High-performance liquid chromatographic determination of a 21-aminosteroid antioxidant in plasma

A novel 21-aminosteroid, 21-[4-(2,6-di-1-pyrrolidinyl-4-pyrimidinyl)-1- piperazinyl]-16 alpha-methylpregna-1,4,9(11)-triene-3,20-dione monomethanesulfonate (I), is under development for the treatment of central nervous system injury in humans. This report describes a reversed-phase high-performance liquid chromatographic (RP-HPLC) method using ultraviolet detection at 254 nm for the determination of I in plasma with low nanogram per milliliter sensitivity. Plasma was deproteinated by mixing with acetonitrile and centrifuging, and I was extracted from the supernatant with disposable bonded-phase columns. The extracts were chromatographed on an octylsilane bonded-phase HPLC column with an acetonitrile-water mobile phase containing triethylammonium acetate, pH5. Quantification was accomplished by peak-height ratio analysis using an analogue of I as the assay internal standard. The method was suitable for the determination of I following a 30 mg/kg intraperitoneal dose in the rat.