Production of Gallic Acid from Swietenia macrophylla Using Tannase from Bacillus Gottheilii M2S2 in Semi-Solid State Fermentation

In this research, an industrially important enzyme tannase and product gallic acid was produced with an inexpensive novel substrate Swietenia macrophylla. Fermentation of S. macrophylla was optimized using a two-step approach: First, the traditional One variable at-a-time technique, and second, the statistical Box-Behnken design for co-production of tannase enzyme and gallic acid. This two-step method of optimization showed the highest tannase activity and gallic acid yield of 0.0497 U/mL and 225 µg/mL respectively which is a 29.5 and 49-fold increase when compared to unoptimized conditions. Further, the partially purified tannase enzyme was characterized and showed optimal tannase activity at pH 4.0 and 30 ℃, and was stable between pH 3.0–6.0 and 4−40 ℃ for 24 h and 10 h, respectively. Also, metal ions such as Ca²⁺, Na⁺, and K⁺ at 1 mM concentration; and organic solvents methanol, and isoamyl alcohol at 20% v/v exhibited the highest activity at optimized reaction conditions. Whereas, Mn²⁺, Zn²⁺, Mg²⁺, Fe²⁺ and Fe³⁺, EDTA, TritonX 100, toluene, and hexane caused the tannase inhibition at higher concentrations. In the end, the fermentative production of gallic acid was verified qualitatively through thin-layer chromatography and Fourier transform infrared spectroscopy. Graphical Abstract: [Figure not available: see fulltext.]