# Microalgae as Potential Sources of Bioactive Compounds for Functional Foods and Pharmaceuticals
## 3 Results and Discussion
### Chemical Characterization of Promising Microalgal Extracts' Fractions  
The most active fractions at each bioactive assay (hexane fractions of POC ET and SKE ET due to neuroprotective activity; dichloromethane fraction of PHA ET for anti-tumoral activity and hexane fraction of TCTP4 ET for anti-inflammatory activity), were analyzed by GC-MS to tentatively identify compounds that may be responsible for the activities tested. The hexane fractions of ethanol extracts from _Porphyridium_ sp. and _Skeletonema_ sp. generated a total of 45 and 29 peaks in the GC-MS chromatogram, with 40 and 23 peaks having been satisfactorily identified, respectively. The analysis of the hexane fraction of ethanol extract of _Tetraselmis striata_ CTP4, revealed 43 compounds with 42 compounds being successfully identified. Finally, the analysis of the dichloromethane fraction from the ethanol extract of _Phaeedactylum tricornutum_ revealed a total of 52 peaks, of which 33 were successfully identified. The complete list and chemical classes of the identified compounds are presented in Figure 3 and Table 8.  
Figure 2: **(A)** Cell Viability (%) of macrophage-differentiated THP-1 (Mac THP-1) cells after incubation with hexane (H) and dichloromethane (DCM) fractions of the ethanol crude extract from _Tetraselmis striata_ CTP4 at 10, 20 and 50 \(\upmu\)g/mL for 48 h, assessed through the MTS assay. **(B)** Anti-inflammatory activity of the crude extract and fractions defined in the panel A in LPS-stimulated Mac THP-1 cells. Evaluation of the inflammatory marker TNF-\(\alpha\) was performed by ELISA in the cell culture media of Mac THP-1 cells treated for 24 h with the crude extract and fractions at 10, 20 and 50 \(\upmu\)g/mL, and then stimulated with LPS (100 ng/mL) for a further 24 h. Dexamethasone (DXM) (2 \(\upmu\)M) was used as a positive anti-inflammatory control and non-stimulated cells (CTR) as controls to LPS stimulation (LPS). Data are representative of three independent experiments, and presented as mean \(\pm\)SD. Two-way ANOVA and multiple comparisons were achieved with the Dunnett’s test and presented relatively to the control untreated cells in **(A)**, and LPS-stimulated cells in **(B)**. Statistical significance was defined as \(p\leq 0.05\) (*), and \(p\leq 0.001\) (***). \begin{table}
\begin{tabular}{c c c c c} \hline \hline  & \multicolumn{4}{c}{**Relative Abundance (\%)**} \\ \hline \multirow{4}{*}{**Compounds**} & \multirow{2}{*}{
\begin{tabular}{c} _Porphylidium sp._ \\ **(POC ET)** \\ **-Hexane Fraction** \\ \end{tabular} } & \multicolumn{2}{c}{_Skeletonema_} & \multicolumn{2}{c}{_Tricontutum_} & \multicolumn{1}{c}{_Tetraselmis striata_} \\  & **(POC ET)** & **sp. (SKE ET)** & **(PHA ET)** & **CTP4 (TCTP4)** \\  & **-Hexane Fraction** & **-Hexane Fraction** & **-Dichloromethane** & **-Hexane Fraction** \\  & & & **Fraction** & \\ \hline Cyclohexanol, 1-butyl & 2.61 & & & \\ \hline
2H-Pyran, & & & & \\
2-[(Scyclopropyl idene \\ pentyl)oxy]tetrahydro & 2.01 & & & & \\ \hline
3,7,11,15-Tetramethyl- & 0.88 & 7.01 & 5.35 & 1.51 \\
2-hexadecen-1-ol & & & & \\ \hline
4-Hexen-3-ol, & & & & \\
2,5-dimethyl & 0.80 & & & \\ \hline
2,3-Dimethyl-undec-1- & & & & \\ en-3-ol & & & & \\ \hline \hline \end{tabular}
\end{table} 
Table 8: Chemical characterization by GC-MS of compounds presents in the most bioactive fractions of the ethanol extracts of _Porphylidium_ sp. (hexane fraction), _Skeletonema_ sp. (hexane fraction), _Phaeodactylum tricontutum_ (dichloromethane fraction) and _Tetraselmis striata_ CTP4 (hexane fraction). Figure 3: Main chemical compound classes identified in the ethanol microalgal extracts by GC/MS. (**A**) Hexane fraction of _Porphylidium_ sp. ; (**B**) Hexane fraction of _Skeletonema_ sp. ; (**C**) Dichloromethane fraction of _Phaeodactylum tricontutum_ and (**D**) Hexane fraction of _Tetraselmis striata_ CTP4. PUFA, polyunsaturated fatty acids; AA, alkanes and alkenes; SFAs, saturated fatty acids; MUFAs, monounsaturated fatty acids.