High selectivity microporous silica membranes for lactic acid dehydration

Lactic acid (LA) has significant market potential for many industries including food, cosmetics, pharmaceuticals, medical and biodegradable materials. Production of LA usually begins with the fermentation of glucose but subsequent stages for the enrichment of lactic acid are complex and energy intensive and could be minimised using water selective membrane technology. In this work, we trialled a highly selective hydrostable carbonised template molecular sieve silica (CTMSS) membrane for the dehydration of a 15 vol% aqueous lactic acid solution with 0.1 vol% glucose. CTMSS membrane films were developed by dip-coating ceramic substrates with silica sols made using the acid catalysed sol-gel process. Permeation was performed by feeding LA/glucose solution to the membrane cell at 18°C in a standard pervaporation setup. The membrane showed selective transport of water from the aqueous feed to the permeate while glucose was not detected. CTMSS membrane permeate flux stabilised at 0.2 kg.m-2.hr-1 in 3.9 hours, and reduced LA to lower than 0.2 vol%. Flux through the CTMSS micropores was activated, displaying increased initial flux to 1.58 kg.m-2.hr-1 at 60°C. To enrich a 1 l.min-1 stream to 85% LA in a single stage, a minimum membrane area of 324 m2 would be required at 18°C. Increased operating temperature to 80°C significantly reduced this area to 24 m2 but LA levels in the permeate stream increased to 0.5 vol%. The highly selective CTMSS membrane technology is an ideal candidate for LA purification. CTMSS membrane systems operate stably in aqueous systems leading to potential cost reductions in LA processing for future markets.

Growth and lactic acid production in batch culture of Lactobacillus rhamnosus in a defined medium

To facilitate metabolic analysis, batch fermentations of Lactobacillus rhamnosus were carried out in a new defined medium. Biomass at 10.5 g/l and lactic acid at 67 g/l with a Y-P/S of 0.84 were achieved. The maximum specific growth rate and the average productivity were 0.49/h and 2.48 g/l.h, respectively. These are comparable to those of this organism and related organisms in complex media. Preliminary amino acid studies were also conducted, highlighting the importance of serine, asparagine, glutamine and cysteine. Kinetic analysis revealed that lactic acid production was predominantly growth-associated with growth associated and non-growth associated lactic acid constants of 0.389 mol/g-cell and 0.0025 mol/g-cell.h, respectively. Finally a kinetic model has been included to describe the fermentation of L. rhamnosus.

Pt/TiO(2)/poly(vinyl sulfonic acid) Layer-by-Layer Films for Methanol Electrocatalytic Oxidation

One major challenge for the widespread application of direct methanol fuel cells (DMFCs) is to decrease the amount of platinum used in the electrodes, which has motivated a search for novel electrodes containing platinum nanoparticles. In this study, platinum nanoparticles were electrodeposited on layer-by-layer (LbL) films from TiO(2) and poly(vinyl sulfonic) (PVS), by immersing the films into a H(2)PtCl(6) solution and applying a 100 mu A current during different electrode position times. Scanning tunnel microscopy (STM) and atomic force microscopy (AFM) images showed increased platinum particle size and electrode roughness for increasing electrodeposition times. The potentiodynamic profile of the electrodes indicated that oxygen-like species in 0.5 mol L(-1) H(2)SO(4) were formed at less positive potentials for the smallest platinum particles. Electrochemical impedance spectroscopy measurements confirmed the high reactivity for the water dissociation and the large amount of oxygen-like species adsorbed on the smallest platinum nanoparticles. This high oxophilicity of the smallest nanoparticles was responsible for the electrocatalytic activity of Pt-TiO(2)/PVS systems for methanol electrooxidation, according to the Langmuir-Hinshelwood bifunctional mechanism. Significantly, the approach used here combining platinum electrodeposition and LbL matrices allows one to both control the particle size and optimize methanol electrooxidation, being therefore promising for producing membrane-electrode assemblies of DMFCs.

Influence of Lactic Acid on Endogenous and Viral RNA-Induced Immune Mediator Production by Vaginal Epithelial Cells

OBJECTIVE: To evaluate the influence of lactic acid on immune mediator release from vaginal epithelial cells. METHODS: The human vaginal epithelial cell line, VK2/E6E7, was cultured in the presence or absence of physiological concentrations of lactic acid, and in the presence or absence of the viral Toll-like receptor 3 agonist, poly (inosinic acid: cytidylic acid). Supernatants were assayed by enzyme-linked immunosorbent assay (ELISA) for interleukin (IL)-1 beta, IL-6, IL-8, IL-23, transforming growth factor (TGF)-beta and secretory leukocyte protease inhibitor. RESULTS: Vaginal epithelial cells spontaneously released IL-1 beta (25.9 pg/mL), IL-8 (1.0 ng/mL), TGF-beta (175 pg/mL), and secretory leukocyte protease inhibitor (33.8 ng/mL). Only TGF-beta production was marginally enhanced (49%) by addition of lactic acid alone. Poly (inosinic acid: cytidylic acid) by itself stimulated the release of IL-6 (305 pg/mL) and enhanced IL-8 production (2.8 ng/mL). The combination of poly (inosinic acid: cytidylic acid) and lactic acid markedly increased IL-8 production (5.0 ng/mL) and induced the release of IL-1 beta (96.2 pg/mL). The poly (inosinic acid: cytidylic acid)-mediated lactic acid effect on IL-1 beta and IL-8 release was abrogated when the lactic acid was neutralized or if acetic acid was substituted for lactic acid. CONCLUSION: Lactic acid enhances the release of selective mediators from vaginal epithelial cells and stimulates antiviral immune responses. (Obstet Gynecol 2011;118:840-6) DOI: 10.1097/AOG.0b013e31822da9e9

New bio-strategies for ochratoxin A detoxification using lactic acid bacteria

The occurrence of mycotoxigenic moulds such as Aspergillus, Penicillium and Fusarium in food and feed has an important impact on public health, by the appearance of acute and chronic mycotoxicoses in humans and animals, which is more severe in the developing countries due to lack of food security, poverty and malnutrition. This mould contamination also constitutes a major economic problem due the lost of crop production. A great variety of filamentous fungi is able to produce highly toxic secondary metabolites known as mycotoxins. Most of the mycotoxins are carcinogenic, mutagenic, neurotoxic and immunosuppressive, being ochratoxin A (OTA) one of the most important. OTA is toxic to animals and humans, mainly due to its nephrotoxic properties. Several approaches have been developed for decontamination of mycotoxins in foods, such as, prevention of contamination, biodegradation of mycotoxins-containing food and feed with microorganisms or enzymes and inhibition or absorption of mycotoxin content of consumed food into the digestive tract. Some group of Gram-positive bacteria named lactic acid bacteria (LAB) are able to release some molecules that can influence the mould growth, improving the shelf life of many fermented products and reducing health risks due to exposure to mycotoxins. Some LAB are capable of mycotoxin detoxification. Recently our group was the first to describe the ability of LAB strains to biodegrade OTA, more specifically, Pediococcus parvulus strains isolated from Douro wines. The pathway of this biodegradation was identified previously in other microorganisms. OTA can be degraded through the hydrolysis of the amide bond that links the L-β-phenylalanine molecule to the ochratoxin alpha (OTα) a non toxic compound. It is known that some peptidases from different origins can mediate the hydrolysis reaction like, carboxypeptidase A an enzyme from the bovine pancreas, a commercial lipase and several commercial proteases. So, we wanted to have a better understanding of this OTA degradation process when LAB are involved and identify which molecules where present in this process. For achieving our aim we used some bioinformatics tools (BLAST, CLUSTALX2, CLC Sequence Viewer 7, Finch TV). We also designed specific primers and realized gene specific PCR. The template DNA used came from LAB strains samples of our previous work, and other DNA LAB strains isolated from elderberry fruit, silage, milk and sausages. Through the employment of bioinformatics tools it was possible to identify several proteins belonging to the carboxypeptidase family that participate in the process of OTA degradation, such as serine type D-Ala-D-Ala carboxypeptidase and membrane carboxypeptidase. In conclusions, this work has identified carboxypeptidase proteins being one of the molecules present in the OTA degradation process when LAB are involved.

Lactic acid bacteria from fresh fruit and vegetables as biocontrol agents of phytopathogenic bacteria and fungi

This study evaluated the efficacy of lactic acid bacteria (LAB) isolated from fresh fruits and vegetables as biocontrol agents against the phytopathogenic and spoilage bacteria and fungi, Xanthomonas campestris, Erwinia carotovora, Penicillium expansum, Monilinia laxa, and Botrytis cinerea. The antagonistic activity of 496 LAB strains was tested in vitro and all tested microorganisms except P. expansum were inhibited by at least one isolate. The 496 isolates were also analyzed for the inhibition of P. expansum infection in wounds of Golden Delicious apples. Four strains (TC97, AC318, TM319, and FF441) reduced the fungal rot diameter of the apples by 20%; only Weissella cibaria strain TM128 decreased infection levels by 50%. Cell-free supernatants of selected antagonistic bacteria were studied to determine the nature of the antimicrobial compounds produced. Organic acids were the preferred mediators of inhibition but hydrogen peroxide was also detected when strains BC48, TM128, PM141 and FF441 were tested against E. carotovora. While previous reports of antifungal activity by LAB are scarce, our results support the potential of LAB as biocontrol agents against postharvest rot. [Int Microbiol 2008; 11(4):231-236]

Evaluation of thermotolerant capacity of lactic acid bacteria isolated from commercial sausages and the effects of their addition on the quality of cooked sausages

The thermotolerant capacity of several lactic acid bacteria strains isolated from cooked commercial sausages was determined. Four strains were positively identified as Lactobacillus plantarum, Lactobacillus curvatus, Pediococcus pentosaceus and Pediococcus acidilacti, after surviving thermal treatment (70 °C during 60 minutes). Thermotolerant strains were inoculated in sausage batters before cooking in order to determine their effect on color, texture, acceptance and inhibition of Enterobacteria during 12 days at 8 °C. No significant effect of the inoculated strains was detected on color parameters. Textural profile parameters, cohesiveness and resilience, were not affected by the inoculation of thermotolerant lactic acid bacteria, but L. curvatus sausages resulted softer than the rest of the treatments. Samples inoculated with L. curvatus also obtained the lowest scores for the sensory attributes evaluated, with the remaining treatments causing no unfavorable effects on sausage acceptance. There was a reduction in enterobacterial counts after 12 days of cold storage in inoculated samples. The performance of inoculated lactic acid bacteria strains can be explained in a similar way as that of starter cultures in dry-fermented sausages, where the growth in nests impairs other pathogenic microorganisms present in the rest of the sausage, since environmental conditions and the early inoculation of these thermotolerant strains favor them to become the dominant flora.

Screening and kinetics of glutaminase and glutamate decarboxylase producing lactic acid bacteria from fermented Thai foods

L-glutaminase and glutamic acid decarboxylase (GAD) catalyzes the hydrolysis of L-glutamine and glutamate, respectively. L-glutaminase widely used in cancer therapy along with a combination of other enzymes and most importantly these enzymes were used in food industries, as a major catalyst of bioconversion. The current investigation was aimed to screen and select L-glutaminase, and GAD producing lactic acid bacteria (LAB). A total of 338 LAB were isolated from fermented meat, fermented fish, fermented soya bean, fermented vegetables and fruits. Among 338 isolates, 22 and 237 LAB has been found to be positive for L-glutaminase and GAD, respectively. We found that 30 days of incubation at 35 ºC and pH 6.0 was the optimum condition for glutaminase activity by G507/1. G254/2 was found to be the best for GAD activity with the optimum condition of pH 6.5, temperature 40 ºC and ten days of incubation. These LAB strains, G507/1 and G254/2, were identified as close relative of Lactobacillus brevis ATCC 14869 and Lactobacillus fermentum NBRC 3956, respectively by 16S rRNA sequencing. Further, improvements in up-stream of the fermentation process with these LAB strains are currently under development.

Studies on exopolysaccharide production by probiotic Lactic acid bacteria

The thesis mainly discussed the isolation and identification of a probiotic Lactobacillus plantarum, fermentative production of exopolysaccharide by the strain, its purification, structural characterisation and possible applications in food industry and therapeutics. The studies on the probiotic characterization explored the tolerance of the isolated LAB cultures to acid, bile, phenol, salt and mucin binding. These are some of the key factors that could satisfy the criteria for probiotic strains . The important factors required for a high EPS production in submerged fermentation was investigated with a collection of statistical and mathematical approach. Chapter 5 of the thesis explains the structural elucidation of EPS employing spectroscopic and chromatographic techniques. The studies helped in the exploration of the hetero-polysaccharide sequence from L. plantarum MTCC 9510. The thesis also explored the bioactivities of EPS from L. plantarum. As majority of chemical compounds identified as anti-cancerous are toxic to normal cells, the discovery and identification of new safe drugs has become an important goal of research in the biomedical sciences. The thesis has explored the anti-oxidant, anti-tumour and immunomodulating properties of EPS purified from Lactobacillus plantarum. The presence of (1, 3) linkages and its molecular weight presented the EPS with anti-oxidant, anti-tumour and immunomodulating properties under in vitro conditions.

Diversity and antimicrobial activity of Lactic Acid Bacteria from the gut of marine fish Rastrelliger kanagurta against fish, shrimp and human pathogens

Emergence of antibiotic resistance among aquaculture pathogens has made it necessary to look into environment friendly, effective and sustainable methods such as probiotic and immunostimulants among others.. In the present study, LAB were isolated from the gut of fish species namely Rastrelliger kanagurta and analyzed for their antibacterial activity against various fish, shrimp and human pathogens. Different LAB species such as Lactobacillus plantarum, L. bulgaricus, L. brevis and L. viridiscens were encountered in the gut of R. kanagurta. Several strains showed good activity against fish, shrimp and human pathogens. LAB from the gut of such marine species may be developed as possible probiont for environment friendly health management of fresh water, estuarine and marine species currently exploited in aquaculture

Lactic acid bacteria as bioprotective agents against foodborne pathogens and spoilage microorganisms in fresh fruits and vegetables

La present tesi doctoral es centra en l'aplicació dels bacteris de l'àcid lactic (BAL) com a agents bioprotectors davant microorganismes patògens i deteriorants.Es van aïllar i seleccionar BAL de fruites i hortalisses fresques i es van assajar in vitro davant 5 microorganismes fitopatògens i 5 patògens humans.Es van realitzar assajos d'eficàcia en pomes Golden Delicious amb tots els aïllats enfront les infeccions causades pel fong Penicillium expansum. La soca més eficaç era Weissella cibaria TM128, que reduïa el diàmetre de les infeccions en un 50%.Les soques seleccionades es van assajar enfront els patògens Salmonella typhimurium, Escherichia coli i Listeria monocytogenes en enciams Iceberg i pomes Golden Delicious.Els BAL interferien eficientment amb el creixemet de S. typhimurium, and L. monocytogenes, però van mostrar poc efecte enfront E. coli.Finalment, es van realitzar assajos dosi-resposta amb les soques Leuconostoc mesenteroides CM135, CM160 and PM249 enfront L. monocytogenes. De totes les soques assajades, la soca CM160 va ser la més efectiva.

A dynamic mechanistic model of lactic acid metabolism in the rumen

Current feed evaluation systems for ruminants are too imprecise to describe diets in terms of their acidosis risk. The dynamic mechanistic model described herein arises from the integration of a lactic acid (La) metabolism module into an extant model of whole-rumen function. The model was evaluated using published data from cows and sheep fed a range of diets or infused with various doses of La. The model performed well in simulating peak rumen La concentrations (coefficient of determination = 0.96; root mean square prediction error = 16.96% of observed mean), although frequency of sampling for the published data prevented a comprehensive comparison of prediction of time to peak La accumulation. The model showed a tendency for increased La accumulation following feeding of diets rich in nonstructural carbohydrates, although less-soluble starch sources such as corn tended to limit rumen La concentration. Simulated La absorption from the rumen remained low throughout the feeding cycle. The competition between bacteria and protozoa for rumen La suggests a variable contribution of protozoa to total La utilization. However, the model was unable to simulate the effects of defaunation on rumen La metabolism, indicating a need for a more detailed description of protozoal metabolism. The model could form the basis of a feed evaluation system with regard to rumen La metabolism.

Susceptibility of Saccharomyces cerevisiae and lactic acid bacteria from the alcohol industry to several antimicrobial compounds

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Lactic acid production by new Lactobacillus plantarum LMISM6 grown in molasses: optimization of medium composition

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)