Abstract: Objective　To establish the glucose oxidase (GOD) method using a new chromogenic substrate N-(2-hydroxy-3-sulfopropyl)-3，5-dimethoxyaniline sodium salt (HDAOS) monohydrate in the detection of glucose in serum.  Methods　　The tetrachlorophenol was replaced by HDAOS, the blue quinone-imine was formed, and it was measured at 600 nm. The GOD method, glucose oxidase-peroxidase-4-aminoantipyrene-phenol (GOD-PAP) and high performance liquid chromatography (HPLC) were used to determine serum hyper lipidemia samples (24 samples), haemolysis samples (24 samples), choloplania samples (24 samples) and normal samples (24 samples), and the results were compared. The methodology evaluation was performed.  Results　There was no statistical significance among the 3 methods, when the normal serum group was measured (P＞0.05). Statistical significance only existed between the GOD method and GOD-PAP, when the serum groups of hyperlipidemia，hemolysis and choloplania were measured (P＜0.05), and there was no statistical significance between the GOD method and HPLC (P＞0.05). The between-run and within-run coefficients of variation (CV) were<0.3%. The GOD method had a good correlation with HPLC (r²=0.995 8), and got to the reaction end point within 5 min. The linear range was 0.20-28.00 mmol/L. When hemoglobin <2.0 g/L, chyle<2.2% and total bilirubin <200 μmol/L, the interference error for the GOD method was <3%. The reference values were 4.10-6.20 mmol/L for males and 4.04-6.15 mmol/L for females，and the maximal absorbing wavelength was 595 nm.  Conclusions　Replacing tetrachlorophenol with HDAOS, the blue quinone-imine is formed. The GOD method eliminates the interference of hyperlipidemia，haemolysis and choloplania through limiting their absorption spectra. The GOD method has a good measurement accuracy in the detection of glucose and performs the same reaction end point.

Key words: Glucose, Glucose oxidase method, N-(2-hydroxy-3-sulfopropyl)-3, 5-dimethoxyaniline sodium salt