5-Hydroxymethylfurfural (HMF) was effectively produced from monosaccharides in a microreactor. A biphasic reaction system was employed to achieve the immediate extraction of produced HMF and suppress the overreaction. A microreactor was utilized to ensure that the reaction occurred under segmented flow to enhance the extraction efficiency. Through many attempts using phosphate buffer saline (PBS) as the reaction phase and 2-sec-butyl phenol (2BP) as the extraction phase, the favorable conditions were determined. By using PBS with pH of 2.0 and 2BP at a volume ratio of 3 to PBS, 80.9 mol % of fructose and 75.7 mol % of glucose were converted into HMF, respectively, at 180 degrees C. By comparing the results obtained through monophasic reactions, it was confirmed that the biphasic system successfully suppressed both the overreaction and the byproducts. The system employed only a simple experimental apparatus and the acid solution and organic solvent reagents without any complex expensive catalyst.