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Xylooligosaccharides (XOS) are non-digestible food ingredients with prebiotic properties for selectively promoting the growth of probiotics, which provide many health benefits and several applications in the food and pharmaceutical industry. The objective of this study was to optimize the concentration of commercial hemicellulases for the production of XOS, with a 2–6 polymerization degree, using a mixture of sugarcane bagasse and straw pretreated with ionic liquid or diluted sulfuric acid. The concentrations of enzymes endo-1,4-xylanase (NS50030, Novozyme®) and α-L-arabinofuranosidase (GH51) (Megazyme®) were optimized using a central composite rotatable design (CCRD). The xylooligosaccharides (XOS) released by hydrolysis were analyzed via capillary electrophoresis and quantified with HPAEC-PAD. The XOS profile obtained from the hydrolisis of the pretreated sugarcane biomass mixture (MPSA …