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A sensitive and specific high performance liquid chromatographic assay for milrinone in rat and human plasma using a
Dion R Brocks1, Tara J Spencer, Anooshirvan Shayeganpour
1Faculty of Pharmacy and Pharmaceutical Sciences, University of Alberta, Edmonton, Alberta, Canada. dbrocks@pharmacy.ualberta.ca
Purpose:
To develop an alternate high performance liquid chromatographic method (HPLC) for the analysis of the positive inotropic agent, milrinone, in rat and human plasma.
Methods:
To plasma samples (0.1 mL), containing milrinone and the commercially available internal standard, amrinone, were added 0.15 mL of water and 0.35 mL of acetonitrile. Tubes were briefly vortex-mixed and centrifuged. The supernatant was transferred to clean tubes and 3 mL of methanol: diethyl ether (5:95) was added. The tubes were vortex mixed, centrifuged, and reconstituted with the mobile phase and injected into the HPLC. Separation was accomplished using a reverse phase chromatography using C18 analytical column, and detection was afforded by monitoring the eluent at an ultraviolet wavelength of 326 nm.
Results:
Standard curves were highly linear over the range 10 to 10000 ng/mL (r2 >0.99). Recovery ranged from 52-69% over a 40-fold range of plasma concentrations from 50 to 2000 ng/mL. Intra- and inter-day coefficient of variation and mean error in were less than 20% at plasma concentrations ranging from 10 to 1000 ng/mL. The utility of the assay was demonstrated in a pharmacokinetic evaluation of milrinone in two rats given intravenous bolus doses.
Conclusion:
The developed assay was sensitive, specific and appropriate for monitoring milrinone in rat or human plasma samples.
