Fermentation of swine waste-corn mixtures for animal feed: Pilot-plant studies

Summary Aerobic culture with solid substrates of fresh swine waste combined with corn resulted in lactic acid fermentation with odor control. Heterofermentative lactic acid bacteria produced lactic and homologous tatty acids from acetic through valeric acid (0.1 meq/dry g) to reduce pH 2 units to 4.2 to 4.6. During the fermentation, lactic acid organisms increased from 10⁷ to 10⁹/dry g. Coliform organisms remained steady in number at 10⁶ organisms/dry g. Pilot-plant scale fermentation produced a product with 21 to 39% more methionine than corn but was still limiting for this amino acid as well as lysine for young pigs. Fermentation product from fresh waste-corn cultures was fed as the major dietary component to young pigs, hens, and sheep. Pigs showed gain and gain/feed diminished by one-third in 13-day trials. Laying hens performed comparably to controls in a 21-day test, and sheep did not discriminate against fermentation product.     

Silage fermentation of swine waste combined with corn

Summary Anaerobic fermentation of feedlot swine waste combined with corn was carried out in a newly designed laboratory silo. A lactic acid fermentation with rapid odor control resulted. Initial numbers of total and lactic acid bacteria (all per dry g) were 10⁷. More than 75% of the initial total population were lactobacilli that increased 27-fold at 24 h and immediately entered a phase of decreasing viable numbers. The lactobacilli were largely the streptobacterium type. Lactobacilli were counted in both plates and roll tubes, and the counts in the roll tubes were as much as 100-fold greater; the difference was attributable to anaerobic lactobacilli. After 38 d, lactobacilli were no longer found in plates, but obligately anaerobic lactobacilli persisted at 10³ through 90 d. Fecal coliform bacteria, initially present at 10⁶, were not detected after 24 h. Yeast cells, starting at 10⁶, decreased 100-fold at 3 d, and clostridia nerver exceeded 82 cells. Both groups were of minor importance in this fermentation. Virtually all the acid produced was lactic, measuring 3.83 (% dry matter) at 2 d and rising to a maximum of 12.45 at 69 d. In response, the starting pH of 6.80 decreased to 4.23 and then remained near 4 thereafter. Fumaric and succinic acid levels nerve exceeded 0.2 (% dry matter). Of the volatile fatty acids measured, acetic was maximum at 0.45 (% dry matter), but n-butyric and propionic were never more than 0.06. Fermenting a swine waste-corn mixture in a laboratory silo conferred preserved properties and diminished disease potential on a moist silage that can serve as a major component in an animal ration.The mention of firm names or trade products does not imply that they are endorsed by the U.S. Department of Agriculture over other firms or similar products not mentioned     

Inhibition of yeast by lactic acid bacteria in continuous culture: nutrient depletion and/or acid toxicity?

Lactic acid was added to batch very high gravity (VHG) fermentations and to continuous VHG fermentations equilibrated to steady state with Saccharomyces cerevisiae. A 53% reduction in colony-forming units (CFU) ml^–1 of S. cerevisiae was observed in continuous fermentation at an undissociated lactic acid concentration of 3.44% w/v; and greater than 99.9% reduction was evident at 5.35% w/v lactic acid. The differences in yeast cell number in these fermentations were not due to pH, since batch fermentations over a pH range of 2.5–5.0 did not lead to changes in growth rate. Similar fermentations performed in batch showed that growth inhibition with added lactic acid was nearly identical. This indicates that the apparent high resistance of S. cerevisiae to lactic acid in continuous VHG fermentations is not a function of culture mode. Although the total amount of ethanol decreased from 48.7 g l^–1 to 14.5 g l^–1 when 4.74% w/v undissociated lactic acid was added, the specific ethanol productivity increased ca. 3.2-fold (from 7.42×10^–7 g to 24.0×10^–7 g ethanol CFU^–1 h^–1), which indicated that lactic acid stress improved the ethanol production of each surviving cell. In multistage continuous fermentations, lactic acid was not responsible for the 83% (CFU ml^–1) reduction in viable S. cerevisiae yeasts when Lactobacillus paracasei was introduced to the system at a controlled pH of 6.0. The competition for trace nutrients in those fermentations and not lactic acid produced by L. paracasei likely caused the yeast inhibition.     

Preservation of crab or shrimp waste as silage for cattle

Five experiments were conducted to either ferment fresh shrimp or crab waste with molasses, molasses and bacterial inoculant, or to preserve it with salt. Experiment 1 was a 4 × 2 factorial arrangement. Crab waste was combined with 0, 5, 10, or 15% liquid molasses, and stored in mini-silos (15 l) with or without lids for 14 days. The addition of molasses slightly decreased pH and offensive odors; mini-silo temperatures without lids were higher than those with lids. Experiment 2 was a 5 × 2 factorial arrangement designed to enhance fermentation. Fresh shrimp waste was combined with 0, 10, 15, 20, or 25% dry molasses and 0 or 1.0 × 10⁸ colony forming bacteria/g inoculant and ensiled for six days. As the level of molasses increased, dry matter and lactic acid increased but, the pH, crude protein, ammonia acetic, butyric, and propionic acid concentrations decreased. Significant molasses by inoculant interactions occurred which were highly variable for each acid. Evidence of fermentation was supported by production of lactic acid at all levels of molasses. The pH decreased from 7.7 in the untreated waste to an average of 7.4 for the 10, 15 and 20% molasses treated wastes to 6.8 in the 25% molasses treated waste. The high pH was an indication that the waste may be unstable with longer storage (> 6 days). Therefore, in Experiment 3, designed as a 2 × 2 factorial arrangement, shrimp waste treated with 15 and 20% molasses, with or without inoculant was ensiled for 21 days to test stability. By day 21, shrimp waste had deteriorated as indicated by a mean pH of 7.5, low lactic acid, and high butyric acid concentration, an unacceptable odor, and the presence of mold on the surface of the samples.In Experiments 4 and 5, shrimp or crab waste was combined with salt at 0, 2.5, 5.0, 7.5, 10.0, and 12.5%. Increasing levels of salt decreased crude protein percent, ammonia concentration, and lactic and volatile fatty acids while increasing the pH and improving the acceptability of the odors in both the shrimp and crab wastes. Treatment of crustacean waste with 7.5% or greater salt was more effective at preserving crude protein and minimizing odor than either dry or liquid molasses.     

Inhibitor effect of products of metabolism on growth of Clostridium acetobutylicum

Summary The growth of Clostridium acetobutylicum was studied by three ways. 1. In batch fermentation, referred to as the control. 2. Fermentation in dialysis which permits elimination of all the products of metabolism: acids, solvents and gases. In order to test the toxic effect of acids, cultures were dialysed against 2 g l^-1 acetic acid or 2 g l^-1 butyric acid. 3. To test the toxic effect of gases only, batch fermentations were carried out under vacuum or with a continuous bubbling of nitrogen. The first method resulted in a productivity of 1.2 g l^-1 dry cell weight and a maximal specific growth rate of 0.2 h^-1; the second, 20 g l^-1 dry cell weight and a constant maximum specific growth rate (μ=0.39 h^-1) between 14 and 20 h. The toxic effect of acetic and butyric acids, starts at low concentrations and about 4 g l^-1 of both acids results in a decrease of 50% of maximal specific growth rate. The third series of experiments showed that gases produced by the bacteria have a high toxic effect, comparable to that of 5 g l^-1 of acid.     

Continuous production ofl-lactic acid from whey permeate by immobilizedLactobacillus casei subsp.casei

Summary The production of l-lactic acid from whey permeate, a waste product of the dairy industry, by fermentation with the lactic acid bacterium Lactobacillus casei subsp. casei was investigated. A fermentation medium consisting of permeate and supplements, which enables exponential growth of the organisms, was developed. A fast method for determination of free and immobilized biomass in solid-rich media, based on measurement of cellular ATP, was evolved. Continuous fermentations in a stirred tank reactor (STR) and in a fluidized bed reactor (FBR) with immobilized biomass were compared. In the STR a volumetric productivity of 5.5 g/l per hour at 100% substrate conversion [dilution rate (D) = 0.22 h^–1] was determined. In the FBR porous sintered glass beads were used for immobilization and a maximum biomass concentration of 105 g/kg support was measured. A productivity of 10 g/l per hour was obtained at D = 0.4 h^–1 (substrate conversion 93%) and of 13.5 g/l per hour at D = 1.0 h^–1 (substrate conversion 50%). Offprint requests to: W. Krischke     

Fermentation studies of stored cocoa beans

Acetic and lactic acid bacteria on fermented cocoa beans were maximally 2.0×10⁶ and 1.9×10⁶ c.f.u./g wet wt, respectively. Acetic and lactic acids were detected on the second and fourth days of fermentation and were maximally 140 and 45 mg/10 g beans, respectively. There was a positive correlation between the sizes of the relevant microbial populations and the amounts of acids produced during fermentation.     

Activity and identity of fermenting microorganisms in full-scale biological nutrient removing wastewater treatment plants

Hydrolysis and fermentation are of key importance in biological nutrient removal (BNR) wastewater treatment plants as they provide polyphosphate-accumulating organisms and denitrifying bacteria with carbon and energy sources (e.g. short chain fatty acids). Little information, however, exists about the microbiology of the microorganisms involved in hydrolysis and fermentation. In this study, fermentation of monosaccharides was found to be a universal process taking place in all full-scale BNR plants investigated, where glucose and other monosaccharides were consumed and fermented during anaerobic conditions. The removal rates of glucose were in the range of 0.05–0.32 mmol gVSS⁻¹ h⁻¹ and only slightly lower than glucose removal under aerobic conditions. The main fermentation products detected were (in descending order) propionic acid, lactic acid, acetic acid and formic acid. The fermentation was diverse, consisting of at least three fermentation metabolisms, including lactic acid (homolactic), mixed acid and propionic acid fermentations. Possible existence of alcohol and/or butyric acid fermentations could not be excluded. Fermentation organisms in Aalborg East treatment plant were identified by using microautoradiography combined with fluorescence in situ hybridization. All microorganisms involved in monosaccharide fermentation belonged to either Gram-positive Firmicutes or Actinobacteria . Most of them were related either to Streptococcus , hybridizing to the oligonucleotide probe Str, or to uncultured Actinobacteria with a phenotype of polyphosphate-accumulating organisms. The fermenting bacteria were widespread in the nine full-scale BNR plants investigated and constituted 3–21% of the total bacterial biovolume.     

Survival of <Emphasis Type="Italic">Listeria monocytogenes</Emphasis> in Wilted and Additive-Treated Grass Silage

Grass was field-dried to 3 different dry matter (DM) levels (200, 430 and 540 g/kg) and inoculated with 10⁶–10⁷ cfu/g of a Listeria monocytogenes strain sharing a phagovar occasionally involved in food-borne outbreaks of listeriosis. Formic acid (3 ml/kg) or lactic acid bacteria (8·10⁵/g) with cellulolytic enzymes were applied only to forages with low and intermediate DM levels. Forages were ensiled in laboratory silos (1700 ml) and were stored at 25°C for 30 or 90 days. After 90 days of storage, L. monocytogenes could not be detected in any silo, except one with the high dry matter grass without additive. After 30 days of storage, between 10² and 10⁶ cfu L. monocytogenes/g silage were isolated from the untreated silages. Increasing the DM content from 200 to 540 g/kg did not reduce listeria counts possibly because of the lower production of fermentation acids (higher pH). In silages treated with additives, counts of L. monocytogenes were always lower than in silages without additive. In wet silages (DM 200 g/kg) both additives were effective, but in the wilted silages (DM 430 g/kg) only the bacterial additive reduced listeria counts below detection level. Listeria counts were highly correlated to silage pH (r = 0.92), the concentration of lactic acid (r = -0.80) and the pooled amount of undissociated acids (r = -0.83).     

The cocoa bean fermentation process: from ecosystem analysis to starter culture development

Cocoa bean fermentation is still a spontaneous curing process to facilitate drying of nongerminating cocoa beans by pulp removal as well as to stimulate colour and flavour development of fermented dry cocoa beans. As it is carried out on farm, cocoa bean fermentation is subjected to various agricultural and operational practices and hence fermented dry cocoa beans of variable quality are obtained. Spontaneous cocoa bean fermentations carried out with care for approximate four days are characterized by a succession of particular microbial activities of three groups of micro‐organisms, namely yeasts, lactic acid bacteria (LAB) and acetic acid bacteria (AAB), which results in well‐fermented fully brown cocoa beans. This has been shown through a plethora of studies, often using a multiphasic experimental approach. Selected strains of several of the prevailing microbial species have been tested in appropriate cocoa pulp simulation media to unravel their functional roles and interactions as well as in small plastic vessels containing fresh cocoa pulp‐bean mass to evaluate their capacity to dominate the cocoa bean fermentation process. Various starter cultures have been proposed for successful fermentation, encompassing both cocoa‐derived and cocoa nonspecific strains of (hybrid) yeasts, LAB and AAB, some of which have been implemented on farms successfully.     

Changes in microbial population during fermentation of feedlot waste with corn

A new process for recycling feedlot waste involves the fermentation of liquid from this waste combined with corn. Changes in the flora of this silage-like fermentation were followed. The fermentation was dominated by lactobacilli and yeasts, which initially constitute 1% or less of the natural flora. The species of yeasts and lactics involved were characterized. The fermentation has two phases. A single heterolactic species multiplied rapidly for the first 24 h until it represented 95% of the lactobacilli and more than 90% of the total microflora. It displaced the betabacterium predominant among lactics of the original waste; the acid produced killed coliforms and other organisms in feedlot waste; and the acetic acid produced probably caused the death of the dominant native yeast Trichosporon cutaneum (de Beurm., Gougerot et Vaucher) Ota. The peak lactobacillus count remained constant (about 2 × 10⁹ organisms/g [wet weight]) throughout the rest of the fermentation. Homolactics dominated the later phase and yeasts increased to 9.5 × 10⁷ organisms/g (wet weight). At 6 days, a stable mixture of three lactobacilli was present, one streptobacterium, one thermobacterium, and one betabacterium. Similarly, yeasts stabilized as a mixture of two Candida sp. and one Pichia sp. The dominant species of lactics were characterized. Information on the sequence of microorganisms provides a basis for enhanced protein synthesis in the fermentation.     

Enhancement and modeling of microparticle-added <Emphasis Type="Italic">Rhizopus oryzae</Emphasis> lactic acid production

Lactic acid has a wide industrial application area and can be produced by fungal strains. However, excessive bulk growth form of fungi during the fermentations is a major problem, which limits the fermentation performance. Microparticles are excellent tools to prevent bulk fungal growth and provide homogenized fermentation broth to increase uniformity and the prediction performance of the models. Therefore, in this study, addition of aluminum oxide and talcum microparticles into fermentations was evaluated to enhance the production of lactic acid by Rhizopus oryzae. The results showed that the bulk fungal growth was prevented and the lactic acid concentration increased from 6.02 to 13.88 and 24.01 g/L, when 15 g/L of aluminum oxide or 10 g/L of talcum was used, respectively, in the shake-flask fermentations. Additionally, substrate concentration, pH, and agitation were optimized in the bioreactors using response surface methodology, and optimum values were determined as 126 g/L of glucose, 6.22 pH, and 387 rpm, respectively. Under these conditions, lactic acid production further increased to 75.1 ± 1.5 g/L with 10 g/L of talcum addition. Also, lactic acid production and glucose consumption in the batch fermentation were successfully modeled with modified Gompertz model and modified logistic model. RMSE and MAE values for lactic acid production were calculated as 2.279 and 1.498 for the modified Gompertz model; 3.6 and 4.056 for the modified logistic model. Additionally, modified logistic model predicted glucose consumption with ?2.088 MAE and 2.868 RMSE, whereas these values were calculated as 2.035 and 3.946 for the modified Gompertz model.     

Efficient conversion of lactic acid to butanol with pH-stat continuous lactic acid and glucose feeding method by Clostridium saccharoperbutylacetonicum

In order to achieve high butanol production by Clostridium saccharoperbutylacetonicum N1-4, the effect of lactic acid on acetone–butanol–ethanol fermentation and several fed-batch cultures in which lactic acid is fed have been investigated. When a medium containing 20 g/l glucose was supplemented with 5 g/l of closely racemic lactic acid, both the concentration and yield of butanol increased; however, supplementation with more than 10 g/l lactic acid did not increase the butanol concentration. It was found that when fed a mixture of lactic acid and glucose, the final concentration of butanol produced by a fed-batch culture was greater than that produced by a batch culture. In addition, a pH-controlled fed-batch culture resulted in not only acceleration of lactic acid consumption but also a further increase in butanol production. Finally, we obtained 15.5 g/l butanol at a production rate of 1.76 g/l/h using a fed-batch culture with a pH-stat continuous lactic acid and glucose feeding method. To confirm whether lactic acid was converted to butanol by the N1-4 strain, we performed gas chromatography–mass spectroscopy (GC-MS) analysis of butanol produced by a batch culture during fermentation in a medium containing [1,2,3-¹³C₃] lactic acid as the initial substrate. The results of the GC-MS analysis confirmed the bioconversion of lactic acid to butanol.     

Production of succinic acid and lactic acid by Corynebacterium crenatum under anaerobic conditions

A new succinic acid and lactic acid production bioprocess by Corynebacterium crenatum was investigated in mineral medium under anaerobic conditions. Corynebacterium crenatum cells with sustained acid production ability and high acid volumetric productivity harvested from the glutamic acid fermentation broth were used to produce succinic acid and lactic acid. Compared with the first cycle, succinic acid production in the third cycle increased 120% and reached 43.4 g/L in 10 h during cell-recycling repeated fermentations. The volumetric productivities of succinic acid and lactic acid could maintain above 4.2 g/(L·h) and 3.1 g/(L·h), respectively, for at least 100 h. Moreover, wheat bran hydrolysates could be used for succinic acid and lactic acid production by the recycled C. crenatum cells. The final succinic acid concentration reached 43.6 g/L with a volumetric productivity of 4.36 g/(L·h); at the same time, 32 g/L lactic acid was produced.     

Effects of Cultivation Parameters on the Morphology of Rhizopus arrhizus and the Lactic Acid Production in a Bubble Column Reactor

The use of filamentous Rhizopus for lactic acid production is facing a challenge due to its low yield mainly caused by the difficulty to control its morphology in submerged fermentation processes. This study was aimed at investigating the impacts of cultivation parameters on the morphology of Rhizopus arrhizus DAR 36017 and lactic acid production using waste potato starch in a laboratory scale bubble column reactor (BCR). The fungal morphology was significantly influenced by carbon sources, process pH, starch concentrations, sparger designs and aeration rates. The favorable morphology for lactic acid production was a freely dispersed small pellet, which was achieved under operation conditions at pH 5.0–6.0, starch concentrations of 60–120 g/L and aeration rates of 0.2–0.8 vvm using a sintered stainless steel disc sparger. Optimal cultivation conditions at pH 6.0 and an aeration rate of 0.4 vvm resulted in the formation of freely dispersed small pellets and 103.8 g/L lactic acid with a yield of 87 % from 120 g/L liquefied potato starch in 48 h. The overall results in terms of lactic acid yield and productivity are comparable to those reported in previous studies using immobilized Rhizopus cells in batch fermentations.     

Oxidation of Metabolites Highlights the Microbial Interactions and Role of Acetobacter pasteurianus during Cocoa Bean Fermentation

Four cocoa-specific acetic acid bacterium (AAB) strains, namely, Acetobacter pasteurianus 386B, Acetobacter ghanensis LMG 23848^T, Acetobacter fabarum LMG 24244^T, and Acetobacter senegalensis 108B, were analyzed kinetically and metabolically during monoculture laboratory fermentations. A cocoa pulp simulation medium (CPSM) for AAB, containing ethanol, lactic acid, and mannitol, was used. All AAB strains differed in their ethanol and lactic acid oxidation kinetics, whereby only A. pasteurianus 386B performed a fast oxidation of ethanol and lactic acid into acetic acid and acetoin, respectively. Only A. pasteurianus 386B and A. ghanensis LMG 23848^T oxidized mannitol into fructose. Coculture fermentations with A. pasteurianus 386B or A. ghanensis LMG 23848^T and Lactobacillus fermentum 222 in CPSM for lactic acid bacteria (LAB) containing glucose, fructose, and citric acid revealed oxidation of lactic acid produced by the LAB strain into acetic acid and acetoin that was faster in the case of A. pasteurianus 386B. A triculture fermentation with Saccharomyces cerevisiae H5S5K23, L. fermentum 222, and A. pasteurianus 386B, using CPSM for LAB, showed oxidation of ethanol and lactic acid produced by the yeast and LAB strain, respectively, into acetic acid and acetoin. Hence, acetic acid and acetoin are the major end metabolites of cocoa bean fermentation. All data highlight that A. pasteurianus 386B displayed beneficial functional roles to be used as a starter culture, namely, a fast oxidation of ethanol and lactic acid, and that these metabolites play a key role as substrates for A. pasteurianus in its indispensable cross-feeding interactions with yeast and LAB during cocoa bean fermentation.     

Production of bioethanol from organic whey using <Emphasis Type="Italic">Kluyveromyces marxianus</Emphasis>

Ethanol production by K. marxianus in whey from organic cheese production was examined in batch and continuous mode. The results showed that no pasteurization or freezing of the whey was necessary and that K. marxianus was able to compete with the lactic acid bacteria added during cheese production. The results also showed that, even though some lactic acid fermentation had taken place prior to ethanol fermentation, K. marxianus was able to take over and produce ethanol from the remaining lactose, since a significant amount of lactic acid was not produced (1–2 g/l). Batch fermentations showed high ethanol yield (~0.50 g ethanol/g lactose) at both 30°C and 40°C using low pH (4.5) or no pH control. Continuous fermentation of nonsterilized whey was performed using Ca-alginate-immobilized K. marxianus. High ethanol productivity (2.5–4.5 g/l/h) was achieved at dilution rate of 0.2/h, and it was concluded that K. marxianus is very suitable for industrial ethanol production from whey.     

Development of alcoholic and malolactic fermentations in highly acidic and phenolic apple musts

This work reports the influence of the high acidity and high phenolic content in apple musts on the development of alcoholic and malolactic fermentations and on the final chemical and microbiological composition of the ciders. Four different musts were obtained by pressing several varieties and proportions of cider apples from the Basque Country (Northern Spain). Specially acidic and phenolic varieties were selected. Three musts were obtained in experimental stations and the fourth one, in a cider factory following usual procedures. The evolution of these musts was monitored during five months by measuring 18 parameters throughout eight samplings. In the most acidic of the three experimental musts, yeasts were added to complete the alcoholic fermentation. In the rest of the musts, alcoholic and malolactic fermentations took place spontaneously due to natural microflora and no chemical was added to control these processes. Malolactic fermentation (MLF) finished before alcoholic fermentation in the three tanks obtained in experimental stations, even in the most acidic and phenolic one (pH 3.18, 1.78 g tannic acid/l). After four months, these ciders maintained low levels of lactic acid bacteria (10(4)CFU/ml) and low content of acetic acid (<0.60 g/l). Both fermentations began simultaneously in the must obtained in the cider factory, but MLF finished 10 days after alcoholic fermentation. Subsequently, this must maintained a high population of lactic acid bacteria (>10(6)CFU/ml), causing a higher production of acetic acid (>1.00 g/l) than in the other ciders. These results show the possible advantages of MLF finishing before alcoholic fermentation.     

Microbial and Physiological Characterization of Weakly Amylolytic but Fast-Growing Lactic Acid Bacteria: a Functional Role in Supporting Microbial Diversity in Pozol, a Mexican Fermented Maize Beverage

Pozol is an acid beverage obtained from the natural fermentation of nixtamal (heat- and alkali-treated maize) dough. The concentration of mono- and disaccharides from maize is reduced during nixtamalization, so that starch is the main carbohydrate available for lactic acid fermentation. In order to provide some basis to understand the role of amylolytic lactic acid bacteria (ALAB) in this fermented food, their diversity and physiological characteristics were determined. Forty amylolytic strains were characterized by phenotypic and molecular taxonomic methods. Four different biotypes were distinguished via ribotyping; Streptococcus bovis strains were found to be predominant. Streptococcus macedonicus, Lactococcus lactis, and Enterococcus sulfureus strains were also identified. S. bovis strain 25124 showed extremely low amylase yield relative to biomass (139 U g [cell dry weight]⁻¹) and specific rate of amylase production (130.7 U g [cell dry weight]⁻¹ h⁻¹). In contrast, it showed a high specific growth rate (0.94 h⁻¹) and an efficient energy conversion yield to bacterial cell biomass (0.31 g of biomass g of substrate⁻¹). These would confer on the strain a competitive advantage and are the possible reasons for its dominance. Transient accumulation of maltooligosaccharides during fermentation could presumably serve as energy sources for nonamylolytic species in pozol fermentation. This would explain the observed diversity and the dominance of nonamylolytic lactic acid bacteria at the end of fermentation. These results are the first step to understanding the importance of ALAB during pozol fermentation.     

In situ production of exopolysaccharides during Sourdough fermentation by cereal and intestinal isolates of lactic acid bacteria

EPS formed by lactobacilli in situ during sourdough fermentation may replace hydrocolloids currently used as texturizing, antistaling, or prebiotic additives in bread production. In this study, a screening of >100 strains of cereal-associated and intestinal lactic acid bacteria was performed for the production of exopolysaccharides (EPS) from sucrose. Fifteen strains produced fructan, and four strains produced glucan. It was remarkable that formation of glucan and fructan was most frequently found in intestinal isolates and strains of the species Lactobacillus reuteri, Lactobacillus pontis, and Lactobacillus frumenti from type II sourdoughs. By the use of PCR primers derived from conserved amino acid sequences of bacterial levansucrase genes, it was shown that 6 of the 15 fructan-producing lactobacilli and none of 20 glucan producers or EPS-negative strains carried a levansucrase gene. In sourdough fermentations, it was determined whether those strains producing EPS in MRS medium modified as described by Stolz et al. (37) and containing 100 g of sucrose liter(-1) as the sole source of carbon also produce the same EPS from sucrose during sourdough fermentation in the presence of 12% sucrose. For all six EPS-producing strains evaluated in sourdough fermentations, in situ production of EPS at levels ranging from 0.5 to 2 g/kg of flour was demonstrated. Production of EPS from sucrose is a metabolic activity that is widespread among sourdough lactic acid bacteria. Thus, the use of these organisms in bread production may allow the replacement of additives.