Accepted Manuscript

Cocoa (Theobroma cacao L.) pod husk: renewable source of bioactive compounds

Rocio Campos-Vega, Karen H. Nieto-Figueroa, B. Dave Oomah

PII: S0924-2244(17)30751-3
DOI: 10.1016/j.tifs.2018.09.022
Reference: TIFS 2329

To appear in: Trends in Food Science & Technology

Received Date: 27 November 2017

Revised Date: 20 September 2018
Accepted Date: 21 September 2018

Please cite this article as: Campos-Vega, R., Nieto-Figueroa, K.H., Oomah, B.D., Cocoa (Theobroma
cacao L.) pod husk: renewable source of bioactive compounds, Trends in Food Science & Technology
(2018), doi: https://doi.org/10.1016/j.tifs.2018.09.022.

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Abstract

Background

Cocoa Pod Husk (CPH) is the main by-product from the coca industry constituting 67-76 %

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of the cocoa fruit weight. This waste represents an important, and challenging, economic,

environmental renewable opportunity, since ten tons of wet CPH are generated for each ton

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of dry cocoa beans.

Scope and Approach

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This review highlights the value that can be added to this industrial co-product to generate

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new pharmaceutical, medical, nutraceuticals or functional food products.
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Key Findings and Conclusions

The quality and functionality of cocoa pod husk (CPH) has being improving through
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processing (fermentation, enzymatic hydrolysis, and combustion, among others), guiding to

their use as source of volatile fragrance compounds, lipase extraction, skin whitening, skin
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hydration and sun screening, ruminants’ food, vegetable gum, organic potash, antibacterial
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and nanoparticles synthesis with antioxidant and larvicidal activities. However, their

exploration to produce high-value-added products, specially for the food industry, is limited
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as well as their potential health benefits. Cocoa pod husk, the main by-product from cacao
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industry (up to 76%), is an abundant, inexpensive, and renewable source of bioactive

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compounds like dietary fiber, pectin, antioxidant compounds, minerals and theobromine,

justifying their valorization. This review highlights the value addition that can be achieved

with this valuable industrial co-product to generate new pharmaceutical, medical,

nutraceuticals or functional food products.

Keywords: Cocoa by-product; Antioxidants; Dietary fiber; Pectin.

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Cocoa (Theobroma cacao L.) pod husk: renewable source of bioactive compounds

Rocio Campos-Vegaa*, Karen H. Nieto-Figueroaa and B. Dave Oomahb

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a
Programa de Posgrado en Alimentos del Centro de la República (PROPAC), Research and
Graduate Studies in Food Science, School of Chemistry, Universidad Autónoma de Querétaro,
76010 Santiago de Querétaro, Qro, Mexico.

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b
Retired, formerly with the National Bioproducts and Bioprocesses Program, Pacific Agri-Food
Research Centre, Agriculture and Agri-Food Canada, Summerland, BC V0H 1Z0, Canada.

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Rocio Campos-Vega: chio_cve@yahoo.com.mx

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B. Dave Oomah: oomahd@gmail.com
Karen H. Nieto-Figueroa: karen.nf@outlook.com
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* Corresponding author: Tel.: (55) 1921200 Ext 5590.
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1 Abbreviations

2 50W Pectin obtained by aqueous extraction in a water bath at 50 °C for 90 min

3 5-ASA 5-aminosalicylic acid

4 ABTS 2,2′-azinobis (3-ethylbenzothiazoline-6-sulfonic acid)

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5 ADF Acid detergent fiber

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6 BHT Butylated hydroxytoluene

7 CHPFR Cocoa hull product from fermented and roasted beans

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8 CKPFR Cocoa kernel product from fermented and roasted beans

9 CLEA Cross-linked enzyme aggregate

10 CPE Cocoa pod extract

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11 CPH Cocoa pod husk

12 CPHE-AgNPs Cocoa pod husk extract synthesized with silver nanoparticles

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13 DA Degree of acetylation
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14 DE Degree of esterification
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15 DPPH 2,2-diphenyl-1-picrylhydrazyl

16 EE Epicatechin equivalent
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17 FRAP Ferric reducing antioxidant power

18 GAE Gallic acid equivalent

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19 HM High methoxy pectin

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20 IC Inhibitory concentration

21 IL Interleukin

22 LDL Low-density lipoprotein

23 LM Low methoxy pectin

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24 MCC Microcrystalline cellulose

25 MIC Minimum inhibitory activity

26 NA-HYP Nitric acid extracted (pH 3.5, 100 °C, 30 min) high performance pectin

27 NDF Neutral detergent fiber

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28 NO Nitric oxide

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29 NSP Non-starch polysaccharides

30 OP Optimized pectin extracted with nitric acid (pH 1.5, 100 °C, 30 min)

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31 ORAC Oxygen radical absorbance capacity

32 Pas Proanthocyanidins

33 PDMS Polydimetylsiloxane
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34 RE Rutin equivalent

35 ROS Reactive oxygen species

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36 SPME Solid-phase microextraction

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37 TcANR Anthocyanidin reductase

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38 TcANS Anthocyanidin synthase

39 TcLAR Leucoanthocyanidin reductase

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40 TDF Total dietary fiber

41 TE Trolox equivalent
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42 TEAC Trolox equivalent antioxidant capacity

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43 TNF-α Tumor necrosis factor-α

44 TSH 565 Trinidad selection hybrid 565

45 WAP Weeks after pollination

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47 1. Introduction

48 Theobroma cacao L. is claimed to be the only commercially cultivated and most prominent in

49 the market among the 22 species of the Theobroma genus (World Agriculture, 2011). The

50 Theobroma cacao tree probably originated from divergent areas in Central and South America;

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51 the Upper Amazon region (10,000 – 15,000 years ago), the upper Orinoco region of north east

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52 Colombia and North West Venezuela, the Andean foothills of North West Colombia, Central

53 America from southern Mexico (Chiapas-Usumacinta) to Guatemala (Young, 1994). However,

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54 Central and South American countries account for only about 14% of the current (2016, latest

55 available data) world cocoa production compared to those from African countries (⅔ of world

56
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production) (FAO, 2018). Three countries, Cote D’Ivoire, Ghana and Indonesia together
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57 cultivate and produce 61 and 67%, respectively of globally traded cocoa, whereas the top ten

58 countries account for ~93% of total world cocoa production (Table 1). Global cocoa production
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59 is estimated at 4.59 million tonnes for 2017/2018 (ICCO, 2018). In 2016, the annual production
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60 of cocoa, in decreasing order, by the eight largest cocoa producing countries were Côte D’Ivoire,
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61 Ghana, Indonesia, Nigeria, Ecuador, Cameroon, Brazil and Malaysia. These countries together

62 produced about 4.23 million tonnes, representing ~ 95% of the world production (ICCO, 2015).
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63 The cocoa bean constitutes one third (33%) of the fruit weight, leaving behind 67% of the fruit as

64 CPH as a waste by-product (Oddoye, Agyente-Badu & Gyedu-Akoto, 2013) (Fig.1). In other
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65 words, ten tons of wet CPH are generated for each ton of dry cocoa beans, thereby representing a
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66 serious disposal problem and an underexploited resource (Vriesmann, Amboni & Petkowicz,

67 2011). Pods are fully developed (100-350 mm long, 0.2-1 kg wet weight) from pollinated

68 flowers after 5-6 months. Three of the major cocoa diseases ((black pod, pod rot and cocoa pod

69 borer), also known as the disease trilogy (Evans, 2007) affect the pod specifically resulting in

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70 significant crop loss (Fowler & Coutel, 2017). Several initiatives have therefore been undertaken

71 to counter the severe crop loss, for example, development of varieties with thicker cuticle that

72 are more resistant to the common black pod rot and/or other pathogens (Fowler & Coutel, 2017).

73 Pod size, type and index (> 60 pods/tree, low disease incidence) are discriminating

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74 morphological characteristics among cocoa genotypes and therefore variation on pod traits

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75 (mainly pod size; length, width, thickness) may be associated with different morpho-geographic

76 groups (Ballesteros, Logos & Ferney, 2015).

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77 The pod has been described as a natural laminated material consisting of three distinctly different

78 layers: epicarp, mesocarp and endocarp (outer, middle and inner pericarp, respectively) (Fig.1).

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The endocarp is a soft whitish tissue protecting the delicate cocoa beans in a well-lubricated
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80 inner chamber; the mesocarp displays a hard-composite structure able to hold the cocoa beans in

81 place even under high impact; and the outermost relatively soft layer is the yellow cover (when
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82 ripe) that is directly exposed to sunshine, after which it turns black indicating rot due to
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83 degradation (Babatope, 2005). These three distinct layers have been analyzed for their chemical
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84 composition and compared to the whole CPH when incorporated as feed component in broiler

85 chick diets (Sobamiwa & Longe, 1994). High proportion of ash (47% CPH), hemicellulose
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86 (50%) and minerals (K, Ca and P) (41-66%) predominated in the epicarp; fiber (crude, NDF and

87 ADF-44-48%) and cellulose (53 %) in the mesocarp; protein (50%), crude fat (50%) and pectin
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88 (59%) in the endocarp (Table 2). The epicarp was the most limiting portion of CPH in the
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89 feeding trial, presumably due to the antagonistic inhibitory effect of lignin and pectin on CPH

90 utilization in broiler diets (Sobamiwa & Longe, 1994).

91 Cocoa pod vary in color (from green [Forastero] to red [Criollo] or variable [Trinitario, the

92 Forastero x Criollo hybrid]) and thickness when ripe depending on their clone. Pod color is a

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93 reflection of the exocarp (the outer 1-3 mm layer fruit tissue of pods harvested 18 weeks after

94 pollination [18WAP]). This exocarp accumulates high levels of soluble and insoluble

95 proanthocyanidins [PAs] (170 and 8 mg/g dw, respectively) compared with flowers, leaves and

96 seeds due to highly expressed PAs synthesis genes [anthocyanidin synthase (TcANS),

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97 anthocyanidin reductase (TcANR) and leucoanthocyanidin reductase (TcLAR)]. Furthermore,

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98 epicatechin and catechin contents in the exocarp (18WAP) were: ~30 and 0.5 mg/g dry weight

99 (dw), respectively (Liu et al., 2013); such information is unavailable for the pericarp.

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100

101 2. Composition

102
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The CPH constitutes 67-76% of the whole fruit by weight. It has been extensively investigated as
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103 poultry and/or livestock feed because of its protein (5.9-9.1%), fiber (22.6-35.7%), crude fat

104 (1.2-10%) and mineral contents, among others (Oddoye, Agyente-Badu & Gyedu-Akpto, 2013).
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105 Fresh manually chopped CPH (~ 1 cm thick) had the following percent composition: organic
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106 matter 87, which includes crude protein, fiber, fat and nitrogen free extract (8.4, 55.7, 2.5 and
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107 20.6%, respectively). The fiber fraction consisted of neutral and acid detergent fibers (80.7 and

108 74.6%), hemicellulose (6.0), cellulose (35.3) and lignin (38.8) with up to 40% in vitro
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109 digestibility (dry and organic matter) (Laconi & Jayanegara, 2015). However, total carbohydrate,

110 total dietary fiber and lignin contents of CPH vary widely (Table 3). Moreover, chemical
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111 composition depends upon the pretreatment used to process CPH, described in the next sections.
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113 2.1. Drying methods

114 The moisture content of fresh CPH is about 90% (Vriesmann, Amboni & Petkowicz, 2011),

115 therefore quick drying is essential to prevent deterioration; this is achieved by slicing the fresh

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116 pods, drying to ~65% moisture, grinding into pellets and drying the pellets to 10% moisture prior

117 to storage (Oddoye, Agyente-Badu & Gyedu-Akpto, 2013). However, the type and conditions of

118 drying can affect CPH composition. For example, dry milled (< 1 mm) CPH has low lipids

119 (1.5% db), high ash (6.7%), protein (8.6%), total dietary fiber (36.6%) composed primarily of

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120 insoluble fiber (74%) and soluble phenolics (4.6%) with carbohydrates and Klason lignin

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121 accounting for 32.3% and 21% of CPH, respectively (Vriesmann, Amboni & Petkowicz, 2011).

122 CPH dried in a circulated air oven (80 °C, 1 day) and ground to a fine powder (22 µm) contained

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123 cellulose (26.4%), lignin (24.2%), hemicellulose (8.7%), carbohydrate (17.5%), ash (9%), crude

124 protein (2.1%), fat (1.5%) and moisture (10.5%) (Chun, Husseinsyah & Yeng, 2016). Oven dried

125
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(40-45 °C) milled (< 1.68 mm) CPH contained ~ 60% total dietary fiber, predominantly non-
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126 starch polysaccharides (NSP) (42%), soluble sugars (13%), proteins (N x 6.25 = 9%), ash (8%),

127 fat (2%), phenolics (7% dw) and minerals (0.5-6%, mainly K, P, Ca and Mg); the reducing
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128 sugars consisted mainly of fructose with some glucose (~ 64% of total soluble sugar). The ratio
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129 of insoluble to soluble dietary fiber (4.2) in addition to high water retention and swelling
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130 capacities (6.5 g/g and 7.3 mL/g) suggests that CPH may be suitable for improving bowel transit

131 time (Yapo et al., 2013). In contrast, air dried, ground (0.40-0.45 mm) CPH had high cellulose
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132 (35.4% db), hemicellulose (37%), ash (12.3%), moisture (14%) and low lignin (14.7%) contents.

133 The high (74%) hollocellulose [cellulose + hemicellulose] of air dried CPH indicates its
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134 potentially excellent quality performance for the pulp and paper industry (Daud et al., 2013).
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135 Hemicellullose and lignin contents increased (27 and 14%, respectively), whereas cellulose

136 content decreased (10.7%) when boiled CPH was compared with directly dehydrated (48 °C, 48

137 h) and milled CPH (< 1 mm). For boiling, pods were washed, chopped into ~2.5 cm pieces,

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138 completely covered with water and heated (98 °C, 10 min), drained and dehydrated (48 °C, 72 h)

139 (Pérez et al., 2015).

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141 2.2. Animal feed quality

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142 Several studies focused on improving CPH feed quality by reducing its high fiber and increasing

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143 protein contents. In this context, fresh anaerobically treated CPH (7 days, 3% molasses with

144 Phanerochaete chrysoporium inoculum that produces lignin degrading enzymes) significantly

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145 increased protein content (19%, p < 0.05) concomitantly reducing crude fiber and lignin contents

146 (18%, p < 0.01). Ammoniated cocoa pod with 1.5% urea increased hemicellulose content the

147
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most (2.2x) and reduced crude fiber content (8%), presumably by reducing its physical strength,
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148 disrupting the silicified cuticular barrier and cleavages of some lignin-carbohydrate bonds. Cow

149 rumen exposure (3% w/w, anaerobic conditions for 7 days) reduced crude fiber, crude fat and
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150 lignin contents (27.5, 92 and 12.6%, respectively) of fresh CPH (Laconi & Jayanegara, 2015).
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151 Ground (< 250 µm) CPH treated with 0.1 M NaOH (solid to liquid ratio 50 g/L, room
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152 temperature, 1 h) incurred 26.6% weight loss, primarily due to lignin removal and changed its

153 structure from compact and smooth to more porous and rough surface increasing its adsorption
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154 capacity (Pua et al., 2013). Such alkalization generally encourages dairy feed intake, increases

155 buffering capacity and reduces the initial acid load of the diet thereby enabling more feed in the
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156 ration without triggering acidosis. CPH crude protein and total soluble carbohydrates increased
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157 (36%) with concomitant reduction in crude fiber, hemicellulose, cellulose, lignin and total

158 tannins (17, 21, 26, 17 and 88%, respectively) on solid-state fermentation (Pleurotus ostreatus

159 with 0.075% MnCl2 [w/w], 5 weeks) (Alemawor et al., 2009a). Solid state fermentation of CPH

160 with Rhizopus stolonifera LAU 07 increased protein (95%) and reduced fiber (7.2%) contents

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161 (Lateef et al., 2008). Ten days’ fermentation with Phanerochaete chrysosporium significantly (p

162 < 0.05) increased crude protein of CPH compared to unfermented CPH enabling the highest

163 lignin degradation (39%) and efficient CPH bioconversion (Suparjo & Nelson, 2017). Other

164 fermentation such as Rhizopus oryzae (28 °C, 6 days) of CPH and rice bran (1:9 ratio mixed

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165 substrate, sterilized at 121 °C, 15 min) also increased CPH protein content (14-15%), and free

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166 amino acids (glutamic acid 34x, aspartic acid 3.7x, alanine 4.6x and valine 1.2x) (Sriherwanto et

167 al., 2016). Furthermore, enzyme cocktails (Pentopan MonoBG + Viscozyme L; Viscozyme L +

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168 Pectinex 5XL and Pentopan MonoBG + Viscozyme L + Pectinex 5XL) effectively maximized

169 sugar release (42-53%) from CPH with a corresponding reduction (7-14%) in crude fiber and

170
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NSP enhancing its use in poultry feed (Alemawor et al., 2009b). In vitro natural fermentation
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171 (washed pods, sun-dried for 14 days and milled into powder; anaerobic fermentation for 3 days

172 at room temperature) increased protein (22-116%), ash (11-31%), fat (68-75%), moisture (49-
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173 142%) and flavonoid (29-101%) contents of pod husk depending on cocoa clones (Criollo,
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174 Forastero, Trinitario). The fermentation improved CPH by reducing crude fiber (59-73%),
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175 phytate (20-81%) and tannin (41-87%) with negligible changes in carbohydrate contents of CPH

176 (Shodehinde & Abike, 2017).

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177 CPH was investigated to develop and transfer appropriate technologies for animal feed

178 production as part of a pilot scale production and commercialization of cocoa by-products in
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179 Ghana in a ten-year project (ICCO/CFC/CRIC, September 1993-July 2003). Full annual
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180 production capacity was about 5,500 tonnes of pelletized animal feed from CPH. The cost

181 benefit and financial viability analysis indicated that the production of animal feed from CPH

182 was unlikely to be economically feasible or profitable (Adomako, 2006). However, a recent

183 study showed that sun dried CPH can be used at high inclusion rate (20%) to achieve similar

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184 feed intake and live weight gains with rabbits fed maize-based diets. In addition, CPH was

185 economically beneficial compared to maize in feeding growing rabbits (Esong et al., 2015).

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187 2.3. Biomass

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188 CPH ash has been extensively investigated particularly for its potential application as partial

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189 replacement of and/or combination with fertilizers, biogas and biofuels (Dias, 2014). However,

190 the high ash content can inhibit the combustion process since oxygen cannot easily penetrate the

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191 ash to reach the burning biomass (Martínez-Ángel, Villamizar-Gallardo& Ortíz-Rodríguez,

192 2015). Potassium (2.5-4% db) is the predominant CPH mineral, often up to 7% (db) in CPH from

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cocoa grown in Ghana accounting for nearly three-quarters (~70%) of the total ash weight
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194 (Donkoh et al., 1991). Although the high potassium concentration has been exploited for soap

195 production, it can lead to potential fouling problems and consequently to formation of deposits
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196 and agglomerations when fired in boilers for combustion or in the gasification process
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197 (Martínez-Ángel, Villamizar-Gallardo& Ortíz-Rodríguez, 2015). Other important CPH minerals

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198 (Ca, 0.3-0.8%; Mg, 0.02-0.06%; P, 0.04-0.12%; S, 0.02-0.05% and Si, 0.5% db) together

199 accounts for 19.9, 21.9 and 27.4% of ash weight (Dias, 2014; Martínez-Ángel, Villamizar-
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200 Gallardo& Ortíz-Rodríguez, 2015). High proportion of ash and minerals (47 and 41-66% of the

201 total pod, respectively) predominate in the epicarp, the most lignified cocoa pod pericarp
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202 (Sobamiwa & Longe, 1994).

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203 The ten-year project (ICCO/CFC/CRIC) also investigated potash (ash) production from CPH for

204 soft soap manufacturing. The potash from CPH is high in potassium carbonate content and as

205 such makes better soap than others in the market. Potash production from CPH was deemed a

206 profitable venture based on economic analysis (Adomako, 2006).

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207

208 2.4. Proteins

209 The protein characteristics of CPH have not been adequately investigated. Reports on the protein

210 characteristics of CPH are scarce and limited probably due to its reduced growth, food utilization

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211 efficiency and/or protein efficiency ratio observed earlier in animal diets (Donkoh, Atuahene,

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212 Wilson & Adomako, 1991; Fagbenro, 1995). One hundred and forty-four proteins were

213 identified from 700 protein spots detected in CPH proteome by MALDI-TOF/TOF MS in

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214 combination with 2-DE analysis (Awang, Karim & Mitsui, 2010). Almost half of the identified

215 proteins (48%) are involved in primary and energy metabolism functioning as housekeeping

216
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proteins. These proteins are involved in general cellular processes related to glycolysis (enolase,
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217 triose-phosphate isomerase, phosphoglycerate kinase), Calvin cycle, and Tricarboxylic acid cycle

218 (malate dehydrogenase). Six proteins including leucoanthocyanidin dioxygenase and

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219 anthocyanidin reductase, cinnamyl alcohol dehydrogenase and polyphenol oxidase pertained to
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220 the phenylpropanoid pathway responsible for the production of secondary metabolites (lignins,
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221 flavonoids, and phytoalexins). Caffeic acid 3-O-methyltransferase and polyphenol oxidase

222 involved in lignin synthesis were differentially expressed in the two clones, LAF17 with smooth
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223 and waxier pod surface and the cocoa pod borer-resistant ICS39 clone with harder and thicker

224 sclerotic layer (Awang, Karim & Mitsui, 2010).

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225
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226 2.4.1 Amino acids

227 The average protein content in CPH (generally ~ 8.5%) is lower than those previously reported

228 in cocoa bean husk (15%) (Bonvehí & Coll, 1999). Data on amino acids is limited to a single

229 report (Donkoh et al., 1991) of CPH compared to an earlier (1984) study. CPH protein has

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230 higher (10-15%) levels of total and essential amino acids and above half (~ 56%) of the total

231 amino acid content of bean husk (Table 4 and 5) due to asparagine, alanine, arginine, glycine,

232 glutamine, lysine, serine, threonine and tyrosine concentrations. CPH had twice the acidic

233 (aspartic + glutamic acids) and basic (arginine + histidine + lysine) amino acids, proline and

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234 valine content of bean husk, whereas the levels of histidine, leucine and methionine were similar

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235 in both husks. CPH protein has higher Fischer ratio and lower content of aromatic amino acids

236 than those of cocoa bean husk. The lysine/arginine ratio, a determinant of the cholesterolaemic

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237 and atherogenic effects of protein, is low for CPH protein, but higher than those of cocoa bean

238 husk. A soluble cocoa fiber product (0.79 PER, contributing 28 g protein/kg diet) obtained by

239
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enzymatic treatment of cocoa bean husk reduced the negative effects of dietary-induced
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240 hypercholesterolemia in an animal model (Ramos et al., 2008).

241 The biological value (37.6%) of bean husk (Bonvehí & Coll, 1999) corresponds to the apparent
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242 digestibility of fresh and sun-dried cocoa pod husk (37.8 and 32.1%, respectively) reflected in
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243 their nitrogen retention (17 and 12.2%, respectively) in sheep rations containing 53% (dm) CPH
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244 (Wong & Hassan, 1988). In addition to protein, CPH also contains theobromine (~ 0.2%), a

245 commercially significant appetite stimulant for ruminants (Trout, Zoumas & Tarka, 1978).
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246

247 2.5. Pectin

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248 2.5.1. Extraction methods and properties

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249 Hot water extracted pectic material from CPH contains galacturonic acid (27%), galactose

250 (21%), rhamnose and arabinose, whereas ammonium oxalate (0.5%) extraction resulted in

251 depolymerized pectin (Blakemore, Dewar & Hodge, 1966). Pectin has also been extracted with

252 mild acetic acid (0.2 N, pH 2.8, 1:3 w/v) from ripe CPH with higher yield (8-11%, db) than those

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253 reported earlier (Dittmar, 1958 cited in Adomako, 1972) from sun dried CPH (5.3-7.1%). The

254 acetic acid extracted pectin had the following composition: ash (8.9-9.8% comprising Ca and K

255 as major components), protein (1.1%), galacturonic acid (62%), galactose (4.65%), rhamnose

256 (2.9%), arabinose (1.7%) and xylose (1.2%) with carbohydrate composition similar to apple

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257 pectin (Adomako, 1972).

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258 Investigation of CPH pectins intensified after a hiatus of almost four decades, particularly in a

259 series of studies by the Brazilian group (Vriesmann et al., 2011-2017). Nitric acid (pH 1.5, 100

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260 °C, 30 min) extraction from CPH produced highly esterified (56.5% degree of esterification

261 [DE]) and acetylated (17.1% degree of acetylation [DA]) pectin (9.8%, db yield) with uronic

262
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acid (66%), homogalacturonan and rhamnogalacturonans; this pectin is considered high methoxy
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263 (HM) pectin (Vriesmann, Teófilo & Petkowicz, 2011). Hot aqueous extraction (50 °C [50W] and

264 100 °C [boiling water], 90 min, 1:25, w/v) was used in a follow up study to obtain 7.5 and 12.6%
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265 pectin from milled and dried CPH (Vriesmann, Amboni & Petkowicz, 2011). The water soluble
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266 pectins were highly acetylated (19.2 and 29% DA), composed of low methoxy
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267 homogalacturonans (37 and 42.3% DE) with rhamnogalacturonan insertions exhibiting non-

268 Newtonian shear-thinning behavior (Vriesmann, Amboni & Petkowicz, 2011). CPH treatment
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269 (aqueous citric acid pH 3, 95 min, 95 °C) yielded 10.1% (db) pectin with low moisture (2.7%),

270 high carbohydrate (64%), protein (13.8%), low phenolics (9.4%), low-methoxy, highly
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271 acetylated homogalaturonan (DE, 40.3%; DA, 15.9%) containing 65% uronic acid and exerting
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272 non-Newtonian shear –thinning behavior (Vriesmann, Teófilo & Petkowicz, 2011). The pectin

273 forms gels under low pH/high sucrose content, suggesting applications as additive in acidic

274 conditions. Similar highly acetylated (DA, 17%) CPH pectin was obtained by optimized nitric-

275 acid-mediated extraction of CPH and the best gel properties were observed at 1.32% galacturonic

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276 acid equivalent concentration, 60% sucrose and pH 2.7, demonstrating its use as a gelling and

277 thickening agent (Vriesmann & Petkowicz, 2013). High yield (10.7%, db) of CPH pectin (DE,

278 41%; DA, 17.6%) was obtained with nitric acid extraction (pH 3.5, 100 °C, 30 min) under reflux

279 with high uronic acid (59.2%), significant proportions of galactose (20.3%) and rhamnose

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280 (11.6%). The large pectic molecule was purported to be primarily responsible for its gelling

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281 properties; forming gels at low pH (< 2.5) and water activity (1% w/v; 60% v/v ethylene glycol)

282 (Vriesmann & Petkowicz, 2017). Pectin extracted with hot acid (nitric acid [pH 3, 100 °C, 30

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283 min]) or citric acid [pH 3, 95 °C, 95 min]) had similar chemical composition (65% uronic acid,

284 8.2% rhamnose, 16.7% galactose, 40.3% DM and 15.9% DA) (Vriesmann & Petkowicz, 2017).

285
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Extraction conditions greatly affect yield and chemical properties of CPH pectins (Table 6).
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286 Thus, pectin yield from cocoa husks varied (3.4-7.6%) with uric acid content ranging from 31-

287 65% and degree of methylation (7-58%) depending on solvent and extraction conditions (Chan
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288 & Choo, 2013). For example, the highest pectin yield (7.6%) is obtained with hot citric acid
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289 (1.25, w/v; pH 2.5, 95 °C, 3h), whereas hot water under similar conditions (1.25 w/v; 95 °C, 3h)
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290 extracts pectin with the highest uronic acid (65%) content. Furthermore, pectin yield increases

291 significantly (p < 0.05) with reduction in substrate –extractant ratio (from 1:25 to 1:10 w/v)
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292 using citric acid at pH 2.5. Water and hydrochloric acid extracted low methylated (LM) pectin

293 (7.2-39.3% DM), whereas citric acid (1:25 w/v, pH 2.5, 1.5 h) produced wide degree of
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294 methylation range and high methylated (HM) pectin (58%) (Chan & Choo, 2013). Similarly,
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295 acidified (1N HNO3) water treatment (1:25 w/v, 75 °C, 90 min) differing in pH (1, 2 or 3)

296 produced CPH pectins varying in yield (3.7-8.6%), galacturonic acid content (51-75%),

297 galactose (5.2-13.9%), total neutral sugars (7.9-24.3%), calcium (48.6-155.7 µmol/g), degree of

298 methylation (37-52%), DA (3.2-9.8%), intrinsic viscosity (162-304 mL/g) and viscosity-average

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299 molecular weight (43-82 kDa) (Yapo & Koffi, 2013). Hydrochloric acid (1.58 M) extraction (62

300 °C, 4.8 h) produced CPH pectin (yield =11.7%) with high DE (58.5%) and galaturonate content

301 (49.9%) (Hutomo, Rahim & Kadir, 2016). The highest CPH pectin yield (23.3% db) was

302 obtained with hot water (1.05 g/mL) extraction in a water bath (50 °C); the pectin had low

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303 moisture (0.2%), ash (1%), high DE (26.8%) and swelling (357, 275 and 360 swelling index in

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304 0.1M HCl, phosphate buffer [pH 6.8], and distilled water, respectively) (Adi-Dako et al., 2016).

305

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306 2.5.2. Applications

307 The hot water soluble pectin had Hausner ratio of 1.17, compressibility index of 14.6% and angle

308
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of repose ~38° indicating good flow properties. It was predominantly rich in potassium (2.27%),
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309 magnesium (0.22%), phosphorous (0.096%) and sulphur (0.094%), copper (10.9% of minor

310 elements), zinc (8.3%) and nickel (3.7%). The pectin exhibited dose-dependent moderate
M

311 antimicrobial activity (minimum inhibitory activity-MIC) against Staphylococcus aureus and
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312 Escherichia coli (MIC: 0.5-1 mg/mL); Pseudomonas aeruginosa, Bacillus subtilis, and
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313 Salmonella typhi (MIC: 1-2 mg/mL); and Shigella spp (Adi-Dako et al., 2016). Freeze-dried hot

314 water extracted CPH pectin (26.8% DE) and 4% hot aqueous citric acid soluble pectin showed
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315 no toxicity on the major haematological indices, bilirubin levels and the spleen when

316 administered up to 71.4 mg/kg in Sprague Dawley rats for 90 days (Adi-Dako et al., 2018). CPH
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317 pectin can therefore be safely incorporated in pharmaceutical formulations as natural polymer
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318 excipients.

319 The high methoxy pectin obtained by nitric acid was used to prepare microparticles containing 5-

320 aminosalicylic acid (5-ASA) for drug delivery to the colon. The microparticles (1:1, 2:1, 3:1 and

321 4:1) obtained by spray drying showed better drug retention with increased pectin content

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322 (Vriesman, 2012). Saponification of this high methoxy pectin promoted distinct increase in

323 biological activities; it triggered the secretion of pro-(NO, TNF-α and IL-12) and anti- (IL-10)

324 inflammatory mediators in peritoneal microphages demonstrating its application as an immune

325 modulator (Amorim et al., 2016).

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326 CPH gums have been described as essentially acidic protein-polysaccharide containing 3-5%

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327 protein with galactose, glucose, rhamnose, arabinose, galacturonic acid, and glucuronic acid in

328 1:0.21:1.9:0.2: 4.2:1.3 molar ratios with high (35-40%) uronic acid content. Crude gum yield

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329 (9.1% of dry CPH) similar to reported polysaccharide yields (8-11%, dw) had high potassium ion

330 (9.6% dw) concentration similar to previously observed CPH ash content (9%) (Figueira, Janick

331
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& BeMiller, 1994). Alcohol extracted gum (5.25%) (sun dried samples were boiled twice with
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332 70% EtOH, then extracted twice with boiling 70% MeOH) from CPH exhibited pseudoplastic

333 behavior obeying the power law flow model with high viscosity (1053 mPas at 5%
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334 concentration) similar to karaya gum (1160 mPas at 3% concentration) (Samuel, 2006). CPH
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335 gum can potentially substitute costly karaya gum, at least partly in many formulations.
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336

337 2.6. Phenolics and antioxidants

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338 2.6.1. Effect of the extraction methods

339 Total phenolic content of CPH ranged between 46 to 57 mg gallic acid equivalent (GAE)/g dry
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340 matter (Karim et al., 2014b). However, lower total phenolic (2.1-3.7 mg GAE/g) has been
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341 reported depending on the location of cocoa growth and solvent system used in extraction. For

342 example, methanol: acetone extracted higher total phenolic of CPH from two locations (Cone

343 and Taura) in Ecuador (3.53 and 3.65 mg GAE/g) than ethanol (2.07 and 2.27 mg GAE/g) that

344 was reflected in their antioxidant activities (38 and 44 µM TE/g ABTS, 34 µM TE/g DPPH and

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345 4.6 µM TE/g FRAP) compared to ethanol (24 µM TE/g ABTS, 18 and 21 µM TE/g DPPH and 2

346 µM TE/g FRAP) (Martínez et al., 2012). Acetone-water (7:3 v/v) and ethanol (70%) extracted

347 CPH phenolics (94.92 and 49.92 mg GAE/g extract, respectively) with the acetone extract

348 exhibiting antifungal activity against Fusarium oxysporum (Mu’nisa, Pagarra & Maulana, 2018).

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349 Pods chopped into 1 cm2, sun dried and ground (1 mm) contained phenolic acid (19.6-49.5 mg

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350 GAE/g) and flavonoid (4-22.4 mg rutin equivalent [RE]/g) depending on the clone (Karim et al.,

351 2014a). Several polyphenolic compounds were identified by LC-MS/MS in CPH extract (80%

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352 aqueous ethanol, 40 °C, 30 min); phenolic acids (protocatechuic acid and its derivatives and p-

353 hydroxybenzoic acid), flavonoids (apigenin, rhamnetin, kaempferol derivatives, flavone

354
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derivatives), luteolin, apigenin and linarin (Karim et al., 2014b). Some of these phenolics
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355 quantified in fresh CPH consisted of catechin (36%), quercetin (21%), epicatechin (21%) and

356 gallic (11.3%), coumaric (6.5%) and protocatechuic (4.5% of total phenolic compounds) acids
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357 (Valadez-Carmona et al., 2017). The total phenolics, flavonoid and flavonol contents of the fresh
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358 CPH was 3.24 mg GAE/g, 0.97 mg and 0.34 mg epicatechin equivalent (EE)/g, dw respectively
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359 (Valadez-Carmona et al., 2017).

360 Aqueous ethanol (80%) extract of the pod exhibited high antioxidant activity (0.26 and 0.57 EC50
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361 for DPPH or 20.35 and 45.26 mg/ml) about four times lower than that of BHT. The extract (1 g)

362 also displays strong ferric reduction potential reducing 80 mol of Fe2+ from Fe3+ (Karim et al.,
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363 2014a). Ethanol extract (50% aqueous ethanol) from a Colombian cocoa clone (TSH 565)
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364 contained epicatechin (0.25-0.35 mg/g) and varied in antioxidant activity (FRAP 137-169 µM

365 TE/g; ABTS 116-230 µM TE/g; ORAC 252-343 µM TE/g) depending on the extraction process

366 (Sotelo, Alvis & Arrázola, 2015). Low- molecular weight soluble phenolics obtained by three

367 step sequential extraction exhibited strong antioxidant activities (85% DPPH inhibition, EC50 =

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368 25 g/g; 52 µM TE/g ABTS and 130 µM TE/g FRAP) significantly higher (p < 0.05) than those of

369 fermented and roasted cocoa hull and kernel products (Yapo et al., 2013). The antioxidant

370 capacity of fresh CPH was 30.6 and 15.1 µM TEAC/g (ABTs and DPPH assays, respectively)

371 similar to those reported earlier (38 and 33 µM TEAC/g for ABTS and DPPH, respectively,

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372 (Martínez et al., 2012).

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373

374 2.6.2. Effect of processing method

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375 Processing affects CPH polyphenolic content. For example, microwave drying (595 W, 11.5

376 min) increased total phenolics (3.4x), flavonoid (5.8x), flavonol (5.8x), phenolic compounds

377
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(catechin 2.2x, quecetin 73%, epicatechin 3.3x, gallic acid 3.8x, coumaric acid 4.9x and
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378 protocatechuic acid 5.9x) and antioxidant capacity (ABTS 2.9x and DPPH 3.9x) of CPH,

379 compared to hot air and freeze-drying (Valadez-Carmona et al., 2017). Pre-treatment to prevent
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380 enzymatic (polyphenol oxidase) oxidation such as soaking (1% citric acid, 15 min) before sun
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381 drying significantly (p < 0.05) increased total phenolic (24%) and flavonoid (13%) contents
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382 compared to unsoaked sun dried CPH (Sartini, Asri & Ismail, 2017). Furthermore, ultrasound

383 (60 Hz, 2 h) extracted significantly (p < 0.05) higher phenolic than those obtained by agitation
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384 (200 rpm, 6 h) (21-23 vs 16-19 mg GAE/g) (Sotelo, Alvis & Arrázola, 2015). Total phenolic

385 content (extracted by 50% (v/v) ethanol solution, 35 °C, 150 rpm, 2h) of dried (60 °C, 24 h) CPH
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386 decreased 42% on sterilization (autoclave 120 °C, 20 min) and increased 37% after solid-state
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387 fermentation with Rhizopus stolonifera NRRL 28169 ((30 °C, pH 5.5, 72 h), presumably due to

388 the generation of new bioactive molecules (Tiburcio, 2017). HPLC of aqueous ethanol extract of

389 CPH revealed the presence of 24 phenolic peaks with 3 major unidentified components

390 accounting for 72% of the total peak area compared to 14 peaks with 3 major peaks accounting

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391 for 81% of the total peak area (33% higher than CPH) in R. stolonifera fermented CPH.

392 Furthermore, CPH displayed higher concentrations of phenolic compounds compared to

393 fermented CPH although the latter exhibited superior antioxidant activity (DPPH and ORAC)

394 than CPH, presumably due to polyphenolic hydrolysis and/or transformation (Tiburcio, 2017).

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395 Industrial fermentation such as those used for cocoa bean (6 days in wooden boxes) from

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396 Mexican cocoa genotypes can be simulated (applied) to increase phenolic content of CPH similar

397 to those reported for cocoa bean husk (Hernández-Hernández et al., 2018). Total polyphenol and

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398 tannin levels decreased by ≥ 11 and ≥ 89%, respectively after fungal treatment (Aspergillus niger

399 or Talaromyces verruculosus 0.15% w/w, moisture 1:3 w/v, 7 days) to eliminate theobromine

400
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and reduce ochratoxin A content of CPH (Oduro-Mensah et al., 2018). Similar reduction in
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401 phenolic concentration occurred on cocoa shell after solid state fermentation with Penicillium

402 roqueforti without affecting flavonol and anthocyanin contents, indicating significant
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403 consumption of these metabolites during fermentation (Lessa et al., 2018). However, CPH
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404 antioxidant capacity generally increased after solid state fermentation by Rhizopus stolonifera
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405 LAU 07. The IC50 of the CPH substrates (fermented/unfermented – 5.5/14.7 mg/mL) increased

406 2.7 folds in DPPH-scavenging capacity (Lateef et al., 2008).

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407

408 2.7. Volatiles

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409 A recent study (Tiburcio, 2017) identified 50 volatile compounds in CPH by SPME (5/95%
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410 Carboxen/Polydimethylsiloxane, 75 µm) followed by desorption and separation on 95% PDMS,

411 5% phenyl GC column (30 m x 0.25 mm, 0.25 µm film thickness). These compounds consisted

412 of 16 alcohols, 11 hydrocarbons, 8 aldehydes, 7 ketones, 5 esters, 2 amines and isovaleric acid.

413 Solid-state fermentation of CPH with Rhizopus stolonifera NRRL 28169 (incubated in a rotatory

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414 shaker 100 rpm, 30 °C, pH 5.5, 7 days) reduced the number (from 50 to 33) of identified volatile

415 compounds. Thirteen volatile compounds [(2S,4S)-(+)-pentanediol, 1-methoxy-2-butanol, 2,3-

416 butanediol, epoxylinalol, phenylethyl alcohol, D-limonene, α-copaene, ethyl iso-allocholate,

417 methyl N-hydroxybenzenecarboximidoate, methyl salicylate, nonanal, octanal, and α-toluic

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418 aldehyde] were present in both fermented and unfermented CPH (Table 7).

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419 Ten volatiles in CPH are present in cocoa, five (2,3-butanediol, phenylethyl alcohol, nonanal, α-

420 toluic aldehyde and methyl salicylate) of which have been identified in both unfermented and

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421 fermented CPH. 2,3-Butanediol imparts sweet chocolate notes reminiscent of the natural odor of

422 cocoa butter, whereas α-toluic acid (2-phenylacetaldehyde) and methyl salicylate confer floral

423
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and nutty sensory perceptions, respectively (Aprotosoaie, Luca & Miron, 2016). Phenylethyl
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424 alcohol (2-phenylethanol) is associated with the typical “floral” volatiles of dark chocolate

425 extracts resulting from microbial actions occurring during fermentation. It is also the most odor-
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426 active compound in dried and fermented cocoa (Álvarez et al., 2016). Linalool, trans-linalool
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427 oxide and isovaleric acid were identified only in nonfermented CPH. Linalool and 2-
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428 phenylethanol are major alcohols in roasted nibs; linalool concentrations in cocoa vary with its

429 origin: West Africa (0-0.5 mg/kg), Malaysia (0-0.2 mg/kg) and Ghana (0.2-0.8 mg/kg). The
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430 linalool/benzaldehyde ratio has been proposed as a flavor index for cocoa. Isovaleric acid has

431 been identified in fermented cocoa beans as a result of sugar metabolism. Benzyl alcohol, also
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432 known as α-methylbenzyl alcohol and 1-hexanol were identified in the fermented CPH; the latter
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433 confers fruity and green notes in cocoa (Aprotosoaie, Luca & Miron, 2016).

434

435 2.8. Other compounds

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436 The methyl ester of linoleic acid (9,12-octadecadenoic acid) is one of the three major

437 components of CPH ethanol (70%) extract (Mu’nisa, Pagarra & Maulana, 2018). Linoleic acid in

438 turn is the predominant (53% of the total peak area) component of bio-oil obtained from sun-

439 dried (1-2 weeks) and ground (< 1 mm) cocoa pod husks by pyrolysis (550-600 °C). The bio-oil

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440 also contained palmitic, carboxymethoxy succinic, linolenic, hexanoic, caproic, pyruvic, azelaic

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441 and caprylic acids (13.2, 5.6, 5.3, 3.8, 3.3, 2.2, and 2.0% of total peak area, respectively) (Adjin-

442 Tetteh et al., 2018). Linoleic acid was also present in petroleum extract (Soxhlet) of sundried (10

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443 days) ground CPH powder along with other fatty acids (arachidic, palmitic, pentadecanoic,

444 stearic acids) and other compounds (Adewole et al., 2013).

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445

446 3. Uses and applications

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447 3.1. Fragrance compounds

448 CPH impregnated with nitrogen sources has been used for the synthesis of volatile fragrance
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449 compounds by a Rhizopus oligosporous strain. Solid-phase microextraction (SPME) coupled

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450 with GC-MS revealed maximum abundance of volatile fragrance components when CPH was

451 fermented for 96 h (1 x 105 inoculum spores/g, 24 °C, 4-5 mm substrate thickness and pH 6.5).
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452 The highest numbers and abundance of volatile compounds was obtained with L-phenylalanine
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453 as the nitrogen source. This demonstrates CPH potential as an inert support for secondary
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454 metabolite bioconversion by fungus strain, i.e. volatile fragrance components in a solid state

455 fermentation system (Norliza, 2006). Another study by the same group (Norliza & Rozita, 2006)

456 showed the release of three compounds, benzaldehyde, phenetyl alcohol and phenyl

457 acetaldehyde when CPH (1 mm) was mixed with L-phenylalanine (2.5, 3.0 and 3.5%, w/w) and

458 fermented with Rhizopus oligosporous for 4 days.

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459 Furthermore, many of the compound identified by Tiburcio (2017) in both fermented and

460 unfermented CPH [1-octen-3-ol, nonanal, octanal, α-terpineol, ionones, methyl salicylate, β-

461 hydroxybutyric acid methyl ester, α-methylbenzyl alcohol and rosefuran] are well-known

462 flavor/aroma compounds (Table 5).

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463

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464 3.2. Skin & hair treatment

465 The skin whitening effect of CPH aqueous ethanol (80%) extract was demonstrated based on in

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466 vitro mushroom tyrosinase assay and sun screening effect (UV-absorbance at 200-400 nm

467 wavelength) (Karim et al., 2014b). Resveratrol and fatty acids, such as linoleic acid, in CPH has

468
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been reported to have in vitro skin whitening properties without adverse effects (Ohguchi et al.,
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469 2003; Parvez et al., 2006). When tyrosinase enzyme activity is inhibited, melanin production is

470 reduced, resulting in a fairer skin (Karim et al., 2014b). The extract induced human fibroblast
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471 cell proliferation at low concentration and displayed UVB sunscreen potential superior than
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472 commercial UV-protecting agents (avenobenzone and octylmethoxycinnamate). It was also

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473 highly effective against collagenase, better than kojic acid (a skin-lightening cosmetic product),

474 although lower against elastase and tyrosinase than pine bark extract (positive control, effective
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475 in maintaining cellular function which affects the skin condition) (Karim et al., 2014b).

476 Application of gel from the CPE extract reduced in vivo skin wrinkles within 3 weeks and after 5
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477 weeks (6.4, and 12.4%, respectively); skin hydration also increased (3%) after 3 weeks’
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478 application (Karim et al., 2016).

479 Cocoa pod ash is a commercial ingredient used as a component of African Black Soap that in

480 turn is used in environmentally-free conditioning cleansing cream (Koiteh, 2013).

481

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482 3.3. Animal appetite stimulant

483 CPH stimulates appetite in ruminants due to the presence of theobromine (0.2-0.25% weight),

484 particularly at low theobromine (0.05-0.1%) inclusion in the feed. This commercially significant

485 appetite stimulation enables weight gain in a short time to bring animals to market sooner due to

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486 increased feed consumption (Trout, Zoumas & Tarka, 1978). CPH contains approximately 0.15-

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487 0.4% (w/w) theobromine limiting its inclusion to 13.7% (w/w) in animal feed due to the 300

488 mg/kg theobromine content limit imposed by the European Union (Oduro-Mensah et al., 2018).

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489

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490 3.4. Texturizing & vegetable gum
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491 The pod may be used as a texturizing agent after drying (30-60% moisture) and grinding to

492 extremely small size (0.05-0.12 inch). Juice from the pods can be separated from the
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493 parenchymatous tissues (cocoa fruit flesh representing 82% of the fruit) to form a pure

494 hydrocolloid derivative that can be used as conventional vegetable gum (Drevici & Drevici,
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495 1980). The juice imparts high viscosity, preserves aroma, odor and color of products and
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496 improves rheological properties due to its carbohydrate composition (galacturonic acid,

497 galactose, rhamnose, arabinose and xylose with galacturonic acid) (Drevici & Drevici, 1980).
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498
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499 3.5. Antibacterial

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500 Some CPH extracts are effective against two major bacteria responsible for most hospital

501 infections: Gram negative Salmonella choleraesuis (1 mg/mL MIC) and Gram positive

502 Staphylococcus epidermidis (2.5 mg/mL MIC). The extract also exhibited strong inhibitory

503 activity against Pseudomonas aeruginosa (Santos et al., 2014). Fractionation (solvent partition)

504 of CPH extracts enabled identification of bioactive molecules such as phenols, steroids, or

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505 terpenes. The extract was prepared by air drying (3 months) chopped pieces of husks (1.5-2.5 cm

506 wide), pouring distilled water over the husks and collecting the aqueous extract that was dried

507 (room temperature, 72 h) and ground (< 1.5 mm). The dry crude water extract represents 0.16%

508 of the fresh CPH (Santos et al., 2014).

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509

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510 3.6. Nanoparticles larvicidal & antioxidant

511 CPH extract synthesized with silver nanoparticles (CPHE-AgNPs) exhibited antioxidant activity

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512 with potent larvicidal activity (70-100% at 10-100 µg/ml; 43.5 µg/ml LC50) against Anopheles

513 mosquito larvae at 10-100 µg/ml concentration. The CPHE-AgNPs exhibited high stability,

514
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antioxidant activity (33-85% and 14-84% DPPH-free radical scavenging and ferric ion reducing
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515 activities at 20-100 µg/ml; 49.7 IC50) and synergistically improved (43-100%) antibacterial

516 activities of cefuroxime and ampicillin. The nanoparticles effectively in vitro inhibited multidrug
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517 resistant Klebsiella pneumonia and Escherichia coli isolates at 40 µg/ml concentration and also
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518 inhibited Streptococcus pyogenes, Staphylococcus aureus, Pseudomonas aeruginosa, Aspergillus

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519 flavus, Aspergillus fumigatus and Aspergillus niger growth in emulsion paint (Lateef et al.,

520 2016).
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521

522 3.7. Encapsulating agent

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523 CPH has also been investigated in green chemistry for the synthesis of nanoparticles against
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524 multidrug resistant clinical bacteria because of its antioxidant, antimicrobial and larvicidal

525 activities (Lateef et al., 2016). Alpha-cellulose was obtained from CPH (26% yield) with 2.74%

526 ash, high crystallinity (27%), molecular weight (63,342) and 390 degree of polymerization

527 [glucose units] (Hutomo et al., 2012). CPH pectin may also serve as a suitable encapsulating

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528 agent alone or in combination with nanolignocellulose as demonstrated for citrus pectin to

529 protect probiotics in low pH fruit juice and gastrointestinal tract (Khorasani & Shojaosadati,

530 2017).

531

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532 3.8. Sweetener-xylitol

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533 Candida boidinii XM02G, isolated from cocoa cultivation areas, has been used to produce

534 xylitol from CPH. Xylitol was obtained in concentrations of 11.34 g.L-1, corresponding to a

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535 yield (Yp/s) of 0.52 g.g-1 with a fermentation efficiency (ε) of 56.6% (Santana et al., 2018).

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536
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537 3.9. Other valuable compounds

538 Ash (average yield 7.2%) from controlled combustion of CPH contained about 75% potash as
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539 potassium carbonate with 1% impurities (iron, calcium and magnesium) that renders the organic

540 potash a premium product suitable for the pharmaceutical and food industries (Woode, 2015).
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541 The high potassium content of CPH enables its use as a catalyst for biodiesel production due to
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542 microstructural development when calcined at 700 °C for 4 h (Betiku et al., 2017). Carbonization
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543 of dried and refined CPH with activated carbon reduces free fatty acid absorption (87%) of waste

544 cooking oil. This CPH can potentially be used as a potassium carbonate catalyst in the
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545 transesterification process for biodiesel production from waste cooking oil (Rachmat, Mawarani
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546 & Risanti, 2018). Microcrystalline cellulose (MCC - 5-10 µm rod-like shape, 11.635 nm

547 diameter and 74% crystallinity) was obtained by subjecting CPH to alkali treatment, bleaching,

548 and hydrochloric acid hydrolysis. This MCC can be used as a reinforcing agent with starch (2:8

549 mass ratio) and glycerol (20%) to produce high tensile strength (0.637 MPa) bioplastics (Lubis et

550 al., 2018).

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551 CPH can be hydrolyzed (1M HCl, 4 h, 75 °C) to yield maximum glucose (30.7%, w/v) that

552 produces ethanol (17.3 %, v/v) on fermentation (26 h) with Saccharomyces cerevisiae (Samah et

553 al., 2011).

554 Lipase extraction from CPH was achieved with the highest enzyme activity using response

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555 surface methodology. Cross-linked enzyme aggregate (CLEA) was produced in the presence of

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556 ammonium sulphate (30 mM) and stabilized/immobilized with glutaraldehyde (70 mM) and

557 bovine serum albumin (0.23 mM) (Yusof et al., 2016). The immobilized lipase had superior

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558 stability in response to temperature (25-60 °C) and pH (5-10), and hydrophilic organic solvents

559 such as methanol (60%) compared to the free enzyme. The highest yield of lipase (11.43 U/ml)

560
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was obtained with 7% CPH and 50 mM sodium phosphate buffer (pH 8) (Khanahmadi et al.,
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561 2015).

562
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563 CPH can be easily converted into valuable products similar to those proposed and commercially
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564 viable for cocoa shells. In this context, the pyrolysis plant (GmbH, Germany) can industrially
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565 produce 3 products: charcoal (65-75%) as the main product of low temperature (250-300 °C)

566 torrefaction/carbonization; pyrolysis fuel (30-50%) and charcoal (25-35%) at medium

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567 temperature (300-550 °C) vaporization; syngas (60-80%) and charcoal (15-20%) at high

568 temperature (600-1000 °C) gasification (Voigt, 2017). The three products, charcoal, pyrolysis
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569 fuel and non-condensable gas account for 38, 32 and 30% of the total output volume,
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570 respectively from the pyrolysis process. Thus, charcoal (280 kg/h) and biofuel (230 L/h) are

571 produced from an infeed of CPH (1 t/h) amounting to 60,000 Euros equivalent to an annual

572 saving of 504 million Euros. Furthermore, the Bühler shell burning technology can save

573 producers up to 50% in total cocoa production costs by CPH combustion system thereby

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574 reducing their carbon footprint by up to 2800 t/a. Fig. 2 summarizes the potential applications of

575 CPH.

576

577 4. Potential health benefits

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578 Notwithstanding the foregoing, the utilization of CPH to enhance the quality and functionality of

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579 pharmaceutical and food products is limited. Moreover, no research is available on the health

580 benefits of bioactive compounds from CPH. Therefore, in this section the health benefits of these

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581 compounds from other plants are presented.

582

583 4.1. Pectins

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584 The readily available CPH can be used to recover value-added compounds such as pectins.

585 Pectins extracted from several plant by-products are widely used as gelling, thickening and
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586 stabilizing agents and have several positive effects on human health, including lowering
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587 cholesterol and serum glucose levels, reducing cancer and stimulating the immune response
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588 (Vriesmann, Amboni & Petkowicz, 2011).

589 Pectin favorably influences cholesterol levels in blood. It helps reduce blood cholesterol in a
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590 wide variety of subjects and experimental conditions as comprehensively reviewed

591 (Sriamornsak, 2001). Consumption of at least 6 g/day of pectin is necessary to have a significant
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592 effect in cholesterol reduction, lower amounts are ineffective (Ginter et al., 1979). Pectin acts as
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593 a natural prophylactic substance against poisoning with toxic cations. It is effective in removing

594 lead and mercury from the gastrointestinal tract and respiratory organs (Kohn, 1982). Pectin’s

595 high water binding capacity provides a feeling of satiety, thus reducing food consumption.

596 Experiments showed a prolongation of the gastric emptying half-time from 23 to 50 minutes of a

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597 meal fortified with pectin (Holt et al., 1979). These attributes of pectin are used in the treatment

598 of overeating-related disorders (Di Lorenzo et al., 1988).

599

600 4.2. Minerals

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601 Four minerals (K, P, Ca and Mg) detected in CPH are quantitatively important (0.5- 6.0%). The

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602 presence of these CPH minerals which are required for maintaining vital functions in living

603 human cells makes it a potential source of those elements (Yapo et al., 2013).

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604 Minerals are essential constituents of skeletal structures such as bones and teeth; play a key role

605 in the maintenance of osmotic pressure, and thus regulate the exchange of water and solutes

606
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within the animal body; serve as structural constituents of soft tissues; they are essential for the
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607 transmission of nerve impulses and muscle contraction; minerals play a vital role in the acid-base

608 equilibrium of the body, and thus regulate the pH of the blood and other body fluids; also serving
M

609 as essential components of many enzymes, vitamins, hormones, and respiratory pigments, or as
D

610 cofactors in metabolism, catalysts and enzyme activators (FAO, 2017).

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611

612 4.3. Dietary fiber

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613 The total dietary fiber in CPH (59%) includes 11 and 48% of soluble and insoluble dietary fiber,

614 respectively (Yapo et al., 2013). CPH contains higher TDF compared to other dietary fiber
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615 sources, such as citrus (35-37%), apple (51%) and banana (33-52%) by-products, but lower than
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616 those of coconut fiber (63%), pea hull (75%) and yellow passion fruit rind (74-82%) by-products

617 (Yapo et al., 2013).

618 High dietary fiber intake increases stool weight and transit time which may deter large bowel

619 disorders such as constipation, diverticulitis and large bowel cancers (Elleuch et al., 2011). Most

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620 non-absorbed carbohydrates have laxative effects, both by increasing bacterial mass or osmotic

621 effects, and by water binding of unfermented fiber. The etiology of cancer involves both

622 inherited and environmental (dietary) factors. The overall evidence for an effect of total fiber

623 intake on the risk of colorectal cancer is not considered sufficient to serve as a basis for

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624 guidelines on dietary fiber intake. However, individuals with lesser fiber intakes may have an

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625 increased risk (Mudgil & Barak, 2013).

626 Recent observational studies consistently show an inverse association between dietary fiber

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627 intake and the risk of coronary heart disease (Mudgil & Barak, 2013). Postulated mechanisms for

628 lower levels of total and low-density lipoprotein (LDL) cholesterol include alterations in

629
U
cholesterol absorption and bile acid re-absorption, and alterations in hepatic metabolism and
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630 plasma clearance of lipoproteins (Theuwissen & Mensink, 2008). In some countries the evidence

631 for the cholesterol-lowering properties of certain viscous fibers, especially β-glucans from oats,
M

632 is considered sufficient for claims on the reduction of the risk of coronary heart disease (Mudgil
D

633 & Barak, 2013).

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634 Some cohort studies inversely associate dietary fiber intake and the risk of developing type 2

635 diabetes (Chandalia, 2000). Furthermore, viscous fibers such as pectin and guar gum delay
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636 gastric emptying, whereas slowly digested starch and resistant starch increase satiety in vivo

637 (Mudgil & Barak, 2013). Presumably other, as yet unidentified substances in such foods can
C

638 explain this; perhaps it is the overall combination of the dietary fiber, nutrients and bioactive
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639 substances acting synergistically, that is critical to health. However, dietary fiber such as

640 resistant starch, non-digestible oligosaccharides and polydextrose may help to prevent and

641 alleviate bowel disorders, and decrease risk factors for coronary heart disease and type 2 diabetes

642 (Kaczmarczyk, Miller & Freund 2012).

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643

644 4.4. Antioxidants compounds

645 Polyphenols are among the other bioactive compounds found in CPH. Polyphenolic compounds

646 usually accumulate in the outer parts of plants, such as shells, skins and husks (Vriesmann,

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647 Amboni & Petkowicz, 2011). The total phenolic content of CPH is estimated at ∼7% and

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648 significantly higher (p < 0.05) than in cocoa hull product from fermented and roasted beans

649 (CHPFR) (∼2-3%) and cocoa kernel product (CKPFR) (Yapo et al., 2013). Therefore, CPH can

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650 be a potential source of antioxidant-dietary fiber-rich food, which may be used to compensate for

651 their shortage or complete lack in refined modern diets currently associated to various free

652
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radical-induced disorders (Yapo et al., 2013).
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653 Free radicals, or Reactive oxygen species (ROS), play a biochemical role in the development of
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654 cancer, multiple sclerosis, Parkinson's disease, rheumatoid and inflammatory diseases. Metabolic

655 disease and environmental contaminants may increase ROS in tissues, leading to increased
D

656 oxidative stress in cells. However; antioxidants are able to ‘scavenge’ free radicals from the
TE

657 body, reducing oxidative stress and the potential development or progression of many disease

658 states (Zumbé, 1998). Studies with procyanidins BI and B3 and (+)-catechin show that these
EP

659 polyphenols are capable of trapping hydrophilic peroxyl radicals in vitro, and their radical

660 scavenging activity increases with polyphenol concentration (Ariga & Hamano, 1990).
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661 Furthermore, antioxidant activity of polyphenols has been demonstrated in vitro to prevent lipid
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662 peroxidation, a type of ROS induced cell injury and low density lipoprotein (LDL) oxidation

663 (Zumbé, 1998).

664 The anti-genotoxic effect of catechin, epicatechin, gallocatechin and epigallocatechin have all

665 been demonstrated in vitro, protecting organisms such as S. typheriurn and E.coli against

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666 mutation following exposure to various known carcinogens. This protective, anti-genotoxic

667 effect of the polyphenols has also been demonstrated in in vitro studies of mammalian cells

668 exposed to such cancer inducers as nitrosamine, benzo[a]pyrene, aflatoxin BI, tobacco smoke

669 and smoked meat extracts. Furthermore, polyphenols prevent DNA damage in vitro and in vivo

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670 following exposure to these types of compound (Agarwal & Mukhtar, 1996; Yang et al., 1996).

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671 Bioactive compounds extracted from CPH can exert potential health benefits and may therefore

672 be an alternative source of these important components (Fig.3).

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673

674 Conclusions

675
U
There is an urgent need for practical and innovative ideas to use this low cost CPH and exploit its
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676 full potential increasing the overall sustainability of the cocoa agro-industry. Since changes

677 towards better efficiency and sustainability can also involve actions to improve the valorization
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678 of by-products and of food related waste, the large amounts of organic compounds (i.e. pectin,
D

679 antioxidants, dietary fiber and minerals) contents in the CPH justify its valorization. CPH is a
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680 good source of nutraceuticals, however their use in food industry is minimal partly due to limited

681 research. CPH potential food applications include extraction of aroma compounds, vegetable
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682 gums, or texturizing agents, among others.

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683 This by-product can be tailored through processing for diverse functionality and bioactivity.
AC

684 Furthermore, CPH should be explored as a functional ingredient generating healthy and

685 innovative functional food, cosmetic and medical products.

686

687 Conflict of interest: The authors declare no competing interests.

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688 Acknowledgements

689 Author Karen Haydeé Nieto Figueroa, was supported by a scholarship from the Consejo

690 Nacional de Ciencia y Tecnología (CONACyT-Mexico) [grant number 854976]. The funding

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691 provided by UAQ (Universidad Autónoma de Querétaro) and CONACyT-Fondos Mixtos

692 (FOMIX-QRO-279751) are appreciated.

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693

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694 REFERENCES

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907 in cacao husks (Theobroma cacao L.), determination of the antioxidant capacity. Revista

908 Colombiana de Ciencias Hortícolas, 9 (1), 124-134.

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909 Sriamornsak, P. (2001). Pectin: The role in health. Journal of Silpakorn University, 21(22), 60-

910 77.

911 Sriherwanto, C., Reksohadiwinoto, B. S., Mahsunah, A.H., Suja’i, I., Toelak, S. & Rusmiyati, M.

912 (2016). Effects of Rhizopus oryzae fermentation of cocoa byproduct on certain amino acid

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913 and theobromine content. Jurnal Bioteknologi & Biosains Indonesia, 3(2), 72-80.

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914 Suparjo, S., & Nelson, N. (2017). Determination length of fermentation of cocoa pod husk with

915 Phanerochaete chrysosporium: chemically evaluation of nutrition quality. Demo-Training,

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916 16- 22.

917 Theuwissen, E., & Mensink, R. P. (2008). Water-soluble dietary fibers and cardiovascular

918
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disease. Physiology & Behavior, 94(2), 285-292.
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919 Tiburcio, P.B. (2017). Solid-state fermentation of Theobroma cacao pod husk using Rhizopus

920 stolonifera - prospection of biomolecules. MSc. Thesis, Universiade Federal do Paraná,

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921 Curitiba, Paraná, Brazil.

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922 Trout, G.A., Zoumas, B.L. & Tarka, S.M. (1978). Method of stimulating appetite in ruminants
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923 and ruminant feed containing appetite stimulant. US Patent 4070487, issued January 24,

924 1978.
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925 Valadez-Carmona, L., Plazola-Jacinto, C.P., Hernández-Ortega, M., Hernández-Navarro, M.D.,

926 Villarreal, F., Necoechea-Mondragón, H., Ortiz-Moreno, A. & Ceballos-Reyes, G. (2017).

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927 Effects of microwaves, hot air and freeze-drying on phenolic compounds, antioxidant
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928 capacity, enzyme activity and microstructure of cocoa pod husks (Theobroma cacao L.).

929 Innovative Food Science and Engineering Technologies, 41, 378-386.

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930 Voigt, M. (2017). Conversion of cocoa shells into valuable products. Connecting markets 2014,

931 GmbH, Germany. http://docplayer.net/25502492-Conversion-of-cocoa-shells-into-

932 valuable-products-mr-matthias-voigt-ceo-gk-energy-gmbh-germany.html

933 Vriesmann, L.C., Amboni, R.D.M.C. & Petkowicz, C.L.O. (2011). Cacao pod husks (Theobroma

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934 cacao L.): composition and hot-water-soluble pectins. Industrial Crops and Products, 34

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935 (1), 1173-1181.

936 Vriesmann, L.C., Teófilo, R.F. & Petkowicz, C.L.O. (2011). Optimization of nitric-mediated

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937 extraction of pectin from cocoa pod husks (Theobroma cacao L.) using response surface

938 methodology. Carbohydrate Polymers, 84(4), 1230-1236.

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Vriesmann, L.C. (2012). Pectinas da casca dos fructose do cacao (Theobroma cacao L.):
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940 optimização da extração e caracterização. PhD Thesis, Universidade Federal do Paraná,

941 Curitiba, Paraná, Brazil.

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942 Vriesmann, L.C. & Petkowicz, C.L.O. (2013). Highly acetylated pectin from cacao pod husks
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943 (Theobroma cacao L.) forms gel. Food Hydrocolloids, 33 (1), 58-65.
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944 Vriesmann, L. C., & Petkowicz, C. L. O. (2017). Cacao pod husks as a source of low-methoxyl,

945 highly acetylated pectins able to gel in acidic media. International Journal of Biological
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946 Macromolecules, 101, 146-152.

947 Woode, M.Y. (2015). Interconnected system and method for purification and recovery of potash.
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948 US Patent 9017426 B2, issued April 28, 2015.

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949 World Agriculture (2011). Systematics, anatomy and morphology of cacao. Monday, August 22,

950 2011, www.agrotechnomarket.com

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951 Yang, C. S., Chen, L., Lee, M. J., & Landau, J. M. (1996). Effects of tea on carcinogenesis in

952 animal models and humans. Dietary Phytochemicals in Cancer Prevention and Treatment,

953 51-61, New York, Plenum Press.

954 Yapo, B. M., Besson, V., Koubala, B. B. & Koffi, K. L. (2013). Adding value to cacao pod

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955 husks as a potential antioxidant-dietary fiber source. American Journal of Food and

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956 Nutrition, 1(3), 38-46.

957 Yapo, B.M. & Koffi, K.L. (2013). Extraction and characterization of gelling and emulsifying

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958 pectin fractions from cocoa pod husk. Journal of Food and Nutrition Research, 1 (4), 46-

959 51.

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Young, A.M. (1994). The Chocolate Tree. A Natural History of Cacao. Washington, DC,
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961 Smithsonian Institution Press.

962 Yusof, F., Khanahmadi, S., Amid, A. & Mahmod, S.S. (2016). Cocoa pod husk, a new source of
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963 hydrolase enzymes for preparation of cross-linked enzyme aggregate. SpringerPlus, 5(1),
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964 57.
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965 Zumbé, A. (1998). Polyphenols in cocoa: are there health benefits? Nutrition Bulletin, 23(1), 94-

966 102.
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Table 1
Cultivation and production of cocoa from top ten countries.
Production Cultivation
Country (Tonnes) %Total (Hectares) %Total
Cote D'Ivoire 1,472,313 32.96 2,851,084 27.96
Ghana 858,720 19.23 1,683,765 16.51

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Indonesia 656,817 14.71 1,701,351 16.69
Cameroon 291,512 6.53 723,853 7.10
Nigeria 236,521 5.30 838,046 8.22

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Brazil 213,843 4.79 720,053 7.06
Ecuador 177,551 3.98 454,257 4.45
Peru 107,922 2.42 125,580 1.23

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Dominican Republic 81,246 1.82 172,940 1.70
Colombia 56,163 1.26 165,844 1.63
Adapted from FAO, 2018.

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Table 2
Structural composition of cocoa pods.
Component (%) CPH Epicarp Mesocarp Endocarp
Moisture 80.2 82.8 64.0 87.1
Ash 9.1 10.1 4.6 6.7
Protein 5.9 5.0 1.9 6.9

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Crude Fiber 22.6 17.3 29.5 15.3
NDF 61.0 62.0 80.0 41.0
ADF 50.0 45.0 70.0 34.0

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Nitrogen-free 62.2 66.8 63.7 70.0
Crude fat (ether extract) 1.2 0.8 0.3 1.1

SC
Cellulose 35.0 30.0 57.5 20.8
Hemicellulose 11.0 17.0 10.0 7.0
Lignin 14.6 15.0 12.0 13.2
Pectin 6.1 5.1 2.1 10.5
Ca 0.32 0.58

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K 3.18 4.61 1.56 2.66
P 0.15 0.16 0.06 0.09
Mg 0.22 0.39 0.10 0.15
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mg/kg
Na 3.1 9.1 6.0 7.2
Zn 40.4 64.9 23.5 30.8
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Fe 90.1 197.1 106.3 112.4

Cu 7.2 13.2 5.6 7.1
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Mn 33.6 103.2 21.3 31.9

Adapted from Sobamiwa & Longe (1994).
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Table 3
Chemical composition (%) of cocoa pod husk.
Total
Reference Moisture Protein Ash Lipid carb TDF Lignin
Vriesmann, Amboni 2011 8.5 8.6 6.7 1.5 32.3 36.6 21.4
Laconi & Jayanegara 2015 NR 8.4 NR 2.5 20.6 55.7 38.8

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Esong et al., 2015 13.0 8.0 13.0 0.6 23.0 50.0 NR
Lateef et al., 2008 NR 8.2 11.3 4.7 NR 18.3 NR
Chun et al., 2016 10.5 2.1 9.0 1.5 17.5 NR 24.24

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Martínez et al., 2012 6.6 4.2 8.4 2.3 29.0 56.0 NR
Yapo et al., 2013 8.5 8.9 7.3 2.3 NR 59.0 19.4

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Range 6.4-14.1 2.1-9.1 5.9-13.0 0.6-4.7 17.5-47.0 18.3-59.0 14.7-38.8
Variance 7.8 5.0 5.8 1.4 901 186 57.7
TDF, total dietary fiber

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Table 4
Amino acid content (g/100 g) of coca pod husk and cocoa husk.
Amino Acids CPH Bean husk
Aspartic acid 0.80 0.74 1.50
Alanine 0.44 0.35 0.80
Argininea 0.22 0.26 0.70

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Cystine 0.09 NR NR
Glycine 0.29 0.34 0.72
Glutamic acid 0.77 0.81 1.87

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Histidinea 0.21 0.18 0.27
Isoleucinea 0.24 0.29 0.48
Leucinea 0.43 0.49 0.45

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Lysinea 0.40 0.35 0.79
Methioninea 0.05 0.04 0.06
Phenylalaninea 0.37 0.30 0.45

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Proline 0.38 0.40 0.20
Serine 0.41 0.35 0.71
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Threoninea 0.30 0.34 0.70
Tryptophan 0.04 NR 0.12
Tyrosine 0.21 0.18 0.42
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Valinea 0.44 0.40 0.25

Total amino acids 6.09 5.82 10.49

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EAI (%) 43.70 45.5 39.56

BCAA 1.11 1.18 1.18
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AAA 0.58 0.48 0.87

Fischer Ratio 1.91 2.46 1.36
Lysine/Arginine 1.82 1.35 1.13
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Arg+Glu+His 1.20 1.25 2.84

Met+Cys 0.14 0.04 0.06
a
Essential amino acids
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EAI, essential amino acid index

BCAA (Val+Leu+Ile)
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AAA (Phe+Tyr)
Fischer ratio (BCAA/AAA)
NR, not reported
Data for CPH from Donkoh, Atuahene, Wilson & Adomako, 1991
Cocoa husk data adapted from Bonvehi & Coll, 1999
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Table 5

Essential and reference amino acid pattern of cocoa pod husk and cocoa husk

mg amino acid/g protein

Cocoa pod huska Cocoa huskb Referencec

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His 27.4 24.3 18 19
Ile 31.3 39.2 32 28
Leu 56.1 66.2 53 66

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Lys 52.2 47.3 53 58
Met + Cys 18.3 5.4 21 25

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Phe + Tyr 75.7 64.9 58 63
Thr 39.2 45.9 47 34
Trp 5.2 - 8 11
Val 57.4 54.1 55 35

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Total 362.8 347.3 345 339
a
Adapted from Donkoh, Atuahene, Wilson & Adomako, 1991
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b
Adapted from Bonvehi & Coll, 1999
c
FAO/WHO/Expert Consultation (1990) reference pattern for 2-5-year old child
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Table 6
Cocoa pod pectin characteristics.
CA-HYP 50W BW NA-HYP OP MOP
Yield 10.1 7.5 12.6 10.7 9.8 8.6
DE 40.3 37.0 42.6 41.0 56.6 20.8
DA 15.9 29.0 19.2 17.6 17.1 9.4

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Uronic 65.1 45.1 44.6 59.2 66.0 56.0
Carbohydrate 64.0 55.8 51.9 60.8 69.9 NR
Protein 13.8 9.6 5.5 14.7 3.6 NR

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Phenolic* 9.4 9.8 8.3 9.9 3.9 NR
Gal 16.7 21.7 25.4 20.3 16.8 19.7
Rha 8.2 21.4 14.4 11.6 10.0 18.0

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Glc 4.1 4.5 5.7 4.9 2.8 3.0
Ara 1.9 4.6 5.0 1.7 2.7 2.5
Xyl 1.4 1.9 1.9 0.5 0.7 0.8

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Man 2.6 0.9 3.1 1.8 tr 1.0
Adapted from Vriesmann (2012) for CA-HYP; Vriesmann, Amboni & Petkowicz (2011) for
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50W and BW and Vriesmann (2012) for OP and MOP; NR-not reported.
CA-HYP and NA-HYP-citric and nitric acids high yield pectin, respectively; OP and MOP -
optimized and modified optimized nitric-acid extracted pectins from cocoa pod husk pectins;
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50W and BW- water soluble pectins extracted from cocoa pod husk flour at 50 and 100 °C,
respectively; DE-degree of esterification; DA-degree of acetylation. Values expressed as %,
m/m, dry base. *Percentage of gallic acid.
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Table 7

Volatile compounds identified in cocoa pod husk (CPH), cocoa pod husk (CPHF) fermentation
in solid-state (with Rhizopus stolonifera ) and cocoa.
Compounds CPH CPHF Cocoa
ALCOHOLS

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5-Methyl-2-hexanol +
(2S,4S)-Pentanediol + +
E-Linalol pyranoxide +

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1,2-Pentanediol +
11-Methyldodecanol +
1-Hexanol + +**

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1-Isopropyl-1-butanol +
1-Methoxy-2-butanol + +
1-Nonanol +

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1-Octen-3-ol +
2,3-Butanediol + + +*, **
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2,6-Dimethylcyclohexanol +
2-Ethyl-1-hexanol +
2-Ethyl-3-pentanol +
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3,4-Dimethylpent-2-en-1-ol +
Epoxylinalol + +
Linalool + +*, **
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n-Tridecanol +
Phenylethyl alcohol (2-Phenylethanol) + + +**
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Trans-linalool oxide + +*’**

α-Hydroxytoluene +
α-Methylbenzyl alcohol (Benzyl alcohol) + +**
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α-Terpinol +
ALDEHYDES
(E,E)-2,4-Heptadienal +
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Heptanal +
Methocycitronellal +
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Nonanal + + +*
Octanal + +
Trans-2-hexenal +
Trans-2-octenal +
α-Toluic aldehyde (2-Phenyl acetaldehyde) + + +**
β-Cyclocitral +
KETONES
n-Amyl methyl ketone +
1-Phenylethanone +
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2-Dodecanone +
Pyrrole-α-methyl ketone + +
Sulcatone +
trans-3-Octen-2-one +
β-Ionone +
ESTERS

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1-Ethylpentyl acetate +
Ethyl iso-allocholate + +
Methyl N-hydroxybenzenecarboximidoate + +

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Methyl salicylate + + +**
Oxalic acid, bis(6-ethyloct-3-yl)ester +
β-Hydroxybutyric acid methyl ester +

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ϒ-Hydroxybutyric acid cyclic ester +
ORGANIC ACIDS
10,12-Tricosadiynoic acid +

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D,L-Mevalonic acid lactone +
Isovaleric acid + +**
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HYDROCARBONS
(-)-Aristolene +
1,2,4-Trimethylcyclopentane +
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13-Phenylpentacosane +
1-Heptadecene +
(+)-δ-Amorphene +
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2,6,10,15-Tetramethylptadecane +
2-Cyclopropylidene-1,7,7-trimethylbicyclo[2,2,1]heptane +
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2-Ethylhexene +
2-Methyl-n-hexacosane +
D-Limone + +
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Eicosane +
Heneicosane +
Isoledene +
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Valerena-4,7(11)-diene +
α-Copane + +
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ϒ-Cadinene +
ϒ-Muurolene +
OTHERS
Octanenitrile +
1,4-Butanediamine +
Rosefuran +
From Tiburcio, 2017, *From Alvarez et al., (2016), **From Aprotosoaie, Luca & Miron (2016)
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Endocarp1

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Mesocarp1

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Pod shell/husk2
67-76%

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Mucilage/pulp2

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8.7-9.9%*

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Bean shell/husk2

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EP 2.1-2.3%

Epicarp1 Bean1
21-23%
(30-40 beans/pod)
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Fig 1. The cocoa fruit structures1 and wastes2 (with information of Babatope, 2005; Oddoye, Agyente-
Badu & Gyedu-Akpto, 2013; Awarikabey et al., 2014; Sobowale et al., 2015; Papalexandratou, & Nielsen, 2016; *By
difference).
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Fig. 2. Potential applications of cocoa pod husk.

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Fig. 3. Potential health benefits of bio-compounds found in cocoa pod husk.

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Highlights

1. CPH is an important industrial waste from which no benefit has been taken yet.
2. The investigation is limited to produce high-value-added products from CPH.

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3. Bio compounds such as dietary fiber, pectins, antioxidants and minerals can be obtained from
CPH.
4. The extraction of this bio compounds may have applications in food industry, among others,

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because their potential health benefit.

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