An Ultra-Sensitive and Selective LC-UV Method for the Simultaneous Determination of Metformin, Pioglitazone, Glibenclamide and Glimepride in API, Pharmaceutical Formulations and Human Serum

摘要

An effective and comprehensive method for the simultaneous quantification of 4 NIDDM drugs (metformin, glimepride, glibenclamide and pioglitazone) was achieved on a Purospher Start C18 (5 μm, 25×0.46 cm) and Supelco C18 column in 2, 3, 7, 9 min respectively. The optimized method involves a C18 column thermostated at 30°C, UV detection at 235 nm, at a flow rate of 1 mL min-1. Good separation of the analytes was achieved by gradient high performance liquid chromatography-UV/visible detector (HPLC-UV/visible) in API, pharmaceutical dosages and serum, mobile phase was a mixture of methanol: water (70:30v/v) the pH of which was adjusted to 3.0 by phosphoric acid.The method exhibited consistent, high-quality recoveries of the four analytes which ranged from 93.8 ± 2.1 to 99.8 ± 1.5 (mean ± RSD) with a high precision for the drug and impurities. Linear regression analysis revealed an excellent correlation between peak responses and concentrations (R2 values of 0.9991–0.9999) for the drug and impurities. Validation under Food and Drug Administration (FDA) guideline of the analytical parameters include: linearity (r2>0.9996), LLODs (0.315, 2.3, 0.2,0.1 ng ml-1), LLOQs (0.95, 0.7, 0.59,0.32 ng-1), intra-day precision (0.001) and inter-day precision 0.9 expressed as relative standard deviation (R.S.D.) and robustness parameters (less than 1.98%) with accuracies between 98% and 102%. The plasma assay was validated for parameters such as specificity, accuracy and extraction recovery. This is the first simultaneous characterization and quantitative determination of multiple NIDDMS. Thus, this method provides a simple, sensitive, selective, accurate and precise assay for the determination of all compounds in active pharmaceutical preparations, dosage formulations and human serum with high percentage of recovery, good accuracy precision (no interference of excepients) and a short run time. The proposed method can be extended for routine analysis of anti-diabetics in pharmaceutical preparations, biological matrices, and clinical laboratories with standard equipment, drug interaction studies and forensic medicine, recoveries ranging from 94 to 99%.