Acetic Acid Production by an Electrodialysis Fermentation Method with a Computerized Control System

In acetic acid fermentation by Acetobacter aceti, the acetic acid produced inhibits the production of acetic acid by this microorganism. To alleviate this inhibitory effect, we developed an electrodialysis fermentation method such that acetic acid is continuously removed from the broth. The fermentation unit has a computerized system for the control of the pH and the concentration of ethanol in the fermentation broth. The electrodialysis fermentation system resulted in improved cell growth and higher productivity over an extended period; the productivity exceeded that from non-pH-controlled fermentation. During electrodialysis fermentation in our system, 97.6 g of acetic acid was produced from 86.0 g of ethanol; the amount of acetic acid was about 2.4 times greater than that produced by non-pH-controlled fermentation (40.1 g of acetic acid produced from 33.8 g of ethanol). Maximum productivity of electrodialysis fermentation in our system was 2.13 g/h, a rate which was 1.35 times higher than that of non-pH-controlled fermentation (1.58 g/h).     

电渗析发酵法生产乳酸的研究

在乳酸发酵过程中,所生成的乳酸对进一步发酵有抑制作用。采用电渗析法从发酵液中及时地分离出产物乳酸,使乳酸的生产量提高到86.4g/L,是不控制pH值发酵时的4倍多。结果表明:电渗析法能有效地消除产物乳酸的抑制作用,提高了乳酸生产率,且简化了提取工艺。     

Antagonism Between Osmophilic Lactic Acid Bacteria and Yeasts in Brine Fermentation of Soy Sauce

Brine fermentation by osmophilic lactic acid bacteria and yeasts for long periods of time is essential to produce a good quality of shoyu (Japanese fermented soy sauce). It is well known that lactic acid fermentation by osmophilic lactic acid bacteria results in the depression of alcoholic fermentation by osmophilic yeasts, but the nature of the interaction between osmophilic lactic acid bacteria and yeasts in brine fermentation of shoyu has not been revealed. The inhibitory effect of osmophilic lactic acid bacteria on the growth of osmophilic yeasts was investigated. It was recognized that osmophilic shoyu yeasts such as Saccharomyces rouxii and Torulopsis versatilis were inhibited by a metabolite produced by osmophilic lactic acid bacteria (belonging to Pediococcus halophilus) in brine fermentation of shoyu. The primary inhibitor was considered to be acetic acid, although lactic acid was slightly inhibitory.     

Integrated bioprocess for the simultaneous production of lactic acid and dairy sewage treatment

An integrated biological process was developed for the conversion of whey lactose to lactic acid. We report about the achievement of maximum COD reduction and thus a substantial unburdening of the environment, combined with the economic production of lactic acid, appropriate for industrial scale. The process – designed for continuous operation – consists of four main steps: (i) Protein recovery by ultrafiltration leading to the first product: protein concentrate. The resulting filtrate is the fermentation substrate acid whey permeate. (ii) Adjustment of the composition of the permeate in the medium preparation step in order to ensure the proper function of the following process steps. (iii) Conversion of the lactose to lactate by fermentation with lactic acid bacteria in a cell recycle reactor, using ceramic microfiltration membranes. (iiii) Conversion of the lactate in the cell-free permeate stream of the fermentation to free lactic acid by bipolar electrodialysis. A stable operation of the process was attained up to more than 2000?hours. Using a new selected strain of lactic acid bacteria, a lactic acid productivity of 17?g?l^?1?h^?1 is achieved at total lactose conversion without any nitrogen supplements like yeast extract. A lactic acid concentration of 190?g?l^?1 is obtained in the acidic cell of the electrodialysis unit and the COD of the remaining sewage is diminished by 92%. As an additional cost reduction item, the neutralization agent of the fermentation is recovered in the caustic cell of the bipolar electrodialysis unit. A cost evaluation for an industrial scale process (100?000?t of whey per year) resulted in a price of 0.66 $ per kg of lactic acid, which under present terms hits the goal of making this process economic for the large scale production of lactic acid as an attractive building block for various purposes in chemical industry.     

Homofermentative production of optically pure <Emphasis Type="SmallCaps">l</Emphasis>-lactic acid from sucrose and mixed sugars by batch fermentation of <Emphasis Type="Italic">Enterococcus faecalis</Emphasis> RKY1

In the present study, lactic acid fermentation was carried out by batch culture of Enterococcus faecalis RKY1 using sucrose and mixed sugars as the major substrate. Maximum lactic acid productivity (5.2 g/L/h) was recorded when 50 and 100 g/L of sucrose were used as a carbon source. Sucrose concentration higher than 150 g/L resulted in the decrease of lactic acid productivity due to inhibition by high substrate concentration, but lactic acid productivity was remained > 3.0 g/L/h until the sucrose used for lactic acid fermentation increased up to 150 g/L. L-Lactic acid content of the total lactic acid produced from sucrose and mixed sugars was higher than 98%. When the fermentation media contained sucrose, the kinetic parameters showing specific rates such as μ, qS, and qP were relatively lower than those of fermentation using glucose as a sole carbon source, which might be due to additional time requirement to induce invertase enzyme for utilization of sucrose. There was no carbon catabolite repression observed when the sugar mixtures containing sucrose, glucose, and/ or fructose were used as a carbon source for lactic acid fermentation by E. faecalis RKY1.     

Extractive lactic acid fermentation with tri-n-decylamine as the extractant

In this study, the possibility of tri-n-decylamine (TDA) was investigated as the extractant in extractive lactic acid fermentation with a recombinant yeast being used. Extractive fermentation with TDA did not provide high l-lactic acid production relative to fermentation without extraction. Through determination of trace components of the TDA, it was found that the TDA contained a high concentration of 1-decylaldehyde, which was confirmed to be toxic to yeast and to have an inhibitory effect on cell growth even at low concentration (40 ppm). When 1-decylaldehyde in TDA was reduced from 700 ppm to 33 ppm, the productivity and total concentration of lactic acid increased by 1.8 times and 2.5 times, respectively.     

Application of metabolically engineered Saccharomyces cerevisiae to extractive lactic acid fermentation

In this study, Saccharomyces cerevisiae OC-2T T165R, metabolically engineered to produce optically pure L(+)-lactic acid, was used to develop a high performance extractive fermentation process. Since the transgenic yeast could produce lactic acid efficiently even at lower than pH 3.5, high extractive efficiency was achieved when tri-n-decylamine (TDA), a tertiary amine, was used as the extractant. Separation of microorganisms by means of a hollow fiber module could not only improve the total amount of lactic acid produced but also increase the lactic acid concentration in the solvent. Moreover, pH had a significant effect on extractive fermentation. The highest rate of recovery of lactic acid could be obtained on pH-uncontrolled fermentation (pH 2.5); however, the lowest amount of lactic acid was produced. Taking into account the trade-off between the fermentation and extraction efficiencies, the optimum pH value was considered to be 3.5, with which the largest amount of lactic acid was produced and the highest lactic acid concentration in the solvent was obtained. The results show promise for the use of the transgenic yeast for extractive fermentation.     

Characterization of ethanol fermentation waste and its application to lactic acid production by Lactobacillus paracasei

In this study, an ethanol fermentation waste (EFW) was characterized for use as an alternative to yeast extract for bulk fermentation processes. EFW generated from a commercial plant in which ethanol is produced from cassava/rice/wheat/barley starch mixtures using Saccharomyces cerevisiae was used for lactic acid production by Lactobacillus paracasei. The effects of temperature, pH, and duration on the autolysis of an ethanol fermentation broth (EFB) were also investigated. The distilled EFW (DEFW) contained significant amounts of soluble proteins (2.91 g/l), nitrogen (0.47 g/l), and amino acids (24.1 mg/l). The autolysis of the EFB under optimum conditions released twice as much amino acids than in the DEFW. Batch fermentation in the DEFW increased the final lactic acid concentration, overall lactic acid productivity, and lactic acid yield on glucose by 17, 41, and 14 %, respectively, in comparison with those from comparable fermentation in a lactobacillus growth medium (LGM) that contained 2 g/l yeast extract. Furthermore, the overall lactic acid productivity in the autolyzed then distilled EFW (ADEFW) was 80 and 27 % higher than in the LGM and DEFW, respectively.     

Lactic acid and animal feeds production from Sophora flavescens residues by Rhizopus oryzae fermentation

In this study, the feasibility of producing lactic acid and animal feeds from Sophora flavescens residues (SFR) by Rhizopus oryzae was explored. Results showed that the simultaneous saccharification and fermentation (SSF) is the optimal fermentation mode, which was simple and high-efficiency. When the inoculation volume of R. oryzae was 10 % and the pH value was adjusted by adding CaCO₃ in stages during SSF, the maximum concentration of lactic acid was 46.78 g/L, and the maximum lactic acid productivity reached 0.97 g/L/h. Results also showed that the protein content of the solid residues after fermentation of R. oryzae reached 12.15 %. This content was 46 times higher than that by the original SFRs and nearly 4.3 times the protein content of the solid residues after fermentation by Enterococcus faecium. In addition, the solid residues after fermentation rich in Fe and Zn could be used as animal feeds or feed additives. Thus, it is expected that this study may provide a novel approach for Chinese medicine residues treatment towards full resource recovery.     

In situ separation of lactic acid from fermentation broth using ion exchange resins

Lactic acid fermentation is an end product inhibited reaction. In situ separation of lactic acid from fermentation broth using ion exchange resins was investigated and compared with conventional fermentation system. Amberlite resin (IRA-400, Cl⁻) was used to separate lactic acid from fermentation broth and pH was controlled online with an automatic pH controller. The effect of process variables on lactic acid production by Lactobacillus casei in whey permeate was studied. The maximum productivity was obtained at pH = 6.1, T = 37 °C and impeller speed = 200 rpm. The maximum concentration of lactic acid at optimum condition was found to be 37.4 g/L after 38 h of fermentation using in situ separation system. The productivity of in situ separation system was five times increased in comparison with conventional system.     

Recovery of lactic acid from fermentation broth by the two-stage process of nanofiltration and water-splitting electrodialysis

A two-stage process of nanofiltration and water-splitting electrodialysis was investigated for lactic acid recovery from fermentation broth. In this process, sodium lactate is isolated from fermentation broth in the first stage of nanofiltration by using an NTR-729HF membrane, and then is converted to lactic acid in the second stage by water-splitting electrodialysis. To determine the optimal operating conditions for nanofiltration, the effects of pressure, lactate concentration, pH and known added impurities were studied. Lactate rejection was less than 5%, magnesium rejection approximated 45%, and calcium rejection was at 40%. In subsequent water-splitting electrodialysis, both the sodium lactate conversion to lactic acid and sodium hydroxide recovery, were about 95%, with a power requirement of 0.9∼1.0 kWh per kg of lactate.     

Utilization of by-products derived from bioethanol production process for cost-effective production of lactic acid

The by-products of bioethanol production such as thin stillage (TS) and condensed distillers solubles (CDS) were used as a potential nitrogen source for economical production of lactic acid. The effect of those by-products and their concentrations on lactic acid fermentation were investigated using Lactobacillus paracasei CHB2121. Approximately, 6.7 g/L of yeast extract at a carbon source to nitrogen source ratio of 15 was required to produce 90 g/L of lactic acid in the medium containing 100 g/L of glucose. Batch fermentation of TS medium resulted in 90 g/L of lactic acid after 48 h, and the medium containing 10 % CDS resulted in 95 g/L of lactic acid after 44 h. Therefore, TS and CDS could be considered as potential alternative fermentation medium for the economical production of lactic acid. Furthermore, lactic acid fermentation was performed using only cassava and CDS for commercial production of lactic acid. The volumetric productivity of lactic acid [2.94 g/(L·h)] was 37 % higher than the productivity obtained from the medium with glucose and CDS.     

Utilization of Molasses Sugar for Lactic Acid Production by Lactobacillus delbrueckii subsp. delbrueckii Mutant Uc-3 in Batch Fermentation

Efficient lactic acid production from cane sugar molasses by Lactobacillus delbrueckii mutant Uc-3 in batch fermentation process is demonstrated. Lactic acid fermentation using molasses was not significantly affected by yeast extract concentrations. The final lactic acid concentration increased with increases of molasses sugar concentrations up to 190 g/liter. The maximum lactic acid concentration of 166 g/liter was obtained at a molasses sugar concentration of 190 g/liter with a productivity of 4.15 g/liter/h. Such a high concentration of lactic acid with high productivity from molasses has not been reported previously, and hence mutant Uc-3 could be a potential candidate for economical production of lactic acid from molasses at a commercial scale.     

Metabolic engineering of Clostridium acetobutylicum for enhanced production of butyric acid

Clostridium acetobutylicum has been considered as an attractive platform host for biorefinery due to its metabolic diversity. Considering its capability to overproduce butanol through butyrate, it was thought that butyric acid can also be efficiently produced by this bacterium through metabolic engineering. The pta-ctfB-deficient C. acetobutylicum CEKW, in which genes encoding phosphotransacetylase and CoA-transferase were knocked out, was assessed for its potential as a butyric acid producer in fermentations with four controlled pH values at 5.0, 5.5, 6.0, and 6.4. Butyric acid could be best produced by fermentation of the CEKW at pH 6.0, resulting in the highest titer of 26.6 g/l, which is 6.4 times higher than that obtained with the wild type. However, due to the remaining solventogenic ability of the CEKW, 3.6 g/l solvents were also produced. Thus, the CEKW was further engineered by knocking out the adhE1-encoding aldehyde/alcohol dehydrogenase to prevent solvent production. Batch fermentation of the resulting C. acetobutylicum HCEKW at pH 6.0 showed increased butyric acid production to 30.8 g/l with a ratio of butyric-to-acetic acid (BA/AA) of 6.6 g/g and a productivity of 0.72 g/l/h from 86.9 g/l glucose, while negligible solvent (0.8 g/l ethanol only) was produced. The butyric acid titer, BA/AA ratio, and productivity obtained in this study were the highest values reported for C. acetobutylicum, and the BA/AA ratio and productivity were also comparable to those of native butyric acid producer Clostridium tyrobutyricum. These results suggested that the simultaneous deletion of the pta-ctfB-adhE1 in C. acetobutylicum resulted in metabolic switch from biphasic to acidogenic fermentation, which enhanced butyric acid production.     

Conversion of acid hydrolysate of oil palm empty fruit bunch to L-lactic acid by newly isolated Bacillus coagulans JI12

Cost-effective conversion of lignocellulose hydrolysate to optically pure lactic acid is commercially attractive but very challenging. Bacillus coagulans JI12 was isolated from natural environment and used to produce L-lactic acid (optical purity?>?99.5 %) from lignocellulose sugars and acid hydrolysate of oil palm empty fruit bunch (EFB) at 50 °C and pH 6.0 without sterilization of the medium. In fed-batch fermentation with 85 g/L initial xylose and 55 g/L xylose added after 7.5 h, 137.5 g/L lactic acid was produced with a yield of 98 % and a productivity of 4.4 g/L?h. In batch fermentation of a sugar mixture containing 8.5 % xylose, 1 % glucose, and 1 % L-arabinose, the lactic acid yield and productivity reached 98 % and 4.8 g/L?h, respectively. When EFB hydrolysate was used, 59.2 g/L of lactic acid was produced within 9.5 h at a yield of 97 % and a productivity of 6.2 g/L?h, which are the highest among those ever reported from lignocellulose hydrolysates. These results indicate that B. coagulans JI12 is a promising strain for industrial production of L-lactic acid from lignocellulose hydrolysate.     

Fermentative lactic acid production with a metabolically engineered yeast immobilized in photo-crosslinkable resins

In this study, the immobilization technique involving photo-crosslinkable resin gels was used for lactic acid production. Saccharomyces cerevisiae OC-2T T165R, a metabolically engineered yeast that produces optically pure l(+)-lactic acid, was immobilized in hydrophilic photo-crosslinked resin gels as a biocatalyst. Three resin gels, TEP 1, TEP 2 and TEP 3, were examined and all of them showed high performance as to lactic acid production. Resin gel TEP 1, which exhibited the highest productivity among the resin gels was used for 15 consecutive batch fermentations without decreases in productivity and mechanical deformation, indicating that it was a suitable carrier for long-term lactic acid fermentation. Moreover, the use of the immobilization technique can improve the productivity of the metabolically engineered yeast in the fermentation with or without extraction, showing promise for using the immobilized engineered yeast for lactic acid production.     

Continuous lactic acid fermentation with concentrated product recovery by ultrafiltration and electrodialysis

Lactose of sweet whey permeate was converted into sodium lactate byLactobacillus helveticus. To increase the, productivity of the lactic acid fermentation and to reduce the amounts of effluents, the bioreactor was coupled with an ultrafiltration module and an electrodialysis unit. Without the electrodialyzer, with total cell recycling and at a dilution rate of 0.88 h^–1, a cellular concentration of 64 gl^–1 and a productivity of 22 gl^–1 h^–1 were obtained. When the electrodialysis unit is coupled, the outlet concentration of lactate was stabilized at 85±5 gl^–1.     

Production of lactic acid using a new homofermentative Enterococcus faecalis isolate

Lactic acid is an intermediate-volume specialty chemical for a wide range of food and industrial applications such as pharmaceuticals, cosmetics and chemical syntheses. Although lactic acid production has been well documented, improved production parameters that lead to reduced production costs are always of interest in industrial developments. In this study, we describe the production of lactic acid at high concentration, yield and volumetric productivity utilizing a novel homofermentative, facultative anaerobe Enterococcus faecalis CBRD01. The highest concentration of 182 g lactic acid l⁻¹ was achieved after 38 h of fed-batch fermentation on glucose. The bacterial isolate utilized only 2–13% of carbon for its growth and energy metabolism, while 87–98% of carbon was converted to lactic acid at an overall volumetric productivity of 5 g l⁻¹ h⁻¹. At 13 h of fermentation, the volumetric productivity of lactate production reached 10.3 g l⁻¹ h⁻¹, which is the highest ever reported for microbial production of lactic acid. The lactic acid produced was of high purity as formation of other metabolites was less than 0.1%. The present investigation demonstrates a new opportunity for enhanced production of lactic acid with potential for reduced purification costs.     

Efficient Open Fermentative Production of Polymer-Grade L-Lactate from Sugarcane Bagasse Hydrolysate by Thermotolerant Bacillus sp. Strain P38

Lactic acid is one of the top 30 potential building-block chemicals from biomass, of which the most extensive use is in the polymerization of lactic acid to poly-lactic-acid (PLA). To reduce the cost of PLA, the search for cheap raw materials and low-cost process for lactic acid production is highly desired. In this study, the final titer of produced L-lactic acid reached a concentration of 185 g·L⁻¹ with a volumetric productivity of 1.93 g·L⁻¹·h⁻¹ by using sugarcane bagasse hydrolysate as the sole carbon source simultaneously with cottonseed meal as cheap nitrogen sources under the open fed-batch fermentation process. Furthermore, a lactic acid yield of 0.99 g per g of total reducing sugars was obtained, which is very close to the theoretical value (1.0 g g⁻¹). No D-isomer of lactic acid was detected in the broth, and thereafter resulted in an optical purity of 100%, which exceeds the requirement of lactate polymerization process. To our knowledge, this is the best performance of fermentation on polymer-grade L-lactic acid production totally using lignocellulosic sources. The high levels of optically pure l-lactic acid produced, combined with the ease of handling and low costs associated with the open fermentation strategy, indicated the thermotolerant Bacillus sp. P38 could be an excellent candidate strain with great industrial potential for polymer-grade L-lactic acid production from various cellulosic biomasses.     

Extractive lactic acid fermentation in poly (ethyleneimine)-based aqueous two-phase system

The potential of an aqueous two-phase system composed of a polycation, poly(ethyleneimine) (PEI), and an uncharged polymer, (hydroxyethyl) cellulose (HEC), for extractive lactic acid fermentation was tested. Batch fermentation with 20 g/L glucose in two-phase medium using Lactococcus lactis without external pH control resulted in 3-4 times higher amount of lactate and biomass produced as compared to that in a conventional one-phase medium. Lactic acid was preferentially partitioned to the PEI-rich bottom phase. However, the cells which favored the HEC-rich top phase in a fresh two-phase medium were partitioned to a significant extent to the bottom phase after fermentation. Addition of phosphate buffer or pH adjustment to 6.5 after fermentation caused fewer cells to move to the bottom phase. With external pH control, fermentation in normal and two-phase medium showed no marked differences in glucose consumption and lactic acid yield, except that about 1.3 times higher cell density was obtained in the two-phase broth, especially at initial glucose concentrations of 50-100 g/L. Use of higher concentration of phosphate during batch fermentation in the two-phase medium with 50 g/L sugar provided a 15% higher yield of lactic acid, but the growth rate of cells was nearly half of the normal, thus affecting the productivity. Continuous fermentation with twice the normal phosphate concentration resulted in higher cell density, product yield, and productivity in two-phase medium than in monophasic medium. (c) 1996 John Wiley & Sons, Inc.