关键词: 三聚氰胺脱氨酶, 酶活性测定, 分光光度法

Abstract: A method to determine melamine deaminase activity by catalytic spectrophotometry was developed based on Berthelot colour reaction. The melamine deaminase activity was calculated by the amount of ammonia released per minute. The reaction conditions for melamine deaminase were optimized by one-factor-at-a-time experiments combined with an orthogonal experiment design L9(34). Optimum coloration reaction condition was obtained when the reaction was allowed to proceed for 40 min in a system composed of 0.3 mL of sodium hypochlorite and 1.2 mL of salicylic acid solution and detection at a wavelength of 650 nm. The optimal enzymatic reaction conditions was found to be 0.2 mL melamine solution, 60 μL crude enzyme and 40 min reaction time. Under the optimized conditions, the melamine deaminase activity was 132.41 U/L with inter-assay and intra-assay coefficients of variation of 0.81%－1.19% and 1.27%－1.69%, respectively.

Key words: melamine deaminase, enzyme activity determination, catalytic spectropotometry