A validated bioanalytical method in mouse, rat, rabbit and human plasma for the quantification of one of the steroid glycosides found in Hoodia gordonii extract

Abstract

Hoodia gordonii extract contains steroid glycosides, fatty acids, plant sterols and polar organic material. Certain steroid glycosides show appetite suppressant activities following oral ingestion. This study describes the validation of a bioanalytical method for the quantification of one of the steroid glycosides, H.g.-12 (∼10% (w/w) of the extract), in mouse, rat, rabbit and human plasma. The method utilises a liquid–liquid extraction with methyl-tert-butyl ether followed by chromatographic separation on a 2.1×50mm C18 Genesis high performance liquid chromatography (HPLC) column and detection on a triple quadrupole mass spectrometer. Detection of H.g.-12 and its stable isotope internal standards is performed using positive TurboIonspray™ ionisation in multiple reaction monitoring mode. The validation procedure demonstrated assay sensitivity, linearity, accuracy, precision and selectivity over the calibration range of 0.5–150ng/mL in human plasma (500μL sample volume), 1.0–100ng/mL in rat and rabbit plasma (150μL sample volume) and 1.0–250ng/mL in mouse plasma (150μL sample volume) with good recoveries (≥77%). H.g.-12 was stable in plasma for ≥6months at −20°C, for up to 4h at ambient temperature (ca22°C) and after 3 freeze–thaw cycles. Plasma extracts were stable for up to 24h at ambient temperature.