Bile Acid Preparation and Analysis

Ultrapure water was obtained using a Milli-Q^® Integral 3 purification system equipped with an EDS-Pak^® Endfilter for the removal of endocrine active substances (Merck Millipore, Burlington, MA, USA). Acetonitrile (HPLC-S Grade) was purchased from Biosolve (Dieuze, France), methanol (CHROMASOLV™ LC-MS grade) from Honeywell (Charlotte, NC, USA), isopropanol (EMSURE^® for analysis) from Merck Millipore, and formic acid (Puriss. p.a. ≥98%) from Sigma-Aldrich (St. Louis, MO, USA). Bile acids and internal standards were purchased from Sigma-Aldrich or Steraloids (Newport, RI, USA) as described recently [28 (link)].