Robust RP-HPLC Method for the Analysis of Domiphen Bromide in Pharmaceuticals with Comprehensive Sustainability Assessment

Purpose

Domiphen bromide (DB), a quaternary ammonium compound, is widely used as an antimicrobial preservative in pharmaceutical formulations. This study aimed to develop and validate a simple, sensitive, and reliable reversed-phase HPLC method for the quantitative determination of DB in pure and commercial preparations, evaluating its stability-indicating capacity and environmental sustainability.

Methods

Chromatographic separation was performed via an Inertsil ODS-3 column with acetonitrile and perchloric acid solution (70:30, v/v) as the mobile phase. Detection was performed at 275 nm with a column temperature of 25 °C. Method validation followed ICH guidelines, assessing linearity, precision, accuracy, robustness, and specificity. A quality by design (QbD) approach was employed with a 2³ full factorial design of experiments (DoE) to optimize critical parameters, including the acetonitrile ratio, flow rate, and column temperature, with statistical analysis (ANOVA) confirming their influence on retention, resolution, and peak shape. Forced degradation studies under acidic, basic, oxidative, thermal, photolytic, and neutral conditions were conducted. The method’s greenness was evaluated via multiple analytical effectiveness and eco-balance metrics. Statistical analysis included one-way ANOVA for batch comparisons.

Results

The method exhibited excellent linearity (1.132–1000 µg/mL, r² >0.999) with exceptional sensitivity (LOD: 0.373 µg/mL, LOQ: 1.132 µg/mL). The RSD values were less than 2% for the intraday and interday analyses. The accuracy ranged from 98.8 to 99.76% across the three concentration levels. Degradation studies revealed the highest susceptibility to basic hydrolysis (26.72%), followed by acid hydrolysis (18.45%) and oxidative stress (15.23%). The method successfully separated DB from all the degradation products, confirming its stability-indicating capacity. ANOVA confirmed that there was no significant batch-to-batch variation in commercial products (F = 0.82, p > 0.05). Solution stability studies confirmed standard/sample integrity for 24 h at 25 °C and 48 h at 4 °C.

Conclusions

The developed RP-HPLC method is robust, accurate, and precise for routine determination and stability assessment of DB in pharmaceutical formulations. This method was further applied to the analysis of a commercial formulation (Maalox^® oral suspension) with no observed interference from excipients. The method aligns with green chemistry principles and is suitable for quality control and regulatory applications.

Graphical Abstract