Peroxyoxalate Chemiluminescence Assay of Hydrogen Peroxide and Glucose Using 2,4,6,8-Tetrathiomorpholuiopyrimido[5,4-d]-pyrimidine as a Fluorescent Component

Abstract

Peroxyoxalate chemiluminescence (CL) assay of hydrogen peroxide (H2O2) or glucose was developed by using 2,4,6,8-tetrathiomorpholinopyrimido[5,4-d]pyrimidine as a fluorescent component and bis(2,4,6-trichlorophenyl)oxalate (TCPO) as an oxalate. Linear relationships between CL intensity and final concentration of H2O2 from 10-8 to 10-4 M were obtained. The detection limit at the ratio of CL intensities for sample and blank (S/B) of 3 was 10 nM. The precision for five replicate measurements at 10-5 and 10-6 M of H2O2 were 17.6 and 15.7% of relative standard deviations, respectively. α-D-Glucose was transformed to β-D-glucose with mutarotase and converted to H2O2 and D-gluconic acid with glucose oxidase, which was detected by using peroxyoxalate CL reaction. A linear calibration graph was obtained up to 1.5×10-4 M of glucose solution. The method was applied to the assay of glucose in human serum. The recovery was 98.2% (N=4). The method correlated well with the conventional colorimetric method (r=0.968).