Determinação de aciclovir em plasma humano através de precipitação de proteínas por UHPLC-ESI-MS/MS: desenvolvimento, validação e aplicação em estudo de bioequivalência

Abstract

A sensitive ultra performance liquid chromatography-electrospray ionization-tandem mass spectrometry (UHPLC-ESI-MS/MS) method for measurements of acyclovir in plasma was developed and validated. A protein precipitation extraction was proposed for the clean up of plasma and entecavir was added as internal standard (IS). The analyses were carried out using a C18 column and mobile phase of 0.1% formic acid:methanol + 0.1% formic acid in gradient mode of proportion. The mass spectrometer equipped with an electrospray ion source in positive mode, was set up in selective reaction monitoring, to detect precursor → product ion with 226 > 152 m/z and 278 > 152 m/z transitions, for analyte and IS, respectively. The linearity was checked between 4 to 1000 ng/mL for acyclovir, with R2=0.998333. Satisfactory selectivity, linearity, precision, accuracy and showed no matrix effect or carryover were obtained for the UPLC-ESI-MS/MS method. The proposed method was successfully applied to a bioequivalence study of healthy human volunteers.