Simultaneous determination of human neutrophil elastase inhibitor (ONO-5046) and its metabolite in plasma and urine by direct injection column-switching HPLC

The direct injection method for the simultaneous determination of a human neutrophil Elastase Inhibitor (ONO-5046) and its metabolite (ONO-EI-601) in plasma and urine has been developed using a column-switching HPLC system. The system was set up with a pre-treatment column, a pre-concentration column and an analytical column which were connected in series via two automatic switching valves. The calibration lines showed a good linearity for concentrations of ONO-5046 and ONO-EI-601 in a range of 156-20000 ng ml-1 in plasma and 1.56-100 micrograms ml-1 in urine, all correlation coefficients being greater than 0.9999. The limit of quantitation of ONO-5046 was 156 ng ml-1 in plasma, that of ONO-EI-601 was 313 ng ml-1 in plasma, and those of ONO-5046 and ONO-EI-601 were 1.56 micrograms ml-1 in urine. The developed method is rapid and sensitive with automated operation, allowing untreated samples to be analysed every 45 min. The application of the present method to the real plasma and urine samples proved to be useful for pharmacokinetic, toxicological and clinical studies.