## 2 Results
### Effect of Enzyme, Substrate and Water Contents  
Herein, Novozym 435(r) was used as the biocatalyst for the reaction that was carried out in the \"2-in-1\" Sorbit DES. Factors impacting the reaction (i.e., enzyme concentration, enzyme dosage and water content in the media) have been investigated to find the optimal parameters for each condition. From Figure 4, it can be discerned that the optimal values were 50 g/L enzyme, 0.5 M vinyl laurate and 5 wt.% water. \begin{table}
\begin{tabular}{c c c c c} \hline \hline
**Formulation Name** & **Reported Activity * & **Formulation Type** &
\begin{tabular}{c} **Reported Optimal** \\ **Temperature Range (°C) * \\ \end{tabular} & **Titer (g/L) ** \\ \hline CalA Immuno 150 & 500 U/g & Immobilized & N.C. & 2.3 \(\pm\) 0.9 \\ CalB Immuno Plus & \textgreater{}9000 PLU/g & Immobilized & 60–80 & 13.3 \(\pm\) 3.0 \\ Lipase CA CLEA & \textgreater{}1.5 U/mg & Cross-linked & \textgreater{}40 & 38.2 \(\pm\) 3.8 \\ Lipase TL CLEA & \textgreater{}25 U/mg & Cross-linked & 40–60 & 9.2 \(\pm\) 2.7 \\ Lipozym 435 & 9000 PLU/g & Immobilized & N.C. & 38.2 \(\pm\) 3.8 \\ Lipozyme CALB L & 5000 LU/g & Liquid & 30–60 & 38.2 \(\pm\) 3.8 \\ Lipozyme RM IM & 275 IUN/g & Immobilized & 30–50 & 13.2 \(\pm\) 2.7 \\ Lipozyme T1 00L & 100 KLU/g & Liquid & 20–50 & 3.3 \(\pm\) 1.2 \\ Lipozyme TL IM & 250 IUN/g & Immobilized & 50–75 & 1.3 \(\pm\) 1.0 \\ NovoCor AD L & 6000 LU/g & Liquid & 30–60 & 3.3 \(\pm\) 1.2 \\ Novozym 40086 & 275 IUN/g & Immobilized & 30–50 & 2.2 \(\pm\) 1.1 \\ Novozym 435 & 10,000 PLU/g & Immobilized & 30–60 & 38.2 \(\pm\) 3.8 \\ Novozym 51032 & 15 KLU/g & Liquid & 35–70 & 4.4 \(\pm\) 1.3 \\ Novozym NS 81356 & N.C. & Immobilized & N.C. & 10.6 \(\pm\) 1.4 \\ Palatase 20000 L & 20,000 LU/g & Liquid & 30–50 & 10.0 \(\pm\) 2.1 \\ Resinase HT & 50 KLU/g & Liquid & \(\leq\)90 & 7.3 \(\pm\) 1.3 \\ \hline \hline \end{tabular}
\end{table} 
Table 2: Titer (g/L) of SL obtained after 48 h with commercially available enzyme formulations. Figure 3: Time course of the reaction under unoptimized conditions: 0.5 M of vinyl laurate, 20 mg, of Novozym 435® and 1.5 mL of Sorbit DES as solvent (sorbitol/choline chloride, 1:1, mR, 5 wt.% water) at 50 \({}^{\circ}\)C.  
Statistical analysis of the different titers reached after 48 h in relation to enzyme concentration revealed significant differences when the amount of enzyme was at least doubled from the 20 g/L starting point (Figure 4A). Drastic differences can be observed in Figure 4B, as 0.5 M of vinyl laurate induces a two-fold increase in titer compared to 0.25 M and 0.75 M. This safely suggests that upon a range of concentration > 0.5 M, there  
Figure 4: Novozym 435(b)-catalyzed transesterification of sorbitol and vinyl laurate in “2-in-1” Sorbit deep eutectic system: Effect of enzyme dosage (**A**); vinyl laurate concentration (**B**); water content in the media (**C**) on the titer after 48 h. a–d show statistically significant differences, at a 0.05 significance level, of the mean values obtained from three independent experiments ran under each condition. was potentially an inhibition of the biocatalyst due to substrate saturation. Above this concentration and in spite of the statistical analysis, comparisons are rendered ambiguous considering the lack of trend and should be interpreted with a certain distance. The results displayed in Figure 4C are rather more ambiguous as well, concerning the water content. Indeed, above 5 wt.% water, it seems that an optimal range of water content was reached. Overall, the results tend to indicate that enzyme and substrate concentrations were predominant factors compared to the water content, to some extent, as the Sorbit DES with a 2.5 wt.% water content ultimately limits the conversion most probably due to its measured 2-fold higher viscosity (Table A1). It also seems that the Sorbit DES with 5 wt.% water provides the optimal A\({}_{\mathrm{w}}\) for the enzymatic activity (-0.08).