Preparative Biochemistry & Biotechnology最新文献

Poly-γ-glutamic acid (γ-PGA) is an extracellular, biodegradable biopolymer synthesized predominantly by Bacillus spp., with reported production yields ranging from 10-115 g/L depending on the strain, substrate, and fermentation conditions. Recent advancements (2020-2024) in microbial engineering, fed-batch optimization, and low-cost agro-residue substrates have significantly improved γ-PGA productivity, reduced downstream processing costs by 15-30%, and enabled the development of high-purity (>90%) fractions suitable for food, pharmaceutical, and biomedical applications. Unlike earlier reviews that primarily catalog applications, the present work provides a cross-sector comparative analysis integrating (1) strain-specific production efficiency, (2) molecular-weight-dependent functional performance, and (3) quantitative application outcomes such as 2-7-fold enhancement in mineral absorption, 40-60% increase in water retention for agricultural soils, and up to 85% reduction in cytotoxicity in biomedical formulations. This review further assesses emerging technologies, such as membrane-assisted purification, genome-edited Bacillus strains, and smart delivery systems; it critically evaluates regulatory challenges, commercial readiness levels, and market trajectories. This review, through the integration of recent data and industrial trends, identifies the major bottlenecks and opportunities that require further research to place γ-PGA as a scalable, microbially sourced, green polymer with high potential for the next-generation sectors biotechnological-driven.

This study aimed to purify and biochemically characterize lactoperoxidase (LPO) enzymes obtained from Jersey bovine milk and starter culture fermented kefir, and to investigate the inhibitory effects of sulfanilamide and sulfaguanidine. LPO enzymes were purified using ammonium sulfate precipitation followed by hydrophobic interaction chromatography (sepharose 4B-L-tyrosine-1-naphthylamine); milk LPO reached 220.8 ± 5.41 fold purification with 24.08 ± 0.30% yield, whereas kefir LPO achieved 399.2 ± 7.51 fold with 39.91 ± 0.45% yield. SDS-PAGE analysis revealed a single band with an approximate molecular weight of 80 kDa for both enzymes. Kinetic studies showed that kefir LPO exhibited a lower K_m (0.088 ± 0.016 mM) and a higher apparent V_max (294 ± 0.9 EU·mL^-1) than milk LPO (K_m 0.166 ± 0.071 mM; V_max 238 ± 0.7 EU·mL^-1). Inhibition assays demonstrated competitive behavior for both inhibitors; by K_i, sulfaguanidine was more potent than sulfanilamide (milk: 5.79 ± 0.78 vs 12.2 ± 1.23 μM; kefir: 5.08 ± 0.82 vs 25 ± 1.18 μM). Molecular docking reproduced the crystallographic pose (redocking RMSD 0.99 Å), and molecular dynamics (MD) simulations indicated more persistent binding of sulfanilamide at the active site.

Debranching enzymes (DBEs) are responsible for cleavage of the α-1, 6, glycosidic linkage (amylopectin) of polysaccharide (starch) chains and commonly used in the starch industries. This enzymatic action generates the debranched starch, that is, short-chain glucans that are directly linked to resistant starch (RS) known to be beneficial for human gut health, glycemic index (GI) controlling, etc. In view of the demand for healthy food choices by consumers, the food industries are looking for low cost and rapid sensors for detecting the DBE activity and the RS content in their new products development. The Nelson-Somogyi (NS) and 3,5-dinitrosalicylic acid (DNS) assays are traditional methods for assessing starch DBE activity. The investigation of high-throughput nanomaterial-based biosensors have great potential as an alternate to the high-cost chromatographic techniques and useful to the industrial and laboratory-scale usage in the food processing sector. This review explores various nanomaterial-based methods for evaluating starch DBE activity and RS estimation.

Growing consumer demand for natural preservatives has increased interest in bacteriocins such as nisin for food preservation, yet the high cost of downstream processing and purification limits widespread use. This study developed a cost-effective antimicrobial powder by producing nisin directly from fermentation broth without prior purification. Nisin production by Lactococcus lactis subsp. lactis N8 was optimized using Response Surface Methodology (RSM) based on a face-centered Central Composite Design, using demineralized whey (DW), soybean meal (SM), and glucose syrup (GS) as substrates. The regression model explained most variation in antimicrobial activity (R² = 0.98, adj-R² = 0.97, pred-R² = 0.87) and identified the optimum. Maximum activity against Micrococcus luteus NCIMB 86100 reached 9,120 IU/mL at 100 g/L DW, 50 g/L SM, and 50 g/L GS. Spray-drying with different maltodextrin and DW levels was then tested to maximize nisin adsorption. The most effective formulation, using 5% maltodextrin, yielded an antimicrobial powder with an activity of 41,000.50 IU/g, representing approximately a 4.5-fold enhancement compared to the optimized fermentation broth. These results show that active nisin can be incorporated into antimicrobial powders through direct fermentation and spray-drying, offering a scalable, economical clean-label approach for food preservation.

Rhodiola species are an integral part of traditional Chinese medicine, with Rhodiola crenulata (R. crenulata) being the only medicinal species officially listed in the 2020 edition of the Chinese Pharmacopeia. Polysaccharides are recognized as key bioactive components of Rhodiola, yet efficient extraction methods have not been systematically reported. In this study, polysaccharides from R. crenulata were extracted using acidic, neutral, and alkaline deep eutectic solvents (DESs), as well as acidic water, deionized water, and alkaline water. Four solvents with distinct pH values, including 0.01 M acidic water (ACW-1), choline chloride:lactic acid deep eutectic solvent (DES-1), neutral water (NW), and 0.01 M alkaline water (ALW-1), were selected for comparative analysis across four Rhodiola species (R. crenulata, R. wallichiana var., R. kirilowii, and R. himalensis). Total sugar content and monosaccharide composition were quantified. DES-1 as the extraction solvent yielded the highest total sugar content across four Rhodiola, with R. himalensis yielding the highest content (67.03 ± 0.81%). Monosaccharide composition indicated that polysaccharides extracted with DES-1 were mainly composed of glucose (Glc) and arabinose (Ara), whereas those from ACW-1, NW, and ALW-1 were rich in galacturonic acid (GalA) and arabinose (Ara). Additionally, density functional theory (DFT) calculations revealed that DES-1 forms a hydrogen-bond network with polysaccharide hydroxyl groups, enhancing solubility and extraction selectivity. In conclusion, this study provides insights into solvent-mediated polysaccharide extraction and elucidates the extraction mechanism of Rhodiola polysaccharides using DES-1.

Cuttlebone (CFB) is a food waste and primarily composed of calcium carbonate. In this study, the chitosan modified CFB (chitosan@CFB) was prepared by the CFB particle coated with chitosan and then employed as an eco-friendly carrier for trypsin immobilization. The immobilized trypsin (trypsin@chitosan@CFB) was characterized by scanning electron microscopy. The optimal pH of trypsin@chitosan@CFB was 9.0 and the optimal temperature was 37 °C. The trypsin@chitosan@CFB exhibited excellent temperature, pH, storage stability, and reusability. The Km of the trypsin@chitosan@CFB was 0.64 mM when casein used as substrate. The trypsin@chitosan@CFB was used for bovine hemoglobin degradation and the result was analyzed by SDS-PAGE. The trypsin@chitosan@CFB demonstrated preferably potential for industrial applications.

The present research investigated the biological activities of zeaxanthin-enriched extract obtained from the microalgae Dunaliella salina. Pigment extraction was carried out using microwave-assisted extraction (MAE) with five different solvent systems to obtain enriched fractions. The bioactive potential of the pigment was assessed through multiple assays. Antioxidant potential was measured using DPPH radical scavenging method, while antimicrobial efficacy was tested against selected pathogenic strains by resazurin dye. The anticancer potential of zeaxanthin-enriched extract was investigated by evaluating cytotoxicity, reactive oxygen species (ROS) and apoptosis in tumor cell lines includes MDA-MB-231 (breast tumor) and A549 (lung tumor). Zeaxanthin-enriched pigment extract from D. salina (ZPDs) exhibited significant cytotoxicity with IC₅₀ values of 242.077 μg/mL in MDA-MB-231 cells & 160.501 μg/mL for A549 cells. Overall, zeaxanthin-enriched extract from D. salina demonstrated notable antioxidant, antimicrobial and anticancer efficacy, highlighting its potential as a natural bioactive compound from therapeutic use. These results support its potential application in the development of nutraceutical, pharmaceutical and cosmetic products. Further research is recommended to scale up extraction and evaluate industrial feasibility.

The cephalopod processing industry generates large quantities of ink sacs, an underutilized yet valuable source of eumelanin. Conventional extraction of melanin, relying on acid or alkali treatments, often leaves residual proteins and carbohydrates, requires multiple purification steps, and risks pigment integrity. This study investigated a polymer-salt-based aqueous two-phase system (ATPS) as an eco-friendly alternative for the selective extraction and purification of melanin. Screening of polyethylene glycols of varying molecular weights with inorganic salts identified the PEG 6000-K₂HPO₄ system as the most effective. The equilibrium characteristic of the system was assessed using Othmer-Tobias and Bancroft correlations, while the partitioning behavior of melanin was evaluated by considering the process parameters, including polymer and salt concentrations, tie-line length, volume ratio, and sample loading. Under optimal conditions (tie-line length of 53.23% (w/w), a volume ratio of 0.14, and a sample loading of 15% (w/w)), melanin recovery reached 99.1% with 95% purity in the PEG-rich phase. These results outperform conventional methods in both yield and product quality, highlighting ATPS as a sustainable and scalable strategy for valorizing cephalopod waste, with promising applications in food, pharmaceutical, and biomedicine.

Gold nanoparticles (AuNPs) have gained considerable attention in biomedical, environmental, and pharmaceutical applications due to their distinct physiochemical attributes. Plant-mediated green synthesis of AuNPs provides low-cost and pollution-free alternatives to traditional chemical synthesis. Leaf extract of Ruellia patula (Rp) was utilized as bioreduction agent and stabilizer for the synthesis of Rp@AuNPs. Spectroscopic peak at 520 nm affirmed the synthesis of Rp@AuNPs, validated using surface plasmon resonance. XRD patterns confirmed the crystallinity of the resultant nanoparticles, while electron microscopic images analyzed their size distribution and morphology, which ranged from 22.47 to 84.57 nm. Subsequently, FTIR analysis denoted the chemically active biomolecules for nanoparticle reduction and stabilization. Large inhibition zones indicated that biosynthesized Rp@AuNPs exhibited significant antibacterial activity toward Streptococcus pyogenes and Pseudomonas aeruginosa. In addition to demonstrating intense antioxidant and antidiabetic activities, the Rp@AuNPs exhibited anti-inflammatory activity by protein denaturation inhibition and HRBC membrane stability assays. The synthesized Rp@AuNPs ranged from 22.47 to 84.57 nm in size and exhibited antibacterial activity with inhibition zones up to 17 mm, antioxidant activity with DPPH and ABTS scavenging efficiencies of 86.7% and 82.9%, respectively, anti-inflammatory activity with 78.5% protein denaturation inhibition, and antidiabetic activity showing IC₅₀ values of 55.6 µg/mL (α-amylase) and 50.8 µg/mL (α-glucosidase).