Exopolysaccharide-producing Lacticaseibacillus paracasei strains isolated from kefir as starter for functional dairy products

Exopolysaccharides (EPS) produced by lactic acid bacteria are molecules of great interest for the dairy food industry. Lacticaseibacillus paracasei CIDCA 8339, CIDCA 83123, and CIDCA 83124 are potentially probiotic strains isolated from kefir grains whose EPS-production on MRS broth is dependent on incubation temperature. The aim of the present work is to evaluate the effect of fermentation temperature on the characteristics of EPS produced in milk by L. paracasei strains and the consequent impact on the rheological properties of the fermented products. Additionally, the protective effect of these EPS against Salmonella infection was evaluated in vitro. Acid gels with each strain were obtained by milk fermentation at 20°C, 30°C, and 37°C evidencing for all the strains a reduction in growth and acidification rate at lower temperature. Lacticaseibacillus paracasei CIDCA 83123 showed low fermentation rate at all temperatures requiring between 3 and 8 days to obtain acids gels, whereas CIDCA 8339 and 83124 needed between 24 and 48 h even when the temperature was 20°C. Fermentation temperature led to changes in crude EPS characteristics of the three strains, observing an increase in the relative amount of the high molecular weight fraction when the fermentation temperature diminished. Additionally, EPS83124 and EPS83123 presented modifications in monosaccharide composition, with a reduction of rhamnose and an increase of amino-sugars as temperature rise. These changes in the structure of EPS83124 resulted in an increase of the apparent viscosity of milks fermented at 20°C (223 mPa.s) and 30°C (217 mPa.s) with respect to acid gels obtained at 37°C (167 mPa.s). In order to deepen the knowledge on EPS characteristics, monosaccharide composition of low and high molecular weight EPS fractions were evaluated. Finally, it was evidenced that the preincubation of intestinal epithelial cells Caco-2/TC-7 with EPS8339 and EPS83124 partially inhibit the association and invasion of Salmonella. In light of these results, it can be concluded that the selection of the EPS-producing strain along with the appropriate fermentation conditions could be an interesting strategy to improve the technological properties of these L. paracasei fermented milks with potential protective effects against intestinal pathogens.The present work was supported by CONICET (PIP 2786), Universidad Nacional de La Plata (UNLP 18/X813), ANPCyT (PICT 2020-03973 and PICT 2020-3239), and the Basque Government (IT1662-22 and PIBA 2020_1_0032)

Producción de tensioactivos con actividad antimicrobiana mediante el empleo de tecnologías amigables con el medio ambiente

Papaína es una endopeptidasa extraída del látex de frutos de Carica papaya ampliamente empleada en las industrias alimentaria y farmacéutica. Es muy utilizada como biocatalizador en la síntesis de péptidos y otros derivados. Los surfactantes derivados de arginina son una familia de tensioactivos catiónicos con propiedades antimicrobianas interesantes como preservativos para formulaciones farmacéuticas y alimentarias.El objetivo del presente trabajo fue la obtención, purificación y determinación del poder antimicrobiano de un compuesto derivado de arginina con potencial actividad tensioactiva, sintetizado mediante el empleo de papaína como biocatalizador.La síntesis se llevó a cabo por condensación del éster etílico de N?-benzoil-arginina con dodecilamina como nucleófilo, utilizando papaína adsorbida sobre poliamida como biocatalizador y acetonitrilo con 0.25% (v/v) de agua como medio de reacción. El producto (Bz-Arg-NHC12) fue purificado mediante cromatografía de intercambio catiónico utilizando como eluyente una mezcla de buffer bórico-borato 0.01 M (pH 8.5)/etanol (1:1) y un gradiente en NaCl. Se ensayó la actividad antimicrobiana mediante el método de microdilución frente a Salmonella enterica, Serratia marcescens, Shigella flexneri, Escherichia coli, Pseudomonas aeruginosa, Bacillus cereus, Bacillus subtilis, Staphylococcus aureus, Enterococcus faecalis, Enterococcus faecium, Kocuria rhizophila, Listeria innocua, Listeria monocytogenes, Pichia anomala, Saccharomyces cereviciae, Kluyveromyces marxianus y Rhodothorula sp., calculándose los porcentajes de inhibición de crecimiento y las concentraciones inhibitoria mínima (CIM) y bactericida mínima (CBM).La síntesis de Bz-Arg-NHC12 catalizada por papaína adsorbida en poliamida tuvo una conversión del 86.2%. Se determinó la actividad antimicrobiana de una solución acuosa de Bz-Arg-NHC12 (60.0 ?g.ml-1) frente a distintas especies de bacterias y levaduras. Los microorganismos Gram positivos resultaron en general inhibidos en más de un 90%, en tanto las bacterias Gram negativas y levaduras ensayadas fueron resistentes.El compuesto puro presentó una CIM de 15.6 ?g.ml-1 para B. cereus, 62.5 ?g.ml-1 para St. aureus y 250.0 ?g.ml-1 para E. coli, en tanto que no afectó el crecimiento de Ps. aeruginosa. Las CBM fueron 250.0 ?g.ml-1, 62.5 ?g.ml-1 y 250.0 ?g.ml-1 para B. cereus, St. aureus y E. coli respectivamente

Síntesis biocatalítica, estudios de toxicidad y caracterización biológica de surfactantes derivados de arginina para su empleo como aditivos en formulaciones farmacéuticas

Tanto la búsqueda como el desarrollo de compuestos bioactivos deben cumplir con requisitos de inocuidad, multifuncionalidad, eficacia, competitividad y bajo impacto ambiental. Una de las estrategias para conseguir tensioactivos ecológicamente aceptables es la preparación de moléculas cuya estructura mimetice la de compuestos naturales, como los lipoaminoácidos. Los surfactantes derivados de arginina son tensioactivos catiónicos con múltiples aplicaciones en la industria y amplia actividad biológica, cuya síntesis puede llevarse a cabo mediante catálisis enzimática. En este sentido, papaína (E.C. 3.4.22.2), una endopeptidasa cisteínica vegetal obtenida del látex de Carica papaya, es una de las enzimas más empleadas en procesos biocatalíticos, entre ellos la síntesis de derivados peptídicos, debido a su robustez y amplia especificidad.Fil: Fait, María Elisa. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Departamento de Ciencias Biológicas. Laboratorio de Investigación de Proteínas Vegetales; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Costa, H.. Universidade Federal do Ceará; BrasilFil: do Santos Queiroz, A.. Universidade Federal do Ceará; BrasilFil: Lorenzo, J.. Universitat Autònoma de Barcelona; EspañaFil: Garrote, Graciela Liliana. Provincia de Buenos Aires. Gobernación. Comisión de Investigaciones Científicas. Centro de Investigación y Desarrollo en Criotecnología de Alimentos. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Centro de Investigación y Desarrollo en Criotecnología de Alimentos. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Centro de Investigación y Desarrollo en Criotecnología de Alimentos; ArgentinaFil: Bakas, L.. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Departamento de Ciencias Biológicas. Laboratorio de Investigación de Proteínas Vegetales; ArgentinaFil: Morcelle del Valle, Susana Raquel. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Departamento de Ciencias Biológicas. Laboratorio de Investigación de Proteínas Vegetales; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin

Application of culture-dependent and culture-independent methods for the identification of Lactobacillus kefiranofaciens in microbial consortia present in kefir grains. Food Microbiol

a b s t r a c t The biological and technological characteristics of kefiran as well as its importance in grain integrity led us to analyze the microbial kefir grain consortium with focus on Lactobacillus kefiranofaciens. The presence of L. kefiranofaciens in the nine kefir grains studied was demonstrated by denaturing gradient gel electrophoresis. By culture dependent methods applying a methodology focused on the search of this species, 22 isolates with typical morphology were obtained and identified applying a combination of SDS-PAGE of whole cell proteins, (GTG) 5 -PCR and sequence analysis of the housekeeping gene encoding the a-subunit of bacterial phenylalanyl-tRNA synthase (pheS). This polyphasic approach allowed the reliable identification of 11 L. kefiranofaciens, 5 Lactobacillus paracasei, 4 Lactobacillus kefiri and 2 Lactobacillus parakefiri isolates. Isolated L. kefiranofaciens strains produced polysaccharide in straindependent concentrations and EPS produced by them also differed in the degree of polymerization. The isolation and accurate identification of L. kefiranofaciens is relevant taking into account the important role of this microorganism in the grain ecosystem as well as its potential application as starter in food fermentations

Local Treatment with Lactate Prevents Intestinal Inflammation in the TNBS-Induced Colitis Model.

International audienceLactate has long been considered as a metabolic by-product of cells. Recently, this view has been changed by the observation that lactate can act as a signaling molecule and regulates critical functions of the immune system. We previously identified lactate as the component responsible for the modulation of innate immune epithelial response of fermented milk supernatants in vitro. We have also shown that lactate downregulates proinflammatory responses of macrophages and dendritic cells. So far, in vivo effects of lactate on intestinal inflammation have not been reported. We evaluated the effect of intrarectal administration of lactate in a murine model of colitis induced by 2,4,6-trinitrobenzenesulfonic acid (TNBS). The increase in lactate concentration in colon promoted protective effects against TNBS-induced colitis preventing histopathological damage, as well as bacterial translocation and rise of IL-6 levels in serum. Using intestinal epithelial reporter cells, we found that flagellin treatment induced reporter gene expression, which was abrogated by lactate treatment as well as by glycolysis inhibitors. Furthermore, lactate treatment modulated glucose uptake, indicating that high levels of extracellular lactate can impair metabolic reprograming induced by proinflammatory activation. These results suggest that lactate could be a potential beneficial microbiota metabolite and may constitute an overlooked effector with modulatory properties