Jackfruit is a sweet tropical fruit with very pleasant aroma, and the ripe seeds are edible. In this study, jackfruit seed proteins were isolated and subjected to trypsin digestion. The resultant protein hydrolysate was then subjected to antioxidant assay-guided purification, using centrifugal filtration, C18 reverse-phase and strong cation exchange (SCX) fractionations. The purified SCX fraction was further analyzed by de novo peptide sequencing, and two peptide sequences were identified and synthesized. Peptide JFS-2 (VGPWQK) was detected with antioxidant potential, with EC50 value comparable to that of commercial GSH antioxidant peptide. Additionally, the identified peptides were tested with protein protection potential, in an albumin protein denaturation inhibitory assay. Concurrently, we also investigated the pH, temperature, and gastrointestinal-digestion stability profiles for the identified peptide. With further research efforts, the identified peptides could potentially be developed into preservative agent for protein-rich food systems or as health-promoting diet supplements.
A modeling technique based on absorbed microwave energy was proposed to model microwave-assisted extraction (MAE) of antioxidant compounds from cocoa (Theobroma cacao L.) leaves. By adapting suitable extraction model at the basis of microwave energy absorbed during extraction, the model can be developed to predict extraction profile of MAE at various microwave irradiation power (100-600 W) and solvent loading (100-300 ml). Verification with experimental data confirmed that the prediction was accurate in capturing the extraction profile of MAE (R-square value greater than 0.87). Besides, the predicted yields from the model showed good agreement with the experimental results with less than 10% deviation observed. Furthermore, suitable extraction times to ensure high extraction yield at various MAE conditions can be estimated based on absorbed microwave energy. The estimation is feasible as more than 85% of active compounds can be extracted when compared with the conventional extraction technique.
This work demonstrated the optimization and scale up of microwave-assisted extraction (MAE) and ultrasonic-assisted extraction (UAE) of bioactive compounds from Orthosiphon stamineus using energy-based parameters such as absorbed power density and absorbed energy density (APD-AED) and response surface methodology (RSM). The intensive optimum conditions of MAE obtained at 80% EtOH, 50mL/g, APD of 0.35W/mL, AED of 250J/mL can be used to determine the optimum conditions of the scale-dependent parameters i.e. microwave power and treatment time at various extraction scales (100-300mL solvent loading). The yields of the up scaled conditions were consistent with less than 8% discrepancy and they were about 91-98% of the Soxhlet extraction yield. By adapting APD-AED method in the case of UAE, the intensive optimum conditions of the extraction, i.e. 70% EtOH, 30mL/g, APD of 0.22W/mL, AED of 450J/mL are able to achieve similar scale up results.
A study on the proximate composition, minerals, vitamins, carotenoids, amino acids, fatty acids profiles and some physicochemical properties of freeze dried Gracilaria changii was conducted. It was discovered that this seaweed was high in dietary fibre (64.74±0.82%), low in fat (0.30±0.02%) and Na/K ratio (0.12±0.02). The total amino acid content was 91.90±7.70% mainly essential amino acids (55.87±2.15mgg(-1)) which were comparable to FAO/WHO requirements. The fatty acid profiles were dominated by the polyunsaturated fatty acids particularly docosahexaenoic (48.36±6.76%) which led to low ω6/ω3, atherogenic, and thrombogenic index. The physicochemical properties of this seaweed namely the water holding and the swelling capacity were comparable to some commercial fibre rich products. This study suggested that G. changii could be potentially used as ingredients to improve nutritive value and texture of functional foods for human consumption.
A higher specific activity of microbial transglutaminase (mTGase) is desirable for a broad range of applications ranging from food industry to biotechnology. Three-dimensional docking simulation of mTGase revealed that residues V65, W69, and Y75 were critical for substrate recognition. A semi-rational mutagenesis approach was applied to each residue to generate three separate mini mutant libraries. A high-throughput screening process identified five mutants that demonstrated improved specific activities than the wild type (WT) mTGase were isolated from the Y75 mini mutant library. Mutant Y75L showed approximately 60% increment in specific activity and improved substrate specificity. Conjugation of two heterologous single-chain fragment variable clones to generate a diabody with mutant Y75L was successfully performed and validated. This work demonstrates the successful application of semi-rational mutagenesis coupled with a high-throughput screening approach to identify mTGase mutants with improved specific activities and specificities which are beneficial for protein-protein conjugation.
Different extraction conditions were applied to investigate the effect of temperature, extraction time and substrate-extractant ratio on pectin extraction from cocoa husks. Pectin was extracted from cocoa husks using water, citric acid at pH 2.5 or 4.0, or hydrochloric acid at pH 2.5 or 4.0. Temperature, extraction time and substrate-extractant ratio affected the yields, uronic acid contents, degrees of methylation (DM) and degrees of acetylation (DA) of the extracted pectins using the five extractants differently. The yields and uronic acid contents of the extracted pectins ranged from 3.38-7.62% to 31.19-65.20%, respectively. The DM and DA of the extracted pectins ranged from 7.17-57.86% to 1.01-3.48%, respectively. The highest yield of pectin (7.62%) was obtained using citric acid at pH 2.5 [1:25 (w/v)] at 95 °C for 3.0 h. The highest uronic acid content (65.20%) in the pectin was obtained using water [1:25 (w/v)] at 95 °C for 3.0 h.
A high-performance polyacid ion exchange (IEX) nanofiber membrane was used in membrane chromatography for the recovery of lysozyme from chicken egg white (CEW). The polyacid IEX nanofiber membrane (P-BrA) was prepared by the functionalization of polyacrylonitrile (PAN) nanofiber membrane with ethylene diamine (EDA) and bromoacetic acid (BrA). The adsorption performance of P-BrA was evaluated under various operating conditions using Pall filter holder. The results showed that optimal conditions of IEX membrane chromatography for lysozyme adsorption were 10% (w/v) of CEW, pH 9 and 0.1 mL/min. The purification factor and yield of lysozyme were 402 and 91%, respectively. The adsorption process was further scaled up to a larger loading volume, and the purification performance was found to be consistent. Furthermore, the regeneration of IEX nanofiber membrane was achieved under mild conditions. The adsorption process was repeated for five times and the adsorption capacity of adsorber was found to be unaffected.
Myosin heavy chain (MHC) isoforms in goat muscles and their possible relationships with meat quality have not been fully elucidated. This study characterized the MHC isoforms in different caprine muscles using sodium dodecyl sulphate glycerol gel electrophoresis (SDS-GGE). The relationships between MHC isoforms, calpain systems and meat quality characteristics of different muscles in goats were examined. Four muscles, namely infraspinatus (IF), longissimus dorsi (LD), psoas major (PM) and supraspinatus (SS) were obtained from ten Boer crossbred bucks (7-10 months old; 26.5 ± 3.5 kg, BW). The percentages of MHC I, MHC IIa and MHC IIx in SS, IF, PM and LD were 47.2, 38.3, 32.1, 11.9; 28.0, 42.1, 33.0, 36.4; and 24.8, 19.6, 34.9 and 51.7, respectively. IF and SS had higher levels of calpastatin, total collagen and insoluble collagen contents than did PM and LD. PM had longer sarcomere length than did other muscles. LD had higher collagen solubility, troponin-T degradation products and glycogen content than did other muscles. These results infer that variable fiber-type composition could account partially for the differences in the physicochemical properties of goat muscles.
The ultra-high-performance liquid chromatography tandem mass spectrometry (UHPLC-MS) method was built to quantify the casein glycomacropeptide (CGMP) in bovine dairy products accurately based on targeted proteomics. Qualitative analysis of theoretical peptides was carried out using high-resolution mass spectrometry (HRMS) and protein software. Isotope-labeled characteristic peptides were acquired via the labeled amino acid condensation method to correct the matrix effects. Peptide MAIPPK was the representative characteristic peptide for distinguishing the CGMP from κ-casein through trypsin digestion. After optimizing the pre-treatment conditions, the final 8% oxidant concentration was selected and the 10% formic acid concentration with 2.5 h oxidation time. Moreover, the results of methodological verification showed that the recovery rate was 103.7%, meanwhile the precision of inter-day and intra-day was less than 5%. In conclusion, the research demonstrated the characteristic peptide MAIPPK could quantitatively applied to detect CGMP in dairy products.
Chili shrimp paste (CSP) is an exotic traditional Southeast Asian condiment prepared using mainly fresh chilies and fermented shrimp paste (belacan) which attributed to strong pungent fishy odor. This study aims to evaluate the effectiveness of electron beam irradiation (EBI) exposure on CSP for microorganisms decontamination, and its physicochemical qualities changes. Changes in capsaicinoid contents and volatile compounds were analyzed using HPLC and GC-MS. Mesophilic bacteria, yeast, mold and pathogenic Enterobacteriaceae decreased as irradiation dose increasing from 0 to 10kGy. EBI at 10kGy effectively decontaminated the samples with no significant effects on phenolic and capsaicinoids contents compared to the fresh samples. From 24 compounds, irradiated CSP retained 23 volatile compounds, while thermally treated CSP has only 19 compounds. EBI at 10kGy is effective for decontamination in CSP with lesser destructive effect on volatile compounds and texture compared to thermal treatment.
Volatile compounds in the peel of calamansi (Citrus microcarpa) from Malaysia, the Philippines and Vietnam were extracted with dichloromethane and hexane, and then analysed by gas chromatography-mass spectroscopy/flame ionisation detector. Seventy-nine compounds representing >98% of the volatiles were identified. Across the three geographical sources, a relatively small proportion of potent oxygenated compounds was significantly different, exemplified by the highest amount of methyl N-methylanthranilate in Malaysian calamansi peel. Principal component analysis and canonical discriminant analysis were applied to interpret the complex volatile compounds in the calamansi peel extracts, and to verify the discrimination among the different origins. In addition, four common hydroxycinnamic acids (caffeic, p-coumaric, ferulic and sinapic acids) were determined in the methanolic extracts of calamansi peel using ultra-fast liquid chromatography coupled to photodiode array detector. The Philippines calamansi peel contained the highest amount of total phenolic acids. In addition, p-Coumaric acid was the dominant free phenolic acids, whereas ferulic acid was the main bound phenolic acid.
Calamansi juices from three countries (Malaysia, the Philippines and Vietnam) were characterised through measuring volatiles, physicochemical properties and non-volatiles (sugars, organic acids and phenolic acids). The volatile components of manually squeezed calamansi juices were extracted using dichloromethane and headspace solid-phase microextraction, and then analysed using gas chromatography-mass spectrometry/flame ionisation detector, respectively. A total of 60 volatile compounds were identified. The results indicated that the Vietnam calamansi juice contained the highest amount of volatiles. Two principal components obtained from principal component analysis (PCA) represented 89.65% of the cumulative total variations of the volatiles. Among the non-volatile components, these three calamansi juices could be, to some extent, differentiated according to fructose and glucose concentrations. Hence, this study of calamansi juices could lead to a better understanding of calamansi fruits.
The objective of this study was to improve product quality of dehydrated fruits (apple, pear, papaya, mango) using combined drying techniques. This involved investigation of bioactivity, colour, and sensory assessment on colour of the dried products as well as the retention of the bio-active ingredients. The attributes of quality were compared in regard to the quality of dehydrated samples obtained from continuous heat pump (HP) drying technique. It was found that for apple, pear and mango the total colour change (ΔE) of samples dried using continuous heat pump (HP) or heat pump vacuum-microwave (HP/VM) methods was lower than of samples dried by other combined methods. However, for papaya, the lowest colour change exhibited by samples dried using hot air-cold air (HHC) method and the highest colour change was found for heat pump (HP) dehydrated samples. Sensory evaluation revealed that dehydrated pear with higher total colour change (ΔE) is more desirable because of its golden yellow appearance. In most cases the highest phenol content was found from fruits dried by HP/VM method. Judging from the quality findings on two important areas namely colour and bioactivity, it was found that combined drying method consisted of HP pre-drying followed by VM finish drying gave the best results for most dehydrated fruits studied in this work as the fruits contain first group of polyphenol compounds, which preferably requires low temperature followed by rapid drying strategy.
The elemental profiles of six honey samples from Malaysia had been constructed using the data obtained from both ICP-AES and ICP-MS. Potassium and sodium were the most abundant minerals covering from 69.3-78.6% and 14.1-28.7%, respectively. The ratio of potassium to sodium was more than one. Even though the minerals and trace elements composition varied dependent on the type of honey samples, there was no statistically significant difference between the analysed honey samples, namely tualang, gelam, acacia and a few forest honeys based on two-factor ANOVA and cluster analysis. The total element content of honey samples were strongly correlated with the electrical conductivity, but only have moderate correlation with the ash content and honey colour based on the regression analysis. PCA result on the available elemental data from worldwide honeys, including honey samples from Malaysia revealed that potassium and sodium were the mineral markers to distinguish honey origin. Both tualang and gelam honey samples from Malaysia have close mineral profile with sesame honeys from Egypt and multifloral honeys from India, whereas forest honeys Malaysia were near to avocado honeys from Spain and multifloral honeys from India.
Both total phenolic content (TPC) and total flavonoid content (TFC) of Labisia pumila extracts were determined spectrophotometrically. L. pumila leaves extracted in 60% methanol (MeOH) were fractionated on C18 cartridge and the antioxidant property of each fraction was determined by measuring free radical 1,1-diphenyl-2-picrylhydrazyl (DPPH) scavenging activity. The 40% MeOH fraction exhibited the highest scavenging activity. Nine flavonols (quercetin, myricetin and kaempferol), two flavanols (catechin and epigallocatechin) and nine phenolic acids were identified from this active fraction by UPLC-ESI-MS/MS, and confirmed by comparison with the mass spectra of standard aglycones, theoretical fragments generated from MS Fragmenter software, and literature values.
This study was aimed to extract bioactive peptides from the white and purple flower varieties of Orthosiphon aristatus leaves. The herb is well known for its pharmacological importance, possibly attributed to its plant proteins. Phenol based extraction was used to extract plant proteins, and then hydrolysed by proteolytic enzymes such as trypsin (serine protease) and pepsin (aspartic protease). MS/MS analysis revealed that 145 and 125 proteins were detected from the white and purple flower varieties, respectively. Trypsin hydrolysates were showed to have a higher degree of hydrolysis (24-33%), resulting in higher antioxidant and antibacterial activities. The white flower of trypsin hydrolysates showed a higher radical scavenging activity which could be attributed to its higher content of stress proteins (19%). However, trypsin hydrolysates from the purple flower showed higher ferric reducing power and bacterial growth inhibition. The performance of hydrolysates was better than ampicillin in inhibiting Acinetobacter baumanni and Staphylococcus aureus.
The adulteration of edible fats is a kind of fraud that impairs the physical and chemical features of the original lipid materials. It has been detected in various food, pharmaceutical and cosmeceutical products. Differential scanning calorimetry (DSC) is the robust thermo-analytical machine that permits to fingerprint the primary crystallisation of triacylglycerols (TAGs) molecules and their transition behaviours. The aims of this study was to assess the cross-contamination caused by lard concentration of 0.5-5% in the mixture systems containing beef tallow (BT) and chicken fat (CF) separately. TAGs species of pure and adulterated lipids in relation to their crystallisation and melting parameters were studied using principal components analysis (PCA). The results showed that by using the heating profiles the discrimination of LD from BT and CF was very clear even at low dose of less than 1%. Same observation was depicted from the crystallisation profiles of BT adulterated by LD doses ranging from 0.1% to 1% and from 2% to 5%, respectively. Furthermore, CF adulterated with LD did not exhibit clear changes on its crystallisation profiles. Consequently, DSC coupled with PCA is one of the techniques that might use to monitor and differentiate the minimum adulteration levels caused by LD in different animal fats.
The method of liquid chromatographic tandem mass spectrometry was utilized and modified to confirm and quantify acrylamide in heating cooking oil and animal fat. Heating asparagine with various cooking oils and animal fat at 180°C produced varying amounts of acrylamide. The acrylamide in the different cooking oils and animal fat using a constant amount of asparagine was measured. Cooking oils were also examined for peroxide, anisidine and iodine values (or oxidation values). A direct correlation was observed between oxidation values and acrylamide formation in different cooking oils. Significantly less acrylamide was produced in saturated animal fat than in unsaturated cooking oil, with 366ng/g in lard and 211ng/g in ghee versus 2447ng/g in soy oil, followed by palm olein with 1442ng/g.
This work investigated the underlying formation of acrylamide from amino acids in frying oils during high temperatures and at different times via modeling systems. Eighteen amino acids were used in order to determine which one was more effective on acrylamide production. Significantly the highest amount of acrylamide was produced from asparagine (5987.5µg/kg) and the lowest from phenylalanine (9.25µg/kg). A constant amount of asparagine and glutamine in palm olein and soy bean oils was heated up in modelling system at different temperatures (160, 180 and 200°C) and times (1.5, 3, 4.5, 6, 7.5min). The highest amount of acrylamide was found at 200°C for 7.5min (9317 and 8511µg/kg) and lowest at 160°C for 1.5min (156 and 254µg/kg) in both frying oils and both amino acids. Direct correlations have been found between time (R2=0.884), temperature (R2=0.951) and amount of acrylamide formation, both at p<0.05.