An efficient lignocellulosic biomass pre-treatment is a crucial step for the valorization of these kind of raw materials. Lignocellulosic biomass is a potentially valuable resource for transformation into biofuels and bio-based products. The use of ionic liquids as media for the biomass pre-treatment is an alternative method that follows the green chemistry concept. This work proposes a new methodology for wheat straw pre-treatment with the ionic liquid (IL) 1-ethyl-3-methylimidazolium acetate ([emim][OAc]), which allowed the production of cellulose, hemicellulose and lignin-rich fractions in a rapid and simple three-step fractionation process. Various temperatures (80–140 °C) and processing times (2–18 h) of the pre-treatment were studied. The quantitative and qualitative analysis of each lignocellulosic biomass fraction was determined by FTIR measurements. The glucan content in recovered cellulose-rich fractions was investigated by enzymatic hydrolysis. The cellulose recovery dependence on the pre-treatment conditions was ascertained through regression analysis. The optimal result for the recovery of the cellulose-rich fraction was obtained at 140 °C during 6 h achieving 37.1% (w/w) of the initial biomass loading. For the same conditions, optimal results were also produced regarding the amount of glucan present (81.1% w/wbiomass) in cellulose-rich fractions, the carbohydrate enrichment in the hemicellulose fraction (96% wt) and the purity of lignin (97% wt). The recovery of IL was performed after each pre-treatment and the obtained yields were up to 86% (w/w). The recovered ILs were analyzed by 13C and 1H NMR. The presence of value-added phenolic compounds in the recovered ILs was analyzed by capillary electrophoresis. Vanillin and its derivatives, as well as other lignin-based products, were identified.
    
         
            
                 
             
                     
                    
                        
                            
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