Determination of the new beta-blocker bisoprolol and of metoprolol, atenolol and propranolol in plasma and urine by high-performance liquid chromatography.

High-performance liquid chromatographic methods using fluorimetric detection have been developed for the determination in plasma and urine of bisoprolol, atenolol, metoprolol and propranolol. Bisoprolol, metoprolol and propranolol were extracted at alkaline pH with dichloromethane, atenolol with n-butanol-dichloromethane (25:75). After evaporation of the organic solvent the compounds were chromatographed on silica gel Si-60 columns (normal phase) using aqueous ammonium phosphate buffer (pH 4) containing 3-7% acetonitrile as eluent (method 1). Alternatively, the compounds were acetylated prior to chromatography on reversed-phase columns (RP-8), using acetonitrile-water mixtures as eluents (method 2). The detection limit was 1-2 ng/ml in plasma and 10 ng/ml in urine for bisoprolol and metoprolol with either method. For atenolol the detection limit was 5 ng/ml in plasma or 50 ng/ml in urine (method 1), for propranolol 1 ng/ml in plasma (method 2).