Bile acids serve as biomarkers for liver function and are indicators for cholestatic and hepatobiliary diseases like hepatitis, cirrhosis, and intrahepatic cholestasis of pregnancy (ICP). Sulfation and renal excretion of bile acids are important elimination steps. The power of ultra high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) allows specific profiling of primary and secondary bile acids as well as their sulfated counterparts.
Twenty-four sulfated and non-sulfated primary and secondary bile acids were quantified in urine with 15 corresponding stable isotope labeled internal standards by using two-dimensional UHPLC-MS/MS. The sample preparation was based on a simple dilution with a methanolic zinc sulfate solution followed by an automated online solid phase extraction clean up.
The validation results of the method fulfilled the criteria of the European Medicine Agency (EMA) “Guideline on bioanalytical method validation”. To verify fitness for purpose, 40 urine samples were analyzed which showed an average of 86% sulfation, 9.1% taurine-conjugation, 14% non-conjugation, and 77% glycine-conjugation rates.
Lossless one-pot sample preparation, automated sample purification, and high number of internal standards are major innovations of the presented profiling method, which may allow diagnostic application of BA profiling in the future.
Research funding: None declared.
Author contributions: All authors have accepted responsibility for the entire content of this manuscript and approved its submission.
Competing interests: Authors state no conflict of interest.
Informed consent: Informed consent was obtained from all individuals included in this study.
Ethical approval: The local Institutional Review Board deemed the study exempt from review.
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The online version of this article offers supplementary material (https://doi.org/10.1515/cclm-2021-1111).
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