Development and Validation of Stability Indicating RP-HPLC Method for the Estimation of Glycopyrrolate and Neostigmine in Bulk and Injection.

ABSTRACT

A stability indicating RP-HPLC method is suggested for determination of Glycopyrrolate-Neostigmine (GLY/NEO) in bulk drugs and injection formulation. GLY/NEO were eluted from a Chromolith High Resolution RP-18e (100 mm×4.6 mm) with buffer solution (pH 3.0) as mobile phase A and a mixture of HPLC grade acetonitrile and water mixture (9010) as mobile phase B. The gradient was optimized with a flowrate of 0.5 mL/min and wavelength of 222 nm. A complete analytical method validation was effectively carried out as per ICH Q2 (R1) guidelines. Recovery studies were performed at 50-150% level of working concentrations, and results were in the range of 99-101%. The linearity was detected in the range of LOQ to 200% of the specification limits i.e., 0.5% each for NEO and GLY, 0.01% for NEO Impurity B and 1.0% for rest of the impurities with respect to the test concentration of the respective components. For stability study, various stress conditions such as acid, base, oxidation and thermal as per ICH guidelines were studied. The high recovery and low relative standard deviation confirm the suitability of proposed method that can be employed for the routine analysis in bulk and pharmaceutical formulation.