Robust and ICH-validated UV-Vis spectrophotometric method for the determination of benfotiamine in bulk substance, formulated products, and plasma using Bratton-Marshall chemistry

Background: Benfotiamine is widely used in the treatment of diabetic neuropathy and metabolic disorders. For the determination of pharmaceutical formulations and biological matrices, Spectrophotometry is widely employed in pharmaceutical analysis due to its simplicity and cost-effectiveness. The Bratton–Marshall reagent is used for Chromogenic reactions involving aromatic amines, producing colored complexes suitable for spectrophotometric quantification. The study aimed to develop and validate a simple, rapid, accurate, and precise UV–Visible spectrophotometric method for quantitative estimation of benfotiamine in bulk drug and human plasma using the Bratton–Marshall reagent. Methods: Benfotiamine was dissolved in 0.1 M hydrochloric acid and reacted with the BM reagent. In this reaction, sodium nitrite in an acidic medium diazotizes the primary aromatic amino group of benfotiamine, producing a pink Chromogenic exhibiting maximum absorbance at 550 nm. The Chromogenic method was further extended to a bioanalytical application for plasma samples. Protein precipitation was carried out using Acetonitrile to remove plasma proteins, followed by vortex mixing and centrifugation at 1000 rpm for 15 minutes. The resulting supernatant was analyzed spectrophotometrically. Results and Discussion: The developed method demonstrated linearity in the concentration range of 3–21 µg/ml with a correlation coefficient (R²) of 0.998. Accuracy ranged between 98.44% and 99.64%. Precision studies indicated %RSD values below 2% and remained within acceptable limits. The recovery of benfotiamine from human plasma was 98.91%, and the matrix effect showed acceptable variability. Conclusion: The proposed method is rapid, precise, and reliable for the quantification of benfotiamine in pharmaceutical and plasma samples.