A New Method for Accurate Determination of Peroxidase Activity Based on Fluorescence Decrease of Guaiacol / 分析化学

ABSTRACT

At present the most common colorimetry for the activity determination of the peroxidase ( POD) is based on the detection of the absorbance of product at 470 nm in a reaction system of the H2 O2/POD/guaiacol ( GA) , but the shortcoming of the method is that the formed product is not stable and there is the serious adsorption phenomenon on the cell. To solve this problem, a new method was established for accurate determination of POD activity based on the fluorescent feature of GA. By using standard solutions of horseradish peroxidase as the test samples and under these optimum conditions such as 0. 5 mmol/L of GA, 0. 5 mmol/L of H2 O2 , pH 6, 0. 05 mol/L of phosphate buffer solution and the reaction temperature of 20℃, the sample volume was only consumed 20 microlitre at a time, the linear response range was 500-60000 U/L ( r=0 . 9993 ) , the detection limit was 385 U/L and relative standard deviation was ≤2 . 4% ( n=11 ) . The comparisons for the determination results of the POD activity in the white radish’s extraction solutions were conducted among our method (9714±132 U/L) and the colorimetric method (9926±352 U/L) as well as the recirculating-catalytic flow analysis ( 9608±456 U/L ) . The results showed that the mutual consistency is better.