The aim of this work was to optimize the glycoside composition of Stevia rebaudiana leaves using supercritical fluid extraction (SFE). A Box-Behnken statistical design was used to evaluate the effect of various values of pressure (150-350bar), temperature (40-80 degrees C) and concentration of ethanol-water mixture (70:30) as co-solvent (0-20%) by CO2 flow rate of 15 g min(-1) for 60 min. The most effective variables were co-solvent concentration (P < 0.005) and temperature (P < 0.005). Evaluative criteria for both dependent variables (stevioside and rebaudioside A yields) in the model was assigned maximum. optimum extraction conditions were elicited as 211 bar, 80 degrees C and 17.4% which yielded 36.66 mg/g stevioside and 17.79 mg/g rebaudioside A. Total glycosides composition were close to those obtained using conventional water extraction (64.49 mg/g) and a little higher than ethanol extraction (48.60 mg/g) demonstrating challenges for industrial scale application of SFE. (C) 2009 Elsevier E.V. All rights reserved.