The aim of this study was to optimize the antioxidant activity of Piper nigrum L. essential oil extracted using the supercritical carbon dioxide (SC-CO₂) technique. Response surface methodology was applied using a three-factor central composite design to evaluate the effects of three independent extraction variables: pressure of 15-30 MPa, temperature of 40-50 °C and dynamic extraction time of 40-80 min. The DPPH radical scavenging method was used to evaluate the antioxidant activity of the extracts. The results showed that the best antioxidant activity was achieved at 30 MPa, 40 °C and 40 min. The extracts were analyzed by GC-FID and GC-MS. The main components extracted using SC-CO₂ extraction in optimum conditions were β-caryophyllene (25.38 ± 0.62%), limonene (15.64 ± 0.15%), sabinene (13.63 ± 0.21%), 3-carene (9.34 ± 0.04%), β-pinene (7.27 ± 0.05%), and α-pinene (4.25 ± 0.06%). The essential oil obtained through this technique was compared with the essential oil obtained using hydro-distillation. For the essential oil obtained by hydro-distillation, the most abundant compounds were β-caryophyllene (18.64 ± 0.84%), limonene (14.95 ± 0.13%), sabinene (13.19 ± 0.17%), 3-carene (8.56 ± 0.11%), β-pinene (9.71 ± 0.12%), and α-pinene (7.96 ± 0.14%). Radical scavenging activity of the extracts obtained by SC-CO₂ and hydro-distillation showed an EC₅₀ of 103.28 and 316.27 µg mL(-1) respectively.
As a novel method of purification, an aqueous organic phase system (AOPS) was employed to purify pectinase from mango waste. The effect of different parameters, such as the alcohol concentration (ethanol, 1-propanol, and 2-propanol), the salt type and concentration (ammonium sulfate, potassium phosphate and sodium citrate), the feed stock crude load, the aqueous phase pH and NaCl concentration, were investigated in the recovery of pectinase from mango peel. The partition coefficient (K), selectivity (S), purification factor (PF) and yield (Y, %) were investigated in this study as important parameters for the evaluation of enzyme recovery. The desirable partition efficiency for pectinase purification was achieved in an AOPS of 19% (w/w) ethanol and 22% (w/w) potassium phosphate in the presence of 5% (w/w) NaCl at pH 7.0. Based on the system, the purification factor of pectinase was enhanced 11.7, with a high yield of 97.1%.
The stone fish (Actinopyga lecanora) ethanolic and methanolic tissue extracts were investigated for total phenolic contents (TPCs) as well as antioxidant activity using 2,2-diphenyl-1-picrylhydrazyl (DPPH(•)) radical scavenging activity and ferric reducing antioxidant power (FRAP) assays. Both extracts showed low amount of phenolics (20.33 to 17.03 mg of gallic acid equivalents/100 g dried sample) and moderate antioxidant activity (39% to 34% DPPH(•) radical scavenging activity and 23.95 to 22.30 mmol/100 mL FeSO4 FRAP value). Enzymatic proteolysis was carried out in order to improve the antioxidant activity using six commercially available proteases under their optimum conditions. The results revealed that the highest increase in antioxidant activity up to 85% was obtained for papain-generated proteolysate, followed by alcalase (77%), trypsin (75%), pepsin (68%), bromelain (68%), and flavourzyme (50%) as measured by DPPH(•) radical scavenging activity, whilst for the FRAP value, the highest increase in the antioxidant activity up to 39.2 mmol/100 mL FeSO4 was obtained for alcalase-generated proteolysate, followed by papain (29.5 mmol/100 mL FeSO4), trypsin (23.2 mmol/100 mL FeSO4), flavourzyme (24.7 mmol/100 mL FeSO4), bromelain (22.9 mmol/100 mL FeSO4), and pepsin (20.8 mmol/100 mL FeSO4). It is obvious that proteolysis of stone fish tissue by proteolytic enzymes can considerably enhance its antioxidant activity.
Bovine gelatin is a biopolymer which has good potential to be used in encapsulating matrices for probiotic candidate Bifidobacterium pseudocatenulatum strain G4 (G4) because of its amphoteric nature characteristic. Beads were prepared by the extrusion method using genipin and sodium alginate as a cross-linking agent. The optimisation of bovine gelatin-genipin-sodium alginate combinations was carried out using face central composition design (FCCD) to investigate G4 beads' strength, before and after exposed to simulated gastric (SGF), intestinal fluids (SIF), and encapsulation yield. A result of ANOVA and the polynomial regression model revealed the combinations of all three factors have a significant effect (p < 0.05) on the bead strength. Meanwhile, for G4 encapsulation yield, only genipin showed less significant effect on the response. However, the use of this matrix remained due to the intermolecular cross-linking ability with bovine gelatin. Optimum compositions of bovine gelatin-genipin-sodium alginate were obtained at 11.21% (w/v), 1.96 mM, and 2.60% (w/v), respectively. A model was validated for accurate prediction of the response and showed no significant difference (p > 0.05) with experimental values.
Penicillium candidum (PCA 1/TT031) synthesizes different types of extracellular proteases. The objective of this study is to optimize polyethylene glycol (PEG)/citrate based on an aqueous two-phase system (ATPS) and Response Surface Methodology (RSM) to purify protease from Penicillium candidum (PCA 1/TT031). The effects of different PEG molecular weights (1500-10,000 g/mol), PEG concentration (9%-20%), concentrations of NaCl (0%-10%) and the citrate buffer (8%-16%) on protease were also studied. The best protease purification could be achieved under the conditions of 9.0% (w/w) PEG 8000, 5.2% NaCl, and 15.9% sodium citrate concentration, which resulted in a one-sided protease partitioning for the bottom phase with a partition coefficient of 0.2, a 6.8-fold protease purification factor, and a yield of 93%. The response surface models displayed a significant (p ≤ 0.05) response which was fit for the variables that were studied as well as a high coefficient of determination (R²). Similarly, the predicted and observed values displayed no significant (p > 0.05) differences. In addition, our enzyme characterization study revealed that Penicillium candidum (PCA 1/TT031) produced a slight neutral protease with a molecular weight between 100 and 140 kDa. The optimal activity of the purified enzyme occurred at a pH of 6.0 and at a temperature of 50 °C. The stability between different pH and temperature ranges along with the effect of chemical metal ions and inhibitors were also studied. Our results reveal that the purified enzyme could be used in the dairy industry such as in accelerated cheese ripening.
This paper reports on the extraction, partial characterization and the potential application of crude polysaccharides from defatted coconut residue as a prebiotic. The coconut residue was defatted and extracted to obtain the crude polysaccharides and its physicochemical properties were determined. The crude polysaccharides were assessed for monosaccharide composition, total carbohydrate content, reducing sugar concentration and protein content determination. The functional group and structural elucidation of crude polysaccharides was also done using Fourier transform infrared spectra analysis. The product was then subjected to artificial human gastric juice treatment to determine digestibility. Finally, an in vitro proliferation and acid production by two probiotic bacteria namely Lactobacillus casei Shirota and Lactobacillus bulgaricus were included in this study. It was found that the defatted coconut residue contained ash (0.54%), moisture (55.42%), protein (1.69%), crude fat (17.26%) and carbohydrate (25.73%). The percentage of crude polysaccharides extracted was 0.73 ± 0.04. The two fractions of monosaccharides obtained were glucose and fructose. Total carbohydrate content of DCR was 13.35% (w/v). The quantitative value of the reducing sugars obtained was 20.71%. Protein content in the crude polysaccharides was 0.009% and the peaks which indicated the presence of protein were observed at around 1640 cm(-1) (amide I) and 1530 cm(-1) (amide II). DCR crude polysaccharides were highly resistant (88%) to hydrolysis when subjected to artificial human gastric juice. The product was found to markedly stimulate two tested probiotics to proliferate and produce organic acids. All the above findings are supportive of the fact that polysaccharides extracted from DCR, an industrial waste, have a vast potential to be exploited as novel prebiotics.
This report shows the partitioning and purification of alkaline extracellular lipase from Penicillium candidum (PCA 1/TT031) by solid-state fermentation (SSF). In the present analysis, some of the important parameters such as PEG concentration, PEG molecular mass, salt concentration and buffer concentration were optimised through the response surface methodology (RSM). The optimum aqueous two-phase systems (ATPS) environment consisted of 13.8% (w/w) phosphate buffer, 9.2% (w/w) PEG-3000 and 3.3% (w/w) NaCl at 25°C. The RSM approach was proved to be the most suitable methodology for the recovery of desired enzymes. In this method, the enzyme partitioned into the top phase of the PEG-buffer-NaCl ATPS. Under this experimental environment, the purification factor was found to be 33.9, the partition coefficient was 4.0 and the yield was found to be 84.0% of lipase. Moreover, the experimental and predicted results were in considerable agreement, which established the reliability and validity of the proposed model. The ATPS methodology is proven to be effective for the primary recovery of lipase at a low cost with a large loading capacity and possibility of linear scale up. In addition to using the existing methodologies for improving enzyme production, the use of statistical optimisation of the constituents of phases through RSM continues to be the basic and practical method.