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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.
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