Publications by authors named "Fernanda Mozzi"

37 Publications

Selenium bio-enrichment of Mediterranean fruit juices through lactic acid fermentation.

Int J Food Microbiol 2021 May 24:109248. Epub 2021 May 24.

Department of Agricultural, Food and Forest Science, Università degli Studi di Palermo, Viale delle Scienze 4, 90128 Palermo, Italy. Electronic address:

This work was carried out to elaborate selenium (Se) bio-enriched fermented Mediterranean fruit juices. To this purpose, pomegranate and table red grape juices were added with sodium selenite (NaSeO) and fermented by Levilactobacillus brevis CRL 2051 and Fructobacillus tropaeoli CRL 2034 individually or combined. To better evaluate the effect of selenite addition and starter strain inoculums on the total bacterial community of the fruit juices, fermentation trials were performed with raw and pasteurized fruit juices. No statistical significant differences were observed for total mesophilic microorganisms (TMM) and rod-shaped lactic acid bacteria (LAB) levels among raw and pasteurized juices inoculated with the starter strains, while significant differences between those juices with and without selenite were registered. LAB cocci, Pseudomonadaceae and yeasts were detected only for the raw juice preparations. The dominance of L. brevis CRL 2051 and F. tropaeoli CRL 2034 was confirmed by randomly amplified polymorphic DNA (RAPD)-PCR analysis. After fermentation, pH dropped for all inoculated trials and control raw juices. The soluble solid content (SSC) levels of the raw juices were higher than the corresponding pasteurized trials. The thermal treatment affected consistently yellowness of grape juice trials and redness of pomegranate juices. No microbial Se accumulation was registered for pomegranate juices, while F. tropaeoli CRL 2034 accumulated the highest amount of Se (65.5 μg/L) in the grape juice. For this reason, only trials carried out with raw grape juices were investigated by metagenomics analysis by Illumina MiSeq technology. Non-inoculated grape juices were massively fermented by acetic acid bacteria while Fructobacillus and Lactobacillus (previous genus name of Levilactobacillus) represented the highest operational taxonomy units (OTUs) relative abundance % of the trials inoculated with the starter strains as confirmed by this technique.
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http://dx.doi.org/10.1016/j.ijfoodmicro.2021.109248DOI Listing
May 2021

Fruits and fruit by-products as sources of bioactive compounds. Benefits and trends of lactic acid fermentation in the development of novel fruit-based functional beverages.

Food Res Int 2021 02 28;140:109854. Epub 2020 Oct 28.

Tecnológico Nacional de México/I.T.Tepic, Av. Tecnológico #2595 Fracc., Lagos del Country, 63175 Tepic Nayarit, Mexico. Electronic address:

Current awareness about the benefits of a balanced diet supports ongoing trends in humans towards a healthier diet. This review provides an overview of fruits and fruit-by products as sources of bioactive compounds and their extraction techniques, and the use of lactic acid fermentation of fruit juices to increase their functionality. Fruit matrices emerge as a technological alternative to be fermented by autochthonous or allochthonous lactic acid bacteria (LAB such as Lactiplantibacillus plantarum, Lacticaseibacillus rhamnosus, and other Lactobacillus species), and also as probiotic vehicles. During fermentation, microbial enzymes act on several fruit phytochemicals producing new derived compounds with impact on the aroma and the functionality of the fermented drinks. Moreover, fermentation significantly reduces the sugar content improving their nutritional value and extending the shelf-life of fruit-based beverages. The generation of new probiotic beverages as alternatives to consumers with intolerance to lactose or with vegan or vegetarian diets is promising for the worldwide functional food market. An updated overview on the current knowledge of the use of fruit matrices to be fermented by LAB and the interaction between strains and the fruit phytochemical compounds to generate new functional foods as well as their future perspectives in association with the application of nanotechnology techniques are presented in this review.
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http://dx.doi.org/10.1016/j.foodres.2020.109854DOI Listing
February 2021

Functional fermented cherimoya (Annona cherimola Mill.) juice using autochthonous lactic acid bacteria.

Food Res Int 2020 12 22;138(Pt A):109729. Epub 2020 Sep 22.

Centro de Referencia para Lactobacilos (CERELA-CONICET), Technological Ecophysiology Laboratory, Chacabuco 145, San Miguel de Tucumán 4000, Tucumán, Argentina; Universidad Nacional de Tucumán (UNT), Faculty of Natural Sciences, Miguel Lillo 205, San Miguel de Tucumán 4000, Tucumán, Argentina. Electronic address:

The biochemical and functional properties of fermented Annona cherimola Mill. (cherimoya) juice using five lactic acid bacteria (LAB) isolated from autochthonous fruits from Northwestern Argentina were studied in this work. Fermentation was carried out at 30 °C for 48 h followed by a 21 day-storage period at 4 °C. The assayed LAB grew well during fermentation (final count of 10 CFU/mL, ΔpH ca. 1 U) and survived after the storage period. All strains consumed fructose and glucose present in cherimoya juice as energy sources, with the consequent synthesis of lactic and/or acetic acids as final metabolic products. However, only two of the five evaluated strains were capable to produce fermented cherimoya juices with a perceptible color change. Due to lactic acid fermentation, a moderate reduction in the total phenolic content (between 13% and 43%) was observed in the majority of the samples, although no change in the antioxidant capacity was detected. The fermented cherimoya juices showed a weak antiplatelet activity when adenosine diphosphate agonist was used. The findings of this study evidenced the potential use of Annona cherimola Mill. fermented juice as a novel matrix for the formulation of stable functional beverages with appealing nutritional and functional properties.
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http://dx.doi.org/10.1016/j.foodres.2020.109729DOI Listing
December 2020

Major role of lactate dehydrogenase D-LDH1 for the synthesis of lactic acid in Fructobacillus tropaeoli CRL 2034.

Appl Microbiol Biotechnol 2020 Sep 14;104(17):7409-7426. Epub 2020 Jul 14.

Centro de Referencia para Lactobacilos (CERELA)-CONICET, Chacabuco 145, 4000, San Miguel de Tucumán, Argentina.

The enzymes D- and L-lactate dehydrogenase are involved in the reduction of pyruvate to D(+)- and L(-)-lactate, respectively. The fig-origin strain Fructobacillus tropaeoli CRL 2034 produces D- and L-lactic acids in a 9:1 ratio. In this work, two D-ldh (ldh1 and ldh2) and one L-ldh (ldh3) genes were found in the CRL 2034 genome. ldh1 and ldh2 are homologous (79% identity) and organized as contiguous operons, each gene containing 996 base pair (bp) and encoding for a 331-amino acid (aa) protein (74% identity). In contrast, ldh3 is a 927-bp gene coding for a 308-aa protein. The identity between ldh1/ldh2 and ldh3 was lower than 48%. To elucidate the role of these genes in the synthesis of lactic acid by the Fructobacillus strain, plasmid insertion mutants in each gene were generated and characterized. The growth kinetic parameters were affected only in CRL2034 ldh1::pRV300 cells, this mutant showing the lowest total lactic acid production (4.50 ± 0.15 versus 6.36 ± 0.67 g/L of wild-type strain), with a D/L ratio of 7.1:2.9. These results showed that the ldh1 gene is primarily responsible for lactic acid production by the studied strain. A comparative analysis among strains of the five Fructobacillus species revealed that the identity of D-LDH proteins was higher than 70%, while the identity of L-LDH was over 60%. Finally, phylogenetic analysis of D- and L-LDHs revealed that only D-LDH phylogeny was consistent to the phylogenetic evolution among Fructobacillus and evolutionarily related genera. Key Points •F. tropaeoli CRL 2034 harbors three ldh genes in its genome. •ldh1 and ldh2 encode D-lactate dehydrogenase; ldh3 encodes L-lactate dehydrogenase. •Gene ldh1 plays the major role in lactic acid production by strain CRL 2034. •Fructobacillus D-LDH phylogeny was consistent to phylogenetic evolution.
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http://dx.doi.org/10.1007/s00253-020-10776-9DOI Listing
September 2020

Exploring the Genome of Fructobacillus tropaeoli CRL 2034, a Fig-Origin Strain that Produces High Levels of Mannitol from Fructose.

Curr Microbiol 2020 Sep 29;77(9):2215-2225. Epub 2020 Jun 29.

Centro de Referencia Para Lactobacilos (CERELA)-CONICET, San Miguel de Tucumán, Tucumán, Argentina.

We report the draft genome sequence of Fructobacillus tropaeoli CRL 2034, a strain isolated from ripe fig in Tucumán province, Argentina. The interest in studying the genome of this fructophilic lactic acid bacterium strain was motivated by its ability to produce high levels of mannitol from fructose. This polyol has multiple industrial applications; however, it is mainly used as low calorie sugar in the food industry. The assembled genome of this strain consists of a 1.66-Mbp circular chromosome with 1465 coding sequences and a G+C content of 44.6%. The analysis of this genome supports the one step reaction of fructose reduction to mannitol by the mannitol 2-dehydrogenase enzyme, which together with a fructose permease, were identified as involved in mannitol synthesis. In addition, a phylogenetic analysis was performed including other Leuconostocaceae members to which the Fructobacillus genus belongs to; according to the 16S rRNA gene sequences, the strain CRL 2034 was located in the Fructobacillus clade. The present genome sequence could be useful to further elucidate regulatory processes of mannitol and other bioactive metabolites and to highlight the biotechnological potential of this fruit-origin Fructobacillus strain.
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http://dx.doi.org/10.1007/s00284-020-02102-3DOI Listing
September 2020

Biotransformation of Selenium by Lactic Acid Bacteria: Formation of Seleno-Nanoparticles and Seleno-Amino Acids.

Front Bioeng Biotechnol 2020 12;8:506. Epub 2020 Jun 12.

Centro de Referencia para Lactobacilos (CERELA-CONICET), Tucumán, Argentina.

Selenium (Se) is an essential micronutrient for the majority of living organisms, and it has been identified as selenocysteine in the active site of several selenoproteins such as glutathione peroxidase, thioredoxin reductase, and deiodinases. Se deficiency in humans is associated with viral infections, thyroid dysfunction, different types of cancer, and aging. In several European countries as well as in Argentina, Se intake is below the recommended dietary Intake (RDI). Some lactic acid bacteria (LAB) can accumulate and bio-transform selenite (toxic) into Se-nanoparticles (SeNPs) and Se-amino acids (non-toxic). The microbial growth, Se metabolite distribution, and the glutathione reductase (involved in selenite reduction) activity of Se-enriched LAB were studied in this work. The ninety-six assayed strains, belonging to the genera , and could grow in the presence of 5 ppm sodium selenite. From the total, eight strains could remove more than 80% of the added Se from the culture medium. These bacteria accumulated intracellularly between 1.2 and 2.5 ppm of the added Se, from which CRL 2034 contained the highest intracellular amount. These strains produced only the seleno-amino acid SeCys as observed by LC-ICP-MS and confirmed by LC-ESI-MS/MS. The intracellular SeCys concentrations were between 0.015 and 0.880 ppm; CRL 2051 (0.873 ppm), CRL 2030 (0.867 ppm), and CRL 2034 (0.625 ppm) were the strains that showed the highest concentrations. Glutathione reductase activity values were higher when the strains were grown in the presence of Se except for the CRL 2034 strain, which showed an opposite behavior. The cellular morphology of the strains was not affected by the presence of Se in the culture medium; interestingly, all the strains were able to form spherical SeNPs as determined by transmission electron microscopy (TEM). Only two strains produced the volatile Se compounds dimethyl-diselenide identified by GC-MS. Our results show that CRL 2051, CRL 2030, and CRL 2034 could be used for the development of nutraceuticals or as starter cultures for the bio-enrichment of fermented fruit beverages with SeCys and SeNPs.
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http://dx.doi.org/10.3389/fbioe.2020.00506DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7303280PMC
June 2020

Survival of selenium-enriched lactic acid bacteria in a fermented drink under storage and simulated gastro-intestinal digestion.

Food Res Int 2019 09 24;123:115-124. Epub 2019 Apr 24.

Centro de Referencia para Lactobacilos (CERELA)-CONICET, Chacabuco 145, San Miguel de Tucumán 4000, Argentina. Electronic address:

Selenium (Se), which is present as SeCys in seleno-proteins, is involved in cancer prevention, thyroid functioning, and pathogen inhibition. Se is incorporated in the diet through Se-containing foods. Some lactic acid bacteria (LAB) can biotransform selenite (toxic) into Se-nanoparticles (SeNPs) and Se-amino acids. To exert their beneficial properties in the host, bacteria should survive the harsh conditions of the gastrointestinal tract and during food storage. We evaluated whether selenization of LAB influenced bacterial growth and survival during gastrointestinal digestion and after storage when present in a fermented fruit juice-milk (FJM) beverage. Lactobacillus brevis CRL 2051 and Fructobacillus tropaeoli CRL 2034 were grown in MRS with and without selenite, and used to inoculate the FJM matrix. Selenization had no effect on LAB growth (9.54-9.9 log CFU/mL) in the FJM drink. The presence of SeNPs was confirmed for both selenized strains in the FJM beverage; however, the highest Se concentration (100 μg/L) was detected for the fermented beverage with selenized L. brevis. Under storage conditions 1.1 log CFU/ml decrease in cell count of selenized cells of L. brevis was observed, while no effect on cell viability was detected for non-selenized L. brevis or both selenized and control cells of F. tropaeoli. Resistance of L. brevis during digestion of the fermented FJM beverage was not affected by selenization. Contrarily, an increase (1 log CFU/mL) in the resistance of F. tropaeoli was observed when cells were selenized. After digestion, Se was detected in the soluble fraction of the beverage fermented by both strains, being higher for L. brevis (23.6 μg/L). Although selenization did not exert a drastic effect on strains´ survival during storage and digestion, microbial selenization previous to food fermentation could be an interesting tool for Se enrichment avoiding thus the addition of toxic Se salts.
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http://dx.doi.org/10.1016/j.foodres.2019.04.057DOI Listing
September 2019

Diversity and Functional Properties of Lactic Acid Bacteria Isolated From Wild Fruits and Flowers Present in Northern Argentina.

Front Microbiol 2019 21;10:1091. Epub 2019 May 21.

Technology and Development Laboratory, Centro de Referencia para Lactobacilos (CERELA)-CONICET, San Miguel de Tucumán, Tucumán, Argentina.

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.
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http://dx.doi.org/10.3389/fmicb.2019.01091DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6536596PMC
May 2019

Silac-based quantitative proteomic analysis of Lactobacillus reuteri CRL 1101 response to the presence of selenite and selenium nanoparticles.

J Proteomics 2019 03 27;195:53-65. Epub 2018 Dec 27.

Department of Analytical Chemistry, Faculty of Chemistry, Complutense University of Madrid, Av. Complutense s/n, 28040 Madrid, Spain. Electronic address:

Stable isotope labeling in cell culture (SILAC) was applied for the first time on a lactic acid bacterium strain (L. reuteri CRL1101) for analyzing differential protein expression associated to selenite(NaSeO) and selenium nanoparticles (SeNPs) exposure. 57 and 47 proteins were found de-regulated by >1,5 fold in presence of selenite and SeNPs, respectively. Only 16 out of 104 proteins differentially expressed were commonly altered by selenite and SeNPs. The use of a clustered heat map allows us to visualize relations between the de-regulated proteins and exposure conditions. We identified a number of proteins involved in diverse functions and biological processes such as metabolism of carbohydrates, selenium and lipids; folding, sorting and degradation; environmental information and processing. In presence of both, selenite and SeNPs, proteins related to selenium metabolism such as cystathione beta-lyase and oxidoreductases (thioredoxine reductase and NAD/FAD oxidoreductase) were over expressed. Interestingly, the over expression of thioredoxin reductase could protect the host from oxidizing compounds. An over expression of phage proteins and chaperones with selenite was observed; this result and the fact that a lower cell count was detected when selenite was added could indicate that this latter Se species has a more deleterious effect than the nanoparticles.
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http://dx.doi.org/10.1016/j.jprot.2018.12.025DOI Listing
March 2019

Determination of size and mass-and number-based concentration of biogenic SeNPs synthesized by lactic acid bacteria by using a multimethod approach.

Anal Chim Acta 2017 Nov 28;992:34-41. Epub 2017 Sep 28.

Departamento de Química Analítica, Facultad de Ciencias Químicas, Universidad Complutense de Madrid, Ciudad Universitaria, 28040 Madrid, Spain. Electronic address:

Selenium nanoparticles (SeNPs) were synthesized by a green technology using lactic acid bacteria (LAB, Lactobacillus acidophilus, L. delbrueckii subsp. bulgaricus and L. reuteri). The exposure of aqueous sodium selenite to LAB led to the synthesis of SeNPs. Characterization of SeNPs by transmission electron microscopy with energy dispersive X-ray spectrum (EDXS) analysis revealed the presence of stable, predominantly monodispersed and spherical SeNPs of an average size of 146 ± 71 nm. Additionally, SeNPs hydrodynamic size was determined by dispersive light scattering (DLS) and nanoparticle tracking analysis (NTA). For this purpose, a methodology based on the use of surfactants in basic medium was developed for isolating SeNPs from the bacterial pellet. The hydrodynamic size values provided by DLS and NTA were 258 ± 4 and 187 ± 56 nm, respectively. NTA measurements of number-based concentration reported values of (4.67±0.30)x10 SeNPs mL with a relative standard deviation lower than 5% (n = 3). The quantitative results obtained by NTA were supported by theoretical calculations. Asymmetrical flow field flow fractionation (AF) on line coupled to the inductively couple plasma mass spectrometry (ICP-MS) and off-line coupled to DLS was further employed to characterize biogenic SeNPs. The distribution of the particle size for the Se-containing peak provide an average size of (247 ± 14) nm. The data obtained by independent techniques were in good agreement and the developed methodology could be implemented for characterizing NPs in complex matrices such as biogenic nanoparticles embedded inside microbial material.
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http://dx.doi.org/10.1016/j.aca.2017.09.033DOI Listing
November 2017

YebC, a putative transcriptional factor involved in the regulation of the proteolytic system of Lactobacillus.

Sci Rep 2017 08 17;7(1):8579. Epub 2017 Aug 17.

Centro de Referencia para Lactobacilos (CERELA-CONICET), Chacabuco 145, 4000, San Miguel de Tucumán, Argentina.

The proteolytic system of Lactobacillus plays an essential role in bacterial growth, contributes to the flavor development of fermented products, and can release bioactive health-beneficial peptides during milk fermentation. In this work, a genomic analysis of all genes involved in the proteolytic system of L. delbrueckii subsp. lactis CRL 581 was performed. Genes encoding the cell envelope-associated proteinase, two peptide transport systems, and sixteen peptidases were identified. The influence of the peptide supply on the transcription of 23 genes involved in the proteolytic system of L. delbrueckii subsp. lactis was examined after cell growth in a chemically defined medium (CDM) and CDM supplemented with Casitone. prtL, oppA , optS, optA genes as well as oppDFBC and optBCDF operons were the most highly expressed genes in CDM; their expression being repressed 6- to 115-fold by the addition of peptides. The transcriptional analysis was confirmed by proteomics; the up-regulation of the PrtL, PepG, OppD and OptF proteins in the absence of peptides was observed while the DNA-binding protein YebC was up-regulated by peptides. Binding of YebC to the promoter region of prtL, oppA , and optS, demonstrated by electrophoretic mobility shift assays, showed that YebC acts as a transcriptional repressor of key proteolytic genes.
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http://dx.doi.org/10.1038/s41598-017-09124-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5561223PMC
August 2017

Enhanced mannitol biosynthesis by the fruit origin strain Fructobacillus tropaeoli CRL 2034.

Appl Microbiol Biotechnol 2017 Aug 3;101(15):6165-6177. Epub 2017 Jul 3.

Centro de Referencia para Lactobacilos (CERELA-CONICET), Chacabuco 145, T4000, Tucumán, Argentina.

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 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.
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http://dx.doi.org/10.1007/s00253-017-8395-1DOI Listing
August 2017

Global Analysis of Mannitol 2-Dehydrogenase in Lactobacillus reuteri CRL 1101 during Mannitol Production through Enzymatic, Genetic and Proteomic Approaches.

PLoS One 2017 6;12(1):e0169441. Epub 2017 Jan 6.

Centro de Referencia para Lactobacilos (CERELA)-CONICET, Chacabuco 145, San Miguel de Tucumán, Tucumán, Argentina.

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.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0169441PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5218481PMC
August 2017

Lactic Acid Bacteria as Cell Factories for the Generation of Bioactive Peptides.

Protein Pept Lett 2017 ;24(2):146-155

Laboratory of Technology, CERELA-CONICET, Chacabuco 145, 4000, San Miguel de Tucuman, Tucuman, Argentina.

There is a growing interest in the incorporation of functional foods in the daily diet to achieve health promotion and disease risk reduction. Numerous studies have focused on the production of biologically active peptides as nutraceuticals and functional food ingredients due to their health benefits. These short peptides, displaying antihypertensive, antioxidant, mineral binding, immunomodulatory and antimicrobial activities are hidden in a latent state within the primary sequences of food proteins requiring enzymatic proteolysis for their release. While microbial fermentation is one of the major and economically most convenient processes used to generate bioactive peptides, lactic acid bacteria (LAB) are widely used as starter cultures for the production of diverse fermented foods. This article reviews the current knowledge on LAB as cell factories for the production of bioactive peptides from a variety of food protein sources. These microorganisms depend on a complex proteolytic system to ensure successful fermentation processes. In the dairy industry, LAB containing cell envelope-associated proteinases (CEPs) are employed as biocatalysts for the first step of casein breakdown releasing bioactive peptides during milk fermentation. A better understanding of the functionality and regulation of the proteolytic system of LAB opens up future opportunities for the production of novel food-derived compounds with potential health-promoting properties.
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http://dx.doi.org/10.2174/0929866524666161123111333DOI Listing
March 2017

S-layer production by Lactobacillus acidophilus IBB 801 under environmental stress conditions.

Appl Microbiol Biotechnol 2016 May 24;100(10):4573-83. Epub 2016 Feb 24.

Institute of Biology Bucharest of the Romanian Academy, Splaiul Independentei No. 296, 060031, Bucharest, Romania.

The ability of microorganisms to synthesize S-layer, the outermost structure of the microbial cell envelope composed of non-covalently bound proteins, has been ascribed to help microorganisms to exert their probiotic properties in the host. In this work, formation of S-layer by the potentially probiotic strain Lactobacillus acidophilus IBB 801 under different stress culture conditions (high incubation temperatures, presence of bile salts or NaCl, and acidic pH) was assayed. A marked S-layer synthesis by L. acidophilus IBB 801 was detected when the strain was grown at 42 °C and in the presence of 0.05 % bile salts or 2.0 % NaCl. The presence of S-layer proteins was further confirmed by transmission electron microscopy and protein identification by MS/MS. The differential expression of the proteome of this strain at 42 °C, when a marked formation of S-layer was detected, revealed the overexpression of six proteins mainly related to general stress and protein biosynthesis and translation, while four proteins detected in lower amounts were involved in DNA repair and energy metabolism. As L. acidophilus IBB 801 produces both a bacteriocin and S-layer proteins, the strain could be of interest to be used in the formulation of functional food products with specific properties.
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http://dx.doi.org/10.1007/s00253-016-7355-5DOI Listing
May 2016

Biopolymers from lactic acid bacteria. Novel applications in foods and beverages.

Front Microbiol 2015 11;6:834. Epub 2015 Sep 11.

Technology Department, Centro de Referencia para Lactobacilos - Consejo Nacional de Investigaciones Científicas y Técnicas, San Miguel de Tucumán Argentina.

Lactic acid bacteria (LAB) are microorganisms widely used in the fermented food industry worldwide. Certain LAB are able to produce exopolysaccharides (EPS) either attached to the cell wall (capsular EPS) or released to the extracellular environment (EPS). According to their composition, LAB may synthesize heteropolysaccharides or homopolysaccharides. A wide diversity of EPS are produced by LAB concerning their monomer composition, molecular mass, and structure. Although EPS-producing LAB strains have been traditionally applied in the manufacture of dairy products such as fermented milks and yogurts, their use in the elaboration of low-fat cheeses, diverse type of sourdough breads, and certain beverages are some of the novel applications of these polymers. This work aims to collect the most relevant issues of the former reviews concerning the monomer composition, structure, and yields and biosynthetic enzymes of EPS from LAB; to describe the recently characterized EPS and to present the application of both EPS-producing strains and their polymers in the fermented (specifically beverages and cereal-based) food industry.
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http://dx.doi.org/10.3389/fmicb.2015.00834DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4566036PMC
October 2015

Whey-derived valuable products obtained by microbial fermentation.

Appl Microbiol Biotechnol 2015 Aug 28;99(15):6183-96. Epub 2015 Jun 28.

Centro de Referencia para Lactobacilos CERELA-CONICET, Chacabuco 145, San Miguel de Tucumán, 4000, Argentina.

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.
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http://dx.doi.org/10.1007/s00253-015-6766-zDOI Listing
August 2015

Efficient mannitol production by wild-type Lactobacillus reuteri CRL 1101 is attained at constant pH using a simplified culture medium.

Appl Microbiol Biotechnol 2015 Oct 18;99(20):8717-29. Epub 2015 Jun 18.

Departamento de Tecnología y Desarrollo, Centro de Referencia para Lactobacilos (CERELA)-CONICET, Chacabuco 145, 4000, San Miguel de Tucumán, Argentina.

Mannitol is a natural polyol with multiple industrial applications. In this work, mannitol production by Lactobacillus reuteri CRL 1101 was studied at free- and controlled-pH (6.0-4.8) fermentations using a simplified culture medium containing yeast and beef extracts and sugarcane molasses. The activity of mannitol 2-dehydrogenase (MDH), the enzyme responsible for mannitol synthesis, was determined. The effect of the initial biomass concentration was further studied. Mannitol production (41.5 ± 1.1 g/l), volumetric productivity (Q Mtl 1.73 ± 0.05 g/l h), and yield (Y Mtl 105 ± 11 %) were maximum at pH 5.0 after 24 h while the highest MDH activity (1.66 ± 0.09 U/mg protein) was obtained at pH 6.0. No correlation between mannitol production and MDH activity was observed when varying the culture pH. The increase (up to 2000-fold) in the initial biomass concentration did not improve mannitol formation after 24 h although a 2-fold higher amount was produced at 8 h using 1 or 2 g cell dry weight/l comparing to the control (0.001 g cell dry weight/l). Finally, mannitol isolation under optimum fermentation conditions was achieved. The mannitol production obtained in this study is the highest reported so far by a wild-type L. reuteri strain and, more interestingly, using a simplified culture medium.
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http://dx.doi.org/10.1007/s00253-015-6730-yDOI Listing
October 2015

Draft Genome Sequence of the Mannitol-Producing Strain Lactobacillus mucosae CRL573.

Genome Announc 2014 Dec 11;2(6). Epub 2014 Dec 11.

Centro de Referencia para Lactobacilos (CERELA) - CONICET, San Miguel de Tucumán, Argentina

Lactobacillus mucosae CRL573, isolated from child fecal samples, efficiently converts fructose and/or sucrose into the low-calorie sugar mannitol when cultured in modified MRS medium at pH 5.0. Also, the strain is capable of producing bacteriocin. The draft genome sequence of this strain with potential industrial applications is presented here.
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http://dx.doi.org/10.1128/genomeA.01292-14DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4263840PMC
December 2014

Lactobacillus delbrueckii subsp. bulgaricus CRL 454 cleaves allergenic peptides of β-lactoglobulin.

Food Chem 2015 Mar 27;170:407-14. Epub 2014 Aug 27.

Centro de Referencia para Lactobacilos (CERELA)-CONICET, Chacabuco 145, 4000 San Miguel de Tucumán, Argentina; Cátedra de Microbiología Superior, Facultad de Bioquímica, Química y Farmacia, Universidad Nacional de Tucumán, Tucumán, Argentina. Electronic address:

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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http://dx.doi.org/10.1016/j.foodchem.2014.08.086DOI Listing
March 2015

Diversity in proteinase specificity of thermophilic lactobacilli as revealed by hydrolysis of dairy and vegetable proteins.

Appl Microbiol Biotechnol 2013 Sep 9;97(17):7831-44. Epub 2013 Jul 9.

Centro de Referencia para Lactobacilos (CERELA)-CONICET, Chacabuco 145, 4000, San Miguel de Tucumán, Argentina.

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 αs-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.
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http://dx.doi.org/10.1007/s00253-013-5037-0DOI Listing
September 2013

Biotechnological and in situ food production of polyols by lactic acid bacteria.

Appl Microbiol Biotechnol 2013 Jun 19;97(11):4713-26. Epub 2013 Apr 19.

Centro de Referencia para Lactobacilos (CERELA)-CONICET, Chacabuco 145, San Miguel de Tucumán 4000, Argentina.

Polyols such as mannitol, erythritol, sorbitol, and xylitol are naturally found in fruits and vegetables and are produced by certain bacteria, fungi, yeasts, and algae. These sugar alcohols are widely used in food and pharmaceutical industries and in medicine because of their interesting physicochemical properties. In the food industry, polyols are employed as natural sweeteners applicable in light and diabetic food products. In the last decade, biotechnological production of polyols by lactic acid bacteria (LAB) has been investigated as an alternative to their current industrial production. While heterofermentative LAB may naturally produce mannitol and erythritol under certain culture conditions, sorbitol and xylitol have been only synthesized through metabolic engineering processes. This review deals with the spontaneous formation of mannitol and erythritol in fermented foods and their biotechnological production by heterofermentative LAB and briefly presented the metabolic engineering processes applied for polyol formation.
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http://dx.doi.org/10.1007/s00253-013-4884-zDOI Listing
June 2013

A putative transport protein is involved in citrulline excretion and re-uptake during arginine deiminase pathway activity by Lactobacillus sakei.

Res Microbiol 2013 Apr 23;164(3):216-25. Epub 2012 Nov 23.

Research Group of Industrial Microbiology and Food Biotechnology (IMDO), Faculty of Sciences and Bioengineering Sciences, Vrije Universiteit Brussel, Pleinlaan 2, B-1050 Brussels, Belgium.

Arginine conversion through the arginine deiminase (ADI) pathway is a common metabolic trait of Lactobacillus sakei which is ascribed to an arc operon and which inquisitively involves citrulline excretion and re-uptake. The aim of this study was to verify whether a putative transport protein (encoded by the PTP gene) plays a role in citrulline-into-ornithine conversion by L. sakei strains. This was achieved through a combination of fermentation experiments, gene expression analysis via quantitative real-time reverse transcription PCR (RT-qPCR) and construction of a PTP knock-out mutant. Expression of the PTP gene was modulated by environmental pH and was highest in the end-exponential or mid-exponential growth phase for L. sakei strains CTC 494 and 23K, respectively. In contrast to known genes of the arc operon, the PTP gene showed low expression at pH 7.0, in agreement with the finding that citrulline-into-ornithine conversion is inhibited at this pH. The presence of additional energy sources also influenced ADI pathway activity, in particular by decreasing citrulline-into-ornithine conversion. Further insight into the functionality of the PTP gene was obtained with a knock-out mutant of L. sakei CTC 494 impaired in the PTP gene, which displayed inhibition in its ability to convert extracellular citrulline into ornithine. In conclusion, results indicated that the PTP gene may putatively encode a citrulline/ornithine antiporter.
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http://dx.doi.org/10.1016/j.resmic.2012.11.004DOI Listing
April 2013

Diversity in growth and protein degradation by dairy relevant lactic acid bacteria species in reconstituted whey.

J Dairy Res 2012 May;79(2):201-8

Centro de Referencia para Lactobacilos (CERELA)-CONICET, Chacabuco 145, 4000 San Miguel de Tucumán, Argentina.

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.
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http://dx.doi.org/10.1017/S0022029912000040DOI Listing
May 2012

Lactobacillus reuteri CRL 1101 highly produces mannitol from sugarcane molasses as carbon source.

Appl Microbiol Biotechnol 2012 Aug 22;95(4):991-9. Epub 2012 Feb 22.

Centro de Referencia para Lactobacilos (CERELA)-CONICET, Chacabuco 145, San Miguel de Tucumán, Argentina.

Mannitol is a natural polyol extensively used in the food industry as low-calorie sugar being applicable for diabetic food products. We aimed to evaluate mannitol production by Lactobacillus reuteri CRL 1101 using sugarcane molasses as low-cost energy source. Mannitol formation was studied in free-pH batch cultures using 3-10% (w/v) molasses concentrations at 37 °C and 30 °C under static and agitated conditions during 48 h. L. reuteri CRL 1101 grew well in all assayed media and heterofermentatively converted glucose into lactic and acetic acids and ethanol. Fructose was used as an alternative electron acceptor and reduced it to mannitol in all media assayed. Maximum mannitol concentrations of 177.7 ± 26.6 and 184.5 ± 22.5 mM were found using 7.5% and 10% molasses, respectively, at 37 °C after 24-h incubation. Increasing the molasses concentration from 7.5% up to 10% (w/v) and the fermentation period up to 48 h did not significantly improve mannitol production. In agitated cultures, high mannitol values (144.8 ± 39.7 mM) were attained at 8 h of fermentation as compared to static ones (5.6 ± 2.9 mM), the highest mannitol concentration value (211.3 ± 15.5 mM) being found after 24 h. Mannitol 2-dehydrogenase (MDH) activity was measured during growth in all fermentations assayed; the highest MDH values were obtained during the log growth phase, and no correlation between MDH activities and mannitol production was observed in the fermentations performed. L. reuteri CRL 1101 successfully produced mannitol from sugarcane molasses being a promising candidate for microbial mannitol synthesis using low-cost substrate.
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http://dx.doi.org/10.1007/s00253-012-3945-zDOI Listing
August 2012

Draft genome sequence of Enterococcus mundtii CRL1656.

J Bacteriol 2012 Jan;194(2):550

Instituto de Biología Molecular y Celular de Rosario and Departamento de Microbiología, Universidad Nacional de Rosario, Suipacha, Rosario, Argentina.

We report the draft genome sequence of Enterococcus mundtii CRL1656, which was isolated from the stripping milk of a clinically healthy adult Holstein dairy cow from a dairy farm of the northwestern region of Tucumán (Argentina). The 3.10-Mb genome sequence consists of 450 large contigs and contains 2,741 predicted protein-coding genes.
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http://dx.doi.org/10.1128/JB.06415-11DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3256672PMC
January 2012

Genome sequence of the bacteriocin-producing Lactobacillus curvatus strain CRL705.

J Bacteriol 2012 Jan;194(2):538-9

Centro de Referencia para Lactobacilos, San Miguel de Tucumán, Argentina.

Lactobacillus curvatus is one of the most prevalent lactic acid bacteria found in fermented meat products. Here, we present the draft genome sequence of Lactobacillus curvatus CRL705, a bacteriocin producer strain isolated from an Argentinean artisanal fermented sausage, which consists of 1,833,251 bp (GC content, 41.9%) and two circular plasmids of 12,342 bp (pRC12; GC, 43.9%) and 18,664 bp (pRC18; GC, 34.4%).
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http://dx.doi.org/10.1128/JB.06416-11DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3256670PMC
January 2012

Mannitol production by heterofermentative Lactobacillus reuteri CRL 1101 and Lactobacillus fermentum CRL 573 in free and controlled pH batch fermentations.

Appl Microbiol Biotechnol 2012 Mar 13;93(6):2519-27. Epub 2011 Oct 13.

Centro de Referencia para Lactobacilos (CERELA)-CONICET, Chacabuco 145, San Miguel de Tucumán, Argentina.

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.
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http://dx.doi.org/10.1007/s00253-011-3617-4DOI Listing
March 2012

Proteolytic action of Lactobacillus delbrueckii subsp. bulgaricus CRL 656 reduces antigenic response to bovine β-lactoglobulin.

Food Chem 2011 Jul 9;127(2):487-92. Epub 2011 Jan 9.

Centro de Referencia para Lactobacilos (CERELA)-CONICET, Chacabuco 145, 4000 San Miguel de Tucumán, Argentina.

The whey protein β-lactoglobulin (BLG) is highly allergenic. Lactic acid bacteria can degrade milk proteins. The capacity of Lactobacillus delbrueckii subsp. bulgaricus CRL 656 to hydrolyse the major BLG epitopes (V41-K60; Y102-R124; L149-I162) and decrease their recognition by IgE of allergic patients was evaluated. The intensity of BLG degradation was analysed by Tricine SDS-PAGE and RP-HPLC. Peptides released were identified by LC-MS/MS and the hydrolysates were tested for their capacity to inhibit IgE binding by ELISA test. L. delbrueckii subsp. bulgaricus CRL 656 degraded BLG (35%, 8h). The sequence analysis of the released peptides indicated that this strain degraded three main BLG epitopes. BLG-positive sera (3-5year old children) were used for testing IgE binding inhibition of BLG hydrolysates from the Lactobacillus strain. The hydrolysates were less immuno-reactive (32%) than the heated BLG. L. delbrueckii subsp. bulgaricus CRL 656 could be used for developing hypoallergenic dairy products.
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http://dx.doi.org/10.1016/j.foodchem.2011.01.029DOI Listing
July 2011

Functional fermented whey-based beverage using lactic acid bacteria.

Int J Food Microbiol 2010 Jun 18;141(1-2):73-81. Epub 2010 Apr 18.

Centro de Referencia para Lactobacilos (CERELA)-CONICET, Chacabuco 145, 4000 San Miguel de Tucumán, Argentina.

Whey protein concentrate (WPC) is employed as functional food ingredient because of its nutritional value and emulsifying properties. However, the major whey protein beta-lactoglobulin (BLG) is the main cause of milk allergy. The aim of this study was to formulate a fermented whey beverage using selected lactic acid bacteria and WPC35 (WPC containing 35% of proteins) to obtain a fermented product with low lactose and BLG contents and high essential amino acid concentration. Cell viability, lactose consumption, lactic acid production, proteolytic activity, amino acid release and BLG degradation by the selected strains Lactobacillus acidophilus CRL 636, Lactobacillus delbrueckii subsp. bulgaricus CRL 656 and Streptococcus thermophilus CRL 804, as single or mixed (SLaB) cultures were evaluated in WPC35 (10%, w/v) incubated at 37 degrees C for 24h. Then, the fermented WPC35 was mixed with peach juice and calcium lactate (2%, w/v) and stored at 10 degrees C for 28 days. During fermentation, single cultures grew 1.7-3.1 log CFU/ml and produced 25.1-95.0 mmol/l of lactic acid as consequence of lactose consumption (14.0-41.8 mmol/l) after 12h fermentation. L. delbrueckii subsp. bulgaricus CRL 656 was the most proteolytic strain (626 microg/ml Leu) and released the branched-chain essential amino acids Leu (16 microg/ml), Ile (27 microg/ml) and Val (43 microg/ml). All strains were able to degrade BLG in a range of 41-85% after 12h incubation. The starter culture SLaB grew 3.0 log CFU/ml, showed marked pH reduction, produced 122.0 mmol/l of lactic acid, displayed high proteolytic activity (484 microg/ml Leu) releasing Leu (13 microg/ml), Ile (18 microg/ml) and Val (35 microg/ml), and hydrolyzed 92% of BLG. The addition of calcium lactate to WPC35 maintained the drink pH stable during shelf life; no contamination was detected during this period. After 28 days, a decrease in cell viability of all strains was observed being more pronounced for L. delbrueckii subsp. bulgaricus CRL 656 and L. acidophilus CRL 636 (2.3 and 1.9 log CFU/ml, respectively). The results showed that WPC fermentation by rationally selected lactic acid bacteria might be used for developing functional beverages with improved characteristics such as reduced BLG content and increased branched-chain essential amino acids.
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http://dx.doi.org/10.1016/j.ijfoodmicro.2010.04.011DOI Listing
June 2010
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