## 5 Conclusions  
Given the current interest in sustainable protein sources, macroalgae have been identified as promising alternatives due to their protein quality and bioactivities. Seaweed proteins and seaweed-derived proteins are still poorly explored, and the development of appropriate and feasible extraction and purification processes is of growing interest in order to incorporate them in pharmaceutical, nutraceutical, cosmeceutical or food and feed applications. The major identified challenges lie on improving the protein yield and on the disruption of the polysaccharide-rich cell wall. Cell disruption techniques play indeed an essential role in the successful extraction and enrichment of algal protein at larger scales. Additionally, the protein contents and quality highly depend on parameters such as the species, harvesting season, location and growing conditions. Conventional protein extraction methods, such as enzymatic hydrolysis and various physical processes, are laborious, time-consuming and costly, and may require the use of solvents. Novel methods, such as PEF, MAE, PLE and SFE, have been predominantly used in the extraction of compounds other than proteins, and are still poorly explored in algae. Table 8 briefly compares relevant aspects of these conventional and novel extraction methods. \begin{table}
\begin{tabular}{c c c c} \hline \hline \multirow{2}{*}{**Species**} & \multicolumn{2}{c}{**Structural Features Searched**} & \multirow{2}{*}{**Method**} & \multirow{2}{*}{**Ref.**} \\  & **and Respective Bands, or** & & \\  & **Identified Proteins/Peptides** & & \\ \hline \multirow{4}{*}{_Codium fragile_} & \multirow{4}{*}{Codiase} & Fibrin(ogen)olytic activity of codiase was & \multirow{4}{*}{[40]} \\  & & examined by FTIR spectroscopy and the & \\  & & molecular weight of codiase was determined by & MALDI-TOF mass spectrometry in linear mode. & \\ \hline \multirow{4}{*}{_Kappaphycus alvarezzi_} & \multirow{4}{*}{704 cm\({}^{-1}\): N—H bending. 616 cm\({}^{-1}\): phosphate group.} & The lyophilized protein concentrate was & \\  & & pelletized with potassium bromide (1/100 ratio & \\  & & pelletized with potassium bromide (1/100 ratio & \\  & & \(w/w\)) and the spectral analysis was carried out & \\  & & using FTIR. & \\ \hline \multirow{4}{*}{_Macrocystis pyrifera_ and _Chondracanthus chamissoi_} & 3281 cm\({}^{-1}\) and 3274 cm\({}^{-1}\): N—H vibrations. & & \\  & 1637 cm\({}^{-1}\) and 1544 cm\({}^{-1}\): C=O & FTIR of dried seaweed and protein extracts & \\  & & vibrations. & of seaweeds. & \\  & 1220 cm\({}^{-1}\) and 1243 cm\({}^{-1}\): S=O & & \\  & & vibrations. & \\ \hline \multirow{4}{*}{_Palmaria palmata_ and _Soliera chordalis_} & \multirow{4}{*}{Bioactive peptides} & Peptide samples were separated by online & \multirow{4}{*}{[49]} \\  & & reversed-phase (RP) nanoscale capillary liquid & \\  & & chromatography (nanoLC) and analyzed by & \\  & & electrospray mass spectrometry (ES MS/MS). & \\ \hline \multirow{4}{*}{_Palmaria palmata, Palmaria_} & \multirow{4}{*}{Mono-iodotyrosine} & Reverse phase high performance liquid & \multirow{4}{*}{[120]} \\  & & chromatography (RP-HPLC) with inductively & \\ \cline{1-1}  & & coupled plasma mass spectrometry (ICP-MS). & \\ \hline \hline \end{tabular}
\end{table} 
Table 7: Protein characterization using spectrometry methods in different seaweed species. To achieve progress in this protein extraction area, future work addressing the applicability of these novel methods is necessary. The same applies to membrane technologies, which also show great promise to pre-purify/concentrate or even isolate (in membrane chromatography) compounds, but are still fully underexploited regarding the protein extraction from macroalgae. Conceptualization, M.M. and M.T.C. ; investigation, M.G., M.M. and M.T.C. ; writing--original draft preparation, M.G. ; writing--review and editing, H.M.P., M.M., M.M.R.d.F. and M.T.C. ; project administration, H.M.P. and M.T.C. ; funding acquisition, M.M.R.d.F. and M.T.C.