Quantitative analysis of 3alpha,6alpha,24-trihydroxy-24,24-di(trifluoromethyl)-5beta-cholane, a potent synthetic steroidal liver X receptor agonist in plasma using liquid chromatography-tandem mass spectrometry.

The steroidal liver X receptor agonist, 3alpha,6alpha,24-trihydroxy-24,24-di(trifluoromethyl)-5beta-cholane (ATI-829) is a potential therapeutic agent for the treatment of atherosclerosis. A sensitive and selective liquid chromatography-tandem mass spectrometry (LC-MS-MS) method for the quantification of ATI-829 in mouse plasma was developed and validated. Proteins in a 25 microL aliquot of mouse plasma were precipitated, and ATI-829 was extracted from the precipitate by the addition of 125 microL methanol. The overall extraction efficiency was greater than 99%. LC-MS-MS with negative ion electrospray and selected reaction monitoring was used for the quantitative analysis of ATI-829. The lower limit of quantitation of ATI-829 corresponded to 5.0 ng/mL (9.7 nM) plasma. Interference from matrix was negligible. The calibration curve was linear over the range 5-2000 ng/mL. The intra-day precision and inter-day precision of the analyses were <4.5% and <6%, respectively, and the accuracy ranged from 92% to 103%. ATI-829 in plasma was stable for at least 6 h at room temperature, 1 week at 4 degrees C, and 3 weeks at -20 degrees C. The validated method was then utilized for pharmacokinetic studies of ATI-829 administered to mice.