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

Abstract

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.

Keywords:

• Bacillus gottheilii
• M2S2
• central composite design
• packed bed reactor
• Partial purification
• polyurethane foam matrix
• tannase

Acknowledgments

The authors are grateful to the Department of Biotechnology, Manipal Institute of Technology, Manipal Academy of Higher Education, India; for providing the facilities to carry out the research work. The authors would also like to show gratitude to Mr. Kannan Natarajan (Assistant Professor-Senior, Manipal Institute of Technology, Manipal) and Dr. Prasanna D. Belur (Assistant Professor, National Institute of Technology, Surathkal, Karnataka) for sharing their insight and expertise with us during the course of this research.