Utility of the chromogenic and fluorogenic properties of benzofurazan for the assay of milnacipran in human urine and plasma

Abstract

Our article presents the development and validation of two simple, very sensitive, and low-cost spectroscopic methods for the assay of milnacipran hydrochloride in bulk form, pharmaceutical tablets and spiked human urine and plasma. Spectroscopic methods (spectrophotometric and spectrofluorimetric techniques) were dependent on the chromogenic and fluorogenic properties of the 4-chloro-7 nitrobenzofurazan (NBD-Cl) reagent. The reaction product, resulting from the interaction between NBD-Cl and milnacipran in the presence of borate buffer pH 8.5, was measured spectrophotometrically at 465 nm and spectrofluorimetrically at 510 nm after excitation at 465 nm. The absorbance–concentration plot was rectilinear over the range of 1.5–12 μg mL⁻¹ with a limit of quantitation 1.09 μg mL⁻¹, while the fluorescence–concentration plot was rectilinear over the range of 0.03–0.5 μg mL⁻¹ with a limit of quantitation 0.02 μg mL⁻¹. Influential parameters affecting the development and stability of the reaction product were studied and optimized. Assurance of the cited drug in its tablets by our proposed methods was successfully completed without obstruction from the presence of the basic excipients with average percentage recoveries of 99.27 ± 1.18 and 99.44 ± 0.69 for the spectrophotometric and spectrofluorimetric methods, respectively. The spectrofluorimetric method was additionally adopted as a preliminary in vitro study for the assay of the cited drug in spiked human urine and plasma with average percentage recoveries of 101.52 ± 1.01 and 100.38 ± 1.57 for spiked urine and plasma, respectively.