Preparative Biochemistry & Biotechnology最新文献

An integrated approach involving response surface methodology (RSM) and artificial neural network-ant-colony hybrid optimization (ANN-ACO) was adopted to develop a bioprocess medium to increase the yield of Bacillus cereus neutral protease under submerged fermentation conditions. The ANN-ACO model was comparatively superior (predicted r² = 98.5%, mean squared error [MSE] = 0.0353) to RSM model (predicted r² = 86.4%, MSE = 23.85) in predictive capability arising from its low performance error. The hybrid model recommended a medium containing (gL^-1) molasses 45.00, urea 9.81, casein 25.45, Ca²⁺ 1.23, Zn²⁺ 0.021, Mn²⁺ 0.020, and 4.45% (vv^-1) inoculum, for a 6.75-fold increase in protease activity from a baseline of 76.63 UmL^-1. Yield was further increased in a 5-L bioreactor to a final volumetric productivity of 3.472 mg(Lh)^-1. The 10.0-fold purified 46.6-kDa-enzyme had maximum activity at pH 6.5, 45-55 °C, with K_m of 6.92 mM, V_max of 769.23 µmolmL^-1 min^-1, k_cat of 28.49 s^-1, and k_cat/K_m of 4.117 × 103 M^-1 s^-1, at 45 °C, pH 6.5. The enzyme was stabilized by Ca²⁺, activated by Zn²⁺ but inhibited by EDTA suggesting that it was a metallo-protease. The biomolecule significantly clarified orange and pineapple juices indicating its food industry application.

Synthetic dyes such as azo dyes are significant pollutants in the wastewater released from various textile industries. The low biodegradability and production from synthetic sources with high shelf life make azo dyes a challenging material for degradation. This study used chemically mutated Aspergillus terrus in the laccase production under solid-state fermentation using sugarcane bagasse. Initially, the wild-type strain produced a laccase activity of 4.12 U/mL. Later, the alkaline pretreatment of sugarcane bagasse showed a significant increase in laccase activity by 38.9%. Further, random mutagenesis treatment with 100 mM EMS generated a hyper laccase-producing strain with a 2.3-fold increment in laccase activity compared to the wild-type strain. The enzyme displayed optimal activity at pH 6.5 and 35 °C. The metal ions such as Fe³⁺ (29.4 U/mL), Fe²⁺ (20.8 U/mL) and Cu²⁺ (18.05 U/mL) showed positive effects on laccase activity. The crude laccase was used to bioremediate Congo red, a prominent azo dye used in textile and pharmaceutical industries. The preliminary studies with a crude enzyme displayed 68.86% dye decolourization after 24 h of incubation. Additionally, with Taguchi orthogonal array optimization experiments, the maximal dye decolorization of 78.24% was achieved by maintaining crude enzyme concentration (20 U), dye concentration (25 mg/L) and pH 4.5.

Since cytoplasmic expression of heterologous proteins with disulfide bonds leads to the formation of inclusion bodies in E. coli, periplasmic production is preferable. The N-terminal signal peptide attached to the secreted protein determines the type of secretory pathway through which the target protein is secreted; Sec, Tat, or SRP. The aim of this study was to design and compare two novel signal peptides for the secretion of recombinant neurturin (as a model) via the Sec and Tat pathways. For this purpose, we aligned the natural signal peptides from E. coli and Bacillus subtilis to identify the conserved amino acids and those with the highest repetition. The SignalP4.1 and TatP1.0 software were used to determine the secretion efficiency of the new signal peptides. The efficiency of new signal peptides was then evaluated and compared experimentally with two naturally used signal peptides. Quantitative analysis of Western blot bands showed that approximately 80% of the expressed neurturin was secreted into the periplasmic space by new signal peptides. Circular dichroism spectroscopy also confirmed the correct secondary structure of the secreted neurturin. In conclusion, these novel signal peptides can be used to secrete any other recombinant proteins to the periplasmic space of E. coli efficiently.

α-L-rhamnosidase (Rha) is ubiquitous in nature and has high feasibility in the food and biotechnology industries. A green and environmentally friendly method was used to improve the activity of Rha. Here, we show that the effects of ultrasound treatment on the Rha. Ultrasonic treatment at 80 W for 10 min yielded the highest enzyme activity. Treatment increased enzyme activity by 26.3% and half-life by 124 min. Further, treatment increased the catalytic efficiency of Rha and increased the substrate conversion rate by 33.88%. These results demonstrate that ultrasound increases the catalytic activity and stability of Rha. Thus, ultrasonic treatment of Rha is cost-effective on the industrial scale.

An efficient method of solid-state fermentation (SSF) is reported for producing bioactive phenolic compounds using soil-isolated fungi. Antioxidant activity using a rapid DPPH (1,1-diphenyl-2-picryl hydrazyl), was employed to screen the 120 fungal isolates from soil. Aspergillus terreus 1, Aspergillus fumigatus, Aspergillus terreus 2, Penicillium citrinum, Aspergillus wentii1, Aspergillus wentii 2, Penicillium expansum and Penicillium granulatum were chosen, concerning their antioxidant activity and total phenolic content. These fungal strains were applied on agro residues viz. sugarcane bagasse, corn cob, rice straw, pea pod and wheat straw, to evaluate the release of phenolic compounds. The fermented extracts from various agro-residues showed good antioxidant activity against DPPH, ferric ion, and nitric oxide radicals. The highest antioxidant activity was observed in fermented extracts of sugarcane bagasse, followed by pea pod. Additionally, the total phenolic content in the fermented extracts positively correlated with antioxidant potential. This study highlights the significant potential of solid substrate fermentation using soil-isolated fungi and agro-residues to produce bioactive phenolic compounds with potent antioxidant properties. The utilization of SSF for the extraction of bioactive compounds from natural sources not only offers a clean and sustainable approach but also contributes to the valorization of agro-industrial residues.

To create tissue-engineered vascular grafts (TEVGs) in vitro, vascular smooth muscle cells (VSMCs) must function effectively and produce sufficient extracellular matrix (ECM) in a three-dimensional space. In this study, we investigated whether the addition of insulin-transferrin-selenium (ITS), a medium supplement, could enhance TEVG formation. PGA fabric was used as the scaffold, and 1% ITS was added to the medium. After two weeks, the tissues were examined using electron microscopy and staining. The ITS group exhibited a denser structure and increased collagen production. VSMCs were cultured in two dimensions with ITS and assessed for collagen production, cell growth, and glucose metabolism. The results showed that ITS supplementation increased collagen production, cell growth, glucose utilization, lactate production, and ATP levels. Furthermore, reducing the amount of fetal bovine serum (FBS) in the medium did not affect the TEVGs or VSMCs when ITS was present. In conclusion, ITS improves TEVG construction by promoting VSMCs growth and reducing the need for FBS.

The removal of hexavalent chromium [Cr (VI)] using non-living cells of Yarrowia lipolytica was investigated. Batch and continuous studies on removal of Cr (VI) achieved 97% and 99% removal from aqueous phase, respectively. The specific uptake values at pH of 2 in batch process were 40.73 ± 1.3 mg/g and 30.09 ± 0.23 mg/g on non-living cells, when 100 and 200 mg/L of metal Cr (VI) concentrations were used. In order to investigate the regulation of Cr (VI) under continuous operation based on reaction volume numerically a new class of feedback controller from structure polynomial was designed. The proposed methodology was used to an experimentally kinetic model for a removal Cr (VI) from Yarrowia lipolytica biomass was showed satisfactory closed-loop performance the proposed controller. Starting from an off-line optimization performed in simulation, we present the controller implementation, focussing on the methodology required to could be suitable for implementation in real time. In our experimental results, we highlight some discrepancies between simulation and reality despite these differences, the controller managed to perform convergence to removal Cr (VI). Finally, the results validated with off-line samples suggest that the proposed control could be suitable for in application in potential scenarios for wastewater treatment.

Given the escalating demand for renewable biofuels amidst the continual consumption of fossil energy, the exploration and identification of microalgal strains for biodiesel production have become crucial. In this study, a microalgal strain named HDMA-12 was isolated from Lake Chenjiadayuan in China to evaluate its biodiesel potential. Phylogenetic analysis of its internal transcribed spacer sequences revealed HDMA-12 as a new molecular record in the genus Coelastrum. When cultivated in BG11 basal medium, HDMA-12 achieved a biomass of 635.7 mg L^-1 and a lipid content of 26.4%. Furthermore, the fatty acid methyl ester profile of HDMA-12 exhibited favorable combustion characteristics. Subjected to 200 mM NaCl stress, HDMA-12 reached its maximum biomass of 751.5 mg L^-1 and a lipid content of 28.9%. These findings indicate the promising prospects of HDMA-12 as a promising microalgal strain for further advancements in biodiesel production.

The Exopolysaccharide (EPS) producing novel strains of Enterococcus previously isolated from the vaginal source of pregnant women were selected based on ropy structure formation. The two selected strains, E.villorum SB-2 and E.rivorum S22-3, were found to be producing 2.87 g/l and 3.14 g/l EPS, respectively, in the minimal media (M17 media) after 24-hour fermentation under anaerobic condition. Both the strains have probiotic properties and have the potential to be used for industrial applications. The production media and fermentation conditions were optimized to enhance the EPS production using the one-factor method, Placket-Burman factorial designing and Central composite design (CCD) of Response surface methodology (RSM). The most relevant factors affecting the EPS yield were sucrose, yeast extract and pH for E.villorum SB2 and sucrose, yeast extract and magnesium sulfate for the E.rivorum S22-3 as determined by Placket-Burman design, whose concentrations were further optimized using CCD. The optimized fermentation conditions gave the total EPS of 9.76 g/l (4 times the initial production) from E.villorum SB-2 and 7.74 g/l (2.5 times the initial production) from E.rivorum S22-3, respectively, after 36-hour incubation at 37 °C. These optimization studies might be helpful during scale-up process for the industrial scale production of these exopolysaccharide.