Hydrogen Peroxide-Induced Deacetylation of Acetyl Resorufin as a Novel Indicator Reaction for Fluorometric Detection of Glucose Using Only Glucose Oxidase

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Hydrogen peroxide(H_2O_2)-induced deacetylation of non-fluorescent acetyl resorufin(1)to fluorescent resorufin(2)as a novel indicator reaction for fluorometric detection of glucose using only glucose oxidase(GOD)is described. When a 1 : 1 : 1 mixture of 1(in CH_3CN), glucose, and GOD(each in pH 7.4 phosphate buffer)was incubated at 25℃ under aerobic conditions, the resulting solution turned yellow to fluorescent pink due to 2. The formation of 2 was markedly retarded on incubation under anaerobic conditions. When a mixture of 1 and H_2O_2 was incubated under aerobic conditions, the formation of 2 was noted as in the case of the enzymatic reaction of 1. These results demonstrated that the observed color change is brought about through deacetylation of 1 to 2 induced by H_2O_2 generated in GOD-catalyzed oxidation of glucose. With regard to the fluorometric traces of the enzymatic reaction with 1(0.2m_M), GOD(0.5mg/ml), and glucose at 25℃, fluorescence intensity exhibited a linear relationship against glucose concentration between 0.2 and 2.0 m_M, with a correlation coefficient of 0.997. Neither ascorbic acid, uric acid, nor bilirubin significantly interfered with the transformation of 1 to 2 through GOD-catalyzed oxidation of glucose.
公益社団法人日本薬学会の論文
2001-03-01