## 3 Results and Discussion  
### Delignification Step via Ultrasound-Assisted Extraction (LIAE)  
In this work, NaDESs were used as efficient, non-toxic solvents for GS delignification in combination with US. Based on the work of [PERSON] et al. for wheat straw biomass delignification [20], and focusing on the development of a novel cascade protocol for biomass valorization, three different choline chloride (ChCl)-based NaDESs (Table 1) were prepared. Regarding ChLevA, and to the best of our knowledge, different ChCl-LevA molar ratios were preliminarily tested (1:1, 1:2, and 1:10) to assess which was the proper combination that would lead to the eutectic depression and best properties. Firstly, a white crystal at room temperature was found when an equimolar proportion was used, also according to the literature [31], thus this mixture was rejected for the experiments. The opposite behavior was observed for 1:10 molar ratio, where a yellowish fluid was easily formed. The reason for using this proportion was to compare the efficiency with LA-based NaDES at the same molar ratio. However, this eutectic mixture showed poor extraction capability, \begin{table}
\begin{tabular}{c c c c} \hline \hline
**NaDESs** & **HBA 1 ** & **HBD 2 ** & **HBA/HBD 1 ** \\ \hline ChLA & ChCl & Lactic acid (LA) & 1:10 \\ ChLAGly & ChCl & Lactic acid (LA) + glycerol (Gly) & 1:1:1 \\ ChLevA & ChCl & Levulinic acid (LevA) & 1:2 \\ \hline \hline \end{tabular}
\end{table} 
Table 1: NaDESs employed. leading to a delignification yield below 10%. Therefore, according to [PERSON] et al. [31], which reported the excellent physical properties, a 1:2 molar ratio was selected between ChCl and LevA for ChLevA. Moreover, for the sake of comparison, an alkaline delignification (NaOH 0.125 M) was tested following the reported data on palm fronds or orange peel waste under US [32; 33]. The operating conditions were chosen according to literature [34], fixing the proper temperature for cavitation at 50 \({}^{\circ}\)C. The range of US frequencies typically used for this kind of processes is 20-100 kHz, in which cell rupture and polymer degradation can be promoted [35]. This value was therefore set at 40 kHz, according to previous studies [20]. Results are shown in Figure 1.  
As observed, NaDESs showed a higher efficiency for US-assisted GS delignification in comparison with the alkaline system, representing a valuable improvement in terms of sustainability. This high extracting power could be attributed to the H-bonding interactions between NaDES components and the phenol groups in lignin structure [20]. Considering alkaline extracts (Figure 1), an increase in terms of sonication time (from 60 to 120 min) led to slightly lower delignification yields (30.05% vs. 25.05%, respectively). Maybe long times under alkaline conditions could promote an aldolic-type recondensation between the lignin monomers previously extracted. [PERSON] et al. [33] reported that a non-significant increase in terms of delignification was observed if the treatment time varies from 1 h to 2 h or 4 h at 50 \({}^{\circ}\)C. In our case, despite this decrease, the difference was not significant (\(p>0.05\)). Concerning NaDESs treatments, longer sonication times might be required to reduce their viscosity and enhance the mass transfer [20]. This fact was confirmed for ChLA and ChLAGly, being only significant for the former (\(p<0.05\)). Particularly, in case of ChLAGly, the presence of glycerol (Gly)--denser than both lactic (LA) and levulinic acid (LevA)--as well as the higher amount of ChCl make this solvent even more viscous and dense, thus hindering the mass transfer also under US irradiation. This fact could explain the lower delignification yields achieved when a US-assisted ChLAGly pretreatment was adopted for GS and the absence of significant differences between 60- and 120-min treatment (\(p>0.05\)), suggesting that even longer times should be required.