Abstract: Objective The purpose of this study was to establish a rapid and efficient liquid chromatography tandem mass spectrometry(LC-MS/MS) for simultaneous determination of 15 bile acids in mouse liver tissues. Methods The activated charcoal was utilized to prepare bile acid-free liver, which served as the biological matrix for the preparation of standard and quality control samples. The mouse liver tissue was homogenized, and a basic acetonitrile solution, including 5% NH₄OH was added to precipitate proteins. The proteins were separated on an Agilent Poroshell 120 EC C18 column (100 mm×4.6 mm,2.7 μm) by using ²H₄-DCA, GUDCA-d₅, and LCA-d₄ as internal standards. The mobile phase is ammonium acetate aqueous solution and methanol acetonitrile mixed solution for gradient elution, the column temperature was 30℃, the flow rate was 0.3mL/min, and the injection volume was 2 μL. The electrospray ion source (ESI) was operated in negative ion mode, and in multiple reaction monitoring (MRM). Results The linearity of the 15 bile acids was good with R² greater than 0.993, the limits of determination were less than 2 ng/mL, and the matrix effects were 90.76%-109.25%; the intra-day and inter-day accuracy and precision were less than 15%, and the stability was good under 4℃ for 24 h, repeated freeze-thaw, and freeze-storage for one month, meeting the analytical requirements of biological samples; the detection of mouse liver tissues showed that both unconjugated BAs and conjugated BAs (G-BAs, T-BAs) were dominated by maternal CA, with the highest content of TCA; the concentration of unconjugated BAs was (723.89±50.65) ng/mL, significantly higher than that of G-BAs [(56.90±11.28) ng/mL, P<0.001]; the concentration of T-BAs was (40322.90±14034.80)ng/mL, significantly higher than unconjugated BAs (P＜0.001), and also significantly higher than G-BAs (P＜0.001). Conclusion The LC-MS/MS method we established is sensitive, accurate, reliable, and suitable for the determination of bile acids concentrations in mouse liver tissues, which might help for further studies.

Key words: Liver tissues, Bile acids, Liquid chromatography-tandem mass spectrometry, Mouse