New Validated Method for the Estimation of Metformin HCl and Nateglinide Using RP-HPLC

Abstract

Special, effective high pressure liquid chromatography method has been developed for the simultaneous
quantification of Metformin HCl and Nateglinide. By using Waters HPLC e-2695 quaternary pump with a PDA
detector of 2998 instrument the chromatographic separation of Metformin HCl and Nateglinide was achieved on
the column of Luna Phenyl Hexyl 150X4.6mm, 3.5µ using an isocratic elution with a buffer containing
0.1percent ortho phosphoric acid and acetonitrile at a rate of 70:30 as a mobile phase with a flow rate of 1
ml/min at ambient temperature. A detector wavelength of 221 nm utilizing the PDA detector were given in the
instrumental settings. The linearity was studied between the concentration range of 5-75 µg/ml of Metformin
HCl and 0.6-9 µg/ml of Nateglinide were injected. The plotted calibration curves were linear with a regression
coefficient of R2> 0.999, indicates that the linearity was with in the limit. As a part of method validation the
parameters like specificity, linearity, accuracy, ruggedness, robustness were determined and the results were
found to be within the allowable limit. The method developed was found to be applicable to routine analysis and
to be used for the measurement of both active pharmaceutical ingredients (i.e, Metformin HCl and Nateglinide).
Validation of the proposed method was carried out according to an International Conference on Harmonization
(ICH) guidelines. Since, there is HPLC method reported in the literature for the estimation of Metformin HCl
and Nateglinide, there is a need to develop quantitative methods under different conditions to achieve
improvement in specificity, selecivity etc.