Combination of simultaneous extraction-hydrolysis and intermittent feeding of tara pod for efficient production of gallic acid

The pods of Caesalpinia spinosa consists of gallotannins, which represent more than 50 % of its weight. The traditional chemical procedure used to produce gallic acid (GA) from tara gallotannins is a high-pollution pro-cess. Although biological procedures, including microbial fermentation and enzymatic hydrolysis, are environ-mentally friendly, insufficient production efficiency significantly hinders industrial application. In this study, we proposed a novel strategy for the simultaneous extraction and enzymatic hydrolysis of tara gallotannins with intermittent feeding. A recombinant thermostable tannase (Tan1) derived from Aspergillus oryzae was first constructed to adapt to the high-temperature simultaneous extraction and enzymatic hydrolysis process. Tan1 showed competent activity yield (83.11 U/mL) and excellent heat resistance, exhibiting the highest activity at 80 degrees C and half-lives at 60 degrees C for more than 6 h. The production efficiencies of GA using different enzymatic hydrolysis strategies were compared. The simultaneous extraction and enzymatic hydrolysis strategy combined with intermittent feeding minimized the inhibition of tara gallotannins on Tan1 activity and resulted in an increased GA yield. Using this strategy, 192 g/L tara powder (intermittent feed for 6 times) was treated with Tan1 at 60 degrees C for 177 min, resulting in 78.5 g/L GA output which was the highest among the currently reported values obtained using biological protocols for GA production.