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

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 × 50 mm 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 0.5–150 ng/mL in human plasma (500 μL sample volume), 1.0–100 ng/mL in rat and rabbit plasma (150 μL sample volume) and 1.0–250 ng/mL in mouse plasma (150 μL sample volume) with good recoveries (≥77%). H.g.-12 was stable in plasma for ≥6 months at −20 °C, for up to 4 h at ambient temperature (ca 22 °C) and after 3 freeze–thaw cycles. Plasma extracts were stable for up to 24 h at ambient temperature.