Production optimization of a heat-tolerant alkaline pectinase from Bacillus subtilis ZGL14 and its purification and characterization

Alkaline pectinase has important applications in the pretreatment of waste water from food processing and in both the fabric and paper industries. In this study, a 2-level factorial design was used to screen significant factors that affect the activity of alkaline pectinase, and the response surface methodology (RSM) with a Box-Behnken design (BBD) was used to optimize their concentrations. Starch, peptone, KH₂PO₄ and K₂HPO₄·3H₂O were found to significantly affect the activity of alkaline pectinase. Their optimal concentrations were as follows: 4.68% starch, 1.6% peptone, 0.26% KH₂PO₄ and 0.68% K₂HPO₄·3H₂O. Under the above conditions, the activity of alkaline pectinase was significantly improved to 734.11 U/mL. Alkaline pectinase was purified to homogeneity with a recovery rate of 9.6% and a specific activity of 52372.52 U/mg. Its optimal temperature and pH were 50°C and 8.6, respectively. The purified enzyme showed strong thermo-stability and good alkali resistance. In addition, the activity of alkaline pectinase was improved in the presence of Mg²⁺. Cu²⁺, Mn²⁺, and Co²⁺ significantly inhibited its activity. This study provides an important basis for the future development and use of a heat-tolerant alkaline pectinase from B. subtilis ZGL14.