The production of 2,3-butanediol (2,3-BDO), a dialcohol of great interest for the food, chemical, and pharmaceutical industry, through the fermentation of biomass, is a sustainable process strategic position for countries with abundant biomass generated by the agribusiness. However, the downstream process of 2,3-BDO is onerous due to the complexity of fermentation broth and the physical-chemical characteristics of the 2,3-BDO. This study investigated the feasibility of 2,3-BDO extraction from model aqueous solutions using aqueous two-phase systems (ATPS). A central composite rotational design (CCRD) was employed to evaluate different ATPS compositions and the influences on the 2,3-BDO recovery and partition coefficient. The polyethylene glycol (PEG) and different concentrations of sodium citrate, ammonium sulfate, and potassium phosphate were investigated. The concentration of salt and PEG in the ATPS was identified as the most significant factors influencing the recovery and partition coefficient of 2,3-BDO. The recovery of 2,3-BDO reached 94.5% and was obtained when the system was composed of 36.22% (w/w) of PEG 4000 and 4.47% (w/w) of potassium phosphate. The results indicate that ATPS based on PEG-salt has a high potential for industrial application, using mild conditions and a simple process for recovering and purifying the 2,3-BDO produced from microbiological synthesis.