Organic solvent enhanced spectrofluorimetric method for determination of laccase activity

摘要

A novel fluorimetric method for determination of laccase activity in organic solvent is proposed. It is based on the oxidation of o-phenylene diamine (1,2-diaminobenzene, OPDA) catalyzed by laccase during a fixed time period yielding the fluorescent product, 2,3-diaminophenazine, which was determined and enhanced fluorimetrically in organic media such as ethanol, 1,4-dioxane or acetone at lambda(ex)/lambda(em)=464 nm/530 nm by a fixed time method. The optimal conditions for laccase determination in an organic medium are 55 degrees C, pH 6.5, 1.0x10(-2) mol l(-1) OPDA, 1.25 mi of ethanol, 1.25 mi of 1,4-dioxane and 1.25 mi of acetone. The linear ranges of the method proposed in ethanol, 1,4-dioxane and acetone media were 0.4-19.3 U, 0.1-20.8 U, 0.4-21.0 U with detection limit (signal:background=3) of 0.09, 0.02 and 0.08 U, respectively. The proposed method has been applied to the determination of laccase activity in real samples with more accuracy and sensitivity than the previous method reported. (C) 1998 Published by Elsevier Science B.V. All rights reserved. [References: 9]