Abstract
A method was developed for the isolation of prostacyclin (PGI 2) from whole blood in a fraction suitable for high pressure liquid chromatography (HPLC) separation of PGI 2 and 6-keto-prostaglandin F 1α (6-K-PGF 1α). Prostacyclin was stabilized in whole blood by rapidly raising the pH to 10 with Na 2CO 3 and cooling the samples to 0°C. Under these conditions, 2.9% hydrolysis was observed after 20 min. Reverse phase extraction columns were used to directly extract both PGI 2 and 6-K-PGF 1α from the alkaline plasma with recoveries of greater than 95% using an acetonitrile/2mM Na 2B 4O 7, pH 10, 40 60 elution solvent mixture. An additional 1.7% hydrolysis was found during the column extraction procedure. Final separation of PGI 2 and 6-K-PGF 1α was performed with HPLC using an alkaline solvent system. This method is capable of rapidly and efficiently extracting and separating PGI 2 and 6-K-PGF 1α from whole blood or plasma. It introduces less than 5% hydrolysis of PGI 2, thus providing a means of applying highly sensitive 6-K-PGF 1α assays to the determination of PGI 2 levels in physiological samples.
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