BIOANALYTICAL METHOD VALIDATION FOR THE DETERMINATION OF ENALAPRIL, ENALAPRILAT AND HYDROCHLOROTHIAZIDE IN HUMAN SERUM BY LC/MS/MS DETECTION

Enalapril and Enalaprilat and Hydrochlorothiazide were extracted from an aliquot of human serum using solid phase extraction method, and then injected into a liquid chromatograph, equipped with mass spectrometry detector. Internal standard method was used for quantitation of Enalapril, Enalaprilat and Hydrochlorothiazide. Linear regression with 1/X 2 weighting was performed to determine the concentration of the drug from serum .Hydroflumethiazide was used as internal standard. A common solid phase extraction procedure for the isolation of drug and its metabolite was developed from serum samples. The samples were analysed on API 3200 Triple quadrapole mass spectrometer using Chromolith, RP-18e column in atmospheric pressure electrospray ionization. The mobile phase composition was a isocratic mixture of 0.01% Ammonia in water : acetonitrile (30:70 %v/v). The method was validated over a linear range of 0.50 – 500 ng/ml and the limit of quantification were 0.50 ng/ml. Recoveries were observed above 70% for all the three analytes. The storage stability of Quality control samples was investigated under various conditions.