Abstract:

Objective Tremella fuciformis polysaccharides’ (TFPs) high molecular weight, high viscosity and other characteristics limit its biological activity and application. The purpose of this study is to screen strains with high yield of TFPs enzyme and optimize its fermentation conditions, so as to achieve the mass production of enzyme, thus effectively reduce the molecular weights of TFPs by enzymatic degradation method, and expand its application potential. Method Based on the characteristics of microbial enzymatic decomposition of TFPs, strains with TFPs as the sole carbon source were isolated from decomposed Tremella fuciformis samples. Combined the determination of TFPs degradation rate with 3,5-dinitrosalicylic acid (DNS) enzyme activity, strains with high yield of TFPs enzyme were screened. Single factor experiments and response surface methodology were used to optimize the fermentation medium (carbon source, nitrogen source, inorganic salts, etc.) and fermentation conditions (temperature, pH, rotation speed, etc.) of high-yield TFPs enzyme strains, and the catalytic characteristics (optimal pH, temperature, and degradation efficiency) of the TFPs enzyme were analyzed. Result The 40 strains isolated from the rotten T. fuciformis samples could use TFPs as sole carbon source, and 10 strains of them had high ability to degrade TFPs. The strain Y3522 with the highest enzyme activity was identified as Mesobacillus by 16S rRNA gene sequencing. The optimized medium components of Mesobacillus sp. Y3522 were as follows: TFPs (molecular weight of 1 250 kD) 8.07 g/L, casein peptone 25.47 g/L, K₂HPO₄ 7.11 g/LNaCl 2.0 g/L,MgSO₄·7H₂O 1.0 g/L. The optimal fermentation conditions were a temperature of 35 ℃, pH of 7.5, inoculation volume of 3%, rotation speed of 250 r/min, and fermentation time of 18-24 h. After optimization, the enzyme activity increased by 60.1%. The optimal catalytic conditions for TFPs enzyme derived from Mesobacillus sp. Y3522 were pH 7.5 and temperature 35 ℃. In addition, 20% (V/V) crude enzyme solution degraded TFPs with a molecular weight of 3 000 kD (0.5%, m/V) to 922, 308, 85, and 18 kD at 30, 60, 90, and 120 min, respectively, demonstrating efficient molecular weight regulation ability. Conclusion The strain Mesobacillus sp. Y3522 producing high yield of TFPS are isolated and identified, its enzyme production fermentation system is optimized. The produced enzyme demonstrates efficient degradation ability on the molecular weights of TFPs.

Key words: Tremella fuciformis polysaccharides enzyme, isolation and identification, Mesobacillus, optimization of fermentation conditions, response surface experiment, molecular weight