Bioethanol production from the shedding bark of Melaleuca leucadendron (Paper-bark Tree, PBT) was studied using subcritical water (SCW) pretreatment at various severities (S-o). High ethanol production was attained by implementing a factorial design on three parameters (S-o, solid loading and enzyme loading) in simultaneous saccharification and fermentation (SSF) mode. Ethanol concentration of 63.2 g L-1 corresponding to ethanol yield of 80.9% were achieved from pretreated biomass (S-o = 2.37) at 0.25 g mL(-1) solid and 16 FPU g(-1) glucan enzyme loadings. Similarly at 0.15 g mL(-1) solid loadings both high ethanol concentration (43.7 g L-1) and high ethanol yield (91.25%) were achieved. Regression analysis of experimental results shows that all process parameters had significant role on maximum ethanol production, glucose solubility, ethanol yield and ethanol volumetric productivity. SSF of SCW treated PBT biomass is economically feasible for production of bioethanol. (C) 2013 Elsevier Ltd. All rights reserved.