•The crude polysaccharides (EPS) from L. helveticus MB2-1 have high viscosity.•Isolation and characterization of three EPS fractions from L. helveticus MB2-1.•Three purified EPS fractions had similar ...average molecular weight (about 2×105Da).•The EPS exhibited high antioxidant and metal ion chelating activities in vitro.•The acidic EPS from L. helveticus MB2-1 had potentials as natural antioxidant.
In the present study, the crude polysaccharides (EPS) from Lactobacillus helveticus MB2-1 were fermented with reconstituted whey medium and further fractionated on DEAE-52 cellulose and Sephadex G-100 chromatography to afford three purified fractions of EPS-1, EPS-2 and EPS-3. Then, their preliminary structures and antioxidant activities in vitro were investigated. Three purified EPS fractions (EPS-1, EPS-2 and EPS-3) had similar molecular weight distributions (about 2×105Da), and it was composed of galactose, glucose and mannose with a molar ratio of 1.33:2.75:1.00, 1.00:1.43:9.34 and 1.17:1.00:2.96, respectively. In addition, the antioxidant activity of EPS-1, EPS-2 and EPS-3 was evaluated with the in vitro scavenging abilities. The results indicated that both crude and purified EPS showed strong scavenging activities on three kinds of radical and chelating activities on ferrous ion, and their antioxidant activities decreased in the order of crude EPS>EPS-3>EPS-2>EPS-1.
Bioactive compounds released during milk fermentation by proteolytic cleavage of milk proteins have a role beyond their nutritional importance. This study assessed the proteolytic activity of ...Lactobacillus helveticus strains ASCC953, ASCC474, ASCC1188, and ASCC1315 and their ability to release bioactive compounds capable of exerting antioxidative and in vitro anticarcinogenic properties during incubation at 37°C in reconstituted skim milk. The performance of these strains was not affected by the pH decline during fermentation. Soluble extracts of fermented milk by L. helveticus 474 showed the highest free radical (1,1-diphenyl-2-picrylhydrazyl) scavenging activity at 12 h of fermentation, followed by a significant reduction of this activity at 24 h compared with the other strains and control (untreated milk). Skim milk fermented by L. helveticus strains contained compounds with anti-colon cancer activity at varied levels during fermentation. The activity (19.03–50.98% growth inhibition) was greatest in the extract obtained after 12 h of fermentation, which markedly declined (5.4–9.94%) at the end of fermentation. Lactobacillus helveticus 1315 released compounds into the skim milk supernatant with a greater growth inhibition (50.98%) on colon cancer HT-29 cell line than the other strains. More importantly, these compounds had no significant inhibition effect on normal, primary colon cells T4056. Whereas these results suggest that milk fermented by L. helveticus strains may release bioactive compounds with important multifunctional properties, the characteristics and activities of these compounds appear highly strain- and fermentation time-dependent.
ABSTRACT
For the present study, we collected 22 Lactobacillus helveticus strains from different dairy (n = 10) and cereal (n = 12) fermentations to investigate their biodiversity and to uncover ...habitat-specific traits. Biodiversity was assessed by comparison of genetic fingerprints, low-molecular-weight subproteomes, metabolic and enzymatic activities, growth characteristics and acidification kinetics in food matrices. A clear distinction between the dairy and cereal strains was observed in almost all examined features suggesting that the different habitats are domiciled by different L. helveticus biotypes that are adapted to the specific environmental conditions. Analysis of the low-molecular-weight subproteome divided the cereal isolates into two clusters, while the dairy isolates formed a separate homogeneous cluster. Differences regarding carbohydrate utilization were observed for lactose, galactose, sucrose and cellobiose as well as for plant-derived glucosides. Enzymatic differences were observed mainly for ß-galactosidase and ß-glucosidase activities. Further, growth temperature was optimal in the range from 33 to 37°C for the cereal strains, whereas the dairy strains showed optimal growth at 40°C. Taken together, adaptation of the various biotypes results in a growth benefit in the particular environment. Acidification and growth tests using either sterile skim milk or a wheat flour extract confirmed these results. Differentiation of these biotypes and their physiological characteristics enables knowledge-based starter culture development for cereal versus dairy products within one species.
Biodiversity assessment of dairy- and cereal-derived Lactobacillus helveticus isolates based on genotypic, phenotypic and physiological analyses revealed significant habitat adaptations that are highly relevant for knowledge-based starter culture development.
•This study aims to evaluate the contribution of chitosan nanoparticles to enhance the antimicrobial activity.•Nisin and Natamycin will be applied either in the free-state or loaded on chitosan ...nanoparticles.•Disc diffusion assay, Lb. helveticus crude CFS recorded the highest potency against all bacterial indicators.•The recorded MIC values were found to be higher in case of tested bacteria than yeast and fungus.•CFS with HS–GC–MS showed that Lb. helveticus produces a wide range of antimicrobial& health-promoting substances.
This research attempted to inspect the contribution of lactic acid bacteria (LAB) with nanoparticle application in antimicrobial enhancement. Seven lactic acid cultures-free supernatants (CFSs) in both free and nanoparticles-loaded states were examined against seven foodborne microorganisms. Lactobacillus helveticus followed by Lactobacillus Plantarum possessed considerable antimicrobial activity. Headspace GC–MS characterization of Lactobacillus helveticus CFS identified a mixture of antimicrobial and health-promoting compounds. Minimal inhibitory concentration (MIC) values for tested Gram-positive bacteria represented 50% of that for Gram-negative bacteria, 20% and 7.35% of those for fungus and yeast respectively. Nanoparticles were prepared through chitosan-tripolyphosphate nanoparticle formation giving nanospheres from in the range from 5 to 10 nm, and narrow size distribution. CFS-loaded chitosan nanoparticles (CS-NPs) significantly enhanced the overall inhibition zone diameter, as well as, the decline in MIC values for Salmonella enterica (50%) and Penicillium chrysogenum (12.5%) was observed. Lactobacillus helveticus CFS, however, displayed lower antimicrobial activity vs. nisin and natamycin, it has both antibacterial and antifungal promising activities.
In a screening for proteolytically active lactic acid bacteria, three strains,
Lactobacillus delbrueckii
ssp.
lactis
92202,
Lactobacillus helveticus
92201, and
Lactobacillus delbrueckii
ssp.
...bulgaricus
92059, showed the highest activities following growth in milk. All three strains degraded α- and β-casein, but did not hydrolyse κ-casein. HPLC analysis of skim milk fermentation revealed increasing amounts of peptides after 5 and 10 h with
Lb. d.
ssp.
bulgaricus
92059. Hydrolysates obtained with
Lb. d.
ssp.
lactis
92202 and
Lb. d.
ssp.
bulgaricus
92059 revealed the highest angiotensin-converting enzyme-inhibitory effect. The effect was dose dependent. Almost no effect (<10%) was seen for
Lb. helveticus
92201. For
Lb. d.
ssp.
bulgaricus
92059, maximal inhibition of approx. 65% was reached after 25 h of fermentation. In an in vitro assay measuring potential immunomodulation, hydrolysates of the three strains yielded anti-inflammatory activities in the presence of TNF-α. However, the effects were more pronounced at lower hydrolysate concentrations. In the absence of TNF-α, slight pro-inflammatory effects were observed. The hydrolysate of
Lb. d.
ssp.
bulgaricus
92059, when purified by means of solid-phase extraction, exhibited pro-inflammatory activity. Sour whey containing
Lb. d.
ssp.
bulgaricus
92059 cells showed pro-inflammatory activity while cell-free sour whey was clearly anti-inflammatory. In the purified hydrolysate, 20 different α- and β-casein (CN)-derived peptides could be identified by LC-MS. Most peptides originated from the central and C-terminal regions of β-casein. Peptide length was between 9 (β-CN(f 59–67)) and 22 amino acids (β-CN(f 117–138)).
Lactobacillus helveticus is widely used in dairy fermentations and produces a range of enzymes, which upon cell lysis can be released into the cheese matrix and impact degradation of proteins, ...peptides and lipids. In our study we set out to explore the potential of Lb. helveticus DSM 20075 for increased autolytic capacity triggered by conditions such as low pH and high salt concentrations encountered in cheese environments. Lb. helveticus DSM 20075 was subjected to varied incubation temperatures (ranging from 37 to 50°C). High-temperature incubation (in the range of 45 to 50°C) allowed us to obtain a collection of six variant strains (V45-V50), which in comparison to the wild-type strain, showed higher growth rates at elevated temperatures (42°C–45°C). Moreover, variant strain V50 showed a 4-fold higher, in comparison to wild type, autolytic capacity in cheese-like conditions. Next, strain V50 was used as an adjunct in lab-scale cheese making trials to measure its impact on aroma formation during ripening. Specifically, in cheeses made with strain V50, the relative abundance of benzaldehyde increased 3-fold compared to cheeses made with the wild-type strain. Analysis of the genome sequence of strain V50 revealed multiple mutations in comparison to the wild-type strain DSM 20075 including a mutation found in a gene coding for a metal ion transporter, which can potentially be linked to intracellular accumulation of Mn2+ and benzaldehyde formation. The approach of high-temperature incubation can be applied in dairy industry for the selection of (adjunct) cultures targeted at accelerated cheese ripening and aroma formation.
•High temperature incubation of Lactobacillus helveticus DSM 20075 produces heat tolerant but salt sensitive mutants•Variant V50 showed a 4-fold higher autolytic capacity in cheese-like conditions•Variant V50 as adjunct culture in cheese trials delivered in 3-fold higher relative abundance of benzaldehyde.
Aims
In this study, we investigated the preventive properties of the supernatant of Lactobacillus helveticus NS8‐fermented milk (NS8‐FS) against UV light‐induced skin oxidative damage and ...hyperpigmentation.
Methods and Results
NS8‐FS exhibited significant radical scavenging activity in tests with ABST+ and DPPH scavenging methods, and as well strongly inhibited 3‐morpholinosydnonimine (Sin‐1)‐induced ROS generation in HaCat keratinocytes. Unexpectedly, NS8‐FS was found to inhibit melanin production in B16F10 melanoma cells and to exhibit inhibitory effects both to the enzymatic activity of tyrosinase (TYR) and the expression of proteins required for melanin synthesis. In SKH‐1 hairless mice, topical application of NS8‐FS alleviated UVB‐induced skin photodamage, including the improvement of the appearance of epidermal thickness, transepidennal water loss and lipid peroxidation levels. In the tanning guinea pig model, the whitening effect of NS8‐FS was demonstrated using Masson‐Fontana staining and TYR staining. Furthermore, NS8‐FS was shown to stimulate the nuclear translocation and activation of the Nrf2 protein, along with recovery of antioxidant enzyme activities.
Conclusion
NS8‐FS exhibits the protective capacities against UV light‐induced skin oxidative damage and hyperpigmentation.
Significance and Impact of the Study
Our findings indicate the potential of cell‐free fermented products of lactic acid bacteria in topical photoprotection.
Lactobacillus helveticus exhibits a remarkable proteolytic system. However, the etiology of these protein hydrolysis characteristics, whether caused by extracellular proteinases (EP) or cell envelope ...proteinases (CEP), has been puzzling researchers. In this study, third-generation Nanopore whole genome sequencing and proteomics analysis were used to unravel the root cause of the aforementioned confusion. The genome of L. helveticus Lh191404 was 2,117,643 bp in length, with 67 secreted proteins were found. Combined with proteomic analysis, it was found that the protein composition of extraction from CEP and EP were indeed the same substance. Bioinformatics analysis indicated that the CEP belonged to the PrtH1 Variant (PrtH1_V) genotype by phylogenetic analysis. The three-dimensional structures of various domains within the PrtH1_V-191,404 had been characterized, providing a comprehensive understanding of its structural features. Results of proteinase activity showed that the optimal reaction temperature was 40 °C, with a pH of 6.50. These findings suggested that the origin of EP in L. helveticus Lh191404 may be due to CEP being released into the substrate after detaching from the cell wall. This research is of guiding significance for further understanding the operational mechanism of the protein hydrolysis system in lactic acid bacteria.
•The protein composition from CEP and EP of L. helveticus Lh191404 were the same substance by proteomic analysis.•The origin of EP may be due to CEP being released into the substrate.•CEP belonged to the PrtH1 Variant (PrtH1_V) genotype by phylogenetic analysis.•The three-dimensional structures of various domains within the PrtH1_V-191,404 were characterized by AlphaFold 2.•The optimal proteinase hydrolysis conditions for PrtH1_V-191,404 were 40 °C and pH 6.5.
The strain Lactobacillus helveticus DPC4571 has emerged as a promising flavor adjunct culture for Cheddar cheese given that it is consistently associated with improved flavor. The availability of the ...complete genome sequence of Lb. helveticus DPC4571 has enabled the search for the presence or absence of specific genes on the genome, in particular those of technological interest. Indeed, this analysis has facilitated a greater understanding into the functioning of lactic acid bacteria as a whole. The biochemical pathways of Lb. helveticus responsible for producing flavor compounds during cheese ripening are poorly understood but now with the availability of a complete genomic sequence are ripe for exploitation. Bioinformatic analysis of the genome of Lb. helveticus DPC4571 has revealed a plethora of genes with industrial potential including those responsible for key metabolic functions that contribute to cheese flavor development such as proteolysis, lipolysis, and cell lysis. In addition, it has been demonstrated that Lb. helveticus has the potential to produce bioactive peptides such as angiotensin converting enzyme inhibitory activity in fermented dairy products, demonstrating the therapeutic value of this species. A most intriguing feature of the genome of Lb. helveticus DPC4571 is the remarkable similarity in gene content with many intestinal lactobacilli, although originating from considerably different environments. Bioinformatic analysis demonstrated that 65 to 75% of genes were conserved between the commensal and dairy lactobacilli, which allowed key niche-specific gene sets to be described. This review focuses on the isolation, characterization, and exploitation of the Lb. helveticus species with particular emphesis on taking into consideration recent genome sequence data for Lb. helveticus and other Lactobacillus species.
In this study the presence and functionality of phage defence mechanisms in Lactobacillus helveticus ATCC 10386, a strain of dairy origin which is sensitive to ΦLh56, were investigated. After ...exposure of ATCC 10386 to ΦLh56, the whole-genome sequences of ATCC 10386 and of a phage-resistant derivative (LhM3) were compared. LhM3 showed deletions in the S-layer protein and a higher expression of the genes involved in the restriction/modification (R/M) system. Genetic data were substantiated by measurements of bacteriophage adsorption rates, efficiency of plaquing, cell wall protein size and by gene expression analysis. In LhM3 two phage resistance mechanisms, the inhibition of phage adsorption and the upregulation of Type I R/M genes, take place and explain its resistance to ΦLh56. Although present in both ATCC 10386 and LhM3 genomes, the CRISPR machinery did not seem to play a role in the phage resistance of LhM3. Overall, the natural selection of phage resistant strains resulted successful in detecting variants carrying multiple phage defence mechanisms in L. helveticus. The concurrent presence of multiple phage-resistance systems should provide starter strains with increased fitness and robustness in dairy ecosystems.
•Phage resistance mechanism of a Lactobacillus helveticus strain was investigated.•Studies on a phage sensitive strain and a resistant derivative were performed.•The presence of two phage resistant mechanisms has been suggested.•CRISPR array did not seem to play a role in the phage resistance.•Importance of the natural selection of phage resistant strains.
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