Specific, sensitive and accurate liquid chromatographic–tandem mass spectrometric method for the measurement of ribavirin in rat and monkey plasma

Ribavirin is a purine nucleoside analog with broad spectrum activity against a spectrum of DNA and RNA viruses. To facilitate pharmacokinetics studies, a LC–MS–MS method for the analysis of ribavirin in rat and monkey plasma was developed and validated. The method involved the addition of acyclovir as an internal standard and protein precipitation with acetonitrile followed by separation by an Intertsil Silica column and quantification by a MS–MS equipped with a positive electrospray ionization in the multiple reaction monitoring mode. The MS–MS reaction was selected to monitor the 245→113 and 226→152 transitions for ribavirin and internal standard, respectively. The calibration curve was linear over a concentration range of 10–5000 ng/ml. The lower limit of quantitation was 10 ng/ml, the coefficient of variation (CV) was 8–11%, and the bias was 1–3%. Intra-day and inter-day analysis of QC samples at 30, 1500 and 3500 ng/ml indicate that the method was precise (CV<18%) and accurate (bias<13%). Ribavirin in rat and monkey plasma was stable at 5 °C for at least 24 h, 0 °C for at least 4 h, and after three freeze–thaw cycles. This specific, accurate and precise assay is useful in the study of the pharmacokinetics of this compound.