The surfactants are emerging as potential nanoparticle stabilizing agents, however, the synthetic surfactants are not economically viable as well as they are not environmentally friendly.Therefore, the biosurfactants are emerging as a green alternate for the synthesis and stabilization ofnanoparticles.In this study, Titanium dioxide (TiO2) nanopaticle were synthesized by water-in-oil micro-emulsion.Sophorolipid biosurfactant and chemical surfactant (Span80) were selected as dispersant in n-hexanol-n-hexane-water system, respectively.The critical micelle contentration (CMC) and the optical moisure content were determined in micro-emulsion, the phase and surface properties of the prepared nanopaticles were also confirmed by X-ray diffraction (XRD) and Brunaner-Emmett-Teller (BET) surface areas respecitvely.On the other hand,the photocatalytic activities of the the prepared nanopaticles were be evaluated as a probe reaction to photocatalytic degradation of methyl orange (MO).The results indicated that biosurfactant sophorolipids may have better solubilization and have lower CMC in micro-emulsion system, resulting in micelles formation easier than that of Span80.The optimal moisture content of microemulsion system is 20%.In addition, the nanoparticles of TiO2 was synthesized with the size of 7-9nm in the sophorolipid micro-emulsion (SME), which possessed smaller particle sizes, higher thermal stability and higher crystal stability, compared with those in the Span 80 micro-emulsion.The nanoparticles size from SME decreased from 9 nm at pH 4 to 7nm at pH 11;Under the optimum condition, 95 % degradation of methyl orange could be achieved within 120 min under UV irradiation with the TiO2 nanopaticles from SME as the photo-catalyst.This method provides a new gateway for the eco-friendly synthesis of nanomaterials in which the nanoparticle can be higher degradation of methyl orange of the solution without adverse effect on the environment.