The analysis of bile acids in human plasma is an important diagnostic tool for the detection of liver disease and can also be used as indicators of potentially harmful side effects of new drugs. There are two main types of bile acids based upon their functional groups: free (or unconjugated) bile acids and conjugated bile acids, primarily glycine- or taurine-bound. Quantitation of bile acids in matrix can be very challenging due to a number of factors. These include structural similarities, varying polarity and stereochemistry, the presence of isomers, limited fragmentation of unconjugated bile acids in a mass spectrometer, high endogenous levels, and matrix effects caused by phospholipids or triglycerides.
Three column chemistries were explored in this study for the analysis of 13 bile acids in plasma in order to resolve a matrix interference. Biphenyl, FluoroPhenyl, and ARC-18 stationary phases were tested on a 100 x 2.1 mm, 2.7 μm column dimension. The ARC-18 stationary phase showed good selectivity for the target analytes. This phase can also resolve all three isomers sets with adequate resolution, and successfully separated the matrix interference from UDCA-D4, which is necessary for accurate quantitation of these analytes.
