In vivo determination of ultratrace amounts of prostaglandin in plasma by high-performance liquid chromatography/laser-induced fluorometry/ultrasensitive laser spectrometry under severe conditions.

Abstract

We succeeded in determining ultratrace prostaglandin amounts in plasma, at the femtomolar level, using laser-induced fluorometry through a complete redesign of the analytical procedures. Practical samples, especially plasma, contain large amounts of admixtures, and prostaglandin in plasma (pg/mL) has been considered to be difficult to detect because the samples and reagents supplied by conventional procedures are neither pure nor stable enough to get good results by ultrasensitive laser spectrometry. We completely redesigned the analytical procedures after careful investigations of the reagent purification and the column separation conditions based on a newly found behavior of the reagent and derivatized prostaglandin in a small quantity of ethanol in the mobile phase. A lower determination limit of 23 pg/mL (65 fmol) was achieved, the variance was 12% at 25 pg/mL, and the recovery rate was 88-89%. This method was applied to in vivo analysis of the concentration of prostaglandin E1 administered as a prodrug of prostaglandin E1 (delta(8)-9-O-butyryl prostaglandin F1 butyl ester, AS-013) by intravenous infusion to beagle dogs. A clear correlation between the change of blood pressure and the prostaglandin E1 concentration was confirmed.