Validated high performance liquid chromatographic method for simultaneous determination of phenytoin, phenobarbital and carbamazepine in human serum.

A high performance liquid chromatographic (HPLC) method for the simultaneous determination of phenytoin (CAS 57-41-0), phenobarbital (CAS 50-06-6, phenobarbitone) and carbamazepine (CAS 298-46-4) is described. The serum was extracted with ethyl acetate, the dried extract was reconstituted in mobile phase and the aliquot was injected. The eluent drugs were detected at 230 nm. The mobile phase consisting of methanol: water: glacial acetic acid mixture (67:33:1, v/v/v) was used at a flow rate of 1 ml/min on C-18 column. The absolute recovery was above 96% of all the three drugs over a concentration range of 0.5-50.0 micrograms/ml. The inter-day and intra-day precision relative standard deviation (RSD) ranged from 0.79-5.13% and 0.11-6.81%, respectively. The method is simple, rapid, accurate and sensitive and is presently used for therapeutic drug monitoring in epileptic patients. The results obtained with this method showed very good clinical correlation.