**3. Antinutritional factors and microbial degradation**

Legumes contain several ANFs, such as raffinose, phytic acid, condensed tannins, saponins, alkaloids, lectins, pyrimidine glycosides, and protease inhibitors [17]. Overall, ANFs decrease the bioaccessibility and bioavailability of other nutrients, and, in some cases, are responsible for adverse reactions to the ingestion.

The content of raffinose-family oligosaccharides (RFOs, raffinose, verbascose, and stachyose) in legumes ranges from 1 to 6% with stachyose as the most abundant compound [18]. While in cereals, it is commonly lower than 1.5%, with raffinose as the sole or the most abundant compound [19, 20]. RFOs are nondigestible oligosaccharides that may result in adverse digestive symptoms when about 15 g/person per day are exceeded [21], a threshold that is readily reached in legume-based diets. Raffinose and RFO are indeed fermented by the intestinal microbiota with abundant gas production, causing discomfort and flatulence.

Phytic acid is the main storage compound for phosphorous and minerals in cereal and legume seeds. In legumes, its concentration can reach 20 g/kg [22, 23]. Phytic acid and divalent minerals (e.g., Ca2+, Zn2+ and iron) form stable complexes (phytates) that are insoluble and not hydrolyzed in the gastrointestinal tract, thus reducing the bioavailability of minerals for the monogastrics. Ca2+ and Zn2+ deficiencies are commonly observed in developing countries, and complexation of dietary minerals by phytates in plant-derived foods contributes to the mineral deficiency [17]. Iron uptake from plant-derived foods is impeded not only by complexation with phytate but also by complexation with condensed tannins [24, 25].

Proanthocyanidins, gallotannins. and ellagitannins, commonly referred to as tannins, are phenolic compounds that occur in a wide variety of plant foods. Their presence in cereals and legumes is dependent on the plant species and the cultivar [26]. Tannins impart bitter taste, reduce protein and starch digestibility by inhibition of pancreatic enzymes, and reduce iron uptake [26, 27]. The presence of tannins reduces the caloric content and the glycemic index of foods [28], but the abundance in diet reduces the supply of macro- and micro-nutrients.

Lectins and specific inhibitors of digestive enzymes (proteases and amylases) further reduce the digestibility of starch and proteins in legumes [26, 29].

Some ANFs are heat-labile (e.g., protease inhibitors and lectins) and easily removed by thermal treatments. Nevertheless, phytic acid, raffinose, tannins, and saponins are rather thermostable. Dehulling, soaking, air classification, extrusion, steaming, and pregelatinization are the main technological options for decreasing

*Fermentation as Strategy for Improving Nutritional, Functional, Technological, and Sensory… DOI: http://dx.doi.org/10.5772/intechopen.102523*

the negative impact of ANF on legume consumption [30–32]. Nevertheless, biological methods such as germination, enzyme treatments, and especially, fermentation seem to be more efficient [30, 31, 33, 34].

Proteolysis, enzyme inhibition due to acidification, acid activation of flour endogenous enzymes (e.g., phytases) and/or microbial enzyme activities (e.g., α-galactosidase, β-glucosidase, phytases, tannases) are responsible for the inactivation of most ANFs.

Raffinose family oligosaccharides are hydrolyzed through the activity of α-galactosidases, levansucrase, and sucrose-phosphorylase activities of lactic acid bacteria [35, 36] or corresponding enzymes of fungal cultures; their removal in legume fermentations has been amply reported [37].

In cereal matrices [22], the phytase activity is often sufficient to degrade phytates, especially in acidic conditions [18, 38]. Therefore, phytate degradation in LAB-fermented matrices spontaneously occurs without microbial enzymes involvement [18]. The optimal pH for the activity of the cereal phytases corresponds to 5.5; nevertheless, phytases are still active at pH levels lower than those commonly reached by sourdough (3.8–4.2) [18]. Sourdough fermentation and other types of traditional bioprocesses involving LAB (e.g., fermentations for production of cereal porridges or beverages) allow the increase of the mineral bioavailability [39]. Compared with that found in cereals, the phytase activity in legumes is poor [22, 40]. Nevertheless, pretreatments and processing conditions including fractionation, germination, soaking, thermal treatments, and fermentation drastically decrease phytate levels in legumes [41]. In many spontaneously fermented legume products, substrate-derived phytases are inactivated, and phytate degradation is achieved by fermentation with bacilli or fungal cultures, for example, *Rhizopus stolonifer* or *Aspergillus oryzae*, which hydrolyze phytate with extracellular enzymes [42, 43].

Metabolism of tannins or other polyphenols by LAB was deeply characterized only in a few fermented plant-derived matrices [44, 45]. *Lactiplantibacillus plantarum*, *Lactiplantibacillus paraplantarum*, *Lactiplantibacillus pentosus* have been identified, among the LAB, as the species that could decrease the tannins concentration through their tannases (tannin acyl hydrolase, EC 3.1.1.20) [46–48]. However, characterization of fermented cassava allowed identifying uncommon tannase producers such as *Weissella cibaria* and *Leuconostoc mesenteroides* ssp. *mesenteroides* [49]. Most of tannase producers were found in fermented vegetables but also in human feces. In *L. plantarum*, tannase is very well characterized. Its activity was demonstrated and characterized by Rodríguez et al. [47], and genetic analysis showed that it constitutes a novel family of tannases [50]. LAB tannases are intracellular. Genes involved in tannins degradation are regulated in a coordinated way and are inducible by tannin and other phenolic compounds [51].

The lactic fermentation of grass pea (*Lathyrus sativus*) with *L. plantarum* lowered the levels of phytic acid and trypsin inhibitory activity [52]. Selected strains of *L. plantarum* and *Levilactobacillus brevis* decreased the content of raffinose up to circa 64% during sourdough fermentation of different legume flours. Sourdoughs made with different legume flours (bean, lentil, pea, grass-pea, chickpea) contained an increased phytase activity compared with the unfermented controls [34]. The combination of legume sprouting and sourdough fermentation decreased the content of phytic acid, condensed tannins and raffinose, and trypsin inhibitory activity [53, 54].

Besides the abovementioned ANFs, faba bean is rich in two glucosidic aminopyrimidine derivatives, vicine and convicine, which, upon hydrolysis of the β-glucosidic bond, generate the aglycones divicine (2,6-diamino-4,5-dihydroxypyrimidine) and isouramil (6-amino-2,4,5-trihydroxypyrimidine), respectively [55]. Divicine and isouramil trigger favism disease in susceptible individuals. Technological processes (air classification, roasting, and boiling) and selection of cultivars with low content of such compounds seemed to be only in part effective [55, 56]. On the contrary, β-glucosidase from LAB effectively degraded the pyrimidine glycosides from faba bean suspension and flour [30]. When used as starter to ferment fava bean flour, *L. plantarum* expressed β-glucosidase activity and decreased the content of vicine and convicine by more than 90%. The degradation was complete after 48 h of fermentation, and aglycone derivatives were not detectable [57]. Similar results were obtained when flours from different faba bean accessions collected from the Mediterranean area were subjected to the LAB fermentation [58]. *Ex-vivo* hemolysis assays on human blood confirmed the lack of toxicity of the fermented fava bean [57].
