A Simple Sample Preparation with HPLC-UV Method for Estimation of Amlodipine from Plasma: Application to Bioequivalence Study

Abstract

A simple, rapid and selective method was developed for the estimation of amlodipine from human plasma. The method involves a simple protein precipitation techniques using nifedipine as internal standard. Chromatographic separa- tion was carried out on a reverse phase C18 column using mixture of 50 mM potassium di hydrogen ortho phosphate (pH 7.5) and acetonitrile (60:40, v/v) at a flow rate of 1.0 mL/min with UV detection at 239 nm. The retention time of am- lodipine and internal standard were 4.12 and 8.31min, respectively. The method was validated and found to be linear in the range of 0.5-50.0 ng/mL. An open, randomized, two-treatment, two period, single dose crossover, bioequivalence study in 24 fasting, healthy, male, volunteers was conducted. After dosing, serial blood samples were collected for the pe- riod of 168.0 h. Various pharmacokinetic parameters including AUC0-t, AUC0-�, Cmax, Tmax, T1/2, and elimination rate constant (Kel) were determined from plasma concentration of both formulations of test (Amlodipine 5 mg tablets) and ref- erence (Amlodipine 5 mg tablets). Log transformed values were compared by analysis of variance (ANOVA) followed by classical 90% confidence interval for Cmax, AUC0-t and AUC0-� and was found to be within the range. These results indi- cated that the analytical method was linear, precise and accurate. Test and reference formulation were found to be bioe- quivalent.