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Determination of Phosphodiesterase Inhibitors Tadalafil, Roflumilast and Roflumilast N-Oxide Using LC-MS in Guinea Pig Plasma.
Journal of Chromatographic Science 2018 November 2
Phosphodiesterases are known as a super-family of 11 isoenzymes, which can exert various functions based on their organ distribution. In this work, a rapid and sensitive ultra-high performance liquid chromatography-tandem mass spectrometry method was developed for quantification of tadalafil (phosphodiesterase five inhibitor), roflumilast (RF) (phosphodiesterase four inhibitor) and its active metabolite, RF N-oxide in guinea pig plasma. Chromatographic separation was carried out on UPLC BEH C18 column (2.1 mm × 50 mm, 1.7 μm) at a flow rate 0.5 mL/min, using 0.2% formic acid in acetonitrile and 0.2% formic acid in water as mobile phases within 4 min. Detection was performed using a triple quadrupole mass spectrometer employing electrospray ionization operated in positive mode using multiple reaction monitoring mode. The method utilized deuterium labeled internal standards, and was validated according to European Medicines Agency guidelines. It showed excellent linearity in the range of 0.5-500.0 ng/mL for all analytes with coefficient of determination >0.99. The intra- and inter-day precisions (relative standard deviation %) were within 6.7%, and the recoveries were greater than 73.4%. Using this method, plasma samples from experiments of phosphodiesterase four, and five inhibitors in a model of ovalbumin-induced allergic inflammation in guinea pigs were analyzed.
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