Development of cocoa pod extract for utilizing in cosmetic products

Suphang Chamniankul
ssuphang@gmail.com
School of Cosmetic Science, Mae Fah Luang University
Nisakorn Saewan
saewan.n@gmail.com
School of Cosmetic Science, Mae Fah Luang University
Abstract
Cocoa husk is a by-product in chocolate production. The aim of this study was to
investigate the potential cosmetic application of cocoa husk. Cocoa husk was extracted with 6
different solvents; ethanol, 50% ethanol in water, water, citric acid solution at pH 2.5, 3.0, and
3.5. The amount of pectin and phenolic content were determined. The suitable solvent for
extraction was citric acid solution pH 3.5 which provided the highest yield of pectin (36.48 %).
Total phenolic content of cocoa husk extracts was in the range of 0.35 – 0.75 mg GAE/ml and the
highest phenolic content was found in 50 % ethanol with 0.75 mg GAE/ml. Antioxidant activity
of cocoa extract was determined by DPPH assay. DPPH radical scavenging activity was 66.06 –
90.18 % and citric acid solution at pH 2.5 showed highest DPPH radical scavenging with 90.18
%. The gel scrub was developed from extracted pectin dispersed in cocoa husk extract and cocoa
husk powder was used as natural scrub. The optimum concentration of pectin was 10 % in the
extract and cocoa husk powder was 12.5 %. The sensory test of cocoa husk gel scrub showed that
this product can be acceptable in texture, viscosity, easily to rinse off, moisture, skin smoothness
and skin brightening effect of product.

Keywords: Cocoa husk/Cocoa husk extract/Pectin/Scrub

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Introduction
Pectin is a polysaccharide consists of 200 – 1000 chains of 1,4-linked α-D-
galacturonic acid which commonly found in the plant cell walls (Seggiani, Puccini, Pierini,
Giovando & Forneris, 2009; Yeoh et al., 2008). Pectin can be divided into 2 types based on
degree of esterification (DE) of the pectin. Pectin with DE higher than 50%, called high methoxyl
pectin (HMP), will form gel with sucrose at pH lower than 3.5, while low methoxyl pectin (LMP;
DE < 50%) will form gel in the presence of calcium ion (Axelos & Thibualt, 1991; Oakenfull,
1991; Ramli & Asmawati, 2011). Pectin is widely used in food, pharmaceutical, and cosmetic
industry as a gelling, thickening, emulsifier, and stabilizer agent. According to the FAO (1969),
pectin is considered to be a safe additive that can be taken daily without limits. Nowadays, there
is no pectin production in Thailand and must be imported from abroad which is insufficient for
usability demand. Thus, the cost of pectin is quite expensive which range from 1,200 – 10,000
baht/kg, depending on source and grade of pectin.
Theobroma cacao L. (Sterculiaceae) or cocoa is an important crop of several tropical
countries. In Thailand, cocoa is cultivated in many provinces such as Chanthaburi, Chumphon,
Suratthani, Nakhonsithammarat, and Rayong. Cocoa beans contain fixed oil 30 – 50 %,
carbohydrate 15 %, protein 15 %, alkaloid, theobromine 1 – 4 %, caffeine 0.07 – 0.36 % and
other substances such as catechin, pyrazine, tyramine and tyrosine. Cocoa beans are used in the
production of cocoa products such as cocoa butter, cocoa powder, chocolate and chocolate-related
products. However, cocoa beans are only 10 % of weight of the cocoa fruit and other parts are
discarded as cocoa waste (mainly cocoa pulp and cocoa husk). Cocoa husk (approximately 52 –
76 % by weight of the cacao fruit) become a serious waste disposal problem (Donkoh, Atuahene,
Wilson & Adomako, 1991). It is interesting to increase economic value and utilization of by-
product from the cocoa industry. Accumulating evidence revealed that cocoa husk is a source of
pectin which can be used in pectin production (Ramli & Asmawati, 2011; Vriesmann, Teofilo &
de Oliveira Petkowicz, 2012).
The aims of this study were to evaluate the impact of solvents on pectin extraction from
cocoa husk and determine total phenolic content and antioxidant activity of cocoa husk extracts.
Moreover, the obtained pectin, cocoa husk powder and cocoa husk extract were used to develop
gel scrub for increasing economic value to cocoa husk waste product.
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Method
Preparation of cocoa husk powder
Residue cocoa husk product from chocolate production process was obtained from
Koh Samui, Suratthani, Thailand. Cocoa husk was washed with water to remove any impurities
and chopped into small pieces. Then, cocoa husk was immersed into sodium metabisulfite to
prevent enzymatic browning reaction. Next, the cocoa husk was dried using hot air oven at 50 °C
for 8 hours. Finally, the dried cocoa husks were ground into powder by using a grinder for use in
next experiment. The preparation process was shown in Figure 1.

Extraction of cocoa husk

The dried powdered cocoa husk was extracted by six extracting solvents; ethanol,
50% ethanol, water, citric acid solution at pH 2.5, 3.0, and 3.5. Briefly, 1 g of cocoa husk powder
was extracted with 20 ml of extracting solvent at 90 °C for 1 hour. Then, the slurry was filtered
and the residue was extracted again. The filtrate of both extraction processes was precipitated
pectin by adding ethanol. Next, pectin was separated with centrifugation at 9,000 rpm for 20 min
and dried at 40 °C for 24 hours. The supernatant was determined total phenolic content and
antioxidant activity.

Fourier Transform Infrared Spectroscopy (FTIR)

The FTIR spectra were used to obtain information on chemical structure of pectin.
FTIR was performed by using KBr diffusion method with wavelengths ranging from 400 – 4,000
cm-1.
Determination of total phenolic content
Total phenolic content was analyzed using the Folin-Ciocalteu assay (Vichit &
Saewan, 2015). Briefly, extract (20 µl) was mixed with 50 µl of deionized water, 20 µl of Folin-
Ciocalteu reagent and 125 µl of 7% sodium carbonate. The mixture was incubated for 60 minutes
at room temperature. The absorbance was measured at 750 nm using a microplate reader. The
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result was determined using gallic acid standard solution and expressed as gallic acid equivalents
(mg GAE/ml extract).

Determination of antioxidant activity

Antioxidant activity was determined by using the DPPH radical scavenging assay
(Vichit & Saewan, 2015). Reaction mixtures containing 5 µl of extract and 195 µl of 0.1 mM
DPPH solution were incubated for 30 minutes at room temperature. Absorbance was measured at
515 nm using a microplate reader. The scavenging activity was derived as follows:
DPPH scavenging activity (%) = [(Acontrol – Asample)/ Acontrol] x 100
Where, Acontrol is the absorbance of the control (without the extract) and Asample is the absorbance in
the presence of the tested sample.

Development of gel scrub product

The gel scrub was developed by using pectin as gelling agent, cocoa husk powder as
abrasive and cocoa husk extract as solvent and active ingredient. After that, the formula was
evaluated the sensory application from 20 volunteers. The questionnaire asked about gel scrub
appearance and feeling of after use product which divided into 5 score. Then, the number of
prefer volunteer (score 4 – 5) was calculated as percentage of preference by using following
equation:
Preference (%) = (Number of prefer volunteer/ Total volunteer) x 100

Statistical analysis
Statistical analysis was performed by using SPSS version 11.5 (SPSS Inc, Chicago,
IL, USA). The one-way ANOVA with Duncan test was performed to evaluate the significant
differences between mean values. The confidence limits used in this study were based on 95% (p
< 0.05).
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Results and Discussion

In chocolate production, cocoa bean was collected from cocoa fruit for producing
chocolate, while, cocoa husk was discarded. In this study, the cocoa husk was chopped into small
pieces, dried, and grinded to powder (Figure 1).

(a) (b) (c) (d)

Figure 1 Appearance of (a) cocoa fruit, (b) cocoa bean, (c) cocoa husk, and
(d) dried cocoa husk powder

Pectin extraction from cocoa husk

Cocoa husk was extracted with 6 solvents including ethanol, 50 % ethanol, water,
citric acid solution at pH 2.5, 3.0, and 3.5. After ethanol adding, pectin from each solvent was
precipitated as light yellow gel. Citric acid solution and water gave high amount of pectin, while,
ethanol cannot observed pectin precipitation (Figure 2).
After that, pectin was separated and dried at 40 °C for 24 hours. The yield of pectin
was calculated based on the weight of cocoa husk powder and varied from 1.00 – 36.48 %,
depending on the extracting solvent used as shown Table 1. The highest yield of pectin was
found in citric acid solution at pH 3.5 with 36.48 %. The result showed that citric acid solutions
were the best solvent for pectin extraction from cocoa husk. This result agreed with those
reported by Mollea, Chiampo & Conti (2007) and Vriesmann, Teofilo, & de Oliveira Petkowicz
(2012), who reported that citric solution is effective in pectin extraction in terms of yield and
physicochemical properties. Moreover, the second pectin extraction was provided higher amount
of pectin than first extraction.
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(a) (b) (c) (d) (e) (f)

Figure 2 Pectin precipitations with 6 solvents including (a) ethanol, (b) 50% ethanol, (c) water,
(d) citric acid solution at pH 2.5, (e) citric acid solution at pH 3.0 and (f) citric acid
solution at pH 3.5

Table 1 Yield of pectin

Extracting solvent 1st extraction 2nd extraction Total yield
(g) (g) (%)
Ethanol 0.002±0.00 0.008±0.002 1.00
50 % ethanol 0.032±0.023 0.014±0.011 4.57
Water 0.055±0.016 0.159±0.015 21.38
Citric acid solution pH 2.5 0.068±0.011 0.205±0.016 27.25
Citric acid solution pH 3.0 0.050±0.006 0.222±0.028 27.25
Citric acid solution pH 3.5 0.066±0.008 0.299±0.034 36.48

Fourier Transform Infrared Spectroscopy (FTIR)

FTIR spectra show the functional groups and provide structural information of the
obtained pectin from citric acid solution at pH 3.5 (Figure 3). Pectin from cocoa husk showed
peak at 1,615 and 1,743 cm-1 which indicated the free and esterified carboxyl groups,
respectively. Moreover, O-H stretching (3420 cm-1), C-H bending of CH3 (1412 cm-1) and C-O
stretching (1000 – 1300 cm-1) were found.
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Figure 3 FTIR spectrum of pectin from cocoa husk

Cocoa husk extract

After pectin separation, cocoa husk extract was obtained as shown in Figure 4.
Cocoa husk extract appeared as clear yellow solution with different color shade depending on
extracting solvent. Then, the extracts were determined for total phenolic content and antioxidant
activity.
Phenolic compounds are a secondary metabolite of plant that have benefit to human
such as antioxidant, antibacterial, antiviral, anti-inflammatory, anticancer, anti-aging,
cardiovascular diseases protection, and diabetes reduction (Cevallos-Casals & Cisneros-Zevallos,
2010; Scalbert, Johnson, & Saltmarsh, 2005; Yao et al., 2004). In this study, total phenolic
content of cocoa husk extract was determined by using the Folin-Ciocalteu assay and expressed
result as gallic acid equivalents (mg GAE/ml extract). As shown in Figure 5, the ethanol extract
of cocoa husk was the highest total phenolic content (0.75 and 0.19 mg GAE/ml for 1st and 2nd
extraction, respectively), followed by water (0.63 and 0.19 mg GAE/ml for 1st and 2nd extraction,
respectively).
Antioxidant activity was determined by using DPPH radical scavenging assay and
expressed the result as percentage of DPPH radical scavenging (Figure 6). The DPPH radical
scavenging activity is one of the most commonly used methods to determine the electron- or
hydrogen-donating ability of a sample, based on the reduction of a purple DPPH radical to
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become a stable, essentially colorless diamagnetic molecule (non-radical) (Gülçin, Huyut,

Elmastaş, & Aboul-Enein, 2010). The result showed that the highest antioxidant activity was
citric acid at pH 2.5 (90.18 and 81.50 %), followed by water (84.13 and 68.59 %). Moreover, the
antioxidant activity found high positive correlation with total phenolic content (r = 0.789).

(a) (b) (c) (d) (e) (f)

Figure 4 Appearance of cocoa husk extract (a) ethanol, (b) 50% ethanol, (c) water, (d) citric acid
solution at pH 2.5, (e) citric acid solution at pH 3.0 and (f) citric acid solution at pH 3.5

1.0 First extraction Second extraction

0.8
mg GAE/ml extract

0.6
0.4
0.2
0.0
pH 2.5 pH 3.0 pH 3.5

Ethanol 50% ethanol Water Citric acid solution

Extracting solvents
Figure 5 Total phenolic content of cocoa husk extract
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100 First extraction Second extraction

% DPPH radical scavenging
80
60
40
20
0
pH 2.5 pH 3.0 pH 3.5

Ethanol 50% ethanol Water Citric acid solution

Extracting solvents
Figure 6 Antioxidant activity of cocoa husk extract

Development of gel scrub product

The gelling properties of pectin from cocoa husk was evaluated in water and aqueous
cocoa husk extract. The result showed that pectin was better formed gel in water than cocoa husk
extract. The increasing concentration of pectin was increase viscosity of product and it could not
completely dissolve when pectin concentration was more than 12 % (Figure 7 – 8). The optimum
concentration of pectin to form suitable viscosity gel in water and cocoa husk extract was 8 and
10 %, respectively.
Gel scrub product was prepared by adding cocoa husk powder in hydrogel (8 %
pectin in water) and extract gel (10 % pectin in cocoa husk extract). The concentration of cocoa
husk powder affect viscosity of product. The optimum concentration of cocoa husk powder was
15 and 12.5 % in hydrogel and extract gel, respectively (Figure 9 – 10). These concentrations
provided good gel scrub which were good viscosity, abrasive and cleaning effect.
The best gel scrub product contained cocoa pectin (10 %) as gelling agent, cocoa
husk powder (12.5 %) as abrasive and cocoa husk extract as solubilizer and active ingredient. The
appearance of gel scrub product was yellow viscos gel. Then, this product was sensory evaluated
by 20 volunteers (Figure 11). The results showed that volunteer prefer gel scrub texture (90 %),
viscosity (80 %) due to easy to use and good holding on skin. However, odor and color of gel
scrub should adjust like a chocolate. After product using, volunteers prefer gel scrub due to its
easily to rinse off (90 %), moisture improvement (80 %), skin smoothness (80 %) and whitening
effect (90 %) of product.
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0% 2% 4% 6%

8% 10% 12%
Figure 7 Appearance of pectin in water at various concentrations

0% 2% 4% 6%

8% 10% 12%
Figure 8 Appearance of pectin in cocoa husk extract at various concentrations
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0.0 % 2.5 % 5% 7.5 %

10.0 % 12.5 % 15.0 % 17.5 %

Figure 9 Appearance of cocoa husk powder in hydrogel at various concentrations

0.0 % 2.5 % 5% 7.5 %

10.0 % 12.5 % 15.0 % 17.5 %

Figure 10 Appearance of cocoa husk powder in extract gel at various concentrations
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Overall
100
Whitening effect 80 Texture
60
Skin smoothness 40 Viscosity
20
0
Miosture Color

Rinse off Odor

Cleansing

Figure 11 Sensory test of gel scrub product

Conclusion
Cocoa husk was extracted with 6 solvents including ethanol, 50 % ethanol, water, pH
2.5, 3.0, and 3.5 citric acid solutions. The highest pectin extraction yield was pH 3.5 citric acid
solution with 36.48 %. While, the highest total phenolic content was found in 50 % ethanol in
water and the highest antioxidant activity was pH 2.5 solution. The development of gel scrub was
achieved by using cocoa pectin as gelling agent (10 %), cocoa husk powder as abrasive (12.5 %)
and cocoa husk extract as solubilizer and active ingredient. The sensory test of cocoa husk gel
scrub showed that this product can be acceptable in texture, viscosity, easily to rinse off, moisture
improvement, skin smoothness and whitening effect. However, odor and color should adjust like
a chocolate. In overall, pectin, powder and extract from cocoa husk can utilize as gelling agent in
cosmetic products.

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