Hydrogen peroxide-induced deacetylation of acetyl resorufin as a novel indicator reaction for fluorometric detection of glucose using only glucose oxidase

Hydrogen peroxide (H2O2)-induced deacetylation of non-fluorescent acetal re sorufin (1) to fluorescent resorufin (2) as a novel indicator reaction for fluorometric detection of glucose using only glucose oxidase (GOD) is descr ibed. When a 1 : 1 : 1 mixture of 1 (in CH3CN), glucose, and GOD (each in p H 7.4 phosphate buffer) was incubated at 25 degreesC under aerobic conditio ns, the resulting solution turned yellow to fluorescent pink due to 2. The formation of 2 was markedly retarded on incubation under anaerobic conditio ns. When a mixture of 1 and H2O2 was incubated under aerobic conditions, th e formation of 2 was noted as in the case of the enzymatic reaction of 1. T hese results demonstrated that the observed color change is brought about t hrough deacetylation of 1 to 2 induced by H2O2 generated in GOD-catalyzed o xidation of glucose. With regard to the fluorometric traces of the enzymati c reaction with 1 (0.2 mM), GOD (0.5 mg/ml), and glucose at 25 degreesC, fl uorescence intensity exhibited a linear relationship against glucose concen tration between 0.2 and 2.0 mM, with a correlation coefficient of 0.997. Ne ither ascorbic acid, uric acid, nor bilirubin significantly interfered with the transformation of 1 to 2 through GOD-catalyzed oxidation of glucose.