3.1 Determination of the molar ratio between ChCl and DHBA
Among the tested molar ratios of ChCl and DHBA, the molar ratio of 2:1 and 3:2 with ChCl and DHBA formed a homogenous and transparent liquid, as shown in Figure S1 . A previous study reported that the DESs with the same components at different molar ratios showed different properties and performances.32 In this study, prior to investigating ternary DESs, two binary ChCl-DHBA DESs with different molar ratios (3:2 and 2:1) were evaluated by applying them to pretreat TG sorghum stover. The pretreatment was carried out at 120 ℃ for 3 h, adapted from our previous study of DES pretreatment with sorghum stover.33 The pretreatment efficiency was determined based on xylan removal, delignification, and fermentable sugar production, as shown in Table 1 . ChCl-DHBA 3:2 DES showed higher xylan removal and delignification than its 2:1 counterpart. Clearly, the composition of DES is an important factor affecting the pretreatment performance. Since the removal of both xylan and lignin could enhance the enzymatic digestibility of the biomass, both ChCl-DHBA DES-pretreated biomass samples were tested for the release of fermentable sugars using a cellulase mixture. The sorghum stover pretreated with ChCl-DHBA 3:2 DES released a higher amount of glucose (391 mg/g-biomass) than that pretreated with ChCl-DHBA 2:1 DES (341 mg/g-biomass); therefore, the 3:2 molar ratio of ChCl and DHBA was selected for subsequent ternary DES testing.
Table 1. Xylan removal, delignification, and glucan production after 72 h of enzymatic hydrolysis of the pretreated transgenic (TG) sorghum stover with ChCl-DHBA at different molar ratios at 120 ℃ for 3 h.