Stability Indicating Method Development and Validation for Quantification of Impurities in Melitracen Hydrochloride, An Antidepressant Drug Using Reverse Phase High-Performance Liquid Chromatography (RP-HPLC)

Anxiolytic and antidepressant uses for melitracen hydrochloride are the main ones. It helps treat anxiety and depression symptoms by adjusting the activity of certain neurotransmitters, mainly norepinephrine and serotonin. To increase the treatment’s therapeutic efficacy, it is frequently used in combination with other drugs. A simple, specific, and accurate stability indication of the RP HPLC method was developed to quantify melitracen hydrochloride (MH) and its associated process impurities, aiming to provide a precise and reliable analytical tool for this bulk drug. This method adheres to the International Conference on Harmonization Guidelines (ICH). The X-Terra RP 18 column from Waters Corporation (15cm × 0.46cm i.d., 5µm particle size) was used to establish an isocratic RP-HPLC on a Shimadzu HPLC system. The mobile phase contained a mixture of buffer, Acetonitrile 35:65; the buffer preparation was prepared weighed 5.23g of potassium phosphate dibasic in 1000 ml of water, stirred well, and the pH was adjusted to 7.0 using orthophosphoric Acid (phosphoric Acid, H₃PO₄). A mobile phase was made (35:65), with Acetonitrile at 35 volumes and buffer at 65 volumes. The peak elution was observed at 220nm, with a 1.2ml/min flow rate; Impurity A 3.89 minutes, impurity B 32.85 minutes, and MH 20.69 minutes were determined to have retention times, respectively. The calibration curve used to assess MH and its related impurities (Impurity A: 10-(3-(dimethylamino) propyl)-9,10-dihydro-9,9-dimethylanthracen-10-ol, and Impurity B: 10,10-dimethyl anthrone), demonstrates a straight line across the concentration range of 5µg/ml to 200µg/ml, with an r2 value of 0.999. The limit of quantification is 29.68µg/ml for MH, 7.95µg/ml for Impurity A, and 17.08µg/ml for Impurity B. Statistical analysis has been conducted on the outcomes obtained from our proposed methodologies. The API drugs were subjected to Acid, base, thermal, UV, sunlight, and peroxide degradation. The degradation studies indicated MH showed degradation only in peroxide conditions. With notable variations in their retention time values, the degradation products from the pure medication were clearly separated. Melitracen hydrochloride can be quantitatively analyzed simultaneously in bulk medications with success when using this approach.