Screening Of Tannase-producing Strains And Optimization Of Culture Conditions

Tanin acyl hydrolase(E.C.3.11.20) commonly referred to as tannase, Tannase catalyzes the hydrolysis of ester and depside bonds in hydrolysable tannins such as tannic acid ,releasing glucose and gallic acid . It produced mainly by fungi., at present Penicillium and Aspergillus niger were studied more than others. In this paper, we carried out a series of experiments, including screening, identifying optimizing fermentation process, and studying characteristics of tannase. The main results are as follows:A tannase producing strain Aspergillus niger A-6 was obtained from soil by assaying hydrolyzed circle on the selective medium and tannase activity under sol1id-state fermentation, the yield of tannase was 8.1U/g.By single-factor experiments and response surface methodology, the culture medium and conditions of the strain A-6 outputing tannase were optimized. The optimum culture composition were as follows: wheat bran 5g and 5mL nutrition solution was taken in 250mL conical flasks,the nutrition solution containing(g/L):sucrose 12, KNO₃ 2.5, corn steep liquor 22.4, gallnut 65.1 , MgSO₄·7H₂O 1.36, CaCl 2·2H₂O 0.2, Sodium Citrate 3.0, NaCl 2.5; the optimal culture conditions were as follows: inoculation 1mL spore suspension(11×10⁹个/mL), temperature 30℃, initial pH value 7.0, solid-liquid ratio1:2 and culture time 108h. Under these optimal conditions, the yield of tannase was 15.81U/g, increased for about 0.94 times more than that of before.Studies on characterization of tannase showed: The crude tannase from the strain A-6 showed well stability at 4℃, the enzyme retained 94% of maximal activity at 4℃after 3 mouths; tannase activity was optimal at 60℃, pH 8.0, and in 1.5mmol/L NaCl solution. The enzyme was stable below 65℃and pH6.0-9.0; The tannase activity was strongly activtied by K⁺, Mn²⁺,, Fe³⁺; and inhibited by Zn2+, Ca²⁺and Fe²⁺.