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 separation 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 amlodipine 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 period of 168.0 h. Various pharmacokinetic parameters including AUC^0-t, AUC^0-∞, C^max, T^max, T^1/2, and elimination rate constant (K^el) were determined from plasma concentration of both formulations of test (Amlodipine 5 mg tablets) and reference (Amlodipine 5 mg tablets). Log transformed values were compared by analysis of variance (ANOVA) followed by classical 90% confidence interval for C^max, AUC^0-t, AUC^0-∞ and was found to be within the range. These results indicated that the analytical method was linear, precise and accurate. Test and reference formulation were found to be bioequivalent.