γ-aminobutyric acid (GABA) is a four-carbon amino acid, that acts as a major inhibitory neurotransmitter in the central nervous system. Among nineteen lactic acid bacteria strains, isolated from ...Andean amaranth (A) and Real Hornillos quinoa (Qr) sourdoughs, Lactobacillus brevis CRL 1942 was the most efficient microorganism for the conversion of 53 mM monosodium glutamate (MSG) to GABA, reaching 50 mM after 96 h cultivation. GABA production was enhanced by optimizing culture conditions, such as incubation temperature, time and MSG concentration. A gradual increase of GABA yield was observed at MSG concentrations rising from 0 to 270 mM. In addition, a higher GABA content was observed at 30 °C. GABA production occurred in a time-dependent manner, and greatest amount (∼255 mM) was yielded after 48 h in cells grown in MRS with 270 mM MSG at 30 °C, with a conversion rate of ∼90%. Cell growth was not affected by MSG addition, implying that the difference in GABA levels could not be attributed to differences in cell numbers. However, addition of glutamate increased viability, indicating a correlation between survival and GABA production. Novel information about LAB with GABA-producing ability is an important breakthrough for the development of health-promoting functional foods.
•Lactobacillus brevis CRL 1942, isolated from quinoa sourdough, was able to produced high levels of GABA.•The optimal conditions for GABA production by Lb. brevis CRL 1942 were 270 mM of MSG and 30 °C for 48 h.•Lb. brevis CRL 1942 showed a conversion rate of ∼90% from the supplied MSG.•The obtained GABA levels are one of the highest among lactobacilli grown in batch culture.
Lactic acid bacteria (LAB) are capable of converting carbohydrate substrates into organic acids (mainly lactic acid) and producing a wide range of metabolites. Due to their interesting beneficial ...properties, LAB are widely used as starter cultures, as probiotics, and as microbial cell factories. Exploring LAB present in unknown niches may lead to the isolation of unique species or strains with relevant technological properties. Autochthonous rather than allochthonous starter cultures are preferred in the current industry of fermented food products, due to better adaptation and performance of autochthonous strains to the matrix they originate from. In this work, the lactic microbiota of eight different wild tropical types of fruits and four types of flowers were studied. The ability of the isolated strains to produce metabolites of interest to the food industry was evaluated. The presence of 21 species belonging to the genera
, and
was evidenced by using culture-dependent techniques. The isolated LAB corresponded to 95 genotypically differentiated strains by applying rep-PCR and sequencing of the 16S rRNA gene; subsequently, representative strains of the different isolated species were studied for technological properties, such as fast growth rate and acidifying capacity; pectinolytic and cinnamoyl esterase activities, and absence of biogenic amine biosynthesis. Additionally, the strains' capacity to produce ethyl esters as well as mannitol was evaluated. The isolated fruit- and flower-origin LAB displayed functional properties that validate their potential use in the manufacture of fermented fruit-based products setting the background for the design of novel functional foods.
Gamma aminobutyric acid (GABA) is a non-protein amino acid that is widely distributed in nature and its physiological importance goes beyond its role as an inhibitory neurotransmitter of the central ...nervous system in mammals. Since microbial fermentation is one of the most promising methods to obtain GABA, the production of this metabolite by several strains of lactic acid bacteria isolated from quinoa and amaranth sourdoughs was investigated. Lactobacillus brevis CRL 2013 produced the highest GABA levels, reaching 265 mM when optimal culture conditions were set up. The fermentative profile showed that CRL 2013 was able to catabolize carbohydrates through the phosphoketolase pathway yielding variable amounts of lactic acid, acetate and ethanol, which depended on the type of carbon source available and the presence of external electron acceptors such as fructose. Enhanced growth parameters and low GABA synthesis were associated to pentose fermentation. This impairment on GABA production machinery was partially overpassed by the addition of ethanol to the culture media. These results support the potential use of L. brevis CRL 2013 as a starter culture for the manufacture of GABA-enriched functional foods and provide further insights to the understanding of the GAD system regulation in lactic acid bacteria.
•L. brevis CRL 2013, isolated from quinoa sourdough, produced high levels of GABA.•Final GABA production is among the highest for lactobacilli grown in batch culture.•Enhanced growth rates and low GABA synthesis were associated to pentose fermentation.•The impairment on GABA production was partially restored by the addition of ethanol.
Mannitol is a natural low-calorie sugar alcohol produced by certain (micro)organisms applicable in foods for diabetics due to its zero glycemic index. In this work, we evaluated mannitol production ...and yield by the fruit origin strain
Fructobacillus tropaeoli
CRL 2034 using response surface methodology with central composite design (CCD) as optimization strategy. The effect of the total saccharide (glucose + fructose, 1:2) content (TSC) in the medium (75, 100, 150, 200, and 225 g/l) and stirring (S; 50, 100, 200, 300 and 350 rpm) on mannitol production and yield by this strain was evaluated by using a 2
2
full-factorial CCD with 4 axial points (
α
= 1.5) and four replications of the center point, leading to 12 random experimental runs. Fermentations were carried out at 30 °C and pH 5.0 for 24 h. Minitab-15 software was used for experimental design and data analyses. The multiple response prediction analysis established 165 g/l of TSC and 200 rpm of S as optimal culture conditions to reach 85.03 g/l 95% CI (78.68, 91.39) of mannitol and a yield of 82.02% 95% CI (71.98, 92.06). Finally, a validation experiment was conducted at the predicted optimum levels. The results obtained were 81.91 g/l of mannitol with a yield of 77.47% in outstanding agreement with the expected values. The mannitol 2-dehydrogenase enzyme activity was determined with 4.6–4.9 U/mg as the highest value found. To conclude,
F. tropaeoli
CRL 2034 produced high amounts of high-quality mannitol from fructose, being an excellent candidate for this polyol production.
Several plants, fungi, algae, and certain bacteria produce mannitol, a polyol derived from fructose. Mannitol has multiple industrial applications in the food, pharmaceutical, and medical industries, ...being mainly used as a non-metabolizable sweetener in foods. Many heterofermentative lactic acid bacteria synthesize mannitol when an alternative electron acceptor such as fructose is present in the medium. In previous work, we reported the ability of Lactobacillus reuteri CRL 1101 to efficiently produce mannitol from sugarcane molasses as carbon source at constant pH of 5.0; the activity of the enzyme mannitol 2-dehydrogenase (MDH) responsible for the fructose conversion into mannitol being highest during the log cell growth phase. Here, a detailed assessment of the MDH activity and relative expression of the mdh gene during the growth of L. reuteri CRL 1101 in the presence of fructose is presented. It was observed that MDH was markedly induced by the presence of fructose. A direct correlation between the maximum MDH enzyme activity and a high level of mdh transcript expression during the log-phase of cells grown in a fructose-containing chemically defined medium was detected. Furthermore, two proteomic approaches (2DE and shotgun proteomics) applied in this study confirmed the inducible expression of MDH in L. reuteri. A global study of the effect of fructose on activity, mdh gene, and protein expressions of MDH in L. reuteri is thus for the first time presented. This work represents a deep insight into the polyol formation by a Lactobacillus strain with biotechnological potential in the nutraceutics and pharmaceutical areas.
The nucleotide sequences of plasmids pRC12 (12,342 bp; GC 43.99%) and pRC18 (18,664 bp; GC 34.33%), harbored by the bacteriocin-producer Lactobacillus curvatus CRL 705, were determined and analyzed. ...Plasmids pRC12 and pRC18 share a region with high DNA identity (> 83% identity between RepA, a Type II toxin-antitoxin system and a tyrosine integrase genes) and are stably maintained in their natural host L. curvatus CRL 705. Both plasmids are low copy number and belong to the theta-type replicating group. While pRC12 is a pUCL287-like plasmid that possesses iterons and the repA and repB genes for replication, pRC18 harbors a 168 amino acid replication protein affiliated to RepB, which was named RepB'. Plasmid pRC18 also possesses a pUCL287-like repA gene but it was disrupted by an 11 kb insertion element that contains RepB', several transposases/IS elements, and the lactocin Lac705 operon. An Escherichia coli / Lactobacillus shuttle vector, named plasmid p3B1, carrying the pRC18 replicon (i.e. repB' and replication origin), a chloramphenicol resistance gene and a pBluescript backbone, was constructed and used to define the host range of RepB'. Chloramphenicol-resistant transformants were obtained after electroporation of Lactobacillus plantarum CRL 691, Lactobacillus sakei 23K and a plasmid-cured derivative of L. curvatus CRL 705, but not of L. curvatus DSM 20019 or Lactococcus lactis NZ9000. Depending on the host, transformation efficiency ranged from 102 to 107 per μg of DNA; in the new hosts, the plasmid was relatively stable as 29-53% of recombinants kept it after cell growth for 100 generations in the absence of selective pressure. Plasmid p3B1 could therefore be used for cloning and functional studies in several Lactobacillus species.
Legumes are an important protein source in developing countries and their flours represent an attractive alternative for the manufacture of gluten free products. In the present study, 4 kidney bean ...varieties (Alubia, Pallar, Black and Red beans) commonly cultivated in northwestern Argentina, were milled and spontaneously fermented in order to isolate and select autochthonous lactic acid bacteria (LAB) with relevant technological and functional properties for usage as starter cultures. Twelve doughs were fermented with daily back-slopping at 37°C for 6days and evolution of total mesophiles, lactic acid bacteria, and yeasts and molds populations were followed by plate counting. A combination of phenotypic and genotypic methods including (GTG)5-based PCR fingerprinting and 16S rRNA gene sequencing were used to differentiate and identify the isolated LAB to species level. LAB counts ranged from around 0.89±0.81 to 8.74±0.03logcfu/g with a pH decline from 6.4 to 3.9 throughout fermentation. Four genera and nine species of LAB: Enterococcus durans, E. faecium, E. mundtii, E. casseliflavus; Lactobacillus rhamnosus, Lactococcus garvieae, Weissella cibaria and W. paramesenteroides were found on kidney beans. Twenty five LAB strains were assessed for their abilities to grow on kidney bean extracts, acidifying capacities (pH and acidification rates), amylolytic, proteolytic, tannase and gallate decarboxylase activities as well as pathogens inhibition by antimicrobials. Based on these properties E. durans CRL 2178 and W. paramesenteroides CRL 2182 were inoculated singly and combined in Alubia kidney bean flour and fermented for 24h at 37°C. LAB strains were beneficial for removing trypsin inhibitors and tannins from sourdoughs and for improving amino acids and phenolics contents, increasing the antioxidant activities of kidney bean matrices. Selected strains have potential as starter cultures for obtaining fermented bean products with high nutritional and functional quality.
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•Argentinean kidney beans were microbiologically analyzed for LAB isolation.•Nine LAB species were identified by Rep-PCR fingerprinting and 16S rDNA sequencing.•Some strains have potential to improve nutritional quality and safety of legumes.•E. durans CRL 2178 and W. paramesenteroides CRL 2182 were selected as legume starters.•Selected strains improved kidney beans flours by fermentation.
, the type species of the genus
, is widely recognized as the primary starter culture in the dairy industry due to its proteolytic activity, which enables it to growth in milk. In this study, a ...comprehensive genomic analysis of the proteolytic system was conducted on
strains. The analysis included 27 genomes of
, with a specific focus on the key enzyme involved in this system, the cell envelope-associated proteinase (CEP). The amino acid sequences, as well as the protein-structure prediction of the CEPs, were compared. Additionally, syntenic analysis of the genomic locus related to the CEPs revealed high conservation in
subsp.
strains, while
subsp.
strains exhibited greater variability, including the presence of insertion sequences, deletions, and rearrangements. Finally, the CEP promoter region and putative regulatory elements responsible for controlling the expression of the proteolytic system in lactobacilli were investigated. Our genomic analysis and
characterization of the CEPs contribute to our understanding of proteolytic activity and the potential applications of these lactic acid bacteria in the dairy industry. Further research in this area will expand our knowledge and potential practical uses of these findings.
Gamma-aminobutyric acid (GABA) plays a key role in mammals as the major inhibitory neurotransmitter of the central nervous system. Although GABA may not be able to cross the human blood-brain ...barrier, it was approved as a food ingredient because of its benefits to the host after oral administration including anti-hypertensive, anti-depressant and anti-inflammatory activities. Considering the current trend toward the development of new functional and natural products and that microbial fermentation is one of the most promising methods to produce this non-protein amino acid, the
production of GABA through fermentation of strawberry and blueberry juices by the efficient GABA producer strain,
(formerly known as
) CRL 2013, was evaluated. A high GABA production (262 mM GABA) was obtained after fermenting strawberry juice supplemented with yeast extract for 168 h, being GABA yield significantly higher in strawberry juices than in the blueberry ones. Thus, GABA-enriched fermented strawberry juice (FSJ) was selected to carry out
and
studies. The
functional analysis of the GABA-enriched FSJ demonstrated its ability to significantly decrease the expression of
gene in LPS stimulated RAW 264.7 macrophages. In addition,
studies in mice demonstrated that both,
CRL 2013 and the GABA-enriched FSJ were capable of reducing the levels of peritoneal, intestinal and serum TNF-α, IL-6, and CXCL1, and increasing IL-10 and IFN-γ in mice exposed to an intraperitoneal challenge of LPS. Of note, the GABA-enriched FSJ was more efficient than the CRL 2013 strain to reduce the pro-inflammatory factors and enhance IL-10 production. These results indicated that the CRL 2013 strain exerts anti-inflammatory effects in the context of LPS stimulation and that this effect is potentiated by fermentation. Our results support the potential use of
CRL 2013 as an immunomodulatory starter culture and strawberry juice as a remarkable vegetable matrix for the manufacture of GABA-enriched fermented functional foods capable of differentially modulating the inflammatory response triggered by TLR4 activation.
•The allergenic whey protein BLG was highly hydrolysed by L. bulgaricus CRL 454.•The strain cleaved the three main epitopes of BLG.•Antioxidant and ACE inhibitor bioactive peptides can be released by ...this strain.•L. bulgaricus CRL454 could be used for formulating novel functional foods.
Whey, a cheese by-product used as a food additive, is produced worldwide at 40.7 million tons per year. β-Lactoglobulin (BLG), the main whey protein, is poorly digested and is highly allergenic. We aimed to study the contribution of Lactobacillus delbrueckii subsp. bulgaricus CRL 454 to BLG digestion and to analyse its ability to degrade the main allergenic sequences of this protein. Pre-hydrolysis of BLG by L. delbrueckii subsp. bulgaricus CRL 454 increases digestion of BLG assayed by an in vitro simulated gastrointestinal system. Moreover, peptides from hydrolysis of the allergenic sequences V41-K60, Y102-R124, C121-L140 and L149-I162 were found when BLG was hydrolysed by this strain. Interestingly, peptides possessing antioxidant, ACE inhibitory, antimicrobial and immuno-modulating properties were found in BLG degraded by both the Lactobacillus strain and digestive enzymes. To conclude, pre-hydrolysis of BLG by L. delbrueckii subsp. bulgaricus CRL 454 has a positive effect on BLG digestion and could diminish allergenic reactions.
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