A sensitive and selective LC-differential mobility-mass spectrometric analysis of allopregnanolone and pregnanolone in human plasma.

ABSTRACT: A sensitive and selective method was developed to quantitate allopregnanolone and its 5? isomer pregnanolone in human plasma using liquid chromatography-differential mobility separation combined with MS/MS detection. The method employed a simple liquid-liquid extraction of 100 ?L plasma with hexane/ethyl acetate. After extraction, the sample was derivatized using a quaternary aminooxy reagent. Separation of allopregnanolone, pregnanolone, and their 3? epimers (epiallopregnanolone and epipregnanolone) was achieved using a Phenomenex Kinetex C18 2.1?×?100-mm 2.6-?m column. A linear calibration curve was obtained over the concentration range from 10 to 25,000 pg/mL, and the inter- and intra-day accuracy of the quality control samples were between 90 and 110 % with the inter- and intra-day precision less than 10 %. The lower limit of quantitation is 50 fg (157 amol) on column for both allopregnanolone and pregnanolone which is 100-fold less than the underivatized compounds. The recovery is above 95 %, and the extracted samples are stable for at least 6 days when stored at 4 °C. Plasma samples from normal, pregnant, and postpartum women were analyzed using this method.

SUBMITTER: Jin W 

PROVIDER: S-EPMC3984615 | biostudies-other | 2013 Nov

REPOSITORIES: biostudies-other