (Maran et al., 2014).The good quality of pectin

includes low moisture and ash contents (Koh et al., 2014).The use of microwave (MW)

also helps in collecting high-quality antioxidant extracts from different plant materials.

This is because the fast and efficient heating of MW transmits heat straight to the

material under consideration releasing the phytochemical compounds readily and rapidly.

It also requires lesser amounts of solvent and time (Dorta et al., 2013).To optimize the

yield of pectin, extraction with numerous parts of the peel with different types of methods

has also been studied like flevedo and albedo of orange peels (Liu et al., 2006). A number

of other factors are equally important to consider while studying the yield, they are effect

of pH, extractant to peel ratio and temperature (Kulkarni and Vijayanand, 2010)

(Zhongdong et al., 2006). The yield obtained from microwave extraction was also

studies by applying statistical tools for the optimization of yield from the novel source of

pectin like apple (Wang et al., 2007). Not only apple but orange was also studied for its

effect of microwave heating on quality and quantity of pectin extracted from it. It was

noted that extraction with the help of microwave increased yield of pectin due to the

rupturing of parenchymal cells which assisted the release of pectin from the tissues.

(Kratchanova et al., 2004). Same source was used to determine the effect on yield of

pectin when pressure was applied along with microwave heating. The study also revealed

that molar mass, size and intrinsic viscosity of extracted pectin also increased as a result

of microwave heating during extraction of pectin from orange peel. This was confirmed

by size-exclusion chromatography (Fishman et al., 1999). Microwave pretreatment was

another method studied for its effect on the quality of novel sources of pectin like orange

lemon and apple (Kratchanova et al., 1996; Kratchanova et al., 1994).

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Enzymatic extraction of pectin

For the augmentation in the yield and to find convenient, less time consuming and

efficient process of extraction of pectin, different types of methods are studied and

investigated. Enzymatic extraction is one of them; use of enzymatic catalyst gave

competitive results (Dominiaket al.,2014). Currently large scale production of pectin

involves potent acids and elevated temperatures, Pectinolytic enzymes also help in the

degradation of pectin and hence used alternate methods for the extraction of pectin

(Mikami et al., 1987). The method is attaining popularity because it is eco-friendly.

Enzymes are excellent catalysts which can break or agitate cell walls and membranes

which aids in extracting considerable amount of functional compounds (Puri et al., 2012).

Polygalacturonase, which is acidic depolymerase, can also be used effectively in the

effective extraction of pectin (Franchi et al., 2014).

The enzymes mode of extraction of pectin involves breaking of pectin linkages lowering

the viscosity of the solution which in turns aids in the process of filtration and

centrifugation. More over enzymatic extraction is eco-friendly as it minimized water

pollution, the main drawback with this type of extraction can be the difficult and

expensive production of enzymes used in this process. The enzymatic extraction should

be monitored with great precision and care so as to stop the degradation of pectin at a

proper time otherwise it may affect the gelling property of pectin (Munarin et

al.,2012).The concentration of enzyme affects the yield of pectin as well as the molar

mass of pectin however no effect on its GalA concentration (Yuliarti et al.,

2011).Celluclast, Econase and Viscoferm are few examples of enzymes which gave good

results (Wikiera et al., 2015).Other new methods like application of high hydrostatic

100

pressure technology for enzymatic extraction has also been reported (Naghshineh et al.,

2013). A study done using microwave assisted flash extraction under control pressures

and temperature claims that microwave assisted extraction (MAE) extracts provides high

molar mass, moderate viscosity pectin in less time using low temperature when

compared to conventional heating (Fishman et al., 2008). Additional methods include

use of microbial enzymes is also reported (Ptichkina et al., 2008).

A number of solvents have been used by different worker to extract pectin, however, the

best results was reported from ethanol. Oxalic acid based extraction also gives good

result when Microwave assisted extraction is used(Tan etal., 2014).The Quantity of

solvent used also has an effect on yield of pectin (Chan and Choo,2013) the ratio of citric

acid required to make the solution pH 2.5 gave better results than with the ratio which

made the solution pH 4.

The use of ionic liquids are also found in past researches and has proved themselves as

an effective alternate solvents in extraction of biologically active compounds with the

help of microwave. They are not only effective but also environmental friendly (Guolin et

al.,2012;Du et al., 2007).The ionic liquids are a combination of organic cations and

inorganic or organic anions (Du,et al., 2007). Whereas the use of new solvents such as

ammonium oxalate (Ismail et al., 2012)disodium phosphate are also investigated (Zhang

and Mu, 2011). Optimization with the help of organic acids like nitric acid has also been

conducted (Vriesmann et al., 2011).

101

Purification of Pectin

For the purification of pectin different methods have been reported, Yapo, and Koffi,

(2014), first purification pectin using 95% ethanol and thus washed pectin precipitates

two-times with 70% ethanol, followed by 95% ethanol and acetone before drying them in

oven. In another study, Shaha et al (2013) washed the precipitated pectin through 95%

ethanol, followed by 55% and then with 70% ethanol. Isopropanol and acetone were also

used for washing pectin for further purification (Ismail et al., 2012) .Different methods

have also been discussed and compared by Sotanaphun, (2011).The authors used dialysis,

Amberlite XAD-16 polystyrene resins for the purification of pectin. Some of the very old

methods used for the purification of pectin include treatment with bromine, chlorine

water or iodine (Krishnamurti andGiri 1949) (Nanji and Chinoy, 1934).Celite also used

for purification since long. It was used in the extraction of pectin to remove impurities

and color that may be present in extracting solution (AOAC, 2012)Patents have also been

granted for celite to be used for the purification of pectin. Celite was used to filter the

solution containing soluble pectin before final precipitation with isopropanol. The celite

was used on a nylon cloth and the solution was filtered on a sieve and also as a filter aid

during vacuum filtration. (Buchholt et al., 2005). Recently a research is being conducted

in which celite 545 is used in the purification of pectin (Dominiaket al., 2014). Figure -13

to 15 shows different lab scale extraction of pectin while Figure- 16 to 18 shows different

industrial scale production of pectin.

102

Figure -14: Extraction of pectin from soy hull (Monsoor and Proctor, 2001).

Soy hull (400 g)

Extraction with 0.05 N HCl

Collect supernatant

Add equal volume of isopropanol

Adjust pH to 3.5

Collect Precipitate

Wash two times with isopropanol

Final wash with 70% isopropanol

Collect precipitate

Disperse with deionized water

Spray dry

103

Figure-15: Extraction of pectin from leaves of Nephrolepis biserrata (Pagarra et al 2014)

Nephrolepis biserrata leaves

Drying of sample

Add aquadest adjusted to 1;40 w/v

Add 0.5 H2SO4

Sample solution at pH 1.5

60; 90; 120 minutes of boiling at 80OC

Filtiration

Add 95% Ethanol (1:1 v/v)

Precipitation, 1 day

Washing with 95% Ethanol

Precipitized

Dry oven 40˚C

Yield of Pectin

104

Figure- 16: Extraction of pectin from raw papaya peel (Boonrod et al., 2006)

Raw papaya peel 15g

Treated material

Pectin solution

Pectin precipitate Pectin

Precipitate

Ethanol-insoluble

pectin

Aluminum-insoluble

pectin

Route

1

Route

2 1\2

Mincing

pH adjustment to 4-4.5

Precipitation with 95% ethanol

for 2 hr

Filtration

Acid-extraction in boiling water bath

with 0.06 M HCl 3 times at volume ratio

of 2:1, 1:5:1 and 1:5:1

Centrifugation at 4,500xg 30

pH adjustment to 3.5

Precipitation with 1.2%

aluminum chloride, pH 4.0 for 2

hr. Filtration

Washing with 70%

ethanol

Oven-drying at 37˚C

Washing with 70%

ethanol

Oven-drying at 37˚C

105

Figure-17: The commercial production of pectin ( Source: ENSYMM)

Citrus Waste

Hydrolysis Reactor

Expansion Tank

Filter

Condense/Decanter

Biogas Digester Precipitator Fermenter

Dryer

Distillation

Sulfuric

Acid Water

Steam

Limestone

Hydrolysat

e Soli

d

Biogas Pectin

Dried Pectin

Ethanol

Stilage

Liquid

Pectin

Depleted

Residue

106

Figure- 18: Commercial production of pectin (Source: Cargill foods)

107

Figure-19: Commercial production of pectin (Source: GENU)

Uses and application of pectin

The family of polysaccharides containing α-(1-4)-linked D-galactopyranosyluronic acid

residue is commonly known as pectin (Yeh et al., 2011). Pectin, a part from cellulose and

hemicellulose is one of the class of polysaccharides involve in the formation of primary

cell wall (Besson et al., 2013). Well known and traditional use of pectin is in food

industries, yet several pharmaceutical uses and applications have also been reported. Use

of Pectin in the production of jams and jellies has provided its additional application in

the manufacturing of many other types of food products, which include dairy products,

desserts and soft drinks (Munarin et al., 2012). Pharmaceutical uses include cholesterol

lowering formulations (Brouns et al., 2011), pectin-derived oligosaccharides considering

as a new source of prebiotics (Gullon et al., 2013), Ophthalmic and nasal formulations

(Mittal and Kaur, 2014 and Morris et al., 2010) and colon specific delivery systems

Pre treatment

of raw material Extraction Filtrations

Water + acid Filter aids

FRUIT

Insoluble material

ion-exchange

(calcium

removal)

Evaporative Concentration

de-

esterficiation

water acid

Precipitation de-esterficiation

and/ or amidation alcohol

washing

drying and

milling blending and

standardization

alkali or

ammonia alcohol

alcohol

alcohol

recover

y Solid waste( alcohol

soluble)

Standardized

Pectin Sugar and other pectin

preparations

Optional

process

Process

aids

Waste/re

cycle

Raw

materials/

products

Standardizing

agents

108

(Kushwaha et al., 2011). Recently, studies on its cosmoceutical applications have also

been undertaken (Suh et al., 2014). New approaches include development of hydrogels in

presence of proteins which can be used in the formulation of low calorie food products

(Wu et al., 2014).

Among many current known pharmaceutical uses pectin is under constant consideration

for finding even more important and beneficial uses. It is studied that both, pectin source

and physico-chemical properties of pectin renders its effect in lowering of cholesterol in

humans. The physico-chemical properties include molecular weight (MW), viscosity and

degree of esterification (DE) (Brouns et al., 2011).Among the advancements in new drug

delivery systems,pecsys is a new technology in which the pectin-based formulation, gels

when it comes in contact with mucosal surface. Such formulations gives rapid absorption,

controlling Cmax, reduces runoff and high bioavailability (Archimedes Pharma, 2011).

Pectin is also studied for the formulation of microencapsulated drug delivery systems in

which it is used in the form of biopolymer particles and are used in both food and

pharmaceutical preparations (Jones et al., 2010).The said drug–delivery systems are

under close observation through clinical trials in order to understand the functionality and

side effects (Watts and Smith, 2009).Pectin are important part of soluble dietary fibers;

these fibers are essential for proper physiological well-being of humans and are used to

treat dyslipidemias and other diseases (Dhingra et al., 2011).The source of pectin used for

pharmaceutical purpose are citrus peels and apple pomace. Different formulations have

been made and studied in connection with the colon-specific drug delivery systems.

Pectin microspheres have been under in-vitro investigations and the drug release from

microspheres are being understood in different conditions which may affect the release of

109

the drug (Kushwaha et al., 2011). The diversified properties of pectin can be associated

with its complex structure that’s the reason pectin is studied for its anti-tumor activities

too (Maxwell et al., 2012). But this association of pectin structure with its bioactivity

cannot be fully explained in literary works as the relationship with origin of pectin and its

possible chemical modification which results in molecular fragmentation are not fully

expressed (Leclere et al., 2013). Recently a work has been done on nanocomposite Pectin

scaffolds which will provide localized therapy in ovarian cancer (Chandran et al., 2013).

Pectin is still in use as antidiarrheal agent in formulations with kaolin and other

ingredients. Some currently marketed products are shown in Figure-20 with their

country/region of availability.

Pectin is commonly used as gelling, thickening and stabilizing agents in foods and to a

certain extent in pharmaceuticals as well. Pectin is mostly used to construct the desired

texture of products which result in controlling the moisture or water in the product

(Imason , 2010). Traditionally pectin was used mainly in food and food preparations, due

to its thickening and jelling properties, jams and jellies are few examples. Pectin is

considered very well tolerated and safe among numerous food additives therefore it is

categorized and recognized in “Acceptable Daily Intake (ADI) levels of “not specified”

products by the FAO/WHO Joint Expert Committee on Food Additives (JECFA). (Food

and Agriculture Organization of the United Nations 2009; Endreb and Christensen,

2009). The stature is equivalent in European Union where Pectin E440 (i) and Amidated

Pectin E440 (ii), both have been accustomed as ADI "not specified" by the Scientific

Committee for Food Specifications. Where as in United States pectin is designated as

GRAS (generally recognized as safe) by the FDA (U S Food and Drug Administration;

110

Endreb and Christensen, 2009). Moreover, it is permissible to use pectin in all non-

standardized foods and in accordance with the Code of Federal Regulations (21CFR

184.1588) Table-3 shows different uses of pectin in food industry. Due to the trust of

consumers and regulatory authorities on inertness and safe nature, pectin is gaining

popularity in making of cosmetics and personal care products. CP Kelco has number of

patents in this field such as GENU pHreshTM pectin which restores skin and maintain a

skin friendly pH has been presented (CP Kelco). Trudsoe et al (2014) describes use of

acidified pectins in personal care products that can be good for human skin. Personal care

products may comprise of a styling gel, moisturizer, lotion, deodorant, toothpaste, body

wash, bath gel, body gel, hand sanitizer, shampoo, conditioner, or combinations.

111

Table - 3: Showing different uses and properties of pectin.

(Source: J.F. Hydrocolloids)

Application/uses of pectin Role of pectin

1. Meat and Poultry

Provides neutral flavor and pH.

Helps in absorption of oil

Provides high content of dietary fiber.

Have more water retention capabilities

2. Fruit and Based (inc. fruit‐based

beverages)

Different types of pectin used having different

setting time

Provides nice and delicate texture.

Enhances body and mouthfeel.

Regulates setting time and temperature.

Helps in flavor release.

Viscosity of beverages is maintained

3. Dairy (inc. dairy‐based beverages) Provides optimum stability of heat treated

dairy drink.

Protects whey separation and casein

precipitation.

Enhances shelf‐life.

Smoother mouthfeel.

Helps flavor release

4. Confectionary Provides nice and delicate texture.

Regulates setting time and temperature.

Helps in flavor release.

5. Bakery Provides delicate and nice texture.

Regulates setting time and temperature.

Enhances flavor release.

112

A

B

c

Figure – 20: A and B available in Middle East while C is available in Australia.

113

Large scale production of pectin

Commercial pectins are largely extracted with acid extraction method from citrus peels

and apple pomace and the yield is usually 25 and 12% pectin, respectively (Ismail Sah et

al., 2012). As far as the structure of commercial pectin is concern they are less complex

as the processes of extraction and purification in industry removes almost all the neutral

sugars in pectin (Seymour, 2002). The pectic skeleton is made up of homoplymer of

galaturonic acid binded with (1→4) linkage with partial methyl esterified carboxyl

groups. According to United Nation’s Food and Agriculture Organization [FAO] and

European Union [EU] commercial pectin must be composed of atleast 65% of

galacturonicacid (Calderon, 2012).Flow chart shows a general procedure of commercially

extracted pectin. Apple pomace and citrus peels are the major raw materials which are the

main source of industrial pectin. But the supply of raw material also dependent upon

available local crop sources. Therefore some other types of raw materials are also used in

different parts of the world which includes potatoes pulps, sugar beet pulp, sunflower

heads etc. although the industrial process for the extraction of pectin is explained by

many scientists but detailed process differ among companies, but the general process is as

follows.

The materials used for the extraction of pectin are heated using reflux technique with

dilute mineral acid for 1 -10 hours at a pH approximately equal to 2 and at temperature of

60-100°C. If the source is apple or peach pomace after separating the hot pectin extract

from solid residue, to hydrolyze starch pectinase-free α-amylase is mixed. The obtained

extract is reduced in volume under vacuum to an approximately 4% pectin material and

114

then it is precipitated with 2- propanol. The precipitated pectin is then washed, dried and

reduced in particle size after grinding. For different types of pectin different mediums are

used for example for low DE values acidic treatments are used and when ammonia is

used amidated pectin were produced. The process is summarized in Figure- 21.

115

Figure -21: Flow chart showing general procedure employed for the extraction of

pectin on commercial scale. Turakhozhaev and Khodzhaev (1993).

Preliminary treatment of raw

material

Protopectin hydrolysis and

pectin extraction

Filtration and concentration

Precipitation and purification

Drying and grinding

Standardization

116

3. MATERIALS AND METHODS

Equipment

The equipment used in this study are as under

Analytical balance

OHAUS (PA 214) USA, Sartorious GmbH; type A 6801

Water bath

Thermostatic Grant Type JB2 water bath. USA

Homogenizer

Panasonic MX-J120P mechanical Blender and grinder.Japan

pH meter

Jenway 3510 pH meter. UK

Microwave oven

National (IEC-705) 700 W microwave. Japan

Freeze Dryer

Tro,science Co, Limited freeze dryer, model number TR-FD-BT-50 Japan

117

Spectrophotometer

A Nicolet Evolution 300, Thermo Electron Corporation, USA,UV-Vis spectrometer 1800

(Shimadzu, Kyoto, Japan)

FT-IR spectrometer

Thermo Nicolet 380, FTIR Instrument. USA

Centrifuge

Laboratory centrifuge shanghai China, model number 800 , Heltich Universal 32R

Tablet Punch Machine

Korsch, Erweka, Germany

Oven

Memmert, Germany

Furnace

Gallen Kamp Scientific Tech. UK

Hardness Tester

Fujiwara, Seisukusho Corporation, Japan

Friabilator

H.Jurgens and Co-Gmbh, D2800, Germany

118

Disintegration apparatus

Eeweka, ZT2, Heusenstamm Germany

Dissolution apparatus

Erweka DT, Heusenstamm, Germany

Refractometer

Hand held ATAGO refractometer, Japan

Viscometer

Brookfield viscometer

Laser spectroscatter

Laser spectroscatter-201 system , RiNA GmbH Berlin, Germany

Mixer

Eisco scientific

119

LIST OF CHEMICALS AND REAGENTS USED

BDH-VWR Laboratories

Industrial methylated spirit, ammonium hydroxide, celite, sodium hexameta- phosphate,

charcol, tartaric acid, phosphorus pentaoxide, sodium tetraborate.

Fisher Scientific

Ammonia, ethylenediamine tetraacetic acid (EDTA).

J T Baker

Hydrochloric acid, sulfuric acid.

Merck

Acetic acid, ethanol, nitric acid, sodium carbonate, sodium chloride, sodium citrate,

sodium hydroxide, m-hydroxydiphenyl, magnesium sulphate, oxalic acid, citric acid,

phenophthelene, phenol red, magnesium sulphate, potassium dihydrogen phosphate,

methanol, lactose,Propyl Paraben (Sodium),

Sigma- Aldrich

Galacturonic acid, pectin, sodium azide, bronopol, Microcrystalline cellulose

Mallinckrodt Speciality Chem Co

Paracetamol, kaolin

120

Fauji Pakistan

Starch

Vijai Minerals

Talc

Taihei Chemical Industrial Co. Japan

Magnesium stearate

Sensient Colors UK

Erythrocine

BASF Aktiengesellschaft, Germany

Bronopol, Ibuprufen

Asian Chemical Company Limited, Japan.

Sodium Saccharin

Nippon Chemical Trading Co. Japan

Sodium carboxy methyl cellulose

Lever Brothers Ltd. – Pakistan

Glycerine

121

Vanderbilt Minerals, LLC

Veegum

Local preparation

Distilled water

Deionized water

122

LIST OF APPARATUS

Knives ( IKEA) , motor and pestle( porceline), kitchen steak hammer (IKEA), kitchen

hand chopper (IKEA), muslin ( local market), burner( made in China) , sieve (80 mesh

sizes), metal dish (5 cm in diameter with lid), sieve ( number 16, 20 and 40) , stainless

steel pan ( local market) , wooden spoon ( Local market) , litmus paper, distillation

assembly , jelly vessels, ice box, burette, pipette ,buchner funnel (porceline), vacuum

pump, porcelain crucible (Yancheng Yongkang Lab Instruments Factory, Jiangsu,

China), reflux condenser (Shanghai Heqi Glass ware Co., Ltd., China), diaphragm

vacuum pump. vernier caliper in mm (CD-6, CSX, Mitutoyo, Japan), brookfield DV-E

viscometer, laboratory glass wares beakers (100ml, 500ml, 1000ml, 3000ml ), glass rods,

glass funnels, test tubes, right arm flask , conical flask (250ml) , volumetric flask (

100ml, 250ml), glass jars with lids, measuring cylinder( 100ml)] (Pyrex, England),

porcelain crucible (Yancheng Yongkang Lab Instruments Factory, Jiangsu, China),

desiccator (Duran, Germany),tripod stand. Centrifuge tubes (30ml), sintered glass

crucible, filter (0.22 mm Millipore, USA), quartz SUPRASIL ®.

123

LIST OF SOLUTIONS USED

Hydrochloric acid (HCl) solutions

0.1N: The solution was prepared by slowly adding 8.212 ml of concentrated HCl

to 250 ml deionized water. The final volume was adjusted to 1000 ml with deionized

water.

1N: The solution was prepared by slowly adding 82.117 ml of concentrated HCl

to 250 ml deionized water. The final volume was adjusted to 1000 ml with deionized

water.

Sodium hydroxide (NaOH) solutions

0.5N: The solution was prepared by dissolving 22 g of NaOH in 1000 ml of deionized

water.

0.1N: The solution was prepared by dissolving 4.5 g of NaOH in 1000 ml of deionized

water.

Sulphuric acid solutions

0.5N: The solution was prepared by slowly adding 14.027 ml of concentrated sulphuric

acid to 250 ml deionized water. The final volume was adjusted to 1000 ml with deionized

water.

124

1N: The solution was prepared by slowly adding 28.024 ml of concentrated sulphuric

to 250 ml deionized water. The final volume was adjusted to 1000 ml with deionized

water.

Nitric acid solution

1N: The solution was prepared by slowly adding 63.704 ml of concentrated nitric acid

to 250 ml deionized water. The final volume was adjusted to 1000 ml with deionized

water.

Oxalic acid solutions

1%: To make 1 % solution, 10 g of oxalic acid was dissolved in 1000 ml of deionized

water.

10%: To make 10% solution, 100 g of oxalic acid was dissolved into 1000 ml of

deionized water.

Tartaric acid solutions

1%: To make 1 % solution, 10 g of tartaric acid was dissolved in 1000 ml of deionized

water.

10%: To make 10% solution, 100 g of tartaric acid was dissolved into 1000 ml of

deionized water.

125

Citric acid solutions

1%: To make 1 % solution, of 10 g of citric acid was dissolved in 1000 ml of deionized

water.

10%: To make 10% solution, 100 g of citric acid was dissolved into 1000 ml of

deionized water.

Citric acid solution for jelly grade

50 g of citric acid was dissolved in distilled water and diluted to 100 ml with deionized

water.

Sodium citrate solution

25 g of sodium citrate was dissolved in distilled water and dilute to 100 ml with distilled

water.

Galacturonic acid stock solution

Galacturonic acid stock solution was prepared by dissolving 100 mg dry galacturonic

acid powder in 100 ml distilled water. This makes the concentration of the solution to be

1mg/ml. The solution was refrigerated. Fresh solution was prepared if the sample is kept

longer than 4 weeks.

M/80 Sodium tetraborate in sulphuric acid (0.0125M solution)

1.192 g of sodium tetraborate was weighed and transferred added into a 250 ml

volumetric flask and the volume was adjusted with concentrated sulphuric acid.

126

Sodium hydroxide (5%)

5 g of sodium hydroxide was transferred in a 100 ml volumetric flask and the volume

adjusted with deionized water.

m-hydroxydiphenyl solution (0.15%)

0.15 g of m-hydroxydiphenyl was weighed and transferred into a 100 ml flask and the

volume was adjusted with the help of 0.5% sodium hydroxide solution and refrigerated.

Phosphate buffer saline pH 7

1.20 g of sodium dihydrogen phosphate and 0.885 g of disodium hydrogen phosphate

were dissolved in 1 liter volume distilled water.

Tris saline buffer

20 mM Tris–HCl buffer was taken in a beaker the pH was adjusted to 7.5with 150mM

sodium chloride (NaCl) and 0.01% sodium azide (NaN3).

127

Extraction of pectin

Processing and preparation of peel

Fruits were purchased from the local market, Karachi, Pakistan and identified. Their

voucher specimen numbers were deposited in the department of Pharmacognosy, Faculty

of Pharmacy and Pharmaceutical Sciences, University of Karachi, Pakistan. All fruits

were washed with distilled water properly before subjecting to any procedure. Excess

water was drained and after air drying, the fruits were peeled off using sharp knife. The

respective peels thus obtained from different fruits were cut into thin slices of few mm in

thickness, weighed at a constant weight and used for extraction.

Screening of seasonal fruits and their wastes for the presence of pectin

contents

The method of extraction of pectin used was originally described by Patel et al (2012).

The modified method used for extraction in the present study is presented in Figure-20.

20 g of thin sliced peels of each fruit were dipped in 100 ml of boiling industrial

methylated sprit (IMS) and each content were boiled for 5 min on a water bath. The IMS

was then carefully decanted in a separate container. The peels obtained after boiling in

IMS were transferred in the jar of a mechanical blender and after adding small quantity of

DI water the contents were grinded for about 30 sec and then transferred the whole

content in a beaker to form loose slurry.

The slurry obtained from mechanical procedure of each fruit peels were decanted

separately into beakers and code numbers were assigned. The volume was raised to 60ml

128

for each test performed. The contents were boiled for 10 min using bunsen burner. The

mixture was then brought to room temperature and DI water was added in each beaker to

adjust the volume, if evaporated during boiling. The pH of the each content was recorded

and adjusted to desired pH 6 ±0.5, either using NH4OH or 0.1N HCl. The solids of each

content were separated, first using a Buchner funnel, followed by centrifugation at 4000

rpm. The supernatants, thus obtained from each content were then further processed by

adding ethanol (95%) in 1:4 ratios which is approximately 240ml. The addition of ethanol

lead to precipitation of pectin in the liquid phase, which is then separated with the help of

centrifugation, freeze dried, weighed and percentage yield was calculated. All freeze

dried materials in glass vials were kept in refrigerator for further study. The list of fruits

which were screened is shown in annexure 1.

Extraction of pectin from selected fruit wastes and their comparative

study

Materials and methods are same as described for the screening of seasonal fruits and their

wastes for the presence of pectin contents. Three fruits were selected in view of screening

results for the identification of new pectin source, ease of availability and cost. Fruits

selected were “Banana”, “Sapodilla” and “Muskmelon”. The fruits were screened for

their maximum pectin release, subjecting them to different mechanical procedure at

different pH using two different heating methods. Same procedure was adopted for

standard (apple and orange) fruits to improve yield in order to get clear yield comparison

between mechanical procedures, pH and heating methods. (Figure-22). A pictorial

representation is given in annexure 2.

129

Mechanical, pH and heating factors used during the study

The extraction procedure for all five fruits was kept similar as used and described in

screening study with slight modification. The peels of the selected fruits after boiling in

IMS were subjected to different mechanical tools, such as, homogenization, grinding,

cutting, chopping and hammering.

Homogenization Grinding Cutting Chopping Hammering

For each mechanical procedure, following techniques were applied:

Homogenization: The peels obtained after boiling in IMS were transferred in the jar of a

mechanical blender and after adding small quantity of DI water the content was grinded

for about 30 sec. and then transferred the whole content in a beaker to form loose slurry.

Grinding: The peels obtained after boiling in IMS were transferred in the jar of a

mechanical grinder and grinded without adding DI water for 30 sec. The grinded peels

were then transferred into a beaker containing required amount of water to form loose

slurry.

Cutting: The peels obtained after boiling in IMS were transferred in the container of a

hand chopper and sized / reduced into small slices by pressing the hand chopper 200

130

times. The processed peels were then transferred into a beaker containing required

amount of water to form loose slurry.

Chopping: The peels obtained after boiling in IMS were transferred in mortar and the

peels were chopped by using a pestle for about 200 times. The processed peels were then

transferred into a beaker containing required amount of water to form loose slurry.

Hammering: The peels obtained after boiling in IMS were transferred on a cutting

board and hammered for about 200 times. The processed peels were then transferred into

a beaker containing required amount of water to form loose slurry.

pH: Five strengths of pH were selected .The pH of the each slurry was maintained at

1,3,5,6 and 7 respectively. The slurry was checked for the initial pH and then HCl (0.1N)

or NH4OH were used to adjust the pH.

Heating methods and time. Two heating methods were used during this procedure

which were microwave heating and bunsen burner. The mixture was boiled for 10 min

using both the heating modules.

131

Figure-22: Showing flowchart of extraction of pectin followed during the current

Fruit peels 20 gms

100 ml IMS

60 ml H₂O

Adjustment of pH with HCL/ NH₄OH

Purification by passing through celite

Filtrate ------------> Add Ethanol 1:4 ratio

Centrifuge at 4000 rpm

Freeze Dryer

Percent Yeild

Boiling for 5 min

Boil for 10 min

Filter with cheeze cloth

132

study

Effects of organic acid and inorganic acids and their strength on pectin

yield from sapodilla

To determine the best extraction procedure, sapodilla fruit was subjected to different

types of extraction procedures using organic and inorganic acids, maintaing pH around

4±0.5and NH4OH to observe the yield at pH 6 to 7. The basic method used for extraction

of pectin was kept similar as used in screening of seasonal fruits and their waste for the

presence of pectin contents. The difference in the type, and concentration, strength of

organic and inorganic acids used are described below.

Extraction with 0.1N HCl, using 5 mechanical procedures, 5 different

pH, two boiling methods and varying time of boiling (10, 20, 40 and

60min)

In this procedure peels of sapodilla were selected and were subjected to the same

extraction procedure as mentioned in preceding text with the exception that the peels

were boiled for 10, 20, 40, and 60 min using both burner and microwave. For the

maintenance of pH 0.1N HCl was used.

Extraction using 1N HCl, using 5 mechanical procedure, 5 different pH,

two boiling methods and varying time of boiling (10, 20, 40 and 60min)

Aforementioned procedure is followed except that for the maintenance of pH 1N HCl

was used.

133

134

Extraction using organic acids

The same foregoing procedure is used except that for pH maintenance three different

organic acids oxalic, citric and tartaric acid were used in 1% and 10% concentrations.

Extraction with different strengths of inorganic acid

In this procedure after following the same extraction method described earlier the pH

was maintained with three different strengths (0.1N, 0.5N, and 1N) of same inorganic

acid (Hydrochloric acid).

Statistical Evaluations

Three factor factorial completely randomized design (CRD) was applied and mean

comparison was done by using Tukey HSD (Steel et al., 1997) at 5% level of significance.

Statistical analysis was performed using SPSS13 and Minitab13.1.

Physical and biochemical characterization of pectin

Percent yield

The percent yield of the dried pectin was calculated as follows

% Yield =Weight of vial with pectin

Weight of empty vial× 100

Identification tests for pectin

135

Identification tests were carried out according to the procedures performed by Sharma et

al (2013).

Stiff gel test

1g of pectin with 9 ml of distilled water was heated on a water bath to get a solution and

cooled

Test with 95% ethanol

To a 1% w/v solution equal volume of ethanol (95%) was added.

Test with potassium hydroxide (KOH)

5ml of 1% w/v solution of pectin sample was prepared and 2% w/v solution of KOH was

added, the solution was left to stand for 15 min.

Iodine test

To a freshly boiled 5ml solution of 2% w/v pectin, 0.15ml of iodine solution was added.

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Biochemical characterization

Preparation of sample

The dried pectin was stored in a cool dry place. The sample is oven dried and used for all

the tests performed.

Qualitative test for ammonia

To 0.5g of dried pectin, 1ml of 0.1N NaOH was added and the sample was heated to

observe the release of ammonia if any (Ranganna, 1986).

Detection of Moisture, ash, methoxy content and equivalent weight

Moisture, ash, equivalent weight and methoxy content of pectin was determined by the

procedure used by Aina et al (2012) and Ranganna (1986) with slight modifications.

Moisture

1g of sample was weighed in a tared metal dish. The sample was dried at 100OC for four

hours after which the dish was cooled in a dessicator.

Ash

1 g of pectin was taken and weighed in an empty tarred crucible. The crucible was heated

at 600OC in a furnace for 24 hrs. The crucible was then taken out of the furnace and set

aside to cool in a desiccators. After cooling the crucible was weighed again and the ash

was calculated with the help of following formula:

137

Ash content (%) = (Weight of ash / Weight of sample) × 100

Equivalent weight

0.5 g of pectin was taken into a 250ml conical flask. The pectin was soaked with 5ml

ethanol and 1.0 g of NaCl was added, 100ml of distilled water and a few drops of phenol

red indicator was also added to the flask to completely dissolve the content. The solution

was then titrated with 0.1M NaOH to a pink color endpoint. Equivalent weight was

calculated using the equation below:

Equivalent weight = (Weight of sample / Volume of alkali (ml) × Molarity of alkali) ×

100%

Methoxyl content

A previously neutralized solution (obtained after the equivalent weight determination)

was used to determine methoxy content. 25ml of 0.25 N sodium hydroxide was added,

shaked vigorously and allowed to stand for 30 min at room temperature. The mixture was

then titrated with 0.1N NaOH to the same end point as used in equivalent weight. The

methoxyl content was calculated with the equation given below:

Methoxyl content % = (Volume of alkali (ml) × Normality of alkali × 3.1/ Weight of

sample (mg)) × 100

Anhydrouronic Acid.

The anhydouronic acid (AUA) content was calculated by the method suggested by Azad,

et al., 2014. The AUA was determined by the following equation:

138

% of AUA= 176x0.1zx100 + 176x 0.1y x100

w x1000 w x1000

When molecular unit of AUA (1 unit) = 176 g

Where,

z = ml (titre) of NaOH from equivalent weight determination.

y = ml (titre) of NaOH from methoxyl content determination.

w = weight of sample

Grading of pectin

Grade of jelly was determined by the method described by Ranganna (1986). 320 ml of

water was taken separately in a kettle, 500 g of sugar was mixed with 3.3 g of pectin

assuming the grade of pectin is 150. To the water in kettle 0.5ml of the citric acid

solution and 1 ml of sodium citrate solution was added. The pectin sugar mixture was

also added later and stirred thoroughly. The mixture was heated rapidly with constant

stirring. The mixture was removed from flame occasionally to avoid the excessive

evaporation of water. The mixture was heated till the weight of solution became 770g.

When the solution reached at the desired weight it was removed from the flame and

allowed to cool for 30 sec at room temperature. 2 ml of citric acid and 0.5ml of sodium

citrate solution was then added. The jelly was transferred into a vessel or any other

desired container and allowed to stand for at least 18 hours at 260 C. After 18 hours the

jelly was checked for overall firmness and compared with standard jelly prepared with

same procedure. The process is repeated with the increased amount of pectin if there is a

difference between the consistencies of standard and sample jellies.

139

Galacturonic acid content

Pectin assay using m-hydroxydiphenyl

Pectin Assay using m-hydroxydiphenyl was used to determination of pectic substance in

the desired samples. The colorimetric assay using m-hyrdoxydiphenyl was used for its

specificity for uronic acids as it can tolerate the presence of sucrose upto 1000 ppm.

Preparation and measurement of samples:

2 ml solution from the galacturonic acid stock solution was taken into a 100 ml

volumetric flask and the volume was made up with deionized water to make 20

microgram / ml of galacturonic acid. Similarly different concentration (40, 60 and 80

micrograms / ml) of galacturonic acid in the sample were also made. Sixteen test tubes

were placed in an ice box to cool before the experiment started. Three tubes were used

for each sample (2 for samples and one is used for blank determination). The sulphuric

acid/ sodium tetraborate solution was also kept in an ice bath throughout the experiment.

1.0 ml of all the test material was then filled in each of the three respective labeled cold

test tubes and was left to cool for a minute. About 6 ml of sodium teteraborate solution

was then added to the test tubes and mixed thoroughly with the help of a test tube stirrer.

The tubes were kept in the ice box until all the samples were prepared. The tubes were

then heated for about 6 minutes after which the tubes were placed again in the ice bath

for cooling. 0.1 ml of m-hydroxydiphenyl was then added to the first tube to develop

color; it was also mixed thoroughly with the stirrer. For blank 0.1ml of 0.5% sodium

hydroxide was taken in the third tube. It was also mix thoroughly and was allowed to

stand for 15–20 min at room temperature to dissipate any bubbles formed during stirring.

140

The absorbance was measured at 520 nm on a spectrophotometer by reading sample

against corresponding blank tube with 0.5% sodium hydroxide. The total absorbance for

each sample was obtained by subtracting absorbance due to m-hydroxydiphenyl and the

absorbance for sample blank. A calibration curve of Absorbance (y-axis) was plotted

against concentration of galacturonic (x-axis). This curve was use for standardization.

Water holding capacity (WHC)

The method used for water holding capacity was measured using Daou and Zhang,

(2011) method with slight modification as the sample used here was dried pectin. 1g

sample was accurately weighed in a graduated test tube. The sample was hydrated with

0.02% sodium azide dissolved in 30ml of deionized water for 18hours. After the assigned

period the supernatant was removed and the sample was drained into already weighed

filter paper which was dried to a constant weight in a forced-air oven at 110OC and

weighed again. WHC was expressed as the amount of water retained per gram dry sample

(g/g dry weight).

WHC (g/g) = (Hydrated residue weight-Dry residue weight)/ Dry residue weight

Water binding capacity (WBC)

The water binding capacity was also determined after slight modification of the method

described by Daou and Zhang (2011). 1g sample was taken in a graduated centrifuge

tube, 0.02% sodium azide dissolved in 30ml of deionized water was added and left at

room temperature for 18 hours. The tubes were centrifuged (3000 rpm) for 20min, the

supernanatent was separated by passing through a sintered funnel under vacuum. The

141

weight of the hydrated sample was noted and the sample was dried at 105OC for 24 hours

to get a constant weight. The value was expressed as the amount of water retained per

gram dry sample

WBC (g/g) = Residue hydrated weight after centrifugation -Residue dry weight / Residue

dry weight

Fat binding capacity (FBC)

The method used for fat binding capacity was previously used by Daou and Zhang,

(2011). 5g sample was taken and added to 20 ml soybean oil in a 50 ml centrifuged tube.

The mixture was then stirred after every 5 min for 30 sec and after 30 min the tubes were

centrifuged at 1600 rpm for 25 min. The free oil was discarded and the absorbed oil was

determined by weight difference and was expressed as ml (oil)/gram sample

FBC (ml/g) = (precipitation weight-Dry weight)/Dry weight

FTIR spectroscopy of selected pectin

FT-IR spectrometer was employed to investigate the characteristic spectra of the

extracted pectin obtained through different variations in extraction procedures. Dried

sample (1 g) was ground and analyzed through Smart Arc attachment, thereafter it was

scanned within the range of 4000-400 cm-1 (Park et al., 2006).

Dynamic Light Scattering (DLS) Analysis

In order to find some basic information regarding the particle size and polydispersity of

pectin. DLS studies were performed as described by Hameed et al., 2014. A laser

142

spectroscatter-201 system with a He–Ne laser providing a 690 nm light source and an

output power in the range of 10–50 mW was used in the study. An auto piloted run of 50

measurements at every 20 s, with a wait time of 1 s was used at 25 C was used for all

measurements. Standard and all other pectin samples was prepared (2% w/v) in Tris–

saline buffer .Samples (20 µl), reagents and buffers were filtered using 0.22 mm filter

(Millipore, USA) before filling into a special quartz SUPRASIL ® cell (light path 1.5

mm) for measurement. The scattered light was recorded at a constant scattering angle of

90o. The autocorrelation functions were evaluated using the CONTIN program to obtain

hydrodynamic radius (RH) distributions. The RH is associated with the diffusion

coefficient by the Einstein–Stokes equation. The data were analyzed using Xtal Concepts

software (XtalConcepts GmbH, Hamburg Germany) present with the instrument.

Optimization of the yield of pectin from sapodilla peel thorough

Response Surface Methodology (RSM).

The sapodilla peel was further studied in order to establish the optimum conditions of

extraction to get the maximum yield of pectin. To determine the optimum extraction

conditions for the extraction of pectin Box-Behnken design was used. A response surface

experimental design, with three independent variables was used to extract pectin from

sapodilla fruit peel. The variables used were time of boiling, temperature at which the

extraction was carried out and pH of the extracting medium. The design of experiment

and regression analysis was performed for response surface methodology by Minitab

(ver. 14) program (Minitab Inc. Quality Plaza, 1829 Pine Hall Rd. State College. PA.

16801. United States).The level of each variable was selected

143

Pharmaceutical preparation

Formulation of tablet using extracted pectin from sapodilla peel

Nine formulations for paracetamol and four formulations for Ibuprufen were designed by

the method described by Menon et al (2011) with slight modifications for direct

compression. The composition of 700mg tablet made by wet granulation method is

mentioned in Table 5 for paracetamol tablet and Table 6 for ibuprofen tablets. The

amount of active ingredient (Paracetamol / Ibuprofen), binder, disintegrant and diluent

were mixed after weighing each amount according to the size of batch. All the

ingredients were mixed and passed through sieve # 40 separately. The ingredients were

then mixed with distilled water as granulating agent and a dough is formed. The dough

was then passed through sieve #16 to get coarse granules which were then dried on 450C

for almost 2 hours. The dried granules were then passed through sieve # 20 to obtain

equal size granules. The obtained granules were then mixed with the weighed amount of

glident and lubricant. The mixture was mixed evenly through tumbler movement. The

finally obtained granules were then compressed through single punch machine to make

tablets of required weight and hardness. A pictorial representation of the process is given

in annexure 3. The comparative effect of binding property of pectin was determined,

control tablets were made with the addition of starch instead of pectin as binding agent.

144

Table - 4: Composition of Paracetamol tablet with different concentrations of pectin used

S.NO Ingredients S1

(mg/tab)

F1

(mg/tab)

F2

(mg/tab)

F3

(mg/tab)

F4

(mg/tab)

F5

(mg/tab)

F6

(mg/tab)

F7

(mg/tab)

F8

(mg/tab)

F9

(mg/tab)

1 Paracetamol 500 500 500 500 500 500 500 500 500 500

2 Microcrystalline

cellulose

30 30 30 30 30 30 30 30 30 30

3 Pectin 10 20 30 40 50 60 80 100 120

4 Starch 10

5 Lactose 153 153 143 133 123 113 103 83 63 43

6 Talc 5 5 5 5 5 5 5 5 5 5

7 Magnesium Sterate 2 2 2 2 2 2 2 2 2 2

Where F1= test formulation one, F2= test formulation two, F3=test formulation three, F4 = test formulation four, F5=test formulation

5, F6=test formulation six, F7 test formulation seven, F8= test formulation eight, F9=test formulation nine

145

Table - 5: Composition of Ibuprufen tablet with different concentrations of pectin used

S.NO Ingredients S2

(mg/tab)

R1

(mg/tab)

R2

(mg/tab)

R3

(mg/tab)

R4

(mg/tab)

1 Ibuprofen 500 500 500 500 500

2 Microcrystalline cellulose 30 30 30 30 30

3 Pectin 50 75 100 125

4 Starch 10

5 Lactose 153 113 88 63 38

6 Talc 5 5 5 5 5

7 Magnesium sterate 2 2 2 2 2

Where R1= TrRal formulation one, R2 = test formulation two, R3 =test formulation three , R4 = test formulation four

91

Evaluation of granules

The granules were evaluated for the flowabilty according to the method given in USP 36/

NF 31, 2013 guidelines, these are as under:

Angle of repose (α)

Angle of repose (α) was determined by funnel method.It is defined as angle possible

between the surface of a pile of the powder and the horizontal plane. (Banerjee and

Singh, 2013).The mixture of granules was carefully passed through a glass funnel. The

granules were passed carefully under the force of gravity on a piece of graph paper or

butter paper. The height of heap or cone (h) was calculated while the radius of the heap

was calculated as (r). The angle of repose was calculated as under:

𝛼 = tan−1 ( ℎ

𝑟)

Bulk density (ρb)

Bulk density of the granules was measured by weighing small amount of granules and

transferring it into a measuring cylinder. The volume of granules occupied in the cylinder

was carefully noted and is called bulk volume (Vb). Weight of the granules is denoted by

(M). The bulk density was calculated as under:

ρb = M/ Vb

92

Tapped density (ρt)

The tapped density was measured by tapping weighed amount of granules in a measuring

cylinder for a specific time. The minimum volume (Vt) occupied by the granules and

weight (M) was used to measure tapped density as under:

ρt = M/ Vt

Compressibility Index

It is measured in % compressibility (C). It can be explained as the measurement of the

free flow property through which a materials flow characteristics can be predicted. It was

measured as under:

C = (ρt –ρb) / ρt * 100

Hausner’s ratio

It can aslo be explined as the easiness of flow of granules and is calculated as under

Hausner’s ratio = ρt / ρb

Loss on drying (LOD)

1 to 2 g of test sample was weighed and kept in a stoppered bottle. The stoppered bottle

was previously weighed and dried under the same conditions for the test sample and

cooled in a desiccator. The test specimen was accurately weighed with stopper and was

93

put in the drying chamber without stoppers. Drying was done till a constant weight was

achieved.

% LOD = D-A x 100

C

Where A = Bottle with cover weight

B = Sample + Bottle with Cover (Initial weight)

C = Sample weight

D = Sample + Bottle with Cover (Final weight.)

Tablet compression

The granules obtained were compressed using a single punch machine in which a convex

shaped punch of diameter 12.38 mm was fitted to get oval shaped tablets each weighing

about 700 mg (±5%). The hardness was set between 6-7 kg compressions ranges. The

compression was done at room temperature and a minimum of fifty tablets were

compressed for each batch.

Tablets testing

The finally compressed tablets were evaluated for quality parameters following the

guidelines of USP 36/ NF 31 2013 and some non- pharmacopeial methods which are

described below

94

Weight variation

The variation in weight of the test formulations and reference tablets were studied by

taking weight of each 20 tablets individually on a Type 1 balance. Mean weight and

standard deviation were calculated

Tablet thickness and diameter

The diameter and thickness of 20 tablets, were determined by a Vernier caliper in mm

Tablet hardness

It is calculated by randomly selected 20 tablets of the test formulations using a hardness

tester .

In-Vitro dissolution studies

The dissolution studies of the test and standard tablets performed by following method

given in USP 32/NF 27, 2009 guidelines by using a USP apparatus II. It was carried out

in 900 ml of phosphate buffer pH 5.8 at 37±5 °C at 50 rpm. An aliquot of 10 ml of

solvent was taken out from vessels at 5, 10, 15, 20, 25, 30, 45, 60, 90 and 120 minutes

and volume was made up again by fresh medium. Spectrometer was used to calculate

drug concentration at 278 nm with dissolution medium taken as blank. The dissolution

profile was also established in distilled water, 0.1 N HCl of pH 1.2 and phosphate buffers

at pH 4.5, 6.8 using the same sampling times as described above to evaluate the release of

drug in the new formulations. Each experiment was repeated in triplicate.

95

Formulation of antidiarrheal preparation (suspension) from extracted

sapodilla pectin

In a 3000 ml container 620 ml distill water was taken. Erythrosine, bronopol, propyl

paraben and sodium saccharin were added one by one. In the next step pectin is added,

followed by veegum, sodium carboxymethyl cellulose. The mixture was stirred and

soaked overnight. More distill water was added with glycerin and sodium citrate at this

stage. The mixture was then continuously stirred and kaolin was added gradually. The

mixture was further mixed thoroughly for an hour and the final volume was adjusted

using DI water. Lastly vanilla flavor was added. The formulation ingredients with their

respective amounts are given in Table - 6

96

Evaluation of Suspension

Color, odor and taste

Organoleptic evaluation of suspension was performed for its color, odor and taste

properties

pH

The pH of the suspension was measured by using pH meter.

Viscosity

The viscosity of suspension was evaluated using Oswald viscometer. The suspension is

filled in the required volume very carefully with the help of a pipette into the viscometer.

The viscometer was then kept into a water bath to achieve the desired temperature. When

the temperature reached to a constant temperature the volume in the viscometer was

adjusted again carefully using pipette. The pressure is then released and the time is noted

for the suspension to reach from one point to the other.

Sedimentation Volume

25ml of suspension was taken in a 50ml stoppered graduated measuring cylinder. The

suspension was then moved upside down two three times and then allowed to settle for

three minutes and the volume of sediment was noted which was considered as the

original volume (H0). The cylinder was then kept stationary for 7 days and the volume of

sediment was noted at 7hrs and 24hrs for consecutive 7 days. This was considered as the

final volume (Hu). The sedimentation volume was calculated as

Sedimentation Volume (F) = Hu/ Ho

97

The height of the solid phase after settling relies on the particle size and the concentration

of solid. For a desirable suspension F should be 0.9 for 1 hr

Redispersibility

For the measurement of redispersity a fixed volume of suspension was kept in a

stoppered cylinder at room temperature for 7 days. The suspension in the cylinder was

moved upside down at regular intervals to remove any sediments present at the bottom of

the cylinder.

98

Table - 6: Composition of antidiarrheal preparation nusing extracted sapodilla

pectin.

S.No Ingredient 1litre 1000 ml

1 Deionized water 0.62 lit 620ml

2 Erythrosine 0.0000248kg 24.8mg

3 Bronopol 0.0001kg 100mg

4 Propylparaben (Sodium) 0.00055kg 550mg

5 Sodium saccharin 0.00075kg 750mg

6 Pectin 0.00434kg 4340mg

7 Veegum 0.004kg 4000 mg

8 Sodium carboxy methyl cellulose 0.0016kg 1600mg

9 Water 0.02 litre 20ml

10 Sodium citrate 0.002kg 2000mg

11 Glycerine 0.1lit 100ml

12 Kaolin 0.2kg 200000mg

13 Flavours 0.004lit 4 ml

Make up the volume to 1 L 1000 ml

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FOOD PREPARATIONS

Preparation of Sapodilla jam

Sapodilla jam was prepared from its fruit and pectin extracted from its peel was used as a

gelling agent. 450g of fruit pulp, 550g of sugar, 5g of citric acid and 5g of pectin was

used in the preparation of jam (Ahmed et al., 2011)

The pulp of sapodilla fruit and sugar was mixed first in a stainless steel pan and cooked

on a medium heat on fire. Pectin was added and the mixture was left for heating with

constant stirring till boiling. The mixture was boiled until the required consistency of jam

was achieved which was checked with the help of spoon test. To check the consistency

stainless steel spoons were kept in refrigerator before the test, a spoon when required was

withdrawn from the refrigerator and little jam was taken out from it. The spoon with the

jam was left aside for a min after which the spoon was held in an upright positon. If the

jam spills from the spoon it means the jam required more quickly or else jam was

removed from the fire. After the cooking the jam were transferred into glass jars and

capped. The bottles were then boiled for 10min in an upside down position in a separate

pan containing water; this process is called processing of jam. The bottles were then

cooled and stored at room temperature for further study. Evaluation of the jam cooked

with the three different concentration and standard pectin was performed and the results

were compared. The ingredients and their amounts are shown in Table-7

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Physico-chemical analysis of jam

Determination of pH

The pH of jam was observed by the method described in AOAC

Determination of viscosity

The viscosity of jam was recorded at 25oC by viscometer using t-shaped spindle at 20rpm

Determination of moisture and ash content

The ash and moisture contents were determined following the methods described by

Nwosu et al., (2014).

Determination of total titratable acidity (TTA)

10g of sample was diluted to 100ml with distilled water and titrated against 0.1N NaOH

solution using phenolphthalein as indicator to a pink color endpoint.(AOAC)

Determination of total soluble sugar

The Reducing and non-reducing sucrose was estimated by lane Eynon method. (Islam et

al., 2012)

Determination of total solids

Total solids were determined by subtracting percentage moisture from hundred as

followed by Shahnawaz et al. (2009)

101

% total solids = 100 - % moisture.

Determination of vitamin C (Ascorbic Acid)

Titrimetric estimation of ascorbic acid was performed by method described by Hussain et

al., (2010)

Sensory Evaluation

The sensory evaluation of jam prepared from extracted and standard pectin was

commenced at PCSIR (Pakistan Council for Scientific and Industrial Research, Karachi -

Pakistan). The jam was judged by 20 panelists which were randomly picked from the

institution. The panelist after tasting the jam marked the different parameters of sensory

evaluation. The jam was assessed on a 9-point hedonic scale, where 1 represented

extremely disliked and 9 extremely liked (Stone and Sidel, 2004)

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Table - 7: Formulation of the sapodilla Jam (1KG) with different

concentration of pectin

Ingredients S

(g)

F1

(g)

F2

(g)

F3

(g)

Fruit pulp 450 450 450 450

Sugar 550 550 550 550

Pectin 5 5 7 10

Citric acid 5 5 5 5

WhereS = Jam with standard pectin, F1 = Jam with 5g pectin, F2 = Jam with 7g pectin,

F3 = Jam with 10g pectin

103

Preparation of pudding from extracted sapodilla pectin

Sample puddings with three different concentration of pectin were prepared. Two

pudding samples were also made as a reference from food grade pectin which has the

same concentration as two among the three sample puddings while one of the puddings

has less quantity of pectin than the reference pudding

Samples of milk pudding were prepared by using different concentration of extracted and

a standard concentration of food grade pectin. The pudding was prepared after mixing

required amount of extracted pectin with 100 g of sugar, 1 egg and approximately 990 ml

of milk (Table- 8) in a saucepan. The mixture was stirred to dissolve sugar and pectin

into the mixture, after 10 min of stirring the mixture was heated till boil. After the

required time of boiling the mixture was kept aside from fire and then poured into the

desired containers for cooling. The cooled pudding mixtures were then kept in

refrigerator for 10 hrs till further analysis. Sensory evaluation of puddings were

performed by the 20 panellist specified for the job.

The sensory evaluation of pudding was performed in by 20 panellist .The testing

procedure chosen for the study was Duo-trio test (Kim et al., 2015). Three milk pudding

samples were provided to the panellist among which one was the reference pudding

sample while among the remaining two one of the sample pudding was same in

concentration as reference pudding in terms of pectin. Among the remaining two samples

one of the sample was same as reference sample while the other was different. The

pudding made from standard pectin was used as reference for the first 10 panellist while

pudding made with extracted pectin in the same quantity as reference for the remaining

104

10 panellist. To avoid any biasness the samples were coded. An evaluation form was

given to each participant to rate the sensory attributes. The example of evaluation form is

given in Annexure 4. Same procedure was followed for the remaining pudding samples.

105

Table-8: Showing different pectin concentration used for each selected extracted

fruit pectin

Material P1 P2 P3 Ps1 Ps2

Pectin ( gram) 5 10 15 10 15

Sugar (gram) 100 100 100 100 100

Milk (ml) 990

(approx.)

990

(approx.)

990

(approx.)

990

(approx.)

990

(approx.)

Egg 1 1 1 1 1

P1, P2 and P3 are sapodilla pectin pudding samples while Ps1 and Ps2 and reference samples while approx is approximately

106

RESULTS

4.1 EXTRACTION AND PURIFICATION OF PECTIN FROM THE SELECTED

FRUIT WASTE

Screening results relating to the determination of pectin content of different seasonal

fruits have been presented in Table - 7. In total of nineteen fruits wastes (peels + exocarp)

were evaluated in the present study to determine the pectin content. A simple standard

procedure based on homogenization of peels followed by extraction with DI water at a

pH maintained between 5-6.5, followed by precipitation of pectin with ethanol was used

in the screening study. The whole process has been shown in Figure -20. Out of nineteen,

twelve fruits indicated presence of pectin while insignificant quantity of pectin was noted

in seven fruits. Among twelve the highest content of pectin (20%) was noted in orange

followed by apple 6%, sapodilla (5.5%) and guava (4%). Rest of the fruits indicated

between 2-3.5% of pectin.

After the screening process three fruits (Sapodilla, Banana and Muskmelon) were

selected for further evaluation while two fruits apple and orange were used for comparing

the extraction process and to observe and compare the difference in the yield of pectin.

Table - 8 represents the yield of pectin from five fruits using different method of

extraction. The highest yield obtained from banana, sapodilla and muskmelon were

10.5%, 4.75% and 2.65% respectively when microwave was used as a heating medium

while when Bunsen burner was used the yield came slightly lesser 8.85% for banana, 4%

for sapodilla and 2.45% for muskmelon. Before further analysis and study, the pectin

obtained from fruits (Banana, Sapodilla and Muskmelon) were subjected to the

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preliminary identification test of pectin which revealed that pectin obtained from

muskmelon and banana fails to form stiff gel which is characteristic of pectin. Jelly grade

test was also performed which further proved that the quality of pectin obtained from

banana was of week nature while muskmelon failed to form any jell. The jelly grade of

banana came out between 80 to 85 while sapodilla jelly grade came higher 100-110

which makes it favorable for further analysis and to figure out best extraction method.

Two strengths of inorganic acids (0.1N and 1N) was selected to check the influence of

the concentration of acids. Table – 25 shows the yield of pectin obtained from 0.1N HCl

while Table - 39 shows results acquired from the use of 1NHCl. The time of boiling was

also taken into account along with difference in pH, mechanical method and mode of

boiling. Among the four different time of boiling the best yield was obtained from 0.1N

HCl after 20 min of boiling (6.35% yield at pH5) using microwave as the mode of boiling

and hammering as the mechanical tool. While almost same results (6.7%) were obtained

after only 10 min of boiling when 1NHCl was used for the extraction of pectin at same

pH (pH5) using microwave as the boiling method but the mechanical method chopping

was proved effective.In quest for understanding the integrated effects of different

variables on the yield of pectin extraction with organic acid was also carried out (Table -

54). The organic acids were used in two strengths (1% and 10%). As in our prior studies

we found that pH between 3-5 was effective for extraction so the pH was maintained in a

range of 3 to 5. 1% oxalic acid gave best result (4.95%) using microwave as the heating

method and homogenizing as mechanical tool. While heating through Bunsen burner

gave best average result also with homogenizing succeeding over the other mechanical

methods gave 3.95% yield with 1% oxalic acid.The last study under the section of

108

extraction was done to learn the effect of the different strength of acids (Table – 65). For

this purpose three different strengths (0.1N, 0.5N and 1N) of same inorganic acid (HCl)

was used. The yield of pectin came out to be best when the strength of acid was

increased, 6.7% of pectin yield was obtained when the strength of acid was increased to

1N HCl. The chopping method was useful in all the strengths of acid while heating with

microwave gave almost good yield in every strength.

4.2 PHYSICAL AND BIOCHEMICAL CHARACTERIZATION OF THE

PURIFIED PECTIN

The pectin isolated from the peels of banana, sapodilla and muskmelon was analyzed for

quality. For this purpose preliminary identification tests were carried which are shown in

Table-75. Banana and sapodilla showed positive results while muskmelon failed to show

desired results hence it was omitted for further investigation. The basic characterization

tests was performed on banana and sapodilla peels and shown in Table- 76 and found that

the pectin obtained from sapodilla peel was better in quality than banana. Hence the rest

of the tests were performed on sapodilla peel pectin only. Water holding, water binding

and fat binding properties of pectin was also assessed and shown in Table-77. The

sapodilla pectin was further tested by subjecting it to FTIR spectroscopy (Table-78)

4.3 OPTIMIZATION OF THE YIELD OF PECTIN FROM SAPODILLA PEEL

THOROUGH RESPONSE SURFACE METHODOLOGY

Response surface methodology with Box Bechen design was applied for determination

of optimal condition employed to get maximum yield. The factors studied were time of

boiling, temperature and pH at the time of extraction and the response observed was yield

109

(Table-79). The experimental design was consisted of fifteen experiments (Table-80).

The analysis of variance (ANOVA) was performed which is used to validate the process

variables for the optimization model. It is also used to investigate and understand the

response interactions of the variables, quality of the fit of the polynomial model

(coefficient of determination (R2), adjusted coefficient of determination (adj-R2 ) and

predicted coefficient of determination (pre-R2) and optimization of process condition

were obtained(Table-81).

4.4 UTILIZATION OF THE PURIFIED PECTIN FOR THE DEVELOPMENT OF

PHARMACEUTICAL AND FOOD PRODUCTS

Sapodilla peel pectin, used as a binder in formulation of paracetamol and ibuprofen tablet

was evaluated for pre compression, micrometrics and dissolution properties. Table-84

and 87 represents the micrometrics properties of granules formulated with different

concentrations of binder of paracetamol and ibuprofen tablets respectively. The weight of

both the types (paracetamol and ibuprofen) remained under the limit of ± 5%. The

diameter and thickness of all the tablets also didn’t exceeded from the required level

(Table- 85and 88).

In paracetamol tablets,10 and 20 mg of pectin were found not suitable to achieve desired

hardness granules when compressed into tablets were soft and thus further concentration

of pectin was increased. When the concentration increased from 30mg/tab desired

hardness was achieved( Table-86) however interestingly as the concentration of pectin

was increased,dissolution was noted to be decreased. The best hardness and dissolution

was achieved with the formulation F4 and F5 when the concentration of 40mg /tab and

110

50 mg/tab were used respectively. Dissolution was recorded as 80.43% and 86.35%

respectively (Table -87). Further increased in pectin from 60 to 120mg not only increased

the hardness of tablet but also noted to decrease the dissolution significantly. While the

ibuprofen tablets showed a similar type of results it is also noted that the higher

concentrations of pectin had detrimental effect on dissolution properties of tablets.

Antidiarrheal preparation formulated from extracted pectin also showed similar results as

compared to the marketed product (Table 93).

The jam was prepared using different concentrations of pectin are shown in Fig.1. The

jam due to different concentrations of pectin appeared to possess different types of

physical and chemical attributes Table- 90. The pH of the jam showed to be in range

from 3.00 to 3.30 .The TTA came out to be in between 1.00 to 1.14. All the formulations

except F3 showed the value of TSS nearer to the desirable level. The Vit C content of jam

came in between 17.6 to 18.3. The moisture found in these formulations was same as that

of previously studied jam. The three formulated jams with different concentration of

pectin were subjected to hedonic testing the results of the sensory evaluation of the three

formulations compared with standards are presented in Table-91

The result of the sensory evaluation of pudding made from extracted pectin through duo

trio test indicated among the 20, 9 correctly identified the difference in sample while 10

failed to identify any difference. To constitute significant differentiation among the

samples 15 correct judgement among 20 were required. The result showed that the

panelist as a whole did not found any significant difference (P> 0.05) between the

pudding made from standard and extracted pectin.

111

Table - 9: List of fruits used in the screening study for the presence of pectin.

(a) is Munarin et al., 2012; (b) is Bhat and Singh, 2014; (c) is Baker, 1997; (d) is Baissise

et al., 2010. (e) is Rasheed, 2008; (f) is Aina et al., 2012.

S no

Fruit

%Yield Reported

amount Common name Parts

used

1 Orange peel 20 6-26% a

2 Apple exocarp 6 2-19%a

3 Grape Fruit peel 3.5 13-32%a

4 Sapodilla peel 5.5 Notfound

5 Muskmelon peel 2 Not found

6 Guava (unripe) exocarp 0 Not found

7 Guava(ripe) exocarp 4 16.8% b

8 Chinese apple (Jungle Fruit) exocarp 0 Notfound

9 Strawberry exocarp 0 0.35–0.44%.C

10 Sweet lime peel 3 6-26%a

11 Peach exocarp 0 4-18%a

12 Apricot exocarp 0 4.97%d

13 Canteloup peel 3 Not found

14 Plum exocarp 0 Not found

15 Watermelon peel 2.35 15.70% e

16 Mango(Unripe) peel 0 Not found

17 Mango (ripe) peel 3.5 9-29%a

18 Banana peel 3.5 2-15%a

19 Lemon peel 3.5 2.7% f

112

Table - 10: Showing percent yield of pectin from five selected fruits

Mechanical

procedure

p

H

Sapodilla Banana Muskmelo

n Apple Orange

B M B M B M B M B M

Homogenizin

g

1 0.5 1.0

5

3.2

5 5.6 0.55 0.55 0.5 1

16.1

5

15.2

5

3 0.7

5 1.6

6.8

5

4.3

5 2 1

1.9

5

2.7

5 18.2 19.8

5 1.2 1.5 8.8

5 8.4 1.45 2.3

2.3

5 3.7 19.4 22.7

6 1.5 2 5.8

5

9.7

5 1.15 1 1.5 1.4

13.9

5 2.85

7 0.5 0.5 5.4

5 3.1 0.55 1.8

0.0

5

0.3

5 21.7 7.5

Grinding

1 0.5

5 0.8

2.4

5 3.2 2.25 2.65

2.4

5 2.5 2.6 3.95

3 4 3.5 4.2 6.9 1.4 1.8 0.4

5

1.1

5 7.8 5.3

5 2.4

5 2.9 3.5 4.5 1.95 2.4 1 1.1 2.3 7.1

6 2 2 3.6

5

2.6

5 0.95 1.6 2.4

2.8

5 6.45 6.6

7 1.9

5 2.3

1.2

5

0.2

5 0.55 1

0.0

5

0.5

5 0.6 0.2

Cutting

1 0.8 0.9 0 2.5 0.5 0.5 2.8

5 3.9 8.1 7

3 1.1 1.5 1.9

5 1.2 2.45 2.55

2.9

5 4.5 8.5 12

5 0.4 1 5 5.5 1 1 2.7 3.7 11.8

5

20.6

5

6 1 1.2 1 5.4 0.7 0.95 1.1

5 2.9 6.95 10.2

7 0.7 1 3 6.5 1.3 2.2 0.0

5 0.7 4 7.4

Chopping

1 2.7 4.7 1.6

5 2.1 0.05 0.7

2.8

5

3.3

5 2.6 3.35

3 3.9 3.4

5 4 3.3 0.05 0.5

1.2

5

2.7

5 7.95

10.0

5

5 3.9

5

4.0

5

5.9

5 6.6 1.7 2.1

0.0

5

1.7

5 10.4 15.8

6 2.3 2.4

5

5.8

5

6.3

5 2.25 1.75 0.1 0.9

15.2

5

17.1

5

7 3.9 2.4 1.8 1.6 0.5 1.05 0.2 0.6 5.35 9.35

113

5 5 5

Hammering

1 1.5 2.0

5 4.5 7 0.65 1.25 1.5 1.7

13.5

5

16.6

5

3 0.6 1.5

5 5.5 5.5 0.5 0.8 2.7 3.5 7.2 8.35

5 0.8 1.9

5 8.5

10.

5 0.65 0.3

3.0

5

4.8

5 7.4

12.1

5

6 0.8 1.4 5 5.6 0.95 0.5 1.2

5

2.0

5 5.55 10.7

7 0.5 0 0.5 0.5 0.45 0.05 1.7 2.0

5 5.5 8.85

B= Boiling on burner, M= heating in microwave

114

Table - 11: Analysis of variance (mean squares) of yield for five selected fruits

Source of variation

Degrees

of

freedom

Mean squares

Sapodilla Banana Muskmelon Apple Orange

Mech. procedure (MP)

pH level

Boiling method (BM)

MP pH

MP BM

pH BM

MP pH x BM

Error

Total

4

4

1

16

4

4

16

100

149

32.6575**

3.3917**

3.2413**

2.5462**

0.3392**

1.0112**

0.6258**

0.0266

51.245**

84.818**

22.349**

16.774**

4.640**

6.332**

4.698**

0.191

4.3869**

1.5751**

2.0184**

2.7688**

0.2814**

0.4112**

0.4141**

0.0112

9.5920**

17.3200**

22.6981**

5.6967**

0.8120**

0.8975**

0.2720**

0.0314

493.729**

142.586**

59.914**

88.941**

81.797**

44.377**

21.750**

1.0100

NS = Non-significant (P>0.05); * = Significant (P<0.05); ** = Highly significant (P<0.01) , Mech = mechanical

115

Table – 12: Means comparison of yield for sapodilla fruit peel (Mechanical procedure pH interaction mean±SE )

pH level Mechanical procedure Mean

Homogenizing Grinding Cutting Chopping Hammering

1 0.78 ± 0.13ij 0.68 ± 0.06j 0.85 ± 0.03hij 3.70 ± 0.47a 1.78 ± 0.14ef 1.56 ± 0.23C

3 1.18 ± 0.19gh 3.75 ± 0.15a 1.30 ± 0.09g 3.68 ± 0.13a 1.08 ± 0.21ghi 2.20 ± 0.24A

5 1.35 ± 0.08g 2.68 ± 0.11c 0.70 ± 0.14j 4.00 ± 0.12a 1.38 ± 0.26g 2.02 ± 0.23B

6 1.75 ± 0.12f 2.00 ± 0.06ef 1.10 ± 0.05ghi 2.38 ± 0.10cd 1.10 ± 0.14ghi 1.67 ± 0.10C

7 0.50 ± 0.01jk 2.13 ± 0.10de 0.85 ± 0.07hij 3.18 ± 0.35b 0.25 ± 0.11k 1.38 ± 0.22D

Mean 1.11 ± 0.10C 2.25 ± 0.19B 0.96 ± 0.05D 3.39 ± 0.16A 1.12 ± 0.12C

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

Table - 13: Means comparison of yield for sapodilla fruit peel (Mechanical procedure boiling method interaction mean±SE)

Boiling Mechanical procedure Mean

Method Homogenizing Grinding Cutting Chopping Hammering

B 0.89 ± 0.11e 2.19 ± 0.30b 0.80 ± 0.07e 3.36 ± 0.20a 0.84 ± 0.09e 1.62 ± 0.14B

M 1.33 ± 0.14c 2.30 ± 0.25b 1.12 ± 0.06d 3.41 ± 0.25a 1.39 ± 0.20c 1.91 ± 0.13A

B= Boiling on burner , M= heating in microwave

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean.

Table - 14: Means comparison of yield for sapodilla fruit peel (Mechanical procedure pH boiling method

interaction mean ± SE)

BM pH Mechanical procedure BM pH

116

Homogenizing Grinding Cutting Chopping Hammering interaction mean

B 1 0.50 ± 0.03qrs 0.55 ± 0.02pqr 0.80 ± 0.04n-r 2.70 ± 0.14ef 1.50 ± 0.06i-l 1.21 ± 0.22E

B 3 0.75 ± 0.03n-r 4.00 ± 0.15bc 1.10 ± 0.03j-o 3.90 ± 0.14bcd 0.60 ± 0.03o-r 2.07 ± 0.41B

B 5 1.20 ± 0.07j-n 2.45 ± 0.10efg 0.40 ± 0.02rs 3.95 ± 0.22bcd 0.80 ± 0.03n-r 1.76 ± 0.35C

B 6 1.50 ± 0.07i-l 2.00 ± 0.10ghi 1.00 ± 0.04l-q 2.30 ± 0.13fg 0.80 ± 0.03n-r 1.52 ± 0.16D

B 7 0.50 ± 0.02qrs 1.95 ± 0.08ghi 0.70 ± 0.03n-r 3.95 ± 0.14bcd 0.50 ± 0.01qrs 1.52 ± 0.36D

M 1 1.05 ± 0.06k-p 0.80 ± 0.03n-r 0.90 ± 0.03m-r 4.70 ± 0.26a 2.05 ± 0.14gh 1.90 ± 0.40BC

M 3 1.60 ± 0.08hij 3.50 ± 0.16cd 1.50 ± 0.06i-l 3.45 ± 0.14d 1.55 ± 0.05h-k 2.32 ± 0.26A

M 5 1.50 ± 0.04i-l 2.90 ± 0.06e 1.00 ± 0.03l-q 4.05 ± 0.14b 1.95 ± 0.11ghi 2.28 ± 0.29A

M 6 2.00 ± 0.09ghi 2.00 ± 0.08ghi 1.20 ± 0.04j-n 2.45 ± 0.16efg 1.40 ± 0.02j-m 1.81 ± 0.13C

M 7 0.50 ± 0.02qrs 2.30 ± 0.12fg 1.00 ± 0.04l-q 2.40 ± 0.08efg 0.00 ± 0.00s 1.24 ± 0.26E

B= Boiling on burner , M= heating in microwave; BM= boiling method

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

.

117

Table – 15: Means comparison of yield for banana fruit peel (Mechanical procedure pH interaction mean±SE)

pH level Mechanical procedure Mean

Homogenizing Grinding Cutting Chopping Hammering

1 4.43 ± 0.55fgh 2.83 ± 0.19j 1.25 ± 0.56klm 1.88 ± 0.12k 5.75 ± 0.60cd 3.23 ± 0.36D

3 5.60 ± 0.57cde 5.55 ± 0.64cde 1.58 ± 0.17kl 3.65 ± 0.19hij 5.50 ± 0.18cde 4.38 ± 0.34C

5 8.63 ± 0.31ab 4.00 ± 0.25ghi 5.25 ± 0.21def 6.28 ± 0.27c 9.50 ± 0.53a 6.73 ± 0.41A

6 7.80 ± 0.90b 3.15 ± 0.26ij 3.20 ± 0.99ij 6.10 ± 0.22cd 5.30 ± 0.19def 5.11 ± 0.42B

7 4.28 ± 0.54gh 0.75 ± 0.23lm 4.75 ± 0.80efg 1.73 ± 0.07k 0.50 ± 0.01m 2.40 ± 0.38E

Mean 6.15 ± 0.41A 3.26 ± 0.33D 3.21 ± 0.40D 3.93 ± 0.37C 5.31 ± 0.55B

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

Table - 16: Means comparison of yield for banana fruit peel (Mechanical procedure boiling method interaction mean±SE)

Boiling Mechanical procedure Mean

Method Homogenizing Grinding Cutting Chopping Hammering

B 6.05 ± 0.50a 3.01 ± 0.29e 2.19 ± 0.46f 3.86 ± 0.50cd 4.80 ± 0.69b 3.98 ± 0.27B

M 6.24 ± 0.67a 3.50 ± 0.59de 4.22 ± 0.55c 3.99 ± 0.57cd 5.82 ± 0.87a 4.75 ± 0.31A

B= Boiling on burner , M= heating in microwave In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

118

Table - 17: Means comparison of yield for banana fruit peel (Mechanical procedure pH boiling method interaction

mean ± SE)

BM pH Mechanical procedure BM pH

Homogenizing Grinding Cutting Chopping Hammering interaction mean

B 1 3.25 ± 0.19l-q 2.45 ± 0.11o-u 0.00 ± 0.00x 1.65 ± 0.12r-w 4.50 ± 0.16h-l 2.37 ± 0.41D

B 3 6.85 ± 0.24def 4.20 ± 0.19i-m 1.95 ± 0.06q-v 4.00 ± 0.19j-n 5.50 ± 0.27e-i 4.50 ± 0.44C

B 5 8.85 ± 0.51b 3.50 ± 0.09l-p 5.00 ± 0.31g-k 5.95 ± 0.22d-h 8.50 ± 0.29b 6.36 ± 0.56B

B 6 5.85 ± 0.26d-h 3.65 ± 0.24k-o 1.00 ± 0.03u-x 5.85 ± 0.31d-h 5.00 ± 0.21g-k 4.27 ± 0.49C

B 7 5.45 ± 0.30e-j 1.25 ± 0.07t-x 3.00 ± 0.12m-s 1.85 ± 0.08q-v 0.50 ± 0.03vwx 2.41 ± 0.46D

M 1 5.60 ± 0.30d-i 3.20 ± 0.16l-q 2.50 ± 0.14o-t 2.10 ± 0.09p-u 7.00 ± 0.43cd 4.08 ± 0.52C

M 3 4.35 ± 0.12i-m 6.90 ± 0.40de 1.20 ± 0.07t-x 3.30 ± 0.15l-q 5.50 ± 0.31e-i 4.25 ± 0.53C

M 5 8.40 ± 0.43bc 4.50 ± 0.23h-l 5.50 ± 0.26e-i 6.60 ± 0.46def 10.50 ± 0.57a 7.10 ± 0.59A

M 6 9.75 ± 0.46ab 2.65 ± 0.14n-t 5.40 ± 0.27f-j 6.35 ± 0.29d-g 5.60 ± 0.21d-i 5.95 ± 0.62B

M 7 3.10 ± 0.09l-r 0.25 ± 0.01wx 6.50 ± 0.32def 1.60 ± 0.07s-w 0.50 ± 0.02vwx 2.39 ± 0.61D

B= Boiling on burner , M= heating in microwave; BM= boiling method

. In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table -18: Means comparison of yield for muskmelon fruit peel (Mechanical procedure pH interaction mean±SE)

pH level Mechanical procedure Mean

119

Homogenizing Grinding Cutting Chopping Hammering

1 0.55 ± 0.02m-p 2.45 ± 0.12a 0.50 ± 0.01n-q 0.38 ± 0.15pqr 0.95 ± 0.14ijk 0.97 ± 0.15D

3 1.50 ± 0.23fg 1.60 ± 0.10ef 2.50 ± 0.05a 0.28 ± 0.10qr 0.65 ± 0.07l-o 1.31 ± 0.15B

5 1.88 ± 0.20cd 2.18 ± 0.13b 1.00 ± 0.02ij 1.90 ± 0.10cd 0.48 ± 0.08o-r 1.49 ± 0.13A

6 1.08 ± 0.05hi 1.28 ± 0.15gh 0.83 ± 0.06jkl 2.00 ± 0.12bc 0.73 ± 0.10k-n 1.18 ± 0.09C

7 1.18 ± 0.28hi 0.78 ± 0.10j-m 1.75 ± 0.21de 0.78 ± 0.13j-m 0.25 ± 0.09r 0.95 ± 0.12D

Mean 1.24 ± 0.11C 1.66 ± 0.12A 1.32 ± 0.14B 1.07 ± 0.15D 0.61 ± 0.06E

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

Table - 19: Means comparison of yield for muskmelon fruit peel (Mechanical procedure boiling method interaction

mean±SE)

Boiling Mechanical procedure Mean

Method Homogenizing Grinding Cutting Chopping Hammering

B 1.14 ± 0.15d 1.42 ± 0.17b 1.19 ± 0.18d 0.91 ± 0.24e 0.64 ± 0.05f 1.06 ± 0.08B

M 1.33 ± 0.17bc 1.89 ± 0.16a 1.44 ± 0.21b 1.22 ± 0.16cd 0.58 ± 0.11f 1.29 ± 0.09A

B= Boiling on burner , M= heating in microwave;

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table -20: Means comparison of yield for muskmelon fruit peel (Mechanical procedure pH boiling method interaction

mean±SE)

BM pH Mechanical procedure BM pH

Homogenizing Grinding Cutting Chopping Hammering interaction mean

B 1 0.55 ± 0.03qr 2.25 ± 0.13bcd 0.50 ± 0.03qr 0.05 ± 0.00s 0.65 ± 0.03pqr 0.80 ± 0.20F

120

B 3 2.00 ± 0.04def 1.40 ± 0.05i-l 2.45 ± 0.06abc 0.05 ± 0.00s 0.50 ± 0.02qr 1.28 ± 0.24BCD

B 5 1.45 ± 0.05h-k 1.95 ± 0.10d-g 1.00 ± 0.04m-p 1.70 ± 0.05f-i 0.65 ± 0.03pqr 1.35 ± 0.13B

B 6 1.15 ± 0.06k-n 0.95 ± 0.03m-p 0.70 ± 0.05opq 2.25 ± 0.10bcd 0.95 ± 0.04m-p 1.20 ± 0.15DE

B 7 0.55 ± 0.02qr 0.55 ± 0.01qr 1.30 ± 0.07j-m 0.50 ± 0.03qr 0.45 ± 0.02qr 0.67 ± 0.09G

M 1 0.55 ± 0.03qr 2.65 ± 0.12a 0.50 ± 0.02qr 0.70 ± 0.04opq 1.25 ± 0.05j-m 1.13 ± 0.22E

M 3 1.00 ± 0.03m-p 1.80 ± 0.09e-h 2.55 ± 0.09ab 0.50 ± 0.02qr 0.80 ± 0.06n-q 1.33 ± 0.20BC

M 5 2.30 ± 0.09a-d 2.40 ± 0.14abc 1.00 ± 0.03m-p 2.10 ± 0.08cde 0.30 ± 0.01rs 1.62 ± 0.22A

M 6 1.00 ± 0.05m-p 1.60 ± 0.08g-j 0.95 ± 0.05m-p 1.75 ± 0.04e-i 0.50 ± 0.02qr 1.16 ± 0.12DE

M 7 1.80 ± 0.11e-h 1.00 ± 0.04m-p 2.20 ± 0.09bcd 1.05 ± 0.07l-o 0.05 ± 0.00s 1.22 ± 0.20CDE

B= Boiling on burner , M= heating in microwave; BM= boiling method

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

121

Table -21: Means comparison of yield for apple fruit peel (Mechanical procedure pH interaction mean±SE)

pH level Mechanical procedure Mean

Homogenizing Grinding Cutting Chopping Hammering

1 0.75 ± 0.12ijk 2.48 ± 0.06d 3.38 ± 0.26bc 3.10 ± 0.15c 1.60 ± 0.05gh 2.26 ± 0.19B

3 2.35 ± 0.19de 0.80 ± 0.16ij 3.73 ± 0.36ab 2.00 ± 0.34ef 3.10 ± 0.20c 2.40 ± 0.22A

5 3.03 ± 0.31c 1.05 ± 0.04i 3.20 ± 0.24c 0.90 ± 0.38i 3.95 ± 0.42a 2.43 ± 0.26A

6 1.45 ± 0.05h 2.63 ± 0.12d 2.03 ± 0.39ef 0.50 ± 0.18jkl 1.65 ± 0.18fgh 1.65 ± 0.16C

7 0.20 ± 0.07l 0.30 ± 0.11l 0.38 ± 0.15kl 0.43 ± 0.08jkl 1.88 ± 0.10fg 0.64 ± 0.12D

Mean 1.56 ± 0.20B 1.45 ± 0.18BC 2.54 ± 0.26A 1.39 ± 0.22C 2.44 ± 0.20A

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

Table - 22: Means comparison of yield for apple fruit peel (Mechanical procedure boiling method interaction mean±SE)

Boiling Mechanical procedure Mean

Method Homogenizing Grinding Cutting Chopping Hammering

B 1.27 ± 0.23e 1.27 ± 0.27e 1.94 ± 0.31c 0.90 ± 0.29f 2.04 ± 0.19c 1.48 ± 0.12B

M 1.84 ± 0.33c 1.63 ± 0.24d 3.14 ± 0.36a 1.87 ± 0.28c 2.83 ± 0.32b 2.26 ± 0.15A

B= Boiling on burner , M= heating in microwave; BM= boiling method

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

Table - 23: Means comparison of yield for apple fruit peel (Mechanical procedure pH boiling method interaction

mean±SE)

BM pH Mechanical procedure BM pH

Homogenizing Grinding Cutting Chopping Hammering interaction mean

122

B 1 0.50 ± 0.02r-u 2.45 ± 0.09fgh 2.85 ± 0.16d-g 2.85 ± 0.13d-g 1.50 ± 0.05i-m 2.03 ± 0.25C

B 3 1.95 ± 0.06hij 0.45 ± 0.02r-u 2.95 ± 0.14c-f 1.25 ± 0.08k-o 2.70 ± 0.15efg 1.86 ± 0.25C

B 5 2.35 ± 0.10gh 1.00 ± 0.05m-r 2.70 ± 0.05efg 0.05 ± 0.00u 3.05 ± 0.15cde 1.83 ± 0.30C

B 6 1.50 ± 0.08i-m 2.40 ± 0.07fgh 1.15 ± 0.06l-p 0.10 ± 0.01u 1.25 ± 0.09k-o 1.28 ± 0.20D

B 7 0.05 ± 0.00u 0.05 ± 0.00u 0.05 ± 0.00u 0.25 ± 0.00tu 1.70 ± 0.09i-l 0.42 ± 0.17F

M 1 1.00 ± 0.06m-r 2.50 ± 0.10e-h 3.90 ± 0.19b 3.35 ± 0.20bcd 1.70 ± 0.04i-l 2.49 ± 0.29B

M 3 2.75 ± 0.16efg 1.15 ± 0.04l-p 4.50 ± 0.21a 2.75 ± 0.14efg 3.50 ± 0.10bc 2.93 ± 0.30A

M 5 3.70 ± 0.15b 1.10 ± 0.04m-q 3.70 ± 0.18b 1.75 ± 0.09ijk 4.85 ± 0.23a 3.02 ± 0.37A

M 6 1.40 ± 0.07j-n 2.85 ± 0.11d-g 2.90 ± 0.06d-g 0.90 ± 0.04n-s 2.05 ± 0.05hi 2.02 ± 0.21C

M 7 0.35 ± 0.02stu 0.55 ± 0.02q-u 0.70 ± 0.04o-t 0.60 ± 0.02p-u 2.05 ± 0.12hi 0.85 ± 0.16E

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

123

Table -24: Means comparison of yield for orange fruit peel (Mechanical procedure pH interaction mean±SE)

pH level Mechanical procedure Mean

Homogenizing Grinding Cutting Chopping Hammering

1 15.70 ± 0.45b 3.28 ± 0.32j 7.55 ± 0.30fgh 2.98 ± 0.19j 15.10 ± 0.88bc 8.92 ± 1.05C

3 19.00 ± 0.91a 6.55 ± 0.58ghi 10.25 ± 0.81d 9.00 ± 0.54def 7.78 ± 0.33e-h 10.52 ± 0.87B

5 21.05 ± 1.02a 4.70 ± 1.09ij 16.25 ± 2.07b 13.10 ± 1.27c 9.78 ± 1.11de 12.98 ± 1.18A

6 8.40 ± 2.51d-g 6.53 ± 0.21ghi 8.58 ± 0.81d-g 16.20 ± 0.61b 8.13 ± 1.17d-g 9.57 ± 0.84C

7 14.60 ± 3.20bc 0.40 ± 0.09k 5.70 ± 0.78hi 7.35 ± 0.92fgh 7.18 ± 0.78fgh 7.05 ± 1.07D

Mean 15.75 ± 1.14A 4.29 ± 0.49C 9.67 ± 0.82B 9.73 ± 0.91B 9.59 ± 0.65B

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

Table -25: Means comparison of yield for orange fruit peel (Mechanical procedure boiling method interaction mean±SE)

Boiling Mechanical procedure Mean

Method Homogenizing Grinding Cutting Chopping Hammering

B 17.88 ± 0.78a 3.95 ± 0.73e 7.88 ± 0.69d 8.31 ± 1.17d 7.84 ± 0.80d 9.17 ± 0.65B

M 13.62 ± 2.02b 4.63 ± 0.67e 11.45 ± 1.35c 11.14 ± 1.34c 11.34 ± 0.83c 10.44 ± 0.68A

B= Boiling on burner , M= heating in microwave; BM= boiling method

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

124

Table -26: Means comparison of yield for orange fruit peel (Mechanical procedure pH boiling method interaction

mean±SE)

BM pH Mechanical procedure BM pH

Homogenizing Grinding Cutting Chopping Hammering interaction mean

B 1 16.15 ± 0.44d-g 2.60 ± 0.14uvw 8.10 ± 0.33m-r 2.60 ± 0.11uvw 13.55 ± 0.48g-j 8.60 ± 1.49DE

B 3 18.20 ± 1.36b-e 7.80 ± 0.21m-r 8.50 ± 0.20l-r 7.95 ± 0.33m-r 7.20 ± 0.41n-s 9.93 ± 1.14CD

B 5 19.40 ± 0.80a-d 2.30 ± 0.10uvw 11.85 ± 0.64i-l 10.40 ± 0.66j-n 7.40 ± 0.20m-r 10.27 ± 1.52BC

B 6 13.95 ± 0.76f-i 6.45 ± 0.40p-t 6.95 ± 0.41o-s 15.25 ± 0.72e-h 5.55 ± 0.24q-u 9.63 ± 1.12B

B 7 21.70 ± 0.72a 0.60 ± 0.03vw 4.00 ± 0.21stu 5.35 ± 0.20r-u 5.50 ± 0.18q-u 7.43 ± 1.97BC

M 1 15.25 ± 0.80e-h 3.95 ± 0.16s-v 7.00 ± 0.17o-s 3.35 ± 0.14t-w 16.65 ± 1.12c-g 9.24 ± 1.52A

M 3 19.80 ± 1.29abc 5.30 ± 0.28r-u 12.00 ± 0.46h-k 10.05 ± 0.51k-o 8.35 ± 0.22m-r 11.10 ± 1.33CD

M 5 22.70 ± 1.36a 7.10 ± 0.39n-s 20.65 ± 1.25ab 15.80 ± 0.54efg 12.15 ± 0.66h-k 15.68 ± 1.55CD

M 6 2.85 ± 0.19uvw 6.60 ± 0.22p-t 10.20 ± 0.66j-o 17.15 ± 0.67c-f 10.70 ± 0.46i-m 9.50 ± 1.28EF

M 7 7.50 ± 0.35m-r 0.20 ± 0.01w 7.40 ± 0.35m-r 9.35 ± 0.43k-p 8.85 ± 0.48k-q 6.66 ± 0.90F

Means sharing similar letter in a row or in a column are statistically non-significant (P>0.05). Small letters represent comparison among interaction means and

capital letters are used for overall mean

125

Table - 27: Percentage yield of pectin from sapodilla fruit peel after using different

physicomechanical procedures (pH, mechanical procedure, boiling method and time of

boiling) with 0.1N HCl

Mechanical procedure

pH %yield 10 Min %yield 20min %yield 40 min %yield 60min

B M B M B M B M

Ham

mer

ing

1 1.5 2.05 0.5 1.1 0.05 0.05 1.2 1.8

3 0.6 1.55 2.2 4.85 0.05 0.1 4.1 3.15

5 0.8 1.95 5.5 6.35 0.05 0.05 4.5 4.6

6 0.8 1.4 3.5 5.2 1.65 1.65 3 3.45

7 0.5 0 2.6 4.35 0.05 0.95 1.75 3.85

Gri

nd

ing

1 0.55 0.8 0.75 1.6 0.1 1.15 0.25 0.35

3 4 3.5 3.55 4 1 1.35 0.05 0.05

5 2.45 2.9 2.05 3.8 2.4 2.75 0.05 2.1

6 2 2 3.4 2.7 2.1 3.1 0.05 0.05

7 1.95 2.3 2.85 3.3 0.95 1.75 0.05 2.3

Cu

ttin

g

1 0.8 0.9 1.45 1.5 0.05 0.05 0.05 0.1

3 1.1 1.5 1.95 3.15 0.45 0.05 0.4 1.4

5 0.4 1 1.5 2.2 1.2 3.95 0.05 0.5

6 1 1.2 1.45 1.85 0.65 1 0.1 0.9

7 0.7 1 1.35 1.8 0.1 1.5 0.05 1.05

Ho

mo

gen

izin

g

1 0.5 1.05 0.05 0.15 0.05 0.05 0.05 0.05

3 0.75 1.6 0.05 1 2.5 1.3 2.75 2.8

5 1.2 1.5 0.05 0.1 3 3.2 2.7 6

6 1.5 2 0.05 0.05 2.25 2.45 2.7 4.5

7 0.5 0.5 0.1 0.05 2.45 2.8 0.15 2.65

Ch

op

pin

g

(mort

ar/

pes

tle

1 2.7 4.7 0.05 0.05 0.05 0.05 1.85 2.75

3 3.9 3.45 4.75 6.5 1.6 1.35 1.25 3

5 3.95 4.05 2.25 3.05 2.05 2.35 1.95 5.6

6 2.3 2.45 2.5 2.8 2.7 3.6 3.25 5

7 3.95 2.4 0.1 0.7 1 2 3.6 3.65

B= Boiling on burner , M= heating in microwave

126

Table -28: Analysis of variance (mean squares) of yield of pectin from sapodilla fruit peel after using different

physicomechanical procedure different fruits with 0.1N HCl

Source of

variation

Degrees of

freedom

Mean squares

Yield 10 min Yield 20 min Yield 40 min Yield 60 min

MP

pH

BM

MP x pH

MP x BM

pH x BM

MP x pH x BM

Error

Total

4

4

1

16

4

4

16

100

149

32.6575**

3.3917**

3.2414**

2.5462**

0.3392**

1.0112**

0.6258**

0.0081

49.7412**

27.4284**

18.6772**

7.5601**

1.7538**

1.4724**

0.4729**

0.0129

12.1446**

20.1921**

6.1206**

2.3355**

1.0438**

1.6176**

0.5655**

0.0070

55.6906**

15.5938**

39.7837**

4.3065**

2.0295**

3.7639**

1.5634**

0.0122

NS = Non-significant (P>0.05); * = Significant (P<0.05); ** = Highly significant (P<0.01). Mech = Mechanical

Table – 29: Means comparison of yield for sapodilla fruit peel after 10 min of boiling using 0.1N HCl (Mechanical procedure

pH interaction mean±SE)

127

pH level Mechanical procedure Mean

Homogenizing Grinding Cutting Chopping Hammering

1 1.78 ± 0.13g 0.68 ± 0.06kl 0.85 ± 0.02k 3.70 ± 0.45b 0.78 ± 0.12k 1.56 ± 0.23D

3 1.08 ± 0.21j 3.75 ± 0.14b 1.30 ± 0.09hi 3.68 ± 0.11b 1.18 ± 0.19ij 2.20 ± 0.24A

5 1.38 ± 0.26h 2.68 ± 0.10d 0.70 ± 0.13k 4.00 ± 0.07a 1.35 ± 0.07hi 2.02 ± 0.23B

6 1.10 ± 0.13j 2.00 ± 0.04f 1.10 ± 0.05j 2.38 ± 0.04e 1.75 ± 0.12g 1.67 ± 0.10C

7 0.25 ± 0.11m 2.13 ± 0.08f 0.85 ± 0.07k 3.18 ± 0.35c 0.50 ± 0.01l 1.38 ± 0.22E

Mean 1.12 ± 0.12C 2.25 ± 0.19B 0.96 ± 0.05D 3.39 ± 0.15A 1.11 ± 0.09C

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table - 30: Means comparison of yield for sapodilla fruit peel after 10 min of boiling using 0.1N HCl (Mechanical procedure

boiling method interaction mean±SE)

Boiling Mechanical procedure Mean

Method Homogenizing Grinding Cutting Chopping Hammering

B 0.84 ± 0.09f 2.19 ± 0.30c 0.80 ± 0.07f 3.36 ± 0.19a 0.89 ± 0.11f 1.62 ± 0.14B

M 1.39 ± 0.20d 2.30 ± 0.24b 1.12 ± 0.06e 3.41 ± 0.24a 1.33 ± 0.14d 1.91 ± 0.13A B= Boiling on burner , M= heating in microwave; BM= boiling method In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table -31: Means comparison of yield for sapodilla fruit peel after 10 min of boiling using 0.1N HCl (Mechanical procedure

pH boiling method interaction mean±SE)

BM pH Mechanical procedure BM pH

Homogenizing Grinding Cutting Chopping Hammering interaction mean

B 1 1.50 ± 0.02ij 0.55 ± 0.01qr 0.80 ± 0.02m-q 2.70 ± 0.05de 0.50 ± 0.01qr 1.21 ± 0.22F

B 3 0.60 ± 0.02pqr 4.00 ± 0.10b 1.10 ± 0.02klm 3.90 ± 0.09b 0.75 ± 0.02n-q 2.07 ± 0.41B

B 5 0.80 ± 0.01m-q 2.45 ± 0.03ef 0.40 ± 0.01r 3.95 ± 0.12b 1.20 ± 0.03jkl 1.76 ± 0.35D

128

B 6 0.80 ± 0.01m-q 2.00 ± 0.05gh 1.00 ± 0.02l-o 2.30 ± 0.05fg 1.50 ± 0.04ij 1.52 ± 0.15E

B 7 0.50 ± 0.02qr 1.95 ± 0.06h 0.70 ± 0.02o-r 3.95 ± 0.11b 0.50 ± 0.01qr 1.52 ± 0.36E

M 1 2.05 ± 0.05gh 0.80 ± 0.02m-q 0.90 ± 0.02l-p 4.70 ± 0.10a 1.05 ± 0.01lmn 1.90 ± 0.39C

M 3 1.55 ± 0.05i 3.50 ± 0.14c 1.50 ± 0.03ij 3.45 ± 0.02c 1.60 ± 0.04i 2.32 ± 0.25A

M 5 1.95 ± 0.05h 2.90 ± 0.05d 1.00 ± 0.01l-o 4.05 ± 0.10b 1.50 ± 0.05ij 2.28 ± 0.29A

M 6 1.40 ± 0.03ijk 2.00 ± 0.08gh 1.20 ± 0.03jkl 2.45 ± 0.04ef 2.00 ± 0.05gh 1.81 ± 0.12CD

M 7 0.00 ± 0.00s 2.30 ± 0.02fg 1.00 ± 0.02l-o 2.40 ± 0.06ef 0.50 ± 0.01qr 1.24 ± 0.26F

B= Boiling on burner , M= heating in microwave; BM= boiling method In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table -32: Means comparison of yield for sapodilla fruit peel after 20 min of boiling using 0.1N HCl (Mechanical procedure

pH interaction mean±SE)

pH level Mechanical procedure Mean

Homogenizing Grinding Cutting Chopping Hammering

1 0.80 ± 0.13k 1.18 ± 0.19j 1.48 ± 0.03i 0.05 ± 0.00m 0.10 ± 0.02m 0.72 ± 0.11E

3 3.53 ± 0.60e 3.78 ± 0.12d 2.55 ± 0.27g 5.63 ± 0.40b 0.53 ± 0.21l 3.20 ± 0.34A

5 5.93 ± 0.19a 2.93 ± 0.40f 1.85 ± 0.16h 2.65 ± 0.19g 0.08 ± 0.01m 2.69 ± 0.37B

6 4.35 ± 0.38c 3.05 ± 0.17f 1.65 ± 0.09hi 2.65 ± 0.08g 0.05 ± 0.00m 2.35 ± 0.28C

7 3.48 ± 0.40e 3.08 ± 0.12f 1.58 ± 0.10i 0.40 ± 0.13l 0.08 ± 0.01m 1.72 ± 0.27D

Mean 3.62 ± 0.35A 2.80 ± 0.19B 1.82 ± 0.10D 2.27 ± 0.38C 0.17 ± 0.05E In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

Table - 33: Means comparison of yield for sapodilla fruit peel after 20 min of boiling using 0.1N HCl (Mechanical procedure

boiling method interaction mean±SE)

Boiling Mechanical procedure Mean

Method Homogenizing Grinding Cutting Chopping Hammering

129

B 2.86 ± 0.44c 2.52 ± 0.28d 1.54 ± 0.06g 1.93 ± 0.47f 0.06 ± 0.01i 1.78 ± 0.18B

M 4.37 ± 0.47a 3.08 ± 0.23b 2.10 ± 0.15e 2.62 ± 0.61d 0.27 ± 0.10h 2.49 ± 0.22A B= Boiling on burner , M= heating in microwave; BM= boiling method

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table - 34: Means comparison of yield for sapodilla fruit peel after 20 min of boiling using 0.1N HCl (Mechanical procedure

pH boiling method interaction mean±SE)

BM pH Mechanical procedure BM pH

Homogenizing Grinding Cutting Chopping Hammering interaction mean

B 1 0.50 ± 0.01ab 0.75 ± 0.01YZa 1.45 ± 0.03UVW 0.05 ± 0.00c 0.05 ± 0.00c 0.56 ± 0.14H

B 3 2.20 ± 0.04OPQ 3.55 ± 0.10GH 1.95 ± 0.01P-S 4.75 ± 0.09D 0.05 ± 0.00c 2.50 ± 0.42C

B 5 5.50 ± 0.07B 2.05 ± 0.03PQR 1.50 ± 0.05TUV 2.25 ± 0.06NOP 0.05 ± 0.00c 2.27 ± 0.48D

B 6 3.50 ± 0.05GHI 3.40 ± 0.13HIJ 1.45 ± 0.03UVW 2.50 ± 0.06MNO 0.05 ± 0.00c 2.18 ± 0.35D

B 7 2.60 ± 0.02MN 2.85 ± 0.09KLM 1.35 ± 0.02VWX 0.10 ± 0.01c 0.10 ± 0.01c 1.40 ± 0.31F

M 1 1.10 ± 0.01WXY 1.60 ± 0.03S-V 1.50 ± 0.05TUV 0.05 ± 0.00c 0.15 ± 0.01bc 0.88 ± 0.18G

M 3 4.85 ± 0.13CD 4.00 ± 0.12EF 3.15 ± 0.06IJK 6.50 ± 0.17A 1.00 ± 0.03XYZ 3.90 ± 0.49A

M 5 6.35 ± 0.05A 3.80 ± 0.12FG 2.20 ± 0.03OPQ 3.05 ± 0.10JKL 0.10 ± 0.01c 3.10 ± 0.55B

M 6 5.20 ± 0.04BC 2.70 ± 0.08LM 1.85 ± 0.05Q-T 2.79 ± 0.10KLM 0.05 ± 0.01c 2.52 ± 0.45C

M 7 4.35 ± 0.14E 3.30 ± 0.10HIJ 1.80 ± 0.05R-U 0.70 ± 0.02Za 0.05 ± 0.00c 2.04 ± 0.43E

B= Boiling on burner , M= heating in microwave; BM= boiling method

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison

among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

Table - 35: Means comparison of yield for sapodilla fruit peel after 40 min of boiling using 0.1N HCl (Mechanical procedure

pH interaction mean±SE)

130

pH level Mechanical procedure Mean

Homogenizing Grinding Cutting Chopping Hammering

1 0.05 ± 0.00l 0.63 ± 0.23ij 0.05 ± 0.00l 0.05 ± 0.00l 0.05 ± 0.00l 0.17 ± 0.06D

3 0.08 ± 0.01kl 1.18 ± 0.08g 0.25 ± 0.09k 1.48 ± 0.06ef 1.90 ± 0.27d 0.98 ± 0.14C

5 0.05 ± 0.00l 2.58 ± 0.08b 2.58 ± 0.62b 2.20 ± 0.08c 3.10 ± 0.09a 2.10 ± 0.23A

6 1.65 ± 0.02e 2.60 ± 0.23b 0.83 ± 0.08h 3.15 ± 0.20a 2.35 ± 0.07c 2.12 ± 0.16A

7 0.50 ± 0.20j 1.35 ± 0.18fg 0.80 ± 0.31hi 1.50 ± 0.23ef 2.63 ± 0.09b 1.36 ± 0.16B

Mean 0.47 ± 0.12D 1.67 ± 0.16B 0.90 ± 0.21C 1.68 ± 0.20B 2.01 ± 0.20A

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table - 36: Means comparison of yield for sapodilla fruit peel after 40 min of boiling using 0.1N HCl ( Mechanical procedure

boiling method interaction mean±SE)

Boiling Mechanical procedure Mean

Method Homogenizing Grinding Cutting Chopping Hammering

B 0.37 ± 0.17f 1.31 ± 0.22d 0.49 ± 0.11e 1.48 ± 0.24c 2.05 ± 0.28a 1.14 ± 0.12B

M 0.56 ± 0.17e 2.02 ± 0.21a 1.31 ± 0.38d 1.87 ± 0.31b 1.96 ± 0.31ab 1.54 ± 0.14A

B= Boiling on burner , M= heating in microwave; BM= boiling method

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table - 37: Means comparison of yield for sapodilla fruit peel after 40 min of boiling using 0.1N HCl (Mechanical procedure

pH boiling method interaction mean±SE)

BM pH Mechanical procedure BM pH

Homogenizing Grinding Cutting Chopping Hammering interaction mean

B 1 0.05 ± 0.00r 0.10 ± 0.00r 0.05 ± 0.00r 0.05 ± 0.00r 0.05 ± 0.01r 0.06 ± 0.01H

B 3 0.05 ± 0.00r 1.00 ± 0.01p 0.45 ± 0.01q 1.60 ± 0.05lm 2.50 ± 0.09fgh 1.12 ± 0.23E

B 5 0.05 ± 0.00r 2.40 ± 0.05h 1.20 ± 0.01op 2.05 ± 0.05j 3.00 ± 0.11cd 1.74 ± 0.28D

131

B 6 1.65 ± 0.02l 2.10 ± 0.02ij 0.65 ± 0.01q 2.70 ± 0.03efg 2.25 ± 0.08hij 1.87 ± 0.19C

B 7 0.05 ± 0.00r 0.95 ± 0.01p 0.10 ± 0.01r 1.00 ± 0.06p 2.45 ± 0.05gh 0.91 ± 0.23F

M 1 0.05 ± 0.00r 1.15 ± 0.01op 0.05 ± 0.00r 0.05 ± 0.01r 0.05 ± 0.00r 0.27 ± 0.12G

M 3 0.10 ± 0.00r 1.35 ± 0.02mno 0.05 ± 0.00r 1.35 ± 0.05mno 1.30 ± 0.04no 0.83 ± 0.17F

M 5 0.05 ± 0.00r 2.75 ± 0.06def 3.95 ± 0.10a 2.35 ± 0.09hi 3.20 ± 0.13c 2.46 ± 0.35A

M 6 1.65 ± 0.03l 3.10 ± 0.09c 1.00 ± 0.01p 3.60 ± 0.08b 2.45 ± 0.07gh 2.36 ± 0.25B

M 7 0.95 ± 0.01p 1.75 ± 0.03kl 1.50 ± 0.03lmn 2.00 ± 0.06jk 2.80 ± 0.07de 1.80 ± 0.16CD

B= Boiling on burner , M= heating in microwave; BM= boiling method

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

132

Table - 38: Means comparison of yield for sapodilla fruit peel after 60 min of boiling using 0.1N HCl Mechanical procedure

pH interaction mean±SE)

pH level Mechanical procedure Mean

Homogenizing Grinding Cutting Chopping Hammering

1 1.50 ± 0.13g 0.30 ± 0.02lm 0.08 ± 0.01mn 2.30 ± 0.21f 0.05 ± 0.00n 0.85 ± 0.17D

3 3.63 ± 0.21c 0.05 ± 0.00n 0.90 ± 0.22j 2.13 ± 0.40f 2.78 ± 0.04e 1.90 ± 0.26C

5 4.55 ± 0.06a 1.08 ± 0.46ij 0.28 ± 0.10lmn 3.78 ± 0.82c 4.35 ± 0.74ab 2.81 ± 0.40A

6 3.23 ± 0.10d 0.05 ± 0.00n 0.50 ± 0.18kl 4.13 ± 0.40b 3.60 ± 0.41c 2.30 ± 0.33B

7 2.80 ± 0.47e 1.18 ± 0.50hi 0.55 ± 0.22k 3.63 ± 0.04c 1.40 ± 0.56gh 1.91 ± 0.27C

Mean 3.14 ± 0.21A 0.53 ± 0.16C 0.46 ± 0.09C 3.19 ± 0.24A 2.44 ± 0.34B In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table - 39: Means comparison of yield for sapodilla fruit peel after 60 min of boiling using 0.1N HCl (Mechanical procedure

boiling method interaction mean±SE)

Boiling Mechanical procedure Mean

Method Homogenizing Grinding Cutting Chopping Hammering

B 2.91 ± 0.34d 0.09 ± 0.02i 0.13 ± 0.04i 2.38 ± 0.24e 1.67 ± 0.34f 1.44 ± 0.17B

M 3.37 ± 0.25b 0.97 ± 0.27g 0.79 ± 0.12h 4.00 ± 0.30a 3.20 ± 0.54c 2.47 ± 0.21A B= Boiling on burner , M= heating in microwave; BM= boiling method

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table - 40: Means comparison of yield for sapodilla fruit peel after 60 min of boiling using 0.1N HCl (Mechanical procedure

pH boiling method interaction mean±SE)

BM pH Mechanical procedure BM pH

Homogenizing Grinding Cutting Chopping Hammering interaction mean

B 1 1.20 ± 0.01qr 0.25 ± 0.01st 0.05 ± 0.00t 1.85 ± 0.05o 0.05 ± 0.00t 0.68 ± 0.19G

B 3 4.10 ± 0.03e 0.05 ± 0.00t 0.40 ± 0.01st 1.25 ± 0.03qr 2.75 ± 0.05l 1.71 ± 0.41E

133

B 5 4.50 ± 0.04d 0.05 ± 0.00t 0.05 ± 0.00t 1.95 ± 0.03no 2.70 ± 0.02l 1.85 ± 0.45D

B 6 3.00 ± 0.06jkl 0.05 ± 0.00t 0.10 ± 0.01t 3.25 ± 0.09hij 2.70 ± 0.10l 1.82 ± 0.38DE

B 7 1.75 ± 0.03op 0.05 ± 0.01t 0.05 ± 0.01t 3.60 ± 0.04fgh 0.15 ± 0.01st 1.12 ± 0.37F

M 1 1.80 ± 0.02o 0.35 ± 0.00st 0.10 ± 0.01t 2.75 ± 0.09l 0.05 ± 0.01t 1.01 ± 0.29F

M 3 3.15 ± 0.05ijk 0.05 ± 0.00t 1.40 ± 0.03pq 3.00 ± 0.12jkl 2.80 ± 0.08kl 2.08 ± 0.32C

M 5 4.60 ± 0.11d 2.10 ± 0.04no 0.50 ± 0.02s 5.60 ± 0.07b 6.00 ± 0.21a 3.76 ± 0.57A

M 6 3.45 ± 0.03ghi 0.05 ± 0.00t 0.90 ± 0.02r 5.00 ± 0.17c 4.50 ± 0.14d 2.78 ± 0.53B

M 7 3.85 ± 0.06ef 2.30 ± 0.08mn 1.05 ± 0.02qr 3.65 ± 0.07fg 2.65 ± 0.11lm 2.70 ± 0.27B

B= Boiling on burner , M= heating in microwave; BM= boiling method In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

134

Table - 41: Percentage yield of pectin from sapodilla fruit peel after using different

physicomechanical procedures (pH, mechanical procedure, boiling method and time of

boling) with IN HCl.

Mechanical

procedure pH

% yield 10 min %Yield 20 min %yield 40 min % yield 60 min

B M B M B M B M

Hammering

1 2.65 3.15 1.5 2.2 3.5 3 0.3 2

3 5.65 6.55 3.45 5.45 4.5 4.5 0.4 2.85

5 2.3 4.6 4.4 4.5 4.2 5.25 0.15 2.3

6 2.15 2.95 2.45 4.3 2 3.4 3.45 2.6

7 2.5 3.1 2.4 2.45 2 2.55 3.5 0.05

Grinding

1 3.1 4.15 1.55 1.65 1.4 3.7 1.4 3

3 5.6 4.8 2.55 5.4 3.25 5.15 3.15 5

5 1.05 6.1 4.2 5.1 3.8 5.5 1 3.3

6 1.45 3.55 4.3 3.65 3.5 3.3 0.8 1.2

7 0.35 2.75 1.3 2.5 2.15 2.6 1.65 1.6

Cutting

1 0.05 0.05 0.5 1.4 3.65 4 0.65 0.05

3 0.05 1.6 1 2.35 2.5 3.5 0.75 0.1

5 2.95 5.3 2.05 2.7 4.1 5.5 0.1 0.05

6 0.15 3.5 3.35 3.7 1.5 3 0 0.6

7 0.05 1.2 2.6 1.8 2 3.25 0.1 0.05

Homogenizing

1 1.55 4.95 2.1 3.6 3.4 4.6 0.6 0

3 2.4 5.35 3.55 5.25 3.65 4.55 0.05 0.15

5 1.7 4.85 3.55 5.9 2.7 3 0.15 3.4

6 0.6 1.5 3.8 3.15 3.9 3.5 0.05 0.25

7 1.2 2.55 1.25 2.2 3.35 3.3 0.1 0.6

Chopping 1 3 5.5 2.5 5.9 1.95 3.2 0.05 3.55

(mortor/pastle 3 3.7 5.4 1.4 6 1.15 4.85 0.2 0.1

5 4.1 6.7 1.25 6.1 4.5 6.5 0.45 0.5

6 3.75 3.8 1.2 4.9 2.5 3.1 0 1

7 0.05 0.25 1.8 2.5 1 3.95 0.1 0.6

B= Boiling on burner , M= heating in microwave

135

Table - 42: Analysis of variance (mean squares) of yield of pectin from sapodilla fruit peel after using different

physicomechanical procedure different fruits with 1N HCl

Source of

variation

Degrees of

freedom

Mean squares

Yield 10 min_NHCl Yield 20 min_NHCl Yield 40 min_NHCl Yield 60 min_NHCl

MP

pH

BM

MP x pH

MP x BM

pH x BM

MP x pH x BM

Error

Total

4

4

1

16

4

4

16

100

149

23.648**

38.736**

106.361**

8.396**

1.944**

4.779**

2.360**

0.022

8.5850**

21.8147**

71.7604**

3.9668**

10.4560**

4.7719**

1.5394**

0.0173

0.5439**

16.3404**

42.5601**

3.5228**

3.5189**

0.9204**

1.2315**

0.0168

21.8563**

0.8645**

14.8838**

4.0399**

2.1424**

4.7014**

3.3080**

0.0054

NS = Non-significant (P>0.05); * = Significant (P<0.05); ** = Highly significant (P<0.01); Mech= mechanical

136

Table - 43: Means comparison of yield for sapodilla fruit peel after 10 min of boiling using 1N HCl ( Mechanical procedure

pH interaction mean±SE)

pH level Mechanical procedure Mean

Homogenizing Grinding Cutting Chopping Hammering

1 2.90 ± 0.12j 3.63 ± 0.24fg 0.05 ± 0.00p 4.25 ± 0.56cd 3.25 ± 0.76i 2.82 ± 0.33C

3 6.10 ± 0.21a 5.20 ± 0.19b 0.83 ± 0.35no 4.55 ± 0.39c 3.88 ± 0.66ef 4.11 ± 0.37A

5 3.45 ± 0.51ghi 3.58 ± 1.13fgh 4.13 ± 0.53de 5.40 ± 0.59b 3.28 ± 0.71hi 3.97 ± 0.34B

6 2.55 ± 0.18k 2.50 ± 0.47k 1.83 ± 0.75lm 3.78 ± 0.07f 1.05 ± 0.20n 2.34 ± 0.24D

7 2.80 ± 0.14jk 1.55 ± 0.54m 0.63 ± 0.26o 0.15 ± 0.05p 1.88 ± 0.30l 1.40 ± 0.22E

Mean 3.56 ± 0.27A 3.29 ± 0.34B 1.49 ± 0.33D 3.63 ± 0.38A 2.67 ± 0.31C In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table - 44: Means comparison of yield for sapodilla fruit peel after 10 min of boiling using 1N HCl ( Mechanical procedure

boiling method interaction mean±SE)

Boiling Mechanical procedure Mean

Method Homogenizing Grinding Cutting Chopping Hammering

B 3.05 ± 0.35d 2.31 ± 0.50e 0.65 ± 0.31g 2.92 ± 0.40d 1.49 ± 0.16f 2.08 ± 0.19B

M 4.07 ± 0.37b 4.27 ± 0.31a 2.33 ± 0.50e 4.33 ± 0.60a 3.84 ± 0.41c 3.77 ± 0.21A

B= Boiling on burner , M= heating in microwave

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison

among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

Table - 45: Means comparison of yield for sapodilla fruit peel after 10 min of boiling using 1N HCl (Mechanical procedure

pH boiling method interaction mean±SE)

BM pH Mechanical procedure BM pH

Homogenizing Grinding Cutting Chopping Hammering interaction mean

B 1 2.65 ± 0.03n-q 3.10 ± 0.05lmn 0.05 ± 0.01w 3.00 ± 0.06mno 1.55 ± 0.03st 2.07 ± 0.31F

137

B 3 5.65 ± 0.12cd 5.60 ± 0.07cd 0.05 ± 0.01w 3.70 ± 0.10jk 2.40 ± 0.08pq 3.48 ± 0.56C

B 5 2.30 ± 0.05pq 1.05 ± 0.03tu 2.95 ± 0.11no 4.10 ± 0.15ij 1.70 ± 0.04rs 2.42 ± 0.28E

B 6 2.15 ± 0.04qr 1.45 ± 0.01st 0.15 ± 0.01vw 3.75 ± 0.08jk 0.60 ± 0.01uv 1.62 ± 0.34G

B 7 2.50 ± 0.05opq 0.35 ± 0.01vw 0.05 ± 0.01w 0.05 ± 0.01w 1.20 ± 0.03st 0.83 ± 0.25H

M 1 3.15 ± 0.09lmn 4.15 ± 0.05ij 0.05 ± 0.00w 5.50 ± 0.08d 4.95 ± 0.12e-h 3.56 ± 0.52C

M 3 6.55 ± 0.08ab 4.80 ± 0.13gh 1.60 ± 0.05s 5.40 ± 0.20de 5.35 ± 0.10def 4.74 ± 0.45B

M 5 4.60 ± 0.02hi 6.10 ± 0.20bc 5.30 ± 0.07d-g 6.70 ± 0.24a 4.85 ± 0.16fgh 5.51 ± 0.22A

M 6 2.95 ± 0.09no 3.55 ± 0.05kl 3.50 ± 0.05klm 3.80 ± 0.13jk 1.50 ± 0.05st 3.06 ± 0.22D

M 7 3.10 ± 0.01lmn 2.75 ± 0.09nop 1.20 ± 0.03st 0.25 ± 0.02vw 2.55 ± 0.05opq 1.97 ± 0.29F

B= Boiling on burner , M= heating in microwave; BM= boiling method

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table - 46: Means comparison of yield for sapodilla fruit peel after 20 min of boiling using 1N HCl Mechanical procedure

pH interaction mean±SE)

pH level Mechanical procedure Mean

Homogenizing Grinding Cutting Chopping Hammering

1 1.82 ± 0.15kl 1.60 ± 0.03l 0.95 ± 0.20m 4.20 ± 0.77cd 2.85 ± 0.34h 2.28 ± 0.27D

3 4.45 ± 0.45abc 3.98 ± 0.64de 1.68 ± 0.30kl 3.70 ± 1.03ef 4.40 ± 0.39bc 3.64 ± 0.32B

5 4.45 ± 0.06abc 4.65 ± 0.21ab 2.38 ± 0.15i 3.68 ± 1.09f 4.73 ± 0.53a 3.98 ± 0.28A

6 3.38 ± 0.41g 3.98 ± 0.15de 3.53 ± 0.10fg 3.05 ± 0.83h 3.48 ± 0.16fg 3.48 ± 0.19C

7 2.43 ± 0.03i 1.90 ± 0.27jk 2.20 ± 0.18i 2.15 ± 0.16ij 1.73 ± 0.21kl 2.08 ± 0.09E

Mean 3.30 ± 0.23BC 3.22 ± 0.27C 2.15 ± 0.18D 3.36 ± 0.37AB 3.44 ± 0.25A

138

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

139

Table - 47: Means comparison of yield for sapodilla fruit peel after 20 min of boiling using 1N HCl ( Mechanical procedure

boiling method interaction mean±SE)

Boiling Mechanical procedure Mean

Method Homogenizing Grinding Cutting Chopping Hammering

B 2.84 ± 0.27d 2.78 ± 0.34d 1.90 ± 0.28f 1.63 ± 0.13g 2.85 ± 0.27d 2.40 ± 0.13B

M 3.77 ± 0.34c 3.66 ± 0.39c 2.39 ± 0.21e 5.08 ± 0.37a 4.02 ± 0.37b 3.78 ± 0.18A

B= Boiling on burner , M= heating in microwave In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

140

Table - 48: Means comparison of yield for sapodilla fruit peel after 20 min of boiling using 1N HCl (Mechanical procedure

pH boiling method interaction mean±SE)

BM pH Mechanical procedure BM pH

Homogenizing Grinding Cutting Chopping Hammering interaction mean

B 1 1.50 ± 0.02qrs 1.55 ± 0.03qrs 0.50 ± 0.01u 2.50 ± 0.06j-m 2.10 ± 0.05mno 1.63 ± 0.18F

B 3 3.45 ± 0.02ghi 2.55 ± 0.05jkl 1.00 ± 0.02t 1.40 ± 0.03q-t 3.55 ± 0.06ghi 2.39 ± 0.28D

B 5 4.40 ± 0.06e 4.20 ± 0.07ef 2.05 ± 0.03nop 1.25 ± 0.02rst 3.55 ± 0.05ghi 3.09 ± 0.33C

B 6 2.45 ± 0.04j-n 4.30 ± 0.04e 3.35 ± 0.05hi 1.20 ± 0.03st 3.80 ± 0.10fg 3.02 ± 0.29C

B 7 2.40 ± 0.05j-n 1.30 ± 0.01rst 2.60 ± 0.09jk 1.80 ± 0.05opq 1.25 ± 0.01rst 1.87 ± 0.15E

M 1 2.13 ± 0.07l-o 1.65 ± 0.01pqr 1.40 ± 0.01q-t 5.90 ± 0.19a 3.60 ± 0.08gh 2.94 ± 0.45C

M 3 5.45 ± 0.08b 5.40 ± 0.13b 2.35 ± 0.02j-n 6.00 ± 0.15a 5.25 ± 0.14bc 4.89 ± 0.35A

M 5 4.50 ± 0.10de 5.10 ± 0.10bc 2.70 ± 0.06j 6.10 ± 0.08a 5.90 ± 0.17a 4.86 ± 0.33A

M 6 4.30 ± 0.03e 3.65 ± 0.06gh 3.70 ± 0.14gh 4.90 ± 0.11cd 3.15 ± 0.07i 3.94 ± 0.16B

M 7 2.45 ± 0.04j-n 2.50 ± 0.08j-m 1.80 ± 0.04opq 2.50 ± 0.05j-m 2.20 ± 0.06k-o 2.29 ± 0.08D

B= Boiling on burner , M= heating in microwave; BM= boiling method

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison

among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

141

Table - 49: Means comparison of yield for sapodilla fruit peel after 40 min of boiling using 1N HCl (Mechanical procedure

pH interaction mean±SE)

pH level Mechanical procedure Mean

Homogenizing Grinding Cutting Chopping Hammering

1 3.25 ± 0.12gh 2.55 ± 0.51k-n 3.83 ± 0.09ef 2.58 ± 0.28j-m 4.00 ± 0.28de 3.24 ± 0.17C

3 4.50 ± 0.05c 4.20 ± 0.43d 3.00 ± 0.23hi 3.00 ± 0.83hi 4.10 ± 0.21de 3.76 ± 0.22B

5 4.73 ± 0.24bc 4.65 ± 0.38bc 4.80 ± 0.32b 5.50 ± 0.46a 2.85 ± 0.08ij 4.51 ± 0.21A

6 2.70 ± 0.32jkl 3.38 ± 0.04g 2.25 ± 0.34o 2.80 ± 0.14ijk 3.70 ± 0.10f 2.97 ± 0.13D

7 2.28 ± 0.13no 2.38 ± 0.10mno 2.63 ± 0.28j-m 2.48 ± 0.66l-o 3.33 ± 0.04g 2.62 ± 0.15E

Mean 3.49 ± 0.20B 3.43 ± 0.22B 3.30 ± 0.20C 3.27 ± 0.31C 3.60 ± 0.11A

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table - 50: Means comparison of yield for sapodilla fruit peel after 40 min of boiling using 1N HCl (Mechanical procedure

boiling method interaction mean±SE)

Boiling Mechanical procedure Mean

Method Homogenizing Grinding Cutting Chopping Hammering

B 3.24 ± 0.29e 2.81 ± 0.24f 2.75 ± 0.26f 2.22 ± 0.34g 3.40 ± 0.11d 2.88 ± 0.12B

M 3.74 ± 0.27c 4.05 ± 0.30b 3.85 ± 0.24c 4.32 ± 0.34a 3.79 ± 0.18c 3.95 ± 0.12A

B= Boiling on burner , M= heating in microwave

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table - 51: Means comparison of yield for sapodilla fruit peel after 40 min of boiling using 1N HCl ( Mechanical procedure

pH boiling method interaction mean±SE)

BM pH Mechanical procedure BM pH

Homogenizing Grinding Cutting Chopping Hammering interaction mean

B 1 3.50 ± 0.05j-n 1.40 ± 0.02uv 3.65 ± 0.05i-m 1.95 ± 0.04t 3.40 ± 0.10k-o 2.78 ± 0.25G

142

B 3 4.50 ± 0.11def 3.25 ± 0.04mno 2.50 ± 0.05rs 1.15 ± 0.02uv 3.65 ± 0.12i-m 3.01 ± 0.30F

B 5 4.20 ± 0.04efg 3.80 ± 0.11g-k 4.10 ± 0.09fgh 4.50 ± 0.04def 2.70 ± 0.10pqr 3.86 ± 0.17C

B 6 2.00 ± 0.04t 3.47 ± 0.03j-n 1.50 ± 0.01u 2.50 ± 0.03rs 3.90 ± 0.05g-j 2.67 ± 0.24G

B 7 2.00 ± 0.04t 2.15 ± 0.03st 2.00 ± 0.03t 1.00 ± 0.03v 3.35 ± 0.08l-o 2.10 ± 0.20H

M 1 3.00 ± 0.11opq 3.70 ± 0.05h-l 4.00 ± 0.10ghi 3.20 ± 0.05no 4.60 ± 0.09de 3.70 ± 0.16D

M 3 4.50 ± 0.04def 5.15 ± 0.14bc 3.50 ± 0.03j-n 4.85 ± 0.09cd 4.55 ± 0.09de 4.51 ± 0.15B

M 5 5.25 ± 0.09bc 5.50 ± 0.07b 5.50 ± 0.08b 6.50 ± 0.19a 3.00 ± 0.05opq 5.15 ± 0.31A

M 6 3.40 ± 0.10k-o 3.30 ± 0.03l-o 3.00 ± 0.04opq 3.10 ± 0.06nop 3.50 ± 0.08j-n 3.26 ± 0.06E

M 7 2.55 ± 0.07rs 2.60 ± 0.05qr 3.25 ± 0.07mno 3.95 ± 0.14ghi 3.30 ± 0.05l-o 3.13 ± 0.14EF

B= Boiling on burner , M= heating in microwave; BM= boiling method

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison

among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

Table - 52: Means comparison of yield for sapodilla fruit peel after 60 min of boiling using 1N HCl (Mechanical procedure

pH interaction mean±SE)

pH level Mechanical procedure Mean

Homogenizing Grinding Cutting Chopping Hammering

1 1.15 ± 0.38fg 2.20 ± 0.36c 0.35 ± 0.13hi 1.80 ± 0.78d 0.30 ± 0.13ij 1.16 ± 0.23B

3 1.63 ± 0.55e 4.08 ± 0.42a 0.43 ± 0.15hi 0.15 ± 0.02jk 0.10 ± 0.02k 1.28 ± 0.31A

5 1.23 ± 0.48f 2.15 ± 0.52c 0.08 ± 0.01k 0.48 ± 0.02h 1.78 ± 0.73de 1.14 ± 0.24B

6 3.03 ± 0.20b 1.00 ± 0.09g 0.30 ± 0.13ij 0.50 ± 0.22h 0.15 ± 0.05jk 1.00 ± 0.21C

7 1.78 ± 0.77de 1.63 ± 0.03e 0.08 ± 0.01k 0.35 ± 0.11hi 0.35 ± 0.11hi 0.84 ± 0.20D

Mean 1.76 ± 0.25B 2.21 ± 0.24A 0.25 ± 0.05E 0.66 ± 0.19C 0.54 ± 0.18D In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table - 53: Means comparison of yield for sapodilla fruit peel after 60 min of boiling using 1N HCl (Mechanical procedure

boiling method interaction mean±SE)

143

Boiling Mechanical procedure Mean

Method Homogenizing Grinding Cutting Chopping Hammering

B 1.56 ± 0.42c 1.60 ± 0.22c 0.32 ± 0.08f 0.16 ± 0.04g 0.19 ± 0.06g 0.77 ± 0.12B

M 1.96 ± 0.27b 2.82 ± 0.36a 0.17 ± 0.06g 1.15 ± 0.33d 0.88 ± 0.34e 1.40 ± 0.17A

B= Boiling on burner , M= heating in microwave

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table - 54: Means comparison of yield for sapodilla fruit peel after 60 min of boiling using 1N HCl (Mechanical procedure

pH boiling method interaction mean±SE)

BM pH Mechanical procedure BM pH

Homogenizing Grinding Cutting Chopping Hammering interaction mean

B 1 0.30 ± 0.01r-u 1.40 ± 0.05kl 0.65 ± 0.03op 0.05 ± 0.01vw 0.60 ± 0.02opq 0.60 ± 0.12E

B 3 0.40 ± 0.02q-t 3.15 ± 0.07de 0.75 ± 0.03o 0.20 ± 0.01t-w 0.05 ± 0.00vw 0.91 ± 0.31D

B 5 0.15 ± 0.01uvw 1.00 ± 0.03mn 0.10 ± 0.01uvw 0.45 ± 0.01p-s 0.15 ± 0.01uvw 0.37 ± 0.09F

B 6 3.45 ± 0.09bc 0.80 ± 0.03no 0.00 ± 0.00w 0.00 ± 0.00w 0.05 ± 0.01vw 0.86 ± 0.36D

B 7 3.50 ± 0.08bc 1.65 ± 0.03j 0.10 ± 0.01uvw 0.10 ± 0.01uvw 0.10 ± 0.01uvw 1.09 ± 0.36C

M 1 2.00 ± 0.05i 3.00 ± 0.08ef 0.05 ± 0.00vw 3.55 ± 0.07b 0.00 ± 0.00w 1.72 ± 0.39B

M 3 2.85 ± 0.05f 5.00 ± 0.10a 0.10 ± 0.01uvw 0.10 ± 0.01uvw 0.15 ± 0.01uvw 1.64 ± 0.53B

M 5 2.30 ± 0.09h 3.30 ± 0.12cd 0.05 ± 0.00vw 0.50 ± 0.02pqr 3.40 ± 0.05bc 1.91 ± 0.37A

M 6 2.60 ± 0.07g 1.20 ± 0.04lm 0.60 ± 0.03opq 1.00 ± 0.05mn 0.25 ± 0.02s-v 1.13 ± 0.22C

M 7 0.05 ± 0.01vw 1.60 ± 0.05jk 0.05 ± 0.00vw 0.60 ± 0.02opq 0.60 ± 0.02opq 0.58 ± 0.15E B= Boiling on burner , M= heating in microwave; BM= boiling method

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

144

Table - 55: Percentage yield of pectin from sapodilla fruit peel after using different

organic acid

Mechanical

procedure Inorganic acid

% of inorganic

acid pH

% yield

B M h

om

og

en

izin

g

Citric acid 1%

3 to 5

2.55 3.25

10% 2.55 4.24

Oxalic acid 1% 3.95 4.95

10% 0.5 3.7

Tartaric acid 1% 1.8 3.25

10% 2.5 2.55

ch

op

pin

g

Citric acid 1% 2.4 4.3

10% 0.05 0.1

Oxalic acid 1% 0.05 1.5

10% 0.15 1.85

Tartaric acid 1% 0.75 1.25

10% 0.35 3.2

Grin

din

g

Citric acid 1% 1.35 2.15

10% 0.05 1.2

Oxalic acid 1% 0.1 0.3

10% 1.25 1.05

Tartaric acid 1% 1.8 2.43

10% 1.65 3.15

Cu

ttin

g

Citric acid 1% 1.4 2.64

10% 0.1 1.1

Oxalic acid 1% 0.05 1.2

10% 0.13 1.32

Tartaric acid 1% 0.2 1.87

10% 0.32 1.45

Ha

mm

erin

g

Citric acid 1% 1.6 1.8

10% 0.54 1.67

Oxalic acid 1% 0.1 1.3

10% 0.43 2

Tartaric acid 1% 0.33 1.4

10% 0.44 2.1

B= Boiling on burner , M= heating in microwave

145

Table - 56 : Analysis of variance (mean squares) of yield of pectin from sapodilla

fruit peel after using different organic acid

Source of

variation

Degrees of

freedom

Mean squares

Yield (citric acid) Yield (oxalic

acid)

Yield (tartaric

acid)

BM

MP

Acid%

BM x MP

BM x Acid%

MP x Acid%

BM x MP x Acid%

Error

Total

1

4

1

4

1

4

4

40

59

14.5829**

7.7555**

21.0278**

0.1237**

0.0049NS

5.6981**

1.0202**

0.0126

23.2877**

14.9075**

0.1882**

1.8114**

0.9077**

5.0161**

0.7483**

0.0090

23.4750**

6.0553**

1.0375**

0.3754**

0.5245**

0.3919**

1.5337**

0.0101

NS = Non-significant (P>0.05); * = Significant (P<0.05); ** = Highly significant (P<0.01) BM= Boiling method. MP= Mechanical procedure

Table - 57: Means comparison of yield for sapodilla fruit peel after using citric acid

(Boiling method x mechanical procedure interaction mean±SE)

Mechanical

procedure

Boiling Method Mean

B M

Homogenizing 2.55 ± 0.05b 3.75 ± 0.23a 3.15 ± 0.21A

Chopping 1.23 ± 0.53e 2.20 ± 0.94c 1.71 ± 0.54B

Grinding 0.70 ± 0.29f 1.68 ± 0.21d 1.19 ± 0.23D

Cutting 0.75 ± 0.29f 1.87 ± 0.34d 1.31 ± 0.27CD

Hammering 1.07 ± 0.24e 1.74 ± 0.04d 1.40 ± 0.15C

Mean 1.26 ± 0.18B 2.25 ± 0.24A

B= Boiling on burner , M= heating in microwave.In a row and column statistically non-significant (P>0.05)

represents mean sharing and are denoted by similar letters. Comparison among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

Table - 58: Means comparison of yield for sapodilla fruit peel after using citric acid

Acid (% x boiling method interaction mean±SE)

Inorganic Boiling Method Mean

146

acid B M

1% 1.86 ± 0.14b 2.83 ± 0.24a 2.34 ± 0.16A

10% 0.66 ± 0.26d 1.66 ± 0.37c 1.16 ± 0.24B B= Boiling on burner , M= heating in microwave Among the two concentrations of acid 1% has found more effective with microwave as heating procedure

Table - 59: Means comparison of yield for sapodilla fruit peel after using citric acid

(Boiling method x Acid% x mechanical procedure interaction mean±SE)

Acid% x MP Boiling Method

Mean B M

1% x MP1 2.55 ± 0.08c 3.25 ± 0.10b 2.90 ± 0.17B

1% x MP2 2.40 ± 0.09cd 4.30 ± 0.16a 3.35 ± 0.43A

1% x MP3 1.35 ± 0.04fg 2.15 ± 0.04d 1.75 ± 0.18D

1% x MP4 1.40 ± 0.02fg 2.64 ± 0.04c 2.02 ± 0.28C

1% x MP5 1.60 ± 0.04ef 1.80 ± 0.05e 1.70 ± 0.05D

10% x MP1 2.55 ± 0.08c 4.24 ± 0.12a 3.40 ± 0.38A

10% x MP2 0.05 ± 0.00i 0.10 ± 0.00i 0.08 ± 0.01G

10% x MP3 0.05 ± 0.00i 1.20 ± 0.02g 0.63 ± 0.26F

10% x MP4 0.10 ± 0.00i 1.10 ± 0.01g 0.60 ± 0.22F

10% x MP5 0.54 ± 0.01h 1.67 ± 0.05ef 1.11 ± 0.25E

BM= Boiling method. MP= Mechanical procedure; B= Boiling on burner , M= heating in microwave

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by

similar letters. Comparison among interaction means are represented by lower case alphabets while capital

alphabets are used for overall mean

Table - 60: Means comparison of yield for sapodilla fruit peel after using oxalic acid

(Boiling method x mechanical procedure interaction mean±SE)

Mechanical

procedure

Boiling Method Mean

B M

Homogenizing 2.23 ± 0.77b 4.33 ± 0.29a 3.28 ± 0.51A

Chopping 0.10 ± 0.02f 1.68 ± 0.08c 0.89 ± 0.24B

Grinding 0.68 ± 0.26e 0.68 ± 0.17e 0.68 ± 0.15C

Cutting 0.09 ± 0.02f 1.26 ± 0.03d 0.68 ± 0.18C

Hammering 0.27 ± 0.07f 1.65 ± 0.16c 0.96 ± 0.22B

Mean 0.67 ± 0.21B 1.92 ± 0.24A

147

B= Boiling on burner , M= heating in microwave; B= Boiling on burner , M= heating in microwave

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by

similar letters. Comparison among interaction means are represented by lower case alphabets while capital

alphabets are used for overall mean

Table - 61: Means comparison of yield for sapodilla fruit peel after using oxalic acid

Acid% x boiling method interaction mean±SE

Inorganic

acid

Boiling Method Mean

B M

1% 0.85 ± 0.42 1.85 ± 0.43 1.35 ± 0.31A

10% 0.49 ± 0.11 1.98 ± 0.25 1.24 ± 0.19B

B= Boiling on burner , M= heating in microwave

Among the two concentrations of acid 1% has found more effective with microwave as

heating procedure

Table - 62: Means comparison of yield for sapodilla fruit peel after using oxalic acid

(Boiling method x Acid% x mechanical procedure interaction mean±SE)

Acid% x MP Boiling Method

Mean B M

1% x MP1 3.95 ± 0.15b 4.95 ± 0.14a 4.45 ± 0.24A

1% x MP2 0.05 ± 0.00h 1.50 ± 0.02d 0.78 ± 0.32E

1% x MP3 0.10 ± 0.01h 0.30 ± 0.01fgh 0.20 ± 0.05F

1% x MP4 0.05 ± 0.00h 1.20 ± 0.01e 0.63 ± 0.26E

1% x MP5 0.10 ± 0.01h 1.30 ± 0.02de 0.70 ± 0.27E

10% x MP1 0.50 ± 0.02f 3.70 ± 0.10b 2.10 ± 0.72B

10% x MP2 0.15 ± 0.01gh 1.85 ± 0.03c 1.00 ± 0.38D

10% x MP3 1.25 ± 0.04de 1.05 ± 0.02e 1.15 ± 0.05CD

10% x MP4 0.13 ± 0.01h 1.32 ± 0.02de 0.73 ± 0.27E

10% x MP5 0.43 ± 0.02fg 2.00 ± 0.05c 1.22 ± 0.35C

BM= Boiling method. MP= Mechanical procedure ; B= Boiling on burner , M= heating in microwave

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by

similar letters. Comparison among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

148

Table - 63: Means comparison of yield for sapodilla fruit peel after using tartaric

acid ( Boiling method x mechanical procedure interaction mean±SE)

Mechanical

procedure

Boiling Method Mean

B M

homogenizing 2.15 ± 0.16b 2.90 ± 0.17a 2.53 ± 0.16A

Chopping 0.55 ± 0.09d 2.23 ± 0.44b 1.39 ± 0.33C

Grinding 1.73 ± 0.04c 2.79 ± 0.17a 2.26 ± 0.18B

Cutting 0.26 ± 0.03e 1.66 ± 0.10c 0.96 ± 0.22D

Hammering 0.39 ± 0.03de 1.75 ± 0.16c 1.07 ± 0.22D

Mean 1.01 ± 0.15B 2.27 ± 0.14A

B= Boiling on burner , M= heating in microwave

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by

similar letters. Comparison among interaction means are represented by lower case alphabets while capital

alphabets are used for overall mean

Table - 64: Means comparison of yield for sapodilla fruit peel after using tartaric

acid (Acid% x boiling method interaction mean±SE)

Inorganic

acid

Boiling Method Mean

B M

1% 0.98 ± 0.19c 2.04 ± 0.20b 1.51 ± 0.17B

10% 1.05 ± 0.24c 2.49 ± 0.18a 1.77 ± 0.20A

B= Boiling on burner , M= heating in microwave

Among the two concentrations of acid 1% has found more effective with microwave as heating procedure

149

Table - 65: Means comparison of yield for sapodilla fruit peel after using tartaric

acid( Boiling method x Acid% x mechanical procedure interaction mean±SE)

Acid% x MP Boiling Method

Mean B M

1% x PM1 1.80 ± 0.03cd 3.25 ± 0.08a 2.53 ± 0.33A

1% x PM2 0.75 ± 0.02g 1.25 ± 0.02f 1.00 ± 0.11E

1% x PM3 1.80 ± 0.05cd 2.43 ± 0.09b 2.12 ± 0.15B

1% x PM4 0.20 ± 0.02h 1.87 ± 0.08cd 1.04 ± 0.38E

1% x PM5 0.33 ± 0.02h 1.40 ± 0.02ef 0.87 ± 0.24E

10% x PM1 2.50 ± 0.07b 2.55 ± 0.12b 2.53 ± 0.06A

10% x PM2 0.35 ± 0.02h 3.20 ± 0.08a 1.78 ± 0.64C

10% x PM3 1.65 ± 0.03de 3.15 ± 0.10a 2.40 ± 0.34A

10% x PM4 0.32 ± 0.02h 1.45 ± 0.02ef 0.89 ± 0.25E

10% x PM5 0.44 ± 0.03h 2.10 ± 0.05c 1.27 ± 0.37D

B= Boiling on burner , M= heating in microwave; BM= Boiling method. MP= Mechanical procedure

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by

similar letters. Comparison among interaction means are represented by lower case alphabets while capital

alphabets are used for overall mean

150

Table - 66: percentage yield of pectin from sapodilla fruit peel using different

strengths of same inorganic acid.

Mechanical

procedure pH

0.1N HCl 0.5N HCl 1N HCl

B M B M B M

Hammering

1 1.5 2.05 0.65 1 2.65 3.15

3 0.6 1.55 0.05 3.3 5.65 6.55

5 0.8 1.95 0.3 2.2 2.3 4.6

6 0.8 1.4 0.45 1.65 2.15 2.95

7 0.5 0 0.25 0.05 2.5 3.1

Grinding

1 0.55 0.8 0.3 4.6 3.1 4.15

3 4 3.5 0.1 2.85 5.6 4.8

5 2.45 2.9 0.1 1 1.05 6.1

6 2 2 3.15 4.05 1.45 3.55

7 1.95 2.3 2.5 2.8 0.35 2.75

Cutting

1 0.8 0.9 3.65 4.2 0.05 0.05

3 1.1 1.5 4 2.25 0.05 1.6

5 0.4 1 4.35 3.75 2.95 5.3

6 1 1.2 0.55 4.75 0.15 3.5

7 0.7 1 1.25 0.5 0.05 1.2

Homogenizing

1 0.5 1.05 4.15 4.95 1.55 4.95

3 0.75 1.6 3.2 4.95 2.4 5.35

5 1.2 1.5 0.15 4.55 1.7 4.85

6 1.5 2 1.8 0.3 0.6 1.5

7 0.5 0.5 3.6 0.6 1.2 2.55

Chopping 1 2.7 4.7 4.4 4.65 3 5.5

(mortor/pastle 3 3.9 3.45 4.8 4.95 3.7 5.4

5 3.95 4.05 2.25 2.65 4.1 6.7

6 2.3 2.45 1.5 0.65 3.75 3.8

7 3.95 2.4 1 0.1 0.05 0.25

BM= Boiling method. MP= Mechanical procedure B= Boiling on burner , M= heating in microwave

133

Table - 67 : Analysis of variance (mean squares) of yield of pectin from sapodilla fruit peel after using different strength of

same inorganic acid

Source of

variation

Degrees of

freedom

Mean squares

Yield_0.1N HCl Yield_0.5N HCl Yield_1N HCl

MP

pH

BM

MP x pH

MP x BM

pH x BM

MP x pH x BM

Error

Total

4

4

1

16

4

4

16

100

149

32.6575**

3.3917**

3.2413**

2.5462**

0.3392**

1.0112**

0.6258**

0.0093

19.2084**

20.5359**

21.2064**

8.7942**

4.8684**

6.8626**

5.2626**

0.0104

24.375**

37.244**

109.739**

7.934**

1.832**

4.535**

2.303**

0.047

NS = Non-significant (P>0.05); * = Significant (P<0.05); ** = Highly significant (P<0.01) B= Boiling on burner , M= heating in microwave; BM= Boiling method. MP= Mechanical procedure

134

Table - 68: Means comparison for yield of pectin from sapodilla fruit peel after using 0.1N HCl ( Mechanical procedure pH

interaction mean±SE )

pH level Mechanical procedure Mean

Homogenizing Grinding Cutting Chopping Hammering

1 0.78 ± 0.12k 0.68 ± 0.06kl 0.85 ± 0.03k 3.70 ± 0.45b 1.78 ± 0.12g 1.56 ± 0.23D

3 1.18 ± 0.19hij 3.75 ± 0.13b 1.30 ± 0.09hi 3.68 ± 0.12b 1.08 ± 0.21j 2.20 ± 0.24A

5 1.35 ± 0.07h 2.68 ± 0.10d 0.70 ± 0.13kl 4.00 ± 0.09a 1.38 ± 0.26h 2.02 ± 0.23B

6 1.75 ± 0.11g 2.00 ± 0.04f 1.10 ± 0.05ij 2.38 ± 0.05e 1.10 ± 0.14ij 1.67 ± 0.10C

7 0.50 ± 0.01l 2.13 ± 0.09f 0.85 ± 0.07k 3.18 ± 0.35c 0.25 ± 0.11m 1.38 ± 0.22E

Mean 1.11 ± 0.09C 2.25 ± 0.19B 0.96 ± 0.05D 3.39 ± 0.15A 1.12 ± 0.12C

B= Boiling on burner , M= heating in microwave; BM= Boiling method. MP= Mechanical procedure

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison

among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

Table - 69: Means comparison for yield of pectin from sapodilla fruit peel after using 0.1N HCl (Mechanical procedure

boiling method interaction mean±SE)

Boiling Mechanical procedure Mean

Method Homogenizing Grinding Cutting Chopping Hammering

B 0.89 ± 0.11e 2.19 ± 0.30b 0.80 ± 0.07e 3.36 ± 0.19a 0.84 ± 0.09e 1.62 ± 0.14B

M 1.33 ± 0.14c 2.30 ± 0.24b 1.12 ± 0.06d 3.41 ± 0.24a 1.39 ± 0.20c 1.91 ± 0.13A

B= Boiling on burner , M= heating in microwave; BM= Boiling method. MP= Mechanical procedure In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table - 70: Means comparison for yield of pectin from sapodilla fruit peel after using 0.1N HCl (Mechanical procedure pH

boiling method interaction mean±SE)

BM pH Mechanical procedure BM pH

Homogenizing Grinding Cutting Chopping Hammering interaction mean

B 1 0.50 ± 0.02qr 0.55 ± 0.01qr 0.80 ± 0.02m-q 2.70 ± 0.05de 1.50 ± 0.04ij 1.21 ± 0.22F

135

B 3 0.75 ± 0.02n-q 4.00 ± 0.10b 1.10 ± 0.03klm 3.90 ± 0.10b 0.60 ± 0.03pqr 2.07 ± 0.41B

B 5 1.20 ± 0.03jkl 2.45 ± 0.04ef 0.40 ± 0.01r 3.95 ± 0.11b 0.80 ± 0.04m-q 1.76 ± 0.35D

B 6 1.50 ± 0.02ij 2.00 ± 0.05gh 1.00 ± 0.03l-o 2.30 ± 0.05fg 0.80 ± 0.03m-q 1.52 ± 0.15E

B 7 0.50 ± 0.01qr 1.95 ± 0.08h 0.70 ± 0.01o-r 3.95 ± 0.10b 0.50 ± 0.01qr 1.52 ± 0.36E

M 1 1.05 ± 0.03lmn 0.80 ± 0.01m-q 0.90 ± 0.02l-p 4.70 ± 0.10a 2.05 ± 0.03gh 1.90 ± 0.39C

M 3 1.60 ± 0.02i 3.50 ± 0.10c 1.50 ± 0.02ij 3.45 ± 0.08c 1.55 ± 0.05i 2.32 ± 0.25A

M 5 1.50 ± 0.03ij 2.90 ± 0.05d 1.00 ± 0.03l-o 4.05 ± 0.15b 1.95 ± 0.03h 2.28 ± 0.29A

M 6 2.00 ± 0.03gh 2.00 ± 0.08gh 1.20 ± 0.02jkl 2.45 ± 0.08ef 1.40 ± 0.05ijk 1.81 ± 0.12CD

M 7 0.50 ± 0.01qr 2.30 ± 0.08fg 1.00 ± 0.01l-o 2.40 ± 0.05ef 0.00 ± 0.00s 1.24 ± 0.26F

B= Boiling on burner , M= heating in microwave; BM= Boiling method. MP= Mechanical procedure

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison

among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

136

Table - 71:Means comparison for yield of pectin from sapodilla fruit peel after using 0.5N HCl (Mechanical procedure pH

interaction mean±SE)

pH level Mechanical procedure Mean

Homogenizing Grinding Cutting Chopping Hammering

1 4.55 ± 0.19b 2.45 ± 0.96fg 3.93 ± 0.13c 4.53 ± 0.08b 0.83 ± 0.08l 3.26 ± 0.32A

3 4.08 ± 0.39c 1.48 ± 0.62i 3.13 ± 0.39e 4.88 ± 0.08a 1.68 ± 0.73i 3.05 ± 0.32B

5 2.35 ± 0.99g 0.55 ± 0.20m 4.05 ± 0.14c 2.45 ± 0.10fg 1.25 ± 0.43j 2.13 ± 0.30C

6 1.05 ± 0.34jk 3.60 ± 0.21d 2.65 ± 0.94f 1.08 ± 0.19jk 1.05 ± 0.27jk 1.89 ± 0.28D

7 2.10 ± 0.67h 2.65 ± 0.07f 0.88 ± 0.17kl 0.55 ± 0.20m 0.15 ± 0.05n 1.27 ± 0.22E

Mean 2.83 ± 0.34B 2.15 ± 0.29D 2.93 ± 0.29A 2.70 ± 0.33C 0.99 ± 0.19E

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

Table - 72: Means comparison for yield of pectin from sapodilla fruit peel after using 0.5N HCl (Mechanical procedure

boiling method interaction mean±SE)

Boiling Mechanical procedure Mean

Method Homogenizing Grinding Cutting Chopping Hammering

B 2.58 ± 0.39c 1.23 ± 0.35e 2.76 ± 0.41b 2.79 ± 0.41b 0.34 ± 0.05f 1.94 ± 0.19B

M 3.07 ± 0.57a 3.06 ± 0.33a 3.09 ± 0.41a 2.60 ± 0.53c 1.64 ± 0.29d 2.69 ± 0.20A B= Boiling on burner , M= heating in microwave; BM= Boiling method. MP= Mechanical procedure In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

Table - 73: Means comparison for yield of pectin from sapodilla fruit peel after using 0.5N HCl (Mechanical procedure pH

boiling method interaction mean±SE)

BM pH Mechanical procedure BM pH

Homogenizing Grinding Cutting Chopping Hammering interaction mean

B 1 4.15 ± 0.09def 0.30 ± 0.02s-v 3.65 ± 0.07h 4.40 ± 0.05cd 0.65 ± 0.03r 2.63 ± 0.48D

B 3 3.20 ± 0.02j 0.10 ± 0.01v 4.00 ± 0.03fg 4.80 ± 0.10ab 0.05 ± 0.01v 2.43 ± 0.53E

B 5 0.15 ± 0.00uv 0.10 ± 0.01v 4.35 ± 0.10cde 2.25 ± 0.05n 0.30 ± 0.01s-v 1.43 ± 0.45H

B 6 1.80 ± 0.02o 3.15 ± 0.07jk 0.55 ± 0.00rst 1.50 ± 0.04op 0.45 ± 0.02r-u 1.49 ± 0.26H

137

B 7 3.60 ± 0.08hi 2.50 ± 0.05mn 1.25 ± 0.03pq 1.00 ± 0.02q 0.25 ± 0.02tuv 1.72 ± 0.32G

M 1 4.95 ± 0.12a 4.60 ± 0.05bc 4.20 ± 0.05def 4.65 ± 0.11abc 1.00 ± 0.03q 3.88 ± 0.39A

M 3 4.95 ± 0.05a 2.85 ± 0.06kl 2.25 ± 0.03n 4.95 ± 0.13a 3.30 ± 0.11ij 3.66 ± 0.30B

M 5 4.55 ± 0.12bc 1.00 ± 0.02q 3.75 ± 0.05gh 2.65 ± 0.08lm 2.20 ± 0.08n 2.83 ± 0.33C

M 6 0.30 ± 0.01s-v 4.05 ± 0.11efg 4.75 ± 0.05ab 0.65 ± 0.03r 1.65 ± 0.03o 2.28 ± 0.48F

M 7 0.60 ± 0.02rs 2.80 ± 0.05lm 0.50 ± 0.01rst 0.10 ± 0.01v 0.05 ± 0.01v 0.81 ± 0.27I B= Boiling on burner , M= heating in microwave; BM= Boiling method. MP= Mechanical procedure

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison

among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

138

Table - 74: Means comparison for yield of pectin from sapodilla fruit peel after using 1N HCl (Mechanical procedure pH

interaction mean±SE)

pH level Mechanical procedure Mean

Homogenizing Grinding Cutting Chopping Hammering

1 3.25 ± 0.76ghi 3.63 ± 0.24fg 0.05 ± 0.00n 4.25 ± 0.56cd 2.90 ± 0.12hij 2.82 ± 0.33B

3 3.88 ± 0.66def 5.20 ± 0.19b 0.83 ± 0.35lm 4.55 ± 0.39c 6.10 ± 0.23a 4.11 ± 0.37A

5 3.28 ± 0.71gh 3.58 ± 1.13fg 4.13 ± 0.53cde 5.40 ± 0.59b 3.45 ± 0.52fg 3.97 ± 0.34A

6 1.05 ± 0.20l 2.50 ± 0.47j 1.83 ± 0.75k 3.78 ± 0.04ef 2.55 ± 0.19j 2.34 ± 0.24C

7 1.88 ± 0.30k 1.55 ± 0.54k 0.63 ± 0.26lm 0.48 ± 0.35mn 2.80 ± 0.14ij 1.47 ± 0.21D

Mean 2.67 ± 0.31C 3.29 ± 0.34B 1.49 ± 0.33D 3.69 ± 0.36A 3.56 ± 0.27A In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

Table - 75: Means comparison for yield of pectin from sapodilla fruit peel after using 1N HCl (Mechanical procedure

boiling method interaction mean±SE)

Boiling Mechanical procedure Mean

Method Homogenizing Grinding Cutting Chopping Hammering

B 1.49 ± 0.16f 2.31 ± 0.50e 0.65 ± 0.31g 2.92 ± 0.40d 3.05 ± 0.35d 2.08 ± 0.19B

M 3.84 ± 0.41c 4.27 ± 0.31ab 2.33 ± 0.50e 4.46 ± 0.55a 4.07 ± 0.37bc 3.79 ± 0.21A

B= Boiling on burner , M= heating in microwave; BM= Boiling method. MP= Mechanical procedure In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

Table - 76: Means comparison for yield of pectin from sapodilla fruit peel after using 1N HCl (Mechanical procedure pH

boiling method interaction mean±SE)

BM pH Mechanical procedure BM pH

Homogenizing Grinding Cutting Chopping Hammering interaction mean

B 1 1.55 ± 0.05rst 3.10 ± 0.03i-m 0.05 ± 0.01w 3.00 ± 0.08j-n 2.65 ± 0.05l-o 2.07 ± 0.31F

139

B 3 2.40 ± 0.02m-p 5.60 ± 0.13bc 0.05 ± 0.01w 3.70 ± 0.10hij 5.65 ± 0.15bc 3.48 ± 0.56C

B 5 1.70 ± 0.03p-s 1.05 ± 0.02s-v 2.95 ± 0.04k-n 4.10 ± 0.09gh 2.30 ± 0.03n-q 2.42 ± 0.28E

B 6 0.60 ± 0.01uvw 1.45 ± 0.03rst 0.15 ± 0.01w 3.75 ± 0.05hi 2.15 ± 0.07o-r 1.62 ± 0.34G

B 7 1.20 ± 0.01stu 0.35 ± 0.01vw 0.05 ± 0.00w 0.05 ± 0.01w 2.50 ± 0.10l-o 0.83 ± 0.25H

M 1 4.95 ± 0.12cde 4.15 ± 0.10fgh 0.05 ± 0.00w 5.50 ± 0.10bcd 3.15 ± 0.08i-l 3.56 ± 0.52C

M 3 5.35 ± 0.12cd 4.80 ± 0.08d-g 1.60 ± 0.05q-t 5.40 ± 0.10bcd 6.55 ± 0.20a 4.74 ± 0.45B

M 5 4.85 ± 0.13def 6.10 ± 0.12ab 5.30 ± 0.10cde 6.70 ± 0.18a 4.60 ± 0.12efg 5.51 ± 0.22A

M 6 1.50 ± 0.03rst 3.55 ± 0.09h-k 3.50 ± 0.09h-k 3.80 ± 0.08hi 2.95 ± 0.10k-n 3.06 ± 0.22D

M 7 2.55 ± 0.09l-o 2.75 ± 0.05l-o 1.20 ± 0.03stu 0.92 ± 0.66tuv 3.10 ± 0.06i-m 2.10 ± 0.26F

B= Boiling on burner , M= heating in microwave; BM= Boiling method. MP= Mechanical procedure In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

140

Table - 77: Identification tests for the presence of pectin from three selected fruits

Test Name Banana Sapodilla Muskmelon

Stiff gel test - + -

Test with 95% Ethanol + + Not clear

Test with Potassium Hydroxide (KOH) + + +

Iodine test + + +

Table - 78: Biochemical Characterization of the purified pectin extracted from

sapodilla pectin at different pH

Test Name Sapodilla

( pH5)

Sapodilla

(pH3)

Sapodilla

(pH1)

FGP*

Quantitative test for ammonia. No

ammonia

No

ammonia

No

ammonia

No ammonia

Moisture (%) 5.29 6.02 5.52 7.02

Ash (%) 5.11 4.3 4.89 1.16

Equivalent weight 1700 2680 2290 1271

Methoxyl Content (%) 5.1 4.4 4.9 8.16

Anhydrouronic Acid. (%) 39.33 31.78 34.56 70.50

Jelly Grade 100 99 98 150

Galacturonic Acid Content (%) 77.7 65.8 60.5 76.19

Protien 1.71 3.36 4.04

Degree of esterificatoion (%) 73.63 72.1 70.2 76.41

*FGP = Food grade pectin

Table - 79: Water holding, water binding and fat binding capacity of sapodilla

pectin

141

Test Name Sapodilla peel

pectin

Apple

pectin

Orange

pectin

Fibers

Apple Orange Grape

Fruit

WBC g/g 7.6 - - 1.62 1.65 2.09

WHC g/g 6.99 16.51 28.07 - - -

FBC g/g 2.2 - - 0.95 1.81 1.52

Table – 80: Showing FTIR spectral values of sample and standard pectin along with

associated functional groups.

Functional groups FOOD

GRADE

Sapodilla

(pH5)

Sapodilla

(pH3)

Sapodilla

(pH1)

O-H stretching 3319.2 3287.3 3271.1 3289.2

C-H stretching , symmetric,

asymmetric 2932.4 2923.6 2929.6 2930.2

C=O esterified 1733.3 1735.6 1738.6 1737.8

COO- asymmetric stretching

1622.2 1611.2 1612.2

COO- symmetric stretching 1426.4 1424.3 1423.3 1425.8

C-H bending 1335.4 1362.0 1363.0

C=O stretching 1234.9 1234.1 1234.5 1228.6

142

Figure – 23: FTIR spectra of food grade pectin

Figure – 24: FTIR spectra of sapodilla peel pectin extracted at pH 5

Figure – 25: FTIR spectra of sapodilla peel pectin extracted at pH 3

403.

3

467.

1

523.

2

679.

9

858.

2

909.

2

995.

2

1052

.6

1234

.9

1273

.913

35.4

1426

.4

1733

.3

1996

.0

2068

.9

2349

.0

2932

.4

3319

.2

3554

.1

*Pectin f ood grade

62

64

66

68

70

72

74

76

78

80

82

84

86

88

90

92

94

96

98

100

102

%T

500 1000 1500 2000 2500 3000 3500 4000

Wavenumbers (cm-1)

143

Figure – 26: FTIR spectra of sapodilla peel pectin extracted at pH 1

DLS studies

Table – 81: Summary the physical characterization of standard pectin and the

different pectin extracted from various source and at different pH by DLS.

144

Sample Extraction at

pH

Time of

extraction RH (nm)

PD (%)

(Đ)

Standard -- -- 73.7-1979.92 8.6-37.9

Apple 5.0 10 94.37-1357 18.4-47.8

Orange 5.0 10 473.07-482.92 36.8-36.9

Sapodilla 3.0 10 464.37-548.53 29.3-30.1

Sapodilla 5.0 10 390.21-421.17 28.1-29.3

RH= Radius of hydration ; PD = polydispersity

145

Standard

Figure – 27:.Physical characterization of standard pecti.by DLS. (A) DLS results of

sstandard pectin.illustrating the experimental conditions i.e., the mean autocorrelation

function (a), monodispersity and radius plot (b-c), respectively. (B) Comparative

corresponding radius distribution of pectin standard (a) and standardpectin (b). All

experiments were performed with an auto–piloted run of 50 measurements at every 20 s,

with a wait time 1 s (at 25 °C).

Figure- 28:.Physical characterization of apple pectin extracted at pH5.0. by DLS. (A)

DLS results of apple pectin extracted at pH5.0. illustrating the experimental conditions

i.e., the mean autocorrelation function (a), monodispersity and radius plot (b-c),

respectively. (B) Comparative corresponding radius distribution of pectin standard (a)

and apple pectin extracted at pH5.0 (b). All experiments were performed with an auto–

piloted run of 50 measurements at every 20 s, with a wait time 1 s (at 25 °C).

146

Figure – 29: Physical characterization of Orange pectin extracted at pH5.0. by DLS. (A)

DLS results of Orange pectin extracted at pH5.0. illustrating the experimental conditions

i.e., the mean autocorrelation function (a), monodispersity and radius plot (b-c),

respectively. (B) Comparative corresponding radius distribution of pectin standard (a)

and Orange pectin extracted at pH5.0 (b). All experiments were performed with an auto–

piloted run of 50 measurements at every 20 s, with a wait time 1 s (at 25 °C).

147

Figure – 30: Physical characterization of sapodilla pectin extracted at pH3.0. by DLS.

(A) DLS results of sapodilla pectin extracted at pH3.0. illustrating the experimental

conditions i.e., the mean autocorrelation function (a), monodispersity and radius plot (b-

c), respectively. (B) Comparative corresponding radius distribution of pectin standard (a)

and sapodilla pectin extracted at pH3.0 (b). All experiments were performed with an

auto–piloted run of 50 measurements at every 20 s, with a wait time 1 s (at 25 °C).

Figure – 31: Physical characterization of sapodilla pectin extracted at pH5.0. by DLS.

(A) DLS results of sapodilla pectin extracted at pH5.0. illustrating the experimental

conditions i.e., the mean autocorrelation function (a), monodispersity and radius plot (b-

c), respectively. (B) Comparative corresponding radius distribution of pectin standard (a)

and sapodilla pectin extracted at pH5.0 (b). All experiments were performed with an

148

auto–piloted run of 50 measurements at every 20 s, with a wait time 1 s (at 25 °C). Also

see Table 5 for details.

Response surface methodology

Table - 82: Box-Bechen experimental design and levels of factors used for

optimization of pectin yield

Variables Symbol Low High

pH X1 1 5

Temperature(oC) X2 50 100

Time ( minutes) X3 10 90

Table - 83: Box-Behnken experimental design and corresponding results for

responses

Std Order Run Order Pt Type Blocks pH Temperature Time Yield

10 1 2 1 3 100 10 1.6

3 2 2 1 1 100 50 1.7

14 3 0 1 3 75 50 1.5

1 4 2 1 1 50 50 1.6

2 5 2 1 5 50 50 3.5

6 6 2 1 5 75 10 2.6

9 7 2 1 3 50 10 1.5

12 8 2 1 3 100 90 1.65

4 9 2 1 5 100 50 2.5

8 10 2 1 5 75 90 3.5

11 11 2 1 3 50 90 1.4

13 12 0 1 3 75 50 2

5 13 2 1 1 75 10 3

15 14 0 1 3 75 50 1.8

149

7 15 2 1 1 75 90 1.5

Table - 84: Analysis of Variance for Yield ( Response surface methodology)

Source DF Adj SS Adj MS F-value Prob.

Model

Linear

pH

Temperature

Time

Square

pH*pH

Temperature*Temperature

Time*Time

2-Way Interaction

pH*Temperature

pH*Time

Temperature*Time

Error

Lack-of-Fit

Pure Error

Total

9

3

1

1

1

3

1

1

1

3

1

1

1

5

3

2

14

7.15996

2.40188

2.31125

0.03781

0.05281

3.00996

2.57694

0.28348

0.00848

1.74813

0.30250

1.44000

0.00563

0.51104

0.38438

0.12667

7.67100

0.79555

0.80063

2.31125

0.03781

0.05281

1.00332

2.57694

0.28348

0.00848

0.58271

0.30250

1.44000

0.00563

0.10221

0.12813

0.06333

7.78*

7.83*

22.61**

0.37

0.52

9.82*

25.21**

2.77

0.08

5.70*

2.96

14.09*

0.06

2.02

0.018

0.025

0.005

0.570

0.504

0.015

0.004

0.157

0.785

0.045

0.146

0.013

0.824

0.348

* = Significant (P<0.05); ** = Highly significant (P<0.01)

S = 0.3197

R² = 93.34%

R²(adj) = 81.35%

R²(pred) = 16.11%

150

Table - 85: Estimated Regression Coefficients for Yield (Box-Bechen experimental

design, response surface methodology)

Term Effect Coef. SE(Coef.) t-value Prob. VIF

Constant

pH

Temperature

Time

pH*pH

Temperature*Temperature

Time*Time

pH*Temperature

pH*Time

Temperature*Time

1.075

-0.137

-0.163

1.671

-0.554

0.096

-0.550

1.200

0.075

1.767

0.538

-0.069

-0.081

0.835

-0.277

0.048

-0.275

0.600

0.038

0.185

0.113

0.113

0.113

0.166

0.166

0.166

0.160

0.160

0.160

9.57**

4.76**

-0.61

-0.72

5.02**

-1.67

0.29

-1.72

3.75*

0.23

0.000

0.005

0.570

0.504

0.004

0.157

0.785

0.146

0.013

0.824

1.00

1.00

1.00

1.01

1.01

1.01

1.00

1.00

1.00

* = Significant (P<0.05); ** = Highly significant (P<0.01)

Yield = 0.76 - 0.947 pH + 0.0784 Temperature - 0.0303 Time + 0.2089 pH² -

0.000443 Temperature² + 0.000030 Time² - 0.00550 pH*Temperature

+ 0.00750 pH*Time + 0.000037 Temperature*Time

Table - 86: Predicted values of yield. (Box-Bechen experimental design, response

surface methodology)

Yield Composite

Solution pH Temperature Time Fit Desirability

1 5 61.1111 90 3.79087 1.00000

2 5 85.2577 86.8718 3.48497 0.99284

3 5 67.7177 17.6639 2.83699 0.68428

151

4 5 67.7177 17.6639 2.83699 0.68428

5 1 90.6592 10 2.79074 0.66226

Figure – 32: Showing the optimal conditions for the extraction of pectin from

sapodilla fruit peel (Box-Bechen experimental design, response surface

methodology)

152

Figure – 33: Response surface graph and contour plot of effect of pH and

temperature on yield of pectin at constant time

Yield = -0.6969-0.5663*x+0.0811*y+0.2079*x*x-0.0055*x*y-0.0004*y*y

> 3.5 < 3.5 < 3 < 2.5 < 2 < 1.5

Yield = Distance Weighted Least Squares

> 4 < 4 < 3.5 < 3 < 2.5 < 2 < 1.5 < 1 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 4.5 5.0 5.5

pH

40

50

60

70

80

90

100

110

Tem

pe

ratu

re

153

Figure- 34: Response surface graph and contour plot of effect of pH and time on

yield of pectin at constant temperature

Yield = 4.0523-1.3913*x-0.0289*y+0.2142*x*x+0.0075*x*y+4.3269E-5*y*y

> 4.5 < 4.5 < 4 < 3.5 < 3 < 2.5 < 2 < 1.5

Yield = Distance Weighted Least Squares

> 4 < 4 < 3.5 < 3 < 2.5 < 2 < 1.5 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 4.5 5.0 5.5

pH

0

10

20

30

40

50

60

70

80

90

100

Tim

e

154

Figure – 35: Response surface graph and contour plot of effect of temperature and

time on yield of pectin at constant pH

Yield = -0.3684+0.0773*x-0.0038*y-0.0005*x*x+3.75E-5*x*y-1.0216E-5*y*y

> 2.2 < 2.2 < 2 < 1.8 < 1.6

Yield = Distance Weighted Least Squares

> 2.5 < 2.5 < 2 < 1.5 < 1 < 0.5 40 50 60 70 80 90 100 110

Temperature

0

10

20

30

40

50

60

70

80

90

100

Tim

e

155

Table – 87: Flow properties of granules made for paracetamol tablets

Test

Formulatio

n

Mass

(g)

Bulk

Volume

(ml)

Tapped

Volume

(ml)

Bulk

Density

(g/ml)

Tapped

Density

(g/ml)

Angle of

Repose

(θ⁻¹)

Compressibilit

y Index

(%)

Hausner

Ratio

-

Standard 2.004±0.00

1

4.5068±0.11

5

4.123±0.12

5

0.446±0.05

1

0.487±0.05

7

17.955±1.15

8 8.41±0.507

1.091±0.00

6

F1 2.010±0.00

5 3.867±0.115

3.067±0.11

5

0.520±0.01

4

0.656±0.02

3

18.747±0.54

3 20.702±0.608

1.261±0.01

0

F2 2.003±0.00

1 3.867±0.115

3.067±0.11

5

0.518±0.01

5

0.654±0.02

4

16.494±1.68

0 20.702±0.608

1.261±0.01

0

F3 2.008±0.00

1 4.067±0.115

3.067±0.11

5

0.494±0.01

4

0.655±0.02

4

16.558±1.79

2 24.603±0.687

1.326±0.01

2

F4 2.008±0.00

1 4.333±0.115

4.133±0.11

5

0.464±0.01

2

0.486±0.01

4

17.545±1.54

0 4.618±0.125

1.048±0.00

1

F5 2.004±0.00

1 3.533±0.306

3.333±0.30

6

0.570±0.05

1

0.605±0.05

7

17.955±1.15

8 5.690±0.507

1.060±0.00

6

F6 2.005±0.00

1 4.133±0.115

3.667±0.46

2

0.485±0.01

4

0.552±0.06

5

17.453±0.17

1 11.032±13.884

1.141±0.16

3

F7 2.016±0.02

1 4.000±0.200

3.333±0.30

6

0.505±0.02

1

0.608±0.05

6

17.987±0.61

8 16.700±5.888

1.204±0.08

2

F8 2.020±0.01 4.067±0.115 3.067±0.11 0.497±0.01 0.659±0.02 21.546±0.44 24.603±0.687 1.326±0.01

156

0 5 4 5 1 2

F9 2.017±0.00

6 4.133±0.115

3.333±0.11

5

0.488±0.01

2

0.605±0.02

0

16.911±0.38

3 19.365±0.550

1.240±0.00

8

F1-F9 = test formulations 1 till 9 Each value is a Mean±SD of three determination

Table - 88: Pharmaceutical characteristics of compressed formulation of paracetamol tablet

Test Formulation

Pharmacopoeial Limits

(USP 32/NF 27)

Wt. Variation

(Mean ± S.D)

(mg)

±5%

Thickness

(Mean ±

S.D)

(mm)

±5%

Diameter

(Mean ± S.D)

(mm)

₋

Hardness

(Mean ± S.D)

(kg)

At least 5 kg

Loss on drying

%

Not more than 1.5%

Standard 702.67± 2.52 5.60±0.20 9.47±0.05 5.22±0.01 4.0%

F1 705.00±5.00 5.22±0.03 9.42±0.03 2.54±0.06 4.0%

F2 672.33±2.52 5.24±0.05 9.39±0.01 1.37±0.07 3.0%

F3 701.33±3.21 5.32±0.05 9.45±0.04 4.16±0.08 4.3%

F4 695±5.00 5.35±0.06 9.44±0.05 5.06±0.21 4.2%

F5 705±4.51 5.21±0.03 9.41±0.03 5.15±0.07 4.6%

F6 701.67±3.97 5.19±0.04 9.42±0.03 5.21±0.07 4.3%

F7 708.3±2.89 5.29±0.04 9.31±0.03 6.07±0.12 4.2%

F8 694.67±4.51 5.27±0.13 9.27±0.06 6.67±0.58 4.5%

157

F9 704.00±3.61 5.27±0.10 9.42±0.04 7.40±0.33 4.5%

F1-F9 = test formulations 1 till 9 Each value is a Mean±SD of three determination.

Table – 89: Dissolution studies of paracetamol tablet

FORMULATION NUMBER

F1 F2 F3 F4 F5 F6 F7 F8 F9

243nm

(Abs)

%

release

243nm

(Abs)

%

release

243nm

(Abs)

%

release

243nm

(Abs)

%

release

243nm

(Abs)

%

release

243nm

(Abs)

%

release

243nm

(Abs)

%

release

243nm

(Abs)

%

release

243nm

(Abs)

%

release

0.304 43.43 0.339 50.64 0.401 58.77 0.269 71.52 0.564 84.17 0.078 23.00 0.084 26.77 0.123 36.73 0.111 32.73

0.291 41.67 0.338 50.49 0.412 60.39 0.283 70.28 0.557 83.12 0.087 25.65 0.080 23.59 0.111 32.73 0.094 27.97

0.362 51.83 0.564 84.25 0.382 55.99 0.331 83.13 0.418 91.93 0.112 33.03 0.093 27.42 0.108 31.85 0.108 31.85

0.372 53.26 0.557 83.20 0.389 57.02 0.333 87.28 0.620 92.53 0.105 30.96 0.096 28.31 0.120 35.39 0.101 29.78

0.313 44.82 0.418 62.44 0.577 84.58 0.285 85.20 0.406 93.13 0.096 28.313 0.076 22.41 0.120 35.39 0.099 29.19

0.323 46.24 0.406 60.65 0.590 86.48 0.312 85.20 0.632 94.32 0.116 34.21 0.091 26.83 0.107 31.55 0.088 25.95

STANDARD FOR FORMULATION

0.692 99.10 0.664 99.10 0.676 99.10 0.478 99.10 0.664 99.10 0.336 99.10 0.336 99.10 0.336 99.10 0.336 99.10

F1-F9 are test formulation while Abs = absorbance and %release = percent release

158

159

F1 F2

F3

Figure -36: Formulated paracetamol tablest ( F1, F2, F3)

160

F4 F5

F6

Figure 37: Formulated paracetamol tablest ( F4, F5, F6)

161

F7 F8

F9

Figure 38: Formulated paracetamol tablest (F7, F8, F9)

162

Table - 90: Flow properties ofgranules for ibuprofen tablet

Test

Formulation

Mass

Bulk

Volume

Tapped

Volume

Bulk

Density

Tapped

Density

Angle of

Repose

Compressibility

Index

Hausner

Ratio

(g) (ml) (ml) (g/ml) (g/ml) (θ⁻¹) (%) ₋

Std 2.02±0.013 4.25±0.015 3.75±0.092 0.475±0.002 0.538±0.020 11.71±0.210 11.71±0.116 1.132±0.061

R1 2.02±0.013 4.20±0.015 3.76±0.093 0.49±0.003 0.55±0.019 15.23±0.208 4.82±0.115 1.13±0.061

R2 2.01±0.016 4.35±0.042 3.61±0.010 0.46±0.005 0.56±0.004 13.83±0.153 9.74±0.344 1.24±0.059

R3 2.01±0.007 4.60±0.080 3.93±0.064 0.42±0.021 0.52±0.010 13.73±0.306 7.40±0.352 1.17±0.028

R4 2.01±0.004 4.51±0.090 4.15±0.050 0.44±0.002 0.47±0.003 11.13±0.153 3.77±0.252 1.09±0.009

R1= Test formulation one, R2= Test formulation two, R3= Test formulation three, R4= Test formulation four. Each value is a Mean±SD of three determination

Table - 91: Pharmaceutical characteristics of compressed formulation of ibuprofen tablet

Test Formulation

Pharmacopoeial Limits

(USP 32/NF 27)

Wt. Variation

(Mean ± S.D)

(mg) ±5%

Thickness

(Mean ± S.D)

(mm) ±5%

Length x width

(Mean ± S.D)

(mm) ₋

Hardness

(Mean ± S.D)

(kg) At least 5 kg

Loss on

Drying %

Standard 833.33±2.08 6.33± 0.06 20 x 9.5 6.13± 0.14 4.5%

R1 843.33±2.08 6.23± 0.06 20 x 9.5 6.23± 0.14 4.9%

R2 848.33±2.89 6.37±0.06 20 x 9.5 8.40±0.40 4.8%

R3 847.67±2.52 6.22±0.03 20 x 9.5 9.23±0.14 4.9%

R4 842.67±2.52 6.27±0.06 20 x 9.5 10.43±0.18 4.7%

163

Table- 92: dissolution studies of ibuprufen tablets

FORMULATION NUMBER

R1 R2 R3 R4

243nm (Abs) % release 243nm (Abs) % release 243nm (Abs) % release 243nm (Abs) % release

0.405 89.6 0.166 36.72 0.092 33.45 -0.018 -6.54

0.415 91.81 0.141 31.190 0.090 32.72 -0.017 -6.182

0.420 92.91 0.147 32.51 0.112 40.72 -0.020 -7.27

0.425 94.02 0.154 34.06 0.113 41.92 0.008 -2.90

0.429 94.95 0.143 31.62 0.030 10.90 -0.008 -2.90

0.430 95.13 0.161 35.60 0.036 13.091 -0.008 -2.90

STANDARD FOR FORMULATION

0.503 111.33 0.503 111.33 0.306 111.33 0.306 111.33

R1-R4 are test formulation while Abs = absorbance and %release = percent release

164

R1 R2

R3 R4

Figure – 39: Formulated ipubrufen tablets ( R1, R2, R3, R4)

91

Table -93: Basic evaluation test of antidiarrheal formulation prepared from

sapodilla pectin

Parameters Suspension made from

sapodilla pectin Comparative suspension *

Color Pinkish white White

Odor vanilla Vanilla

Taste sweet sweet

pH 6.1 5.56

Viscosity 14.14 13.13

Sedimentation rate 0.3 0.1

Redispersity +++ +++

WHC 32.69 33.56

+ denotes the number of times the cylinder was moved. * Keptin antidiarrheal preparation

92

Figure- 40 Formulated antidiarrheal preparation

93

Table - 94: Effect of different concentration of sapodilla pectin on the chemical

properties of the jam samples

Sample PH

TSS

(mg)

TTA

(ml)

MC

(%)

Vit C

(mg)

Ash

(%)

TS

(mg)

Viscosity

(Cp)

S 3.06 61.9 1.14 35.9 18.3 1.78 64.1 71

F1 3.16 56.7 1.08 41 17.6 1.69 59 56

F2 3.25 62.6 1.01 35.7 18.1 1.73 64.3 63

F3 3.25 62.4 1.03 35.1 17.9 1.71 64.9 67

WhereS = Jam with standard pectin, F1 = Jam with 2g pectin, F2 = Jam with pectin in same concentration

as in standard, F3 = Jam with 10g pectin while MC is moisture content . TSS total soluble solids, TTA

total titratable acid and TS is total solid.

Table - 95: Scores for sensory parameters as judged by twenty (20) panelists.

Sample Appearance Taste Aroma Spreadability Texture Mouth

feel

Overall

acceptability

S 7.7 7.9 7.9 8.2 7.8 7.5 7.8

F1 7.5 7.3 7.2 7.3 7.2 7 7.2

F2 7.5 7.6 7.7 7.8 7.8 7.5 7.6

F3 6.2 6.4 6.2 5.6 6.3 5.5 6

WhereS = Jam with standard pectin, F1 = Jam with 2g pectin, F2 = Jam with pectin in same concentration

as in standard, F3 = Jam with 10g pectin

(S)=Jam made from 5g standard pectin (F1) Jam made from 5g pectin

94

(F2) Jam made from 7 g pectin (F3) Jam made from 10g pectin

Figure -41 formulation of Jam made from extracted pectin

Figure -42: formulation of pudding made from extracted pectin.

95

Table - 9: List of fruits used in the screening study for the presence of pectin

(a) is Munarin et al., 2012; (b) is Bhat and Singh, 2014; (c) is Baker, 1997; (d) is Baissise

et al., 2010. (e) is Rasheed, 2008; (f) is Aina et al., 2012.

S no

Fruit

%Yield Reported

amount Common name Parts

used

1 Orange peel 20 6-26% a

2 Apple exocarp 6 2-19%a

3 Grape Fruit peel 3.5 13-32%a

4 Sapodilla peel 5.5 Notfound

5 Muskmelon peel 2 Not found

6 Guava (unripe) exocarp 0 Not found

7 Guava(ripe) exocarp 4 16.8% b

8 Chinese apple (Jungle Fruit) exocarp 0 Notfound

9 Strawberry exocarp 0 0.35–0.44%.C

10 Sweet lime peel 3 6-26%a

11 Peach exocarp 0 4-18%a

12 Apricot exocarp 0 4.97%d

13 Canteloup peel 3 Not found

14 Plum exocarp 0 Not found

15 Watermelon peel 2.35 15.70% e

16 Mango(Unripe) peel 0 Not found

17 Mango (ripe) peel 3.5 9-29%a

18 Banana peel 3.5 2-15%a

19 Lemon peel 3.5 2.7% f

96

Table - 10: Showing percent yield of pectin from five selected fruits

Mechanical

procedure

p

H

Sapodilla Banana Muskmelo

n Apple Orange

B M B M B M B M B M

Homogenizin

g

1 0.5 1.0

5

3.2

5 5.6 0.55 0.55 0.5 1

16.1

5

15.2

5

3 0.7

5 1.6

6.8

5

4.3

5 2 1

1.9

5

2.7

5 18.2 19.8

5 1.2 1.5 8.8

5 8.4 1.45 2.3

2.3

5 3.7 19.4 22.7

6 1.5 2 5.8

5

9.7

5 1.15 1 1.5 1.4

13.9

5 2.85

7 0.5 0.5 5.4

5 3.1 0.55 1.8

0.0

5

0.3

5 21.7 7.5

Grinding

1 0.5

5 0.8

2.4

5 3.2 2.25 2.65

2.4

5 2.5 2.6 3.95

3 4 3.5 4.2 6.9 1.4 1.8 0.4

5

1.1

5 7.8 5.3

5 2.4

5 2.9 3.5 4.5 1.95 2.4 1 1.1 2.3 7.1

6 2 2 3.6

5

2.6

5 0.95 1.6 2.4

2.8

5 6.45 6.6

7 1.9

5 2.3

1.2

5

0.2

5 0.55 1

0.0

5

0.5

5 0.6 0.2

Cutting

1 0.8 0.9 0 2.5 0.5 0.5 2.8

5 3.9 8.1 7

3 1.1 1.5 1.9

5 1.2 2.45 2.55

2.9

5 4.5 8.5 12

5 0.4 1 5 5.5 1 1 2.7 3.7 11.8

5

20.6

5

6 1 1.2 1 5.4 0.7 0.95 1.1

5 2.9 6.95 10.2

7 0.7 1 3 6.5 1.3 2.2 0.0

5 0.7 4 7.4

Chopping

1 2.7 4.7 1.6

5 2.1 0.05 0.7

2.8

5

3.3

5 2.6 3.35

3 3.9 3.4

5 4 3.3 0.05 0.5

1.2

5

2.7

5 7.95

10.0

5

5 3.9

5

4.0

5

5.9

5 6.6 1.7 2.1

0.0

5

1.7

5 10.4 15.8

6 2.3 2.4

5

5.8

5

6.3

5 2.25 1.75 0.1 0.9

15.2

5

17.1

5

7 3.9 2.4 1.8 1.6 0.5 1.05 0.2 0.6 5.35 9.35

97

5 5 5

Hammering

1 1.5 2.0

5 4.5 7 0.65 1.25 1.5 1.7

13.5

5

16.6

5

3 0.6 1.5

5 5.5 5.5 0.5 0.8 2.7 3.5 7.2 8.35

5 0.8 1.9

5 8.5

10.

5 0.65 0.3

3.0

5

4.8

5 7.4

12.1

5

6 0.8 1.4 5 5.6 0.95 0.5 1.2

5

2.0

5 5.55 10.7

7 0.5 0 0.5 0.5 0.45 0.05 1.7 2.0

5 5.5 8.85

B= Boiling on burner, M= heating in microwave

98

Table - 11: Analysis of variance (mean squares) of yield for five selected fruits

Source of variation Degrees of

freedom

Mean squares

Sapodilla Banana Muskmelon Apple Orange

Mech. procedure (MP)

pH level

Boiling method (BM)

MP pH

MP BM

pH BM

MP pH x BM

Error

Total

4

4

1

16

4

4

16

100

149

32.6575**

3.3917**

3.2413**

2.5462**

0.3392**

1.0112**

0.6258**

0.0266

51.245**

84.818**

22.349**

16.774**

4.640**

6.332**

4.698**

0.191

4.3869**

1.5751**

2.0184**

2.7688**

0.2814**

0.4112**

0.4141**

0.0112

9.5920**

17.3200**

22.6981**

5.6967**

0.8120**

0.8975**

0.2720**

0.0314

493.729**

142.586**

59.914**

88.941**

81.797**

44.377**

21.750**

1.0100

NS = Non-significant (P>0.05); * = Significant (P<0.05); ** = Highly significant (P<0.01) , Mech = mechanical

99

Table – 12: Means comparison of yield for sapodilla fruit peel (Mechanical procedure pH interaction mean±SE )

pH level Mechanical procedure Mean

Homogenizing Grinding Cutting Chopping Hammering

1 0.78 ± 0.13ij 0.68 ± 0.06j 0.85 ± 0.03hij 3.70 ± 0.47a 1.78 ± 0.14ef 1.56 ± 0.23C

3 1.18 ± 0.19gh 3.75 ± 0.15a 1.30 ± 0.09g 3.68 ± 0.13a 1.08 ± 0.21ghi 2.20 ± 0.24A

5 1.35 ± 0.08g 2.68 ± 0.11c 0.70 ± 0.14j 4.00 ± 0.12a 1.38 ± 0.26g 2.02 ± 0.23B

6 1.75 ± 0.12f 2.00 ± 0.06ef 1.10 ± 0.05ghi 2.38 ± 0.10cd 1.10 ± 0.14ghi 1.67 ± 0.10C

7 0.50 ± 0.01jk 2.13 ± 0.10de 0.85 ± 0.07hij 3.18 ± 0.35b 0.25 ± 0.11k 1.38 ± 0.22D

Mean 1.11 ± 0.10C 2.25 ± 0.19B 0.96 ± 0.05D 3.39 ± 0.16A 1.12 ± 0.12C

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

Table - 13: Means comparison of yield for sapodilla fruit peel (Mechanical procedure boiling method interaction mean±SE)

Boiling Mechanical procedure Mean

Method Homogenizing Grinding Cutting Chopping Hammering

B 0.89 ± 0.11e 2.19 ± 0.30b 0.80 ± 0.07e 3.36 ± 0.20a 0.84 ± 0.09e 1.62 ± 0.14B

M 1.33 ± 0.14c 2.30 ± 0.25b 1.12 ± 0.06d 3.41 ± 0.25a 1.39 ± 0.20c 1.91 ± 0.13A

B= Boiling on burner , M= heating in microwave

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean.

Table - 14: Means comparison of yield for sapodilla fruit peel (Mechanical procedure pH boiling method

interaction mean ± SE)

BM pH Mechanical procedure BM pH

100

Homogenizing Grinding Cutting Chopping Hammering interaction mean

B 1 0.50 ± 0.03qrs 0.55 ± 0.02pqr 0.80 ± 0.04n-r 2.70 ± 0.14ef 1.50 ± 0.06i-l 1.21 ± 0.22E

B 3 0.75 ± 0.03n-r 4.00 ± 0.15bc 1.10 ± 0.03j-o 3.90 ± 0.14bcd 0.60 ± 0.03o-r 2.07 ± 0.41B

B 5 1.20 ± 0.07j-n 2.45 ± 0.10efg 0.40 ± 0.02rs 3.95 ± 0.22bcd 0.80 ± 0.03n-r 1.76 ± 0.35C

B 6 1.50 ± 0.07i-l 2.00 ± 0.10ghi 1.00 ± 0.04l-q 2.30 ± 0.13fg 0.80 ± 0.03n-r 1.52 ± 0.16D

B 7 0.50 ± 0.02qrs 1.95 ± 0.08ghi 0.70 ± 0.03n-r 3.95 ± 0.14bcd 0.50 ± 0.01qrs 1.52 ± 0.36D

M 1 1.05 ± 0.06k-p 0.80 ± 0.03n-r 0.90 ± 0.03m-r 4.70 ± 0.26a 2.05 ± 0.14gh 1.90 ± 0.40BC

M 3 1.60 ± 0.08hij 3.50 ± 0.16cd 1.50 ± 0.06i-l 3.45 ± 0.14d 1.55 ± 0.05h-k 2.32 ± 0.26A

M 5 1.50 ± 0.04i-l 2.90 ± 0.06e 1.00 ± 0.03l-q 4.05 ± 0.14b 1.95 ± 0.11ghi 2.28 ± 0.29A

M 6 2.00 ± 0.09ghi 2.00 ± 0.08ghi 1.20 ± 0.04j-n 2.45 ± 0.16efg 1.40 ± 0.02j-m 1.81 ± 0.13C

M 7 0.50 ± 0.02qrs 2.30 ± 0.12fg 1.00 ± 0.04l-q 2.40 ± 0.08efg 0.00 ± 0.00s 1.24 ± 0.26E

B= Boiling on burner , M= heating in microwave; BM= boiling method

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

.

101

Table – 15: Means comparison of yield for banana fruit peel (Mechanical procedure pH interaction mean±SE)

pH level Mechanical procedure Mean

Homogenizing Grinding Cutting Chopping Hammering

1 4.43 ± 0.55fgh 2.83 ± 0.19j 1.25 ± 0.56klm 1.88 ± 0.12k 5.75 ± 0.60cd 3.23 ± 0.36D

3 5.60 ± 0.57cde 5.55 ± 0.64cde 1.58 ± 0.17kl 3.65 ± 0.19hij 5.50 ± 0.18cde 4.38 ± 0.34C

5 8.63 ± 0.31ab 4.00 ± 0.25ghi 5.25 ± 0.21def 6.28 ± 0.27c 9.50 ± 0.53a 6.73 ± 0.41A

6 7.80 ± 0.90b 3.15 ± 0.26ij 3.20 ± 0.99ij 6.10 ± 0.22cd 5.30 ± 0.19def 5.11 ± 0.42B

7 4.28 ± 0.54gh 0.75 ± 0.23lm 4.75 ± 0.80efg 1.73 ± 0.07k 0.50 ± 0.01m 2.40 ± 0.38E

Mean 6.15 ± 0.41A 3.26 ± 0.33D 3.21 ± 0.40D 3.93 ± 0.37C 5.31 ± 0.55B

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

Table - 16: Means comparison of yield for banana fruit peel (Mechanical procedure boiling method interaction mean±SE)

Boiling Mechanical procedure Mean

Method Homogenizing Grinding Cutting Chopping Hammering

B 6.05 ± 0.50a 3.01 ± 0.29e 2.19 ± 0.46f 3.86 ± 0.50cd 4.80 ± 0.69b 3.98 ± 0.27B

M 6.24 ± 0.67a 3.50 ± 0.59de 4.22 ± 0.55c 3.99 ± 0.57cd 5.82 ± 0.87a 4.75 ± 0.31A

B= Boiling on burner , M= heating in microwave In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

102

Table - 17: Means comparison of yield for banana fruit peel (Mechanical procedure pH boiling method interaction mean ±

SE)

BM pH Mechanical procedure BM pH

Homogenizing Grinding Cutting Chopping Hammering interaction mean

B 1 3.25 ± 0.19l-q 2.45 ± 0.11o-u 0.00 ± 0.00x 1.65 ± 0.12r-w 4.50 ± 0.16h-l 2.37 ± 0.41D

B 3 6.85 ± 0.24def 4.20 ± 0.19i-m 1.95 ± 0.06q-v 4.00 ± 0.19j-n 5.50 ± 0.27e-i 4.50 ± 0.44C

B 5 8.85 ± 0.51b 3.50 ± 0.09l-p 5.00 ± 0.31g-k 5.95 ± 0.22d-h 8.50 ± 0.29b 6.36 ± 0.56B

B 6 5.85 ± 0.26d-h 3.65 ± 0.24k-o 1.00 ± 0.03u-x 5.85 ± 0.31d-h 5.00 ± 0.21g-k 4.27 ± 0.49C

B 7 5.45 ± 0.30e-j 1.25 ± 0.07t-x 3.00 ± 0.12m-s 1.85 ± 0.08q-v 0.50 ± 0.03vwx 2.41 ± 0.46D

M 1 5.60 ± 0.30d-i 3.20 ± 0.16l-q 2.50 ± 0.14o-t 2.10 ± 0.09p-u 7.00 ± 0.43cd 4.08 ± 0.52C

M 3 4.35 ± 0.12i-m 6.90 ± 0.40de 1.20 ± 0.07t-x 3.30 ± 0.15l-q 5.50 ± 0.31e-i 4.25 ± 0.53C

M 5 8.40 ± 0.43bc 4.50 ± 0.23h-l 5.50 ± 0.26e-i 6.60 ± 0.46def 10.50 ± 0.57a 7.10 ± 0.59A

M 6 9.75 ± 0.46ab 2.65 ± 0.14n-t 5.40 ± 0.27f-j 6.35 ± 0.29d-g 5.60 ± 0.21d-i 5.95 ± 0.62B

M 7 3.10 ± 0.09l-r 0.25 ± 0.01wx 6.50 ± 0.32def 1.60 ± 0.07s-w 0.50 ± 0.02vwx 2.39 ± 0.61D

B= Boiling on burner , M= heating in microwave; BM= boiling method

. In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table -18: Means comparison of yield for muskmelon fruit peel (Mechanical procedure pH interaction mean±SE)

pH level Mechanical procedure Mean

103

Homogenizing Grinding Cutting Chopping Hammering

1 0.55 ± 0.02m-p 2.45 ± 0.12a 0.50 ± 0.01n-q 0.38 ± 0.15pqr 0.95 ± 0.14ijk 0.97 ± 0.15D

3 1.50 ± 0.23fg 1.60 ± 0.10ef 2.50 ± 0.05a 0.28 ± 0.10qr 0.65 ± 0.07l-o 1.31 ± 0.15B

5 1.88 ± 0.20cd 2.18 ± 0.13b 1.00 ± 0.02ij 1.90 ± 0.10cd 0.48 ± 0.08o-r 1.49 ± 0.13A

6 1.08 ± 0.05hi 1.28 ± 0.15gh 0.83 ± 0.06jkl 2.00 ± 0.12bc 0.73 ± 0.10k-n 1.18 ± 0.09C

7 1.18 ± 0.28hi 0.78 ± 0.10j-m 1.75 ± 0.21de 0.78 ± 0.13j-m 0.25 ± 0.09r 0.95 ± 0.12D

Mean 1.24 ± 0.11C 1.66 ± 0.12A 1.32 ± 0.14B 1.07 ± 0.15D 0.61 ± 0.06E

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

Table - 19: Means comparison of yield for muskmelon fruit peel (Mechanical procedure boiling method interaction

mean±SE)

Boiling Mechanical procedure Mean

Method Homogenizing Grinding Cutting Chopping Hammering

B 1.14 ± 0.15d 1.42 ± 0.17b 1.19 ± 0.18d 0.91 ± 0.24e 0.64 ± 0.05f 1.06 ± 0.08B

M 1.33 ± 0.17bc 1.89 ± 0.16a 1.44 ± 0.21b 1.22 ± 0.16cd 0.58 ± 0.11f 1.29 ± 0.09A

B= Boiling on burner , M= heating in microwave;

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table -20: Means comparison of yield for muskmelon fruit peel (Mechanical procedure pH boiling method interaction

mean±SE)

BM pH Mechanical procedure BM pH

Homogenizing Grinding Cutting Chopping Hammering interaction mean

B 1 0.55 ± 0.03qr 2.25 ± 0.13bcd 0.50 ± 0.03qr 0.05 ± 0.00s 0.65 ± 0.03pqr 0.80 ± 0.20F

104

B 3 2.00 ± 0.04def 1.40 ± 0.05i-l 2.45 ± 0.06abc 0.05 ± 0.00s 0.50 ± 0.02qr 1.28 ± 0.24BCD

B 5 1.45 ± 0.05h-k 1.95 ± 0.10d-g 1.00 ± 0.04m-p 1.70 ± 0.05f-i 0.65 ± 0.03pqr 1.35 ± 0.13B

B 6 1.15 ± 0.06k-n 0.95 ± 0.03m-p 0.70 ± 0.05opq 2.25 ± 0.10bcd 0.95 ± 0.04m-p 1.20 ± 0.15DE

B 7 0.55 ± 0.02qr 0.55 ± 0.01qr 1.30 ± 0.07j-m 0.50 ± 0.03qr 0.45 ± 0.02qr 0.67 ± 0.09G

M 1 0.55 ± 0.03qr 2.65 ± 0.12a 0.50 ± 0.02qr 0.70 ± 0.04opq 1.25 ± 0.05j-m 1.13 ± 0.22E

M 3 1.00 ± 0.03m-p 1.80 ± 0.09e-h 2.55 ± 0.09ab 0.50 ± 0.02qr 0.80 ± 0.06n-q 1.33 ± 0.20BC

M 5 2.30 ± 0.09a-d 2.40 ± 0.14abc 1.00 ± 0.03m-p 2.10 ± 0.08cde 0.30 ± 0.01rs 1.62 ± 0.22A

M 6 1.00 ± 0.05m-p 1.60 ± 0.08g-j 0.95 ± 0.05m-p 1.75 ± 0.04e-i 0.50 ± 0.02qr 1.16 ± 0.12DE

M 7 1.80 ± 0.11e-h 1.00 ± 0.04m-p 2.20 ± 0.09bcd 1.05 ± 0.07l-o 0.05 ± 0.00s 1.22 ± 0.20CDE

B= Boiling on burner , M= heating in microwave; BM= boiling method

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

105

Table -21: Means comparison of yield for apple fruit peel (Mechanical procedure pH interaction mean±SE)

pH level Mechanical procedure Mean

Homogenizing Grinding Cutting Chopping Hammering

1 0.75 ± 0.12ijk 2.48 ± 0.06d 3.38 ± 0.26bc 3.10 ± 0.15c 1.60 ± 0.05gh 2.26 ± 0.19B

3 2.35 ± 0.19de 0.80 ± 0.16ij 3.73 ± 0.36ab 2.00 ± 0.34ef 3.10 ± 0.20c 2.40 ± 0.22A

5 3.03 ± 0.31c 1.05 ± 0.04i 3.20 ± 0.24c 0.90 ± 0.38i 3.95 ± 0.42a 2.43 ± 0.26A

6 1.45 ± 0.05h 2.63 ± 0.12d 2.03 ± 0.39ef 0.50 ± 0.18jkl 1.65 ± 0.18fgh 1.65 ± 0.16C

7 0.20 ± 0.07l 0.30 ± 0.11l 0.38 ± 0.15kl 0.43 ± 0.08jkl 1.88 ± 0.10fg 0.64 ± 0.12D

Mean 1.56 ± 0.20B 1.45 ± 0.18BC 2.54 ± 0.26A 1.39 ± 0.22C 2.44 ± 0.20A

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

Table - 22: Means comparison of yield for apple fruit peel (Mechanical procedure boiling method interaction mean±SE)

Boiling Mechanical procedure Mean

Method Homogenizing Grinding Cutting Chopping Hammering

B 1.27 ± 0.23e 1.27 ± 0.27e 1.94 ± 0.31c 0.90 ± 0.29f 2.04 ± 0.19c 1.48 ± 0.12B

M 1.84 ± 0.33c 1.63 ± 0.24d 3.14 ± 0.36a 1.87 ± 0.28c 2.83 ± 0.32b 2.26 ± 0.15A

B= Boiling on burner , M= heating in microwave; BM= boiling method

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

Table - 23: Means comparison of yield for apple fruit peel (Mechanical procedure pH boiling method interaction

mean±SE)

BM pH Mechanical procedure BM pH

Homogenizing Grinding Cutting Chopping Hammering interaction mean

106

B 1 0.50 ± 0.02r-u 2.45 ± 0.09fgh 2.85 ± 0.16d-g 2.85 ± 0.13d-g 1.50 ± 0.05i-m 2.03 ± 0.25C

B 3 1.95 ± 0.06hij 0.45 ± 0.02r-u 2.95 ± 0.14c-f 1.25 ± 0.08k-o 2.70 ± 0.15efg 1.86 ± 0.25C

B 5 2.35 ± 0.10gh 1.00 ± 0.05m-r 2.70 ± 0.05efg 0.05 ± 0.00u 3.05 ± 0.15cde 1.83 ± 0.30C

B 6 1.50 ± 0.08i-m 2.40 ± 0.07fgh 1.15 ± 0.06l-p 0.10 ± 0.01u 1.25 ± 0.09k-o 1.28 ± 0.20D

B 7 0.05 ± 0.00u 0.05 ± 0.00u 0.05 ± 0.00u 0.25 ± 0.00tu 1.70 ± 0.09i-l 0.42 ± 0.17F

M 1 1.00 ± 0.06m-r 2.50 ± 0.10e-h 3.90 ± 0.19b 3.35 ± 0.20bcd 1.70 ± 0.04i-l 2.49 ± 0.29B

M 3 2.75 ± 0.16efg 1.15 ± 0.04l-p 4.50 ± 0.21a 2.75 ± 0.14efg 3.50 ± 0.10bc 2.93 ± 0.30A

M 5 3.70 ± 0.15b 1.10 ± 0.04m-q 3.70 ± 0.18b 1.75 ± 0.09ijk 4.85 ± 0.23a 3.02 ± 0.37A

M 6 1.40 ± 0.07j-n 2.85 ± 0.11d-g 2.90 ± 0.06d-g 0.90 ± 0.04n-s 2.05 ± 0.05hi 2.02 ± 0.21C

M 7 0.35 ± 0.02stu 0.55 ± 0.02q-u 0.70 ± 0.04o-t 0.60 ± 0.02p-u 2.05 ± 0.12hi 0.85 ± 0.16E

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

107

Table -24: Means comparison of yield for orange fruit peel (Mechanical procedure pH interaction mean±SE)

pH level Mechanical procedure Mean

Homogenizing Grinding Cutting Chopping Hammering

1 15.70 ± 0.45b 3.28 ± 0.32j 7.55 ± 0.30fgh 2.98 ± 0.19j 15.10 ± 0.88bc 8.92 ± 1.05C

3 19.00 ± 0.91a 6.55 ± 0.58ghi 10.25 ± 0.81d 9.00 ± 0.54def 7.78 ± 0.33e-h 10.52 ± 0.87B

5 21.05 ± 1.02a 4.70 ± 1.09ij 16.25 ± 2.07b 13.10 ± 1.27c 9.78 ± 1.11de 12.98 ± 1.18A

6 8.40 ± 2.51d-g 6.53 ± 0.21ghi 8.58 ± 0.81d-g 16.20 ± 0.61b 8.13 ± 1.17d-g 9.57 ± 0.84C

7 14.60 ± 3.20bc 0.40 ± 0.09k 5.70 ± 0.78hi 7.35 ± 0.92fgh 7.18 ± 0.78fgh 7.05 ± 1.07D

Mean 15.75 ± 1.14A 4.29 ± 0.49C 9.67 ± 0.82B 9.73 ± 0.91B 9.59 ± 0.65B

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

Table -25: Means comparison of yield for orange fruit peel (Mechanical procedure boiling method interaction mean±SE)

Boiling Mechanical procedure Mean

Method Homogenizing Grinding Cutting Chopping Hammering

B 17.88 ± 0.78a 3.95 ± 0.73e 7.88 ± 0.69d 8.31 ± 1.17d 7.84 ± 0.80d 9.17 ± 0.65B

M 13.62 ± 2.02b 4.63 ± 0.67e 11.45 ± 1.35c 11.14 ± 1.34c 11.34 ± 0.83c 10.44 ± 0.68A

B= Boiling on burner , M= heating in microwave; BM= boiling method

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

108

Table -26: Means comparison of yield for orange fruit peel (Mechanical procedure pH boiling method interaction

mean±SE)

BM pH Mechanical procedure BM pH

Homogenizing Grinding Cutting Chopping Hammering interaction mean

B 1 16.15 ± 0.44d-g 2.60 ± 0.14uvw 8.10 ± 0.33m-r 2.60 ± 0.11uvw 13.55 ± 0.48g-j 8.60 ± 1.49DE

B 3 18.20 ± 1.36b-e 7.80 ± 0.21m-r 8.50 ± 0.20l-r 7.95 ± 0.33m-r 7.20 ± 0.41n-s 9.93 ± 1.14CD

B 5 19.40 ± 0.80a-d 2.30 ± 0.10uvw 11.85 ± 0.64i-l 10.40 ± 0.66j-n 7.40 ± 0.20m-r 10.27 ± 1.52BC

B 6 13.95 ± 0.76f-i 6.45 ± 0.40p-t 6.95 ± 0.41o-s 15.25 ± 0.72e-h 5.55 ± 0.24q-u 9.63 ± 1.12B

B 7 21.70 ± 0.72a 0.60 ± 0.03vw 4.00 ± 0.21stu 5.35 ± 0.20r-u 5.50 ± 0.18q-u 7.43 ± 1.97BC

M 1 15.25 ± 0.80e-h 3.95 ± 0.16s-v 7.00 ± 0.17o-s 3.35 ± 0.14t-w 16.65 ± 1.12c-g 9.24 ± 1.52A

M 3 19.80 ± 1.29abc 5.30 ± 0.28r-u 12.00 ± 0.46h-k 10.05 ± 0.51k-o 8.35 ± 0.22m-r 11.10 ± 1.33CD

M 5 22.70 ± 1.36a 7.10 ± 0.39n-s 20.65 ± 1.25ab 15.80 ± 0.54efg 12.15 ± 0.66h-k 15.68 ± 1.55CD

M 6 2.85 ± 0.19uvw 6.60 ± 0.22p-t 10.20 ± 0.66j-o 17.15 ± 0.67c-f 10.70 ± 0.46i-m 9.50 ± 1.28EF

M 7 7.50 ± 0.35m-r 0.20 ± 0.01w 7.40 ± 0.35m-r 9.35 ± 0.43k-p 8.85 ± 0.48k-q 6.66 ± 0.90F

Means sharing similar letter in a row or in a column are statistically non-significant (P>0.05). Small letters represent comparison among interaction means and

capital letters are used for overall mean

109

Table - 27: Percentage yield of pectin from sapodilla fruit peel after using different

physicomechanical procedures ( pH, mechanical procedure, boiling method and time of

boiling) with 0.1N HCl

Mechanical procedure

pH %yield 10 Min %yield 20min %yield 40 min %yield 60min

B M B M B M B M

Ham

mer

ing

1 1.5 2.05 0.5 1.1 0.05 0.05 1.2 1.8

3 0.6 1.55 2.2 4.85 0.05 0.1 4.1 3.15

5 0.8 1.95 5.5 6.35 0.05 0.05 4.5 4.6

6 0.8 1.4 3.5 5.2 1.65 1.65 3 3.45

7 0.5 0 2.6 4.35 0.05 0.95 1.75 3.85

Gri

nd

ing

1 0.55 0.8 0.75 1.6 0.1 1.15 0.25 0.35

3 4 3.5 3.55 4 1 1.35 0.05 0.05

5 2.45 2.9 2.05 3.8 2.4 2.75 0.05 2.1

6 2 2 3.4 2.7 2.1 3.1 0.05 0.05

7 1.95 2.3 2.85 3.3 0.95 1.75 0.05 2.3

Cu

ttin

g

1 0.8 0.9 1.45 1.5 0.05 0.05 0.05 0.1

3 1.1 1.5 1.95 3.15 0.45 0.05 0.4 1.4

5 0.4 1 1.5 2.2 1.2 3.95 0.05 0.5

6 1 1.2 1.45 1.85 0.65 1 0.1 0.9

7 0.7 1 1.35 1.8 0.1 1.5 0.05 1.05

Ho

mo

gen

izin

g

1 0.5 1.05 0.05 0.15 0.05 0.05 0.05 0.05

3 0.75 1.6 0.05 1 2.5 1.3 2.75 2.8

5 1.2 1.5 0.05 0.1 3 3.2 2.7 6

6 1.5 2 0.05 0.05 2.25 2.45 2.7 4.5

7 0.5 0.5 0.1 0.05 2.45 2.8 0.15 2.65

Ch

op

pin

g

(mort

ar/

pes

tle

1 2.7 4.7 0.05 0.05 0.05 0.05 1.85 2.75

3 3.9 3.45 4.75 6.5 1.6 1.35 1.25 3

5 3.95 4.05 2.25 3.05 2.05 2.35 1.95 5.6

6 2.3 2.45 2.5 2.8 2.7 3.6 3.25 5

7 3.95 2.4 0.1 0.7 1 2 3.6 3.65

B= Boiling on burner , M= heating in microwave

110

Table -28: Analysis of variance (mean squares) of yield of pectin from sapodilla fruit peel after using different

physicomechanical procedure different fruits with 0.1N HCl

Source of

variation

Degrees of

freedom

Mean squares

Yield 10 min Yield 20 min Yield 40 min Yield 60 min

MP

pH

BM

MP x pH

MP x BM

pH x BM

MP x pH x BM

Error

Total

4

4

1

16

4

4

16

100

149

32.6575**

3.3917**

3.2414**

2.5462**

0.3392**

1.0112**

0.6258**

0.0081

49.7412**

27.4284**

18.6772**

7.5601**

1.7538**

1.4724**

0.4729**

0.0129

12.1446**

20.1921**

6.1206**

2.3355**

1.0438**

1.6176**

0.5655**

0.0070

55.6906**

15.5938**

39.7837**

4.3065**

2.0295**

3.7639**

1.5634**

0.0122

NS = Non-significant (P>0.05); * = Significant (P<0.05); ** = Highly significant (P<0.01). Mech = Mechanical

Table – 29: Means comparison of yield for sapodilla fruit peel after 10 min of boiling using 0.1N HCl (Mechanical procedure

pH interaction mean±SE)

111

pH level Mechanical procedure Mean

Homogenizing Grinding Cutting Chopping Hammering

1 1.78 ± 0.13g 0.68 ± 0.06kl 0.85 ± 0.02k 3.70 ± 0.45b 0.78 ± 0.12k 1.56 ± 0.23D

3 1.08 ± 0.21j 3.75 ± 0.14b 1.30 ± 0.09hi 3.68 ± 0.11b 1.18 ± 0.19ij 2.20 ± 0.24A

5 1.38 ± 0.26h 2.68 ± 0.10d 0.70 ± 0.13k 4.00 ± 0.07a 1.35 ± 0.07hi 2.02 ± 0.23B

6 1.10 ± 0.13j 2.00 ± 0.04f 1.10 ± 0.05j 2.38 ± 0.04e 1.75 ± 0.12g 1.67 ± 0.10C

7 0.25 ± 0.11m 2.13 ± 0.08f 0.85 ± 0.07k 3.18 ± 0.35c 0.50 ± 0.01l 1.38 ± 0.22E

Mean 1.12 ± 0.12C 2.25 ± 0.19B 0.96 ± 0.05D 3.39 ± 0.15A 1.11 ± 0.09C

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table - 30: Means comparison of yield for sapodilla fruit peel after 10 min of boiling using 0.1N HCl (Mechanical procedure

boiling method interaction mean±SE)

Boiling Mechanical procedure Mean

Method Homogenizing Grinding Cutting Chopping Hammering

B 0.84 ± 0.09f 2.19 ± 0.30c 0.80 ± 0.07f 3.36 ± 0.19a 0.89 ± 0.11f 1.62 ± 0.14B

M 1.39 ± 0.20d 2.30 ± 0.24b 1.12 ± 0.06e 3.41 ± 0.24a 1.33 ± 0.14d 1.91 ± 0.13A B= Boiling on burner , M= heating in microwave; BM= boiling method In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table -31: Means comparison of yield for sapodilla fruit peel after 10 min of boiling using 0.1N HCl (Mechanical procedure

pH boiling method interaction mean±SE)

BM pH Mechanical procedure BM pH

Homogenizing Grinding Cutting Chopping Hammering interaction mean

B 1 1.50 ± 0.02ij 0.55 ± 0.01qr 0.80 ± 0.02m-q 2.70 ± 0.05de 0.50 ± 0.01qr 1.21 ± 0.22F

B 3 0.60 ± 0.02pqr 4.00 ± 0.10b 1.10 ± 0.02klm 3.90 ± 0.09b 0.75 ± 0.02n-q 2.07 ± 0.41B

B 5 0.80 ± 0.01m-q 2.45 ± 0.03ef 0.40 ± 0.01r 3.95 ± 0.12b 1.20 ± 0.03jkl 1.76 ± 0.35D

112

B 6 0.80 ± 0.01m-q 2.00 ± 0.05gh 1.00 ± 0.02l-o 2.30 ± 0.05fg 1.50 ± 0.04ij 1.52 ± 0.15E

B 7 0.50 ± 0.02qr 1.95 ± 0.06h 0.70 ± 0.02o-r 3.95 ± 0.11b 0.50 ± 0.01qr 1.52 ± 0.36E

M 1 2.05 ± 0.05gh 0.80 ± 0.02m-q 0.90 ± 0.02l-p 4.70 ± 0.10a 1.05 ± 0.01lmn 1.90 ± 0.39C

M 3 1.55 ± 0.05i 3.50 ± 0.14c 1.50 ± 0.03ij 3.45 ± 0.02c 1.60 ± 0.04i 2.32 ± 0.25A

M 5 1.95 ± 0.05h 2.90 ± 0.05d 1.00 ± 0.01l-o 4.05 ± 0.10b 1.50 ± 0.05ij 2.28 ± 0.29A

M 6 1.40 ± 0.03ijk 2.00 ± 0.08gh 1.20 ± 0.03jkl 2.45 ± 0.04ef 2.00 ± 0.05gh 1.81 ± 0.12CD

M 7 0.00 ± 0.00s 2.30 ± 0.02fg 1.00 ± 0.02l-o 2.40 ± 0.06ef 0.50 ± 0.01qr 1.24 ± 0.26F

B= Boiling on burner , M= heating in microwave; BM= boiling method In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table -32: Means comparison of yield for sapodilla fruit peel after 20 min of boiling using 0.1N HCl (Mechanical procedure

pH interaction mean±SE)

pH level Mechanical procedure Mean

Homogenizing Grinding Cutting Chopping Hammering

1 0.80 ± 0.13k 1.18 ± 0.19j 1.48 ± 0.03i 0.05 ± 0.00m 0.10 ± 0.02m 0.72 ± 0.11E

3 3.53 ± 0.60e 3.78 ± 0.12d 2.55 ± 0.27g 5.63 ± 0.40b 0.53 ± 0.21l 3.20 ± 0.34A

5 5.93 ± 0.19a 2.93 ± 0.40f 1.85 ± 0.16h 2.65 ± 0.19g 0.08 ± 0.01m 2.69 ± 0.37B

6 4.35 ± 0.38c 3.05 ± 0.17f 1.65 ± 0.09hi 2.65 ± 0.08g 0.05 ± 0.00m 2.35 ± 0.28C

7 3.48 ± 0.40e 3.08 ± 0.12f 1.58 ± 0.10i 0.40 ± 0.13l 0.08 ± 0.01m 1.72 ± 0.27D

Mean 3.62 ± 0.35A 2.80 ± 0.19B 1.82 ± 0.10D 2.27 ± 0.38C 0.17 ± 0.05E In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

Table - 33: Means comparison of yield for sapodilla fruit peel after 20 min of boiling using 0.1N HCl (Mechanical procedure

boiling method interaction mean±SE)

Boiling Mechanical procedure Mean

Method Homogenizing Grinding Cutting Chopping Hammering

113

B 2.86 ± 0.44c 2.52 ± 0.28d 1.54 ± 0.06g 1.93 ± 0.47f 0.06 ± 0.01i 1.78 ± 0.18B

M 4.37 ± 0.47a 3.08 ± 0.23b 2.10 ± 0.15e 2.62 ± 0.61d 0.27 ± 0.10h 2.49 ± 0.22A B= Boiling on burner , M= heating in microwave; BM= boiling method

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table - 34: Means comparison of yield for sapodilla fruit peel after 20 min of boiling using 0.1N HCl (Mechanical procedure

pH boiling method interaction mean±SE)

BM pH Mechanical procedure BM pH

Homogenizing Grinding Cutting Chopping Hammering interaction mean

B 1 0.50 ± 0.01ab 0.75 ± 0.01YZa 1.45 ± 0.03UVW 0.05 ± 0.00c 0.05 ± 0.00c 0.56 ± 0.14H

B 3 2.20 ± 0.04OPQ 3.55 ± 0.10GH 1.95 ± 0.01P-S 4.75 ± 0.09D 0.05 ± 0.00c 2.50 ± 0.42C

B 5 5.50 ± 0.07B 2.05 ± 0.03PQR 1.50 ± 0.05TUV 2.25 ± 0.06NOP 0.05 ± 0.00c 2.27 ± 0.48D

B 6 3.50 ± 0.05GHI 3.40 ± 0.13HIJ 1.45 ± 0.03UVW 2.50 ± 0.06MNO 0.05 ± 0.00c 2.18 ± 0.35D

B 7 2.60 ± 0.02MN 2.85 ± 0.09KLM 1.35 ± 0.02VWX 0.10 ± 0.01c 0.10 ± 0.01c 1.40 ± 0.31F

M 1 1.10 ± 0.01WXY 1.60 ± 0.03S-V 1.50 ± 0.05TUV 0.05 ± 0.00c 0.15 ± 0.01bc 0.88 ± 0.18G

M 3 4.85 ± 0.13CD 4.00 ± 0.12EF 3.15 ± 0.06IJK 6.50 ± 0.17A 1.00 ± 0.03XYZ 3.90 ± 0.49A

M 5 6.35 ± 0.05A 3.80 ± 0.12FG 2.20 ± 0.03OPQ 3.05 ± 0.10JKL 0.10 ± 0.01c 3.10 ± 0.55B

M 6 5.20 ± 0.04BC 2.70 ± 0.08LM 1.85 ± 0.05Q-T 2.79 ± 0.10KLM 0.05 ± 0.01c 2.52 ± 0.45C

M 7 4.35 ± 0.14E 3.30 ± 0.10HIJ 1.80 ± 0.05R-U 0.70 ± 0.02Za 0.05 ± 0.00c 2.04 ± 0.43E

B= Boiling on burner , M= heating in microwave; BM= boiling method

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison

among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

Table - 35: Means comparison of yield for sapodilla fruit peel after 40 min of boiling using 0.1N HCl (Mechanical procedure

pH interaction mean±SE)

114

pH level Mechanical procedure Mean

Homogenizing Grinding Cutting Chopping Hammering

1 0.05 ± 0.00l 0.63 ± 0.23ij 0.05 ± 0.00l 0.05 ± 0.00l 0.05 ± 0.00l 0.17 ± 0.06D

3 0.08 ± 0.01kl 1.18 ± 0.08g 0.25 ± 0.09k 1.48 ± 0.06ef 1.90 ± 0.27d 0.98 ± 0.14C

5 0.05 ± 0.00l 2.58 ± 0.08b 2.58 ± 0.62b 2.20 ± 0.08c 3.10 ± 0.09a 2.10 ± 0.23A

6 1.65 ± 0.02e 2.60 ± 0.23b 0.83 ± 0.08h 3.15 ± 0.20a 2.35 ± 0.07c 2.12 ± 0.16A

7 0.50 ± 0.20j 1.35 ± 0.18fg 0.80 ± 0.31hi 1.50 ± 0.23ef 2.63 ± 0.09b 1.36 ± 0.16B

Mean 0.47 ± 0.12D 1.67 ± 0.16B 0.90 ± 0.21C 1.68 ± 0.20B 2.01 ± 0.20A

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table - 36: Means comparison of yield for sapodilla fruit peel after 40 min of boiling using 0.1N HCl ( Mechanical procedure

boiling method interaction mean±SE)

Boiling Mechanical procedure Mean

Method Homogenizing Grinding Cutting Chopping Hammering

B 0.37 ± 0.17f 1.31 ± 0.22d 0.49 ± 0.11e 1.48 ± 0.24c 2.05 ± 0.28a 1.14 ± 0.12B

M 0.56 ± 0.17e 2.02 ± 0.21a 1.31 ± 0.38d 1.87 ± 0.31b 1.96 ± 0.31ab 1.54 ± 0.14A

B= Boiling on burner , M= heating in microwave; BM= boiling method

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table - 37: Means comparison of yield for sapodilla fruit peel after 40 min of boiling using 0.1N HCl (Mechanical procedure

pH boiling method interaction mean±SE)

BM pH Mechanical procedure BM pH

Homogenizing Grinding Cutting Chopping Hammering interaction mean

B 1 0.05 ± 0.00r 0.10 ± 0.00r 0.05 ± 0.00r 0.05 ± 0.00r 0.05 ± 0.01r 0.06 ± 0.01H

B 3 0.05 ± 0.00r 1.00 ± 0.01p 0.45 ± 0.01q 1.60 ± 0.05lm 2.50 ± 0.09fgh 1.12 ± 0.23E

B 5 0.05 ± 0.00r 2.40 ± 0.05h 1.20 ± 0.01op 2.05 ± 0.05j 3.00 ± 0.11cd 1.74 ± 0.28D

115

B 6 1.65 ± 0.02l 2.10 ± 0.02ij 0.65 ± 0.01q 2.70 ± 0.03efg 2.25 ± 0.08hij 1.87 ± 0.19C

B 7 0.05 ± 0.00r 0.95 ± 0.01p 0.10 ± 0.01r 1.00 ± 0.06p 2.45 ± 0.05gh 0.91 ± 0.23F

M 1 0.05 ± 0.00r 1.15 ± 0.01op 0.05 ± 0.00r 0.05 ± 0.01r 0.05 ± 0.00r 0.27 ± 0.12G

M 3 0.10 ± 0.00r 1.35 ± 0.02mno 0.05 ± 0.00r 1.35 ± 0.05mno 1.30 ± 0.04no 0.83 ± 0.17F

M 5 0.05 ± 0.00r 2.75 ± 0.06def 3.95 ± 0.10a 2.35 ± 0.09hi 3.20 ± 0.13c 2.46 ± 0.35A

M 6 1.65 ± 0.03l 3.10 ± 0.09c 1.00 ± 0.01p 3.60 ± 0.08b 2.45 ± 0.07gh 2.36 ± 0.25B

M 7 0.95 ± 0.01p 1.75 ± 0.03kl 1.50 ± 0.03lmn 2.00 ± 0.06jk 2.80 ± 0.07de 1.80 ± 0.16CD

B= Boiling on burner , M= heating in microwave; BM= boiling method

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

116

Table - 38: Means comparison of yield for sapodilla fruit peel after 60 min of boiling using 0.1N HCl Mechanical procedure

pH interaction mean±SE)

pH level Mechanical procedure Mean

Homogenizing Grinding Cutting Chopping Hammering

1 1.50 ± 0.13g 0.30 ± 0.02lm 0.08 ± 0.01mn 2.30 ± 0.21f 0.05 ± 0.00n 0.85 ± 0.17D

3 3.63 ± 0.21c 0.05 ± 0.00n 0.90 ± 0.22j 2.13 ± 0.40f 2.78 ± 0.04e 1.90 ± 0.26C

5 4.55 ± 0.06a 1.08 ± 0.46ij 0.28 ± 0.10lmn 3.78 ± 0.82c 4.35 ± 0.74ab 2.81 ± 0.40A

6 3.23 ± 0.10d 0.05 ± 0.00n 0.50 ± 0.18kl 4.13 ± 0.40b 3.60 ± 0.41c 2.30 ± 0.33B

7 2.80 ± 0.47e 1.18 ± 0.50hi 0.55 ± 0.22k 3.63 ± 0.04c 1.40 ± 0.56gh 1.91 ± 0.27C

Mean 3.14 ± 0.21A 0.53 ± 0.16C 0.46 ± 0.09C 3.19 ± 0.24A 2.44 ± 0.34B In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table - 39: Means comparison of yield for sapodilla fruit peel after 60 min of boiling using 0.1N HCl (Mechanical procedure

boiling method interaction mean±SE)

Boiling Mechanical procedure Mean

Method Homogenizing Grinding Cutting Chopping Hammering

B 2.91 ± 0.34d 0.09 ± 0.02i 0.13 ± 0.04i 2.38 ± 0.24e 1.67 ± 0.34f 1.44 ± 0.17B

M 3.37 ± 0.25b 0.97 ± 0.27g 0.79 ± 0.12h 4.00 ± 0.30a 3.20 ± 0.54c 2.47 ± 0.21A B= Boiling on burner , M= heating in microwave; BM= boiling method

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table - 40: Means comparison of yield for sapodilla fruit peel after 60 min of boiling using 0.1N HCl (Mechanical procedure

pH boiling method interaction mean±SE)

BM pH Mechanical procedure BM pH

Homogenizing Grinding Cutting Chopping Hammering interaction mean

B 1 1.20 ± 0.01qr 0.25 ± 0.01st 0.05 ± 0.00t 1.85 ± 0.05o 0.05 ± 0.00t 0.68 ± 0.19G

B 3 4.10 ± 0.03e 0.05 ± 0.00t 0.40 ± 0.01st 1.25 ± 0.03qr 2.75 ± 0.05l 1.71 ± 0.41E

117

B 5 4.50 ± 0.04d 0.05 ± 0.00t 0.05 ± 0.00t 1.95 ± 0.03no 2.70 ± 0.02l 1.85 ± 0.45D

B 6 3.00 ± 0.06jkl 0.05 ± 0.00t 0.10 ± 0.01t 3.25 ± 0.09hij 2.70 ± 0.10l 1.82 ± 0.38DE

B 7 1.75 ± 0.03op 0.05 ± 0.01t 0.05 ± 0.01t 3.60 ± 0.04fgh 0.15 ± 0.01st 1.12 ± 0.37F

M 1 1.80 ± 0.02o 0.35 ± 0.00st 0.10 ± 0.01t 2.75 ± 0.09l 0.05 ± 0.01t 1.01 ± 0.29F

M 3 3.15 ± 0.05ijk 0.05 ± 0.00t 1.40 ± 0.03pq 3.00 ± 0.12jkl 2.80 ± 0.08kl 2.08 ± 0.32C

M 5 4.60 ± 0.11d 2.10 ± 0.04no 0.50 ± 0.02s 5.60 ± 0.07b 6.00 ± 0.21a 3.76 ± 0.57A

M 6 3.45 ± 0.03ghi 0.05 ± 0.00t 0.90 ± 0.02r 5.00 ± 0.17c 4.50 ± 0.14d 2.78 ± 0.53B

M 7 3.85 ± 0.06ef 2.30 ± 0.08mn 1.05 ± 0.02qr 3.65 ± 0.07fg 2.65 ± 0.11lm 2.70 ± 0.27B

B= Boiling on burner , M= heating in microwave; BM= boiling method In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

118

Table - 41: Percentage yield of pectin from sapodilla fruit peel after using different

physicomechanical procedures (pH, mechanical procedure, boiling method and time of

boling) with IN HCl.

Mechanical

procedure pH

% yield 10 min %Yield 20 min %yield 40 min % yield 60 min

B M B M B M B M

Hammering

1 2.65 3.15 1.5 2.2 3.5 3 0.3 2

3 5.65 6.55 3.45 5.45 4.5 4.5 0.4 2.85

5 2.3 4.6 4.4 4.5 4.2 5.25 0.15 2.3

6 2.15 2.95 2.45 4.3 2 3.4 3.45 2.6

7 2.5 3.1 2.4 2.45 2 2.55 3.5 0.05

Grinding

1 3.1 4.15 1.55 1.65 1.4 3.7 1.4 3

3 5.6 4.8 2.55 5.4 3.25 5.15 3.15 5

5 1.05 6.1 4.2 5.1 3.8 5.5 1 3.3

6 1.45 3.55 4.3 3.65 3.5 3.3 0.8 1.2

7 0.35 2.75 1.3 2.5 2.15 2.6 1.65 1.6

Cutting

1 0.05 0.05 0.5 1.4 3.65 4 0.65 0.05

3 0.05 1.6 1 2.35 2.5 3.5 0.75 0.1

5 2.95 5.3 2.05 2.7 4.1 5.5 0.1 0.05

6 0.15 3.5 3.35 3.7 1.5 3 0 0.6

7 0.05 1.2 2.6 1.8 2 3.25 0.1 0.05

Homogenizing

1 1.55 4.95 2.1 3.6 3.4 4.6 0.6 0

3 2.4 5.35 3.55 5.25 3.65 4.55 0.05 0.15

5 1.7 4.85 3.55 5.9 2.7 3 0.15 3.4

6 0.6 1.5 3.8 3.15 3.9 3.5 0.05 0.25

7 1.2 2.55 1.25 2.2 3.35 3.3 0.1 0.6

Chopping 1 3 5.5 2.5 5.9 1.95 3.2 0.05 3.55

(mortor/pastle 3 3.7 5.4 1.4 6 1.15 4.85 0.2 0.1

5 4.1 6.7 1.25 6.1 4.5 6.5 0.45 0.5

6 3.75 3.8 1.2 4.9 2.5 3.1 0 1

7 0.05 0.25 1.8 2.5 1 3.95 0.1 0.6

B= Boiling on burner , M= heating in microwave

119

Table - 42: Analysis of variance (mean squares) of yield of pectin from sapodilla fruit peel after using different

physicomechanical procedure different fruits with 1N HCl

Source of

variation

Degrees of

freedom

Mean squares

Yield 10 min_NHCl Yield 20 min_NHCl Yield 40 min_NHCl Yield 60 min_NHCl

MP

pH

BM

MP x pH

MP x BM

pH x BM

MP x pH x BM

Error

Total

4

4

1

16

4

4

16

100

149

23.648**

38.736**

106.361**

8.396**

1.944**

4.779**

2.360**

0.022

8.5850**

21.8147**

71.7604**

3.9668**

10.4560**

4.7719**

1.5394**

0.0173

0.5439**

16.3404**

42.5601**

3.5228**

3.5189**

0.9204**

1.2315**

0.0168

21.8563**

0.8645**

14.8838**

4.0399**

2.1424**

4.7014**

3.3080**

0.0054

NS = Non-significant (P>0.05); * = Significant (P<0.05); ** = Highly significant (P<0.01); Mech= mechanical

120

Table - 43: Means comparison of yield for sapodilla fruit peel after 10 min of boiling using 1N HCl ( Mechanical procedure

pH interaction mean±SE)

pH level Mechanical procedure Mean

Homogenizing Grinding Cutting Chopping Hammering

1 2.90 ± 0.12j 3.63 ± 0.24fg 0.05 ± 0.00p 4.25 ± 0.56cd 3.25 ± 0.76i 2.82 ± 0.33C

3 6.10 ± 0.21a 5.20 ± 0.19b 0.83 ± 0.35no 4.55 ± 0.39c 3.88 ± 0.66ef 4.11 ± 0.37A

5 3.45 ± 0.51ghi 3.58 ± 1.13fgh 4.13 ± 0.53de 5.40 ± 0.59b 3.28 ± 0.71hi 3.97 ± 0.34B

6 2.55 ± 0.18k 2.50 ± 0.47k 1.83 ± 0.75lm 3.78 ± 0.07f 1.05 ± 0.20n 2.34 ± 0.24D

7 2.80 ± 0.14jk 1.55 ± 0.54m 0.63 ± 0.26o 0.15 ± 0.05p 1.88 ± 0.30l 1.40 ± 0.22E

Mean 3.56 ± 0.27A 3.29 ± 0.34B 1.49 ± 0.33D 3.63 ± 0.38A 2.67 ± 0.31C In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table - 44: Means comparison of yield for sapodilla fruit peel after 10 min of boiling using 1N HCl ( Mechanical procedure

boiling method interaction mean±SE)

Boiling Mechanical procedure Mean

Method Homogenizing Grinding Cutting Chopping Hammering

B 3.05 ± 0.35d 2.31 ± 0.50e 0.65 ± 0.31g 2.92 ± 0.40d 1.49 ± 0.16f 2.08 ± 0.19B

M 4.07 ± 0.37b 4.27 ± 0.31a 2.33 ± 0.50e 4.33 ± 0.60a 3.84 ± 0.41c 3.77 ± 0.21A

B= Boiling on burner , M= heating in microwave

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison

among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

Table - 45: Means comparison of yield for sapodilla fruit peel after 10 min of boiling using 1N HCl (Mechanical procedure

pH boiling method interaction mean±SE)

BM pH Mechanical procedure BM pH

Homogenizing Grinding Cutting Chopping Hammering interaction mean

B 1 2.65 ± 0.03n-q 3.10 ± 0.05lmn 0.05 ± 0.01w 3.00 ± 0.06mno 1.55 ± 0.03st 2.07 ± 0.31F

121

B 3 5.65 ± 0.12cd 5.60 ± 0.07cd 0.05 ± 0.01w 3.70 ± 0.10jk 2.40 ± 0.08pq 3.48 ± 0.56C

B 5 2.30 ± 0.05pq 1.05 ± 0.03tu 2.95 ± 0.11no 4.10 ± 0.15ij 1.70 ± 0.04rs 2.42 ± 0.28E

B 6 2.15 ± 0.04qr 1.45 ± 0.01st 0.15 ± 0.01vw 3.75 ± 0.08jk 0.60 ± 0.01uv 1.62 ± 0.34G

B 7 2.50 ± 0.05opq 0.35 ± 0.01vw 0.05 ± 0.01w 0.05 ± 0.01w 1.20 ± 0.03st 0.83 ± 0.25H

M 1 3.15 ± 0.09lmn 4.15 ± 0.05ij 0.05 ± 0.00w 5.50 ± 0.08d 4.95 ± 0.12e-h 3.56 ± 0.52C

M 3 6.55 ± 0.08ab 4.80 ± 0.13gh 1.60 ± 0.05s 5.40 ± 0.20de 5.35 ± 0.10def 4.74 ± 0.45B

M 5 4.60 ± 0.02hi 6.10 ± 0.20bc 5.30 ± 0.07d-g 6.70 ± 0.24a 4.85 ± 0.16fgh 5.51 ± 0.22A

M 6 2.95 ± 0.09no 3.55 ± 0.05kl 3.50 ± 0.05klm 3.80 ± 0.13jk 1.50 ± 0.05st 3.06 ± 0.22D

M 7 3.10 ± 0.01lmn 2.75 ± 0.09nop 1.20 ± 0.03st 0.25 ± 0.02vw 2.55 ± 0.05opq 1.97 ± 0.29F

B= Boiling on burner , M= heating in microwave; BM= boiling method

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table - 46: Means comparison of yield for sapodilla fruit peel after 20 min of boiling using 1N HCl Mechanical procedure

pH interaction mean±SE)

pH level Mechanical procedure Mean

Homogenizing Grinding Cutting Chopping Hammering

1 1.82 ± 0.15kl 1.60 ± 0.03l 0.95 ± 0.20m 4.20 ± 0.77cd 2.85 ± 0.34h 2.28 ± 0.27D

3 4.45 ± 0.45abc 3.98 ± 0.64de 1.68 ± 0.30kl 3.70 ± 1.03ef 4.40 ± 0.39bc 3.64 ± 0.32B

5 4.45 ± 0.06abc 4.65 ± 0.21ab 2.38 ± 0.15i 3.68 ± 1.09f 4.73 ± 0.53a 3.98 ± 0.28A

6 3.38 ± 0.41g 3.98 ± 0.15de 3.53 ± 0.10fg 3.05 ± 0.83h 3.48 ± 0.16fg 3.48 ± 0.19C

7 2.43 ± 0.03i 1.90 ± 0.27jk 2.20 ± 0.18i 2.15 ± 0.16ij 1.73 ± 0.21kl 2.08 ± 0.09E

Mean 3.30 ± 0.23BC 3.22 ± 0.27C 2.15 ± 0.18D 3.36 ± 0.37AB 3.44 ± 0.25A

122

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

123

Table - 47: Means comparison of yield for sapodilla fruit peel after 20 min of boiling using 1N HCl ( Mechanical procedure

boiling method interaction mean±SE)

Boiling Mechanical procedure Mean

Method Homogenizing Grinding Cutting Chopping Hammering

B 2.84 ± 0.27d 2.78 ± 0.34d 1.90 ± 0.28f 1.63 ± 0.13g 2.85 ± 0.27d 2.40 ± 0.13B

M 3.77 ± 0.34c 3.66 ± 0.39c 2.39 ± 0.21e 5.08 ± 0.37a 4.02 ± 0.37b 3.78 ± 0.18A

B= Boiling on burner , M= heating in microwave In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

124

Table - 48: Means comparison of yield for sapodilla fruit peel after 20 min of boiling using 1N HCl (Mechanical procedure

pH boiling method interaction mean±SE)

BM pH Mechanical procedure BM pH

Homogenizing Grinding Cutting Chopping Hammering interaction mean

B 1 1.50 ± 0.02qrs 1.55 ± 0.03qrs 0.50 ± 0.01u 2.50 ± 0.06j-m 2.10 ± 0.05mno 1.63 ± 0.18F

B 3 3.45 ± 0.02ghi 2.55 ± 0.05jkl 1.00 ± 0.02t 1.40 ± 0.03q-t 3.55 ± 0.06ghi 2.39 ± 0.28D

B 5 4.40 ± 0.06e 4.20 ± 0.07ef 2.05 ± 0.03nop 1.25 ± 0.02rst 3.55 ± 0.05ghi 3.09 ± 0.33C

B 6 2.45 ± 0.04j-n 4.30 ± 0.04e 3.35 ± 0.05hi 1.20 ± 0.03st 3.80 ± 0.10fg 3.02 ± 0.29C

B 7 2.40 ± 0.05j-n 1.30 ± 0.01rst 2.60 ± 0.09jk 1.80 ± 0.05opq 1.25 ± 0.01rst 1.87 ± 0.15E

M 1 2.13 ± 0.07l-o 1.65 ± 0.01pqr 1.40 ± 0.01q-t 5.90 ± 0.19a 3.60 ± 0.08gh 2.94 ± 0.45C

M 3 5.45 ± 0.08b 5.40 ± 0.13b 2.35 ± 0.02j-n 6.00 ± 0.15a 5.25 ± 0.14bc 4.89 ± 0.35A

M 5 4.50 ± 0.10de 5.10 ± 0.10bc 2.70 ± 0.06j 6.10 ± 0.08a 5.90 ± 0.17a 4.86 ± 0.33A

M 6 4.30 ± 0.03e 3.65 ± 0.06gh 3.70 ± 0.14gh 4.90 ± 0.11cd 3.15 ± 0.07i 3.94 ± 0.16B

M 7 2.45 ± 0.04j-n 2.50 ± 0.08j-m 1.80 ± 0.04opq 2.50 ± 0.05j-m 2.20 ± 0.06k-o 2.29 ± 0.08D

B= Boiling on burner , M= heating in microwave; BM= boiling method

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison

among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

125

Table - 49: Means comparison of yield for sapodilla fruit peel after 40 min of boiling using 1N HCl (Mechanical procedure

pH interaction mean±SE)

pH level Mechanical procedure Mean

Homogenizing Grinding Cutting Chopping Hammering

1 3.25 ± 0.12gh 2.55 ± 0.51k-n 3.83 ± 0.09ef 2.58 ± 0.28j-m 4.00 ± 0.28de 3.24 ± 0.17C

3 4.50 ± 0.05c 4.20 ± 0.43d 3.00 ± 0.23hi 3.00 ± 0.83hi 4.10 ± 0.21de 3.76 ± 0.22B

5 4.73 ± 0.24bc 4.65 ± 0.38bc 4.80 ± 0.32b 5.50 ± 0.46a 2.85 ± 0.08ij 4.51 ± 0.21A

6 2.70 ± 0.32jkl 3.38 ± 0.04g 2.25 ± 0.34o 2.80 ± 0.14ijk 3.70 ± 0.10f 2.97 ± 0.13D

7 2.28 ± 0.13no 2.38 ± 0.10mno 2.63 ± 0.28j-m 2.48 ± 0.66l-o 3.33 ± 0.04g 2.62 ± 0.15E

Mean 3.49 ± 0.20B 3.43 ± 0.22B 3.30 ± 0.20C 3.27 ± 0.31C 3.60 ± 0.11A

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table - 50: Means comparison of yield for sapodilla fruit peel after 40 min of boiling using 1N HCl (Mechanical procedure

boiling method interaction mean±SE)

Boiling Mechanical procedure Mean

Method Homogenizing Grinding Cutting Chopping Hammering

B 3.24 ± 0.29e 2.81 ± 0.24f 2.75 ± 0.26f 2.22 ± 0.34g 3.40 ± 0.11d 2.88 ± 0.12B

M 3.74 ± 0.27c 4.05 ± 0.30b 3.85 ± 0.24c 4.32 ± 0.34a 3.79 ± 0.18c 3.95 ± 0.12A

B= Boiling on burner , M= heating in microwave

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table - 51: Means comparison of yield for sapodilla fruit peel after 40 min of boiling using 1N HCl ( Mechanical procedure

pH boiling method interaction mean±SE)

BM pH Mechanical procedure BM pH

Homogenizing Grinding Cutting Chopping Hammering interaction mean

B 1 3.50 ± 0.05j-n 1.40 ± 0.02uv 3.65 ± 0.05i-m 1.95 ± 0.04t 3.40 ± 0.10k-o 2.78 ± 0.25G

126

B 3 4.50 ± 0.11def 3.25 ± 0.04mno 2.50 ± 0.05rs 1.15 ± 0.02uv 3.65 ± 0.12i-m 3.01 ± 0.30F

B 5 4.20 ± 0.04efg 3.80 ± 0.11g-k 4.10 ± 0.09fgh 4.50 ± 0.04def 2.70 ± 0.10pqr 3.86 ± 0.17C

B 6 2.00 ± 0.04t 3.47 ± 0.03j-n 1.50 ± 0.01u 2.50 ± 0.03rs 3.90 ± 0.05g-j 2.67 ± 0.24G

B 7 2.00 ± 0.04t 2.15 ± 0.03st 2.00 ± 0.03t 1.00 ± 0.03v 3.35 ± 0.08l-o 2.10 ± 0.20H

M 1 3.00 ± 0.11opq 3.70 ± 0.05h-l 4.00 ± 0.10ghi 3.20 ± 0.05no 4.60 ± 0.09de 3.70 ± 0.16D

M 3 4.50 ± 0.04def 5.15 ± 0.14bc 3.50 ± 0.03j-n 4.85 ± 0.09cd 4.55 ± 0.09de 4.51 ± 0.15B

M 5 5.25 ± 0.09bc 5.50 ± 0.07b 5.50 ± 0.08b 6.50 ± 0.19a 3.00 ± 0.05opq 5.15 ± 0.31A

M 6 3.40 ± 0.10k-o 3.30 ± 0.03l-o 3.00 ± 0.04opq 3.10 ± 0.06nop 3.50 ± 0.08j-n 3.26 ± 0.06E

M 7 2.55 ± 0.07rs 2.60 ± 0.05qr 3.25 ± 0.07mno 3.95 ± 0.14ghi 3.30 ± 0.05l-o 3.13 ± 0.14EF

B= Boiling on burner , M= heating in microwave; BM= boiling method

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison

among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

Table - 52: Means comparison of yield for sapodilla fruit peel after 60 min of boiling using 1N HCl (Mechanical procedure

pH interaction mean±SE)

pH level Mechanical procedure Mean

Homogenizing Grinding Cutting Chopping Hammering

1 1.15 ± 0.38fg 2.20 ± 0.36c 0.35 ± 0.13hi 1.80 ± 0.78d 0.30 ± 0.13ij 1.16 ± 0.23B

3 1.63 ± 0.55e 4.08 ± 0.42a 0.43 ± 0.15hi 0.15 ± 0.02jk 0.10 ± 0.02k 1.28 ± 0.31A

5 1.23 ± 0.48f 2.15 ± 0.52c 0.08 ± 0.01k 0.48 ± 0.02h 1.78 ± 0.73de 1.14 ± 0.24B

6 3.03 ± 0.20b 1.00 ± 0.09g 0.30 ± 0.13ij 0.50 ± 0.22h 0.15 ± 0.05jk 1.00 ± 0.21C

7 1.78 ± 0.77de 1.63 ± 0.03e 0.08 ± 0.01k 0.35 ± 0.11hi 0.35 ± 0.11hi 0.84 ± 0.20D

Mean 1.76 ± 0.25B 2.21 ± 0.24A 0.25 ± 0.05E 0.66 ± 0.19C 0.54 ± 0.18D In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table - 53: Means comparison of yield for sapodilla fruit peel after 60 min of boiling using 1N HCl (Mechanical procedure

boiling method interaction mean±SE)

127

Boiling Mechanical procedure Mean

Method Homogenizing Grinding Cutting Chopping Hammering

B 1.56 ± 0.42c 1.60 ± 0.22c 0.32 ± 0.08f 0.16 ± 0.04g 0.19 ± 0.06g 0.77 ± 0.12B

M 1.96 ± 0.27b 2.82 ± 0.36a 0.17 ± 0.06g 1.15 ± 0.33d 0.88 ± 0.34e 1.40 ± 0.17A

B= Boiling on burner , M= heating in microwave

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table - 54: Means comparison of yield for sapodilla fruit peel after 60 min of boiling using 1N HCl (Mechanical procedure

pH boiling method interaction mean±SE)

BM pH Mechanical procedure BM pH

Homogenizing Grinding Cutting Chopping Hammering interaction mean

B 1 0.30 ± 0.01r-u 1.40 ± 0.05kl 0.65 ± 0.03op 0.05 ± 0.01vw 0.60 ± 0.02opq 0.60 ± 0.12E

B 3 0.40 ± 0.02q-t 3.15 ± 0.07de 0.75 ± 0.03o 0.20 ± 0.01t-w 0.05 ± 0.00vw 0.91 ± 0.31D

B 5 0.15 ± 0.01uvw 1.00 ± 0.03mn 0.10 ± 0.01uvw 0.45 ± 0.01p-s 0.15 ± 0.01uvw 0.37 ± 0.09F

B 6 3.45 ± 0.09bc 0.80 ± 0.03no 0.00 ± 0.00w 0.00 ± 0.00w 0.05 ± 0.01vw 0.86 ± 0.36D

B 7 3.50 ± 0.08bc 1.65 ± 0.03j 0.10 ± 0.01uvw 0.10 ± 0.01uvw 0.10 ± 0.01uvw 1.09 ± 0.36C

M 1 2.00 ± 0.05i 3.00 ± 0.08ef 0.05 ± 0.00vw 3.55 ± 0.07b 0.00 ± 0.00w 1.72 ± 0.39B

M 3 2.85 ± 0.05f 5.00 ± 0.10a 0.10 ± 0.01uvw 0.10 ± 0.01uvw 0.15 ± 0.01uvw 1.64 ± 0.53B

M 5 2.30 ± 0.09h 3.30 ± 0.12cd 0.05 ± 0.00vw 0.50 ± 0.02pqr 3.40 ± 0.05bc 1.91 ± 0.37A

M 6 2.60 ± 0.07g 1.20 ± 0.04lm 0.60 ± 0.03opq 1.00 ± 0.05mn 0.25 ± 0.02s-v 1.13 ± 0.22C

M 7 0.05 ± 0.01vw 1.60 ± 0.05jk 0.05 ± 0.00vw 0.60 ± 0.02opq 0.60 ± 0.02opq 0.58 ± 0.15E B= Boiling on burner , M= heating in microwave; BM= boiling method

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

128

Table - 55: Percentage yield of pectin from sapodilla fruit peel after using different

organic acid

Mechanical

procedure Inorganic acid

% of inorganic

acid pH

% yield

B M h

om

og

en

izin

g

Citric acid 1%

3 to 5

2.55 3.25

10% 2.55 4.24

Oxalic acid 1% 3.95 4.95

10% 0.5 3.7

Tartaric acid 1% 1.8 3.25

10% 2.5 2.55

ch

op

pin

g

Citric acid 1% 2.4 4.3

10% 0.05 0.1

Oxalic acid 1% 0.05 1.5

10% 0.15 1.85

Tartaric acid 1% 0.75 1.25

10% 0.35 3.2

Grin

din

g

Citric acid 1% 1.35 2.15

10% 0.05 1.2

Oxalic acid 1% 0.1 0.3

10% 1.25 1.05

Tartaric acid 1% 1.8 2.43

10% 1.65 3.15

Cu

ttin

g

Citric acid 1% 1.4 2.64

10% 0.1 1.1

Oxalic acid 1% 0.05 1.2

10% 0.13 1.32

Tartaric acid 1% 0.2 1.87

10% 0.32 1.45

Ha

mm

erin

g

Citric acid 1% 1.6 1.8

10% 0.54 1.67

Oxalic acid 1% 0.1 1.3

10% 0.43 2

Tartaric acid 1% 0.33 1.4

10% 0.44 2.1

B= Boiling on burner , M= heating in microwave

129

Table - 56 : Analysis of variance (mean squares) of yield of pectin from sapodilla

fruit peel after using different organic acid

Source of

variation

Degrees of

freedom

Mean squares

Yield (citric acid) Yield (oxalic

acid)

Yield (tartaric

acid)

BM

MP

Acid%

BM x MP

BM x Acid%

MP x Acid%

BM x MP x Acid%

Error

Total

1

4

1

4

1

4

4

40

59

14.5829**

7.7555**

21.0278**

0.1237**

0.0049NS

5.6981**

1.0202**

0.0126

23.2877**

14.9075**

0.1882**

1.8114**

0.9077**

5.0161**

0.7483**

0.0090

23.4750**

6.0553**

1.0375**

0.3754**

0.5245**

0.3919**

1.5337**

0.0101

NS = Non-significant (P>0.05); * = Significant (P<0.05); ** = Highly significant (P<0.01) BM= Boiling method. MP= Mechanical procedure

Table - 57: Means comparison of yield for sapodilla fruit peel after using citric acid

(Boiling method x mechanical procedure interaction mean±SE)

Mechanical

procedure

Boiling Method Mean

B M

Homogenizing 2.55 ± 0.05b 3.75 ± 0.23a 3.15 ± 0.21A

Chopping 1.23 ± 0.53e 2.20 ± 0.94c 1.71 ± 0.54B

Grinding 0.70 ± 0.29f 1.68 ± 0.21d 1.19 ± 0.23D

Cutting 0.75 ± 0.29f 1.87 ± 0.34d 1.31 ± 0.27CD

Hammering 1.07 ± 0.24e 1.74 ± 0.04d 1.40 ± 0.15C

Mean 1.26 ± 0.18B 2.25 ± 0.24A

B= Boiling on burner , M= heating in microwave.In a row and column statistically non-significant (P>0.05)

represents mean sharing and are denoted by similar letters. Comparison among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

Table - 58: Means comparison of yield for sapodilla fruit peel after using citric acid

Acid (% x boiling method interaction mean±SE)

Inorganic Boiling Method Mean

130

acid B M

1% 1.86 ± 0.14b 2.83 ± 0.24a 2.34 ± 0.16A

10% 0.66 ± 0.26d 1.66 ± 0.37c 1.16 ± 0.24B B= Boiling on burner , M= heating in microwave Among the two concentrations of acid 1% has found more effective with microwave as heating procedure

Table - 59: Means comparison of yield for sapodilla fruit peel after using citric acid

(Boiling method x Acid% x mechanical procedure interaction mean±SE)

Acid% x MP Boiling Method

Mean B M

1% x MP1 2.55 ± 0.08c 3.25 ± 0.10b 2.90 ± 0.17B

1% x MP2 2.40 ± 0.09cd 4.30 ± 0.16a 3.35 ± 0.43A

1% x MP3 1.35 ± 0.04fg 2.15 ± 0.04d 1.75 ± 0.18D

1% x MP4 1.40 ± 0.02fg 2.64 ± 0.04c 2.02 ± 0.28C

1% x MP5 1.60 ± 0.04ef 1.80 ± 0.05e 1.70 ± 0.05D

10% x MP1 2.55 ± 0.08c 4.24 ± 0.12a 3.40 ± 0.38A

10% x MP2 0.05 ± 0.00i 0.10 ± 0.00i 0.08 ± 0.01G

10% x MP3 0.05 ± 0.00i 1.20 ± 0.02g 0.63 ± 0.26F

10% x MP4 0.10 ± 0.00i 1.10 ± 0.01g 0.60 ± 0.22F

10% x MP5 0.54 ± 0.01h 1.67 ± 0.05ef 1.11 ± 0.25E

BM= Boiling method. MP= Mechanical procedure; B= Boiling on burner , M= heating in microwave

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by

similar letters. Comparison among interaction means are represented by lower case alphabets while capital

alphabets are used for overall mean

Table - 60: Means comparison of yield for sapodilla fruit peel after using oxalic acid

(Boiling method x mechanical procedure interaction mean±SE)

Mechanical

procedure

Boiling Method Mean

B M

Homogenizing 2.23 ± 0.77b 4.33 ± 0.29a 3.28 ± 0.51A

Chopping 0.10 ± 0.02f 1.68 ± 0.08c 0.89 ± 0.24B

Grinding 0.68 ± 0.26e 0.68 ± 0.17e 0.68 ± 0.15C

Cutting 0.09 ± 0.02f 1.26 ± 0.03d 0.68 ± 0.18C

Hammering 0.27 ± 0.07f 1.65 ± 0.16c 0.96 ± 0.22B

Mean 0.67 ± 0.21B 1.92 ± 0.24A

131

B= Boiling on burner , M= heating in microwave; B= Boiling on burner , M= heating in microwave

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by

similar letters. Comparison among interaction means are represented by lower case alphabets while capital

alphabets are used for overall mean

Table - 61: Means comparison of yield for sapodilla fruit peel after using oxalic acid

Acid% x boiling method interaction mean±SE

Inorganic

acid

Boiling Method Mean

B M

1% 0.85 ± 0.42 1.85 ± 0.43 1.35 ± 0.31A

10% 0.49 ± 0.11 1.98 ± 0.25 1.24 ± 0.19B

B= Boiling on burner , M= heating in microwave

Among the two concentrations of acid 1% has found more effective with microwave as

heating procedure

Table - 62: Means comparison of yield for sapodilla fruit peel after using oxalic acid

(Boiling method x Acid% x mechanical procedure interaction mean±SE)

Acid% x MP Boiling Method

Mean B M

1% x MP1 3.95 ± 0.15b 4.95 ± 0.14a 4.45 ± 0.24A

1% x MP2 0.05 ± 0.00h 1.50 ± 0.02d 0.78 ± 0.32E

1% x MP3 0.10 ± 0.01h 0.30 ± 0.01fgh 0.20 ± 0.05F

1% x MP4 0.05 ± 0.00h 1.20 ± 0.01e 0.63 ± 0.26E

1% x MP5 0.10 ± 0.01h 1.30 ± 0.02de 0.70 ± 0.27E

10% x MP1 0.50 ± 0.02f 3.70 ± 0.10b 2.10 ± 0.72B

10% x MP2 0.15 ± 0.01gh 1.85 ± 0.03c 1.00 ± 0.38D

10% x MP3 1.25 ± 0.04de 1.05 ± 0.02e 1.15 ± 0.05CD

10% x MP4 0.13 ± 0.01h 1.32 ± 0.02de 0.73 ± 0.27E

10% x MP5 0.43 ± 0.02fg 2.00 ± 0.05c 1.22 ± 0.35C

BM= Boiling method. MP= Mechanical procedure ; B= Boiling on burner , M= heating in microwave

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by

similar letters. Comparison among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

132

Table - 63: Means comparison of yield for sapodilla fruit peel after using tartaric

acid (Boiling method x mechanical procedure interaction mean±SE)

Mechanical

procedure

Boiling Method Mean

B M

homogenizing 2.15 ± 0.16b 2.90 ± 0.17a 2.53 ± 0.16A

Chopping 0.55 ± 0.09d 2.23 ± 0.44b 1.39 ± 0.33C

Grinding 1.73 ± 0.04c 2.79 ± 0.17a 2.26 ± 0.18B

Cutting 0.26 ± 0.03e 1.66 ± 0.10c 0.96 ± 0.22D

Hammering 0.39 ± 0.03de 1.75 ± 0.16c 1.07 ± 0.22D

Mean 1.01 ± 0.15B 2.27 ± 0.14A

B= Boiling on burner , M= heating in microwave

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by

similar letters. Comparison among interaction means are represented by lower case alphabets while capital

alphabets are used for overall mean

Table - 64: Means comparison of yield for sapodilla fruit peel after using tartaric

acid (Acid% x boiling method interaction mean±SE)

Inorganic

acid

Boiling Method Mean

B M

1% 0.98 ± 0.19c 2.04 ± 0.20b 1.51 ± 0.17B

10% 1.05 ± 0.24c 2.49 ± 0.18a 1.77 ± 0.20A

B= Boiling on burner , M= heating in microwave

Among the two concentrations of acid 1% has found more effective with microwave as heating procedure

133

Table - 65: Means comparison of yield for sapodilla fruit peel after using tartaric

acid (Boiling method x Acid% x mechanical procedure interaction mean±SE)

Acid% x MP Boiling Method

Mean B M

1% x PM1 1.80 ± 0.03cd 3.25 ± 0.08a 2.53 ± 0.33A

1% x PM2 0.75 ± 0.02g 1.25 ± 0.02f 1.00 ± 0.11E

1% x PM3 1.80 ± 0.05cd 2.43 ± 0.09b 2.12 ± 0.15B

1% x PM4 0.20 ± 0.02h 1.87 ± 0.08cd 1.04 ± 0.38E

1% x PM5 0.33 ± 0.02h 1.40 ± 0.02ef 0.87 ± 0.24E

10% x PM1 2.50 ± 0.07b 2.55 ± 0.12b 2.53 ± 0.06A

10% x PM2 0.35 ± 0.02h 3.20 ± 0.08a 1.78 ± 0.64C

10% x PM3 1.65 ± 0.03de 3.15 ± 0.10a 2.40 ± 0.34A

10% x PM4 0.32 ± 0.02h 1.45 ± 0.02ef 0.89 ± 0.25E

10% x PM5 0.44 ± 0.03h 2.10 ± 0.05c 1.27 ± 0.37D

B= Boiling on burner, M= heating in microwave; BM= Boiling method. MP= Mechanical procedure

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by

similar letters. Comparison among interaction means are represented by lower case alphabets while capital

alphabets are used for overall mean

134

Table - 66: percentage yield of pectin from sapodilla fruit peel using different

strengths of same inorganic acid.

Mechanical

procedure pH

0.1N HCl 0.5N HCl 1N HCl

B M B M B M

Hammering

1 1.5 2.05 0.65 1 2.65 3.15

3 0.6 1.55 0.05 3.3 5.65 6.55

5 0.8 1.95 0.3 2.2 2.3 4.6

6 0.8 1.4 0.45 1.65 2.15 2.95

7 0.5 0 0.25 0.05 2.5 3.1

Grinding

1 0.55 0.8 0.3 4.6 3.1 4.15

3 4 3.5 0.1 2.85 5.6 4.8

5 2.45 2.9 0.1 1 1.05 6.1

6 2 2 3.15 4.05 1.45 3.55

7 1.95 2.3 2.5 2.8 0.35 2.75

Cutting

1 0.8 0.9 3.65 4.2 0.05 0.05

3 1.1 1.5 4 2.25 0.05 1.6

5 0.4 1 4.35 3.75 2.95 5.3

6 1 1.2 0.55 4.75 0.15 3.5

7 0.7 1 1.25 0.5 0.05 1.2

Homogenizing

1 0.5 1.05 4.15 4.95 1.55 4.95

3 0.75 1.6 3.2 4.95 2.4 5.35

5 1.2 1.5 0.15 4.55 1.7 4.85

6 1.5 2 1.8 0.3 0.6 1.5

7 0.5 0.5 3.6 0.6 1.2 2.55

Chopping

(mortor/pastle

1 2.7 4.7 4.4 4.65 3 5.5

3 3.9 3.45 4.8 4.95 3.7 5.4

5 3.95 4.05 2.25 2.65 4.1 6.7

6 2.3 2.45 1.5 0.65 3.75 3.8

7 3.95 2.4 1 0.1 0.05 0.25

BM= Boiling method. MP= Mechanical procedure B= Boiling on burner , M= heating in microwave

133

Table - 67: Analysis of variance (mean squares) of yield of pectin from sapodilla fruit peel after using different strength of

same inorganic acid

Source of

variation

Degrees of

freedom

Mean squares

Yield_0.1N HCl Yield_0.5N HCl Yield_1N HCl

MP

pH

BM

MP x pH

MP x BM

pH x BM

MP x pH x BM

Error

Total

4

4

1

16

4

4

16

100

149

32.6575**

3.3917**

3.2413**

2.5462**

0.3392**

1.0112**

0.6258**

0.0093

19.2084**

20.5359**

21.2064**

8.7942**

4.8684**

6.8626**

5.2626**

0.0104

24.375**

37.244**

109.739**

7.934**

1.832**

4.535**

2.303**

0.047

NS = Non-significant (P>0.05); * = Significant (P<0.05); ** = Highly significant (P<0.01) B= Boiling on burner , M= heating in

microwave; BM= Boiling method. MP= Mechanical procedure

134

Table - 68: Means comparison for yield of pectin from sapodilla fruit peel after using 0.1N HCl (Mechanical procedure pH

interaction mean±SE )

pH level Mechanical procedure Mean

Homogenizing Grinding Cutting Chopping Hammering

1 0.78 ± 0.12k 0.68 ± 0.06kl 0.85 ± 0.03k 3.70 ± 0.45b 1.78 ± 0.12g 1.56 ± 0.23D

3 1.18 ± 0.19hij 3.75 ± 0.13b 1.30 ± 0.09hi 3.68 ± 0.12b 1.08 ± 0.21j 2.20 ± 0.24A

5 1.35 ± 0.07h 2.68 ± 0.10d 0.70 ± 0.13kl 4.00 ± 0.09a 1.38 ± 0.26h 2.02 ± 0.23B

6 1.75 ± 0.11g 2.00 ± 0.04f 1.10 ± 0.05ij 2.38 ± 0.05e 1.10 ± 0.14ij 1.67 ± 0.10C

7 0.50 ± 0.01l 2.13 ± 0.09f 0.85 ± 0.07k 3.18 ± 0.35c 0.25 ± 0.11m 1.38 ± 0.22E

Mean 1.11 ± 0.09C 2.25 ± 0.19B 0.96 ± 0.05D 3.39 ± 0.15A 1.12 ± 0.12C

B= Boiling on burner , M= heating in microwave; BM= Boiling method. MP= Mechanical procedure

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison

among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

Table - 69: Means comparison for yield of pectin from sapodilla fruit peel after using 0.1N HCl (Mechanical procedure

boiling method interaction mean±SE)

Boiling Mechanical procedure Mean

Method Homogenizing Grinding Cutting Chopping Hammering

B 0.89 ± 0.11e 2.19 ± 0.30b 0.80 ± 0.07e 3.36 ± 0.19a 0.84 ± 0.09e 1.62 ± 0.14B

M 1.33 ± 0.14c 2.30 ± 0.24b 1.12 ± 0.06d 3.41 ± 0.24a 1.39 ± 0.20c 1.91 ± 0.13A

B= Boiling on burner, M= heating in microwave; BM= Boiling method. MP= Mechanical procedure In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are

represented by lower case alphabets while capital alphabets are used for overall mean

Table - 70: Means comparison for yield of pectin from sapodilla fruit peel after using 0.1N HCl (Mechanical procedure pH

boiling method interaction mean±SE)

BM pH Mechanical procedure BM pH

Homogenizing Grinding Cutting Chopping Hammering interaction mean

B 1 0.50 ± 0.02qr 0.55 ± 0.01qr 0.80 ± 0.02m-q 2.70 ± 0.05de 1.50 ± 0.04ij 1.21 ± 0.22F

135

B 3 0.75 ± 0.02n-q 4.00 ± 0.10b 1.10 ± 0.03klm 3.90 ± 0.10b 0.60 ± 0.03pqr 2.07 ± 0.41B

B 5 1.20 ± 0.03jkl 2.45 ± 0.04ef 0.40 ± 0.01r 3.95 ± 0.11b 0.80 ± 0.04m-q 1.76 ± 0.35D

B 6 1.50 ± 0.02ij 2.00 ± 0.05gh 1.00 ± 0.03l-o 2.30 ± 0.05fg 0.80 ± 0.03m-q 1.52 ± 0.15E

B 7 0.50 ± 0.01qr 1.95 ± 0.08h 0.70 ± 0.01o-r 3.95 ± 0.10b 0.50 ± 0.01qr 1.52 ± 0.36E

M 1 1.05 ± 0.03lmn 0.80 ± 0.01m-q 0.90 ± 0.02l-p 4.70 ± 0.10a 2.05 ± 0.03gh 1.90 ± 0.39C

M 3 1.60 ± 0.02i 3.50 ± 0.10c 1.50 ± 0.02ij 3.45 ± 0.08c 1.55 ± 0.05i 2.32 ± 0.25A

M 5 1.50 ± 0.03ij 2.90 ± 0.05d 1.00 ± 0.03l-o 4.05 ± 0.15b 1.95 ± 0.03h 2.28 ± 0.29A

M 6 2.00 ± 0.03gh 2.00 ± 0.08gh 1.20 ± 0.02jkl 2.45 ± 0.08ef 1.40 ± 0.05ijk 1.81 ± 0.12CD

M 7 0.50 ± 0.01qr 2.30 ± 0.08fg 1.00 ± 0.01l-o 2.40 ± 0.05ef 0.00 ± 0.00s 1.24 ± 0.26F

B= Boiling on burner , M= heating in microwave; BM= Boiling method. MP= Mechanical procedure

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison

among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

136

Table - 71:Means comparison for yield of pectin from sapodilla fruit peel after using 0.5N HCl (Mechanical procedure pH

interaction mean±SE)

pH level Mechanical procedure Mean

Homogenizing Grinding Cutting Chopping Hammering

1 4.55 ± 0.19b 2.45 ± 0.96fg 3.93 ± 0.13c 4.53 ± 0.08b 0.83 ± 0.08l 3.26 ± 0.32A

3 4.08 ± 0.39c 1.48 ± 0.62i 3.13 ± 0.39e 4.88 ± 0.08a 1.68 ± 0.73i 3.05 ± 0.32B

5 2.35 ± 0.99g 0.55 ± 0.20m 4.05 ± 0.14c 2.45 ± 0.10fg 1.25 ± 0.43j 2.13 ± 0.30C

6 1.05 ± 0.34jk 3.60 ± 0.21d 2.65 ± 0.94f 1.08 ± 0.19jk 1.05 ± 0.27jk 1.89 ± 0.28D

7 2.10 ± 0.67h 2.65 ± 0.07f 0.88 ± 0.17kl 0.55 ± 0.20m 0.15 ± 0.05n 1.27 ± 0.22E

Mean 2.83 ± 0.34B 2.15 ± 0.29D 2.93 ± 0.29A 2.70 ± 0.33C 0.99 ± 0.19E

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

Table - 72: Means comparison for yield of pectin from sapodilla fruit peel after using 0.5N HCl (Mechanical procedure

boiling method interaction mean±SE)

Boiling Mechanical procedure Mean

Method Homogenizing Grinding Cutting Chopping Hammering

B 2.58 ± 0.39c 1.23 ± 0.35e 2.76 ± 0.41b 2.79 ± 0.41b 0.34 ± 0.05f 1.94 ± 0.19B

M 3.07 ± 0.57a 3.06 ± 0.33a 3.09 ± 0.41a 2.60 ± 0.53c 1.64 ± 0.29d 2.69 ± 0.20A B= Boiling on burner , M= heating in microwave; BM= Boiling method. MP= Mechanical procedure In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

Table - 73: Means comparison for yield of pectin from sapodilla fruit peel after using 0.5N HCl (Mechanical procedure pH

boiling method interaction mean±SE)

BM pH Mechanical procedure BM pH

Homogenizing Grinding Cutting Chopping Hammering interaction mean

B 1 4.15 ± 0.09def 0.30 ± 0.02s-v 3.65 ± 0.07h 4.40 ± 0.05cd 0.65 ± 0.03r 2.63 ± 0.48D

B 3 3.20 ± 0.02j 0.10 ± 0.01v 4.00 ± 0.03fg 4.80 ± 0.10ab 0.05 ± 0.01v 2.43 ± 0.53E

B 5 0.15 ± 0.00uv 0.10 ± 0.01v 4.35 ± 0.10cde 2.25 ± 0.05n 0.30 ± 0.01s-v 1.43 ± 0.45H

B 6 1.80 ± 0.02o 3.15 ± 0.07jk 0.55 ± 0.00rst 1.50 ± 0.04op 0.45 ± 0.02r-u 1.49 ± 0.26H

137

B 7 3.60 ± 0.08hi 2.50 ± 0.05mn 1.25 ± 0.03pq 1.00 ± 0.02q 0.25 ± 0.02tuv 1.72 ± 0.32G

M 1 4.95 ± 0.12a 4.60 ± 0.05bc 4.20 ± 0.05def 4.65 ± 0.11abc 1.00 ± 0.03q 3.88 ± 0.39A

M 3 4.95 ± 0.05a 2.85 ± 0.06kl 2.25 ± 0.03n 4.95 ± 0.13a 3.30 ± 0.11ij 3.66 ± 0.30B

M 5 4.55 ± 0.12bc 1.00 ± 0.02q 3.75 ± 0.05gh 2.65 ± 0.08lm 2.20 ± 0.08n 2.83 ± 0.33C

M 6 0.30 ± 0.01s-v 4.05 ± 0.11efg 4.75 ± 0.05ab 0.65 ± 0.03r 1.65 ± 0.03o 2.28 ± 0.48F

M 7 0.60 ± 0.02rs 2.80 ± 0.05lm 0.50 ± 0.01rst 0.10 ± 0.01v 0.05 ± 0.01v 0.81 ± 0.27I B= Boiling on burner , M= heating in microwave; BM= Boiling method. MP= Mechanical procedure

In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison

among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

138

Table - 74: Means comparison for yield of pectin from sapodilla fruit peel after using 1N HCl (Mechanical procedure pH

interaction mean±SE)

pH level Mechanical procedure Mean

Homogenizing Grinding Cutting Chopping Hammering

1 3.25 ± 0.76ghi 3.63 ± 0.24fg 0.05 ± 0.00n 4.25 ± 0.56cd 2.90 ± 0.12hij 2.82 ± 0.33B

3 3.88 ± 0.66def 5.20 ± 0.19b 0.83 ± 0.35lm 4.55 ± 0.39c 6.10 ± 0.23a 4.11 ± 0.37A

5 3.28 ± 0.71gh 3.58 ± 1.13fg 4.13 ± 0.53cde 5.40 ± 0.59b 3.45 ± 0.52fg 3.97 ± 0.34A

6 1.05 ± 0.20l 2.50 ± 0.47j 1.83 ± 0.75k 3.78 ± 0.04ef 2.55 ± 0.19j 2.34 ± 0.24C

7 1.88 ± 0.30k 1.55 ± 0.54k 0.63 ± 0.26lm 0.48 ± 0.35mn 2.80 ± 0.14ij 1.47 ± 0.21D

Mean 2.67 ± 0.31C 3.29 ± 0.34B 1.49 ± 0.33D 3.69 ± 0.36A 3.56 ± 0.27A In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

Table - 75: Means comparison for yield of pectin from sapodilla fruit peel after using 1N HCl (Mechanical procedure

boiling method interaction mean±SE)

Boiling Mechanical procedure Mean

Method Homogenizing Grinding Cutting Chopping Hammering

B 1.49 ± 0.16f 2.31 ± 0.50e 0.65 ± 0.31g 2.92 ± 0.40d 3.05 ± 0.35d 2.08 ± 0.19B

M 3.84 ± 0.41c 4.27 ± 0.31ab 2.33 ± 0.50e 4.46 ± 0.55a 4.07 ± 0.37bc 3.79 ± 0.21A

B= Boiling on burner , M= heating in microwave; BM= Boiling method. MP= Mechanical procedure In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

Table - 76: Means comparison for yield of pectin from sapodilla fruit peel after using 1N HCl (Mechanical procedure pH

boiling method interaction mean±SE)

BM pH Mechanical procedure BM pH

Homogenizing Grinding Cutting Chopping Hammering interaction mean

B 1 1.55 ± 0.05rst 3.10 ± 0.03i-m 0.05 ± 0.01w 3.00 ± 0.08j-n 2.65 ± 0.05l-o 2.07 ± 0.31F

139

B 3 2.40 ± 0.02m-p 5.60 ± 0.13bc 0.05 ± 0.01w 3.70 ± 0.10hij 5.65 ± 0.15bc 3.48 ± 0.56C

B 5 1.70 ± 0.03p-s 1.05 ± 0.02s-v 2.95 ± 0.04k-n 4.10 ± 0.09gh 2.30 ± 0.03n-q 2.42 ± 0.28E

B 6 0.60 ± 0.01uvw 1.45 ± 0.03rst 0.15 ± 0.01w 3.75 ± 0.05hi 2.15 ± 0.07o-r 1.62 ± 0.34G

B 7 1.20 ± 0.01stu 0.35 ± 0.01vw 0.05 ± 0.00w 0.05 ± 0.01w 2.50 ± 0.10l-o 0.83 ± 0.25H

M 1 4.95 ± 0.12cde 4.15 ± 0.10fgh 0.05 ± 0.00w 5.50 ± 0.10bcd 3.15 ± 0.08i-l 3.56 ± 0.52C

M 3 5.35 ± 0.12cd 4.80 ± 0.08d-g 1.60 ± 0.05q-t 5.40 ± 0.10bcd 6.55 ± 0.20a 4.74 ± 0.45B

M 5 4.85 ± 0.13def 6.10 ± 0.12ab 5.30 ± 0.10cde 6.70 ± 0.18a 4.60 ± 0.12efg 5.51 ± 0.22A

M 6 1.50 ± 0.03rst 3.55 ± 0.09h-k 3.50 ± 0.09h-k 3.80 ± 0.08hi 2.95 ± 0.10k-n 3.06 ± 0.22D

M 7 2.55 ± 0.09l-o 2.75 ± 0.05l-o 1.20 ± 0.03stu 0.92 ± 0.66tuv 3.10 ± 0.06i-m 2.10 ± 0.26F

B= Boiling on burner , M= heating in microwave; BM= Boiling method. MP= Mechanical procedure In a row and column statistically non-significant (P>0.05) represents mean sharing and are denoted by similar letters. Comparison among interaction means are represented by lower case alphabets while capital alphabets are used for overall mean

140

Table - 77: Identification tests for the presence of pectin from three selected fruits

Test Name Banana Sapodilla Muskmelon

Stiff gel test - + -

Test with 95% Ethanol + + Not clear

Test with Potassium Hydroxide (KOH) + + +

Iodine test + + +

Table - 78: Biochemical Characterization of the purified pectin extracted from

sapodilla pectin at different pH

Test Name Sapodilla

( pH5)

Sapodilla

(pH3)

Sapodilla

(pH1) FGP*

Quantitative test for ammonia. No

ammonia

No

ammonia

No

ammonia No ammonia

Moisture (%) 5.29 6.02 5.52 7.02

Ash (%) 5.11 4.3 4.89 1.16

Equivalent weight 1700 2680 2290 1271

Methoxyl Content (%) 5.1 4.4 4.9 8.16

Anhydrouronic Acid. (%) 39.33 31.78 34.56 70.50

Jelly Grade 100 99 98 150

Galacturonic Acid Content (%) 77.7 65.8 60.5 76.19

Protien 1.71 3.36 4.04

Degree of esterificatoion (%) 73.63 72.1 70.2 76.41

*FGP = Food grade pectin

141

Table -79: Water holding, water binding and fat binding capacity of sapodilla pectin

Test Name Sapodilla peel pectin Apple

pectin

Orange

pectin

Fibers

Apple Orange Grape

Fruit

WBC g/g 7.6 - - 1.62 1.65 2.09

WHC g/g 6.99 16.51 28.07 - - -

FBC g/g 2.2 - - 0.95 1.81 1.52

Table – 80: Showing FTIR spectral values of sample and standard pectin along with

associated functional groups.

Functional groups FOOD

GRADE

Sapodilla

(pH5)

Sapodilla

(pH3)

Sapodilla

(pH1)

O-H stretching 3319.2 3287.3 3271.1 3289.2

C-H stretching , symmetric,

asymmetric 2932.4 2923.6 2929.6 2930.2

C=O esterified 1733.3 1735.6 1738.6 1737.8

COO- asymmetric stretching

1622.2 1611.2 1612.2

COO- symmetric stretching 1426.4 1424.3 1423.3 1425.8

C-H bending 1335.4 1362.0 1363.0

C=O stretching 1234.9 1234.1 1234.5 1228.6

142

Figure – 23: FTIR spectra of food grade pectin

Figure – 24: FTIR spectra of sapodilla peel pectin extracted at pH 5

403.

3

467.

1

523.

2

679.

9

858.

2

909.

2

995.

2

1052

.6

1234

.9

1273

.913

35.4

1426

.4

1733

.3

1996

.0

2068

.9

2349

.0

2932

.4

3319

.2

3554

.1

*Pectin f ood grade

62

64

66

68

70

72

74

76

78

80

82

84

86

88

90

92

94

96

98

100

102

%T

500 1000 1500 2000 2500 3000 3500 4000

Wavenumbers (cm-1)

143

Figure – 25: FTIR spectra of sapodilla peel pectin extracted at pH 3

Figure – 26: FTIR spectra of Sapodilla peel pectin extracted at pH 1

DLS studies

Table – 81: Summary the physical characterization of standard pectin and the

different pectin extracted from various source and at different pH by DLS

144

Sample Extraction at

pH

Time of

extraction RH (nm)

PD (%)

(Đ)

Standard -- -- 73.7-1979.92 8.6-37.9

Apple 5.0 10 94.37-1357 18.4-47.8

Orange 5.0 10 473.07-482.92 36.8-36.9

Sapodilla 3.0 10 464.37-548.53 29.3-30.1

Sapodilla 5.0 10 390.21-421.17 28.1-29.3

RH= Radius of hydration ; PD = polydispersity

145

Standard

Figure – 27: Physical characterization of standard pectin by DLS. (A) DLS results of

standard pectin illustrating the experimental conditions i.e., the mean autocorrelation

function (a), monodispersity and radius plot (b-c), respectively. (B)Comparative

corresponding radius distribution of pectin standard (a) and standard pectin (b). All

experiments were performed with an auto–piloted run of 50 measurements at every 20 s,

with a wait time 1 s (at 25 °C).

Figure- 28: Physical characterization of apple pectin extracted at pH5.0. by DLS. (A)

DLS results of apple pectin extracted at pH5.0. Illustrating the experimental conditions

i.e., the mean autocorrelation function (a), monodispersity and radius plot (b-c),

respectively. (B) Comparative corresponding radius distribution of pectin standard (a)

and apple pectin extracted at pH5.0 (b). All experiments were performed with an auto–

piloted run of 50 measurements at every 20 s, with a wait time 1 s (at 25 °C).

146

Figure – 29: Physical characterization of Orange pectin extracted at pH5.0. by DLS. (A)

DLS results of Orange pectin extracted at pH5.0. Illustrating the experimental conditions

i.e., the mean autocorrelation function (a), monodispersity and radius plot (b-c),

respectively. (B) Comparative corresponding radius distribution of pectin standard (a)

and Orange pectin extracted at pH5.0 (b). All experiments were performed with an auto–

piloted run of 50 measurements at every 20 s, with a wait time 1 s (at 25 °C).

147

Figure – 30: Physical characterization of sapodilla pectin extracted at pH3.0. by DLS.

(A) DLS results of sapodilla pectin extracted at pH3.0. illustrating the experimental

conditions i.e., the mean autocorrelation function (a), monodispersity and radius plot (b-

c), respectively. (B) Comparative corresponding radius distribution of pectin standard (a)

and sapodilla pectin extracted at pH3.0 (b). All experiments were performed with an

auto–piloted run of 50 measurements at every 20 s, with a wait time 1 s (at 25 °C).

Figure – 31: Physical characterization of sapodilla pectin extracted at pH5.0. by DLS.

(A) DLS results of sapodilla pectin extracted at pH5.0. illustrating the experimental

conditions i.e., the mean autocorrelation function (a), monodispersity and radius plot (b-

c), respectively. (B) Comparative corresponding radius distribution of pectin standard (a)

and sapodilla pectin extracted at pH5.0 (b). All experiments were performed with an

148

auto–piloted run of 50 measurements at every 20 s, with a wait time 1 s (at 25 °C). Also

see Table 5 for details.

Response surface methodology

Table - 82: Box-Bechen experimental design and levels of factors used for

optimization of pectin yield

Variables Symbol Low High

pH X1 1 5

Temperature(oC) X2 50 100

Time ( minutes) X3 10 90

Table - 83: Box-Behnken experimental design and corresponding results for

responses

Std Order Run Order Pt Type Blocks pH Temperature Time Yield

10 1 2 1 3 100 10 1.6

3 2 2 1 1 100 50 1.7

14 3 0 1 3 75 50 1.5

1 4 2 1 1 50 50 1.6

2 5 2 1 5 50 50 3.5

6 6 2 1 5 75 10 2.6

9 7 2 1 3 50 10 1.5

12 8 2 1 3 100 90 1.65

4 9 2 1 5 100 50 2.5

8 10 2 1 5 75 90 3.5

11 11 2 1 3 50 90 1.4

13 12 0 1 3 75 50 2

5 13 2 1 1 75 10 3

15 14 0 1 3 75 50 1.8

149

7 15 2 1 1 75 90 1.5

Table - 84: Analysis of Variance for Yield (Response surface methodology)

Source DF Adj SS Adj MS F-value Prob.

Model

Linear

pH

Temperature

Time

Square

pH*pH

Temperature*Temperature

Time*Time

2-Way Interaction

pH*Temperature

pH*Time

Temperature*Time

Error

Lack-of-Fit

Pure Error

Total

9

3

1

1

1

3

1

1

1

3

1

1

1

5

3

2

14

7.15996

2.40188

2.31125

0.03781

0.05281

3.00996

2.57694

0.28348

0.00848

1.74813

0.30250

1.44000

0.00563

0.51104

0.38438

0.12667

7.67100

0.79555

0.80063

2.31125

0.03781

0.05281

1.00332

2.57694

0.28348

0.00848

0.58271

0.30250

1.44000

0.00563

0.10221

0.12813

0.06333

7.78*

7.83*

22.61**

0.37

0.52

9.82*

25.21**

2.77

0.08

5.70*

2.96

14.09*

0.06

2.02

0.018

0.025

0.005

0.570

0.504

0.015

0.004

0.157

0.785

0.045

0.146

0.013

0.824

0.348

* = Significant (P<0.05); ** = Highly significant (P<0.01)

S = 0.3197

R² = 93.34%

R²(adj) = 81.35%

R²(pred) = 16.11%

150

Table - 85: Estimated Regression Coefficients for Yield (Box-Bechen experimental

design, response surface methodology)

Term Effect Coef. SE(Coef.) t-value Prob. VIF

Constant

pH

Temperature

Time

pH*pH

Temperature*Temperature

Time*Time

pH*Temperature

pH*Time

Temperature*Time

1.075

-0.137

-0.163

1.671

-0.554

0.096

-0.550

1.200

0.075

1.767

0.538

-0.069

-0.081

0.835

-0.277

0.048

-0.275

0.600

0.038

0.185

0.113

0.113

0.113

0.166

0.166

0.166

0.160

0.160

0.160

9.57**

4.76**

-0.61

-0.72

5.02**

-1.67

0.29

-1.72

3.75*

0.23

0.000

0.005

0.570

0.504

0.004

0.157

0.785

0.146

0.013

0.824

1.00

1.00

1.00

1.01

1.01

1.01

1.00

1.00

1.00

* = Significant (P<0.05); ** = Highly significant (P<0.01)

Yield = 0.76 - 0.947 pH + 0.0784 Temperature - 0.0303 Time + 0.2089 pH² -

0.000443 Temperature² + 0.000030 Time² - 0.00550 pH*Temperature

+ 0.00750 pH*Time + 0.000037 Temperature*Time

Table - 86: Predicted values of yield. (Box-Bechen experimental design, response

surface methodology)

Yield Composite

Solution pH Temperature Time Fit Desirability

1 5 61.1111 90 3.79087 1.00000

2 5 85.2577 86.8718 3.48497 0.99284

3 5 67.7177 17.6639 2.83699 0.68428

151

4 5 67.7177 17.6639 2.83699 0.68428

5 1 90.6592 10 2.79074 0.66226

Figure – 32: Showing the optimal conditions for the extraction of pectin from

sapodilla fruit peel (Box-Bechen experimental design, response surface

methodology)

152

Figure – 33: Response surface graph and contour plot of effect of pH and

temperature on yield of pectin at constant time

Yield = -0.6969-0.5663*x+0.0811*y+0.2079*x*x-0.0055*x*y-0.0004*y*y

> 3.5 < 3.5 < 3 < 2.5 < 2 < 1.5

Yield = Distance Weighted Least Squares

> 4 < 4 < 3.5 < 3 < 2.5 < 2 < 1.5 < 1 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 4.5 5.0 5.5

pH

40

50

60

70

80

90

100

110

Tem

pe

ratu

re

153

Figure- 34: Response surface graph and contour plot of effect of pH and time on

yield of pectin at constant temperature

Yield = 4.0523-1.3913*x-0.0289*y+0.2142*x*x+0.0075*x*y+4.3269E-5*y*y

> 4.5 < 4.5 < 4 < 3.5 < 3 < 2.5 < 2 < 1.5

Yield = Distance Weighted Least Squares

> 4 < 4 < 3.5 < 3 < 2.5 < 2 < 1.5 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 4.5 5.0 5.5

pH

0

10

20

30

40

50

60

70

80

90

100

Tim

e

154

Figure – 35: Response surface graph and contour plot of effect of temperature and

time on yield of pectin at constant pH

Yield = -0.3684+0.0773*x-0.0038*y-0.0005*x*x+3.75E-5*x*y-1.0216E-5*y*y

> 2.2 < 2.2 < 2 < 1.8 < 1.6

Yield = Distance Weighted Least Squares

> 2.5 < 2.5 < 2 < 1.5 < 1 < 0.5 40 50 60 70 80 90 100 110

Temperature

0

10

20

30

40

50

60

70

80

90

100

Tim

e

155

Table – 87: Flow properties of granules made for paracetamol tablets

Test

Formulatio

n

Mass

(g)

Bulk

Volume

(ml)

Tapped

Volume

(ml)

Bulk

Density

(g/ml)

Tapped

Density

(g/ml)

Angle of

Repose

(θ⁻¹)

Compressibilit

y Index

(%)

Hausner

Ratio

-

Standard 2.004±0.00

1

4.5068±0.11

5

4.123±0.12

5

0.446±0.05

1

0.487±0.05

7

17.955±1.15

8 8.41±0.507

1.091±0.00

6

F1 2.010±0.00

5 3.867±0.115

3.067±0.11

5

0.520±0.01

4

0.656±0.02

3

18.747±0.54

3 20.702±0.608

1.261±0.01

0

F2 2.003±0.00

1 3.867±0.115

3.067±0.11

5

0.518±0.01

5

0.654±0.02

4

16.494±1.68

0 20.702±0.608

1.261±0.01

0

F3 2.008±0.00

1 4.067±0.115

3.067±0.11

5

0.494±0.01

4

0.655±0.02

4

16.558±1.79

2 24.603±0.687

1.326±0.01

2

F4 2.008±0.00

1 4.333±0.115

4.133±0.11

5

0.464±0.01

2

0.486±0.01

4

17.545±1.54

0 4.618±0.125

1.048±0.00

1

F5 2.004±0.00

1 3.533±0.306

3.333±0.30

6

0.570±0.05

1

0.605±0.05

7

17.955±1.15

8 5.690±0.507

1.060±0.00

6

F6 2.005±0.00

1 4.133±0.115

3.667±0.46

2

0.485±0.01

4

0.552±0.06

5

17.453±0.17

1 11.032±13.884

1.141±0.16

3

F7 2.016±0.02

1 4.000±0.200

3.333±0.30

6

0.505±0.02

1

0.608±0.05

6

17.987±0.61

8 16.700±5.888

1.204±0.08

2

F8 2.020±0.01 4.067±0.115 3.067±0.11 0.497±0.01 0.659±0.02 21.546±0.44 24.603±0.687 1.326±0.01

156

0 5 4 5 1 2

F9 2.017±0.00

6 4.133±0.115

3.333±0.11

5

0.488±0.01

2

0.605±0.02

0

16.911±0.38

3 19.365±0.550

1.240±0.00

8

F1-F9 = test formulations 1 till 9 Each value is a Mean±SD of three determination

Table - 88: Pharmaceutical characteristics of compressed formulation of paracetamol tablet

Test Formulation

Pharmacopoeial Limits

(USP 32/NF 27)

Wt. Variation

(Mean ± S.D)

(mg)

±5%

Thickness

(Mean ±

S.D)

(mm)

±5%

Diameter

(Mean ± S.D)

(mm)

₋

Hardness

(Mean ± S.D)

(kg)

At least 5 kg

Loss on drying

%

Not more than 1.5%

Standard 702.67± 2.52 5.60±0.20 9.47±0.05 5.22±0.01 4.0%

F1 705.00±5.00 5.22±0.03 9.42±0.03 2.54±0.06 4.0%

F2 672.33±2.52 5.24±0.05 9.39±0.01 1.37±0.07 3.0%

F3 701.33±3.21 5.32±0.05 9.45±0.04 4.16±0.08 4.3%

F4 695±5.00 5.35±0.06 9.44±0.05 5.06±0.21 4.2%

F5 705±4.51 5.21±0.03 9.41±0.03 5.15±0.07 4.6%

F6 701.67±3.97 5.19±0.04 9.42±0.03 5.21±0.07 4.3%

F7 708.3±2.89 5.29±0.04 9.31±0.03 6.07±0.12 4.2%

F8 694.67±4.51 5.27±0.13 9.27±0.06 6.67±0.58 4.5%

157

F9 704.00±3.61 5.27±0.10 9.42±0.04 7.40±0.33 4.5%

F1-F9 = test formulations 1 till 9 Each value is a Mean±SD of three determination.

Table – 89: Dissolution studies of paracetamol tablet

FORMULATION NUMBER

F1 F2 F3 F4 F5 F6 F7 F8 F9

243nm

(Abs)

%

release

243nm

(Abs)

%

release

243nm

(Abs)

%

release

243nm

(Abs)

%

release

243nm

(Abs)

%

release

243nm

(Abs)

%

release

243nm

(Abs)

%

release

243nm

(Abs)

%

release

243nm

(Abs)

%

release

0.304 43.43 0.339 50.64 0.401 58.77 0.269 71.52 0.564 84.17 0.078 23.00 0.084 26.77 0.123 36.73 0.111 32.73

0.291 41.67 0.338 50.49 0.412 60.39 0.283 70.28 0.557 83.12 0.087 25.65 0.080 23.59 0.111 32.73 0.094 27.97

0.362 51.83 0.564 84.25 0.382 55.99 0.331 83.13 0.418 91.93 0.112 33.03 0.093 27.42 0.108 31.85 0.108 31.85

0.372 53.26 0.557 83.20 0.389 57.02 0.333 87.28 0.620 92.53 0.105 30.96 0.096 28.31 0.120 35.39 0.101 29.78

0.313 44.82 0.418 62.44 0.577 84.58 0.285 85.20 0.406 93.13 0.096 28.313 0.076 22.41 0.120 35.39 0.099 29.19

0.323 46.24 0.406 60.65 0.590 86.48 0.312 85.20 0.632 94.32 0.116 34.21 0.091 26.83 0.107 31.55 0.088 25.95

STANDARD FOR FORMULATION

0.692 99.10 0.664 99.10 0.676 99.10 0.478 99.10 0.664 99.10 0.336 99.10 0.336 99.10 0.336 99.10 0.336 99.10

F1-F9 are test formulation while Abs = absorbance and %release = percent release

158

159

F1 F2

F3

Figure -36: Formulated paracetamol tablest ( F1, F2, F3)

160

F4 F5

F6

Figure 37: Formulated paracetamol tablest ( F4, F5, F6)

161

F7 F8

F9

Figure 38: Formulated paracetamol tablest (F7, F8, F9)

162

Table - 90: Flow properties ofgranules for ibuprofen tablet

Test Formulation

Mass Bulk Volume Tapped Volume Bulk Density Tapped Density Angle of Repose Compressibility Index Hausner Ratio

(g) (ml) (ml) (g/ml) (g/ml) (θ⁻¹) (%) ₋

Std 2.02±0.013 4.25±0.015 3.75±0.092 0.475±0.002 0.538±0.020 11.71±0.210 11.71±0.116 1.132±0.061

R1 2.02±0.013 4.20±0.015 3.76±0.093 0.49±0.003 0.55±0.019 15.23±0.208 4.82±0.115 1.13±0.061

R2 2.01±0.016 4.35±0.042 3.61±0.010 0.46±0.005 0.56±0.004 13.83±0.153 9.74±0.344 1.24±0.059

R3 2.01±0.007 4.60±0.080 3.93±0.064 0.42±0.021 0.52±0.010 13.73±0.306 7.40±0.352 1.17±0.028

R4 2.01±0.004 4.51±0.090 4.15±0.050 0.44±0.002 0.47±0.003 11.13±0.153 3.77±0.252 1.09±0.009

R1= Test formulation one, R2= Test formulation two, R3= Test formulation three, R4= Test formulation four. Each value is a Mean±SD of three determination

Table - 91: Pharmaceutical characteristics of compressed formulation of ibuprofen tablet

Test Formulation

Pharmacopoeial Limits

(USP 32/NF 27)

Wt. Variation

(Mean ± S.D)

(mg) ±5%

Thickness

(Mean ± S.D)

(mm) ±5%

Length x width

(Mean ± S.D)

(mm) ₋

Hardness

(Mean ± S.D)

(kg) At least 5 kg

Loss on

Drying %

Standard 833.33±2.08 6.33± 0.06 20 x 9.5 6.13± 0.14 4.5%

R1 843.33±2.08 6.23± 0.06 20 x 9.5 6.23± 0.14 4.9%

R2 848.33±2.89 6.37±0.06 20 x 9.5 8.40±0.40 4.8%

R3 847.67±2.52 6.22±0.03 20 x 9.5 9.23±0.14 4.9%

R4 842.67±2.52 6.27±0.06 20 x 9.5 10.43±0.18 4.7%

Table- 92: dissolution studies of ibuprufen tablets

163

FORMULATION NUMBER

R1 R2 R3 R4

243nm (Abs) % release 243nm (Abs) % release 243nm (Abs) % release 243nm (Abs) % release

0.405 89.6 0.166 36.72 0.092 33.45 -0.018 -6.54

0.415 91.81 0.141 31.190 0.090 32.72 -0.017 -6.182

0.420 92.91 0.147 32.51 0.112 40.72 -0.020 -7.27

0.425 94.02 0.154 34.06 0.113 41.92 0.008 -2.90

0.429 94.95 0.143 31.62 0.030 10.90 -0.008 -2.90

0.430 95.13 0.161 35.60 0.036 13.091 -0.008 -2.90

STANDARD FOR FORMULATION

0.503 0.503 111.33 0.306 111.33 0.306 111.33

R1-R4 are test formulation while Abs = absorbance and %release = percent release

164

R1 R2

R3 R4

Figure – 39: Formulated ipubrufen tablets ( R1, R2, R3, R4)

165

Table -93: Basic evaluation test of antidiarrheal formulation prepared from

sapodilla pectin

Parameters Suspension made from

sapodilla pectin Comparative suspension *

Color Pinkish white White

Odor vanilla Vanilla

Taste sweet sweet

pH 6.1 5.56

Viscosity 14.14 13.13

Sedimentation rate 0.3 0.1

Redispersity +++ +++

WHC 32.69 33.56

+ denotes the number of times the cylinder was moved. * Keptin antidiarrheal preparation

166

Figure- 40 Formulated antidiarrheal preparation

167

Table - 94: Effect of different concentration of sapodilla pectin on the chemical

properties of the jam samples

Sample PH

TSS

(mg)

TTA

(ml)

MC

(%)

Vit C

(mg)

Ash

(%)

TS

(mg)

Viscosity

(Cp)

S 3.06 61.9 1.14 35.9 18.3 1.78 64.1 71

F1 3.16 56.7 1.08 41 17.6 1.69 59 56

F2 3.25 62.6 1.01 35.7 18.1 1.73 64.3 63

F3 3.25 62.4 1.03 35.1 17.9 1.71 64.9 67

WhereS = Jam with standard pectin, F1 = Jam with 2g pectin, F2 = Jam with pectin in same concentration

as in standard, F3 = Jam with 10g pectin while MC is moisture content . TSS total soluble solids, TTA

total titratable acid and TS is total solid.

Table - 95: Scores for sensory parameters as judged by twenty (20) panelists.

Sampl

e

Appearanc

e

Tast

e

Arom

a

Spreadabilit

y

Textur

e

Mout

h feel

Overall

acceptabilit

y

S 7.7 7.9 7.9 8.2 7.8 7.5 7.8

F1 7.5 7.3 7.2 7.3 7.2 7 7.2

F2 7.5 7.6 7.7 7.8 7.8 7.5 7.6

F3 6.2 6.4 6.2 5.6 6.3 5.5 6

WhereS = Jam with standard pectin, F1 = Jam with 2g pectin, F2 = Jam with pectin in same concentration as in standard, F3 = Jam with 10g pectin

168

(S)=Jam made from 5g standard pectin (F1) Jam made from 5g

pectin

(F2) Jam made from 7 g pectin (F3) Jam made from 10g

pectin

Figure -41 formulation of Jam made from extracted pectin

169

Figure -42: formulation of pudding made from extracted pectin.

170

DISCUSSION

Appropriate utilization of waste products after fruit processing is one of the most

propitious field which needs comprehensive research, evaluation and interpretation by the

scientists of related discipline. Surprisingly, wastes from food processing industries are

among the largest mass produced all over the world as a result of pre-consumer

procedures. It is sometimes around 50% of the total processing waste and are very

difficult to manage for further developments because of improper utilization techniques,

available resources, cost and regulatory issues. Making useful functional compounds

from fruit wastes and their proper utilization can be beneficial economically and will also

help in reducing risk to the changing global environment. Initial studies on pectin has

been reported in some articles published in 1750 with the utilization to develop food

products specially apple jelly (Kertesz, 1951). Although extraction and identification of

crude pectin has been reported after this study, however the first commercial extraction

process started in Germany in 1908 and subsequently the process was granted a patent

(U.S. Patent no. 1,082.682) in 1913 (Canteri, 2012). This has triggered various

researchers as well as food industries to develop new and improved method of extractions

of pectin to obtain considerable quantities of pectin to utilize in developing various food

and pharmaceutical products. The commercial production of pectin and its utilization

needs proper handling selection of fruits and processing of waste products to achieve the

desired results. This demands a very comprehensive and dedicated research, evaluation

and interpretation of the results by the scientist working in the particular field. In the

initial phase most studies on pectin production and its utilization have been reported in

western world because of the resources and techniques available with them. However

171

with the passage of time this has moved to under developed or developing countries and

as a result a large number of people have been noted to be engaged in such studies to

develop an economical extraction procedure for pectin. The research in this field is still

continued and despite huge commercial production by some of the well reputed

companies (such as Copenhagen pectin, Denisco pectin Mexico, Sanofi bio- industries

and Cargill and CP Kelco ) researchers are still trying to find new methods modified

resources and optimization of extraction conditions to increase pectin yield.

Pakistan is primarily classified under agricultural based country where huge quantity of

fruits are produced and sold in the market. However the processing industries are lacking

which results in the commercial production of various food products with the utilization

of pectin. Every year considerable quantity of fruits are spoiled and discarded due to

improper handling and non-availability of basic manufacturing units which can utilize

these fruits and their waste to convert into valuable products. Ultimately Pakistan is

mostly dependent upon the imports of such basic raw materials including pectin to cater

its food and pharmaceutical industries to utilize and develop products for commercial and

therapeutic applications consequently a huge quantity of pectin is imported in Pakistan

against the expenditure of valuable foreign exchange currency. The June 2015 to October

2016 import figures were 1,120,029.15US $ import of pectin and most important

exporting countries include Brazil, China, Czech Republic, Germany, Denmark

and Spain. With this aim and objective present studies were organized and conducted to

evaluate an economical and easily available source of fruit for developing process to

extract pectin and its utilization in some food and pharmaceutical formulations. The

integral part in the production of high quality pectin is based on the method of extraction

172

used to recover pectin from the desired source. The selection of fruits as well as the

nature of waste and the experimental design used for the extraction of pectin greatly

affect the physico-mechanical properties and yield. Recent studies indicated successful

recovery of pectin from the lime biomass using enzyme cellulose obtained from

Penicillium funiculosum and designated as Laminex CK2 (Dominiak et al., 2014). The

current study focus on a number of methods which may affect pectin yield, were studied

and repeated as a new system in order to acquire an efficient extraction procedure from

fruits wastes. Method for extraction and evaluation also moves around various physico-

mechanical factors for optimizing the yield of pectin from a new source which were not

reported earlier from this part of the world and also its utilization in pharmaceutical

preparations and as a thickening agent in food products.

The different sources investigated in this study are mentioned in Table - 9 with their

respective percent yield of pectin. During the preliminary study acid extraction was

mainly used because it generally results to acquire a good yield of pectin. Hydrochloric

acid for pH 3-5, ammonium hydroxide for 6 and above with conventional boiling

methods (Bunsen burner) and 10 min of boiling was applied for the extraction process.

The pH of the medium varied with the origin of the material used for pectin extraction

(May, 1990). The study conducted exibited a variable response in terms of yield and were

compared with reported values. Orange, apple, banana and lemon showed yield similar to

the range presented in previous studies ( Aina et al., 2012 ; Munarin et al., 2012 ) while

grapefruit (Munarin et al., 2012), guava (Bhat and Singh, 2014), sweet lime (Munarin et

al., 2012), watermelon (Rasheed, 2008) and mango (Munarin et al., 2012) presented

lesser yield, while there were few fruits which failed to produce any results, although a

173

considerable amount of pectin was reported from them in earlier studies. The fruits were

peach (Munarin et al., 2012) and apricot (Baissise et al., 2010). Few fruits showed a

good quantity of pectin, although there was no previous study found on them, the fruits

were sapodilla, muskmelon and cantaloupe. Moreover, there were fruits which were

unable to present any resulted pectin and no previous study was found on them such

as guava (unripe), jungle fruit and mango (unripe). The difference in yield of pectin from

the screened fruits from their past studies was due to the fact that that the

physicochemical properties of pectin depend upon the breed, characteristic features and

atmospheric conditions during growth of the fruit and fruit sources (El-Nawawi and

Shehata, 1987; Chan and Choo., 2013).

Following the preliminary screening, three fruits were selected for further evaluation

while two already explored fruits; apple and orange were studied for their comparing

effects. The extraction conditions used during the isolation of pectin was designed in a

way that all the preliminary yield-affecting variables of pectin were studied very

carefully. pH, boiling method and mechanical methods are factors which can affect the

yield from fruit as described earlier. The extraction regime selected for the study was pH

(1 to 7), boiling method (Bunsen burner and microwave), and mechanical procedures

(homogenising, grinding, chopping, cutting and hammering). The overall objective was

to explore the fruits for better yields of pectin after applying different phyicomechanical

conditions which later can identify a single source for further evaluation to enhance yield

of pectin from it. The data was also analyzed statistically to study the significant impact

of all the variables (pH, boiling method and mechanical method) used during the study.

Table -10 shows the yield of pectin achieved from five selected fruits (sapodilla, banana,

174

muskmelon, apple and orange) and indicates difference in yield after applying different

combination of variables. Statistically analyzed by Analysis of variance (ANOVA),

results indicated that different variables like mechanical procedure (MP), pH and boiling

method (BM) have significant impact on the yield of all five fruits at 1% level of

significance (Table - 11 ). The difference among the means regarding different factors

during evaluation of all five fruits was also determined by Tukey’s test and are shown

under respective tables of investigated fruit.

The yields recorded from different fruits proved to be dependent upon their natural pectin

content and efficiency of the process used and are showed in Table- 10. Previous study

showed that sapodilla fruit was investigated for total dietary fibers and pectin contents

(0.35g ± 0.01/100g fruit) in its edible portion (Mahattanatawee et al., 2006). However, in

the present study, pectin was extracted from peel of sapodilla fruit and was best extracted

(4% yield) through grinding at pH 3, using conventional heating method. A further

increase in yield was observed (4.7%) at lower pH (pH 1) when heating mode was

changed to microwave (Table-10), with the indication that intensity of heating has more

impact on pectin yield from sapodilla on that specific pH. The difference among the

means regarding different factors using Tukey’s test are given in Table – 12-14 for

sapodilla peel. The overall mean for mechanical procedure, pH and boiling method

recommended chopping, pH 3 (Table - 12) and microwave boiling (Table -13) has a

significant impact (P<0.05) on the yield of pectin from sapodilla fruit. Table 12 and 13

also show the status of remaining mechanical procedure, pH and boiling method and their

order of influence is given from most to least effective. Alphabet A was considered the

most and subsequent alphabets lesser in effectiveness than the preceding alphabet.

175

Overall interaction effect of boiling method with pH (BM x pH) on sapodilla peel showed

(Table - 14) that M x 3 and M x 5 (Heating using microwave at pH 3 and 5) were

significantly non-significant with each other, giving significantly (P<0.05) highest yield

while yield at B x 1 ( Heating using Bunsen burner at pH1) was the lowest.

Table- 10 also showed the comparison of yields form banana fruit peel. The microwave

heating was also noted as a better option to obtain good yield using all mechanical

methods. The banana peels gave good yield of pectin and was observed to be very much

close to the yield of pectin as reported in an earlier study (Christy et al., 2014).

Homogenizing was the best mechanical procedure, as it gave the maximum yield (8.85%

yield) at pH 5, using conventional method. For microwave heating extraction, hammering

was noted as the best method (10.5% yield) for banana peel at the same pH (pH 5). The

difference among the means regarding different factors using Tukey’s test are given in

Table – 15-17 for banana peel. The overall mean for mechanical procedure, pH and

boiling method recommended homogenising, pH 5 (Table - 15) and microwave boiling

(Table - 16) has a significant impact (P<0.05) on the yield of pectin from banana fruit.

Table 15 and 16 also show the status of remaining mechanical procedure, pH and boiling

method and their order of influence is given from most to least effective. Alphabet A was

considered the most and subsequent alphabets lesser in effectiveness than the preceding

alphabet. The Overall interaction effect of boiling method and pH (BM X pH) on the

yield of banana showed that M x 5 (Heating using microwave at pH 5) gave highest yield

and significantly (P<0.05) different from other levels of BM x pH. Yield at B x 1

(Heating using Bunsen burner) was lowest which was statistically significantly different

from all other levels (Table -16).

176

Muskmelon was also studied previously for total dietary fibers (Mahattanatawee et

al., 2006). For muskmelon peels, pH 3 using conventional heating and cutting as

mechanical process was noted as the best option to achieve high yield (2.45%) of pectin.

However, using microwave technique, extraction after grinding as mechanical process at

pH 1 provided best yield (2.65%) (Table-10). The difference among the means regarding

different factors using Tukey’s test are given in Table – 18 - 20 for muskmelon peel. The

overall mean for mechanical procedure, pH and boiling method recommended grinding,

pH 5 (Table 18) and microwave boiling (Table 19) has a significant impact (P<0.05) on

the yield of pectin from muskmelon fruit. Table 18 and 19 also show the status of

remaining mechanical procedure, pH and boiling method and their order of influence is

given from most to least effective. Alphabet A was considered the most and subsequent

alphabets lesser in effectiveness than the preceding alphabet. Overall interaction effect of

BM x pH showed that M x 5 (Heating using microwave at pH 5) gave highest yield and

significantly (P<0.05) different from other levels of BM x pH while yield at B x 7

(Heating using Bunsen burner at pH1) was lowest (Table-20).

Although literature contains the amount of pectin that can be extracted from apple

pomace (10 to 15%) (Ziari et al., 2010), it was not comparable with the present study,

since to synchronize the study, apple peels, instead of pomace was used. For apple peels

the best mechanical method was found hammering for both the heating methods at pH 5

(3.05% to 4.85% yield) Table – 10. The difference among the means regarding different

factors using Tukey’s test are given in Table – 21 -23 for apple peel. The overall mean

for mechanical procedure, pH and boiling method recommended cutting and hammering,

pH 3 and 5 (Table - 21) and microwave boiling (Table - 22) has a significant impact

177

(P<0.05) on the yield of pectin from apple fruit. Table - 21 and 22 also show the status of

remaining mechanical procedure, pH and boiling method and their order of influence is

given from most to least effective. Alphabet A was considered the most and subsequent

alphabets lesser in effectiveness than the preceding alphabet. Overall interaction effect of

boiling method and pH (BM x pH) showed that M x 3 and M x 5 (Heating using

microwave at pH 3 and 5) gave highest yield and significantly (P<0.05) different from

other levels of BM x pH while yield at B x 7 ( Heating using Bunsen burner at pH7) was

lowest Table – 23.

The extracted pectin yield from orange peels was also comparable with previously

extracted pectin (Sayah et al., 2014). For orange peels grinding was found producing

highest yield at pH=7 in conventional method (21.7 %) and slightly higher yield with

microwave method (22.7%) at lower pH 5 (Table -10). The difference among the means

regarding different factors using Tukey’s test are given in Table – 24-26 for orange peel.

The overall mean for mechanical procedure, pH and boiling method recommended

homogenising, pH 1 (Table - 24) and microwave boiling (Table - 25) has a significant

impact (P<0.05) on the yield of pectin from orange fruit. Table 24 and 25 also show the

status of remaining mechanical procedure, pH and boiling method and their order of

influence is given from most to least effective. Alphabet A was considered the most and

subsequent alphabets lesser in effectiveness than the preceding alphabet The overall

interaction effect of boiling method and pH (BM x pH) showed that M x 1 (Heating using

microwave at pH 1) gave highest yield and significantly (P<0.05) different from other

levels of BM x pH while yield at M x 7 (Heating using Bunsen burner at pH7) was lowest

Table – 26.

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In earlier studies, the effect of particle size reduction was also reported to have significant

impact, based on mechanical procedure for yield of pectin. This is because reduction in

particle size enhances protopectin release which ultimately increases the yield of pectin

when precipitated with ethanol (Canteri-Schemin et al., 2005). Likewise, pH of the

extracting medium also affects significantly on the yield of pectin (Ziari et al., 2010). In

the present study the effect of interaction between variable combination such as

mechanical procedure with pH, mechanical procedure with boiling methods, pH with

boiling methods and also the interaction of all mechanical procedures, pH and boiling

method on yield of all five fruits were also investigated. Results highlighted in Table -8

showed that interactions of all have positive impact on the yield of pectin from all five

fruits.

Preliminary investigation pointed towards a new source sapodilla fruit peel from which

extraction of pectin has not been investigated to date. Sapodilla ranked top among the

high pulp fruit with about 85% as edible portion. But the fruit has been neglected and did

not achieve the position it deserved. The current research can highlight one of the many

advantages of this special fruit grown in large areas of Pakistan and rest of the world. The

extraction of pectin from sapodilla fruit peel were conducted on the aforementioned

variables used for the study of five selected fruits (Table-8) with the exception that time

of boiling was added ( 10, 20, 40 and 60 min). The objective of adding time of boiling

was to evaluate the best condition for maximum yield that could be employed in the

commercialization of sapodilla peel pectin. The added extraction technique was applied

after using two strengths of inorganic acid (0.1N and 1N HCl).

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Time of boiling has a profound effect on pectin yield which was studied among the other

affecting variable on yield by different scientists (Kliemann et al 2009, Khan et al

2015), while the mode of boiling also has a positive effect (Guo et al 2012). The highest

yield (6.5%) obtained at pH 3, chopping as mechanical procedure after 20 min of boiling

in Microwave (Table- 27). As reported earlier, the extraction results obtained after

3 hrs of conventional heating were the same with yield obtained by 15 min of extraction

in microwave (Salam et al., 2012). The maximum yield obtained by conventional heating

was 5.5% but at pH 5 and hammering as mechanical procedure (Table – 27). To

understand the overall effects of each variable, the data was also analyzed statistically by

the same software and procedure as used in aforestated five fruit analysis. Analysis of

variance (ANOVA) in the Table - 28 shows that the mechanical procedures have a

significant (P<0.01) impact on yield on all time of boiling selected for the study.

Similarly, pH level and Boiling method also significantly (P<0.01) impact on yield when

using different times of boiling. All the two way interactions i.e. mechanical procedure

and pH, (MPxpH), mechanical procedure and boiling method (MPxBM), pH and boiling

method (pHxBM) and three factor interaction of mechanical procedure, pH and boiling

method MPxpHxBM showed significant (P<0.01) impact on yield. Table 29- 31 shows

the overall mean comparisons and their effect after 10min of boiling.

The difference among the means regarding different factors using Tukey’s test are given

in Table – 29- 31 for 10 min of boiling. The overall mean for mechanical procedure, pH

and boiling method showed chopping, pH 3 (Table - 29) and microwave boiling (Table -

30) has a significant impact (P<0.05) on the yield of pectin after 10 min of boiling of

peels. Table 29 and 30 also show the status of remaining mechanical procedure, pH and

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boiling method and their order of response is given from most to least effective. Alphabet

A was considered the most and subsequent alphabets lesser in effectiveness than the

preceding alphabet. Overall interaction effect in Table – 31 shows interaction of BM x

pH showed that M x 3 and M x 5 (Heating in microwave at pH 3and 5) gave highest yield

and significantly (P<0.05) different from other levels of BM x pH while yield at B x 1

and Bx7 (Heating on Bunsen burner) was lowest.

For 20 min of boiling the difference among the means regarding different factors using

Tukey’s test are given in Table – 32-34. The overall mean for mechanical procedure, pH

and boiling method showed homogenising pH 3 (Table - 32) and microwave boiling

(Table - 33) has a significant impact (P<0.05) on the yield of pectin after 20 min boiling

of peels. Table-32 and 33 also show the status of remaining mechanical procedure, pH

and boiling method and their order of response is given from most to least effective.

Alphabet A was considered the most and subsequent alphabets lesser in effectiveness

than the preceding alphabet. Overall interaction effect in Table – 34 shows interaction of

BM x pH showed that M x 3 and M x 5 (Heating in microwave at pH 3and 5) gave

highest yield and significantly (P<0.05) different from other levels of BM x pH while

yield at B x 1 and Bx7 (Boiling on Bunsen burner at pH 1 and 7) was lowest.

The difference among the means regarding different factors using Tukey’s test are given

in Table – 35-37 for 40 min of boiling. The overall mean for mechanical procedure, pH

and boiling method showed hammering pH 5 and 6 (Table 35) and microwave boiling

(Table 36) has a significant impact (P<0.05) on the yield of pectin after 40 min of boiling

of peels. Table-35 and 36 also show the status of remaining mechanical procedure, pH

and boiling method and their order of response is given from most to least effective.

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Alphabet A was considered the most and subsequent alphabets lesser in effectiveness

than the preceding alphabet. Overall interaction effect in Table – 37 shows interaction of

BM x pH showed M x 5 (Heating on microwave at pH 5) gave highest yield and

significantly (P<0.05) different from other levels of BM x pH while yield at B x 1

(Heating on Bunsen burner at pH1) was lowest.

The difference among the means regarding different factors using Tukey’s test are given

in Table – 38- 40 for 60 min of boiling. The overall mean for mechanical procedure, pH

and boiling method showed chopping and homogenising, pH 5 (Table-38) and

microwave boiling (Table-39) has a significant impact (P<0.05) on the yield of pectin

after 60 min of boiling of peels. Table-38 and 39 also show the status of remaining

mechanical procedure, pH and boiling method and their order of response is given from

most to least effective. Alphabet A was considered the most and subsequent alphabets

lesser in effectiveness than the preceding alphabet. Overall interaction effect of BM x pH

level showed that M x 5 (Heating on microwave at pH 5) gave highest yield and

significantly (P<0.05) different from other levels of BM x pH and yield at B x 1(Heating

on Bunsen burner at pH1) was the lowest (Table-40).

In the subsequent part extraction was carried out by same procedure as followed in

foregoing study with the exception of increasing the strength of acid from 0.1N to 1N

HCl. Strength of acid showed to influence positively on the yield of pectin in numerous

earlier studies (Yapo 2009, Kalapathy and Proctor 2001) and was tested in current study

for extraction of pectin from sapodilla fruit peel. Table - 41 presented the yield acquired

by the use of increasing strength of acid. It was observed that maximum yield was

achieved from chopped peels only after 10 min of boiling in microwave (6.7%) at pH 5.

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Whereas 0.1 N HCl showed lesser highest yield 6.5% after 20 min of boiling at pH3.

Table - 42 shows analysis of variance (mean squares) of yield for different fruits. The

Table showed that the mechanical procedures has a significant (P<0.01) effect on yield

on all time of boiling selected for the study. Similarly, pH and boiling method have

significantly (P<0.01) affected the yield when using different times of boiling. All the

two way interactions i.e. mechanical procedure and pH (MPxpH), mechanical procedure

and boiling method (MPxBM), Ph and boiling method (pHxBM) and three factor

interaction mechanical procedure, pH and boiling method (MPxpHxBM) showed

significant (P<0.01) effect on yield. The difference among the means regarding different

factors using Tukey’s test are given in Table – 43-45 for 10 min of boiling using 1NHCl.

The overall mean for mechanical procedure, pH and boiling method showed chopping

and homogenising, pH 3 (Table - 43) and microwave boiling (Table - 44) has a

significant impact (P<0.05) on the yield of pectin after 10 min of boiling of peels using

1N HCl. Table- 43 and 44 also show the status of remaining mechanical procedure, pH

and boiling method and their order of response is given from most to least effective.

Alphabet A was considered the most and subsequent alphabets lesser in effectiveness

than the preceding alphabet. Overall interaction effect of boiling method and pH (BM x

pH) showed that M x 5 (Heating on microwave at pH 5) gave highest yield and

significantly (P<0.05) different from other levels of BM x pH while yield at B x 7

(Heating on Bunsen burner at pH 7) was lowest Table - 45.

The difference among the means regarding different factors using Tukey’s test are given

in Table – 46-48 for 20 min of boiling using 1NHCl. The overall mean for mechanical

procedure, pH and boiling method showed hammering, pH 5 (Table 46) and microwave

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boiling (Table 47) has a significant impact (P<0.05) on the yield of pectin after 20 min of

boiling of peels using 1N HCl. Table- 46 and 47 also show the status of remaining

mechanical procedure, pH and boiling method and their order of response is given from

most to least effective. Alphabet A was considered the most and subsequent alphabets

lesser in effectiveness than the preceding alphabet. Overall interaction effect of boiling

method and pH (BM x pH) showed that M x 3 and M x 5 (Heating on microwave at pH 3

and pH 5) gave highest yield and significantly (P<0.05) different from other levels of BM

x pH while yield at B x 1 (Heating on Bunsen burner at pH1) was lowest (Table – 48).

The difference among the means regarding different factors using Tukey’s test are given

in Table – 49-51 for 40 min of boiling using 1NHCl. The overall mean for mechanical

procedure, pH and boiling method showed hammering, pH 5 (Table - 49) and microwave

boiling (Table - 50) has a significant impact (P<0.05) on the yield of pectin after 40 min

of boiling of peels using 1N HCl. Table- 49 and 50 also show the status of remaining

mechanical procedure, pH and boiling method and their order of response is given from

most to least effective. Alphabet A was considered the most and subsequent alphabets

lesser in effectiveness than the preceding alphabet. Overall interaction effect of boiling

method and pH (BM x pH) showed that M x 5 (Heating in microwave at pH 5) gave

highest yield and significantly (P<0.05) different from other levels of BM x pH while

yield at B x 7 (Heating on Bunsen burner at pH at 7) was lowest Table – 51.

The difference among the means regarding different factors using Tukey’s test are given

in Table – 52 -54 for 60 min of boiling using 1NHCl. The overall mean for mechanical

procedure, pH and boiling method showed grinding, pH 3 (Table - 52) and microwave

boiling (Table - 53) has a significant impact (P<0.05) on the yield of pectin after 60 min

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of boiling of peels using 1N HCl. Table- 50 and 51 also show the status of remaining

mechanical procedure, pH and boiling method and their order of response is given from

most to least effective. Alphabet A was considered the most and subsequent alphabets

lesser in effectiveness than the preceding alphabet. Overall interaction effect of boiling

method and pH (BM x pH) showed that M x 5 (Heating in microwave at pH 5) gave

highest yield and significantly (P<0.05) different from other levels of BM x pH and yield

at B x 5 (Heating on Bunsen burner at pH5) was lowest (Table - 54).

Organic acids are also widely used for optimizing the yield of pectin from various fruits.

The use of organic acid instead of inorganic was also studies in the current study. These

include citric, oxalic and tartaric acid. Oxalic and citric acid gave comparable high yield

of pectin when used in the study. Organic acids like oxalic and citric acid have been used

to extract pectin and gave similar results 4.95% and 4.24% yield respectively after 10 min

of extraction time (Table - 55). Chan and Choo, 2013 studied effect on yield of pectin

after using citric acid and extraction time from 1.5 to 3.0 h and reported increase in yield

with increase extraction time. While Vriesmann et al. (2012) and Canteri-Schemin et al.

(2005) at the end of their study drawn conclusion that extraction time can increase yield

of pectin from coca husk and apple pomace. However the most effective model for

extraction of pectin using organic acid was after using 1% oxalic acid with homogenizing

as a mechanical tool in the present study. Two different concentration of organic acid 1%

and 10% were used in the present study. Low concentration of all the organic acids (1%)

has been found more effective with microwave as heating procedure. Table – 56 shows

Analysis of variance (mean squares) of yield for pectin from sapodilla fruit using organic

acid. The table shows that the mechanical procedures has a significant (P<0.01) effect on

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yield using different types of acid (citric acid, oxalic acid and tartaric acid) selected for

the study. Similarly, the concentration of organic acids (% age) and boiling method was

also been noted to have significant effect (P<0.01) on the yield of pectin. The two way

interactions i.e. boiling method (BM x MP) and mechanical procedure and acid

concentration (MP x Acid %) and three factor interaction boiling method, mechanical

procedure and acid concentration (BM x MP x Acid %) showed significant (P<0.01)

effect on yield. Only boiling method and acid concentration (BM x Acid %) has no

significant (P>0.05) effect on yield.

The difference among the mean values related to various factors (e.g. mechanical

procedure and method of boiling using Tukey’s test are given in Table – 57 -59 for yield

of pectin using citric acid from sapodilla fruit. The overall mean for mechanical

procedure, indicated homogenising and 1% strength of organic acid (Table – 57- 58) has

a significant impact (P<0.05) on the yield of pectin. Table- 57 and 58 also show the

status of remaining mechanical procedure,% strength and boiling method and their order

of response is given from most to least effective. Alphabet A was considered the most

and subsequent alphabets lesser in effectiveness than the preceding alphabet. While

overall interaction for 1% citric acid and mechanical procedure 2 (Chopping) and for

10% mechanical procedure 1 (Homogenizing) gave most effective results as compare to

others (Table – 59).

The difference among the mean values related to various factors (e.g. mechanical

procedure and method of boiling using Tukey’s test are given in Table –60 -62 for yield

of pectin using oxalic acid from sapodilla fruit. The overall mean for mechanical

procedure, indicated homogenising and 1% strength of organic acid (Table – 60 -61) has

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a significant impact (P<0.05) on the yield of pectin. Table- 60 and 61 also show the status

of remaining mechanical procedure, % strength and boiling method and their order of

response is given from most to least effective. Alphabet A was considered the most and

subsequent alphabets lesser in effectiveness than the preceding alphabet. While overall

interaction for 1% oxalic acid with mechanical procedure, mechanical procedure

1(Homogenizing) gave most effective results as compare to others Table – 62.

The difference among the mean values related to various factors (e.g. mechanical

procedure and method of boiling using Tukey’s test are given in Table – 63 - 65 for yield

of pectin using tartaric acid from sapodilla fruit. The overall mean for mechanical

procedure, indicated homogenising (Table- 63 ) and 10% strength of organic acid (Table

- 64 ) has a significant impact (P<0.05) on the yield of pectin Table- 63 and 64 also show

the status of remaining mechanical procedure,% strength and boiling method and their

order of response is given from most to least effective. Alphabet A was considered the

most and subsequent alphabets lesser in effectiveness than the preceding alphabet. While

overall interaction of mechanical procedure strength of organic acid and boiling method,

1% tartaric acid with mechanical procedure 1 ( Homogenizing) , 10% tartaric acid with

mechanical procedure 1 and 10% tartaric acid with mechanical procedure 3 (Grinding) is

equally effective for better yield from sapodilla fruit as compare to others (Table – 65).

Finally the effect of strength of inorganic acid on the yield of pectin is investigated using

three strength of inorganic acid 0.1, 0.5 and 1N HCl (Table - 66). The yield under 0.1N

HCL and 1N HCl (Table – 27 and 41 respectively) has already been discussed in

previous part of this discussion hence a statistical analysis was performed. Table - 67

shows Analysis of variance (mean squares) of yield for pectin from sapodilla fruit using

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different strength inorganic acid (HCl). The table shows that the mechanical procedures

has a significant (P<0.01) effect on yield using different strength of acid selected for the

study. Similarly pH and Boiling method have significantly (P<0.01) affected the yield

when using different strength of acids. The two way interactions i.e. boiling method and

pH (BM x MP), mechanical procedure and pH (MP x pH) and three factor interaction

boiling method, mechanical procedure and pH (BM x MP x pH) and boiling method and

pH (BM x pH) showed significant (P<0.01) effect on yield.

The difference among the mean values related to various factors (e.g. mechanical

procedure and method of boiling using Tukey’s test are given in Table – 68 - 70 for

pectin yield from sapodilla fruit using 0.1NHCl. The overall mean for mechanical

procedure, pH and boiling method showed chopping, pH 3 (Table - 68) and microwave

boiling (Table - 69) has a significant impact (P<0.05) on the yield of pectin after using

0.1N HCl. Table- 68 and 69 also show the status of remaining mechanical procedure, pH

and boiling method and their order of response is given from most to least effective.

Alphabet A was considered the most and subsequent alphabets lesser in effectiveness

than the preceding alphabet. Overall interaction effect of boiling method and pH (BM x

pH) showed that M x 5 (heating in microwave at pH 5) gave highest yield and

significantly (P<0.05) different from other levels of BM x pH (Table - 70)

The difference among the mean values related to various factors (e.g. mechanical

procedure and method of boiling using Tukey’s test are given in Table – 71 -73 for pectin

yield from sapodilla fruit using 0.5 NHCl. The overall mean for mechanical procedure,

pH and boiling method showed cutting, pH 1 (Table - 71) and microwave boiling (Table -

72) has a significant impact (P<0.05) on the yield of pectin after using0.5N HCl. Table-

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71 and 72 also show the status of remaining mechanical procedure, pH and boiling

method and their order of response is given from most to least effective. Alphabet A was

considered the most and subsequent alphabets lesser in effectiveness than the preceding

alphabet. Overall interaction effect of boiling method and pH (BM x pH) showed that M

x 1 (Heating in microwave at pH 1) showed highest yield and significantly (P<0.05)

different from other levels of BM x pH (Table - 73)

The difference among the mean values related to various factors (e.g. mechanical

procedure and method of boiling using Tukey’s test are given in Table – 74 -76 for pectin

yield from sapodilla fruit using 1N HCl. The overall mean for mechanical procedure, pH

and boiling method showed chopping, pH 3 and 5 (Table - 74) and microwave boiling

(Table - 75) has a significant impact (P<0.05) on the yield of pectin after using 1N HCl.

Table- 74 and 75 also show the status of remaining mechanical procedure, pH and boiling

method and their order of response is given from most to least effective. Alphabet A was

considered the most and subsequent alphabets lesser in effectiveness than the preceding

alphabet. Overall interaction effect of boiling method and pH (BM x pH) showed that M

x 5 (heating in microwave at pH 51) showed highest yield and significantly (P<0.05)

different from other levels of BM x pH (Table - 76).

In the conducted study, maximum yield of pectin was obtained in acidic medium,

indicating it as an important factor to be considered while extracting pectin. Earlier

studies also suggested positive effect of acidic medium (low pH) upon yield of pectin and

a drastic effect on pectin yield with the increase in pH (Canteri-Schemin et al., 2005

and Yapo et al., 2007). The lower value of pH supports the theory of the disruption of the

cell wall, release of protopectin containing cellulose pectin in the medium and its

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hydrolysis, thus releasing high quantity of pectin in the liquid phase which can thus be

recovered by precipitation. It was also observed that among the five fruit peels

investigated in the study, sapodilla peels gave considerably good yield in basic medium

too that is pH 6.5 and 7, which is quite interesting but in accordance with an earlier

published report (Kirtchev et al., 1989) in which authors suggested that both acidic and

alkaline medium with elevated temperatures can help the rupture of cell

walls to release protopectin and its hydrolysis to release pectin.

The data showed that mechanical method does have effect on yield as shown in Table -

10. Different kinds of techniques have been used for their specific advantages by various

workers while working on pectin extraction process optimization. Cutting of peels into

smaller pieces, help to increase surface area of the material for better extraction

(Rudolph and Petersen, 2012). Likewise, mortar and pestle facilitated the size reduction

of the raw material selected for the extraction of pectin (Poovaiah and Nukaya, 1979).

Hammer mill is extensively used in studies to ground cell wall material to extract pectin

( Besson et al., 2013), while grinding in mechanical blender and in mills procedure has

also been termed a successful procedure to extract pectin (Loyola et al., 2011).

Homogenizing is also an effective way of forming loose slurry used in extraction of

pectin (Slavov et al., 2013). Different fruits indicated different yield when subjected to

various mechanical methods adopted in the study. The effectiveness of a mechanical

process in increasing pectin yield depends upon its capability of disrupting the cell wall

of the peels to release pectic substance. One mechanical process may not be suitable for

different fruit peels because of the structure and composition of the cell wall. Chopping

with mortar and pestle has been used for size reduction of raw materials with or without

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adding solvents since long in the extraction processes (Lamotte et al., 1969; Poovaiah and

Nukaya, 1979).

With the conventional or direct heating procedure there is a chance of pectin

degradation. Therefore, microwave heating was studies in the present study to learn the

effect of extraction time and to improve the quality of pectin. Among the two boiling

methods, the overall yield was recorded better with microwave. The mineral acid used in

the extraction of pectin was used at low strengths (0.1 till 1N HCl), in order to keep the

process environmental friendly because of corrosive nature of HCl (Vriesmanna and

Petkowicz, 2013). The methods used in the present study were kept simple, in order to

get the required preliminary knowledge about the fruits like sapodilla, muskmelon and

banana, while comparing it at the same time with known high content pectin fruits, in

order to get an authentic and reproducible results for more effective and economical way

for the better extraction of pectin.

Apparently it is recognized that physicochemical properties of pectin influence their

functional properties consequently affecting its application in food and pharmaceutical

systems. In order to study the basic character of sapodilla pectin basic identification and

bio characterization tests were performed. Table - 77 describes the basic identification

test based on USP for isolated sapodilla, banana and muskmelon pectin. The basic

identification tests of the current study showed positive response from sapodilla and

banana and affirms the isolated pectin has all the basic characteristics of being a

carbohydrate material (Yadav and Agarwala 2011). However, banana pectin failed to

form stiff gel which is the basic and most important functional character of pectin.

Whereas muskmelon also failed to form gel and tests performed for identification. Hence

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the two fruits (banana and muskmelon) were omitted for further analysis and sapodilla

peel pectin was given full consideration. The preceding extraction analysis revealed the

impact of pH on the yield of pectin from sapodilla fruit peel. Recent studies on isolation

of pectin from different fruits have also indicated that pH has a key role both on the yield

and the functionality of pectin (Ziari et al., 2011; Chan and Choo 2013; Kaya et al.,

2014). Hence further studies were also involved the sapodilla peel pectin extracted at

different pH (Table-78).

As mentioned earlier the basic functional property of pectin is to form gel with a

specified system. The gel formation capacity of pectin depends upon many factors which

were also included in the studies. Sapodilla being a new source was studied for its basic

bio-character and are shown in Table - 78. The dried pectin was stored in a cool dry

place. Before analysis it was necessary to confirm that the dried pectin is free from

ammonia. Presence of ammonia may obstruct with the other tests used for the

characterization of pectin. If ammonium is detected, the sample was washed with

acidified 60% alcohol, followed by neutral alcohol to remove the acid. The presence of

ammonia can be identified by the distinctive smell of ammonia (Ranganna, 1986). In the

present study all the samples were observed free rom ammonia.

The moisture content of pectin appeared in the range of 5.52% to 6.02% .The high

moisture content is likely to increase the susceptibility of dried pectin for the growth of

microorganisms in it and may also lead to the formation of pectinase enzyme which also

has a supplementary role in affecting the quality of pectin (Muhamadzadeh et al.,

2010).In the present study the moisture in the extracted pectin was in accordance with the

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former studies as done by Acikgoz, 2011 and it is also under the required standard limit.(

Brejnholt , 2010).

The standard ash content of the extracted pectin reported to be less than 10% on free

moisture basis. Low values of ash content is among the judge for the purity of pectin

along with other important chemical parameters like galacturonic acid content which

should be 65% (Acikgoz, 2011).The ash content in the current study exhibited in the

range of 4.3% till 5.11% for extracted sapodilla pectin .The equivalent weight of the

extracted pectin showed to be little higher than the food grade pectin for sapodilla. The

pectin extracted at pH3 and pH1 indicated higher equivalent weight (1700 at pH3 and

1975 at pH1) as compared to the pH5 which is 1325. The equivalent weight at pH5 was

comparable with the apple (833.33 to 1666.30, Kumar & Chauhan, 2010) and

commercial pectin (1271±18.4) equivalent weights. Equivalent weight also plays a key

role in the jelling property of pectin, the good jelling ability pectin have higher molecular

weight. (Vaclavik and Christian, 2008). This is due to the fact that equivalent weight can

be influenced by number of free acid and maturity of fruit and the low values of

equivalent weight might be due to some degradation of pectin (Ramli and Asmawati,

2011). It was learned further that increased temperature may results in greater yield of

pectin but it affects negatively on the methoxy content and equivalent weigh of pectin

(Kulkarni and Vijayanand, 2010).

Methoxyl content of the extracted pectin appeared to be 4.4% - 5.1%. Methoxyl content

is an important factor in controlling the setting time of pectins and the ability of the

pectin to form gels (Constenla and Lozano, 2003). Table - 78 shows that the sapodilla

pectin at pH 5 gave higher methoxyl content (10.25%) followed by pH3 (4.4%) and then

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at pH1 (4.9%). These values were approximately similar to earlier reported values as

found for peel of mango (7.33%), pomelo peel (8.57%), Lime (9.92%), (Madhav and

Pushpalatha, 2002), passion (8.81%-9.61%) but higher than dragon fruit pectin (2.98% to

4.34%) (Ismail et al., 2012). Methoxyl content of extracted pectin vary from 0.2-12%

depending on the source and mode of extraction, the percentage methoxyl content

obtained falls within the range. Since all the values obtained experimentally were below

or almost 7%, for sapodilla means that the pectin were of low ester characterization

indicating that the pectin is good in terms of quality .Aina, 2012.

Estimation of anhydrouronic acid content is essential to determine the purity and degree

of esterification, and to evaluate the physical properties. It was found in a range from

61.78% to 69.33% in case of sapodilla which is the desired value in an extracted pectin as

indicated by the food chemical codex 1996 which states that AUA should not be less than

65%. Similar values were observed in apple pomace pectin, commercial apple pectin and

dragon fruit pectin which were 59.52% to 70.50%, (Kumar & Chauhan, 2010), 61.72%

and 45.25% to 52.45% (Ismail et al., 2012) respectively. Low value of AUA means that

the extracted pectin might have a high amount of protein (Ismail et al., 2012).

The gelling capacity of extracted pectin was also evaluated and observed to be in range of

99 to 100 for the extracted pectin. The jelling capacity of fruit pectin was calculated low

which can be due to numerous gelling affecting property for a pectin. One of the reason is

the presence of acetyl group and presence of neutral side chains of sugars which in case

of sugar beet pectin hinders in making gel (Fishman et al 2010). The other factor such as

ash cannot be the cause here as it was already in required limits and it is learned that

lower ash values aids in forming better gels (Ismail et al., 2011).

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The GA contents of sapodilla pectin was observed in the range of 60.5% to 77.7%. The

difference in the galacturonic acid is also due to many factors as Garna et al (2007)

reported that pH and temperature at which the extraction is done influenced the

galacturonic acid content of apple pectin. Also it may be a result of the side sugar chain

hydrolysis of pectin which in turn is a reaction of high temperature extraction. (Fraeye et

al., 2007). However, this could also be the attribute of the type of acid used, as when

citric acid was added for extraction opposite phenomenon was observed and low

temperature resulted in high contents of uronic acid. (Garna et al., 2004). The

hydrochloric acid used in extraction can also have an effect on the galacturonic acid

content as it was also learned that apple when only hot water used during extraction gave

better GA contents compared to the extraction using hydrochloric acid (Hwang, Kim, and

Kim (1998). Moreover increased extraction time also influenced uronic acid value as

studied by Vriesmann et al. (2011). Protein contents are necessary to determine in order

to assess the purity of extracted pectin ( Alba et al 2015) the gelling property of pectin is

also influenced by protein content as in case of sugar beet pectin ( Yapo et al., 2007). The

pectin content for the sapodilla pectin at different pH showed smaller values (1.71- 4.04),

and within limit which also has an effect on its gelling property.

Water holding capacity is also an important parameter in order to understand the physical

characteristic of a compound. The affinity of a particular compound to binds with water

determine its water holding capacity which consequently exerts great influence in

determining the physiological property the compound may carry. The water binding

capacity was greater than the reported values of apple and orange fiber (7.6 for sapodilla)

(Table- 79) while the water holding capacity was less than the reported values of apple

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and orange pectin (6.99 for sapodilla) (Lecumberri et al., 2007) while the fat binding

capacity was quite comparable with the apple, orange and grape fruit fibers (Figuerola et

al., 2005). Physicochemical properties such as water holding, water binding and fat

binding capacities are important to understand as structural information alone cannot

predict the functional properties of dietary fibers and their health benefits. Hence the

physiological effects of fibers rely on inter related combination of physical, chemical and

structural properties (Blackwood at al., 2000). Due to the many reported advantages of

dietary fibers present day researchers and industries are giving it great attention. Dietary

fibers, including pectin have proved affective against the control of cardiovascular, blood

cholesterol, diabetes and colon cancer. With the comprehensive knowledge about the

nutritional as well as functional properties it is assumed that new value added foods and

convenience food products can be developed which are high in demand along with

functional properties of the foods.

The Fourier Transform Infra-Red (FT-IR) study was also carried out to understand the

structural similarities of extracted and food grade pectin available in market. The

absorbance (400-4000 cm-1) of sapodilla peel pectin extracted at different pH were

presented in Figure- 23 to 26.The absorbance region of important peaks are tabulated in

Table - 80. The spectra revealed no prominent structural differences in commercial and

pectin extracted from peel of sapodilla at different pH. The IR absorption peaks achieved

in between 1000-2000 cm-1 comprised of the main functional groups in pectin as these

field of spectra are commonly employed in describing different types of pectin

(Kalapathy and Proctor, 2001). Similar spectral peaks are seen in the absorption range of

1300-1000 cm-1. The spectra also revealed C=O stretching in a range of 1640-1750 cm-1.

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These peaks are of particular interest when degree of esterification (DE) values of pectin

are calculated the reason being the esterified carboxylic group shows an increase in

intensities and band regions as the DE value increases. DE was more than the standard

pectin in all the samples and appeared similar as in C limon which exhibited strong C=O

stretch at 1710-1665 cm-1 Kanmani, P 2014.The C-H stretching of the CH3 group was

also observed in all samples of pectin including the food grade . These O-H stretching

vibrations appeared within a broad range of frequencies (Singthong et al. 2004), followed

by the absorption band of C-H stretching of CH, CH2 or CH3 roughly at 2900 cm-1

(Singthong et al. 2004; Tamaki et al. 2007). The moisture in the samples typically

broadens the band from 2400-3600 which mainly form due to the stretching of the

hydroxyl group. Among the samples least moisture was found in food grade while pectin

at pH 5 has the most. Similar sharp and strong peaks were observed by Kanmanim, P.,

2014 when studying C.limon at 3595.31

The presence of peaks at 1728.22 cm_1 and 1242.16 cm_1 shows the existence of α, β-

unsaturated esters and aliphatic amine functional groups. These absorption peaks are

usually the result of inter and intra molecular hydrogen bonding of galacturonic acid

backbone. The finger print region (1300e800 cm_1) can also identify variations in

monosaccharide composition of pectin (Kamnev et al., 1998; Monsoor et al., 2001),

however, some researchers believe that it is challenging to interpret because of spectrum

overlap (Gnanasambandam& Proctor, 2000), this was also found in the present study and

it was difficult to assign the peaks of fingerprint region to corresponding chemical

analysis results. However, some symbolic characteristic peaks of pectin, for example, 882

cm_1 (pyranose ring), 1273 cm_1 (CeO dilatation vibration), 1453 cm_1 (eCH3

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antisymmetric deformation or the eCH2e symmetric deformation), were observed. The

weak peaks of amide bands (amide I: 1670 cm_1; amide II: 1588 cm_1) in samples

indicated that small amount of protein existed, which also corresponds with chemical

analysis (Table - 80).

Particle size determination was carried though dynamic light scattering studies (DLS)

which is one of most common methods use for particle size. It is used to explore

polydispersity index and size of different extracted and standard pectin. For the detailed

study of particle size apple and orange pectin was also extracted and used in the study.

The apple and orange pectin particle size study gave an insight of other extracted pectin

extracted in the same conditions (Ahmed and others 2016). Table - 81 summarizes the

results obtained after DLS studies, while the illustration of standard and other extracted

pectin are shown from Figure - 28 till Figure - 31. In the figure the autocorrelation

function (ACF) is shown in the top panel denoted by letter A (a). The ACF are usually in

the form of a curve which shows the size of the particle and which is formed after

plotting values of ACF against time. The curve formed specifically after the movement of

particles according to their size, greater the particle size, slower will be its movement and

vice versa. The decay of ACF, with time, is due to the faster agitation of the smaller

particles that leads to the faster decorrelation of scattered light intensity. The central

panel depicts the intensity of radius distribution of the particles denoted by A(b) and are

in the range of 1 nm to 10 μm. The solid line peak indicated the result of 50 experiments

mean calculated by the help of software, while the dotted lines is the basic data received

after the 50 measurements. The bottom panel A(c)) shows the radius plot of all the 50

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measurements which were taken over a period of 1000 s with measurement of every 20 s

and a delay time of 1 s in between.

It represents the differences in the particle size and monodispersity of pectin in each

individual results in 50 measurements. While in B it illustrate the final results of standard

(a) and sapodilla fruit pectin (b). Table -81 demonstrated the radius of hydration (RH)

which is the average particle size and percent polydispersity (% PD) characterized by the

DLS, of standard and the pectin extracted from various other fruits during the study.

Table - 81, presents the average particle size which is expressed in radius of hydration

(RH) and the polydispersity (% PD) which were characterized by DLS studies. It is

observed that most of the extracted pectin (orange, sapodilla extracted at pH 3 and 5)

acquired in relatively smaller and same sizes (~400 - 550 nm), while apple and standard

pectin has quite comparable large sizes (~70 - 1980 nm). While the percent polydispersity

were approximately in identical range for all samples (~20-45%). However it is

suggested that particle size decreases when the pH increases when pectin was

investigated in an aqueous medium (Gbassi and others 2013). It also determined that size

is an essential criteria in assessing the stability and chemical behavior of a particle in a

particular solution or formulation (Gruy 2011). The solubility enhances with the lowering

of particle size in a solution. The dispersion of smaller particle sizes in suspension

dominates the gravitational force which results in keeping the homogeneousness of

suspension intact (Gbassi and others 2013). Observations collected are quite comparable

with a previous study which reported identical particle size data of pectin, while a

contrasting estimation of polydispersity was achieved (Nguyen and others 2011).

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During the studies conducted under extraction of pectin from sapodilla fruit was noted

that a number of factors can influence the yield of pectin from the peel. When there are

many factors influencing a certain response, RSM or response surface methodology is an

effective tool to determine the optimal response (Sun et al., 2011). Box-Behnken design

was used in the RSM study to produce the experimental design that is shown in Tables 82

and 83.

The analysis depicting the variance for yield (ANOVA) is shown in Table- 84. This table

shows that the fitted model is in full quadratic model where the p-value of lack of fit is

0.348 which proposes that the data is adequately fitting for the model. Examination and

evaluation of the variance table shows the linear, quadratic and two-way interactions

between variables. Overall, the model is statistically significant, as indicated by the p-

value of 0.018 for the model. In addition, the small p-values for linear terms (i.e.

p=0.025), quadratic terms (i.e. p=0.015) and two-way interactions (i.e. p=0.045) indicates

a curvature in the response surface. These effects are statistically significant as indicated

by p-values for the pH by time interaction (p = 0.017) and pH squared (p = 0.004). The

value of R-square shows that the fitted model explain 81.35% variation due to pH, time

and temperature that is good but the predicted R-value explain 16.11% variation which is

not proper in the new dataset. R-square (adjusted) refers to the total variation in yield due

to pH, time and temperature. The error term, s = 0.3197. The F-value (22.61) indicates

that pH has a highly significant impact on the yield of pectin and when pH is changed

slightly it has a greater effect on the yield of pectin. Thus pH is an important factor

influencing yield of pectin from sapodilla fruit peel. Similar studies are carried which

also showed that pH has a major influence on yield of pectin (Oliveira and others 2016).

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In addition, the interaction of the pH with the time of heating or boiling is another

important factor that has a significant impact on the yield of pectin.

The coefficients with their standard error, t value and probability for all the terms in the

model are all illustrated in Table -84. In this case, an orthogonal design model was

exercised which led to each effect being assessed independently from each other. Value

for VIFs (variance influence factor) can all be rounded-off to 1, which shows that the

predictors are not interrelated, thus there is no multicollinearity present in the model. The

coefficients in Table-85 indicated the direction of individual variable impact on yield.

According to findings, pH, interaction of pH x pH and pH x time have significant (p <

0.05) impact on yield. All the three terms (i.e. the pH, pH2 and pH x time) denotes the

positive impact of independent variables on yield since they have the positive coefficient

value.

Using approximated coefficients for yield, the following predicted equation or regression

equation was derived which gave the yield of pectin from sapodilla fruit peel.

Yield = 0.76 – 0.947 pH + 0.0784 Temperature – 0.0303 Time + 0.2089 pH2 – 0.000443

Temperature2 + 0.000030 Time2 – 0.00550 pH*Temperature + 0.00750 pH* Time +

0.000037 Temperature*Time

The three dimensional response surface is plotted and illustrated in figure 33-35, where

the vertical axis represents the yield and the two horizontal axes represents any two of the

three affecting variables (i.e. pH, temperature and time). The response surface aids to

understand the optimal level of every variable which affects the pectin yield and also

indicates the interaction of variables that affect the response. The effect on pectin yield

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by the interaction of temperature and pH, the fact that pH has a substantial influence on

the pectin yield and that maximum yield of pectin can be reached at a pH greater than 5

are all shown in Figure-33. Figure -34 represents the effect on pectin yield influenced by

the interaction of pH and time at constant temperature. The result concludes that the

highest yield can be attained when the extraction is done after a heating span of 100

minutes at a pH greater than 5. Figure - 35 demonstrates the effect of the interaction of

temperature and time on the yield of pectin at constant pH. However, the graph shows

that no linear relationship exists between the two quantities and demonstrates that the

impact of the interaction of time and temperature is unresponsive and has no significant

effect on the yield.

In order to determine the optimized response lowest yield (1.4) and target yield (3.5) was

given to generate the best solution to optimize yield. The predicted values are given in

Table – 86 According to response optimization predicted model, the best extraction

variables to extract pectin is to keep a constant pH of 5 at 61.11oC for a heating span of

90 minutes will contribute a 3.7% yield. Pectin was extracted from sapodilla fruit peel

through the predicted model and it was verified, since a value close to the approximated

value of yield (i.e. 3.5%) was extracted when the predicted values of variables (pH,

temperature and time) were followed.

Sapodilla crude pectin extracted was used in the preparation of two common

pharmaceutical solid oral formulations, paracetamol and ibuprofen tablets .The extracted

sapodilla pectin was used as a binding agent in both formulations and were evaluated for

different physico-mechanical and micromeritics properties. As sapodilla pectin is a new

source of pectin which has not been reported in earlier studies in tablet formulation

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therefore, it is important to determine the micromeritics of granules .The nature and

concentration of a binder can effect on the compression, mechanical strength,

consolidation, flow and mechanical strength of a tablet (Rao et al., 2012). Table - 87 and

90 represents the micrometrics properties of granules formulated with different

concentrations of binder which is pectin in this case, in paracetamol and ibuprofen tablets

respectively. The compressibility index of paracetamol came out to be in range from

4.618 to 24.603 %. Among the different formulations F4 and F5 has excellent

compressibility index (4.618% and 5.690 % respectively) and Hausner ratio (F4=1.048

and F5=1.060), F6 was good for both compressibility index and Hausner ratio and F1,

F2, F7 and F9 were fair formulations while F3 and F8 were passable .The angle of repose

came out to be under 21.546. While for ibuprofen carr’s index was excellent for all four

formulations (Table - 90) and excellent to fair Hausner ratio for the four formulations. R1

showed excellent Hausner ratio (1.09) and compressibility index (3.77) while angle of

repose was also better for all four formulations which came between 11.13 to 15.25.

The weight variation of tablet is a credible sign of the uniformity of constituents present

in the formulation which is a basic requirement of good manufacturing practice as well as

important for keeping a constant size of the tablets( Nasrin et al., 2011). Uniformity in

weight of tablet is also required as the table contains the active compound or drug which

should be present in a specific ratio, weight is an important indicator that the drug in the

tablet is in required limit or not. The weight of both the types (paracetamol and ibuprofen

) remained under the limit of ± 5%. The diameter and thickness of all the tablets also

didn’t exceeded from the required level (Table - 88 and 91). Hardness is also an

important parameter which is useful to control chipping, abrasion or breakdown of tablets

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during storage and transportation of tablets (Bano, 2011). Therefore, the method to check

hardness is the breaking of tablet by applying reasonable pressure on it. Hardness is also

an important parameter to ascertain the quality of a drug as it also effects disintegration

and dissolution of a tablet (Nasrin et al., 2011)

In paracetamol tablets,10 and 20 mg of pectin were found unsuitable to achieve desired

hardness granules when compressed into tablets were soft and thus further concentration

of pectin was increased. When the concentration increased from 30mg/tab desired

hardness was achieved (Table-88) however interestingly as the concentration of pectin

was increased upto a certain limit, dissolution was noted to decreased. The best hardness

and dissolution was achieved with the formulation F4 and F5 when the concentration of

40mg /tab and 50 mg/tab were used respectively. Dissolution was recorded as 80.43%

and 86.35% respectively. Further increased in pectin from 60 to 120mg not only

increased the hardness of tablet but also noted to decrease the dissolution significantly.

While the ibuprofen tablets showed a similar type of results. It is also noted that the

higher concentrations of pectin had detrimental effect on dissolution properties of tablets.

Among ibubrufen tablets 50mg of pectin gave the best dissolution results as compared to

the rest of the formulations containing 75mg, 100mg and 125 mg of pectin as a binder. It

was also observed that the hardness of the tablet was successfully increased by increasing

the binder but it significantly affected the tablets dissolution property of ibuprofen tablet.

The bulk and tap density results selected formulation shows that granules have good flow

and compressibility property and same observation supported with the Carr’s index and

Hausner ration results.Tablet hardness usually required between 6 - 14 KP and is

primarily based on the friability less than 0.1% and DT not more than 5 minutes results

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which a drug formulation targets to achieve. The particle distribution result shows good

combination of cores and fine granules for weight variation control during compression.

The thickness of the tablets are also found consistent, indicating reasonable justification

to use blister packaging very common for these ( active pharmaceutical ingredients

(APIs) in Pakistan. Uniformity in blend shows good distribution of API while dissolution

result also support selection of compression parameters.

An antidiarrheal preparation was also formulated with the extracted sapodilla pectin .The

basic evaluation tests (Table- 93) of suspension indicated no major difference between

the physical attributes of suspension made from sapodilla pectin and marketed products

(Figure – 40). Pectin is an effective antidiarrheal agent and together with kaolin it acts as

adsorbent resulting in more solid form stool and it also has affinity to attach the digestive

mucus and toxins hence aids in reducing water loss (Abdullah and Firmansyah, 2013;

Smith, 2013). The use of kaolin-pectin antidiarrheal preparation is still common in

Middle east and Gulf (MOH Saudi Arabia ; HAAD) South Africa (Westhuizen, A. V.

(2010), Indonesia (Abdullah and Firmansyah, 2013) and Australia (Bis pectin,2016)

Although FDA has discontinued the use of kaolin /pectin suspension early 2000, the use

of kaolin pectin as adsorbent has proved effective in the pharmacological treatment of for

noninfectious HIV-associated diarrhea (Dikman et al., 2015). The primary function of

pectin based antidiarrheal is to provide water soluble fibers capable of stimulating

epithelial growth in the colon helping in the reduction of diarrheal frequency. Apart from

this, pectin possess strong water holding capacity, water and fat binding properties which

ultimately helps to change the consistency of stool from watery to soft and also helps in

eliminating excess mucus formed in GI tract. In the present study the water holding

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capacity of extracted pectin was noted as 6.99 g/g while the formulated suspension

indicated 32.69 g/g which was quite comparable with the reference sample (Kepect)

showing value of 33.96 g/g.

For the food preparation one of the formulation was sapodilla jam. The jam was prepared

with three different concentrations (5, 7 and 10 g) of extracted sapodilla pectin. A

standard jam preparation was also made with equal quantity of pectin as F1 of the sample

preparations. It was clearly seen through the results that the concentration of pectin

affected the physical and chemical attributes of jam. A slightly increased amount of

extracted sapodilla pectin was needed to prepare jam which was comparable with the

standard pectin jam.( Figure - 41). The pH of jam should be under limit specified by the

Food and Agriculture Organization of the United Nations to minimize any bacterial

growth which might contaminate the food product. The pH of all the samples prepared

showed with in the specified range (3.00 to 3.30) as shown is Table -94. The pH has also

some influence on the viscosity of finally prepared jam. The gel firmness also depends

upon pH of jam, its range should be between 3 to 3.5 for best results. However the

viscosity depends upon other factors too which makes it difficult to predict for the final

end product. The other factors which influence viscosity of jam are temperature, degree

of methylation and concentration of pectin or any alkaline earth salts which may be

present in it (Panda, 2011). Pectin concentration should increase the viscosity of jam and

was proved during the current study when the viscosity of jam increased with the

increasing concentration of pectin. This is also in accordance with some previous

studies.(Nwson et al.,2014 ; Kar and Arsalan, 1999). Moisture content is another

important determinant in assessing the shelf life or life span of a food product. It also

206

gives basic information for how long a product can be stored safely and effectively

(Ashaye and Adeleke, 2009). The higher the amount of moisture content greater will be

affinity for microbial growth specially fungus and molds. Pectin has a key role in

stabilizing the moisture content present in a finished product (Nwson et al., 2014). The

moisture content also came into the specified range which was reported in earlier studies

(Ajenifujah-Solebo and Aina, 2011).The moisture content was also observed six month

after the formulation of the test samples and no bacterial growth was found in it. Table –

94 also shows the value of ash present in the test as well as standard formulations which

is also an indicator of stability criteria of the formulated product. The increased value of

ash enhances the vulnerability of product in terms of stability. The test samples exhibits

ash values ranging from 1.69 to 1.73% which is same in value of the standard product

and hence showed to be a stable product.

Another important in jam processing is the level of acidity in fruit pulp which also affects

the gel formation. The total titratable acidity (TTA) of the jam under investigation also

observed to be within the range of standard values (1.01 to 1.14, Table - 94) which also

predicts that the jam prepared was of good quality. Total soluble sugar (TSS) was also

measured which is an important observation during the storage of jam for assessing its

quality. It was reported in earlier studies that with a 5% level of pectin and pH ranging

from 3.0 to 3.5 the TSS should be 65%. (Ndabikunze et al., 2011) Among all the test

formulation only F3 showed the values of TSS to be in desirable range. It was also

learned that TSS decreases if the product is not refrigerated and kept at room temperature

after formulation ( Sindumathi and Amutha, 2014). Vitamin c (ascorbic acid) contents

were also determined during the study which is also shown in Table 94. The values were

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in the range from 17.6 to 18.3. Vitamin C is a natural antioxidant which determines the

quality of a product. As pectin is used as a gelling agent and helps the jam in setting

faster at lower temperature also aids in preserving the heat sensitive nutrients present in

the jam (Nwson et al., 2014).

Sensory evaluation of sample as well as standard jam was performed on a nine point

hedonic scale as shown in Table - 95. The data collected from the test showed that the

amount of pectin used in each formulation affects positively the sensory attributes of jam.

Among the three formulations F1 showed good scores for appearance, aroma and

spreadibility (7.5, 7.2 and 7.3 respectively) but a moderate score for taste and mouth feel

( 7.3 and 7.0). The results can be linked to the amount of pectin present in the

formulation. The formulation F2 exhibited highest score and was comparable with the

scores achieved for standard formulation. Score for appearance, taste, mouth feel and

aroma were almost same as that of standard jam (7.5, 7.6 7.5, and 7.7 respectively).

While spreadibility of F2 jam was slightly lesser (7.8 for F2 and 8.2 for S). The least

acceptable formulation was F3 in which the highest amount of pectin was used. It was

observed that the high amount of pectin affected the formulation negatively and got the

least scores. The jam also became harder and hence the overall acceptability score was

observed as 6.03. The cooking of the jam usually caramelize the sugar and hydrolyzes

pectin in it which can be one reason of low scores of mouth feel and taste in formulation

F1 and F3 (Fishman and Jen, 1986). Pectin is used as a thickening agent in formulations

hence the increased amount in formulation F3 resulted in the thickening of jam more than

it is desired which also affects the texture, spreadibility and mouth feel of jam (Broomes

and Badrie 2010).

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Another preparation which is used to investigate the functional property of pectin was the

formulation of pudding with the extracted and standard commercial pectin. Textural

property of pudding was the main attribute which is studied here as it is the main desired

quality a good pudding should possess. The failure of the participant to establish any

difference in the pudding texture made from two sample concentration of sapodilla

pectin points towards the similar physical attributes which all the pudding possessed. The

study also gives a slight insight of the role of pectin in milk products. It was studied

earlier that pectin in milk preparations aids in minimizing the agglomeration of protein

which helps to keep the phase in the milk product intact and prevents separation. Pectin

due to its thickening property also has a role in the specific type of mouthfeel which is

required for a particular product (www.herbstreith-fox 2016) According to a some

details published on a company website (Cargill foods 2016). HM pectins are good

stabilizers in acid milk drinks and probably the two puddings pass the tests made from

sapodilla pectin due to this property. It was also understood from the site that HM pectins

creates a coat on the casein particles of milk products. The reason for making this food

formulation was to determine the effect of extracted sapodilla pectin in a formulation

other than the conventional use that is gel formulation.

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Conclusion

Comprehensive knowledge on carbohydrate chemistry and skills in hydrocolloids are the

basic and prerequisite for developing new pectin source as well as to ascertain the

quantity and quality of pectin along with its applications. On the basis of sound

background on both the disciplines, one can rely on the selection of raw material as a

starting source of pectin, establishment of extraction protocol and the bio-characterization

of the end product to find its proper utilization. In view of this, current study was

designed to investigate possible new indigenous source (fruits) of pectin, utilization of

different physico-mechanical procedures for the extraction of pectin from fruit wastes

and to explore one best fruit for in-depth studies. The best identified fruit in this study is

observed as Sapodilla (Manilkara zapota) and thus a comprehensive plan of work was

designed and attention was driven towards the pectin extracted from sapodilla for

physicochemical characterization and its application in food and pharmaceutical products

development. In conclusion, the whole study and suggestions for future studies have been

summarized below.

(i) Mechanical procedure noted to be a potential tool for the extraction of pectin.

Out of five mechanical procedures and five different pH, Chopping was

observed the best mechanical procedure at pH 3 for sapodilla fruit peel when

microwave was used for extraction of pectin. For future studies, suggest to

develop a pilot scale setup to optimize the selected procedure for possible

commercial exploitation.

210

(ii) The bio-characterization along with FTIR and DLS were found excellent

methodologies to study type, structure and particle size of extracted pectin.

However, suggest to utilize more sophisticated spectroscopic analysis such as

GCMS to detect the monosaccharide composition for more knowledge of the

fine structure of pectin and for its specific application.

(iii) Since process optimization has significant effect on the yield of pectin,

therefore various parameters were utilized for high yield of pectin and results

were authenticated through RSM studies. For future studies, suggest to use

RSM technique to authenticate the mechanical procedure at different pH as

well as in the bio-characterization of pectin such as degree of methylation and

degree of acetylation.

(iv) The formulation of solid oral pharmaceutical (tablets) is a very vast field and

needs good and cost effective binders in developmental work. The extracted

Sapodilla pectin was observed a good binder to be utilized in some tablet

formulation. Although dissolution studies were conducted, however it is

suggested to perform kinetic studies as well in future for more authentic data

and effect of Sapodilla pectin as a binder.

(v) Antidiarrheal preparation formulated in the present studies were compared

with the marketed product showed similar results, especially when tested for

its water holding capacity. In addition, the water binding and fat binding

capacities of extracted pectin indicated a promising results. This confidently

pointed towards the potential use of sapodilla pectin in antidiarrheal

211

preparation. However, in vivo procedures can add more data if performed in

future studies.

(vi) Pectin has great food applications. Present study demonstrated potential use of

extended sapodilla pectin in the preparation of Jam and pudding. However,

the present formulation needs further optimization for reproducible results

despite the fact that both formulation showed similar results when compared

with the formulation prepared from standard pectin. Yogurt and milk

preparations should also be formulated and tested to find more specific role of

sapodilla pectin in future.

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