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An evaluation of a new lactic acid polymer drug delivery system: a preliminary report.

A new drug delivery system, made of a mixture of DL-lactic acid polymer and a copoly (L-lactic acid/delta-valerolactone) polymer (LAP) containing varying concentrations of Ofloxacin, has been evaluated. Results revealed that a LAP drug delivery system containing 0.03% Ofloxacin inhibited the in vitro growth of clinically derived Staphylococcus aureus and Pseudomonas aeruginosa. Further, in an in vivo situation in rats, at 1 week after the wound was inflicted, no infection was macroscopically seen in split-thickness wounds that had been covered by LAP drug delivery system containing 0.5 or 0.03% Ofloxacin, and more than 80% of the wound area had epithelialised.

Animals↗

Growth and Metabolism of Lactic Acid Bacteria during and after Malolactic Fermentation of Wines at Different pH.

Commercially produced red wines were adjusted to pH 3.0, 3.2, 3.5, 3.7, or 4.0 and examined during and after malolactic fermentation for growth of lactic acid bacteria and changes in the concentrations of carbohydrates, organic acids, amino acids, and acetaldehyde. With one exception, Leuconostoc oenos conducted the malolactic fermentation in all wines and was the only species to occur in wines at pH below 3.5. Malolactic fermentation by L. oenos was accompanied by degradation of malic, citric, and fumaric acids and production of lactic and acetic acids. The concentrations of arginine, histidine, and acetaldehyde also decreased at this stage, but the behavior of hexose and pentose sugars was complicated by other factors. Pediococcus parvulus conducted the malolactic fermentation in one wine containing 72 mg of total sulfur dioxide per liter. Fumaric and citric acids were not degraded during this malolactic fermentation, but hexose sugars were metabolized. P. parvulus and species of Lactobacillus grew after malolactic fermentation in wines with pH adjusted above 3.5. This growth was accompanied by the utilization of wine sugars and production of lactic and acetic acids.

Journal Article↗

Biodiversity of lactic acid bacteria in Romanian dairy products.

Traditionally fermented dairy products are still a very important part of the daily food in Romania, especially for people living in the countryside. To study the biodiversity of lactic acid bacterium strains of these products, 110 samples (raw and fermented milk, sour cream, and cheese) were collected from farm houses, monasteries, and local markets throughout Romania. Lactic acid bacteria (LAB) were isolated using six different cultivation conditions. All 599 isolates were tested for their Gram reaction, catalase activity, and morphology. A rep-PCR fingerprinting technique with the (GTG)5 primer and, in some cases SDS-PAGE of total cell proteins and 16S rRNA gene sequencing were used to cluster and/or identify the LAB. The biodiversity of the isolated strains was correlated with the type of product and/or technology applied. The most frequent LAB found in Romanian raw milk and fermented dairy products were Lactococcus lactis, Leuconostoc spp., and Enterococcus spp. Among the latter, a new species E. saccharominimus was found.

Bacteria↗

Characteristics and antagonistic activity of lactic acid bacteria isolated from chilled fish products.

In fish products, contrary to other foods, the commercial use of starter cultures is relatively unexploited. In this study, 61 isolates of lactic acid bacteria from lightly preserved, chilled fish products were characterized with special emphasis on their antagonistic activity and their possible use as starter cultures. The strains were phenotypically identified and characterized with regard to gas production, carbohydrate fermentations, production of off odours, H2O2-production, antagonistic activity and growth at 2, 5 and 10 degrees C. Growth at 10 degrees C was detected for all strains within 4 days. At 5 degrees C, 90% of the isolates were growing within 3-9 days. At 2 degrees C only 33% of the strains were growing within 9 days. Fourteen strains were selected for further testing of antagonistic activity against spoilage- and pathogenic bacteria at different pH in a disc assay. Two strains, a Leuconostoc spp. (V 6) and a Lactobacillus plantarum (LKE 5), differed from the other lactic acid bacteria by a very wide inhibitory spectrum for V 6 and by very distinct inhibition zones for LKE 5. The mechanisms of their inhibitory effects are discussed.

Cold Temperature↗

Lactic acid kinetics in respiratory alkalosis.

OBJECTIVE: To evaluate the impact of respiratory alkalosis on the elimination of intravenously infused lactate. DESIGN: Prospective, randomized, crossover study. SETTING: Medical ICU of a university hospital. PATIENTS: Eight patients treated by ventilatory support for neurologic or neuromuscular diseases. INTERVENTIONS: Patients were investigated on two occasions: during normoventilation (pH 7.42 +/- 0.1, PCO2 41 +/- 2 torr [5.5 +/- 0.2 kPa]) and during respiratory alkalosis (pH 7.59 +/- 0.1, PCO2 27 +/- 2 torr [3.6 +/- 0.2 kPa]) induced by controlled hyperventilation. To evaluate lactate elimination kinetics, 1 mmol/kg body weight of L-lactic acid was infused over 5 mins. MEASUREMENTS AND MAIN RESULTS: Arterial lactate concentrations and blood gas values were determined before and repeatedly after the infusion. Lactate elimination variables were calculated from the plasma curve by using a two-compartment model. Respiratory alkalosis increased plasma lactate from 1.56 +/- 0.1 to 2.49 +/- 0.2 mmol/L (p less than .001). The lactate elimination half-life increased from 4.57 +/- 0.2 mins at pH 7.42, to 9.96 +/- 1.1 mins during pH 7.59 (p less than .01), and beta half-life increased from 12.2 +/- 1.9 to 44.1 +/- 1 mins (p less than .01). Whole-body clearance decreased 40% from 24.2 +/- 2.9 to 14.3 +/- 2.0 mL/kg body weight-min (p less than .01). CONCLUSIONS: Respiratory alkalosis increases the basal concentration of plasma lactate and decreases clearance of infused lactic acid. These findings provide further evidence of the adverse effects of alkalosis.

Aged↗

Role of DL-lactic acid as an intermediate in rumen metabolism of dairy cows.

The role of DL-lactic acid as an intermediate in the rumen of a Friesian X Holstein dairy cow adapted to a diet of hay ad libitum plus 12 kg of a concentrate mixture was studied in vitro and in vivo. Concentrations of soluble sugars in the rumen fluid became maximal at 30 min postfeeding, but at 90 min no sugars were detectable. The DL-lactate concentration increased very rapidly to about 30 mm at 30 min after feeding, whereas the maximum total VFA concentration was reached 15 min later. More than 80% of the DL-lactate fermented to VFA was converted by Megasphaera elsdenii. Whereas only 16% of L-lactate was fermented to propionate, 75% of the D-lactate was converted to propionic acid. When all soluble sugars had been fermented, the participation of M. elsdenii to lactate fermentation declined and fermentation patterns for D- and L-lactate became similar yielding mostly acetate. Except for a brief period immediately after feeding, DL-lactate did not appear to be an important precursor of VFA in the rumen of a cow adapted to concentrate feeding. DL-lactate may become a more important intermediate in rumen fermentation temporarily when dairy cows are gradually changed from a hay diet to a diet including concentrates. The first 30 d after parturition, when the changeover takes place, is an unstable period, during which the microbial population is changing to fit the new environment.

Animals↗

Anti-bacterial activity of Lactobacillus plantarum strain SK1 against Listeria monocytogenes is due to lactic acid production.

AIMS: The aim of this research was to investigate the potential of Lactobacillus plantarum strain SK1 for use as a biological control agent against Listeria monocytogenes and determine its mechanism of anti-listerial activity. METHODS AND RESULTS: Co-growth of Lact. plantarum SK1 and L. monocytogenes UMCC98 in MRS broth showed that anti-listerial activity of Lact. plantarum SK1 occurred during late log/early stationary phase of growth. This coincided with a reduction in broth pH to 4.26. Evidence obtained from the analysis of cell-free culture filtrates of strain SK1 grown in MRS broth using thin-layer chromatography and growth of L. monocytogenes in pH-adjusted culture filtrates suggested that the anti-listerial activity was due to lactic acid production alone. Trials of Lact. plantarum SK1 on radishes stored at 5 degrees C showed that it had statistically significant (P < 0.05) anti-listerial activity. CONCLUSIONS: The anti-listerial activity of Lact. plantarum SK1 was due to lactic acid production alone. A small-scale trial on radishes stored at 5 degrees C showed it to have significant anti-listerial activity in planta. SIGNIFICANCE AND IMPACT OF THE STUDY: This organism has potential as a biological control agent for L. monocytogenes.

Antibiosis↗

Comparative analysis of conserved genetic markers and adjacent DNA regions identified in beer-spoilage lactic acid bacteria.

AIMS: To conduct an inter-species comparative study on the nucleotide sequences of the conserved DNA regions surrounding ORF5, a genetic marker for differentiating beer-spoilage lactic acid bacteria. METHODS AND RESULTS: The conserved DNA regions surrounding ORF5 were examined by PCR analysis, using three beer-spoilage strains, Lactobacillus brevis ABBC45C, L. paracollinoides LA2T and Pediococcus damnosus ABBC478. As a result, the DNA regions containing ORF1-7, originally found in ABBC45C, appeared to be conserved among the three strains, while the downstream region was not found in L. paracollinoides LA2T and P. damnosus ABBC478. The sequencing analysis of the conserved DNA regions of LA2T and ABBC478 revealed ca 99% nucleotide sequence identities with that of ABBC45C. CONCLUSIONS: The nucleotide sequences of the ca 8.2 kb DNA regions containing ORF1-7 were virtually identical among the three strains belonging to different species. The internal organizations of the ORFs were found to be remarkably similar. SIGNIFICANCE AND IMPACT OF THE STUDY: The level of nucleotide sequence identities suggests the DNA regions surrounding ORF5 were horizontally acquired by these beer-spoilage strains belonging to the three different species of lactic acid bacteria.

Beer↗

Effect of spray-dried yogurt and lactic acid bacteria on the initiation and promotion stages of chemically induced skin carcinogenesis in mice.

The effects of dietary spray-dried yogurt powder product (YPP) and two strains of lactic acid bacteria on the initiation and promotion stages of carcinogenesis were investigated using the 7,12-dimethylbenz[a]anthracene (DMBA)-12-O-tetradecanoylphorbol-13-acetate (TPA) mouse skin carcinogenesis model. In two independent studies, Sencar mice were fed a diet containing 86%, 43%, or 0% YPP or the 0% YPP diet supplemented with viable cultures of Lactobacillus acidophilus or bifidobacteria. Animals were initiated with a single topical application of DMBA (10 nmol). Promotion began three weeks later with twice weekly treatment of TPA (1 microgram/200 microliters acetone). During the initiation study (Study 1) the experimental diets were fed for four weeks before and one week after DMBA treatment. All mice were then switched to the AIN-76 diet. For the promotion study (Study 2) the experimental diets were begun one week after initiation and fed during the remainder of the study. Gross appearance of tumors was assessed weekly. No statistically significant differences in body weight or food disappearance were observed among the diet groups during the studies. For Studies 1 and 2, final histologically verified papilloma incidence and multiplicity and carcinoma incidence were not statistically different. These data suggest that different levels of YPP or lactic acid bacteria fed during the initiation or promotion stage of carcinogenesis do not significantly affect chemically induced skin tumor development.

9,10-Dimethyl-1,2-benzanthracene↗

Selection and characterization of mixed starter cultures for lactic acid fermentation of carrot, cabbage, beet and onion vegetable mixtures.

An evaluation of various lactic acid bacteria (LAB) for the fermentation of cabbage, carrot and beet-based vegetable products was carried out. As part of a screening process, the growth of 15 cultures in a vegetable juice medium (VJM) was characterized by automated spectrophotometry. Acidification patterns as well as viability during storage of the LAB were also established. There were greater differences between the pure cultures than the mixed ones with respect to growth in VJM and viability during storage. Reductions in viable cell counts during storage of the fermented VJM occurred more rapidly with a Leuconostoc strain than for pediococci or lactobacilli. Inoculation of vegetables was carried out with cultures of Lactobacillus plantarum NK-312, Pediococcus acidilactici AFERM 772 and Leuconostoc mesenteroides BLAC which were rehydrated in a brine. This rehydration procedure was not detrimental to viability. During fermentation of a carrot/cabbage vegetable mix, sugar metabolism was characterized by the assimilation of both glucose and fructose, but sugars remained in the fermented vegetables when acidification stopped. The pH in the LAB-inoculated vegetables after 72 h at 20 degrees C was significantly lower (by 0.2 units) than the uninoculated control. Inoculation with LAB designed for silage fermentation resulted in the inhibition of acetic acid production, and reduced the production of ethanol during fermentation. The selection process on VJM enabled the preparation of a mixed culture that was more rapid than the silage inoculants in acidifying the medium and was more effective in reducing the production of gas during the fermentation and storage of the fermented vegetables.

Colony Count, Microbial↗

Body distribution of fully biodegradable [14C]-poly(lactic acid) nanoparticles coated with albumin after parenteral administration to rats.

Fully biodegradable polylactic acid (PLA) nanoparticles (90-250 nm) coated with human serum albumin (HSA) were prepared by high-pressure emulsification and solvent evaporation, using the protein as surfactant. A new analytical tool was developed, based on Mie's law and size exclusion chromatography, to establish that, after evaporation of the solvent, the protein saturates the surface of the nanoparticles, masking the PLA core. According to this technique, no HSA is encapsulated in the polymer matrix. A radiolabelled [14C]-PLA50 was synthesized to follow the fate of this new drug carrier after i.v. administration to rats. The time necessary to clear the albumin-coated nanoparticles from the plasma was significantly longer than for the uncoated ones but not extended enough to target cells other than mononuclear phagocytes. As deduced from whole-body autoradiography and quantitative distribution experiments, the 14C-labelled polymer is rapidly captured by liver, bone marrow, lymph nodes, spleen and peritoneal macrophages. Nanoparticle degradation was addressed following 14C excretion. The elimination of the 14C was quick on the first day (30% of the administered dose) but then slowed down. In fact, if the metabolism of the PLA proceeds to lactic acid which is rapidly converted into CO2 via the Krebs cycle (80% of the total excretion was fulfilled by the lungs), anabolism from the lactic acid may also have taken place leading to long-lasting radioactive remnants, by incorporation of 14C into endogenous compounds.

Animals↗

Isolation, identification and changes in the composition of lactic acid bacteria during the malting of two different barley cultivars.

Malt has a complex microbial population, which changes as the malting process commences. Little is known about the proliferation of lactic acid bacteria (LAB) in each of the malting phases. In this study, we determined the number of LAB present in the different phases of malting with Clipper and Prisma barley cultivars. The strains were identified to species level by using numerical analysis of total soluble cell protein patterns, RAPD-PCR banding patterns and 16S rRNA sequencing. The number of viable LAB in the barley before steep was higher in Prisma than Clipper (7.6 x 10(4) and 1.2 x 10(3) cfu/g, respectively). Despite this, the number of viable cells recorded in the first steep water was slightly higher for Clipper (9.0 x 10(5) cfu/g) than Prisma (5.5 x 10(5) cfu/g). More or less, the same cell numbers were recorded for the two barley cultivars after the first and second dry stands. Both cultivars displayed more or less the same cell numbers (3.7 x 10(7) for Clipper and 3.2 x 10(7) cfu/g for Prisma) after the third day of germination. However, a higher number of LAB were detected in the kilned Prisma malt (6.9 x 10(4) cfu/g) than the Clipper malt (1.5 x 10(4) cfu/g). Leuconostoc argentinum, Le. lactis and Weissella confusa were the most predominant in both cultivars. A few strains were identified as W. paramesenteroides (four strains), Lactobacillus casei (five strains), Lactococcus lactis (five strains) and Lb. rhamnosus (two strains). Lb. casei and Lb. rhamnosus were not isolated from the Prisma cultivar, while W. paramesenteroides and Lc. lactis were absent in the Clipper cultivar. Kilned malt of the Clipper cultivar contained predominantly Le. argentinum, whereas the Prisma cultivar contained mainly Lc. lactis. To our knowledge. this is the first report of LAB in Clipper and Prisma barley and the various malting phases. The influence of the various groups of lactic acid bacteria on the fermenting ability of brewers' yeast is currently being determined.

Colony Count, Microbial↗

Biocompatibility and effect on osteogenesis of poly(ortho ester) compared to poly(DL-lactic acid).

Implantation of demineralized bone induces new bone formation by the action of contained growth factors, of which bone morphogenetic proteins are of prime importance. A biodegradable polymer may be used as a carrier for demineralized bone particles or recombinant bone growth factors to prevent displacement of the implant, preserve its volume and shape, and assure sustained release of the incorporated active components. A polymer for this use should be biocompatible and completely absorbed without interfering with the osteogenesis. We investigated the host-tissue response and effect on demineralized bone-induced bone formation by two biodegradable polymers, a poly(ortho ester) and an amorphous low-molecular poly(DL-lactic acid). Both polymers had a plastic consistency, could easily be molded, and adhered well to the demineralized bone particles. Demineralized bone particles were implanted alone and in combination with each of the polymers in the abdominal muscles of 45 male Wistar rats. Four weeks after the operation the implants were recovered and subjected to (85)Sr uptake analysis to quantify bone formation and histologic examination. The poly(ortho ester) provoked little inflammation; it was largely absorbed by 4 weeks, and no qualitative or quantitative effect on bone formation was found. The poly(DL-lactic acid) provoked a chronic inflammation with multinuclear giant cells, macrophages with engulfed material, and proliferating fibroblasts; part of the material was still present, and the bone formation was inhibited.

Animals↗

Streptococcus alactolyticus is the dominating culturable lactic acid bacterium species in canine jejunum and feces of four fistulated dogs.

Canine intestinal lactic acid bacterium (LAB) population in four fistulated dogs was cultured and enumerated using MRS agar. LAB levels ranging from 1.4x10(6) to 1.5x10(7) CFU ml(-1) were obtained in jejunal chyme. In the fecal samples 7.0x10(7) and 2.0x10(8) CFU g(-1) were detected. Thirty randomly selected isolates growing in the highest sample dilutions were identified to species level using numerical analysis of 16S and 23S rDNA restriction fragment length polymorphism patterns (ribotyping) and 16S rDNA sequence analysis. According to these results, Streptococcus alactolyticus was the dominant culturable LAB species in both feces and jejunal chyme. In addition, Lactobacillus murinus and Lactobacillus reuteri were detected.

Animals↗

Comparative study of the efficiency of some additives in protecting lactic acid bacteria against freeze-drying.

Cultures of 14 lactic acid bacteria species were freeze-dried in 10 or 20% non-fat skim milk and in distilled water containing one of the following additives: bovine albumin, glycogen, dextran, polyethylene glycol (PEG) 1000, PEG 4000, PEG 6000, glycerol, beta-glycerophosphate, sodium glutamate, asparagine, or cysteine. Each of the potential protective agents tested exhibited marked variations in the protection afforded to different species, none of them was effective for the preservation of viability of thermophilic lactobacilli. However, glycerol provided effective protection for L. leichmannii ATCC 4797 (90% survival), while L. bulgaricus ATCC 11842 reached a viability of 78% with 0.04 M cysteine.

Asparagine↗

Isolation and characterization of bacteriocin-producing lactic acid bacteria from ready-to-eat food products.

Lactic acid bacteria isolated from a range of foods sold in ready-to-eat form were screened for bacteriocin production. Twenty-two bacteriocin-producing cultures were isolated from 14 of the 41 foods sampled. Bacteriocin-producing isolates from meat, fish and dairy products were Lactobacillus and Leuconostoc species typically found associated with these products. Most of these isolates gave only a narrow inhibitory spectrum although two showed activity against Listeria monocytogenes. Fruit and vegetable products gave a broader range of organisms but most of the bacteriocin-producing cultures were found to be strains of Lactococcus. Several lactococci produced a nisin-like activity, and showed a broad inhibitory spectrum against the indicator strains tested. The ease with which bacteriocin-producing strains could be isolated implies that they are already being safely consumed in food, and highlights the potential for using bacteriocin-producing cultures for biopreservation, especially in association with minimally processed products.

Bacteriocins↗

Development of plasmid vector and electroporation condition for gene transfer in sporogenic lactic acid bacterium, Bacillus coagulans.

Bacillus coagulans is a sporogenic lactic acid bacterium that ferments glucose and xylose, major components of plant biomass, a potential feedstock for cellulosic ethanol. The temperature and pH for optimum rate of growth of B. coagulans (50 to 55 degrees C, pH 5.0) are very similar to that of commercially developed fungal cellulases (50 degrees C; pH 4.8). Due to this match, simultaneous saccharification and fermentation (SSF) of cellulose to products by B. coagulans is expected to require less cellulase than needed if the SSF is conducted at a sub-optimal temperature, such as 30 degrees C, the optimum for yeast, the main biocatalyst used by the ethanol industry. To fully exploit B. coagulans as a platform organism, we have developed an electroporation method to transfer plasmid DNA into this genetically recalcitrant bacterium. We also constructed a B. coagulans/E. coli shuttle vector, plasmid pMSR10 that contains the rep region from a native plasmid (pMSR0) present in B. coagulans strain P4-102B. The native plasmid, pMSR0 (6823bp), has 9 ORFs, and replicates by rolling-circle mode of replication. Plasmid pNW33N, developed for Geobacillus stearothermophilus, was also transformed into this host and stably maintained while several other Bacillus/Escherichia coli shuttle vector plasmids were not transformed into B. coagulans. The transformation efficiency of B. coagulans strain P4-102B using the plasmids pNW33N or pMSR10 was about 1.5x10(16) per mole of DNA. The availability of shuttle vectors and an electroporation method is expected to aid in genetic and metabolic engineering of B. coagulans.

Amino Acid Sequence↗

Potential of bacteriocin-producing lactic acid bacteria for improvements in food safety and quality.

Lactic acid bacteria (LAB) have been used for centuries in the fermentation of a variety of dairy products. The preservative ability of LAB in foods is attributed to the production of anti-microbial metabolites including organic acids and bacteriocins. Bacteriocins generally exert their anti-microbial action by interfering with the cell wall or the membrane of target organisms, either by inhibiting cell wall biosynthesis or causing pore formation, subsequently resulting in death. The incorporation of bacteriocins as a biopreservative ingredient into model food systems has been studied extensively and has been shown to be effective in the control of pathogenic and spoilage microorganisms. However, a more practical and economic option of incorporating bacteriocins into foods can be the direct addition of bacteriocin-producing cultures into food. This paper presents an overview of the potential for using bacteriocin-producing LAB in foods for the improvement of the safety and quality of the final product. It describes the different genera of LAB with potential as biopreservatives, and presents an up-to-date classification system for the bacteriocins they produce. While the problems associated with the use of some bacteriocin-producing cultures in certain foods are elucidated, so also are the situations in which incorporation of the bacteriocin-producer into model food systems have been shown to be very effective.

Bacteria↗