Simultaneous determination of tacrine and 1-hydroxy-, 2-hydroxy-, and 4-hydroxytacrine in human plasma by high-performance liquid chromatography with fluorescence detection.

An analytical method was developed for the simultaneous reversed-phase (cyano) high-performance liquid chromatographic determination with fluorescence detection of tacrine and 1-hydroxy-, 2-hydroxy-, and 4-hydroxytacrine in human plasma. Alkalinized human plasma samples were extracted with a mixture of chloroform/l-propanol (90:10, v/v). Extraction recoveries generally ranged from 68% to 83% for all four compounds and did not appear to be concentration dependent over the range examined. Calibration curves were constructed over the following ranges: 0.5-30.0 ng/mL for tacrine (THA) and 4-hydroxytacrine (4-OH-THA), 1.0-30.0 ng/mL for 2-hydroxytacrine (2-OH-THA), and 0.925-46.2 ng/mL for 1-hydroxytacrine (1-OH-THA). The intra- and interassay precision (RSD) for the quality control specimens analyzed during validation were < or = 11.8% and < or = 9.28%, respectively. The intra- and interassay accuracy (RE) for the quality control specimens analyzed during validation ranged from 14.1% to -7.5% and 12.1% to -3.33%, respectively, for all four compounds. The limit of quantitation was estimated as the level of the lowest concentration in the calibration curve for each compound based on acceptable interassay precision and accuracy statistics generated at this concentration. The sensitivity of the method was adequate for the determination of THA, 1-OH-THA, 2-OH-THA, and 4-OH-THA following oral administration of Cognex (40 mg single dose) to normal volunteers.