Stereoselective analysis of nadolol in human plasma

A stereoselective method, involving a single liquid-liquid extraction step, was developed and validated for the analysis of nadolol in human plasma. The assay involved extraction of nadolol and desmethyl-nadolol (as internal standard (IS)) from alkalinized plasma into dichloromethane. The organic solvent was separated and evaporated under nitrogen at 40 degrees C. A chiral derivatization scheme with (R)-(-)-napthylethylisocyanate (50 microL of 0.1% solution in dichloromethane for 60 min) was employed to convert the enantiomers of nadolol into the corresponding diastereomeric derivatives. The residue was reconstituted in the mobile phase and injected onto a C-18 column. The mobile phase was a mixture of methanol:tetrahydrofuran:water (52:7:41 by vol) containing about 0.001% v/v of both phosphoric acid and tetramethylethylenediamine. Fluorimetric detection was performed at excitation 230 and emission 330 nm. The assay was specific for the enantiomers of nadolol and the lower limit of quantitation was 2 ng/mL for each of the enantiomers. Analysis of quality control samples resulted in precision estimates of 7% RSD for inter-assay and 10.1% RSD for intra-assay and the predicted concentrations deviated less than 9.4% of the nominal values for the four enantiomers. The extraction recoveries of the individual enantiomer was about 70%. Stability of nadolol enantiomers were established for four freeze/thaw cycle periods and in the autosampler at 5 degrees C for at least 116 h.