ESTIMATION OF METOPROLOL IN HUMAN PLASMA BY HPLC METHOD

Abstract

Objective: The current project was undertaken to develop and validate a simple method for estimation of metoprolol in human plasma using HPLC. Before implementing the method on human plasma it was developed from scratch and validated, further human plasma was spiked with drug to achieve considerable results.

Methods: From UV absorption studies, λmax of metoprolol was found to be 262 nm. The mobile phase consisted of acetonitrile-potassium phosphate (pH 3.0) buffer in the ratio of 60:40 v/v. Propanolol was used as working internal standard. Standardization of metoprolol was performed on Grace C₁₈ ODS (250 mm X 4.6 mm) reversed-phase column packed with 5 µm spherical ODS packing. Isocratic elution with acetonitrile-potassium phosphate (pH 3.0) buffer (60:40 v/v) was performed at a flow rate of 1.0 ml/min at ambient temperature. Shimadzu SPD 10 AT VP variable wavelength detectors and injection loop of 50 μl were used and metoprolol detection was carried out at 262 nm. The same method was implemented on human plasma spiked with the drug.

Results: Retention time of metoprolol was found to be 4.07±0.02 min and propranolol was found to be 3.98±0.05 min and when the plasma was spiked with the drug retention time of human plasma was 2.20±0.01 min, retention time of internal standard (Propranolol) was 3.89±0.05 min and retention time of metoprolol was 4.14±0.02 min respectively. The method developed was found to comply with standards LOD (Limit of Detection), LOQ (Limit of Quantification), Precision and Accuracy (Intra- and Inter-day variation), Specificity, Selectivity and Recovery/Extraction efficiency. Various validation parameters were analyzed and found to be complying well within the range.

Conclusion: Thus it can be concluded that the method developed is compatible for the estimation of the drug in human plasma and the same can be utilized to estimate the concentrations of the drug in patients on metoprolol therapy.