Assay of flecainide in serum by high-performance liquid chromatography after microscale protein precipitation

Flecainide is a new drug that is effective for the treatment of ventricular arrhythmias. Optimal therapeutic response is obtained with serum levels maintained in the 200-1000 ng/ml range. To measure flecainide in this concentration range, a high-performance liquid chromatography procedure was devised using the rapid and convenient microscale protein precipitation procedure described previously by Lam et al. to prepare the specimen for chromatography. The drug in the supernatant was injected into a Nucleosil-5,C18 reversed phase column and eluted in 5 min with a mobile phase containing 300 ml of acetonitrile, 1 ml of phosphoric acid, and 0.5 ml of diethylamine in 1 L of distilled water at a flow rate of 2 ml/min. Flecainide was detected with a Perkin Elmer 650-10 LC fluorescence spectrophotometer at 370 nm upon excitation at 285 nm. It was resolved from the endogenous compounds found in serum and from the common cardiac drugs, which eluted close to the solvent front. Peak height was proportional to flecainide levels from 120 to 1200 ng/ml. A detection limit of 30 ng/ml with signal-to-noise level better than 5 is achieved. A coefficient of variance of less than 5.0% is obtained without an internal standard.