Preparative Biochemistry & Biotechnology最新文献

Chikungunya virus (CHIKV) is a mosquito-borne pathogen responsible for recurring outbreaks, highlighting the need for rapid and accurate diagnostic tools. The E2 glycoprotein of CHIKV is a promising target for antibody-based detection. This study aims to determine the optimal expression conditions of recombinant single-chain variable fragment (scFv) fused with biotin acceptor domain (BAD) specific to the CHIKV E2 glycoprotein in Escherichia coli Origami B (DE3). The optimization was performed using the Box-Behnken design of response surface methodology (RSM), with the IPTG inducer concentration, induction time, and induction temperature as the independent variables. The optimal conditions were identified as 0.2 mM IPTG, 2 hours induction at 37 °C, resulting in a total protein concentration of 0.658 mg/mL. The soluble fraction of scFv-BAD was successfully purified using Ni²⁺-NTA affinity chromatography, with a purity of 91.11%. ELISA confirmed that the recombinant scFv-BAD was biotinylated and retained its ability to bind the CHIKV E2 antigen. The optimized scFv-BAD construct demonstrates potential for use in various immunoassay platforms, including rapid diagnostic tests for CHIKV detection.

Bioactive peptides derived from natural sources have attracted significant attention for their antioxidative and anticancer potential. This study explores the potential of antioxidative peptides from Solanum lycopersicum var. Win Win, a new Malaysian tomato hybrid and their correlation with anticancer activity. Peptides were extracted using ultrasound-assisted extraction under optimized conditions: 4 mins extraction time, 57% amplitude, and 60% ethanol concentration, yielding 4.50 ± 0.11 mg/mL. The antioxidant activity of the peptide extracts was validated using DPPH and FRAP assays. Peptide identification through LC-MS/MS and PEAKS revealed 570 peptides. Identified peptides underwent in silico screening to refine lists for anticancer and antioxidant potential. This process also evaluated peptides that are lacking hemolytic, allergenic, and cytotoxic properties. Top five antioxidative peptides underwent molecular docking to evaluate the interaction with VEGFR2, a key target in cancer cells proliferation. Among them, YGYMC and RWYHNQ demonstrated better docking orientations with VEGFR2, suggesting strong binding affinity. These findings highlight the potential of tomato-derived peptides in cancer therapy development. Further in vitro validation is recommended to confirm their clinical applicability and efficacy.

Phycobiliproteins (PBPs) are one of the prominent pigments produced by Cyanobacteria, and phycocyanin (PC) is a major pigment. Rhodophytes, cryptophytes, and cyanobacteria all contain the blue pigment CPC, which has antioxidative and fluorescent qualities. The PC production is mainly done through open ponds, which increases the risk of contamination. The factors influencing the production of PC including the various strains, light intensity, photoperiod, nutrient sources and modes of production (photoautotrophic, mixotrophic, and heterotrophic modes) is critically discussed. The productivity and safety of PC production have improved under photobioreactors and a heterotrophic production system. It has been demonstrated that altering the genetic and metabolic structure of algae strains can boost PC output and enhance its quality. This paper highlights specific methods for extracting PC, such as freeze-thaw methods, high-pressure homogenizations, ionic liquids, supercritical fluids, pulse electric fields, and ultrasound-assisted extraction. Mainly review articles focus on production stracegies of cyanobacterial biomass and less review has been published production approach of CPC with their extraction extraction etsrategies. Additionally, this review article contains information on current insights concerning the source and production of CPC, along with in-depth details regarding the genetic regulation of CPC biosynthesis and its techno-economic feasibility and sustainability.

Introduction: Plant-derived polyphenols are increasingly sought as multi-target options against oxidative stress, metabolic disorders and cancer.

Methods: An ultrasound-assisted water extract of Sideritis montana subsp. montana was chemically profiled and tested for antioxidant, enzyme-inhibitory, and cytotoxic activities.

Results: The extract contained 30.08 mg GAEs/g phenolics and 22.18 mg/RE/g flavonoids; LC-ESI-MS/MS identified chlorogenic acid as the dominant metabolite (4120 µg/g). Antioxidant tests gave EC₅₀ values of 1.59 mg/mL (phosphomolybdenum), 2.03 mg/mL (CUPRAC) and 0.72 mg/mL (FRAP); radical-scavenging IC₅₀ values were 4.63 mg/mL (DPPH) and 2.80 mg/mL (ABTS). Enzyme assays showed inhibition of tyrosinase (IC₅₀ 1.92 mg/mL), BChE (3.39 mg/mL) and AChE (4.36 mg/mL), with weaker effects on α-amylase and α-glucosidase. In A549 lung-carcinoma cells the extract lowered viability to about 50% between 250 and 500 µg/mL, and exposure at this midpoint raised TNF-α from 3.39 to 18.23 ng/mL (∼5.4-fold) and TGF-β from 12.71 to 58.18 pg/mL (∼4.6-fold).

Conclusions: Overall, the hydroxycinnamate-rich matrix exhibits moderate redox, enzyme-modulating and cytostatic activities, suggesting potential in cosmetic depigmentation or adjunct neuroprotection. Future fraction-guided isolation, broader cell panels and in vivo studies are needed to pinpoint active constituents, clarify cytokine implications and assess translational value.

In this study, thermophilic, spore-forming bacteria from the families Bacillaceae and Anoxybacillaceae were analyzed for their ability to produce silver nanoparticles (AgNPs). Anoxybacillus sp. D401a was selected due to its high AgNP synthesis potential. The culture conditions (NaCl-free nutrient broth, 60 °C, pH 8.5, 48 hr) and synthesis parameters (1.5 mM AgNO₃, 48 hr at 60 °C, pH 8.5) were optimized, resulting in a sevenfold increase in AgNP yield. Scale-up production yielded 139.4 mg of dry AgNPs with strong antimicrobial activity that exhibited significant minimum inhibitory concentration (MIC) values against Gram-positive, Gram-negative and fungal pathogens, outperforming commercial AgNPs. Sub-MIC concentrations of the biosynthesized AgNPs also inhibited biofilm formation and quorum sensing in Chromobacterium violaceum ATCC 12472. Characterization by ultraviolet-visible spectroscopy revealed a surface plasmon resonance peak at 400-450 nm, and Fourier-transform infrared spectroscopy indicated the presence of organic residues that stabilized the particles. Transmission electron microscopy analysis showed predominantly spherical AgNPs (24-57 nm). MTT assays showed a dose-dependent cytotoxicity against human keratinocytes (HaCaT), fibroblasts (HDF, L929), and cancer cells (HT-29, MCF-7), indicating a therapeutic window compared to commercial AgNPs. These results emphasize the potential of Anoxybacillus-derived AgNPs as a sustainable alternative for biomedical applications.

Black glutinous rice bran (BGRB) serves as a plentiful natural pigment source. However, limited research has been conducted on the extraction, purification, and characterization of the anthocyanins present in BGRB. This work aims to provide an eco-friendly process for industrial anthocyanin extraction and purification. Following green chemistry principles, safe solvents and recyclable purification materials were used in the experiment. And waste production and energy consumption were limited. Optimal extraction conditions: ethanol-water ratio 40:60 (v/v, pH 2.5), solvent-feed ratio 80:1 (mL/g), 50 °C, and 40 min yielded 19.79 ± 0.14 mg/g anthocyanin, and 32 anthocyanins were identified in the BGRB anthocyanin extract. Optimal purification conditions: employing a D101/AB-8 mixed resin (7:3 w/w), the volume, pH value, and flow rate of BGRB anthocyanin extract were 1.5 Bed Volumes (BV), pH 3.0, and 0.5 BV/hr, respectively; desorption was performed successively with 40:60 (v/v, pH 2.5) ethanol-water of 6 BV, 0.5 BV/hr. Purification achieved 65.97% anthocyanin recovery. After purification, total anthocyanins (67.70 ± 5.43 mg/g to 171.27 ± 7.99 mg/g) and color value (33.48 ± 2.91 to 84.58 ± 1.41) were both 2.5 times higher than in the extract. The metabolite expression of purified anthocyanins showed 4.2 times increase by liquid chromatography-mass spectrometry (LC-MS).

Microbial infestation related food loss poses a major threat to the global food sector. Both bacterial and fungal infestations play a crucial role in the food deterioration and various health issues. This has led to the increased demand for the development of active packaging materials. In the current study, selected essential oils were subjected to MIC, MBC and MFC analysis against both bacterial and fungal agents. Further to this, PVA based bionanocomposites were developed by incorporating the selected essential oils along with ZnONPs. By the FTIR analysis, the interactions among individual components of the developed thin films could be confirmed. Interestingly, the incorporation of nanoparticles was found to have modulatory effect on the release of essential oil components from the films as evidenced by the GC-MS analysis. Further to this, the films were also demonstrated to have enhanced mechanical properties and low moisture content which favor its application as promising packaging material. The incorporation of ZnONPs was also observed to positively modulate the antibacterial and antifungal activity of the films without compromising its microbial barrier efficacy. The findings of the current study thus indicate the potential of the developed bionanocomposite films as antimicrobial packaging systems.

Accurate detection of bioethanol is essential for optimizing biofuel production. Although precise, current industrial methods such as Gas chromatography (GC) and High-performance liquid chromatography (HPLC) are expensive, time-consuming, and unsuitable for on-site monitoring. This study explores the development of a cost-effective, rapid electrochemical method using nickel oxide (NiO)-modified glassy carbon electrodes for detecting bioethanol produced via fermentation using Saccharomyces cerevisiae. Fermentation was optimized with varying sucrose concentrations (20-100 g/L), achieving a maximum ethanol yield of 15.69 g/L at 40 g/L sucrose with an agitation of 100 rpm at 48 h. Distilled ethanol was characterized by GC and HPLC. Cyclic voltammetry (CV) was employed for electrochemical detection, with NiO nanoparticles synthesized and characterized using X-ray diffraction (XRD), confirming their crystalline structure and electroactive surface area. The CV analysis demonstrated clear anodic and cathodic peaks for standard and fermented bioethanol, with a cathodic peak at 0.43 V and an anodic peak at 0.52 V. The oxidation peak current showed a linear relationship with the scan rate, confirming a diffusion-controlled process. These findings confirm NiO-modified electrodes as a dependable ethanol detection method. As a scalable and sustainable substitute for traditional methods, the technique exhibits potential for non-enzymatic bioethanol monitoring in industrial settings.

Lignin peroxidase (LiP) is a haem-containing microbial ligninolytic enzyme that has gained much attention for its potential application to degrade industrial effluents. In the current study, seven fungi (SLF1-7) showing ligninolytic activity were isolated from Similipal forest soil, and screened for LiP activity. SLF3 showed the highest LiP activity (3.351 U/mL.min) at un-optimized conditions and was identified as P. chrysogenum using 18S rRNA sequencing. The production of enzyme was optimized using Response Surface Methodology. LiP activity of P. chrysogenum was enhanced to 9.38 U.mL^-1.min (2.79 folds) at pH 6, temperature 37.5 °C, substrate concentration 1.75%, and time 108 h. The LiP enzyme was purified by ammonium sulfate precipitation followed by dialysis and Sephadex G-100 column chromatography. The purified protein showed a single band of approximately 40 kDa by SDS-PAGE analysis. The purified LiP also showed the highest catalytic efficiency with K_m of 0.316 M and V_max of 147.059 U/mL.min. The purified LiP reduced brilliant green color (69.53 ± 0.003%) after 80 min of incubation at 37 °C and pH 6.0. It is apparent from the study that the strain P. chrysogenum has significant potential in degrading brilliant green, which can be explored for bioremediation applications of industrial dyes.

With numerous advantages over conventional techniques, ultrasound-assisted extraction (UAE) has become a viable method for the effective extraction of biomolecules from prokaryotic and eukaryotic cells. The fundamentals and workings of UAE are examined in this review, focusing on current developments, including how these impact the extraction of proteins, lipids, enzymes, and other bioactive compounds. UAE not only enhances cell disruption and mass transfer, leading to improved extraction yields, but also preserves the integrity of the extracted bioactive molecules under optimized conditions, making it a preferred choice in Biochemistry and Biotechnology. Additionally, this review explores recent innovative approaches that combine ultrasound with other techniques like enzymatic digestion, supercritical CO₂, deep eutectic solvents, and Three-Phase Partitioning (UA-TPP) etc, to further enhance extraction efficiency. The differences in extraction effectiveness between prokaryotic and eukaryotic cells are attributed to cellular structure and ultrasonic conditions. Overall, this review highlights UAE's promise as a viable and efficient substitute for biomolecule extraction concerning prokaryotic and eukaryotic cells while bringing up areas that need additional research and development.