A New Method of Measuring Bilirubin in Serum by Vanadic Acid

Abstract

A new method for measuring bilirubin using vanadic acid as oxidizing agent was studied. A solution of a cation surfactant, cetyltrimethylammonium bromide (CTAB) as the reaction accelerator and that of a reducing agent, hydroxylamine as the reaction inhibitor in citrate or tartarate buffer, pH 3 (the first reagent) were used for measuring the concentration of total bilirubin and direct bilirubin, respectively. A solution of vanadic acid (4 mmol/l) containing ethylenediamine tetraacetic acid (EDTA) or hydroxyethan-di-phosphonic acid, pH 7 was used as the second reagent. The bilirubin concentrations were calculated by a two point method measuring absorbances at a specific wavelength before and after oxidation reaction. Under these experimental conditions, the oxidation reaction was accomplished within 2 to 3 minutes.
This new method had fundamental abilities for the determination, such as an appropriate dynamic range, a high reproducibility and the good correlation with the diazo method. Furthermore, the results were affected very little by coexisting substances in serum.
In addition, such liquid type reagents are very convenient without being prepared for every determination and will contribute to the correction of the day-to-day differences and save works in the laboratories.