DETERMINATION OF EBROTIDINE AND ITS METABOLITES IN HUMAN URINE BY REVERSED-PHASE ION-PAIR HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY

Ebrotidine is a new H-2-receptor antagonist with powerful antisecretor y activity, demonstrated gastroprotection and the ability to inhibit p rotease and lipase activities of Helicobacter pylori. As a tool in the clinical pharmacokinetic study of ebrotidine, an analytical method fo r the simultaneous determination of ebrotidine an its metabolites in h uman urine was developed. An ion-pair reversed-phase HPLC separation u sing 1-hexanesulfonic acid and acetonitrile as mobile phase with gradi ent elution was optimized. In addition, several procedures of preconce ntration and clean-up were tested, including solid-phase and liquid-li quid extraction, the mixture dichloromethane-2-propanol (9:1, v/v) at pH 11 being the most efficient. The quality parameters of the whole an alytical method were established, the calibration curves were linear o ver the range studied (1-200 mu g/ml) and the reproducibility of the m ethod was high (inter-day R.S.D. values lower than 4.4%). The limits o f detection were between 26 and 110 ng/ml of urine for ebrotidine and its metabolites. The method was applied to the analysis of urine colle cted from two volunteers during 96 h following oral administration of ebrotidine at a dose of 400 mg.