Highly sensitive determination of saquinavir in biological samples using liquid chromatography–tandem mass spectrometry

A selective and sensitive method for the determination of the HIV protease inhibitor saquinavir in human plasma, saliva, and urine using liquid–liquid extraction and LC–MS–MS has been developed, validated, and applied to samples of a healthy individual. After extraction with ethyl acetate, sample extracts were chromatographed isocratically within 5 min on Kromasil RP-18. The drug was detected with tandem mass spectrometry in the selected reaction monitoring mode using an electrospray ion source and 2H5-saquinavir as internal standard. The limit of quantification was 0.05 ng/mL. The accuracy of the method varied between −1 and +10% (SD within-batch) and the precision ranged from +4 to +10% (SD batch-to-batch). The method is linear at least within 0.05 and 87.6 ng/mL. After a regular oral dose (600 mg) saquinavir concentrations were detectable for 48 h in plasma and were well correlated with saliva concentrations (r2=0.9348, mean saliva/plasma ratio 1:15.1). The method is well suited for low saquinavir concentrations in different matrices.