Identification and Optimization of Enzymes Extracted from Solid-State Fermention

Abstract

Microbial enzymes produced through solid-state fermentation are essential source of numerous microbial strains due to their higher stability, production rate, biochemical versatility, and availability. These enzymes, especially the thermophilic ones, are capable of withstanding harsh environments, high temperatures, and chemicals used in various industrial processes. This study involved multiple steps. Firstly, bacteria were identified using 16S rRNA gene analysis. Subsequently, solid-state fermented (SSF) enzymes, including amylase, protease, cellulase, and xylanase, were screened using their respective assays. Prior to optimization, the conditions affecting the extraction were evaluated using a one-factor-at-a-time (OFAT) approach. The extraction of SSF enzymes was then optimized using a Face Centered Central Composite Design (FCCCD). Bacterial enzyme identification was performed using Next-generation sequencing, and the following strain families were found: Enterobacteriaceae, Klebsiella, Aneurinibacillaceae, Atopobiaceae, Bacillaceae, Burkholderiaceae, Clostridiaceae, Lactobacillaceae, Peptostreptococcaceae, Staphylococcaceae, Streptococcaceae, and Streptomycetaceae. The highest protein yield was achieved using the one-factor-at-a-time (OFAT) method, with a protein concentration of 6.07 mg/m obtained from 10g of SSF material (in 90 ml of sodium phosphate buffer) at pH 9. The extraction process involved a temperature of 65°C, a duration of 2 hours, and an incubation speed rotation of 250 rpm. Under these optimized conditions, the activities of the SSF enzymes were determined as follows: protease (11.04 U/ml), cellulase (11.59 U/ml), xylanase (174.13 U/ml), and amylase (11.01 U/ml). This research will further contribute to the extraction of palm oil, offering high yield and a promising solution across various fields by replacing the expensive industrial enzymes.