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Whey, the main by-product of the cheese industry, is considered as an important pollutant due to its high chemical and biological oxygen demand. Whey, often considered as waste, has high nutritional value and can be used to obtain value-added products, although some of them need expensive enzymatic synthesis. An economical alternative to transform whey into valuable products is through bacterial or yeast fermentations and by accumulation during algae growth. Fermentative processes can be applied either to produce individual compounds or to formulate new foods and beverages. In the first case, a considerable amount of research has been directed to obtain biofuels able to replace those derived from petrol. In addition, the possibility of replacing petrol-derived plastics by biodegradable polymers synthesized during bacterial fermentation of whey has been sought. Further, the ability of different organisms to produce metabolites commonly used in the food and pharmaceutical industries (i.e., lactic acid, lactobionic acid, polysaccharides, etc.) using whey as growth substrate has been studied. On the other hand, new low-cost functional whey-based foods and beverages leveraging the high nutritional quality of whey have been formulated, highlighting the health-promoting effects of fermented whey-derived products. This review aims to gather the multiple uses of whey as sustainable raw material for the production of individual compounds, foods, and beverages by microbial fermentation. This is the first work to give an overview on the microbial transformation of whey as raw material into a large repertoire of industrially relevant foods and products.

Quinoa fermentation by lactic acid bacteria (LAB) is an interesting alternative to produce new bakery products with high nutritional value; furthermore, they are suitable for celiac patients because this pseudo-cereal contains no gluten. Growth and lactic acid production during slurry fermentations by Lactobacillus plantarum CRL 778 were greater in quinoa (9.8 log cfu/mL, 23.1 g/L) than in wheat (8.9 log cfu/mL, 13.9 g/L). Lactic fermentation indirectly stimulated flour protein hydrolysis by endogenous proteases of both slurries. However, quinoa protein hydrolysis was faster, reaching 40–100 % at 8 h of incubation, while wheat protein hydrolysis was only 0–20 %. In addition, higher amounts of peptides (24) and free amino acids (5 g/L) were determined in quinoa compared to wheat. Consequently, greater concentrations (approx. 2.6-fold) of the antifungal compounds (phenyllactic and hydroxyphenyllactic acids) were synthesized from Phe and Tyr in quinoa by L . plantarum CRL 778, an antifungal strain. These promising results suggest that this LAB strain could be used in the formulation of quinoa sourdough to obtain baked goods with improved nutritional quality and shelf life, suitable for celiac patients.

The consumers’ demand for food with high nutritional quality and free of chemical additives increases the need to look for new products and preservation strategies. Quinoa ( Chenopodium quinoa ) is an Andean pseudocereal highly appreciated because of its nutritional properties. Moreover, it is an optimal substrate for growing and production of improved amounts of antifungal compounds by Lactobacillus plantarum CRL 778. The aim of this work was to optimize a lactic ferment for packaged breads with improved nutritional value and prolonged shelf life by applying a statistical experimental design model. The addition of 30 % quinoa to the wheat semiliquid ferment (QWF) could highly improve the amino acids release (4.3 g/L) during fermentation. Moreover, this quinoa proportion was sufficient to obtain the same concentration of the antifungal compounds, phenyllactic and hydroxiphenyllactic acids (PLA and OH-PLA) as with 100 % quinoa (ca. 36 and 51 mg/L, respectively). Statistical model analysis showed that citrate and skimmed milk enhanced significantly all evaluated parameters specially PLA (ca. 71 mg/L), HO-PLA (ca. 75 mg/L), and lactate (27 g/L) with a p value <0.005. The synergic effects of higher antifungal compounds production, acid release, and pH decrease allowed lowering the amount (about 50 %) of the chemical preservative calcium propionate commonly added to bread. Moreover, these breads show increased shelf life.

In this work, the thermotolerance of Lactobacillus rhamnosus CRL1505, an immunobiotic strain, was studied as a way to improve the tolerance of the strain to industrial processes involving heat stress. The strain displayed a high intrinsic thermotolerance (55°C, 20 min); however, after 5 min at 60°C in phosphate buffer a two log units decrease in cell viability was observed. Different heat shock media were tested to improve the cell survival. Best results were obtained in the mediumcontaining inorganic salts (KH2PO4, Na2HPO4, MnSO4, and MgSO4) likely as using 10% skim milk. Flow cytometry analysis evinced 25.0% live cells and a large number of injured cells (59.7%) in the inorganic salts medium after heat stress. The morphological changes caused by temperature were visualized by transmission electronic microscopy (TEM). In addition, TEM observations revealed the presence of polyphosphate (polyP) granules in the cells under no-stress conditions. A DAPI-based fluorescence technique, adjusted to Gram-positive bacteria for the first time, was used to determine intracellular polyP levels. Results obtained suggest that the high initial polyP content in L. rhamnosus CRL 1505 together with the presence of inorganic salts in the heat shock medium improve the tolerance of the cells to heat shock. To our knowledge, this is the first report giving evidence of the relationship between polyP and inorganic salts in thermotolerance of lactic acid bacteria.

We have previously demonstrated that Lactobacillus reuteri CRL1098 soluble factors were able to reduce TNF-α production by human peripheral blood mononuclear cells. The aims of this study were to determine whether L. reuteri CRL1098 soluble factors were able to modulate in vitro the inflammatory response triggered by LPS in murine macrophages, to gain insight into the molecular mechanisms involved in the immunoregulatory effect, and to evaluate in vivo its capacity to exert anti-inflammatory actions in acute lung injury induced by LPS in mice. In vitro assays demonstrated that L. reuteri CRL1098 soluble factors significantly reduced the production of pro-inflammatory mediators (NO, COX-2, and Hsp70) and pro-inflammatory cytokines (TNF-α, and IL-6) caused by the stimulation of macrophages with LPS. NF-kB and PI3K inhibition by L. reuteri CRL1098 soluble factors contributed to these inhibitory effects. Inhibition of PI3K/Akt pathway and the diminished expression of CD14 could be involved in the immunoregulatory effect. In addition, our in vivo data proved that the LPS-induced secretion of the pro-inflammatory cytokines, inflammatory cells recruitment to the airways and inflammatory lung tissue damage were reduced in L. reuteri CRL1098 soluble factors treated mice, providing a new way to reduce excessive pulmonary inflammation.

We have demonstrated previously that a soluble factor (LrS) produced by Lactobacillus (L.) reuteri CRL 1098 modulates the inflammatory response triggered by lipopolysaccharide. In this study, the production of LrS by L. reuteri CRL 1098 was realized through two steps: i) bacterial biomass production, ii) LrS production, where the bacterial biomass was able to live but did not proliferate. Therefore, the simultaneous evaluation of the effect of different factors on the growth and LrS production was performed. Biomass production was found to be dependent mainly on culture medium, while LrS production with anti-inflammatory activity depended on culture conditions of the biomass such as pH, agitation and growth phase. The L. reuteri CRL 1098 biomass and LrS production in the optimized culture media designed for this work reduced the complete process cost by approximately 95%, respectively to laboratory scale cost.

The high nutritional value of whey makes it an interesting substrate for the development of fermented foods. The aim of this work was to evaluate the growth and proteolytic activity of sixty-four strains of lactic acid bacteria in whey to further formulate a starter culture for the development of fermented whey-based beverages. Fermentations were performed at 37°C for 24 h in 10 and 16% (w/v) reconstituted whey powder. Cultivable populations, pH, and proteolytic activity (o-phthaldialdehyde test) were determined at 6 and 24 h incubation. Hydrolysis of whey proteins was analysed by Tricine SDS-PAGE. A principal component analysis (PCA) was applied to evaluate the behaviour of strains. Forty-six percent of the strains grew between 1 and 2 Δlog CFU/ml while 19% grew less than 0·9 Δlog CFU/ml in both reconstituted whey solutions. Regarding the proteolytic activity, most of the lactobacilli released amino acids and small peptides during the first 6 h incubation while streptococci consumed the amino acids initially present in whey to sustain growth. Whey proteins were degraded by the studied strains although to different extents. Special attention was paid to the main allergenic whey protein, β-lactoglobulin, which was degraded the most by Lactobacillus acidophilus CRL 636 and Lb. delbrueckii subsp. bulgaricus CRL 656. The strain variability observed and the PCA applied in this study allowed selecting appropriate strains able to improve the nutritional characteristics (through amino group release and protein degradation) and storage (decrease in pH) of whey.

Certain lactic acid bacteria, especially heterofermentative strains, are capable to produce mannitol under adequate culture conditions. In this study, mannitol production by Lactobacillus reuteri CRL 1101 and Lactobacillus fermentum CRL 573 in modified MRS medium containing a mixture of fructose and glucose in a 6.5:1.0 ratio was investigated during batch fermentations with free pH and constant pH 6.0 and 5.0. Mannitol production and yields were higher under constant pH conditions compared with fermentations with free pH, the increase being more pronounced in the case of the L. fermentum strain. Maximum mannitol production and yields from fructose for L. reuteri CRL 1101 (122 mM and 75.7 mol%, respectively) and L. fermentum CRL 573 (312 mM and 93.5 mol%, respectively) were found at pH 5.0. Interestingly, depending on the pH conditions, fructose was used only as an alternative external electron acceptor or as both electron acceptor and energy source in the case of the L. reuteri strain. In contrast, L. fermentum CRL 573 used fructose both as electron acceptor and carbon source simultaneously, independently of the pH value, which strongly affected mannitol production by this strain. Studies on the metabolism of these relevant mannitol-producing lactobacilli provide important knowledge to either produce mannitol to be used as food additive or to produce it in situ during fermented food production. [PUBLICATION ABSTRACT]

PURPOSE OF WORK: To clone, express and characterize uroporphyrinogen III synthase/methyltransferase gene (cobA/hemD) from Lactobacillus reuteri. Some strains of Lb. reuteri produce cobalamin (vitamin B12). Cobalamin biosynthesis relies on the sequential action of more than 25 enzymes in a complex metabolic pathway. We have cloned, expressed and characterized the gene in Lb. reuteri that codes for the S-adenosy L-methionine uroprophyrinogen III methyltransferase/synthase (CobA/HemD), a key bifunctional enzyme in the biosynthesis of cobalamin and other tetrapyrrols.

Ability of industrially relevant species of thermophilic lactobacilli strains to hydrolyze proteins from animal (caseins and β-lactoglobulin) and vegetable (soybean and wheat) sources, as well as influence of peptide content of growth medium on cell envelope-associated proteinase (CEP) activity, was evaluated. Lactobacillus delbrueckii subsp. lactis (CRL 581 and 654), L. delbrueckii subsp. bulgaricus (CRL 454 and 656), Lactobacillus acidophilus (CRL 636 and 1063), and Lactobacillus helveticus (CRL 1062 and 1177) were grown in a chemically defined medium supplemented or not with 1 % Casitone. All strains hydrolyzed mainly β-casein, while degradation of α s -caseins was strain dependent. Contrariwise, κ-Casein was poorly degraded by the studied lactobacilli. β-Lactoglobulin was mainly hydrolyzed by CRL 656, CRL 636, and CRL 1062 strains. The L. delbrueckii subsp. lactis strains, L. delbrueckii subsp. bulgaricus CRL 656, and L. helveticus CRL 1177 degraded gliadins in high extent, while the L. acidophilus and L. helveticus strains highly hydrolyzed soy proteins. Proteinase production was inhibited by Casitone, the most affected being the L. delbrueckii subsp. lactis species. This study highlights the importance of proteolytic diversity of lactobacilli for rational strain selection when formulating hydrolyzed dairy or vegetable food products.