R,R-2,3-Butanediol(R,R-2,3-BD) can been produced through one-step fermentation of inulin extract from Jerusalem artichoke tubers using Paenibacillus polymyxa ZJ-9.The pretreatment of fermentation broth by chitosan flocculation was investigated.The results were as following:the optimum flocculation was obtained using 0.75 g/L of chitosan with 43.5 ku molecular weight,0.125 g/L of sodium alginate,pH 5.0 and stirring for 15 minutes.The flocculation rate of cells and proteins reached 89.46% and 78.93%,respectively,and the retained ratio of R,R-2,3-butanediol reached 98.54%.Then,the different aqueous two-phase extraction systems were used to further investigate extraction of R,R-2,3-BD from the pretreated fermentation broth and the isopropanol/ammonium sulphate system was chosen as optimum system.The optimum phase composition was 33%(w/w) of isopropanol and 30.0%(w/w) of ammonium sulphate,and the partition coefficient and recovery of R,R-2,3-BD reached 7.96 and 89.40%,respectively.Furthermore,the isopropanol/ammonium sulphate aqueous two-phase extraction system could be used to separate different concentration R,R-2,3-BD from the pretreated fermentation broth by chitosan flocculation.