Screening and Identification of High Chitinase Producing Strains, Optimization of Fermentation Conditions and Analysis of Enzymatic Properties

To enhance the value-added utilization of chitin derived from crustacean waste, this study collected samples from the bottom of a shrimp farm in Guangxi. High-yield chitinase-producing strains were isolated through preliminary and secondary screening, followed by taxonomic identification. The optimal fermentation conditions were determined using single-factor experiments combined with orthogonal designs. Subsequently, the enzymatic properties and hydrolysis products of the chitinase produced by the selected strain were characterized. Results showed that initial screening on chitin-containing agar plates yielded nine strains capable of forming clear zones, among which strain GXUN-J3 exhibited the highest chitinase activity (297.24±10.77 U/L) and was identified as belonging to the genus Streptomyces sp.. Under optimized fermentation conditions—6% (w/v) chitin, 0.25% (w/v) bean powder, an initial pH of 7.0, a fermentation temperature of 25 ℃, a liquid volume of 40 mL per 150 mL flask, and an inoculum size of 5% (v/v)—the chitinase activity reached 1026.72±18.69 U/L, representing a remarkable increase of 3.51-fold compared to the original process. Enzyme characterization revealed optimal activity at 50 ℃ and pH5.0. Among the metal ions tested, Mg²⁺ and Fe³⁺ significantly enhanced enzyme activity, whereas Fe²⁺, Mn²⁺, and Cu²⁺ exhibited significant inhibitory effects (P<0.05). Additionally, the chemical agents of EDTA, SDS, and methanol markedly suppressed enzymatic activity (P<0.05). Thin-layer chromatography (TLC) analysis demonstrated that the hydrolysis products primarily consisted of chitin dimer and N-acetyl-D-glucosamine, suggesting that this enzyme functions as an exo-chitinase. This study successfully established a process for producing chitinase and preparing chitin oligosaccharide using Streptomyces strain GXUN-J3, offering a promising approach for high-value utilization of crustacean waste.