Use of ‘dilute-and-shoot’ liquid chromatography-high resolution mass spectrometry in preclinical research: application to a DMPK study of perhexiline in mouse plasma.

By Esposito, Simone; Bracacel, Elena; Nibbio, Martina; Speziale, Roberto; Orsatti, Laura; Veneziano, Maria; Monteagudo, Edith; Bonelli, Fabio
From Journal of Pharmaceutical and Biomedical Analysis (2016), 118, 70-80. Language: English, Database: CAPLUS, DOI:10.1016/j.jpba.2015.10.004

This work describes a simple, sensitive and rapid liq. chromatog.-high resoln. mass spectrometry method for the quantitation of perhexiline and the simultaneous detection of perhexiline metabolites in C57bl/6 mice plasma. Only 5 μL of plasma was used for anal. Pretreatment was limited to a 100-fold diln. (‘dil.-and-shoot’). The analyte was detected by high resoln. mass spectrometry (Orbitrap technol.). Three scan events were performed over the entire chromatogram. Targeted single ion monitoring with data dependent acquisition was employed for perhexiline quantitation and confirmation, while full scan was used to perform untargeted detection of perhexiline phase I and phase II circulating metabolites. The calibration curve was linear (r2 = 0.990) ranging from 0.305 ng/mL (LLOQ) to 10000 ng/mL. Matrix effect was limited to 6.1%. The method was applied to a pharmacokinetic study of perhexiline in mouse plasma and the results obtained were compared to a std. sample prepn. method based on protein pptn. and liq. chromatog.-tandem mass spectrometry (MRM mode) detection. The new approach provided comparable results in terms of pharmacokinetics parameters est. with a high sensitivity, addnl. information on perhexiline circulating metabolites and a low consumption of biol. sample. The combination of the ‘dil.-and-shoot’ approach together with HRMS targeted and untargeted detection represents a suitable alternative to classic bioanal. approaches in preclin. research.