Eco-friendly simultaneous determination of oxytetracycline HCl, sulfadimethoxine sodium, and trimethoprim glutamate in pharmaceutical formulations using HPLC

The development of environmentally sustainable analytical methods for simultaneous determination of multiple active pharmaceutical ingredients (APIs) is critical for advancing pharmaceutical quality control. This study introduces a novel, green reversed-phase high-performance liquid chromatography (RP-HPLC) method for the concurrent quantification of oxytetracycline (OTC), sulfadimethoxine sodium (SFM), and trimethoprim glutamate (TMG) in veterinary formulations. Using a Hypersil BDS C18 column (250 mm x 4.6 mm, 5 mu m) with an isocratic mobile phase (phosphate buffer pH 7: methanol, 55:45 v/v) at 1.0 mL/min and detection at 280 nm, the method achieved baseline separation of all analytes within 6 min. Validation per ICH guidelines demonstrated excellent linearity (r2 >= 0.9994) across ranges of 10-200 mu g/mL (SFM), 30-300 mu g/mL (OTC), and 6-40 mu g/mL (TMG). Sensitivity was confirmed with limits of detection (LOD) of 2.34, 8.33, and 1.05 mu g/mL and limits of quantification (LOQ) of 7.09, 25.24, and 3.20 mu g/mL for SFM, OTC, and TMG, respectively. Accuracy (98.40-102.05 % recovery) and precision (%RSD < 2 %) met regulatory standards. The method's environmental sustainability was rigorously evaluated using AGREE (score: 0.82), NEMI, ComplexGAPI, and AMVI tools, highlighting reduced solvent consumption (AMVI = 4.67 vs. 350 for conventional methods) and compliance with green chemistry principles. Additionally, the BAGI tool affirmed its practicality (score: 87.5/100), underscoring cost-effectiveness and ease of implementation. Successful application to a commercial veterinary powder, containing oxytetracycline HCl, sulfadimethoxine sodium, and trimethoprim glutamate, confirmed label claims (99.54-100.59 % recovery) with no matrix interference. This method represents the first simultaneous analysis of OTC, SFM, and TMG, offering a robust, eco-friendly alternative to pharmacopeial methods while enhancing efficiency in pharmaceutical quality control.