Spectrophotometric Approach for the Determination of Dobutamine Hydrochloride Using Diazotization-Coupling Reaction

Abstract

A simple, rapid, and sensitive spectrophotometric method has been suggested for the determination of Dobutamine hydrochloride (DB) in pure forms and pharmaceutical formulations using an azo-coupling reaction with an innovative reagent derived from methotrexate. The developed method is based on the diazotization process of methotrexate using NaNO2 in an acidic medium at (0–5°C), followed by a coupling step with Dobutamine in an alkaline medium (NH4OH) to yield a stable yellow azo dye showing a maximum absorbance at 466 nm. Experimental factors—including the type and volume of acid, diazotizing agent, base, reaction duration, solvent selection, and order of addition—were practically optimized. Under the established optimal conditions, the method demonstrated linearity in the concentration range 2–60 µg/mL, with a correlation coefficient (R²) of 0.9943 and a molar absorptivity of 1.10 × 10⁵ L·mol⁻¹·cm⁻¹. The limits of detection (LOD) value and quantification (LOQ) value were determined to be 0.0231 µg/mL and 0.0702 µg/mL, respectively. Accuracy and precision of the proposed spectrophotometric method were confirmed through recovery studies, yielding a mean recovery of 100.68% and a relative standard deviation value (RSD) of less than 6.42%. The proposed approach was successfully applied to the quantitative analysis of Dobutamine in pharmaceutical formulations (Injection), demonstrating excellent accuracy and negligible interference from common excipients. A stoichiometric ratio of 1:1 (Dobutamine –Methotrexate) was confirmed.