Determination of nitroglycerin and its dinitrate metabolites in human plasma by high-performance liquid chromatography with thermal energy analyzer detection

Abstract

A highly selective and sensitive high-performance liquid chromatographic assay employing a thermal energy analyzer as the detector for nitroglycerin and its dinitrate metabolites in human plasma has been developed. Prior to chromatography the method employs a simple one-stage extraction step. Nitroglycerin and its dinitrate metabolites are then chromatographed on a 10-micron nitrile bonded phase column using an internal-external standard method. The nitroglycerin and its 1,2,3-propanetriol-1,3- and -1,2-dinitrate metabolites (glyceryl-1,3- and -1,2-dinitrate) have a retention time of 8.5, 10.5, and 11.5 min, respectively at a flow rate of 2.0 mL/min for a mobile phase of 5% v/v acetone in n-hexane. The limits of sensitivity were 0.05 ng/mL for nitroglycerin and 0.25 ng/mL for the dinitrate metabolites. Linearity of response was observed over the 0.1-2.0-ng/mL range for nitroglycerin and 0.5-10.0-ng/mL range for the dinitrate metabolites. Blood level data from a pilot study with human volunteers in receipt of an oral form of nitroglycerin is presented.