Cross-linked enzyme aggregate (CLEA) is easily prepared from crude enzyme and has many advantages to the environment and it is considered as an economic method in the context of industrial biocatalysis compared to free enzyme. In this work, a highly active and stable CLEA-lipase from cocoa pod husk (CPH) which is a by-product after removal of cocoa beans, were assayed for their hydrolytic activity and characterized under the optimum condition successfully. Face centered central composite design (FCCCD) under response surface methodology (RSM) was used to get the optimal conditions of the three significant factors (concentration of ammonium sulfate, concentration of glutaraldehyde and concentration of additive) to achieve higher enzyme activity of CLEA. From 20 runs, the highest activity recorded was around 9.407U (83% recovered activity) under the condition of using 20% saturated ammonium sulfate, 60mM glutaraldehyde as cross-linker and 0.17mM bovine serum albumin as feeder. Moreover, the optimal reaction temperature and pH value in enzymatic reaction for both crude enzyme and immobilized were found to be 45°C at pH 8 and 60°C at pH 8.2, respectively. A systematic study of the stability of CLEA and crude enzyme was taken with regards to temperature (25-60°C) and pH (5-10) value and in both factors, CLEA-lipase showed more stability than free lipase. The Km value of CLEA was higher compared to free enzyme (0.55mM vs. 0.08mM). The CLEA retained more than 60% of the initial activity after six cycles of reuse compared to free enzyme. The high stability and recyclability of CLEA-lipase from CPH make it efficient for different industrial applications.
Chocolate is a popular food and its consumption has long been associated with enjoyment and pleasure. The effect of chocolate on mood too has long been recognised. Chocolate is thought to have interactions with neurotransmitters which contribute to mood modulation and appetite regulation. However, the evidence in chocolate and mood studies remains highly controversial. As more is known about the influence of chocolate on mood, the reasons for these effects appear increasingly complex and inter-related.
Cocoa-specific aroma precursors and methylpyrazines in underfermented cocoa beans obtained from fermentation induced by indigenous carboxypeptidase have been investigated. Fermentation conditions and cocoa bean components were analyzed during 0 to 3 d of fermentation. Underfermented cocoa beans were characterized as having hydrophilic peptides and free hydrophobic amino acids much higher than unfermented ones. These 2 key components of cocoa aroma precursors may be produced from the breakdown of proteins and polypeptides by endogenous carboxypeptidase during the fermentation process. The enzyme was activated during fermentation. Polypeptides of 47, 31, and 19 kDa were observed in the samples throughout the 3-d fermentation period; however, only the first 2 polypeptides were remarkably reduced during fermentation. Since the 1st day of fermentation, underfermented cocoa beans contained methylpyrazines, a dominant group of cocoa-specific aroma. This might be due to microbial activities during fermentation, observed through a decrease of pH value and an increase of temperature of cocoa beans. The concentration of tetramethylpyrazines was significantly increased during the 3 d of fermentation. This may increase the cocoa-specific flavor to the beans.
Chemical transformations of Amadori compounds are responsible for the formation of aroma volatiles at the end of the Maillard reaction cascade, which in turn contributes to unique organoleptic characteristics of chocolate. A large amount of short peptides reported in fermented cocoa suggests the existence of a much larger variety of these flavor precursors than previously suspected. An HPLC-MS-MS study was performed on dried Malaysian cocoa beans to identify novel Amadori and Heyns compounds. In total, 34 species were found, including 26 previously unknown derived from di- and tripeptides. We illustrate how the structures were elucidated via tandem MS experiments, as well as present a comparative study on their relative quantities in samples coming from 11 countries of origin. There were significant differences between them, and discrimination was possible by principal component analysis based on Amadori content alone. However, the PCA separation could be a result of various post-harvest practices exerted among said countries.
Reported here is a summary of the proceedings of the Symposium on Plant Polyphenols: Nutrition, Health and Innovations, which was cosponsored by the Southeast Asia Region branch of the International Life Sciences Institute and the Nutrition Society of Malaysia in Kuala Lumpur, Malaysia, June 22-23, 2009. The symposium provided a timely update of research regarding the protective effects of polyphenols in chronic diseases, such as cardiovascular disease and cancer, as well as the development of innovative polyphenol-containing food products with enhanced nutritive and health properties. Presentations covered polyphenols from a wide range of food sources such as tea, coffee, nuts and seeds, cocoa and chocolate, soy, and Asian fruits, vegetables, and spices. The symposium was attended by a large and diverse group of nutritionists, dietitians, researchers and allied health professionals, as well as management, research and development, and marketing personnel from the food and beverage industry. Their enthusiastic participation was a testament to the increasing awareness and interest in polyphenols in the prevention and control of chronic diseases. Presented here are some of the highlights and important information from the symposium.
Food and fuel prices have soared in recent years affecting most adversely the poor and those with fixed incomes. Since 2000, wheat price in the international market has more than tripled and maize prices have more than doubled. The price of rice, the staple of billions in Asia, has tripled in the past year. The surge of food prices has been blamed on multiple factors including higher energy and fertilizer costs, greater global demand, drought, the loss of arable land to biofuel crops and price speculation. In light of the spiraling rise in food prices, there is the prospect of increasing rates of under-nutrition worldwide. As it is, 800 million are estimated to be suffering from chronic malnourishment, with another 2.1 billion people living close to subsistence levels on less than US$2 a day. Some perspectives of the food production experience of Malaysia are shared here as a case of a country that has built up capabilities and resources through high level of foreign and domestic investment leading to a diversified economy. In response to the recent surge in the price of rice, the Malaysian government announced the setting up of a dedicated fund amounting to US$1.25 billion to increase production of food including fruits and vegetables, and targeting 100% self-sufficiency in rice, by growing rice on a massive scale in Sarawak. During the current five-year development plan for the period of 2006-2010, (Ninth Malaysia Plan), the role of the agriculture sector is considerably enhanced to be the third pillar of economic growth, after manufacturing and services. Among the measures taken, are those aimed at increasing incomes of smallholders and fishermen mainly through improving productivity. These measures include encouraging more rice farmers to participate in mini-estates and group farming, providing financial assistance to rehabilitate cocoa, pepper and sago smallholdings, enhancing the capabilities of coastal fishermen, and setting up of a special program to assist poor households in the agriculture sector to diversify their sources of income. The various socio-economic programs in Malaysia that have been put in place over the years may have cushioned to some extent so far the severity of the dramatic hikes in food prices.
Obesity and related metabolic diseases (e.g., type 2 diabetes, cardiovascular diseases, and hypertension) are the most prevailing nutrition-related issues in the world. An emerging feature of obesity is their relationship with chronic inflammation that begins in white adipose tissue and eventually becomes systemic. One potential dietary strategy to reduce glucose intolerance and inflammation is consumption of polyphenol-rich cocoa-like cocoa or their by-products. In vitro as well as in vivo data indicate that cocoa polyphenols (CPs) may exhibit antioxidant and anti-inflammatory properties. Polyphenols commonly found in cocoa have been reported to regulate lipid metabolism via inducing metabolic gene expression or activating transcription factors that regulate the expression of numerous genes, many of which play an important role in energy metabolism. Currently, several molecular targets (e.g., nuclear factor Kappa B, activated protein-1, peroxisome proliferator-activated receptors, liver X receptors, and adiponectin gene) have been identified, which may explain potential beneficial obesity-associated diseases effects of CPs. Further studies have been performed regarding the protective effects of CPs against metabolic diseases by suppressing transcription factors that antagonize lipid accumulation. Thus, polyphenols-rich cocoa products may diminish obesity-mediated metabolic diseases by multiple mechanisms, thereby attenuating chronic inflammation.
The aims of this study were to determine the antioxidant and antiproliferative activity of the following Theobroma cacao plant part methanolic extracts: leaf, bark, husk, fermented and unfermented shell, pith, root, and cherelle. Antioxidant activity was determined using 2,2-diphenyl-2-picrylhydrazyl (DPPH), thiobarbituric acid-reactive substances (TBARS), and Folin-Ciocalteu assays; the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium (MTT) assay was used to determine antiproliferative activity. The root extract had the highest antioxidant activity; its median effective dose (EC50) was 358.3±7.0 µg/mL and total phenolic content was 22.0±1.1 g GAE/100 g extract as compared to the other methanolic plant part extracts. Only the cherelle extract demonstrated 10.4%±1.1% inhibition activity in the lipid peroxidation assay. The MTT assay revealed that the leaf extract had the highest antiproliferative activity against MCF-7 cells [median inhibitory concentration (IC50)=41.4±3.3 µg/mL]. Given the overall high IC50 for the normal liver cell line WRL-68, this study indicates that T. cacao methanolic extracts have a cytotoxic effect in cancer cells, but not in normal cells. Planned future investigations will involve the purification, identification, determination of the mechanisms of action, and molecular assay of T. cacao plant extracts.
In this present study, we investigated the effects of cocoa extract containing polyphenols and methylxanthines prepared from cocoa powder on the biochemical parameters of obese-diabetic (Ob-db) rats. Obese-diabetic (Ob-db) rats were developed using a high-fat diet (49% fat, 32% carbohydrate, and 19% protein from total energy, kcal) for 3 months, followed by a low dose (35 mg/kg body weight) streptozotocin (STZ) injection. Cocoa extract (600 mg/kg body weight/day) was given to the rats for 4 weeks. The results indicated that there were no significant differences in fasting plasma glucose and insulin level after 4 weeks of cocoa extract administration. Oral glucose tolerance test revealed that cocoa supplementation in Ob-db rats significantly (p < 0.05) reduced plasma glucose at 60 and 90 min compared to unsupplemented Ob-db rats. Plasma free fatty acid and oxidative stress biomarker (8-isoprostane) were significantly (p < 0.05) reduced after cocoa supplementation. Superoxide dismutase activity was enhanced in Ob-db compared to that in nonsupplemented rats. However, no change was observed in catalase activity. The results showed that cocoa supplementation had an effect on postprandial glucose control but not for long term (4 weeks). Moreover, cocoa supplementation could reduce circulating plasma free fatty acid and 8-isoprostane and may enhance the antioxidant defense system.
Cocoa beans are rich in numbers of beneficial bioactive compounds such as phenolics and
phytosterols, which benefits to human being. The suitable extraction method is needed to
produce high quality and quantity of cocoa butter and other bioactive compounds. There are
many extraction method to extract these compounds such as Soxhlet extraction, supercritical
fluid extraction, ultrasound extraction method and others. The objective of this study is to
determine the effectiveness of the different extraction methods producing high yields of cocoa
butter, lower oxidative value, stable phytosterols and antioxidant content. The cocoa beans were
subjected to different extraction methods such as Soxhlet extraction (SE), Ultrasonic extraction
method (USE), Supercritical carbon dioxide (SCO2
) and Supercritical carbon dioxide with cosolvent
(SCO2
-Ethanol). Cocoa butter extracted using SCO2
-Ethanol has significantly (p
This study was carried out to extract and compare the characteristic ability of globulins from cottonseed, alfalfa seed, pea seed, mung bean and French bean with cocoa seeds to produce cocoa-specific aroma precursors. The extracted globulins were compared through SDS PAGE, amino acid and oligopeptide profiles. A very low recovery was obtained during globulin extraction from different seeds ranging from 0.5% to 2.7%. Cottonseed produced the highest total protein (13.90 mg/g), followed by cocoa seed (11.91 mg/g), whereas alfalfa seed, mung bean, pea seed and French bean produced 7.86, 4.77, 4.59 and 3.89 mg/g respectively. Two distinctive bands of 51.1 and 33.0 kDa were observed for cocoa vicilin-class globulin (VCG) from SDS PAGE. More than three bands were shown for other seed globulins. Comparative HPLC analyses of the obtained peptide mixtures revealed different and complex patterns of predominantly hydrophobic peptides. A similar high content of amides (glutamic acids-glutamine, aspartic acid- asparagine and arginine) and low concentrations of lysine were observed in all seeds globulin.
Colorectal cancer (CRC) is the third most common malignancy in males and the second most common cancer worldwide. Chronic colonic inflammation is a known risk factor for CRC. Cocoa contains many polyphenolic compounds that have beneficial effects in humans. The objective of this study is to explore the antioxidant properties of cocoa in the mouse model of azoxymethane (AOM)/dextran sulfate sodium (DSS)-induced colitis-associated cancer, focusing on the activation of Nrf2 signaling. Mice were treated with AOM/DSS and randomized to receive either a control diet or a 5 and 10% cocoa diet during the study period. On day 62 of the experiment, the entire colon was processed for biochemical and histopathological examination and further evaluations. Increased levels of malondialdehyde (MDA) were observed in AOM/DSS-induced mice; however, subsequent administration of cocoa decreased the MDA. Enzymatic and nonenzymatic antioxidants, such as superoxide dismutase, catalase, glutathione peroxidase and glutathione reductase, were decreased in the AOM/DSS mice. Cocoa treatment increases the activities/levels of enzymatic and nonenzymatic antioxidants. Inflammatory mediators, such as inducible nitric oxide synthase (iNOS) and cyclooxygenase (COX)-2, were elevated during AOM/DSS-induction, and treatment with 5 and 10% cocoa effectively decreases the expression of iNOS and COX-2. The NF-E2-related factor 2 and its downstream targets, such as NQO1 and UDP-GT, were increased by cocoa treatment. The results of our study suggest that cocoa may merit further clinical investigation as a chemopreventive agent that helps prevent CAC.
Fat, alkaloid and polyphenol contents of two clones of cocoa (UIT1 and PBC 140) were removed and the remaining powder was autolyzed at pH 3.5 and 5.2. Based on the results, autolysates of UIT produced at pH 3.5 exhibited the highest ability to inhibit α-amylase activity. However, no α-glucosidase inhibition activity was observed under the conditions specified. Autolysates produced under pH 3.5 caused the highest amount of insulin secretion. In streptozotocin-diabetic rats, all cocoa autolysates significantly decreased blood glucose at 4h. To assure that the results from the assays were not due to the polyphenols of cocoa autolysates qualitative and quantitative tests were applied. According to their results cocoa autolysates were found to be free from polyphenols. Analysis of amino acid composition revealed that cocoa autolysates were abundant in hydrophobic amino acids. It can be suggested that besides other compounds of cocoa, its peptides and amino acids could contribute to its health benefits.
Cocoa and cocoa products have received much attention due to their significant polyphenol contents. Cocoa and cocoa products, namely cocoa liquor, cocoa powder and chocolates (milk and dark chocolates) may present varied polyphenol contents and possess different levels of antioxidant potentials. For the past ten years, at least 28 human studies have been conducted utilizing one of these cocoa products. However, questions arise on which of these products would deliver the best polyphenol contents and antioxidant effects. Moreover, the presence of methylxanthines, peptides, and minerals could synergistically enhance or reduce antioxidant properties of cocoa and cocoa products. To a greater extent, cocoa beans from different countries of origins and the methods of preparation (primary and secondary) could also partially influence the antioxidant polyphenols of cocoa products. Hence, comprehensive studies on the aforementioned factors could provide the understanding of health-promoting activities of cocoa or cocoa products components.
Chocolate contains a wide range of antioxidants that includes soluble phenolic compounds (phenolic acids, catechin, epicatechin, and proanthocyanidins), insoluble polymeric phenolics and methylxanthines. The objective of this study was to determine phenolic and theobromine contents in dark (DC), milk (MC), and white (WC) chocolates commonly found in the Malaysian marketplace. Total phenolic and flavonoids were determined by means of a spectrometric assay, while catechin, epicatechin and theobromine were quantified using a reverse-phase HPLC method. Dark chocolates exhibited the highest phenolics and flavonoids contents, followed by milk and white chocolates. Catechin and epicatechin were major flavonoids detected in dark chocolates. Theobromine was detected in dark and milk chocolates, but not in white chocolates. A high correlation (r= 0.93) between total phenolic and flavonoid contents, indicating that the major phenolic compounds in dark chocolates belong to the flavonoid class. When nutrition and health promotion are of concern, dark chocolates would be recommended over milk and white chocolates owing to their higher contents of antioxidant phenolic compounds.
Studies indicated that cocoa-based products effectively mitigate the risks associated with metabolic syndrome (MetS), however, the effect varies based on cocoa types, dosages, and study durations. This review aimed to determine the flavanol-rich cocoa consumption on MetS outcomes within the last decade (2013-2023), adhering to Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines. Seven randomized-controlled trials (RCTs) used cocoa-based products containing 0.3-1680 mg flavanol monomers and 3.5-1270 mg procyanidins. Cocoa-based products beneficially reduced glycemic response, blood pressure and lipid profiles. However, this review highlights little evidence pinpointing the best cocoa products type and required dosage for the observed effects. Further intervention aiming to improve MetS should justify the selection and concentration of flavanols (monomers and procyanidins). A robust study design should consider registering the trials before study commencement, consider multicenter RCT trials, and adjust for potential covariates that might "masked" the outcomes.
A new patent pending process is proposed in this study to produce edible film directly from seaweed (Kappaphycus alvarezii). Seaweed together with other ingredients has been used to produce the film through casting technique. Physical and mechanical tests were performed on the edible films to examine the thickness, colour, transparency, solubility, tensile strength,
elongation at break, water permeability rate, oxygen permeability rate and surface morphology. Produced film was transparent, stretchable, sealable and have basic properties as a film for food packaging. This study suggests that the edible film could be used as novel materials in food industry as sachet/pouch/bag for instant coffee, breakfast cereals drinks, seasoning powder,
candies etc; as wrapper for seasoning cube and chocolate; as interleaf for frozen foods such as burger patties to avoid the patties from sticking together; and also as material for edible logo in bakeries products. Other than that, the edible film also could be used in pharmaceutical industry as functional strips such as oral freshener strips and drug strips. In cosmetic and toiletries industries, the edible film could be used to produce facial mask and bag for pre-portioned detergent. Compared with edible film developed earlier using alginate and carrageenan, film developed in this research used seaweed directly. The developed film reduced the need to extract the alginate and carrageenan, making material preparation easier and cheaper.
Acid sulfate soils having a pH of less than 3.5 are widespread in Malaysia. Some of these soils are planted to cocoa, but the yield is reported to be low due to soil infertility related to Al toxicity. Cocoa growth is sensitive to the presence of Al in the soil. To a certain extent, Al toxicity in soils can be reduced by organic matter application and to a greater extent in iron-poor acid sulfate soil. A study was conducted to determine the efficacy of various types of organic materials easily available in the country to ameliorate acid sulfate soil infertility for growing cocoa seedlings. The treatments were control (nil), lime (3 t/ha), peat (10% w/w), peat plus green manure (10% w/w), peat plus rice straw (10% w/w), peat plus chicken dung (10% w/w) and peat plus POMS (Palm oil mill sludge) (10% w/w). The growth of cocoa seedlings was affected significantly by the presence of Al in the cocoa tissues. As the amount of Al in the leaves increased, the relative top dry weight of cocoa seedlings decreased. Likewise, the relative plant height was negatively correlated with Al in the leaves. Peat as well as peat in combination with green manure, rice straw, chicken dung or palm oil mill sludge was able to reduce Al toxicity in acid sulfate soil; the highest top dry weight of cocoa seedlings were obtained in the peat plus green manure treatment. The best cocoa seedlings root growth was found for the peat treatment alone. The relative top dry weight of cocoa seedlings was negatively correlated with Al(3+) as well as Al(3+)+Al(OH)(2+)+Al(OH)(2)(+) activity in the soil solution. The critical values for Al(3+) and the combination of Al(3+)+Al(OH)(2+)+Al(OH)(2)(+) activity in the soil solution were 10 microM and 15 microM, respectively.
The previously identified female sex pheromone of cocoa pod borer, Conopomorpha cramerella, was re-evaluated for its attractive activity in different field conditions. It was found that lures containing 100-mug of synthetic sex pheromone blend, (E,Z,Z)- and (E,E,Z)-4,6,10-hexadecatrienyl acetates, and the corresponding alcohols in a ratio of 40:60:4:6 in a polyethylene vial attracted male C. cramerella moths in Sabah and peninsular Malaysia and in Sumatra and Sulawesi, Indonesia, suggesting that the same pheromone strain existed in a wide stretch of the Indo-Malayan archipelago. Of the three kinds of trap designs tested, the Delta traps were more effective than Pherocon V scale traps. Male captures were not significantly different among traps baited with 100-, 300-, or 1,000-mug doses of sex pheromone. A release rate study of pheromone formulation conducted in the laboratory showed that volatile active ingredients were desorbed from polyethylene vials following first-order kinetics, which indicates a satisfactory "half-life time" of a 100-mug loading is approximately 6 wk under laboratory conditions. A satisfactory attractiveness of the lure with a 100-mug loading was approximately 1-2 mo in the fields.