A Validated HPTLC Method for the Estimation of Amitriptyline HCL in Bulk and Its Tablet Dosage Form

Abstract

A simple, rapid, reliable and accurate HPTLC method has been developed for the quantitative determination of Amitriptyline HCL in bulk and tablets. Various aliquots of the sample solution were spotted automatically by means of camag ATS 4 applicator on precoated silica gel 60 F₂₅₄ on aluminium sheet as stationary phase pre washed with methanol using Toluene: Methanol: Acetone: Ammonia (5:3:2:0.2)v/v/v/v as mobile phase. The spots were scanned at 254 nm. The R_f value of AMITRIPTYLINE HCL was 0.66 ± 0.02. Calibration curves were linear in the range of 67.5 - 472.5 ng/band. The limit of detection and limit of quantification were found to be 9.37 ng/band and 2.80 ng/band respectively. The suitability of this method for the quantitative determination of compound was proved by validation in accordance with requirements of pharmaceutical regulatory standards.

Objectives: This study aimed to enhance the stability indicating chromatographic method for determining Amitriptyline hydrochloride in the presence of degradation products and impurities, ensuring purity and stability of the bulk drug using HPTLC, in compliance with ICH recommendations.

Methods: The study used a pure gift sample of Amitriptyline HCL from Unichem Laboratories Ltd, GOA, India, and prepared a mobile phase with methanol, toluene, acetone, and ammonia. The substance's melting point was determined, and solubility was tested in various solvents. The ideal solvent was methanol, as it was completely soluble, stable, and economical. A standard stock solution of 0.1 mg/ml of Amitriptyline HCL was prepared by dissolving 10 mg in methanol. A sample was prepared by powdering 20 tablets, adding 50mL of methanol, sonicating, and filtering the solution.Chromatography was conducted on aluminium packed silica gel 60 F254 HPTLC plates, which were washed and dried before use. Samples were applied as 6 mm bands, and ascending development was performed at 25°C with a mobile phase of Toluene, Methanol, Acetone, and Ammonia. Densitometric scanning was performed using a Camag TLC scanner 4 with Wincats software. The following validation parameters are typically monitored for HPTLC method : Linearity, Sensitivity, Specificity, Precision, Data of repeatability, Interday Precision Data and Recovery. The mean weight of 20 tablets was determined, and a finely powdered powder equivalent to 1 tablet of Amitriptyline HCL was prepared, with a drug content of 99.53.

Results: The HPTLC method was validated according to ICH guidelines and was found to be linear, accurate, and precise for Amitriptyline HCL. Its sensitivity was found to be low, with a sensitivity coefficient of 0.999 and a coefficient of variation of 0.72%.

Conclusions: The developed HPTLC technique is precise, specific and accurate. The advantages lie in the simplicity of sample preparation and the low cost of reagents used. Statistical analysis proves that the method is suitable for the analysis of Amitriptyline HCL as bulk drug and in Pharmaceutical formulation without any interference from the excipient. Hence this HPTLC method can be used for routine drug analysis.