Solid state fermentation of Bacillus gottheilii M2S2 in laboratory-scale packed bed reactor for tannase production

Production of tannase was performed in packed bed reactor filled with an inert support polyurethane foam (PUF) using Bacillus gottheilii M2S2. The influence of process parameters such as fermentation time (24-72 h), tannic acid concentration (0.5-2.5% w/v), inoculum size (7-12% v/v), and aeration rate (0-0.2 L/min) on tannase production with PUF were analyzed using one variable at a time (OVAT) approach. The outcome of OVAT was optimized by central composite design. Based on the statistical investigation, the proposed mathematical model recommends 1% (w/v) of tannic acid, 10% (v/v) of inoculum size and 0.13 L/min of aeration rate for maximum production (76.57 ± 1.25 U/L). The crude enzyme was purified using ammonium sulfate salt precipitation method followed by dialysis. The biochemical properties of partially purified tannase were analyzed and found the optimum pH (4.0), temperature (40 °C) for activity, and K_m (1.077 mM) and V_max (1.11 µM/min) with methyl gallate as a substrate. Based on the SDS-PAGE analysis, tannase exhibited two bands with molecular weights of 57.5 and 42.3 kDa. Briefly, the partially purified tannase showed 4.2 fold increase (63 ± 1.60 U/L) in comparison to the submerged fermentation and the production of tannase was validated by using NMR spectrometer.