*6.1. Chemistry and Obtaining of Prebiotic Compounds from Seaweeds*

Some prebiotics (or prebiotic candidates) naturally occurring in seaweeds are saccharides presenting a degree of polymerization (DP) within 2 and 9 (sometimes within 8 and 20). Some researchers consider up to 25 sugar residues [122]. Such oligo/polysaccharides include galacto-oligosaccharides (GOS), agarose-derived oligosaccharides (AGAROS), xylo-oligosaccharides (XOS), neoagaro-oligosaccharides (NAOS), alginate-derived oligosaccharides (ALGOS), arabinoxylans, galactans and glucans [114]. Although not all of them fulfill all the criteria requested for a compound to be considered as prebiotic (i.e., be refractingto hydrolysis and absorption in the upper part of the gastro-intestinal tract, selective promotion of bifidobacteria and/or lactobacilli in the colon, beneficial effect of their fermentation products, stability towards technological processes when incorporated into food products) [121], none of them is degraded by enzymes in the first part of the gastro-intestinal tract. Therefore, they achieve at least one of the requested criteria and take part of the dietary fiber.

Prebiotic oligosaccharides from seaweeds can be obtained by the hydrolysis of naturally occurring polysaccharides. This process has encouraged the development of new extraction techniques, as algae polysaccharides have a wide variety of chemical bonds and conformations, including α or β bonds, *cis* or *trans* configurations, d or l, or *R* or *S* chiral centers. This issue is of great importance since certain bonds can be hydrolyzed with enzymes, whereas others require other techniques as no natural enzymes are able to hydrolyze them [120,123]. Therefore, it is necessary to look for other techniques to obtain oligosaccharides from polysaccharides. Some authors [122] summarized the characteristics of different techniques, as well as their suitability to be used for the hydrolysis of some polysaccharides. Hence, the main non-enzymatic hydrolytic techniques used at an industrial level are ultrasound (in the case of carrageenans, agarose and xylan), microwaves (for hydrolysis of exopolysaccharides obtained

from *Porphyridium cruentum*), use of free radicals (for fucoidan and algae galactan), or the application of diluted acids (for example, phosphoric acid) [124–130].

In the case of acids, they can be used as hydrolytic agents only for neutral polysaccharides, namely fucoidans, carrageenans, or galactans. In addition, certain components can be lost during the process and most of the glycosidic bonds are not specifically hydrolyzed, thus giving rise to various low molecular weight derivatives. In spite of that, phosphoric acid is capable of effectively hydrolyzing polysaccharides of some algae species, such as *Chlorella vulgaris* and *Spirulina platensis* [128], mostly composed of uronic acids and giving rise to interesting oligosaccharides with potential prebiotic properties [122].

The use of free radicals is another effective technique for the hydrolysis of polysaccharides, since it does not affect the structure of the compounds when it is correctly performed [122]. Other authors have demonstrated that this technique can be applied for the hydrolysis of fucoidans in low molecular weight compounds (≈8 kDa), although then it is necessary to apply other procedures for the purification of the obtained products [129]. Additionally, the obtained products demonstrated a better anticoagulant capacity with respect to the original polysaccharide [129].

The physical technique of microwaves also gives rise to glucidic compounds of low molecular weight (≈12 kDa) from natural polysaccharides extracted from algae. The obtained products do not undergo structural changes and have a surprisingly enhanced immunomodulatory and anticancer capacities with regard to the original substrate. The higher solubility of the products (with lower molecular weight) was remarked as a possible explanation for such observations [131]. Therefore, the use of this technique offers several clear advantages: it is economical, easy to use, non-toxic and is green, since the energetic and time consumption are not environmentally harmful [132]. All these techniques can be optimized to guarantee the highest results [133].
