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  1. Muhamad H, Zainudin BH, Abu Bakar NK
    Food Chem, 2012 Oct 15;134(4):2489-96.
    PMID: 23442715 DOI: 10.1016/j.foodchem.2012.04.095
    Solid phase extraction (SPE) and dispersive solid-phase extraction (d-SPE) were compared and evaluated for the determination of λ-cyhalothrin and cypermethrin in palm oil matrices by gas chromatography with an electron capture detector (GC-ECD). Several SPE sorbents such as graphitised carbon black (GCB), primary secondary amine (PSA), C(18), silica, and florisil were tested in order to minimise fat residues. The results show that mixed sorbents using GCB and PSA obtained cleaner extracts than a single GCB and PSA sorbents. The average recoveries obtained for each pesticide ranged between 81% and 114% at five fortification levels with the relative standard deviation of less than 7% in all cases. The limits of detection for these pesticides were ranged between 0.025 and 0.05 μg/g. The proposed method was applied successfully for the residue determination of both λ-cyhalothrin and cypermethrin in crude palm oil samples obtained from local mills throughout Malaysia.
  2. Mohamed R, Zainudin BH, Yaakob AS
    Food Chem, 2020 Jan 15;303:125392.
    PMID: 31446362 DOI: 10.1016/j.foodchem.2019.125392
    In this article, an easy and quick method based on microwave assisted acid digestion technique prior to quantification using inductively coupled plasma mass spectrometry for the analysis of heavy metals in cocoa beans, cocoa powder and chocolate was established and validated for arsenic (As), cadmium (Cd), lead (Pb), and antimony (Sb). Limit of quantification for all elements were product dependent and varies from 7.84 to 194.52 µg/kg. The recoveries of the heavy metals at 250 and 1000 µg/kg spiking levels were ranged between 96.27-108.75%, 90.43-101.97% and 89.72-106.26% for cocoa beans, cocoa powder, and chocolate, respectively. Relative standard deviation values obtained were all below 20% and the expanded uncertainty measurements for the elements were less than 25%. The analysis of real samples found that the concentration level is far from the national alarming level except for cadmium in cocoa beans.
  3. Zainudin BH, Salleh S, Mohamed R, Yap KC, Muhamad H
    Food Chem, 2015 Apr 1;172:585-95.
    PMID: 25442595 DOI: 10.1016/j.foodchem.2014.09.123
    An efficient and rapid method for the analysis of pesticide residues in cocoa beans using gas and liquid chromatography-tandem mass spectrometry was developed, validated and applied to imported and domestic cocoa beans samples collected over 2 years from smallholders and Malaysian ports. The method was based on solvent extraction method and covers 26 pesticides (insecticides, fungicides, and herbicides) of different chemical classes. The recoveries for all pesticides at 10 and 50 μg/kg were in the range of 70-120% with relative standard deviations of less than 20%. Good selectivity and sensitivity were obtained with method limit of quantification of 10 μg/kg. The expanded uncertainty measurements were in the range of 4-25%. Finally, the proposed method was successfully applied for the routine analysis of pesticide residues in cocoa beans via a monitoring study where 10% of them was found positive for chlorpyrifos, ametryn and metalaxyl.
  4. Zainudin BH, Salleh S, Yaakob AS, Mohamed R
    Food Chem, 2021 Aug 06;368:130778.
    PMID: 34391100 DOI: 10.1016/j.foodchem.2021.130778
    Multiresidue quantitative and qualitative screening method for the analysis of pesticide residues in dried cocoa beans was validated and applied to imported and domestic cocoa beans samples. The quantitative method comprises of 15 pesticides while the screening method covers 110 pesticides of different chemical classes. The method was based on modified QuEChERS (Quick Easy Cheap Efficient Rugged Safe) extraction and detection using triple quadrupole (QQQ-MS) and ion mobility quadrupole time of flight mass spectrometry (IMS-QTOF). The method was quantitatively validated in terms of linearity, limit of quantification (LOQ), specificity, selectivity, accuracy, and precision. On the other hand, screening detection limits were established for 110 pesticides. Finally, the optimized strategy was successfully applied for the routine analysis of pesticide residues in 137 cocoa bean samples and 32% of the total samples were found positive for ametryn, chlorpyrifos, isoprocarb, and metalaxyl.
  5. Karim AA, Azlan A, Ismail A, Hashim P, Abd Gani SS, Zainudin BH, et al.
    BMC Complement Altern Med, 2014 Oct 07;14:381.
    PMID: 25292439 DOI: 10.1186/1472-6882-14-381
    BACKGROUND: Cocoa pod is an outer part of cocoa fruits being discarded during cocoa bean processing. Authors found out that data on its usage in literature as cosmetic materials was not recorded in vast. In this study, cocoa pod extract was investigated for its potential as a cosmetic ingredient.

    METHODS: Cocoa pod extract (CPE) composition was accomplished using UHPLC. The antioxidant capacity were measured using scavenging assay of 1,2-diphenyl-2-picrylhydrazyl (DPPH), β-carotene bleaching assay (BCB) and ferric reducing antioxidant power (FRAP). Inhibiting effect on skin degradation enzymes was carried out using elastase and collagenase assays. The skin whitening effect of CPE was determined based on mushroom tyrosinase assay and sun screening effect (UV-absorbance at 200-400 nm wavelength).

    RESULTS: LC-MS/MS data showed the presence of carboxylic acid, phenolic acid, fatty acid, flavonoids (flavonol and flavones), stilbenoids and terpenoids in CPE. Results for antioxidant activity exhibited that CPE possessed good antioxidant activity, based on the mechanism of the assays compared with ascorbic acid (AA) and standardized pine bark extract (PBE); DPPH: AA > CPE > PBE; FRAP: PBE > CPE > AA; and BCB: BHT > CPE > PBE. Cocoa pod extract showed better action against elastase and collagenase enzymes in comparison with PBE and AA. Higher inhibition towards tyrosinase enzyme was exhibited by CPE than kojic acid and AA, although lower than PBE. CPE induced proliferation when tested on human fibroblast cell at low concentration. CPE also exhibited a potential as UVB sunscreen despite its low performance as a UVA sunscreen agent.

    CONCLUSIONS: Therefore, the CPE has high potential as a cosmetic ingredient due to its anti-wrinkle, skin whitening, and sunscreen effects.

  6. Abdul Karim A, Azlan A, Ismail A, Hashim P, Abd Gani SS, Zainudin BH, et al.
    J Cosmet Dermatol, 2016 Sep;15(3):283-95.
    PMID: 27041391 DOI: 10.1111/jocd.12218
    OBJECTIVE: Cocoa pods are abundant waste materials of cocoa plantation, which are usually discarded onto plantation floors. However, due to poor plantation management, the discarded cocoa pods can create suitable breeding ground for Phytophthora palmivora, which is regarded as the causal agent of the black pod disease. On the other hand, cocoa pods potentially contain antioxidant compounds. Antioxidant compounds are related to the protection of skin from wrinkles and can be used as functional cosmetic ingredients. Therefore, in this study, cocoa pods were extracted and to be used as active ingredients for antiwrinkles.

    METHODS: The active compounds in cocoa pod extracts (CPE) were screened using liquid chromatography-mass spectrometry (LC-MS). Fibroblast cells were used to determine the effective concentration of CPE to maintain the viability for at least 50% of the cells (EC50 ). The gel was tested by 12 panelists to determine the efficacy of CPE in gel form using Visioscan to reduce skin wrinkles and improve skin condition.

    RESULTS: CPE was detected to contain malic acid, procyanidin B1, rosmarinic acid, procyanidin C1, apigenin, and ellagic acid, all of which may contribute to functional cosmetic properties of CPE. The EC50 value of cocoa pod extracts was used to calculate the amount of CPE to be incorporated into gel so that the formulated product could reach an effective concentration of extract while being nonintoxicant to the skin cell. The results showed that CPE is potential ingredient to reduce wrinkles. Skin wrinkles reduced at 6.38 ± 1.23% with the application of the CPE gel within 3 weeks and significantly improved further (12.39 ± 1.59%) after 5 weeks. The skin hydration increased (3.181 ± 1.06%) after 3 weeks of the CPE gel application.

    CONCLUSION: Flavonoid compounds in CPE contributed to the functional cosmetic properties of CPE. The CPE which is nontoxic to skin cells help to reduce wrinkles on skin after 3 weeks of application. CPE can be used as the active ingredients in antiwrinkle products, and prolonged application may result in significant visual changes to the naked eyes.

  7. Md Yusof AH, Abd Gani SS, Zaidan UH, Halmi MIE, Zainudin BH
    Molecules, 2019 Feb 16;24(4).
    PMID: 30781448 DOI: 10.3390/molecules24040711
    This study investigates the ultrasound-assisted extraction of flavonoids from Malaysian cocoa shell extracts, and optimization using response surface methodology. There are three variables involved in this study, namely: ethanol concentration (70⁻90 v/v %), temperature (45⁻65 °C), and ultrasound irradiation time (30⁻60 min). All of the data were collected and analyzed for variance (ANOVA). The coefficient of determination (R²) and the model was significant in interaction between all variables (98% and p < 0.0001, respectively). In addition, the lack of fit test for the model was not of significance, with p > 0.0684. The ethanol concentration, temperature, and ultrasound irradiation time that yielded the maximum value of the total flavonoid content (TFC; 7.47 mg RE/g dried weight (DW)) was 80%, 55 °C, and 45 min, respectively. The optimum value from the validation of the experimental TFC was 7.23 ± 0.15 mg of rutin, equivalent per gram of extract with ethanol concentration, temperature, and ultrasound irradiation time values of 74.20%, 49.99 °C, and 42.82 min, respectively. While the modelled equation fits the data, the T-test is not significant, suggesting that the experimental values agree with those predicted by the response surface methodology models.
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