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  • A new continuous spectrophotometric assay method for DOPA oxidase activity of tyrosinase.

A new continuous spectrophotometric assay method for DOPA oxidase activity of tyrosinase.

Journal of protein chemistry (2003-12-24)
Yong-Doo Park, Jae-Rin Lee, Kyung-Hee Park, Hwa-Sun Hahn, Myong-Joon Hahn, Jun-Mo Yang
ABSTRACT

Sensitive assay methods for tyrosinase are essential not only for the understanding the process of pigment production but also for the development of effective inhibitors of tyrosinase. To develop an efficient assay method, we applied thymol blue to reaction mixtures. The enzyme kinetic study revealed that DOPA oxidase activity of tyrosinase in thymol blue-applied reaction system was more sensitively measured, even under lower enzyme units compared with the previous report with significant enhancement of Vmax while affinity change on substrate was not observed. To test whether this method could be applicable to the inhibition and the inactivation kinetic study of tyrosinase, the effect of kojic acid, a well-known tyrosinase inhibitor, and sodium chloride respectively, have been studied. Conclusively, thymol blue method can assay tyrosinase activity with sensitivity and is applicable to the inhibition and the inactivation study of tyrosinase.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Thymol Blue, ACS reagent
Sigma-Aldrich
Thymol Blue sodium salt, ACS reagent, Dye content 95 %
Supelco
Thymolphthalein, ACS reagent, indicator (pH 8.8-10.5)
Sigma-Aldrich
Thymolphthalein, ACS reagent, Dye content 95 %
Sigma-Aldrich
Thymol Blue, JIS special grade
Sigma-Aldrich
Thymolphthalein, JIS special grade