Optimization of fermentation conditions for the production of acidophilic β-glucosidase by Trichoderma reesei S12 from mangrove soil

Abstract

Vegetative biomass contains a large number of macromolecular substances such as cellulose and hemicellulose, which can be used by microorganisms to produce biofuels and other chemical byproducts. In this study, a filamentous fungus with high β-glucosidase activity was isolated from mangrove soil in Hainan Province, China. Through morphological identification and internal transcribed spacer (ITS) sequence analysis, the strain was identified as Trichoderma reesei. Fermentation conditions were optimized with the response surface method to improve β-glucosidase activity. Inoculum, pH and liquid volume in flask were found to be the key factors. We further examined the optimal range of the three factors using steepest ascent path, and optimum conditions were further investigated according to a Box-Behnken design. We calculated the optimized fermentation conditions to be: carbon source (microcrystalline cellulose (MCC)) 1%, nitrogen source (yeast extract) 0.5%, pH 3.68, inoculum of 3.22 × 105 cfu/mL, temperature 28 °C, shaking speed of 160 rpm, and liquid volume in flask of 84.74 mL/250 mL. These conditions increased the β-glucosidase activity 6-fold to 1.13 U/mL compared to that before optimization. The resulting β-glucosidase had an optimum pH of 5.0 and an optimum temperature of 45 °C, and showed high thermal stability at 30-60 °C.