生化学と分析生化学

概要

Analytical Method Development and Validation for Determination of Assay of Antibacterial Drugs Besifloxacin Hydrochloride and Phenoxyethanol in Gel Formulation

Pradeep Kumar, Neelam Pawar*, Neha Minocha, Kavita Bahmani, Asha Poonia

Aim: The aim of the present work is to develop an analytical method and validate it for determination of assay of an antibacterial dug in gel formulation.

Background: Analytical method validation is a process involving confirmation studies that procedure/method/ system/analyst provides precise and reproducible outcome recognized by research laboratory studies that the performance features of the technique follows the necessities required for the analytical applications.

Objective: To improve the conditions and parameters which should be followed in the development and validation by developing a new sensitive and accurate RP-HPLC method? Validating the proposed newly developed methods in accordance with the analytical parameters mentioned in the IP, USP, BP and ICH guidelines.

Methods: HPLC method was validated to indicate that the analytical Procedure used is suitable for intended use by using various parameters like specificity, linearity, LOD, LOQ, precision, accuracy, range, robustness, stability in analytical solution and system suitability.

Results: The Retention times for Drug Besifloxacin standard were found to be 7.781 min and sample was at 7.731 respectively. The Area of standard besifloxacin was 1828547 and sample area was 1825315. The Assay of sample was 98%. The Retention times for Drug Phenoxyethanol standard were found to be 2.010 min and sample was at 2.004 respectively. The Area of standard Phenoxyethanol was 438025 and sample area was 438103. The Assay of sample was 97.04%. In System suitability, The RSD for 5 replicate injections for each peak is 0.33%. In specificity peaks of Diluent, Placebo and Impurities are not interfering with the Besifloxacin peaks. Peaks of Besifloxacin were found to be pure. Degradation products were found to be well separated from besifloxacin peak. The peak purity factor was NLT 0.9995.In precision study System Precision RSD of the Retention time for Besifloxacin obtained from six replicate injections 0.33%. The RSD of the Area of Besifloxacin obtained from six replicate injections is 0.46%. Method precision RSD was calculated on 6 determinations assay value of Drug besifloxacin is 0.56%. The RSD calculated on 6 determinations for assay value of Drug besifloxacin is 0.50%. In Intermediate precision RSD was calculated on 6 determinations for assay value of Drug besifloxacin is 0.50%. The RSD calculated on 12 determinations (Method precision and Intermediate precision) for assay value is 0.50%. Stability in analytical solution for the standard and sample, the area difference of besifloxacin peak were found to be within ± 2.0% from initial Linearity. The correlation coefficient and regression coefficient (R square) should be not less than 0.995 for Besifloxacin Correlation Coefficient is 0.998 Regression coefficient is 1.000. The % intercept should be within ± 5.0% of the response at 100% level Precision at 50% and 150% level: the RSD is 0.01%. Precision at 50% and 200% level: the RSD found to be NMT 2.0%. Accuracy mean % recovery at each level found to be between 98 to 101% of drug besifloxacin. RSD on 9 (3 levels x3) determinations is 1.2 which is following the criteria of NMT 2% Range. Correlation was found 1.0% for the accuracy and linearity parameters.

Conclusion: The recovery is in between 98% to 101% and the % RSD for all recovery values is 1.41% which is in limits. In HPLC method, the conditions were optimized to obtain an adequate separation of eluted compounds.