In the present study, the optimization of extractive desulfurization in the microchannels was investigated using a deep eutectic solvent (DES). The influence of some operating and microchannel geometrical parameters, including the initial sulfur concentration of fuel, the length of the microchannel, the DES flow rate, and the ratio of fuel to DES flow rate (RF/D), on sulfur removal was investigated using response surface methodology (RSM). Moreover, the flow pattern map in the microchannels was investigated. In this regard, different flow patterns of slug, lengthy slug, smooth annular, throat annular, wavy annular, and droplet were observed in the microchannels applied in the extractive desulfurization (EDS) process. Besides, the effects of the inertial force, interfacial tension force, and viscosity force on the different flow patterns were discussed by analyzing the capillary number, the Weber number, and the Reynolds number. It was found that the most effective regime in the EDS process in the microchannel is the slug flow pattern in which the interfacial tension force (6 × 10 −5 < Weber number < 0.82767) is dominant. Based on the analysis of the RSM proposed model, it was also found that the RF/D is the main influential parameter on sulfur removal, and 73.2% sulfur removal was achieved in the microchannel under the optimum conditions in a very short residence time of 2.5 s. The results also confirmed the capability of this EDS process for sulfur removal from real diesel fuel.