Malaysian Cocoa Journal, 8/2014

ANTIOXIDANT PROPERTIES OF COCOA PODS AND SHELLS

Azila Abdul Karim1, Azrina Azlan2 3, Amin Ismail2, Puziah Hashim3, Nur Azilah Abdullah1
1
Cocoa Innovation & Technology Centre, Malaysian Cocoa Board, PT12621, Nilai Industrial Area, 71800
Nilai, Negeri Sembilan Malaysia; e-mail: aziela@koko.gov.my
2
Department of Nutrition and Dietetics, Faculty of Medicine and Health Sciences, Universiti Putra
Malaysia, UPM Serdang, Selangor, Malaysia;
3
Halal Product Development Institute, Universiti Putra Malaysia, UPM Serdang, Selangor, Malaysia

Malaysian Cocoa J. 8: 49 – 56 (2014)

ABSTRACT - This study was conducted to compare antioxidant properties of cocoa pods and shells, in
terms of its level and activities. The antioxidant level was determined by total phenolic and total flavonoid
contents using spectrophotometric method by Follin-Ciocalteau’s reagent and aluminum chloride solution,
respectively. Antioxidant activity of these extracts were measured using DPPH and FRAP assays. Cocoa
pods contained promising amount of total phenolic and flavonoid content in comparison to cocoa shells.
Antioxidant activity found using DPPH assay showed that cocoa pods exhibited highest activity in
comparison with the cocoa shells although lower than gallic acid standard. Significant high antioxidant
activity of cocoa pod extract was clearly exhibited using FRAP assay in comparison with cocoa shells.
Therefore, cocoa pods are potential waste materials that can be used as raw materials for other beneficial
application including cosmeceutical product.

Keywords: cocoa pod, cocoa shell, total phenolic content, total flavonoid content, scavenging effect

INTRODUCTION solution in industrial line (Meunier, 2003). In

other application, cocoa shells were used in
Abundant of waste from cocoa areas can be particleboard making as its surface layer (Hii,
turned into value-added end products. It was 2006) and as a source of pectin (Mollea, 2008).
estimated that only 25% from total weight of Cocoa shells, also known as cocoa hulls, showed
fresh cocoa fruits were used to produce cocoa positive antioxidant activity when evaluated in
beans for cocoa and chocolate processing. Other vitro (Arlorio, 2005). Total phenolic content
parts including the pods and shells are usually (TPC) of non-roasted cocoa husks (cocoa shells)
discarded. Despite of being thrown away, cocoa have been measured by Ramos (2008) using
pods can also be used as sources of fertilizer or catechin as standard, of 70% methanol-acidic
animal feed and activated carbon production extract after enzymatic hydrolysis treatment to
(Fisal, 2005). Other potential use of cocoa pods obtain dietary fiber. Strong antioxidant activity
are in producing gum (Samuel, 2006), potash in was exhibited by cocoa shells (Azizah, 1999).
soap-making as well as food colorant (Figueira, Recumberri (2007) also evaluated the total
1993). The pods contained 45.6–46.4 mg gallic antioxidant of cocoa shells along with the
acid equivalent of soluble phenolic; 32.3% determination of dietary fiber composition in
carbohydrate, 21.44% lignin, 19.2% sugars, cocoa shells. Amin (2006) determined that total
8.6% protein and 27.7% minerals (Vriesmann, phenolic content of cocoa shells was 112.9 + 0.6
2011). Arsel (1992) and Fapohunda (2012) mg GAE/g extract and scavenging activity at
reported that cocoa pods contained phenolic acid, 95.9 + 2.0%, which was extracted using 70%
including caffeic acid, but no specific antioxidant ethanol. In contrast, extract of 80% ethanol
measurement was carried out. results in total phenolic content at 23.36 + 1.59
mg GAE/g extract and 2.23 + 0.90 mg CE/g total
Cocoa shells, which is enclosed the cocoa seeds flavonoid using catechin as standard (John,
is endocarp (Bewley, 2006) and must be 2012). Cocoa shells and cocoa pods extract also
removed after roasting process to obtain the exhibited antimicrobial activities in reduction of
cocoa nibs. Cocoa shells are a potential source Streptococcus mutans and mouth plaque
of dietary fibre (Maria, 1994) and fertilizer (Srikanth, 2008).
(Chung, 2003) due to its mono &
oligosaccharides, starch, pentosans & cellulose Mechanism of skin wrinkles formation was
content, and removal of heavy metal from acidic extensively reported by Mukherjee (2011) and
Malaysian Cocoa Journal, 8/2014

bioactives from plants that exhibited anti- Board) in Hilir Perak, Perak and Jengka, Pahang.
wrinkles effects including catechin, epicatechin For comparison purpose, cocoa pods and shells
and gallic acid. These compounds have high were collected by clones, namely, MCB C1 (C1),
antioxidant activity. Addition of extract from MCB C3 (C3), MCB C4 (C4), MCB C5 (C5)
plants can delay the oxidation process by radical and PBC123 (PBC). In addition, a collection of
oxygen species (ROS) which is responsible for cocoa pods were labeled MIX representing cocoa
the formation of wrinkles on the skin. Evans pods which has been collected in bulk due to the
(2010) reported that phenolic compounds such as practices of plantation where the pods where
catechin, epicatechin, quercetin and kaempferol discarded despite their clones. Pods collected
with antioxidant activity reduced oxidative stress were washed thoroughly with tap water to
including skin wrinkles and photoaging (Jung, remove debris and chopped into about 1cm2 in
2007). Garbisa (2011) proved that quercetin and size and sun-dried. Chopped pieces of pods
kaempferol inhibit MMP-1 activity, in-vitro, and were placed on the net and exposed to direct
topically (Casagrande, 2006). MMP-1 is a sunlight until dried. The dried cocoa pod husks
collagenase enzyme involve in degradation of were ground into powder form using warring
collagen resulting the wrinkle formation (Kim, blender. Ground pod were sieved using
2006). Report by Aburjai (2003) revealed that automatic sieve to mesh size of 1 mm. Sample
catechin and epicatechin can treat dry skin. of the shells was collected after roasting the
Polyphenolics substances delayed skin aging, in- cocoa beans of fermented said beans as usually
vitro (Rutter, 2003) and exhibited UV-radiation practices in the grinding industry. In this
protection (Nichols, 2010). Antioxidant activity experiment, cocoa shells, irrespective the clones,
of plant phenolic extract offered an alternative in was collected from the industry and labeled
producing natural anti-wrinkles cream MIX. The pod and shell was labeled as P for pod
(Baumann, 2007 & Masaki, 2010). and S for shell, respectively.

This study was carried out to determine total Extraction

phenolic and flavonoid content of cocoa pods in Extraction was carried out using aqueous ethanol
comparison with the shells. The antioxidant (80% v/v). The method described by Catherine
activity was also measured using DPPH and (2003) was used for the extraction procedure,
FRAP assays. Results of this experimental will where 80% ethanol (Sanbogi, 1998) was
be used for the selection of materials for anti- selected. One gram of pod powder was soaked
wrinkles cosmetic formulation. in 20 ml of aqueous ethanol in 50 ml flask with
cap. The flask was placed in water bath shaker
for 30 minutes, 35oC at 120 rpm. Decanted
portion was then filtered using Whatmann filter
MATERIALS AND METHODS No.1 to remove remaining floating materials.
Solvent was removed using rotary vacuum
Reagents evaporator to dryness. Crude extract was re-
Denatured ethanol (95%) from Hmbg, anhydrous dissolved in 1-2ml aqueous ethanol (80% v/v).
sodium carbonate from AR, Gallic acid (3,4,5- Extracted portion were again filtered and filtered
Trihydroxybenzoic acid) from Sigma-Aldrich, portion was subjected to the analysis. The
Folin-Ciocalteau (FC) reagent from Merck, extraction was carried out in triplicate.
aluminum chloride (AlCl3.6H2O, MW 241.43
g/mol) from FriendemannSchmidt, Rutin Total Phenolic Content (TPC) Determination
(rutinoside: 2-(3,4-dihydroxyphenyl)-5,7- Kaur (2002) method was used with several
dihydroxy-4-oxo-4H1-benzopyran-3-yl modifications to measure the total phenolic
rutinoside, C27H30O16) from Powder Spectrum content in cocoa pod using Folin-Ciocalteau (FC)
and Methanol from Merck were used in this reagent. Total phenolic content of the samples
experimentation. In addition, 1,2-diphenyl-2- was obtained by the calibration curve of standard
picrylhydrazyl (DPPH, C18H8N5O6, MW gallic acid measured. Hundred (100) µl of
394.23g/mol) was purchased from Aldrich and extracted sample (or standard) was pipette into
dimethylsulfoxide (DMSO) from Merck for total vial. Seven point nine millimeters (7.9ml) of
antioxidant activities measurement. distilled water was added to the sample (or
standard). Five hundred (500) µl of FC was
Sample collection added to the diluted sample and mix for 8
Cocoa pod were collected from Cocoa Research seconds. Then, 1500 µl of sodium carbonate
& Development Centre (Malaysian Cocoa solution (20% w/v) was added and left in the
Malaysian Cocoa Journal, 8/2014

dark at 40oC for 1 hour. Measurement was method from Benzie (1996), Szollosi (2002),
carried out with UV-Spectrophotometer at Katalinic (2004), Arnous (2002), Pulido (2000)
765nm, twice, against gallic acid standard. The and Bub (2000). In brief, ferric chloride solution
results obtain were expressed in mg Gallic Acid was made by dissolving 3 mM of Ferric chloride
Equivalent (GAE)/g sample. in 5 mM citric acid with distilled water. TPTZ
solution was prepared by dissolving 1 mM of
Total Flavonoid Content (TFC) Determination TPTZ in 0.05M hydrochloric acid. Standard
Measurement of total flavonoids content was calibration curve was obtained by different
carried out using microplate reader at wavelength concentration of FeSO4.7H2O (1.0 – 0.1 mM
of 405 nm using aluminium chloride colorimetric Fe(II)). To measure the FRAP value, 15μl of
assay. Method by Chang (2002) was adapted. sample or standard was added to 270 μl of TPTZ
Solution containing 2% (w/v) of aluminium solution and measure at 620nm for initial reading
chloride was prepared by dissolving 2g of using microplate reader. Ferric chloride solution
aluminium chloride with 100 ml of methanol. at 15μl was added and measurement was carried
Extract of sample 0r standard was pipetted at 100 out at immediate. The sample was then
µl into 96-well microplate and 100 µl aluminium incubated for 30 minutes (30-37oC) with interval
chloride solution was added. Incubate the measurement for 2-3 minutes. The result was
sample for 10 minutes and measure the compared with gallic acid or ascorbic acid. EC 1
absorbance at 405 nm. Total flavonoids content is effective concentration of the extract required
was obtained using calibration curve of rutin to react with 1 mol Fe (II)/L. Higher value
standard. The samples were expressed in mg indicated lower antioxidant activity, which
Rutin Equivalent per gram sample (mg RE/g). means high concentration of extract was needed
to reduce 1 mol of Fe (III) to Fe (II) referring to
the mechanism of antioxidant compound to
Determination of DPPH Scavenging Effect and FRAP assay.
EC50
Antioxidant activity was be carried out using
DPPH assay as described by Patil (2010), Young Statistical analysis
(2008), Mustafa (2006) and Kim (2009) methods Standard calibration curves were accepted when
with some modifications. In brief, the DPPH the R-squared of the linear regression was higher
stock solution was prepared by dissolving 0.2M than 0.999. One-way ANOVA was used to
of DPPH in ethanol. Solution of DPPH was screen the difference value of overall TPC and
made by diluting 1.2ml of DPPH stock solution TFC content of cocoa pod and shell extract from
with 3ml ethanol and 0.5ml DMSO. For clones. Two sample t-test was used to indicate
antioxidant activity, samples were made into significant difference between experimental data
serial dilution of 1000-7.81 ppm and measure for in this study and from literature, if any. This test
comparison. Then, 270µl of DPPH solution was was also carried out to see the difference on
added to 30µl sample at different concentration, DPPH scavenging effect. Level of significant in
shake and left it to react for at least 10 minutes this study was at level p<0.05, using Minitab
before measuring at absorbance 550nm. Gallic Software version 14.
acid standard was used as positive control.
Ethanol with the addition of DPPH solution was
used as blank (Lee, 2003). Calculation of EC50 RESULTS & DISCUSSION
(the concentration needed to reduce DPPH by
50%) was obtained using linear regression Total phenolic content
(Marxen, 2007) on the graph plotted of Calibration curve for standard gallic acid at
remaining DPPH percentage versus 765nm was plotted and regression analysis was
concentration ratio of sample to DPPH. Small carried out, where the equation (1) was obtained
value EC50 indicates low concentration of sample (R2=0.999) and used to predict the total phenolic
required to exhibit antioxidant activity by this content (mg GAE/g) of the samples with the
assay. dilution factor.
Determination of FRAP and EC1
Method of conducting FRAP assay for cocoa pod
extract was carried out with modification of
A765 = -0.003800 + 0.000817*concentration (1)
Malaysian Cocoa Journal, 8/2014

Where A765 is absorbance value at wavelength 765nm and concentration is in part per millions (ppm) of
total phenolic content.

Figure 1: Total Phenolic Content

total phenolic of cocoa shell and pod
60

50
shell pod

mg GAE/g extract
40

30

20

10

0
C1 C3 C4 C5 MIX PBC

Number of sample taken was repeated several shells was varied depending on the source of
times for different samples to obtain low relative collection as reported by Amin (2006) and John
standard deviation (RSD < 25%) indicating the (2012). Weather, soils and environmental factors
data are representative. It was shown in Figure 1 contributed to the chemical and physical content
that pod labeled MIX (49.54 + 3.69 mg GAE/g of the plants. It was also shown that between
extract, n=6) has the highest total phenolic pod and shell, within clones, total phenolic
content among the selected clones, equally to C4 content was significantly higher except for C1,
(47.57 + 3.23 mg GAE/g extract, n=6). Cocoa where the TPC value was almost equal. Since
pod extract labeled C5 had 43.96 + 1.79 mg both type of samples from C1 and C3 has low
GAE/g extract (n=14) followed by PBC (27.57 + TPC values, measurement of total flavonoid
2.89 mg GAE/g extract, n=16), C3 (27.43 + 4.67 content was not being carried out for these
mg GAE/g extract, n=5) and C1 (19.59 + 2.69 samples.
mg GAE/g extract, n=4). High TPC of cocoa
pod MIX was not significantly different from Total flavonoid content
value obtained by Vriesmann (2011). Calibration curve for rutin standard with known
concentration measured at 405nm obtained
For shells, C4 (41.35 + 2.23 mg GAE/g extract, equation (2) with R2= 0.994 and used to predict
n=6) showed significantly highest total phenolic the total flavonoids (mg RE/g) of the samples.
content within the groups, followed by C5 (37.90 A405 = 0.0670 + 0.00776 concentration
+ 2.40 mg GAE/extract, n=12), PBC (35.91 + (2)
1.73 mg GAE/g extract, n=12), MIX (32.59 + Where A405 is absorbance of extract at 405
0.28 mg GAE/g extract, n=6), C1 (18.63 + 0.67 wavelength and concentration is in part per
mg GAE/g extract, n=12) and C3 (16.30 + 1.04 million (ppm) of total flavonoids content. Figure
mg GAE/g extract, n=12). TPC value of cocoa 2 summarized the TFC of cocoa pods and shells.
Malaysian Cocoa Journal, 8/2014

Figure 2: Total Flavonoid Content

Total flavonoid content vs clones
35

30
SHELL POD
25

mg RE/g extract
20

15

10

0
C4 C5 MIX PBC

The results showed that pod flavonoid content

was significantly lower than in the shell for C4, DPPH scavenging effect & EC50
C5 and PBC. However, pod labeled MIX (22.42 Determination scavenging effect was carried out
+ 0.99, n=6) contained significantly highest to potential extract of cocoa pods of C4 and MIX
amount of total flavonoids, followed by C4 as well as their shell extract. Scavenging effect
(14.18 + 2.93, n=8), C5 (4.52 + 0.91, n=9) and of cocoa pods and shells in comparison with
PBC (3.96 + 0.84, n=9). For shells, C4 (27.44 + gallic acid was summarized in Figure 3. At low
2.23, n=5) had the highest amount of total concentration (less than 125ppm), cocoa pods
flavonoid followed by PBC (17.47 + 3.28, n=6), extract exhibited higher scavenging effect from
MIX (10.54 + 0.92, n=6) and C5 (7.15 +1.13, shells, especially MIX, although lower than
n=5). Statistically, between pods and shells, the gallic acid, significantly. At 250ppm and
total flavonoid was different, within clones. 500ppm, both cocoa shells extract has higher
From this result, it was shown that cocoa pods scavenging effect on DPPH, but were not
and shells of C5 and PBC has low value of TFC, significantly different from the cocoa pod extract
therefore these samples were not tested for their labeled MIX, which however significant at
antioxidant activity as extract with high phenolic 1000ppm. Cocoa pod extract of C4 has lowest
compounds contribute to antioxidant activity scavenging effect at these concentrations.
(Sies, 2004).

Figure 3: Scavenging Effect of Cocoa Waste Extract (mean value, n=3)

DPPH Scavenging (%) vs Concentration (ppm)

100.0
90.0
80.0
70.0
DPPH Scavenging %

60.0
50.0
40.0
30.0
20.0
10.0
0.0
7.8 15.5 31.1 62.3 125 250 500 1000
ppm
P/C4 7.0 18.3 26.7 73.0 58.2 67.3 72.3 77.6
P/MIX 11.4 34.2 76.5 87.1 85.0 86.8 84.5 79.6
S/C4 3.9 6.8 13.3 28.2 40.8 87.1 88.2 88.0
S/MIX 3.9 7.2 11.9 23.7 24.3 88.3 84.3 88.3
GALLIC ACID 46.7 88.8 91.7 93.1 92.9 94.5 95.0 96.1
Malaysian Cocoa Journal, 8/2014

Effective concentration was determined to FRAP value & EC1

understand the antioxidant activity of the sample FRAP value of cocoa pod extract and shells at
using DPPH assay. Table 1 shows the summary different concentration were summarized in
of the effective concentration (EC50) of the Figure 4. At low concentration of samples, high
samples. Cocoa pod MIX has the lowest EC50 FRAP values were obtained and comparable with
among the samples. Although the EC50 the value ascorbic and gallic acid as standards. High value
was two-folds from the gallic acid, it exhibited of FRAP was exhibited by the cocoa pods and
antioxidant activity. High antioxidant activity shells, significantly, from these standards was
was exhibited in cocoa pods due coloring probably due to the synergistically effect of
pigment that might exist in the extract, which antioxidant component in the extracts, yet to be
will be determined later. Coloring pigment in measured using high performance liquid
the fruit pods can be the results of bioactive chromatography (HPLC) in the future. In order
compound such as chlorophyll, carotenoids and to elaborate more on FRAP value obtained by
phenolics (Lancaster, 1997). Lowest EC50 was shells and pods extract, the EC1 was determined
exhibited by cocoa shells MIX which is almost (Table 1). Pod extract exhibited lowest EC1
20-folds of gallic acid. The antioxidant activity value in comparison with gallic acid and shell
was reduced due to the roasting and fermentation extract. It showed that 79.96 mol Fe(II) are
process that might degrade the potentially active being reduced from Fe(III) in the presence of one
compound that exhibited antioxidant activity gram of cocoa pod extract, while only 33.60 mol
using DPPH as an assay. When calculating the of Fe(II) produced with one gram of cocoa shell
exact concentration needed to scavenging the extract. These values were even higher in
DPPH radical species by the extract, it was comparison with gallic acid as a standard (13.66
indicated that high concentration of cocoa shells mol Fe(II)) as well as ascorbic acid (1.5 mol
MIX was needed compared to cocoa pods of Fe(II)).
MIX and C4.

Figure 4: FRAP Value for Cocoa Pod and Shell Extract

180

160
FRAP value
140
ascorbic acid
mM Fe (II)/g extract

120
gallic acid
100 pod
80 shell
60

40

20

0
ppm 7.86 15.56 31.13 62.5 125 250 500 1000

Table 1: Effective Concentration

DPPH FRAP
Sample
EC50 Concentration (mg/ml) at EC50 EC1(ppm) mol Fe(II)/g extract at EC1
Gallic acid 0.106 8.36 73.19 13.66
Pod/MIX 0.258 20.35 12.67 79.96
Pod/C4 0.574 45.26 n.a* n.a*
Shell/C4 1.799 141.87 n.a* n.a*
Shell/MIX 1.938 152.83 29.76 33.60
*n.a not available
Malaysian Cocoa Journal, 8/2014

CONCLUSIONS Chang, C.C., et al. (2002). Estimation of total

flavonoid content in propolis by two
Cocoa pods and shells exhibited antioxidant complementary colorimetric methods. J.
activity with promising amount of total phenolic Food Drug Anal., 10(3); 178-182.
and flavonoid contents. Cocoa pods have higher Chung, B.Y., et al. (2003). Compositional
antioxidant activity in comparison with shells. characterization of cacao (Theobroma cacao
The antioxidant activities of the extracts were L.) hull. Agric. Chem. Biotehcnol., 46(1);
contributed of several bioactive compounds that 12-16.
acts synergistically which suggested to be Fapohunda, S., Afolayan, A. (2012).
confirmed by High Performance Liquid Fermentation of cocoa beans and
Chromatography (HPLC). antimicrobial potentials of the pod husk
phytochemicals. J Phys Pharm Adv, 2(3);
ACKNOWLEDGEMENT 158-164.
Fisal, A. et al. (2005). Activated Carbons from
The author would like to thank Director General Cocoa Pod Husks Under Chemical
of Malaysian Cocoa Board (MCB) for his Activation with H3PO4: Effect of CO2 as
permission to publish this paper. Our Oxidizing Agent. Abstracts Malaysian
appreciation goes to the MCB’s staff who had Cocoa International Cocoa Conference
directly or indirectly contributed in carrying out 2005, 114.
this experiment. Hii, C.L., Chiow, F.S. (2006). Development of
Particle Board from Cocoa Shells.
REFERENCES Malaysian Cocoa Journal, 2:22-27.
Amin, I., Chew, L.Y. (2006). Antioxidant John, N.A., et al. (2012). Quantification of total
effects of extracts of cocoa shell, roselle polyphenolic content and antimicrobial
seeds and a combination of both extracts on activity of cocoa (Theobroma cacao L.) bean
the susceptibility of cooked beef to lipid shells. Pakistan Journal of Nutrition, 11(7);
oxidation. J. Food Technol., 4(1); 10-15. 574-579.
Arlorio, M., et al. (2005). Antioxidant and Katalinic, V., et al. (2006). Screening of 70
biological activity of phenolic pigments medicinal plant extracts for antioxidant
from Theobroma cacao hulls extracted with capacity and total phenols. Food Chem., 94;
supercritical CO2. Food Res Int., 38(8-9); 550–557.
1009-1014. Kaur, C., Kapoor, H.C. (2002). Anti-oxidant
Arnous, A., et al. (2002). Correlation of pigment activity and total phenolic content of some
and flavonoid content with antioxidant Asian vegetables. Int. J. Food Sci. Technol.,
properties in selected aged regional wines 37; 153-161.
from Greek. J. Food Comp. Anal., 15; 655- Kim, J.H., et al. (2009). Compounds with
665. elastase inhibition and free radical
Arsel, O. (1992). Study of the phenolic acids of scavenging activities from Callistemon
cocoa stems, fruit stalks and beans. In lanceolatus. J Med Plants Res, Vol.3(11);
Study of the phenolic acids of cocoa stems. 914-920.
Record Number: 19936796013. pp.30. Kim, Y.H., et al. (2008). Anti-wrinkle activity of
Azizah, A.H., et al. (1999). Extraction and Ziyuglycoside I isolated from a
characterization of antioxidant from cocoa Saonguisorba officinalis root extract and its
by-products. Food Chem., 64; 199-202. application as a cosmeceutical ingredient.
Benzie, I.F.F., Strain, J.J. (1996). The ferric Biosci. Biotechnol. Biochem., 72(2); 303-
reducing ability of plasma (FRAP) as a 311.
measure of “antioxidant power”: The FRAP Lancaster, J.E, et al. (1997). Influence of
assay. Anal. Biochem., 239; 70-76. pigment composition on skin color in a wide
Bewley, J.D., et al. (2006). The encyclopedia of range of fruit and vegetable. J. Amer. Soc.
seeds: science, technology & uses. CAB Hort. Sci., 122(4); 594-598.
International. Lee, K. W., et al. (2003). Cocoa has more
Bub, A., et al. (2000). Moderate Intervention phenolic phytochemicals and a higher
with Carotenoid-Rich Vegetable Products antioxidant capacity than teas and red wine.
Reduces Lipid Peroxidation in Men. The J. Agric. Food Chem., 51; 7292-7295.
Journal of Nutrition, 2200-2206. Maria, A.M., et al. (1994). Cocoa Hull: A
Catherine, A.R., Lester, P. (2003). Flavonoids in Potential Source of Dietary Fibre. J. Sci.
Health and Disease. U.S.A: Marcel Dekker. Food Agric., 66:307-311.
Malaysian Cocoa Journal, 8/2014

Marxen, K., et al. (2007). Determination of hot-water-soluble pectins. Ind. Crops &
DPPH radical Oxidation caused by Prod., 34; 1173-1181.
methanolic extracts of some microalgae
species by linear regression analysis of
spectrophotometric measurements. Sensors,
7; 2080-2095.
Meunier, N., et al. (2003). Cocoa shells for
heavy metal removal from acidic solutions.
Bioresource Technol., 90:255-263.
Mollea, C., et al. (2008). Extraction and
characterization of pectins from cocoa
husks: A preliminary study. Food Chem.,
107; 1353-1356.
Mustafa, O., et al. (2006). Modified 2,2-Azino-
bis-3-ethylbenzothiazoline-6-sulfonic acid
(ABTS) method to measure antioxidant
capacity of selected small fruits and
comparison to ferric reducing antioxidant
power (FRAP) and 2,2’-diphenyl-1-
picrylhydrazyl (DPPH) method. J. Agric.
Food Chem., 54; 1151-1157.
Patil, Y., et al. (2010). Evaluation of In Vitro
Antioxidant Activity of Herbage of
Aromatic Plants. J. Cell Tissue Res., Vol.
10(1); 2125-2129.
Pulido, R., et al. (2000). Antioxidant activity of
dietary polyphenols as determined by
modified ferric reducing/antioxidant power
assay. J. Agric. Food Chem., 48; 3396-
3402.
Ramos, S., et al. (2008). Hypolipidemic effect in
cholesterol-fed rats of a soluble fiber-rich
product obtained from cocoa husks. J.
Agric. Food Chem., 56; 6985–6993.
Recumberri, E., et al. (2007). Dietary fibre
composition, antioxidant capacity and
physic-chemical properties of fibre-rich
product from cocoa (Theobroma cacao, L.).
Food Chem., 104; 948-954.
Samuel, Y.K.C. (2006). Crude Gum from Cocoa
of Malaysian Origin: Part 1: Rheological
Properties. Malaysian Cocoa Journal, 2:28-
31.
Sanbogi, C., et al. (1998). Antioxidative
polyphenols isolated from Theobroma
cacao. J. Agric. Food Chem., 46; 454-457.
Sies, H., Stahl, W. (2004). Nutritional protection
against skin damage from sunlight. Annu.
Rev. Nutr., 24; 173-200.
Szollosi, R., Varga, I.S. (2002). Total
antioxidant power in some species of
Labiatae (Adaption of FRAP method). Acta
Biologica Szegediensis, 46(3-4); 125-127.
In Proceedings of the 7th Hungarian
Congress on Plant Physiology, S2-P24.
Vriesmann, L.C., et al. (2011). Cacao pod husks
(Theobroma cacao L.): composition and