Production and Optimization of γ-PGA from Bacillus subtilis Using Rice Husk as a Cheap Biomass

Abstract

Aims: We optimized poly γ-glutamic acid (γ-PGA) production from Bacillus subtilis isolate utilizing rice husk in solid-state fermentation (SSF). The effects of various factors, including medium components and process conditions, on γ-PGA yield, with a focus on enhancing production efficiency were assessed. Study Design: The study employed response surface methods to design experiments, incorporating a definitive screening design (DSD) and a full central composite design (FCCD). These designs allowed for the evaluation and optimization of significant factors affecting γ-PGA production, considering a range of variables such as temperature, citric acid concentration, glycerol concentration, and 6-diazo-5-oxo-L-norluecine (DON) concentration. Place and Duration of Study: The study was conducted in the laboratory setting over a specified duration, utilizing resources and facilities conducive to microbial fermentation experiments. The exact location and timeframe of the study are as per the laboratory facilities and research schedule. Methodology: Solid-state fermentation (SSF) was employed for γ-PGA production using Bacillus subtilis and rice husk as the substrate. Response surface methods, including DSD and FCCD, were utilized to design experiments and optimize production conditions. The impact of factors such as temperature, citric acid concentration, glycerol concentration, and DON concentration on γ-PGA yield was assessed. Confirmation experiments were conducted to validate the model, with analysis techniques including amino acid analysis, FT-IR spectroscopy, and SDS-PAGE for molecular weight determination. Results: The results indicated that temperature, citric acid concentration, glycerol concentration, and DON concentration significantly influenced γ-PGA yield. Optimum conditions for γ-PGA production were determined as follows: DON concentration of 25.0 μg/100g, citric acid concentration of 5.68% (w/w), and glycerol concentration of 9.50% (v/w). Confirmation experiments confirmed the validity of the model, with a correlation coefficient of 0.9554. Analysis techniques revealed distinctive peaks of glutamic acid in amino acid analysis, characteristic bands in FT-IR spectra, and molecular weights of 35, 44, and 96 kDa respectively in SDS-PAGE. Conclusion: This study underscores the potential of Bacillus subtilis for γ-PGA production using rice husk as a substrate via solid-state fermentation. Furthermore, it highlights the inhibitory effect of DON on γ-glutamyl transpeptidase (γ-GTP), leading to enhanced γ-PGA yield. These findings contribute to the optimization of γ-PGA production processes and offer insights into the utilization of agricultural by-products for biopolymer synthesis, with implications for various industrial applications.