INTRODUCTION: Bile acids are the final products of cholesterol catabolism. They are synthesized in the liver and stored in the gallbladder where they conjugate with either glycine or taurine to create bile salts. Total bile acid levels are a sensitive marker of liver function and they are often used in diagnosis of hepatic, intestinal and biliary diseases. They are also useful in diagosing hepatic diseases in pregnant women. OBJECTIVES: The aim of this study was to determine the concentrations of bile salts in different samples to enable the optimisation of the bile salt concentration so we could apply them in the method of in vitro digestion. STUDY DESIGN: Cross sectional study MATERIALS AND METHODS: We used the Total bile acids 21 FS commercial kit to measure the bile salt concentrations. Absorbance was measured on 10 different samples, the calibrator and the blank sample. They were measured on the spectrophotometer with the wavelenght set to 405 nanometers. The samples included two commercial serums- Unitrol I and Unitrol II and two work serums. Each sample was created in two copies and each copy was measured three times with one minute intervals between measurements. The kit is based on an enzymatic cycling method which is based on the activity of the 3-α-hydroxysteroid dehydrogenase enzyme. RESULTS: Most of the samples follow a decreasing trendline in absorbances depending on time that passed. However, the Unitrol I serum, serum K, the blank and the calibrator defy that trendline. CONCLUSION: A further evaluation of the kit is required due to it being very new to the market. Our research did not provide definite results on the methods lower limit of detection and measured concentrations in the samples.