Rapid high-performance liquid chromatography–tandem mass spectrometry method for determination of pentoxifylline and its active metabolites M1 and M5 in human plasma and its application in bioavailability study

A new rapid, sensitive and selective liquid chromatography coupled with mass spectrometry method was developed and validated for the simultaneous quantification of pentoxifylline (PTX) and two major active metabolites in human plasma (M1 and M5). After a deproteinization step, chromatographic separation of the selected analytes was performed on a RP-C18 column. The detection of target compounds was in multiple reaction monitoring mode using an ion trap mass spectrometer equipped with an electrospray ion source. The method was validated and proved to be linear, accurate and precise over the range 5.08–406.14, 10.08–806.40 and 20.15–1611.60 ng/mL in case of PTX, M1 and M5, respectively. The major advantages of this method are the small sample volume, simple sample processing technique, the high sensitivity and the very good selectivity guaranteed by the MS/MS (in case of PTX) or MS/MS/MS (in case of M1 and M5) detection. The validated method has been successfully applied to a bioequivalence study.