Bacterial cellulose used for many membrane applications such as artificial dura mater membrane. This research aimed to determine the effects of glycerol concentration variation as plasticizer on physical and biological characteristics of bacterial cellulose-chitosan biocomposite membrane.Bacterial cellulose-glycerol membrane was soaked for six hours in 0.5% chitosan solution together with glycerol at variuos concentrations of 0.5%, 1%, 1.5%, and 2% using freeze dry method. Based on results of FTIR test, there is a crosslinking between-OH groups in bacterial cellulose and chitosan-glycerol. Meanwhile, results of the physical characteristic tests showed tensile strength value of 29.40 MPa for the control sample and 142.24 MPa for samples of bacterial cellulose-chitosan added 2% glycerol. It indicates that the addition of glycerol can increase tensile strength value. Results of morphological test, depicted pore size of 93.30-216 nm. Results of the degradation test demonstrated that samples added 2% glycerol degraded as much as 50.15% for 2 weeks. Meanwhile, results of the cytotoxicity test indicated that all of samples were not toxic. Finally, it can be concluded that the best result of the characterization analysis was found on the samples added 2% glycerol although they have not yet fulfilled the standard value of artificial dura mater.