3.1.1. Antitumor Activity

Naturally occurring compounds have been tested for antiproliferative/cytotoxic, pro-apoptotic, anti-metastatic, and anti-neoplastic activities, among others [52–54].

Screening of antiproliferative activity is the most common approach to evaluate antitumor potential of complex lipids. Several cancer cell lines have been used including hepato [55,56], cervix [57], breast [56,58], leukemia [58,59], colon [58,60], lung [58,61], melanoma [62], and prostate and ovarian cancer [58]. The majority of these studies used lipid fractions enriched in a specific lipid group or class, obtained by using silica gel columns and solvents with different polarities. This approach was performed, for example, to evaluate the PLs fraction of the brown seaweed *Sargassum marginatum* inhibiting promyelocytic cells (HL-60) [59]. There is a huge variety of classes within PLs group that can contribute for bioactivity of these fractions; thus, the analysis of enriched lipid fractions solely provides a partial interpretation of results. Fractions enriched in GLs classes were isolated, allowing the identification of inhibitory activity against several cancer cells lines in digalactosyldiacylglycerol (DGDG) [60] and sulfoquinovosyldiacylglycero (SQDG) [55–57,60] enriched fractions (Table 1).

Few works have evaluated bioactivities of isolated lipid classes. The monogalactosyldiacylglycerol (MGDG) (MGDG 14:0\_16:1) from the red seaweed *Solieria chordalis* and DGDG (14:0\_18:3) from the green seaweed *Ulva armoricana* were found to have activity against NSCLC-N6 cancer cells [61]. However, to the best of our knowledge, the authors only identified the most abundant lipid species in the fraction, undervaluing other unidentified lipid species. Therefore, the antiproliferative activity of previous GLs molecular species could be incorrectly attributed.

There are very few studies that achieved the isolation and identification of pure compounds, such as 1-*O*-(5Z, 8Z, 11Z, 14Z, 17Z-eicosapentanoyl)-2-*O*-(6Z,9Z,12Z,15Zoctadecatetraenoyl)-3-*O*-β-D-galactopiranosyl-*sn*-glycerol, (MGDG (20:5/18:4)) (Figure 9A) from the brown seaweed *Fucus evanescence* [62] with activity against malignant melanoma (SK-MEL-28), and 1-*O*-(palmitoyl)-2-*O*-(5Z, 8Z, 11Z, 14Z-eicosatetraenoyl)-3-*O*-β-Dgalactopyranosylglycerol, (MGDG 20:4/16:0) (Figure 9B) from the red seaweed *Hydrolithon reinboldii*, which demonstrated inhibitory activity against a range of 12 cancer cell lines [58].

Along with the assessment of cell viability and the antiproliferative effect of lipid extracts, several biochemical approaches have also been developed in order to interrupt the cancer cells progression, including the inhibition of enzymes and disruption of mitotic process. The inhibition of DNA polymerases α was achieved by GLs species identified as galactosyldiacylglycerol esterified with the FAs C18:1 and C16:0 (GDG(18:1/16:0)) (Figure 9C) isolated from the brown seaweed *Petalonia bingbamiae* [63]. Likewise, the inhibition of MYT1 kinase by two GLs lipid species from unknown seaweed species were reported and these bioactive GLs species were identified as *sn*-1,2-dipalmitoyl-3- (*N*-palmitoyl-6-deoxy-6-amino-α-D-glucosyl)-glycerol and *sn*-1-palmitoyl-2-myristoyl-3- (*N*-stearyl-6-deoxy-6-aminoglucosyl)-glycerol (Figure 9D) [64]. The total synthesis of 1,2 dipalmitoyl-3-(*N*-palmitoyl-6 -amino-6 -deoxy-α-D-glucosyl)-*sn*-glycerol based on previous study [64], was achieved by Göllner and co-authors that confirmed those GLs lipids species as bioactive [65].

The species of GL isolated from the green seaweed *Avrainvillea nigricans*, named Nigricanoside A (Figure 9E), showed the capacity to arrest MCF-7 cells in mitosis, stimulating the polymerization of pure tubulin in vitro and thus inhibiting the proliferation of MCF-7 and HCT-116 cells [66]. The potent antimitotic activity of Nigricanoside A was seen without precedent among previously known GL.

#### 3.1.2. Anti-Inflammatory Activity

Inflammation is a multifactorial condition ubiquitously present in most diseases and particularly in non-communicable diseases. It involves a large number of identified mediators, comprising leukocyte cells that release specialized substances such as

pro-inflammatory cytokines [67] and high levels of nitric oxide (NO) in response to the inflammatory process [68].

NO is a potent pro-inflammatory mediator in over inflammation conditions [69]. On a small scale, and for research purposes, inhibition of NO, represents a protective effect of several anti-inflammatory compounds. The reduction in NO production from immune cells is assessed as a first step in the anti-inflammatory potential of natural products. Using this approach, several studies evaluated the anti-inflammatory activity of isolated and characterized seaweed lipid molecules (Table 2) including (2S)-1- *O*-eicosapentaenoyl-2-*O*-myristoyl-3-*O*-(6-sulfo-*α*-D-quinovopyranosyl)-glycerol SQDG (20:5/14:0), (2S)-1-*O*-eicosapentaenoyl-2-*O*-palmitoyl-3-*O*-(6-sulfo-*α*-D-quinovopyranosyl) glycerol SQDG(20:5/16:0), 1-*O*-eicosapentaenoyl-2-*O*-*trans*-3-hexadecanoyl-3-phospho-(1 glycerol)-glycerol PG(20:5/*trans*-16:1), 1-*O*-eicosapentaenoyl-2-*O*-palmitoyl-3-phospho-(1 glycerol)-glycerol PG(20:5/16:1), and 1,2-*bis*-*O*-eicosapentanoylglycero-3-phosphocholine PC(20:5/20:5) (Figure 10(A1–A3)) from the red seaweed *Palmaria palmata* [70]; and isolated galactolipid species from the red seaweed *Chondrus crispus*, such as (2S)-1,2-*bis*-*O*eicosapentaenoyl-3-*O*-*β*-D-galactopyranosylglycerol MGD(20:5/20:5), (2S)-1-*O*-eicosapentaenoyl-2-*O*-arachidonoyl-3-*O*-*β*-D-galactopyranosylglycerolMGDG(20:5/20:4), (2S)-1-*O*-eicosapentaenoyl-2-*O*-palmitoyl-3-*O*-*β*-D-galactopyranosylglycerol MGDG(20:5/16:0), (2S)-1-*O*-eicosapentaenoyl-2-*O*-palmitoyl-3-*O*-(*β*-D-galactopyranosyl-6-1-*α*-D-galactopyranosyl)-glycerol DGDG(20:5/16:0), (2S)-1,2-*bis*-*O*-arachidonoyl-3-*O*-*β*-D-galactopyranosylglycerol MGDG(20:4/20:4), (2S)-1-*O*arachidonoyl-2-*O*-palmitoyl-3-*O*-*β*-D-galactopyranosylglycerol MGDG(20:4/16:0), (2S)-1- *O*-arachidonoyl-2-*O*-palmitoyl-3-*O*-(*β*-D-galactopyranosyl-6-1*α*-D-galactopyranosyl)-glycerol DGDG(20:4/16:0), and (2S)-1-*O*-(6Z,9Z,12Z,15Z-octadecatetranoyl)-2-*O*-palmitoyl-3-*O*-*β*-D-galactopyranosylglycerol MGDG(18:4/16:0) (Figure 10(B1–B3)) [71], which showed significant NO inhibition through down-regulation of inducible Nitric Oxide Synthase (iNOS). PUFA side chains, mainly EPA and arachidonic acid (AA), esterified to polar lipid structure seem to be relevant for their potent NO inhibition. Curiously, isolated PUFA, such as EPA, AA, and DHA, showed less NO inhibitory activity when compared to their esterified forms in polar lipid [70,71].

**Figure 9.** Chemical structures of bioactive complex lipids reported with antitumor activity. (**A**) 1-*O*-(5Z, 8Z, 11Z, 14Z, 17Zeicosapentanoyl)-2-*O*-(6Z,9Z,12Z,15Z-octadecatetraenoyl)-3-*O*-*β*-D-galactopiranosyl-*sn*-glycerol MGDG (20:5/18:4) (brown seaweed *Fucus evanescence*); (**B**) 1-*O*-(palmitoyl)-2-*O*-(5Z, 8Z, 11Z, 14Z eicosatetraenoyl)-3-*O*-*β*-D-galactopyranosyl-glycerol MGDG (20:4/16:0) (red seaweed *Hydrolithon reinboldii*); (**C**) GDG (16:0, 18:1) (brown seaweed *Petalonia bingbamiae*) [63]; (**D**) *sn*-1,2-dipalmitoyl-3-(*N*-palmitoyl-6-deoxy-6-amino-*α*-D-glucosyl)-glycerol (1) and *sn*-1-palmitoyl-2-myristoyl-3-(*N*stearyl-6-deoxy-6-aminoglucosyl)-glycerol (2); (**E**) Nigricanoside A (green seaweed *Avrainvillea nigricans*).


**Table 1.** Lipid species extracted

 from seaweeds

 with

antitumor

 activities.

Extraction

 and

characterization

methodologies

 and cell lines used in in vitro

assays are


The capacity to inhibit phospholipase A2 (PLA2) has been linked to the efficacy for the treatment of inflammatory processes, since PLA2 hydrolyze membrane phospholipids releasing AA, the precursor of the pro-inflammatory mediators prostaglandins, thromboxanes, and leukotrienes [72,73]. Inhibition of PLA2 is the pharmacological mechanism of action of corticosteroids, a group of drugs with potent anti-inflammatory properties. The 7-methoxy-9-methylhexadeca-4,8-dienoic acid (MMHDA) (Figure 10C) isolated from the brown seaweed *Ishige okamurae* was tested in vitro for inhibition of PLA2 activity, and in vivo on edema and erythema induced in rat models. In both models, it demonstrated potent inhibitor of PLA2 activity and inflammation, with IC50 concentrations lower than the ones reported for rutin, a flavonoid model [74].

**Figure 10.** Chemical structures of bioactive complex lipids reported with anti-inflammatory activity: (**A1**) (2S)-1-*O*eicosapentaenoyl-2-*O*-myristoyl-3-*O*-(6-sulfo-*α*-D-quinovopyranosyl)-glycerol SQDG (20:5/14:0) (1); (2S)-1-*O*-eicosapentaenoyl-2-*O*-palmitoyl-3-*O*-(6-sulfo-*α*-D-quinovopyranosyl)-glycerol SQDG(20:5/16:0) (2); (**A2**) 1-*O*-eicosapentaenoyl-2-*O*-*trans*-3 hexadecanoyl-3-phospho-(1 -glycerol)-glycerol PG(20:5/*trans*-16:1) (3); 1-*O*-eicosapentaenoyl-2-*O*-palmitoyl-3-phospho-(1 glycerol)-glycerol PG(20:5/16:1) (4); (**A3**) 1,2-*bis*-*O*-eicosapentanoylglycero-3-phosphocholine PC(20:5/20:5) (red seaweed *Palmaria palmata*); (**B1**) (2S)-1,2-*bis*-*O*-eicosapentaenoyl-3-*O*-*β-*D-galactopyranosylglycerol MGDG(20:5/20:5) (1); (2S)-1- *O*-eicosapentaenoyl-2-*O*-arachidonoyl-3-*O-β-*D-galactopyranosylglycerol MGDG(20:5/20:4) (2); (2S)-1-*O*-eicosapentaenoyl-2-*O*-palmitoyl-3-*O-β-*D-galactopyranosylglycerol MGDG(20:5/16:0) (3); (2S)-1-O-eicosapentaenoyl-2*-O*-palmitoyl-3-*O*-(*β-*D-galactopyranosyl-6-1-*α*-D-galactopyranosyl)-glycerol DGDG (20:5/16:0)(4); (**B2**) (2S)-1,2*-bis*-*O*-arachidonoyl-3-*O-β-*Dgalactopyranosylglycerol MGDG(20:4/20:4) (5); (2S)-1-*O*-arachidonoyl-2-*O*-palmitoyl-3-*O-β-*D-galactopyranosylglycerol MGDG(20:4/16:0) (6); (2S)-1-*O*-arachidonoyl-2-*O*-palmitoyl-3-*O*-(*β-*D-galactopyranosyl-6-1-*α*-D-galactopyranosyl)-glycerol DGDG(20:4/16:0) (7); (**B3**) (2S)-1-*O*-(6Z,9Z,12Z,15Z-octadecatetranoyl)-2-*O*-palmitoyl-3-*O*-*β-*D-galactopyranosylglycerol MGDG (18:4/16:0) (red seaweed *Chondrus crispus*); (**C**) 7-methoxy-9-methylhexadeca-4,8-dienoic acid (MMHDA) (brown seaweed *Ishige okamurae*).

