A VALIDATED REVERSED PHASE HPLC ASSAY FOR THE DETERMINATION OF OMEPRAZOLE IN HUMAN PLASMA

Rajaa F. Hussein, Nada H. Binhashim, Syed N. Alvi and Muhammad M. Hammami*

ABSTRACT

A simple and precise reversed-phase high performance liquid chromatographic (HPLC) assay for the measurement of omeprazole level in human plasma, using lansoprazole as an internal standard (IS), was developed and validated. Plasma samples containing omeprazole were spiked with the IS then extracted with tert. butyl methyl ether and reconstituted in mobile phase. The compounds of interest were efficiently separated on Atlantis dC18 column at room temperature and detected with photodiode array detector set at 302 nm. The mobile phase consisted of 0.05 M dibasic sodium phosphate (pH = 7.0, adjusted with phosphoric acid) and acetonitrile (60:40, v:v) and was delivered at a flow rate of 1.0 ml/min. The relationship between omeprazole concentration in plasma and peak height ratio of omeprazole/ IS was linear (R2  0.9992) in the range of 0.01 – 1.20 μg/ml and the intra- and inter- day coefficient of variations (CV) and accuracy were ≤5.0% and ≤10.1%, ≥90% and ≥92%, respectively. Mean extraction recovery of omeprazole and the IS from plasma samples both was 91% and 91%, respectively. In processed samples, omeprazole was stable for at least 24 hours at room temperature (≥91%) and 48 hours at -20 º C (≥89%). In unprocessed samples it was stable at least 12 weeks at -20ºC (≥96%) and after 3 cycles of freeze and thaw (≥96%). Further, it was successfully applied to measure omeprazole level in samples obtained from a healthy volunteer.

Keywords: Omeprazole, Lansoprazole, Human plasma, HPLC.