Effect of protein on the determination of total bile acids in serum

Abstract

We measured total serum bile acids on a fluorescence-light-scattering micro centrifugal analyzer by the direct enzymatic method with 3α-hydroxysteroid dehydrogenase (EC 1.1.1.50) and with resazurin as a fluorogenic electron acceptor. We found that serum protein has an inhibitory effect on the measurement of bile acids, but this effect was eliminated by adding bovine serum albumin to the reaction mixture in a final protein concentration (12.2 g/L) that was high compared with that contributed by a normal serum specimen. The assay is a sensitive method that reaches equilibrium in 5 min. The method is microscale (5 μL of sample, 150 μL of working reagent), is easy to perform, and is accurate (analytical recovery = 104.1%) and precise (CV = 11.1 and 5.7% on specimens with bile acid concentrations of 7.6 and 35.4 μol/L, respectively). Normal values are 1-12 and <9 μol/L on nonfasting and fasting individuals, respectively. Pure 3α-hydroxysteroid dehydrogenase must be used: we found several enzyme preparations that gave falsely high values for bile acid.