Hydrophilic interaction liquid chromatography–tandem mass spectrometric approach for simultaneous determination of safingol and D-erythro-sphinganine in human plasma

A simple and specific hydrophilic interaction liquid chromatography-tandem mass spectrometry (HILIC-MS/MS) method was developed for the simultaneous determination of C18-L-threo-sphinganine (safingol, an anti-neoplastic in phase I trials) and its diastereomer, C18-D-erythro-sphinganine (sphinganine), in human plasma. Sample pretreatment involved a protein precipitation with methanol using 25 μL aliquots of plasma. Chromatographic separation of the diastereomers and C17-D-erythro-sphinganine, an internal standard, was achieved on a Xbridge HILIC (3.5 μm, 100 × 2.1 mm) using isocratic elution with the mobile phase of 2 mM ammonium bicarbonate in water (pH 8.3) and acetonitrile at a flow rate of 0.3 mL/min. Electrospray ionization (ESI) mass spectrometry was operated in the positive ion mode with multiple reaction monitoring (MRM). The calibration curves obtained were linear over the concentration range of 0.2–100 ng/mL with a lower limit of quantification of 0.2 ng/mL. The relative standard deviation of intra-day and inter-day precision was below 8.27%, and the accuracy ranged from 92.23 to 110.06%. The extraction recoveries were found to be higher than 93.22% and IS-normalized matrix effect was higher than 90.92%. The analytes were stable for the durations of the stability studies. The validated method was successfully applied to the analyses of pharmacokinetic samples from patients treated with safingol and all-trans-N-(4-hydroxyphenyl)retinamide; (fenretinide, 4-HPR) in a current phase I clinical trial (SPOC-2010-002, ClinicalTrials.gov Identifier: NCT01553071).