Determination of prostaglandin E 1 in plasma with picogram per millilitre sensitivity by double antibody extraction and column switching high-performance liquid chromatography

Abstract

Prostaglandin E 1 (PGE 1) is a vaso- and arteriodilator that is used for the experimental treatment of a variety of vascular diseases. A high-performance liquid chromatography (HPLC) method was developed for the purpose of quantifying PGE 1 in the plasma of patients undergoing constant infusion therapy. Plasma (0.5 ml) was extracted with a double antibody precipitation technique and the PGE 1-immunoprecipitate was isolated and washed by repeated resuspension and centrifugation. PGE 1 was recovered from the precipitate by extraction with acetonitrile and derivatized with panacyl bromide for determination on a heteromodel column switching HPLC system with fluorescence detection. The content of PGE 1 in immunoextracts was determined by HPLC peak height comparison against a standard curve of PGE 1 peak height vs amount derivatized. The result was corrected for the plasma extraction efficiency (determined with radiolabelled PGE 1) to obtain the plasma concentration. Standard curves were linear from 25 to 400 pg ( r > 0.99) and the y-intercepts were not significantly different from zero ( p < 0.05). The immunoextraction recovery from six human donors was 63.5 ± 2.0% (S.D., n = 18). The quantification limit of the method was 50 pg ml −1 (signal-to-noise ratio 3:1), at which the estimated assay relative standard deviation was 18%. The utility of the method for the measurement of PGE 1 plasma levels during constant intravenous infusion was demonstrated in a dog study.