Separation and analysis of delta 9-tetrahydrocannabinol in biological fluids by high-pressure liquid chromatography...

High-pressure liquid chromatographic (HPLC) systems were developed to separate quantitatively delta 9-tetrahydrocannabinol from heptane-extractable lipoidal and other endogenous substances in biological fluids. These substances interfered with the quantitation by flame-ionization GLC of the unmodified compound and by electron-capture GLC of the pentafluorobenzoyl derivative. Reversed-phase HPLC elution, with 47% acetonitrile in water, and normal-phase HPLC with 25% chloroform in heptane separated delta 9-tetrahydrocannabinol from 11-hydroxy-delta 9-tetrahydrocannabinol and other monohydroxylated tetrahydrocannabinols. These systems also purified stock solutions of delta 9-tetrahydrocannabinol from accompanying contaminants. The various monohydroxylated tetrahydrocannabinols were resolved from each other in the normal phase, 80% chloroform in heptane. The delta 8- and delta 9-tetrahydrocannabinols were separable in the normal phase with 5% tetrahydrofuran in hexane. The GLC analysis of pentafluorobenzoylated delta 9-tetrahydrocannabinol had a sensitivity of 1 ng/ml of plasma, with an estimated 5% standard error with the developed extraction and GLC procedures. Radiochemical analysis of the HPLC-separated fraction had a sensitivity of 0.2 ng/ml of plasma, with an estimated 2% standard error. There was no significant difference between the liquid scintillation and electron-capture GLC assays of the HPLC-separated delta 9-tetrahydrocannabinol obtained from the plasma of dogs administered the drug. Radiolabeled compounds can be added to plasma samples as internal standards to determine the recovery efficiencies of the several procedures in the analysis of unlabeled tetrahydrocannabinol.