Purification of lipase from Burkholderia metallica fermentation broth in a column chromatography using polymer impregnated resins.

In this work, porous glass beads grafted with polyethylene glycol (PEG) were used as an adsorbent to purify lipase from Burkholderia metallica in column chromatography. The purification parameters viz. salt stability, types and concentrations of PEG and salt, pH of the binding solution, and flow rate were studied to determine the performance of the purification system in an XK16/20 column. The crude lipase was mixed with different types and concentrations of salts 1-5% (w/w) (sodium citrate, potassium citrate, and sodium acetate) and subjected to the column containing the polymeric glass bead. One-variable-at-a-time experimentation revealed that 20% (w/w) PEG 6000 g/mol impregnated glass beads with a binding solution of 5% sodium citrate at pH 7.7, a flow rate of 1.0 mL/min and extraction time of 10 min resulted in the highest purification factor and recovery yield at 3.67 and 88%, respectively. The purified lipase has 55 ∼ 60 kDa molecular mass. The outcome of the study showed PEG could be applied to modify the inert glass beads into polymeric form, providing a biocompatible and mild separation condition for lipase. Thus, PEG could be successfully applied for the purification of lipase from B. metallica fermentation broth using column chromatography.

Bioseparation of phycocyanin from Phormidium tergestinum using an aqueous two-phase system.

This study aimed at purification of phycocyanin (PC) from Phormidium tergestinum using an aqueous two-phase system (ATPS) comprised of polyethylene glycol (PEG) and salts. The partitioning efficiency of PC in ATPS and the effect of phase composition, pH, crude loading, and neutral salts on purification factor and yield were investigated. Results showed that PC was selectively partitioned toward bottom phase of the system containing potassium phosphate. Under optimum conditions of 20% (w/w) PEG 4000, 10% (w/w) potassium phosphate, 20% (v/v) crude load at pH 7, with addition of 0.5% (w/w) NaCl, PC from P. tergestinum was partially purified up to 5.34-fold with a yield of 87.8%. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that the molecular weight of PC was ∼19 kDa. Results from this study demonstrated ATPS could be used as a potential approach for the purification of PC from P. tergestinum.

Recovery of a Bacteriocin-Like Inhibitory Substance from Lactobacillus bulgaricus FTDC 1211 Using Polyethylene-Glycol Impregnated Amberlite XAD-4 Resins System.

Lactobacillus bulgaricus is a LAB strain which is capable of producing bacteriocin substances to inhibit Staphylococcus aureus. The aim of this study was to purify a bacteriocin-like inhibitory substance (BLIS) produced by L. bulgaricus FTDC 1211 using an aqueous impregnated resins system consisting of polyethylene-glycol (PEG) impregnated on Amberlite XAD4. Important parameters influencing on purification of BLIS, such as the molecular weight and concentration of PEG, the concentration and pH of sodium citrate and the concentration of sodium chloride, were optimized using a response surface methodology. Under optimum conditions of 11% (w/w) of PEG 4000 impregnated Amberlite XAD4 resins and 2% (w/w) of sodium citrate at pH 6, the maximum purification factor (3.26) and recovery yield (82.69% ± 0.06) were obtained. These results demonstrate that AIRS could be used as an alternate purification system in the primary recovery step.

Thermoplastic starch (TPS) bioplastic, the green solution for single-use petroleum plastic food packaging - A review.

Plastic throughout the years is now one of the biggest world commodities and also the largest pollution to have an environmental impact, accumulating in landfills and also leaching into water systems and oceans. Especially with the shift to single-use disposable plastic, evermore positions plastics as the number one novel entity that pollutes the earth. This shift is also consistent in the food packaging industry. Managing plastic waste is still an issue at large, while the process of pyrolysis incineration still requires an obscene amount of energy that also does not resolve the problems with its environmental impact, the cost of mechanical-chemical degradation even outweighs the cost of producing the materials, and biodegradation process is a very slow and long process. Converting to bioplastics is one of the potential solutions to the global plastic issue. This review covers the potentials, limitations, challenges, progress and advancements of bioplastics, especially thermoplastic starch (starch-based bioplastic) in their efforts to replace petroleum plastics in food packaging and smart food packaging, especially for single-use (disposable) food packaging.

Characterization of alcohol/salt aqueous two-phase system for optimal separation of gallic acids.

Gallic acid (GA) is a hydrophilic polyphenol which is noteworthy for strong antioxidant capacity. The drawbacks of conventional extraction approaches such as time-consuming and high processing cost are often viewed as a hurdle to extract GA from plant sources in industrial scale. Aqueous two-phase system (ATPS) is a separation approach which can be employed as an alternative to the conventional approaches. The partition behaviour of GA in an alcohol/salt ATPS was investigated in this study to aid the development of industrial scale ATPS to extract GA from natural sources. The separation of GA was characterized by determining the types of alcohol and salt, phase composition, sample load, pH of the system and addition of adjuvants applied in the alcohol/salt ATPS construction. The hydrophilic GA was targeted to the salt-rich phase of the alcohol/salt ATPS with a partition coefficient (KGA) of 25.00 ± 0.00. The optimum condition of ATPS for the maximum partition of GA was achieved in ATPS comprised of 24% (w/w) 1-propanol and 22% (w/w) phosphate salt at pH 8 with 5% (w/w) of 1 mg/mL sample loading and 2% (w/w) NaCl addition. The findings suggest that ATPS can be applied for separation of GA from various natural sources.

Isolation and characterization of Enterococcus faecium DSM 20477 with ability to secrete antimicrobial substance for the inhibition of oral pathogen Streptococcus mutans UKMCC 1019.

Streptococcus mutans and Candida albicans are the main oral pathogens which contribute to dental caries that affects all ages of human being. OBJECTIVES: This study focuses on the potential of crude cell free supernatant (CCFS) from lactic acid bacteria (LAB) to inhibit of the growth of S. mutans UKMCC 1019. DESIGN: A total of 61 CCFS from LAB strains were screened for their inhibitory ability against S. mutans UKMCC 1019 by broth microdilution method. The selected LAB with highest antimicrobial activity was identified and its CCFS was characterized for pH stability, temperature tolerance, enzyme sensitivity, metabolism of carbohydrates, enzymatic activities and antimicrobial activity against S. mutans UKMCC 1019 and C. albicans UKMCC 3001 by well diffusion assay. The effect of CCFS on cell structure of S. mutans UKMCC 1019 was observed under transmission electron microscopy (TEM). RESULTS: The CCFS from isolate CC2 from Kimchi showed the highest inhibition against S. mutans UKMCC 1019, which was 76.46 % or 4406.08 mm2/mL and it was identified to be most closely related to Enterococcus faecium DSM 20477 based on 16 s rRNA sequencing. The CCFS of E. faecium DSM 20477 had high tolerance to acidic and alkaline environment as well as high temperature. It also shows high antifungal activities against C. albicans UKMCC 3001 with 2362.56 mm2/mL. Under TEM, the cell walls and the cytoplasm membrane of S. mutans UKMCC 1019 were disrupted by the antimicrobial substance, causing cell lysis. CONCLUSIONS: Hence, the CCFS from E. faecium DSM 20477 is a potential bacteriocin in future for the treatment of dental caries.

Comprehensive studies on optimization of cellulase and xylanase production by a local indigenous fungus strain via solid state fermentation using oil palm frond as substrate.

The high cost of cellulases remains the most significant barrier to the economical production of bio-ethanol from lignocellulosic biomass. The goal of this study was to optimize cellulases and xylanase production by a local indigenous fungus strain (Aspergillus niger DWA8) using agricultural waste (oil palm frond [OPF]) as substrate. The enzyme production profile before optimization indicated that the highest carboxymethyl cellulose (CMCase), filter paper (FPase), and xylanase activities of 1.06 U/g, 2.55 U/g, and 2.93 U/g were obtained on day 5, day 4, and day 5 of fermentation, respectively. Response surface methodology was used to study the effects of several key process parameters in order to optimize cellulase production. Of the five physical and two chemical factors tested, only moisture content of 75% (w/w) and substrate amount of 2.5 g had statistically significant effect on enzymes production. Under optimized conditions of 2.5 g of substrate, 75% (w/w) moisture content, initial medium of pH 4.5, 1 × 106 spores/mL of inoculum, and incubation at ambient temperature (±30°C) without additional carbon and nitrogen, the highest CMCase, FPase, and xylanase activities obtained were 2.38 U/g, 2.47 U/g, and 5.23 U/g, respectively. Thus, the optimization process increased CMCase and xylanase production by 124.5 and 78.5%, respectively. Moreover, A. niger DWA8 produced reasonably good cellulase and xylanase titers using OPF as the substrate when compared with previous researcher finding. The enzymes produced by this process could be further use to hydrolyze biomass to generate reducing sugars, which are the feedstock for bioethanol production.

Purification of ß-mannanase derived from Bacillus subtilis ATCC 11774 using ionic liquid as adjuvant in aqueous two-phase system.

The partitioning of ß-mannanase derived from Bacillus subtilis ATCC 11774 in aqueous two-phase system (ATPS) was studied. The ATPS containing different molecular weight of polyethylene glycol (PEG) and types of salt were employed in this study. The PEG/salt composition for the partitioning of ß-mannanase was optimized using response surface methodology. The study demonstrated that ATPS consists of 25% (w/w) of PEG 6000 and 12.52% (w/w) of potassium citrate is the optimum composition for the purification of ß-mannanase with a purification fold (PF) of 2.28 and partition coefficient (K) of 1.14. The study on influences of pH and crude loading showed that ATPS with pH 8.0 and 1.5% (w/w) of crude loading gave highest PF of 3.1. To enhance the partitioning of ß-mannanase, four ionic liquids namely 1-butyl-3-methylimidazolium tetrafluoroborate ([Bmim]BF4), 1-ethyl-3-methylimidazolium tetrafluoroborate ([Emim]BF4), 1-butyl-3-methylimidazolium bromide ([Bmim]Br), 1-ethyl-3-methylimidazolium bromide ([Emim]Br) was added into the system as an adjuvant. The highest recovery yield (89.65%) was obtained with addition of 3% (w/w) of [Bmim]BF4. The SDS-PAGE analysis revealed that the ß-mannanase was successfully recovered in the top phase of ATPS with the molecular size of 36.7kDa. Therefore, ATPS demonstrated a simple and efficient approach for recovery and purification of ß-mannanase from fermentation broth in one single-step strategy.

Aqueous two-phase flotation for primary recovery of bacteriocin-like inhibitory substance (BLIS) from Pediococcus acidilactici Kp10.

An aqueous two-phase flotation (ATPF) system based on polyethylene glycol (PEG) and sodium citrate (NaNO3C6H5O7·2H2O) was considered for primary recovery of bacteriocin-like inhibitory substance (BLIS) from Pediococcus acidilactici Kp10. The effects of ATPF parameters namely phase composition, tie-line length (TLL), volume ratio between the two phases (VR), amount of crude load (CL), pH, nitrogen gas flow rate (FR) and flotation time (FT) on the performance of recovery were evaluated. BLIS was mainly concentrated into the upper PEG-rich phase in all systems tested so far. The optimum conditions for BLIS purification, which composed of PEG 8000/sodium citrate, were: TLL of 42.6, VR of 0.4, CL of 22% (w/w), pH 7, average FT of 30min and FR of 20mL/min. BLIS was partially purified up to 5.9-fold with a separation efficiency of 99% under this optimal conditions. A maximum yield of BLIS activity of about 70.3% was recovered in the PEG phase. The BLIS from the top phase was successfully recovered with a single band in SDS-gel with molecular weight of about 10-15kDa. ATPF was found to be an effective technique for the recovery of BLIS from the fermentation broth of P. acidilactici Kp10.

Primary recovery of a bacteriocin-like inhibitory substance derived from Pediococcus acidilactici Kp10 by an aqueous two-phase system.

A polymer-salt aqueous two-phase system (ATPS) consisting of polyethylene-glycol (PEG) with sodium citrate was developed for direct recovery of a bacteriocin-like inhibitory substance (BLIS) from a culture of Pediococcus acidilactici Kp10. The influences of phase composition, tie-line length (TLL), volume ratio (VR), crude sample loading, pH and sodium chloride (NaCl) on the partition behaviour of BLIS was investigated. Under optimum conditions of ATPS, the purification of BLIS was achieved at 26.5% PEG (8000)/11% sodium citrate with a TLL of 46.38% (w/w), VR of 1.8, and 1.8% crude load at pH 7 without the presence of NaCl. BLIS from P. acidilactici Kp10 was successfully purified by the ATPS up to 8.43-fold with a yield of 81.18%. Given that the operation of ATPS is simple, environmentally friendly and cost-effective, as it requires only salts and PEG, it may have potential for industrial applications in the recovery of BLIS from fermentation broth.