PHYSICO-CHEMICAL CHARACTERIZATION AND STABILITY STUDIES OF ANTHOCYANINS EXTRACTS

FROM JUSTICIA carnea HOOKER LEAVES, AND ITS POTENTIAL USES IN SELECTED FOOD SYSTEMS

  

EMEKA NNAEMEZIE(172488)

 DEPARTMENT OF FOOD TECHNOLOGY

FACULTY OF TECHNOLOGYUNIVERSITY OF IBADAN

  

NOVEMBER, 2014.

INTRODUCTION

Color is one of the most important quality attributes affecting the consumer's acceptance of food since many convenience foods would thus appear undesirable without the inclusion of colorants (Hirunpanish et al., 2006).

Anthocyanins are naturally occurring water soluble plant pigments

The interest in natural colorants has increased significantly recently (Huck and Wilkes, 1996).

Of special interest to the food industry is the limited availability of red pigments (Lauro and Francis, 2000; Azza et al., 2011).

Introduction Continues

Red beet pigments have been extensively used worldwide; the red-violet anthocyanin pigments from Roselle have been employed also in food colorants (Schnetler and Breene, 1994; Hendry and Houghton, 1996). Other sources include berries, red onions, grapes (Rein, 2005).

Till date the supply of natural colorants is still insufficient, therefore other sources are yet to be explored

therefore in this study, anthocyanins of Justicia carnea H. leaves which has not much been documented on was explored.

Justicia carnea Hooker is grown in large commercial quantity in the eastern part of Nigeria where the plant infusion serves as beverage.

STUDY OBJECTIVE

The objective of this study includes;

to extract, freeze dry, and store pigments while monitoring likely changes in physicochemical indices particularly color intensity and stability

to characterize the Anthocyanin compounds and to evaluate possible changes during storage

In addition, to study color intensity and stability in selected food systems, as well as to consider consumer acceptance.

Plate 2.1. Justicia carnea Hooker plant. Source: The virtual field Herbarium (Hawthorne et al., 2001).

MATERIALS AND METHODS

Materials and Sourcing Justicia carnea Hooker leaves were used as source of the natural pigment (extracts) in this research study. Justicia carnea H. leaves were identified in the Herbarium of the Botany Department, University of Ibadan and were collected from the University’s Botany Garden.

Pre-processingDrying of leave samples (Oven and Air drying)Milling into powder

Extraction of Pigment

Justicia carnea H. leaves pigments were extracted with 1% Citric acidified Ethanol, 2% Citric acid solution and Distilled water (60oC).

Physicochemical Analysis

The following physicochemical analyses of the freeze-dried pigments were analyzed at weekly intervals for the period of eight (8) weeks to determine

Color Intensity

Determination of pH

Determination of Ascorbic Acid

Total Pigment Content, etc.

RESULTS AND DISCUSION

Physicochemical properties of freeze dried pigments extracts of oven dried and ambient room dried samples of Justicia carnea Hooker (leaves) extracted with varying extraction media are shown table 4.1 below.

Table 4.1: Physicochemical properties of samples extracts of J. carnea H. leaves

PROPERTIES ODJP ADJP ODEE ADEE ODCE ADCE ODWE ADWE E129

COLORL*a*b*

22.23±0.05g

6.04±0.04b

2.78±0.02e

23.99±0.05f

5.08±0.02c

2.44±0.03e

27.30±0.02d

5.88±0.01c

3.80±0.02d

24.23±0.01e

1.19±0.05f

-0.46±0.05g

51.35±0.05b

8.82±0.01a

15.09±0.04a

55.78±0.03a

4.63±0.03d

10.36±0.01b

35.29±0.04c

3.65±0.01e

5.71±0.01c

51.61±0.03b

1.99±0.04f

3.75±0.02d

18.295.430.64

Solubility (%)

4.14±0.03e 4.53±0.05e 5.18±0.05d 5.21±0.04d 6.16±0.08c 6.20±0.03c 7.04±0.08b 7.00±0.00b 50.00±0.01a

Moisture Content (%)

10.03±0.01e 12.20±0.04d 12.96±0.02d 13.11±0.10c 12.89±0.04d 14.01±0.11b 16.00±0.16a 16.62±0.02a 0.20±0.02f

pH 3.47±0.01c 3.31±0.05c 2.72±0.02d 2.66±0.04d 3.07±0.00c 3.14±0.05c 8.65±0.02a 8.65±0.02a 6.20±0.22b

% Titratable Acidity

2.84±0.11c 2.91±0.08c 3.07±0.01b 3.20±0.00b 5.74±0.02a 5.92±0.14a 0.13±0.02d 0.16±0.03d _

Total Soluble Solids(0Brix)

5.22±0.04e 5.24±0.05e 6.20±0.00d 5.30±0.00e 15.87±0.00a 15.76±0.00a 9.89±0.01c 10.51±0.00b _

Ascorbic Acid(mg/100g)

110±0.05c 107±0.02d 135.96±0.12b

130.10±1.10b

148.21±0.23a

141.81±0.21a

108.10±0.02d

99.32±0.08e _

Anthocyanins(mg/100g)

1880.04±1.14a

1826.00±0.56a

1329.15±0.02b

1035.57±0.04d

437.38±0.11e

452.69±0.07e

1393.55±1.20b

1044.08±0.02d

_

OH

HO

OH

OH

OH

OCH3

HOOH

HO

OCH3

OH

OH

Structures of Anthocyanins Present In Justicia carnea Hooker Extract

Figure 4.1: Delphinidin3, 4’-diglucoside

OH

HO

OH

OH

OH

HO

OH

OCH3

HO

OH

HO

OCH3

Figure 4.2: Malvidin 3,5-diglucoside

H3CO

H3CO

oHOCH3

OCH3

Figure 4.3: Malvidin 3-(2”- glucosyl – (3-oxalate) -6”- (6-ferulic)), 5-glucoside

Type Concentration (%)

Malvidin 3,5-diglucoside

Delphinidin 3,4’-diglucoside

Malvidin 3-(2”-glucosyl-(3-oxalate)-

6”-(6-ferulic)),5-diglucoside

10.09

2.96

11.61

Total 24.66

Table 4.2: Types and concentrations of Anthocyanins found in Justicia carnea H. extracts

1 2 3 4 5 6 70

1

2

3

4

5

6

7

8

9

10

ODAEEODCAEODWIE

Time (Weeks)

Chro

ma

valu

es (a

*)

Figure 4.4: Variation in Color Intensity (Chroma , a*) of oven dried samples of Justicia Carnea H. extracts. 

1 2 3 4 5 6 70

1

2

3

4

5

6

7

8

ADAEEADCAEADWIE

Time (Weeks)

Chr

oma

valu

es (a

*)

Figure 4.5: Variation in Color Intensity (Chroma a*) of Air dried samples of Justicia Carnea H. extracts.

Figure 4.6: Variation in pH of oven dried samples of Justicia Carnea H. extracts.extracts.

1 2 3 4 5 6 70

1

2

3

4

5

6

7

8

9

10

ODAEEODCAEODWIE

Time (Weeks)

pH

Figure 4.7: Variation plot for pH readings of Air dried sample extracts of Justicia carnea H..

1 2 3 4 5 6 70

20

40

60

80

100

120

140

160

ODAEEODCAEODWIE

Time (Weeks)

Ascu

rbic

Acid

(mg/

100g

)

Figure 4.8: Variation in Ascorbic acid content of Oven dried sample extracts.

1 2 3 4 5 6 70

20

40

60

80

100

120

140

160

ADAEEADCAEADWIE

Time (Weeks)

Asco

rbic

Acid

(mg/

100g

)

Figure 4.9: Variation in Ascorbic acid content of Air dried sample extracts.

Figure 4.10: Variation in Anthocyanin content of the oven dried Justicia carnea H. sample extracts.

1 2 3 4 5 6 70

500

1000

1500

2000

2500

ODAEEODCAEODWIE

Time (Weeks)

Anth

ocya

nins

(mg/

100g

)

Figure 4.11: Variation in Anthocyanin content of the Air dried Justicia carnea H. sample extracts.

1 2 3 4 5 6 70

500

1000

1500

2000

2500

ADAEEADCAEADWIE

Time (weeks)

Anth

ocyn

ins (

mg/

100g

)

Figure 4.12: Variation in Color Intensity (Chroma (a*)) of Spirit-Mixed Drink samples.

1 2 3 4 5 60

5

10

15

20

25

30

35

40

DRINK BDRINK CDRINK D

Time (Week)

Chro

ma

(a*)

Plate 1: From left to right – Spirit-mixed drink colored with synthetic colorant (E129), Spirit-mixed drink colored with Ethanol pigment extracts, Spirit-mixed drink with no colorant, and Spirit-mixed drink colored with Water-infused pigment extracts.

Figure 4.13: Variation in Color Intensity (Chroma (a*)) of colored Watermelon Juice samples.

1 2 3 4 5 60

5

10

15

20

25

JUICE AJUICE BJUICE CJUICE D

Time (weeks)

Chro

ma

(a*)

Plate 2: Watermelon Juices made with the inclusion of colorants, natural and synthetic.

Juice made with synthetic colorant Juice without colorant

Juice made with ethanol extract colorantJuice made with water-infused colorant

Figure 4.14: Changes in Color Intensity (Chroma (a*)) of colored Butter-Icing samples.

1 2 3 4 5 60

5

10

15

20

25

ICING AICING BICING CICING D

Time (weeks)

Chro

ma

(a*)

Plate 3: Cake-Butter Icing. Left part is colored with citric acid extract and right part is colored with ethanol extract of Justicia Carnea H. pigments.

Values with different superscript letters in each row are significantly different (p<0.05)

Table 4.3: Sensory result of Spirit-Mixed Drinks

SAMPLES DRINK A DRINK B DRINK C DRINK D

COLOR 7.00±1.03c 7.90±0.70a 7.36±0.48b 7.90±0.78a

MOUTH FEEL 7.20±0.72c 7.34±0.77bc 7.68±0.68a 7.56±0.57ab

FLAVOR 7.70±0.58ab 7.58±1.10b 7.96±0.80a 7.86±0.72ab

TASTE 7.12±0.79c 7.52±1.18b 8.02±0.82a 7.94±0.71a

OVERALL ACCEPTABILITY 7.04±0.75b 8.16±0.73a 7.30±0.88b 8.22±0.61a

SAMPLES DRINK A DRINK B DRINK C DRINK D

COLOR 6.74±0.87c 7.62±0.80a 7.08±0.27b 7.58±0.88a

MOUTH FEEL 6.88±0.47c 7.06±0.65bc 7.38±0.69a 7.24±0.47ab

FLAVOR 7.18±0.56b 7.18±1.06b 7.58±1.03a 7.36±0.63ab

TASTE 7.02±0.79a 6.96±1.14a 7.14±1.06a 7.34±1.08a

OVERALLACCEPTABILITY 6.76±0.71c 7.64±0.72a 7.12±0.77b 7.48±0.86a

Table 4.4: Sensory result of Spirit-Mixed Drinks after storage

Values with different superscript letters in each row are significantly different (p<0.05)

SAMPLES JUICE A JUICE B JUICE C JUICE D

COLOR 7.84±0.73a 7.56±1.18a 7.88±0.77a 7.82±0.71a

MOUTH FEEL 7.14±0.83c 6.40±0.94d 7.68±0.55a 7.54±0.57b

FLAVOR 6.86±0.92c 7.24±0.89b 7.70±0.67a 7.54±0.61ab

TASTE 7.14±1.74bc 6.92±0.85c 7.72±0.60a 7.48±0.67ab

OVERALL ACCEPTABILITY 8.02±0.65a 7.80±0.67a 7.98±0.74a 7.74±0.69a

Table 4.5: Sensory result of colored Watermelon Juices

Values with different superscript letters in each row are significantly different (p<0.05)

SAMPLES JUICE A JUICE B JUICE C JUICE D

COLOR 6.66±0.74b 7.28±0.83a 7.16±0.37a 7.38±1.08a

MOUTH FEEL 6.62±0.60b 6.88±0.77ab 7.14±0.88a 7.02±0.68a

FLAVOR 7.06±0.68ab 6.84±0.99b 7.24±1.13a 7.06±0.66ab

TASTE 6.88±0.68b 6.82±1.06b 6.86±0.85b 7.34±1.08a

OVERALL ACCEPTABILITY 6.72±0.72c 7.60±0.83a 7.10±0.76b 7.28±0.70b

Table 4.6: Sensory result of colored Watermelon Juices after storage

Values with different superscript letters in each row are significantly different (p<0.05)

SAMPLES ICING A ICING B ICING C ICING D

COLOR 6.20±0.80c 8.13±0.68a 7.33±0.75b 7.53±0.77b

MOUTH FEEL 7.23±0.93b 7.80±0.96a 7.13±0.83b 6.90±1.21b

TEXTURE 6.46±1.07a 6.43±0.97a 6.13±0.86a 6.70±1.41a

TASTE 5.86±1.00c 7.36±1.44b 7.23±0.97b 8.10±1.12a

OVERALL ACCEPTABILITY 6.36±0.96c 8.03±1.09a 6.86±1.00bc 7.26±1.01b

Table 4.7: Sensory result of colored Cake Butter Icing

Values with different superscript letters in each row are significantly different (p<0.05)

SAMPLES ICING A ICING B ICING C ICING D

COLOR 7.56±1.04a 7.86±1.30a 7.33±0.84a 7.30±0.98a

MOUTH FEEL 5.23±0.82a 5.50±0.82a 5.10±1.02a 4.20±0.55b

TEXTURE 4.23±0.62a 4.40±0.88a 4.56±1.10a 4.13±0.50a

TASTE 5.46±1.10b 6.66±1.06a 6.86±1.27a 5.26±0.88b

OVERALL ACCEPTABILITY 5.50±0.81b 6.83±1.28a 5.63±0.88b 5.23±0.50b

Table 4.8: Sensory result of colored Cake Butter Icing after 6weeks of storage

Values with different superscript letters in each row are significantly different (p<0.05)

Conclusion and Recommendation

From this study which explored yet another potential source of anthocyanin type pigment, it was observed that the total anthocyanins content derived from Justicia carnea H. extracts, could serve as a good source of natural colorant with potentials as food coloring additive since the pigments retained till the end of storage, substantial percentage of the original concentration and color intensity which compares well with reports given on anthocyanins content of other sources (Walford, 1980; Thakur, 1989; Rein, 2005). Of which Malvidin was highest in concentration till the end of storage. This high concentration of malvidin makes research importance, the individual extraction and isolation of this compound from Justicia carnea H. as it is basically the pigment responsible for the retained color intensity. However, molecular copigmentation and encapsulation of anthocysnin pigments has proven to preserve better, the color intensity of the pigments during shelf storage.

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Appendix VII

Plate 1: From left to right – Spirit-mixed drink colored with synthetic colorant (E129), Spirit-mixed drink colored with Ethanol pigment extracts, Spirit-mixed drink with no colorant, and Spirit-mixed drink colored with Water-infused pigment extracts.

Plate 2: Watermelon Juices made with the inclusion of colorants, natural and synthetic.

Juice made with synthetic colorant Juice without colorant

Juice made with ethanol extract colorantJuice made with water-infused colorant

Plate 3: Cake-Butter Icing. Left part is colored with citric acid extract and right part is colored with ethanol extract of Justicia Carnea H. pigments.