RESUMO
The aim of this work was to evaluate the suitability of incorporating Fe3O4 (magnetite, M) NPs into water kefir (wKef) beverages. Magnetite NPs were synthesized and coated with pectins (cM), and incorporated into wKef beverages obtained by fermentation of a muscovado sugar solution with wKef grains. FeSO4, usually employed as fortifier, was used as a control. Four different beverages were analyzed: wKef, wKef-cM, wKef-M, wKef-FeSO4, indicating wKef beverages fortified with cM, M or FeSO4, respectively. Their stability was assessed by determining the viability of total lactic acid bacteria and yeasts, and the composition of saccharides along storage at 4 °C for up to 30 days. The toxicity of M and cM was evaluated in an in vivo model of Artemia salina. The absorption of iron was quantified by determining ferritin values on intestinal Caco-2/TC7 cells, and its internalization mechanisms, by employing inhibitors of endocytic pathways and quantifying ferritin. M and cM were non-toxic on Artemia salina up to 500 µg/mL, a toxicity even lower than that of FeSO4, which showed a LD50 of 304.08 µg/mL. After 30 days of storage, no significant decrease on yeasts viability was observed, and bacteria viability was above 6 log CFU/mL for the four beverages. In turn, sucrose decreased to undetectable values, concomitantly to an increase in the concentrations of glucose and fructose. Both wKef-M and wKef-cM led to a significant increase in the ferritin values (up to 2 folds) with regard to the basal state. The internalization of M NPs occurred via clathrins and caveolin pathways, whereas that of cM, by macropinocytosis. Safely incorporating M and cM NPs into wKef beverages appear as an innovative strategy for providing bioavailable iron aiming to ameliorate the nutritional status of populations at risk of iron deficiency (e.g., vegans).
Assuntos
Kefir , Nanopartículas de Magnetita , Células CACO-2 , Humanos , Ferro , ÁguaRESUMO
AIMS: This study evaluated whether by-products from industrial processing of acerola (Malpighia glabra L.; AB) and guava (Psidium guajava L.; GB) fruit may stimulate the growth and metabolism of probiotic Lactobacillus and Bifidobacterium and induce changes in human colonic microbiota. METHODS AND RESULTS: The ability of non-digested and digested AB or GB to stimulate the growth ad metabolism of Lactobacillus acidophilus LA-05, Lactobacillus casei L-26 and Bifidobacterium animalis subsp. lactis BB-12 was evaluated. Changes in populations of distinct bacterial groups of human colonic microbiota induced by digested AB and GB were evaluated using an in vitro colonic fermentation system. Non-digested and digested AB and GB favoured probiotic growth. No difference among counts of probiotics in media with glucose, fructooligosaccharides and non-digested and digested AB and GB was found during a 48-h cultivation. Cultivation of probiotics in media with non-digested and digested AB and GB resulted in decreased pH, increased organic acid production and sugar consumption over time. Digested AB and GB caused overall beneficial changes in abundance of Bifidobacterium spp., Lactobacillus-Enterococcus, Eubacterium rectall-Clostridium coccoides and Bacteroides-Provotella populations, besides to decrease the pH and increase the short-chain fatty acid production during a 24-h in vitro colonic fermentation. CONCLUSION: AB and GB could be novel prebiotic ingredients because they can stimulate the growth and metabolism of probiotics and induce overall beneficial changes in human colonic microbiota. SIGNIFICANCE AND IMPACT OF THE STUDY: AB and GB stimulated the growth and metabolism of probiotics, in addition to induce beneficial alterations in human colonic microbiota composition and increase short-chain fatty acid production. These results characterize AB and GB as potential prebiotic ingredients and fruit processing by-products as sources of added-value compounds.
Assuntos
Bifidobacterium animalis/crescimento & desenvolvimento , Colo/microbiologia , Lactobacillus/crescimento & desenvolvimento , Malpighiaceae/metabolismo , Prebióticos/análise , Probióticos/análise , Psidium/metabolismo , Resíduos/análise , Bifidobacterium animalis/metabolismo , Clostridiales , Ácidos Graxos Voláteis/metabolismo , Fermentação , Frutas/química , Frutas/metabolismo , Microbioma Gastrointestinal , Humanos , Lactobacillus/metabolismo , Lactobacillus acidophilus/crescimento & desenvolvimento , Malpighiaceae/química , Oligossacarídeos/análise , Oligossacarídeos/metabolismo , Probióticos/metabolismo , Psidium/químicaRESUMO
Phospholipids are building blocks of biological membranes having a key role in cellular functionality. The presence of unsaturated fatty acids in their conformation makes them prompt to oxidation reactions, leading to dysfunctions of living cells or to instability of lipid containing food products. The aim of this review is to gather together the latest advances on the understanding on lipids' peroxidation, using liposomes as model systems, including the main available analytical methods to monitor peroxidation reactions, with special emphasis on Fourier Transform Infrared (FTIR) and Raman spectroscopies. Lipid peroxidation is the most widely studied free radical chain reaction, which occurs in three steps: initiation, propagation and termination, making difficult to determine peroxidation products. Using liposomes as model membrane systems provides a useful tool to investigate the effects of free radicals. Different analytical methods enable the determination of peroxidation primary or secondary products. In particular, FTIR and Raman spectroscopies allow the simultaneous determination of peroxidation products in a non-destructive and easy-to-use manner. A quick monitoring of both reagents and products provides a reliable method for the quality control of industrial products or even for diagnostics, thus underlying the strong potential of vibrational spectroscopic based techniques.
Assuntos
Lipossomos , Fosfolipídeos , Radicais Livres , Peroxidação de Lipídeos , OxirreduçãoRESUMO
Eukaryotic antimicrobial peptides (AMPs) interact with plasma membrane of bacteria, fungi and eukaryotic parasites. Noteworthy, Lactobacillus delbrueckii subsp. lactis (CIDCA 133) and L. delbrueckii subsp. bulgaricus (CIDCA 331) show different susceptibility to human beta-defensins (ß-sheet peptides). In the present work we extended the study to α-helical peptides from anuran amphibian (Aurein 1.2, Citropin 1.1 and Maculatin 1.1). We studied the effect on whole bacteria and liposomes formulated with bacterial lipids through growth kinetics, flow cytometry, leakage of liposome content and studies of peptide insertion in lipid monolayers. Growth of strain CIDCA 331 was dramatically inhibited in the presence of all three peptides and minimal inhibitory concentrations were lower than those for strain CIDCA 133. Flow cytometry revealed that AMPs lead to the permeabilization of bacteria. In addition, CIDCA 331-derived liposomes showed high susceptibility, leading to content leakage and structural disruption. Accordingly, peptide insertion in lipid monolayers demonstrated spontaneous interaction of AMPs with CIDCA 331 lipids. In contrast, lipids monolayers from strain CIDCA 133 were less susceptible. Summarizing we demonstrate that the high resistance of the probiotic strain CIDCA 133 to AMPs extends to α helix peptides Aurein, Citropin and Maculatin. This behavior could be ascribed in part to differences in membrane composition. These findings, along with the previously demonstrated resistance to ß defensins from human origin, suggest that strain CIDCA 133 is well adapted to host innate immune effectors from both mammals and amphibians thus indicating conserved mechanisms of interaction with key components of the innate immune system.
Assuntos
Antibacterianos/farmacologia , Lactobacillus/efeitos dos fármacos , Lipossomos , Peptídeos/farmacologia , Sequência de Aminoácidos , Antibacterianos/química , Testes de Sensibilidade Microbiana , Peptídeos/química , Conformação Proteica em alfa-HéliceRESUMO
Iron deficiency is the most common nutritional deficit worldwide. The goal of this work was to obtain iron-pectin beads by ionic gelation and evaluate their physiological behavior to support their potential application in the food industry. The beads were firstly analyzed by scanning electronic microscopy, and then physical-chemically characterized by performing swelling, thermogravimetric, porosimetry, Mössbauer spectroscopy and X-ray fluorescence analyses, as well as by determining the particle size. Then, physiological assays were carried out by exposing the beads to simulated gastric and intestinal environments, and determining the iron absorption and transepithelial transport into Caco-2/TC7 cells. Iron-pectin beads were spherical (diameter 1-2â¯mm), with high density (1.29â¯g/mL) and porosity (93.28%) at low pressure, indicating their high permeability even when exposed to low pressure. Swelling in simulated intestinal medium (pH 8) was higher than in simulated gastric medium. The source of iron [FeSO4 (control) or iron-pectin beads] did not have any significant effect on the mineral absorption. Regarding transport, the iron added to the apical pole of Caco-2/TC7 monolayers was recovered in the basal compartment, and this was proportional with the exposure time. After 4â¯h of incubation, the transport of iron arising from the beads was significantly higher than that of the iron from the control (FeSO4). For this reason, iron-pectin beads appear as an interesting system to overcome the low efficiency of iron transport, being a potential strategy to enrich food products with iron, without altering the sensory properties.
Assuntos
Sistemas de Liberação de Medicamentos , Intestinos/citologia , Ferro/administração & dosagem , Ferro/metabolismo , Pectinas/química , Células CACO-2 , Humanos , Ferro/química , Tamanho da Partícula , Propriedades de SuperfícieRESUMO
Oil-in-water (O/W) emulsions of okara oil-caseinate (1:2; 1:3 and 1:4 O/W ratios) were used to encapsulate Lactobacillus plantarum CIDCA 83114. Once encapsulated, microorganisms were freeze-dried or spray-dried, and observed by scanning electronic and confocal microscopies. A physical characterization of the dehydrated capsules was carried out by determining their moisture content, water activity, particle size, polydispersity index and zeta potential. Determining the induction times and peroxide values provided information about their susceptibility to oxidation. In turn, bacterial stability was analyzed by plate counting before and after freeze-drying and spray-drying, and during storage at 4°C. Spray-dried emulsions had lower Z-sizes and polydispersity indexes, higher induction times and lower peroxide values than the freeze-dried ones, thus resulting better systems to protect L. plantarum CIDCA 83114. In addition, the culturability of spray-dried bacteria did not decrease neither after spray-drying nor up to 60days of storage at 4°C. The results showed that the better physical-chemical stability of spray-dried capsules determined the greater stability of microorganisms. This demonstrates the importance of defining adequate emulsions' formulations for an efficient encapsulation of microorganisms, with promising applications in the development of novel functional foods.
Assuntos
Emulsões/análise , Indústria Alimentícia/métodos , Liofilização , Glycine max/química , Lactobacillus plantarum/crescimento & desenvolvimento , Óleos/química , Probióticos/administração & dosagem , Cápsulas , Caseínas , Contagem de Colônia Microbiana , Dessecação , Composição de Medicamentos/métodos , Alimento Funcional , Humanos , Resíduos Industriais , Viabilidade Microbiana , Microscopia/métodos , Oxirredução , Peróxidos/metabolismo , ÁguaRESUMO
Liposomes are generally used as delivery systems, as they are capable of encapsulating a wide variety of molecules (i.e. plasmids, recombinant proteins, therapeutic drugs). However, liposomal drug delivery have to fulfill different requirements, such as the effective internalization by the target cells and avoidance of the degradative activity of the intracellular compartments. The use of polymer lipid complexes (PLCs), by including different polymers in the liposome formulation, could improve internalization and intracellular release of drugs. The aim of the present work is to study the mechanisms of cellular uptaking and the intracellular trafficking of PLCs formed with cholesterol-poly(2-(dimethylamino)ethyl methacrylate) CHO-PDMAEMA and lecithin (LC CHO-PD). Calcein-loaded liposomes were used to determine cellular uptake and intracellular localization by flow cytometry and confocal microscopy. Incorporation of CHO-PDMAEMA to lecithin liposomes enhanced the internalization capacity of PLCs. Internalization of PLCs by human epithelial-like cells (HEK-293) diminished at 4°C, suggesting uptake by endocytosis. PLCs showed no co-localization with acidic compartments after internalization. Experiments with endocytosis inhibitors and co-localization of liposomes and albumin, suggested the caveolae endocytic pathway as the most probable route for intracellular trafficking of PLCs. In this work, we demonstrated an efficient uptake of LC CHO-PDs by human epithelial-like cells (HEK-293) through the non-degradative caveolae endocytic pathway. The mode of internalization and the intracellular fate of liposomes under study, suggest a promising use of LC CHO-PDs as drug delivery systems.
Assuntos
Colesterol/metabolismo , Endocitose , Lipossomos , Metacrilatos/metabolismo , Nylons/metabolismo , Células HEK293 , HumanosRESUMO
S-layers are paracrystalline bidimensional arrays of proteins or glycoproteins that overlay the cell surface of several genus and species of bacteria and archaea. As the outermost layer of several genus and species of microorganisms, S-layer proteins (SLP) are in direct contact with bacterial environment and thus may be involved in many of their surface properties, including adherence to various substrates, mucins and eukaryotic cells, aggregation and coaggregation with yeasts and other bacteria. In addition, SLP have been reported to be responsible for the bacterial protection against detrimental environmental conditions and to play an important role in surface recognition or as carriers of virulence factors. In this mini-review, we bring together the latest evidences about functional and mechanical properties of bacterial SLP from two different perspectives: (A) their role on bacterial adherence to different substrates and surfaces, and (B) their role as mechanical barriers in bacterial harmful environments.
Assuntos
Bactérias/metabolismo , Aderência Bacteriana/fisiologia , Proteínas da Membrana Bacteriana Externa/metabolismo , Glicoproteínas de Membrana/metabolismo , Bactérias/crescimento & desenvolvimento , Proteínas da Membrana Bacteriana Externa/imunologia , Biofilmes/crescimento & desenvolvimento , Membrana Celular/metabolismo , Glicoproteínas de Membrana/biossíntese , Glicoproteínas de Membrana/imunologia , Fatores de Virulência/metabolismoRESUMO
The aim of this work was to evaluate the changes due to acclimation to ethanol on the fatty acid composition of three oenological Lactobacillus plantarum strains and their effect on the resistance to ethanol and malic acid consumption (MAC). Lactobacillus plantarum UNQLp 133, UNQLp 65.3 and UNQLp 155 were acclimated in the presence of 6 or 10% v/v ethanol, for 48 h at 28°C. Lipids were extracted to obtain fatty acid methyl esters and analysed by gas chromatography interfaced with mass spectroscopy. The influence of change in fatty acid composition on the viability and MAC in synthetic wine was analysed by determining the Pearson correlation coefficient. Acclimated strains showed a significant change in the fatty composition with regard to the nonacclimated strains. Adaptation to ethanol led to a decrease in the unsaturated/saturated ratio, mainly resulting from an increase in the contribution of short-length fatty acid C12:0 and a decrease of C18:1. The content of C12:0 was related to a higher viability after inoculation of synthetic wine. The MAC increased at higher contents in saturated fatty acid, but its efficiency was strain dependent.
Assuntos
Etanol/farmacologia , Ácidos Graxos/análise , Lactobacillus plantarum/química , Lactobacillus plantarum/efeitos dos fármacos , Malatos/análise , Vinho/microbiologia , Adaptação Fisiológica , Cromatografia Gasosa , Farmacorresistência Bacteriana , Etanol/análise , Ácidos Graxos Insaturados/análise , Viabilidade MicrobianaRESUMO
AIMS: The aim of this work was to evaluate the effect of acclimation on the viability, membrane integrity and the ability to consume malic acid of three oenological strains of Lactobacillus plantarum. METHODS AND RESULTS: Cultures in the stationary phase were inoculated in an acclimation medium (Accl.) containing 0, 6 or 10% v/v ethanol and incubated 48 h at 28°C. After incubation, cells were harvested by centrifugation and inoculated in a synthetic wine, containing 14% v/v ethanol and pH 3.5 at 28°C. Viability and membrane integrity were determined by flow cytometry (FC) using carboxyfluorescein diacetate (cFDA) and propidium iodide. Bacterial growth and malic acid consumption were monitored in a synthetic wine during 15 days. In nonacclimated strains, the damage of bacterial membranes produced a dramatic decrease in microbial viability in synthetic wine. In contrast, survival of strains previously acclimated in Accl. with 6 and 10% v/v ethanol was noticeable higher. Therefore, acclimation with ethanol increased the cultivability in synthetic wine and consequently, the consumption of l-malic acid after 15 days of growth. CONCLUSION: Acclimation of oenological strains in media containing ethanol prior to wine inoculation significantly decreases the membrane damage and improves viability in the harsh wine conditions. The role of membrane integrity is crucial to warrant the degradation of l-malic acid. SIGNIFICANCE AND IMPACT OF THE STUDY: The efficiency of multiparametric FC in monitoring viability and membrane damage along with the malic acid consumption has a strong impact on winemaking because it represents a useful tool for a quick and highly reliable evaluation of oenological parameters.