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PubMed2006

Optimization of L-(+)-lactic acid production using pelletized filamentous Rhizopus oryzae NRRL 395.

Lactic acid is used as a food additive for flavor and preservation and a precursor in the development of poly-lactic acid, a product used to make biodegradable plastics and textiles. Rhizopus oryzae NRRL 395 is known to be a strain that produces optically pure L-(+)-lactic acid. The morphology of Rhizopus cultures is complex, forming filamentous, clumps, and pellet mycelia. Different morphology growth has significant effects on lactic acid production. In bioreactors, the filamentous or clump mycelia increase the viscosity of the medium, wrap around impellers, and block the nutrient transportation, leading to a decrease in production efficiency and bioreactor performance. Growing fungi in pellet form can significantly improve these problems. In this study, factors that affect lactic acid production in pelletized flask cultures using R. oryzae NRRL 395 were investigated in detail. Completely randomized designs were used to determine the influence of culture temperature, time, concentration of glucose, and inoculum size. Lactic acid fermentation using clump and pellet morphologies were performed in a 5 L fermentor at the optimal values obtained from flask culture. Finally, fed-batch culture was used to enhance the lactate concentration in broth. The final lactate concentration of fed-batch culture reached 92 g/L. The data presented in the article can provide useful information on optimizing lactic acid production using alternative source materials.

Biofilms↗

PubMed2001

Anaerobic conversion of lactic acid to acetic acid and 1, 2-propanediol by Lactobacillus buchneri.

The degradation of lactic acid under anoxic conditions was studied in several strains of Lactobacillus buchneri and in close relatives such as Lactobacillus parabuchneri, Lactobacillus kefir, and Lactobacillus hilgardii. Of these lactobacilli, L. buchneri and L. parabuchneri were able to degrade lactic acid under anoxic conditions, without requiring an external electron acceptor. Each mole of lactic acid was converted into approximately 0.5 mol of acetic acid, 0.5 mol of 1,2-propanediol, and traces of ethanol. Based on stoichiometry studies and the high levels of NAD-linked 1, 2-propanediol-dependent oxidoreductase (530 to 790 nmol min(-1) mg of protein(-1)), a novel pathway for anaerobic lactic acid degradation is proposed. The anaerobic degradation of lactic acid by L. buchneri does not support cell growth and is pH dependent. Acidic conditions are needed to induce the lactic-acid-degrading capacity of the cells and to maintain the lactic-acid-degrading activity. At a pH above 5.8 hardly any lactic acid degradation was observed. The exact function of anaerobic lactic acid degradation by L. buchneri is not certain, but some results indicate that it plays a role in maintaining cell viability.

Acetic Acid↗

PubMed2004

Differential inactivation of Listeria monocytogenes by D- and L-lactic acid.

AIMS: To determine inactivation of Listeria monocytogenes by the two lactic acid isomers. METHODS AND RESULTS: The survival of four strains with varying sensitivity to acid was determined following treatment with L- or D-lactic acid at 100 mmol l(-1) (pH 3.7) or HCl at pH 3.37. There was some, but not complete, similarity in the relative sensitivity of the four strains to the two types of acid. All strains were most sensitive to D-lactic acid, which gave 0.6-2.2 log units greater reduction than L-lactic acid midway in the inactivation curves. Even very low concentrations of the two isomers had an immediate effect on pH(i) which was identical for the two isomers. CONCLUSIONS: The results show that L. monocytogenes is more sensitive to D- than to L-lactic acid; however, this difference is less than the strain variation in L-lactic acid sensitivity. SIGNIFICANCE AND IMPACT OF THE STUDY: This work has implications for the application of lactic acid for food preservation as well as for the understanding of the antibacterial mechanisms of weak organic acids.

Colony Count, Microbial↗

PubMed2000

Elimination of amino acid interferences in the chiral ligand-exchange chromatographic analysis of lactic acid enantiomers in wine.

Chiral ligand-exchange liquid chromatography is used to identify and quantitate lactic acid enantiomers in wines that have or have not undergone malolactic fermentation. The stationary phase is (R)-penicillamine, which is bound lipophilically to a C18 bonded silica matrix. The mobile phase is 1mM copper sulfate, and the detection mode is ultraviolet. Serious interference from (S)-aspartic acid and other amino acids is eliminated by the use of propanesulfonic acid-type cation exchange solid-phase extraction cartridges prior to chromatographic analysis. Lactic acid enantiomers in wine are quantitated in the range of 10 to 500 mg/L. The detection limit is 3 mg/L. The method is also successful in the determination of lactic acid enantiomers in certain beers (e.g., lambic beers), kim-chi, sauerkraut, and various yogurts.

Amino Acids↗

PubMed1997

Effect of lactic acid on L-glutamate uptake in cultured astrocytes: mechanistic considerations.

Elevated levels of lactic acid can be deleterious to CNS tissue. Lactic acid is known to cause astroglial swelling and since glial swelling has been shown to inhibit L-glutamate (L-Glu) uptake, we examined whether one of the actions of lactic acid is to inhibit L-Glu uptake. Astrocyte cultures treated with lactic acid (25 mM; pH 6.1) showed an inhibition of L-Glu uptake by 65%. HCl (pH 6.1) also inhibited L-Glu uptake and this inhibition was potentiated by sodium lactate (25 mM). The inhibitory effect of lactic acid on L-Glu uptake was partially reversible and the reversibility was enhanced by hypothermia. Blocking glial swelling with D-mannitol, or treatment with antioxidants or hypothermia did not inhibit the effect of lactic acid on L-Glu uptake, indicating that swelling per se or free radicals, were not the factors in L-Glu uptake inhibition. Lactic acid induced a four-fold enhancement of L-Glu release and a seven-fold increase of K+ release. Our results suggest that lactic acid, by direct effect on pH, brings about a stimulation of K+ and L-Glu release which may be a factor in the inhibition of L-Glu uptake by lactic acid in astrocytes.

Animals↗

PubMed2001

Receptor-mediated delivery of all trans-retinoic acid to hepatocyte using poly(L-lactic acid) nanoparticles coated with galactose-carrying polystyrene.

All trans-retinoic acid (RA)-loaded poly(L-lactic acid) (PLA) nanoparticles coated with galactose-carrying polymer, as hepatocyte-specific targeting material using galactose ligands as recognition signals to asialoglycoprotein receptors were prepared by the diafiltration method. Effects of released RA from its loaded nanoparticles on morphology and DNA synthesis of hepatocytes were studied. Receptor-mediated endocytosis of the nanoparticles was checked by fluorescence and confocal laser microscopy. It was found that the shapes of most hepatocytes attached onto polystyrene dish precoated with collagen solution were flat and spreading at low concentration of RA for the RA-loaded nanoparticles, whereas their shapes were round at even low concentration of RA when RA was mixed with the nanoparticles. From the fluorescence and confocal laser microscopic studies, it was suggested that the nanoparticles coated with galactose-carrying polymers were internalized by the hepatocytes through the receptor-mediated mechanism. The RA-loaded nanoparticles were more potent stimulators of hepatocyte DNA synthesis than the free RA system in the presence of epidermal growth factor (EGF) owing to the controlled release of RA from the RA-loaded nanoparticles.

Animals↗

PubMed1997

Modelling the growth rate of Escherichia coli as a function of pH and lactic acid concentration.

The growth rate responses of Escherichia coli M23 (a nonpathogenic strain) to suboptimal pH and lactic acid concentration were determined. Growth rates were measured turbidimetrically at 20 degrees C in the range of pH 2.71 to 8.45. The total concentration of lactic acid was fixed at specific values, and the pH was varied by the addition of a strong acid (hydrochloric) or base (sodium hydroxide) to enable the determination of undissociated and dissociated lactic acid concentrations under each condition. In the absence of lactic acid, E. coli grew at pH 4.0 but not at pH 3.7 and was unable to grow in the presence of > or = 8.32 mM undissociated lactic acid. Growth rate was linearly related to hydrogen ion concentration in the absence of lactic acid. In the range 0 to 100 mM lactic acid, growth rate was also linearly related to undissociated lactic acid concentration. A mathematical model to describe these observations was developed based on a Bĕlehrádek-like model for the effects of water activity and temperature. This model was expanded to describe the effects of pH and lactic acid by the inclusion of novel terms for the inhibition due to the presence of hydrogen ions, undissociated lactic acid, and dissociated lactic acid species. Preliminary data obtained for 200 and 500 mM total lactic acid concentrations show that the response to very high lactic acid concentrations was less well described by the model. However, for 0 to 100 mM lactic acid, the model described well the qualitative and quantitative features of the response.

Cell Division↗

PubMed2003

Effects of lactic acid and NaCl on creatine kinase from rabbit muscle.

The lactic acid induced unfolding and the salt-induced folding of creatine kinase (CK) were studied by enzyme activity, fluorescence emission spectra, circular dichroism spectra, and native polyacrylamide gel electrophoresis. The results showed that the kinetics of CK inactivation was a monophase process. Lactic acid caused inactivation and unfolding of CK with no aggregation during CK denaturation. The unfolding of the whole molecule and the inactivation of CK in solutions of different concentration of lactic acid were compared. Much lower lactic acid concentration values were required to bring about inactivation than were required to produce significant conformational changes of the enzyme molecule. At higher concentrations of lactic acid (more than 0.2 mM) the CK dimers were partially dissociated, as proved by native polyacrylamide gel electrophoresis. NaCl induced the molten globule state with a compact structure after CK was denatured with 0.8 mM lactic acid, and the increasing of anions led to a tight side-chain. The above results suggest that the effect of lactic acid differed from that of other denaturants such as guanidine hydrochloride, HCI, or urea during CK folding, and the molten globule state indicates that intermediates exist during CK folding.

Animals↗

PubMed1998

Incorporation of urea nitrogen into fecal protein and plasma protein amino acids in elderly human volunteers after ingestion of lactic acid bacteria.

Health effects of fermented milks have been associated with the metabolic activity of lactic acid bacteria in the gastrointestinal tract. It has been proposed that an increased excretion of urea nitrogen via microbial protein may decrease the workload on kidneys and liver. Therefore, a study was carried out in healthy elderly human subjects to investigate the incorporation of [15N2]urea nitrogen into plasma and fecal proteins and amino acids. Over a period of 10 d 13 healthy elderly subjects ingested daily a freeze-dried microbial preparation which contained different genera of lactic acid bacteria and is used to produce fermented milk products. One of the strains was originally isolated from stool samples of elderly people from the Caucasus region (Lactobacillus plantarum). No stimulation of fecal protein-nitrogen excretion and no increase in 15N-abundances in fecal protein was measured following the administration of the viable microbial preparation and a [15N2]urea bolus. Tentatively, it was concluded that this may have been caused by the inability of the microbial culture to survive the gastro-intestinal passage and (or) by the absence of additional fermentable carbohydrates in the diet as energy source for bacterial protein synthesis in the large intestine. However, using a highly sensitive GC-C-IRMS method we observed a significant incorporation of 15N into plasma protein amino acids. 15N-Enrichments in single amino acids were found according to their participation in transamination reactions. The slight enrichment of lysine which is not transaminated in mammalian tissues may indicate a microbial synthesis and absorption of bacterial lysine.

Aged↗

PubMed2001

Effect of lactic acid administration in the drinking water during preslaughter feed withdrawal on Salmonella and Campylobacter contamination of broilers.

The crop is a known source of Salmonella and Campylobacter contamination. We evaluated the use of selected organic acids (0.5% acetic, lactic, or formic) in drinking water during a simulated 8-h pretransport feed withdrawal (FW). Salmonella typhimurium was recovered from 53/100 control crops and from 45/100 of crops from acetic acid-treated broilers. However, treatment with lactic acid (31/100) or formic acid (28/76) caused significant (P < 0.05) reduction in incidence. Reductions of recovered incidence were also associated with reduced numbers of S. typhimurium recovered (e.g., control, log 1.45 cfu/crop; lactic acid, 0.79 cfu/crop). In an additional commercial farm study, broilers were provided 0.44% lactic acid during a 10-h FW (4 h on the farm and 6 h transport) and pre-FW crop, post-FW crop, and pre-chill carcass wash samples were collected for Campylobacter and Salmonella detection. Crop contamination with Salmonella was significantly reduced by lactic acid treatment (6/175) as compared with controls (29/175). Importantly, Salmonella isolation incidence in prechill carcass rinses was significantly reduced by 52.4% with the use of lactic acid (26/175 vs. 55/176). Crop contamination with Campylobacter was significantly reduced by lactic acid treatment (62.3%) as compared with the controls (85.1%). Lactic acid also reduced the incidence of Campylobacter found on pre-chill carcass rinses by 14.7% compared with the controls. These studies suggest that incorporation of lactic acid in the drinking water during pretransport FW may reduce Salmonella and Campylobacter contamination of crops and broiler carcasses at processing.

Animals↗

PubMed2003

The effect of sodium acetate on the activity of L- and D-lactate dehydrogenases in Lactobacillus sakei NRIC 1071(T) and other lactic acid bacteria.

The effect of sodium acetate on the production of stereoisomers of lactic acid produced by Lactobacillus sakei NRIC 1071(T) and other lactic acid bacteria was studied. L. sakei NRIC 1071(T) started producing L-lactic acid at the early logarithmic phase and d-lactic acid at the late logarithmic phase. The activity of L-lactate dehydrogenase [EC 1.1.1.27, L-LDH] from the resting cells of L. sakei NRIC 1071(T) appeared at the early stage of the logarithmic phase during the growth, and the activity of D-lactate dehydrogenase [EC 1.1.1.28, D-LDH] at the late stage of the logarithmic phase. The resting cells and cell-free extracts of L. sakei NRIC 1071(T) did not produce DL-lactic acid from L- or D-lactic acid. Stained bands of L-LDH and D-LDH appeared in the cell-free extracts from the cells of L. sakei NRIC 1071(T). Consequently, L. sakei conclusively produced L- and D-lactic acid by the action of L-LDH and D-LDH. This finding leads to the conclusion that lactate racemase [EC 5.1.2.1] does not exist in this strain. When the specific activity of LDHs (the total activity of L-LDH plus D-LDH) from the cells cultivated in the presence of sodium acetate is compared with that cultivated in its absence, the ratio of the activity between the cells cultivated in the former condition and those in the latter fell from 1.7 on the cell-free extracts to 1.3 on the preparation of the QAE-Toyopearl 550c chromatography. This result indicates that the amount of LDHs in the cells of L. sake NRIC 1071(T) cultivated in the presence of 50 mM sodium acetate was much more than that in the cells cultivated in the absence of sodium acetate. The shift of the type of stereoisomers of lactic acid from the DL-type to the L-type is discussed in the case of L. sakei strains.

Bacteria↗

PubMed1991

Selection of Rhizopus strains for L(+)-lactic acid and gamma-linolenic acid production.

The production of L(+)-lactic acid and formation of gamma-linolenic acid by 50 Rhizopus strains growing on saccharidic substrates were investigated. Formation of acids was observed on solid cultivation media but mainly during submerged fermentation. Strains with the highest selectivity of both L(+)-lactic acid production and gamma-linolenic acid formation were tested in a laboratory fermenter. The best producer was treated by UV irradiation to increase the fatty acid content in the biomass, especially that of gamma-linolenic acid. The conversion of 10% saccharidic substrate by this newly prepared strain Rhizopus arrhizus CCM 8109 results in more than 95% of theoretical yield of L(+)-lactic acid and permits a volume productivity of 0.4 g gamma-linolenic acid per liter.

Biotechnology↗

PubMed2001

High-rate continuous production of lactic acid by Lactobacillus rhamnosus in a two-stage membrane cell-recycle bioreactor.

It is important to produce L(+)-lactic acid at the lowest cost possible for lactic acid to become a candidate monomer material for promising biodegradable polylactic acid. In an effort to develop a high-rate bioreactor that provides high productivity along with a high concentration of lactic acid, the performance of membrane cell-recycle bioreactor (MCRB) was investigated via experimental studies and simulation optimization. Due to greatly increased cell density, high lactic acid productivity, 21.6 g L(-1) h(-1), was obtained in the reactor. The lactic acid concentration, however, could not be increased higher than 83 g/L. When an additional continuous stirred tank reactor (CSTR) was attached next to the MCRB a higher lactic acid concentration of 87 g/L was produced at significant productivity expense. When the two MCRBs were connected in series, 92 g/L lactic acid could be produced with a productivity of 57 g L(-1) h(-1), the highest productivity among the reports of L(+)-lactic acid that obtained lactic acid concentration higher than 85 g/L using glucose substrate. Additionally, the investigation of lactic acid fermentation kinetics resulted in a successful model that represents the characteristics of lactic acid fermentation by Lactobacillus rhamnosus. The model was found to be applicable to most of the existing data with MCRBs and was in good agreement with Levenspiel's product-inhibition model, and the Luedeking-Piret equation for product-formation kinetics appeared to be effective in representing the fermentation kinetics. There was a distinctive difference in the production potential of cells (cell-density-related parameter in Luedeking-Piret equation) as lactic acid concentration increases over 55 g/L, and this finding led to a more precise estimation of bioreactor performance.

Bioreactors↗

PubMed2005

Poly-L-lactic acid as a facial filler.

Poly-L-lactic acid is a filler recently approved by the US FDA for the correction of facial lipoatrophy in patients infected with the human immunodeficiency virus (HIV). Currently, poly-L-lactic acid, sold under the brand name Sculptratrade mark (Dermik), is the only product approved by the FDA specifically for this indication. The market for poly-L-lactic acid will likely be larger than the HIV-infected population, as physicians use poly-L-lactic acid off-label to correct lipoatrophy associated with the normal aging process in non-HIV-infected patients. The benefits of poly-L-lactic acid are limited by the fact that multiple treatments are necessary to achieve the desired correction; its results are temporary and its cost is high.

Biocompatible Materials↗

PubMed2003

Effects of intermittent addition of cellulase for production of L-lactic acid from wastewater sludge by simultaneous saccharification and fermentation.

An attempt was made to create L-lactic acid, a precursor of poly-lactic acid, which is a biodegradable plastic, from wastewater sludge from the paper-manufacturing industry. The sludge contained a high percentage of cellulose and needed to be hydrolyzed to glucose by the action of the cellulase before being treating with lactic acid bacteria. Therefore, a method involving simultaneous saccharification and fermentation (SSF) was carried out. The optimum pH of the SSF for production of the lactic acid by the newly isolated lactic acid bacterium with a high selectively of L-lactic acid was found out to be around pH = 5.0, and the optimum temperature to be approximately 40 degrees C. On the basis of the measurement of the cell density changes in the lactic acid bacteria, it was ascertained that the bacterial activity could continue at a high level for a relatively long period of time, and that the L-lactic acid productivity was diminished by the rapid deactivation of the cellulase. With the intermittent addition of cellulase once daily for the sake of compensating for the cellulase deactivation, the L-lactic acid attained a maximum concentration of 16.9 g/L, i.e., a 72.2% yield based on the potential glucose contained in the sludge under optimum pH and temperature conditions.

Biodegradation, Environmental↗

PubMed1984

The role of increased lactic acid concentration in neovascularizations.

The important role of a prolonged increase in lactic acid concentration in the mechanism of neovascularization has been shown by the facts as follows. The lactic acid concentration is increased in vascularizing tissues. Increasing the lactic acid concentration of avascular tissues by lactic acid injections leads to vascularization. L-Lactate induces more intensive corneal vascularization than the D-lactate foreign to the organism. The lactic acid concentration of the cornea is low in cases of avascular swelling. Lactic acid promotes the proliferation of other mesenchymal cells as well. A prolonged increase in lactic acid concentration seems to be the most important condition for neovascularization. This is promoted by increased anaerobic or aerobic glycolysis and drainage difficulties of metabolites. Such an affected tissue fails to vascularize only if its compactness acts as a mechanical barrier to vessel growth.

Animals↗

PubMed2004

Production of lactic acid from paper sludge by simultaneous saccharification and fermentation.

Production of lactic acid from paper sludge has been performed by simultaneous saccharification and fermentation (SSF). The SSF process design was based on experimental data obtained from cellulose hydrolysis and fermentation. The SSF process was employed to avoid an excessively dense solution when the sludge content of the feed was higher than 15%; this is one of several benefits of SSF. The enzyme system used for hydrolysis of paper sludge for production of glucose was optimized. CMCase and beta-glucosidase with activities of 2.5 and 10 U mL(-1), respectively, were found to be optimum for hydrolyzing 5% sludge. In batch SSF 16 g L(-1) lactic acid was produced from 5% paper sludge with an yield of 80%. Paper sludge which served as a feed seemed to have a buffering effect during SSF, probably because of the inorganic ash component in the sludge. The final product concentration by SSF was observed to be limited by the cellulose content of the system, which can probably be resolved by intermittent feeding of the paper sludge. SSF of paper sludge fed in batch mode, with intermittent feeding, produced lactic acid at 162 g L(-1), with a yield of 74% and a productivity of 1.4 g L(-1) h(-1). The lactic acid production performance of the modified bioreactor improved after removal of indigestible solid materials from the upper compartment, which enabled the feed of paper sludge to be increased. A mathematical model is described which predicts glucose and subsequent lactic acid production on the basis of the rate expressions of each step of the SSF process. Saccharification kinetics were determined by experiments on enzymatic cellulose hydrolysis, by use of a Michaelis-Menten equation; growth kinetics of L. rhamnosus were determined by use of a Monod expression which incorporated lactic acid inhibition. The kinetic model is expected to predict the performance of the SSF process. For further use of the lactic acid, i.e. polylactic acid, it must be recovered and purified. Results from application of the simulated moving-bed (SMB) process for separation of lactic acid and acetic acid are given, as are several methods of lactic acid purification.

Bioreactors↗

PubMed1991

Biodegradation and antitumour effect of adriamycin-containing poly(L-lactic acid) microspheres.

Adriamycin-containing poly (L-lactic acid) microspheres were prepared to develop a slow-releasing and long-acting adriamycin delivery system. An almost constant release of adriamycin from the adriamycin-containing poly(L-lactic acid) was achieved in Tris buffer and adriamycin disappeared within 20 d. Adriamycin was not detected in serum for up to 14 d, when the suspension of the adriamycin-containing poly(L-lactic acid) microspheres was injected into lung parenchyma, the femoral muscles of rabbits or the peritoneal cavity of mice. However, adriamycin remained in the rabbit muscles for up to 10 d under formation of scar tissue. When free adriamycin was added to P815 tumour cells in culture, the cell survival rate decreased with the exposure time. The treatment with the adriamycin-containing poly(L-lactic acid) microspheres showed a higher survival rate for mice bearing P815 tumour cells than with free adriamycin. In addition, the systemic side effects were insignificant when the adriamycin-containing poly(L-lactic acid) microspheres were given to mice instead of free adriamycin.

Animals↗