Preparative Biochemistry & Biotechnology最新文献

Botrytis cinerea, an airborne plant pathogen, holds the potential to synthesize sesquiterpenes, which have been used for the industrial production of abscisic acid. Previously, through our genetic technology, we obtained strain ZX2, whose main product 1´,4´-trans-ABA-diol is physiologically active in plants. In this study, 50 L of fed fermentation was carried out with ZX2 strain to study the stability of expression of TUA, TUB, ATC, EF-1, GAPDH, UCE and GTP genes. Four kinds of software (GeNorm, NormFinder, BestKeeper and Delta Ct) were used to analyze the expression stability of candidate genes, and finally the best reference gene was screened by RefFinder. Based on the results, the ACT was the most stable gene. It was used to normalize the expression levels of two genes related to 1´,4´-trans-ABA-diol production (hmgr and bcaba3) when fed-batch fermentation. Guide the selection of appropriate internal reference genes during the fermentation process to accurately quantify the relative transcription levels of target genes in B.cinerea ZX2.

Investigating the biotechnological potential of wild microorganisms is paramount for optimizing bioprocesses. Given this premise, we looked for yeasts in Brazilian native stingless bees, considering the recognized potential of pollinating insect-associated microorganisms for the production of volatile organic compounds (VOCs). Two yeast strains of the species Meyerozyma caribbica were isolated from bees Scaptotrigona postica and evaluated for their fermentative capacity. Both yeasts were capable of fermenting sucrose (the main sugar used in the Brazilian ethanol industry) with over 90% efficiency and yields of up to 0.504 g/g. Through an experimental design analysis (CCD), it was verified that the ethanol productivity of these yeasts can also benefit from high concentrations of sucrose and low pH values, desirable traits for microorganisms in this biofuel production. At the same time, CCD analyses also showed the great capacity of these M. caribbica strains to produce another alcohol of broad biotechnological interest, 2-phenylethanol. Interestingly, the statistical analyses demonstrated that greater production of this compound can occur at high sugar concentrations and low availability of nitrogen sources, which can be easily achieved using residual low-cost feedstocks. Thus, our results suggest that these M. caribbica strains may be efficiently used in both ethanol and 2-phenylethanol production.

This study investigated the ultrasound-assisted extraction (UAE) techniques used to enhance the polyphenols and antioxidants of mango peel extract (MPE). Additionally, it explored the bacteriostatic activity of MPE against various microorganisms. The UAE method was optimized using response surface methodology (RSM) at different times, temperatures, and ratios, with optimal conditions found to be 35 minutes, 45 °C, and a 1:35 ratio. The optimized yield results for total polyphenol content (TPC) were 17.33 ± 1.57 mg GAE/g, total flavonoid content (TFC) was 12.14 ± 0.29 mg QE/g, and radical scavenging activity (RSA) was 72.11 ± 2.19%. These response models were extremely significant with p-values less than 0.05. MPE showed selective effectiveness against Bacillus cereus, Geobacillus stearothermophilus, and Escherichia coli (E. coli). The results highlight the potential of mango peel as a sustainable source of bioactive compounds, contributing to waste reduction in the food industry and the development of natural antimicrobial agents. This study contributes to further research on the application of MPE in processed foods.

Traditional Chinese medicine (TCM) is often composed of a variety of natural medicines. Its composition is complex, and many of its components can not be analyzed and identified. The first step in the rational application of TCM is to successfully separate the effective components which is also a great inspiration for the development of new drugs. Among the many separation technologies of TCM, the traditional heating concentration separation technology has high energy consumption and low efficiency. As a new separation technology, membrane separation technology has the characteristics of simple operation, high efficiency, environment-friendly and so on. The separation effect of high molecular weight difference solution is better. The applications of several main membrane separation technologies such as microfiltration, nanofiltration, ultrafiltration and reverse osmosis are reviewed, the methods of restoring membrane flux after membrane fouling are discussed, and their large-scale industrial applications in the future are prospected and summarized.

In the [Bmim]Cl reaction medium, five different acidic ionic liquids were used as catalysts to study the effects of reaction time, reaction temperature, system water content, catalyst dosage, microwave power, and other factors on cellulose hydrolysis under microwave irradiation. The results showed that in the [Bmim]Cl reaction system, using N-methylpyrrolidone methylsulfonic acid salt as a catalyst, controlling the microwave reaction time at 10 min, reaction temperature at 130 °C, catalyst dosage at 1 g/g (cellulose), water addition at 0.756 μL/g ([Bmim]Cl), and microwave power at 480 W, resulted in the best cellulose hydrolysis effect with a glucose yield of 74.49%. Compared to conventional heating, the glucose yield increased by 24% and the hydrolysis time was reduced by 77%. Microwave irradiation significantly enhances the cellulose hydrolysis process in an ionic liquid medium.

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.