## 4 Biodegradable Solvents for Protein Extraction
### Solvent Recovery Options  
From an industrial point of view, recycling and reusing the solvents is essential to achieve a cost-efficient process [120]. However, it is also vital to obtain the product (proteins) without any impurity of solvent or by-product. Thus far, research has focused on transferring the proteins from the matrix to the biodegradable solvent. However, several issues such as solvent recycling, protein recovery from the biodegradable solvent, \begin{table}
\begin{tabular}{c c c c c c} \hline \hline
**Microalgae** & **DES/IL Used** & **Salt/Polymer** & **Extraction Method** & **Yield** & **Reference** \\
**Strain** & **(Molar Ratio)** & **Used (in Case of ATPS)** & **and Conditions** & **Yield** & **Reference** \\ \hline \multirow{4}{*}{_Haematococcus_ _pluvialis_} & Trihexyl[tetradecyl] & \multirow{4}{*}{None} & Single IL extraction, aided with & \multirow{4}{*}{Protein recovery: \(>\)90\%} & \multirow{4}{*}{[82]} \\  & Phosphonium bis- & & & & \\ \cline{3-5}  & 325 trifamide & & & & \\ \cline{3-5}  & & & & & \\ \cline{3-5}  & & & & & \\ \cline{3-5}  & & & & & \\ \hline \multirow{4}{*}{_Chlorella vulgaris_} & Cholinium 2- & \multirow{4}{*}{None} & Single IL extraction & \multirow{4}{*}{Protein recovery: \(>\)90\%} & \multirow{4}{*}{[83]} \\  & hydroxy-3-morpho & & & & \\ \cline{3-5}  & linopropanesulfonate & & & & \\ \cline{3-5}  & & & & & \\ \cline{3-5}  & & & & & \\ \hline \multirow{4}{*}{_Naechloris_ _oleabundans_} & Tributylmethyl & \multirow{4}{*}{None} & Biomass:Solvent concentration: \(<\)6.67 g L\({}^{-1}\) & Protein recovery: \(<\)80\% & \multirow{4}{*}{[84]} \\  & phosphonium methyl sulfate & & & & \\ \cline{3-5}  & (40\% solution) & & & & \\ \cline{3-5}  & & & & & \\ \cline{3-5}  & & & & & \\ \hline \multirow{4}{*}{_Naechloris_ _oleabundans_} & Cholinium & Polypropylene & IL-ATPS extraction & \multirow{4}{*}{Protein recovery: \(<\)80\%} & \multirow{4}{*}{[86]} \\  & dihydrogen & glycol 440 & & & \\ \cline{3-5}  & phosphate (14 wt\%) & (40 wt\%) & & & \\ \cline{3-5}  & & & & & \\ \cline{3-5}  & & & & & \\ \hline \multirow{4}{*}{_Isochrysis_ _galbana_} & 1-decyl-3- & \multirow{4}{*}{K\({}_{3}\)PO\({}_{4}\)} & IL-ATPS extraction: 10,000 rpm, & Protein separation: \(<\)100\% in the & \multirow{4}{*}{[87]} \\  & methylimidazolium & (20 wt\%) & & & \\ \cline{3-5}  & chloride (15 wt\%) & & Temperature: RT & top phase \\ \hline \multirow{4}{*}{_Naechloris_ _oleabundans_} & \multirow{4}{*}{Iollity 221 PG} & \multirow{4}{*}{Potassium citrate} & IL-ATPS extraction: 14,000 rpm and 30 min & Protein extraction efficiencies: \(<\)75–85\% & \multirow{4}{*}{[88]} \\  & & & & & \\ \cline{3-5}  & & & & & \\ \cline{3-5}  & & & & & \\ \cline{3-5}  & & & & & \\ \hline \multirow{4}{*}{_Scenedesmus_ _dimorplus_ (UTEX 1237)} & \multirow{4}{*}{1,2-propanediol; \(<\)6.67} & Single DES extraction: 250 rpm & \multirow{4}{*}{None} & Single \\  & chloine chloride; \(<\)1:1:1 & & & & \\ \cline{3-5}  & & & & & \\ \cline{3-5}  & & & & & \\ \cline{3-5}  & & & & & \\ \cline{3-5}  & Octanoic acid and & & & & \\ \cline{3-5}  & dodecanoic acid & & & & \\ \cline{3-5}  & (1:1), plus a dilute & & & & \\ \cline{3-5}  & (5\%) aqueous & & & & \\ \cline{3-5}  & solution of Jeffamine & & & & \\ \cline{3-5}  & D-230 & & & & \\ \hline \hline \end{tabular}
\end{table} 
Table 2: Cont. and potential solvent regeneration are not usually addressed and remain to be solved. Mainly because of the low vapor pressure of the solvents used, the isolation of the proteins from ILs or DESs by vaporization (the most extended method) is complicated, implying protein degradation [100]. New approaches have been attempted, such as the use of resins, ultrafiltration, adsorption [120, 121]. Liquid-liquid re-extraction can be an efficient approach to isolate the biocompatibility and recover the solvent [122], although organic solvents could be involved, limiting the greenness of the process. The liquid-liquid re-extraction (or back-extraction) method is preferred for studies concerning protein recovery.