Abstract
An automatic flow-injection stopped-flow kinetic spectrophotometric method for determination of laccase activity is proposed. It is based on the oxidation of o-phenylenediamine (1,2-diaminobenzene) catalyzed by laccase during the stopped-flow period, yielding 2,3-diaminophenazine, which is determined spectrophotometrically. The substrate solution (o-phenylenediamine) and Na2HPO4-HAc (pH 6.20) buffer with sample (laccase) are pumped into the flow manifold via two separated channels at the same flow velocity and merged in the flow-through cell, then stopped for measuring variation of absorbance at 420 nm with time at a constant temperature of 55°C. The linear range of the method is over 0-2 U, and the detection limitation is 0.012 U. The proposed method has been applied to the analysis of laccase activity of real samples at 30 samples per hour. © 1996 John Wiley & Sons, Inc.
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