Quantification of limonin in human urine using solid-phase extraction by LC–MS/MS

A highly sensitive liquid chromatography-tandem mass spectrometry method was developed and validated for the determination of limonin in human urine using podophyllotoxin as internal standard. The analyte and IS were extracted with solid-phase extraction and separated by a rapid isocratic elution with 1% formic acid/methanol (v:v, 40:60) on an C18 column (150 mm × 2.1 mm I.D.). The detection was performed by mass spectrometry in the multi-reaction-monitoring mode. The precursor to product ion transitions of m/z 471.3 → 161.2 and m/z 397.2 → 313.1 were used to measure the analyte and the IS. The assay was linear over the concentration range of 0.0783–10 ng/mL for limonin in human urine. The lower limit of quantification was 0.0783 ng/mL and the extraction recovery was larger than 76.7% for limonin. The inter- and intra-day precision of the method at three concentrations was less than 7.4%. The method was successfully applied to pharmacokinetic study of limonin in humans.

► This new method has been successfully applied to the evaluation of limonin in human urine after oral administration for the first time.
► The lower limit of quantification (LLOQ) of developed LC–MS/MS method was lower than the previous report.
► The method is simple, sensitive and specific, with a short analysis time (5 min).
► The work presented here used a solid-phase (SPE) extraction.