Acetic acid production from corn straw via enzymatic degradation using putative acetyl esterase from the metagenome assembled genome.

This chapter provides a review on recent advances in the development of bioprocesses for butyric and acetic acids production, highlighting metabolic engineering in strain development and novel bioreactor and separation technologies in process development leading to their commercial applications in the near future. Acetic acid is produced naturally as the sole or main fermentation product by anaerobic homoacetogens and aerobic acetic acid bacteria. Metabolic engineering has been widely used in strain development for industrial fermentation to produce bio-based chemicals and biofuels. Conventional batch fermentation processes for acetic and butyric acid production are limited by low reactor productivity, low product yield, and low final product concentration, making product recovery difficult and the process uneconomical. Current petroleum-based manufacturing processes for acetic acid and butyric acid production use distillation for their recovery and purification.

We previously reported that the aldehyde dehydrogenase encoded by ALD3 but not ALD6 was responsible, in part, for the increased acetic acid found in Icewines based on the expression profile of these genes during fermentation. We have now completed the expression profile of the remaining yeast aldehyde dehydrogenase genes ALD2, ALD4 and ALD5 during these fermentations to determine their contribution to acetic acid production. The contribution of acetaldehyde stress as a signal to stimulate ALD expression during these fermentations was investigated for all ALD genes. The expression of glycerol-3-phosphate encoded by GPD2 was also followed during these fermentations to determine its role in addition to the role we already identified for GPD1 in the elevated glycerol produced during Icewine fermentation. Icewine juice (38.5 degrees Brix, 398 +/- 5 g l(-1) sugar), diluted Icewine juice (20.8 degrees Brix, 196 +/- 4 g l(-1) sugar) and the diluted juice with sugar levels equal to the original Icewine juice (36.6 degrees Brix, 395 +/- 6 g l(-1) sugar) were fermented in duplicate using the commercial wine yeast K1-V1116. Acetic acid and glycerol production increased 8.4- and 2.7-fold in the Icewine vs the diluted juice fermentation, respectively, accompanied by a fourfold transient increase in acetaldehyde in the Icewine condition during the first week. Both mitochondrial aldehyde dehydrogenases encoded by ALD4 and ALD5 were expressed, with ALD5 expression highest at the start of all fermentations and ALD4 expression increasing during the first week of each condition. ALD2, ALD4, ALD5 and GPD2 showed no differential expression between the three fermentation conditions indicating their lack of involvement in elevating acetic acid and glycerol in Icewine. When yeast fermenting the diluted fermentation was exposed to exogenous acetaldehyde, the transient spike in acetaldehyde increased the expression of ALD3 but this response alone was not sufficient to cause an increase in acetic acid. Expression of the other aldehyde dehydrogenases was unaffected by the acetaldehyde addition. The aldehyde dehydrogenases encoded by ALD2, ALD4 and ALD5 do not contribute to the elevated acetic acid production during Icewine fermentation. Expression of GPD2 was not upregulated in high sugar fermentations and does not reflect the elevated levels of glycerol found in these wines. Acetaldehyde at a concentration produced during Icewine fermentation stimulates the expression of ALD3, but has no impact on the expression of ALD2, -4, -5 and -6. Upregulation of ALD3 alone in the dilute fermentation is not sufficient to increase acetic acid in wine and requires additional responses found in cells under hyperosmotic stress. This work confirms that increased acetic acid and glycerol production during Icewine fermentation follows upregulation of ALD3 and GPD1 respectively, but upregulation of ALD3 alone is not sufficient to increase acetic acid production. Additional responses of cells under osmotic stress are required to increase acetic acid in Icewine.

This study investigated the effect of select non-Saccharomyces yeast strains on Hanseniaspora uvarum growth and acetic acid and ethyl acetate production during prefermentation cold soak. We tested commercially available non-Saccharomyces yeasts for their ability to reduce H. uvarum growth and acetic acid production during a simulated cold soak in a grape juice-based medium. All tested non-Saccharomyces yeast reduced H. uvarum growth and acetic acid production, with some yeast having a greater impact than others. Following the screening of non-Saccharomyces yeast, we tested 14 different H. uvarum isolates against a selected non-Saccharomyces yeast, Metschnikowia fructicola, and found that all H. uvarum isolates had reduced growth and acetic acid production when grown in co-culture with M. fructicola, with variation between isolates noted. Finally, we evaluated the effect of M. fructicola on H. uvarum during prefermentation cold soak of Pinot noir grapes. Pinot noir grapes were inoculated with a combination of H. uvarum and M. fructicola and cold soaked for six days at 8°C. At the end of cold soaking, treatments inoculated with M. fructicola contained lower populations of H. uvarum and significantly lower acetic acid and ethyl acetate concentrations compared with treatments not inoculated with M. fructicola. After the completion of alcoholic fermentation, wines where M. fructicola was added contained significantly lower ethyl acetate but no differences in acetic acid concentration. These results suggest that adding select non-Saccharomyces yeast may be another method to reduce the risk of spoilage by H. uvarum during prefermentation cold soaking.

With the enormous amount of oil palm frond (OPF) left unused in an oil palm plantation, this study proposed to develop the process for ethanol and acetic acid production from OPF juice (pH 4.83) using Saccharomyces cerevisiae TISTR5055 and Acetobacter aceti at room temperature (30±2 °C) for 72 h. Among three different fermentation modes tested, single- and two-stage fermentation produced only ethanol, while simultaneous fermentation exhibited both ethanol and acetic acid production. The yields of 0.39 g ethanol/g glucose and 0.88 g acetic acid/g ethanol were achieved with 76% and 68% of the theoretical yields. Simultaneous fermentation of OPF juice under different shaking speeds and aeration rates revealed no shaking (static) and no aeration were the optimum conditions for ethanol production (19.59 g/L). Shaking at 180 rpm and aeration at 0.5–1.0 vvm exhibited the highest acetic acid production with yields of 0.95–1.16 g acetic acid/g ethanol (73–89 % of the theoretical yields). In addition, the aeration rate had a profound effect on acetic acid production rate with 9.0-fold increase as the aeration rate increased from 0 to 0.5 vvm. The culture filtrate from acetic acid production was analyzed to contain 15 compounds that could be chemically grouped into alcohols, alkenes, aromatic hydrocarbons, and alkanes, with acetic acid as the most abundant (54.6%). Interestingly, the culture filtrate had the efficacy to suppress the growth of Ceratocystis paradoxa TT1, causing black seed rot disease in oil palm with the effective dose (100% inhibition) at 0.20% and 0.50% (v/v) acetic acid in liquid and solid culture, respectively. Therefore, OPF juice without nutrient supplementation could be used as an alternative fermentation medium for ethanol and acetic acid production with high potential exploitation of the acetic acid culture filtrate as an antifungal agent against black seed rot disease.

The influence of the oxygen supply on the growth, acetic acid and ethanol production by Brettanomyces bruxellensis in a glucose medium was investigated with different air flow rates in the range 0-300 l h(-1 ) x (0-0.5 vvm). This study shows that growth of this yeast is stimulated by moderate aeration. The optimal oxygen supply for cellular synthesis was an oxygen transfer rate (OTR) of 43 mg O(2) l(-1) x h(-1). In this case, there was an air flow rate of 60 l h(-1) (0.1 vvm). Above this value, the maximum biomass concentration decreased. Ethanol and acetic acid production was also dependent on the level of aeration: the higher the oxygen supply, the greater the acetic acid production and the lower the ethanol production. At the highest aeration rates, we observed a strong inhibition of the ethanol yield. Over 180 l h(-1) x (0.3 vvm, OTR =105 mg O(2) l(-1) x h(-1)), glucose consumption was inhibited and a high concentration of acetic acid (6.0 g x l(-1)) was produced. The ratio of "ethanol + acetic acid" produced per mole of consumed glucose using carbon balance calculations was analyzed. It was shown that this ratio remained constant in all cases. This makes it possible to establish a stoichiometric equation between oxygen supply and metabolite production.

The objective of this study was to determine the effect of increasing juice soluble solids above 40 degrees Brix on wine yeast's ability to grow and ferment the juice, with particular focus on acetic acid production, titratable acidity (TA) changes and the maximum amount of sugar consumed by the yeast. Riesling Icewine juices at 40, 42, 44 and 46 degrees Brix were inoculated with K1- V116 at 0.5 g 1(-1) and fermented at 17 degrees C until sugar consumption ceased. Increasing soluble solids showed strong negative linear correlations with yeast growth, sugar consumption and ethanol production (r = -0.999, -0.997 and 0.984, P < 0.001, respectively). Acetic acid, glycerol and TA production normalized to sugar consumed showed strong positive correlations to the initial juice concentration (r = 0.992, 0.963, and 0.937, P < 0.001 respectively) but no correlation was found for ethanol production. The acetic acid produced as a function of sugar consumed was positively correlated to the glycerol produced (r = 0.970, P < 0.001). The final TA of the wines ranged between 11.8 and 13.7 g 1(-1) tartaric acid, increasing by 2.3-3 g 1(-1) over the starting juice. The increase in TA was positively correlated to the increase in acetic acid produced after normalizing the data to the amount of sugar consumed (r =0.975, P < 0.001). The acid equivalents resulting from the increase in acetic acid accounted for 80-100% of the TA increase when converted to units of tartaric acid. In the final Icewines, acetic acid represented 19-20% of wine TA. Increasing Icewine juice concentration from 40 to 46 degrees Brix increases the proportion of yeast sugar metabolism towards glycerol and acetic acid production to cope with the increased osmotic stress by decreasing yeast growth, sugar consumption rate, the total amount of sugar consumed and the total amount of ethanol produced. The high proportional contribution of acetic acid to titratable acidity in Riesling Icewine may affect acidity perception. We have determined that 10% v/v ethanol would not be achievable with initial juice concentrations above 42 degrees Brix and that Riesling Icewine juice above 52.5 degrees Brix would be theoretically unfermentable. The high proportional contribution of acetic acid to TA may be an important factor in the organoleptic balance of these Icewines.

Some microorganisms, such as Escherichia coli, harbor transhydrogenases that catalyze the interconversion between NADPH and NADH. However, such transhydrogenase genes have not been found in the genome of a glutamic acid-producing bacterium Corynebacterium glutamicum. In this study, the E. coli transhydrogenase genes udhA and pntAB were introduced into the C. glutamicum wild-type strain ATCC 13032, and the metabolic characteristics of the recombinant strains under aerobic and microaerobic conditions were examined. No major metabolic changes were observed following the introduction of the E. coli transhydrogenase genes under aerobic conditions. Under microaerobic conditions, significant metabolic change was not observed following the introduction of the udhA gene. However, the specific production rates of lactic acid, acetic acid, and succinic acid, and the overall production levels of acetic acid and succinic acid were increased by introducing the E. coli pntAB gene. Moreover, the NADH/NAD(+) ratio was increased by introduction of pntAB. Our results suggest that the E. coli PntAB transhydrogenase enhances the conversion of NADPH to NADH in C. glutamicum under microaerobic conditions, and the increased NADH/NAD(+) ratio results in increased succinic acid production. In addition, acetic acid production might be enhanced to supply ATP to the anaplerotic reaction catalyzed by pyruvate carboxylase.

Swine manure contains a host of chemical and biological constituents which make it desirable for amending lignocellulosic biomass in storage for year round processing in a biorefinery. Application of swine manure in an integrated biomass storage and conversion system was investigated to determine the potential for improved conversion of corn stover to organic acids and soluble carbohydrates during ensiling. Corn stover- swine manure mixtures were prepared containing swine manure at rates of 0%, 15%, 30%, 45%, and 60% while simultaneously being adjusted to 65% moisture on a wet basis and ensiled for 0, 1, 7, and 21 days. Samples were analyzed for pH, dry matter, water-soluble carbohydrates, and organic acids. All treatments, with the exception of the 60% manure substrate, produced a pH less than 4.5, which is sufficient for stable storage. Water-soluble carbohydrates were highest in the control treatment, producing a level of 3.0% DM at day 21. Lactic acid production was unaffected by the rate of manure, with a concentration of 2.8% DM reached at day 21. Acetic acid production was improved with the manure substrates. Manure amendment rates of 30%, 45%, and 60% produced the highest acetic acid concentration of 1.8% DM. Treatments of 0%, 15%, 30%, and 45% swine manure would be acceptable substrates for use in this system; however, if preservation of fermentable carbohydrates is a higher priority than organic acid production, then the pure corn stover substrate would be the most appropriate material to use.

Formation of lactic, acetic, succinic, propionic, formic and butyric acid by lactic acid bacteria

The biomass ensilage conversion (BEC) system is an integrated process for production of organic acids, mainly lactic and acetic acid, and pretreatment and storage of high lignocellulosic materials. Iodoform has been found to selectivity inhibit certain microbes and may help to inhibit undesirable silage microbes. Manure presents a source of nutrients for amending biomass. The objectives of this thesis were to examine the effect of iodoform and manure on fermentation in the BEC system for corn stover. Iodoform treatment rates of 0, 0.03, 0.06, 0.11, and 0.23 g/kg dry matter (DM) were applied to a swine manure-corn stover substrate containing 60 % manure and ensiled for 21 days. A substantial decrease in pH was observed in all treatments, but none of the treatments reached a pH of 4.5, which is sufficient for stable storage. Lactic and acetic acid production was increased with application of iodoform at 0.23 g/kg DM. Iodoform was also found to inhibit butyric fermentation and presumably clostridia activity when applied at a rate of 0.23 g/kg DM. Corn stover-manure mixtures were prepared containing manure at rates of 0, 15, 30, 45, and 60 % while and ensiled for 21 days. All treatments, with the exception of the 60% manure substrate, produced a ph less than 4.5, which is sufficient for stable storage. Water soluble carbohydrates were highest in the control treatment, producing a level of 3.0 % DM at day 21. Lactic acid production was unaffected by the rate of manure, with a concentration of 2.8 % DM reached at day 21. Acetic acid production was improved with the manure substrates, with the 30, 45, and 60 % manure rates producing the highest concentration of 1.8 % DM. These results indicate that iodoform can improve fermentation in this hybrid ensilage system by increasing lactic acid production and inhibiting butyric fermentation. Treatments of 0, 15, 30, and 45% manure would be acceptable substrates for use in this system, however if pretreatment is a higher priority than organic acid production the pure corn stover substrate would be the most appropriate material to use.

To determine acetic acid, acetaldehyde and glycerol production by wine yeast throughout Icewine fermentation. The expression of yeast cytosolic aldehyde dehydrogenases (ALD3 and ALD6) and glycerol-3-phosphate dehydrogenase (GPD1) were followed to relate metabolites in the wines to expression patterns of these genes. Icewine juice (38.8 degrees Brix, 401 +/- 7 g l(-1) sugar), diluted Icewine juice (21.3 degrees Brix, 211 +/- 7 g l(-1) sugar) and the diluted juice with sugar levels equal to the original Icewine juice (35.6 degrees Brix, 402 +/- 6 g l(-1) sugar) were fermented in triplicate using the commercial wine yeast K1-V1116. Acetic acid production increased 7.1-fold and glycerol production increased 1.8-fold in the Icewine fermentation over that found in the diluted juice fermentation. ALD3 showed a 6.2-fold induction and GPD1 showed a 2.5-fold induction during Icewine vs the diluted fermentation. ALD3 was not glucose repressed when additional sugar was added to diluted juice, but was upregulated 7.0-fold. The NAD+-dependant aldehyde dehydrogenase encoded by ALD3 appears to contribute to acetic acid production during Icewine fermentation. Expression of GPD1 was upregulated in high sugar fermentations and reflects the elevated levels of glycerol. Solutes in Icewine juice in addition to sugar contribute to the yeast metabolic response. This work represents the first descriptive analysis of the fermentation of Canadian Icewine, the expression patterns of yeast genes involved in metabolite production, and their impact on Icewine quality. A role for ALD3 in acetic acid production during Icewine fermentation was found.

Half of U.S. acetic acid production is used in manufacturing vinyl acetate monomer (VAM) and is economical only in very large production plants. Nearly 80% of the VAM is produced by methanol carbonylation, which requires high temperatures and exotic construction materials and is energy intensive. Fermentation-derived acetic acid production allows for small-scale production at low temperatures, significantly reducing the energy requirement of the process. The goal of the project is to develop a scaleable production and separation process for fermentation-derived acetic acid. Synthesis gas (syngas) will be fermented to acetic acid, and the fermentation broth will be continuously neutralized with ammonia. The acetic acid product will be recovered from the ammonium acid broth using vapor-based membrane separation technology. The process is summarized in Figure 1. The two technical challenges to success are selecting and developing (1) microbial strains that efficiently ferment syngas to acetic acid in high salt environments and (2) membranes that efficiently separate ammonia from the acetic acid/water mixture and are stable at high enough temperature to facilitate high thermal cracking of the ammonium acetate salt. Fermentation - Microbial strains were procured from a variety of public culture collections (Table 1). Strains were incubated and grown in the presence of the ammonium acetate product and the fastest growing cultures were selected and incubated at higher product concentrations. An example of the performance of a selected culture is shown in Figure 2. Separations - Several membranes were considered. Testing was performed on a new product line produced by Sulzer Chemtech (Germany). These are tubular ceramic membranes with weak acid functionality (see Figure 3). The following results were observed: (1) The membranes were relatively fragile in a laboratory setting; (2) Thermally stable {at} 130 C in hot organic acids; (3) Acetic acid rejection > 99%; and (4) Moderate ammonia flux. The advantages of producing acetic acid by fermentation include its appropriateness for small-scale production, lower cost feedstocks, low energy membrane-based purification, and lower temperature and pressure requirements. Potential energy savings of using fermentation are estimated to be approximately 14 trillion Btu by 2020 from a reduction in natural gas use. Decreased transportation needs with regional plants will eliminate approximately 200 million gallons of diesel consumption, for combined savings of 45 trillion Btu. If the fermentation process captures new acetic acid production, savings could include an additional 5 trillion Btu from production and 7 trillion Btu from transportation energy.

Since the large occurrence of Brettanomyces yeasts in strict anaerobiosis environments (sparkling wines) has been found without an increase in acetic acid content, we evaluated the influence of the oxygen concentration on acetic acid production. Results showed that the oxygen concentration exerted a strong influence on both growth and acetic acid production by Brettanomyces yeasts in winemaking. Full aerobiosis lead to a large production of acetic acid causing a block of metabolic activity. Semi-aerobiosis resulted in the best condition for alcoholic fermentation (Custers effect) combined with acetic acid production. In anaerobic condition Brettanomyces yeasts did not result in high acetic acid production and a pure, even if slow, alcoholic fermentation occurred. The absence of an increase in acetic acid in wines, does not exclude the active presence of Brettanomyces yeast since the characteristic 'high acetic producer' in Brettanomyces yeast is linked to the presence of oxygen. ©1997 SCI

Effect of copper nanoparticles on nitrification in a soil–plant system

The production of vinegar on an industrial scale from different raw materials is subject to constraints, notably the low tolerance of acetic acid bacteria (AAB) to high temperatures and high ethanol concentrations. In this study, we used 25 samples of different fruits from seven Moroccan biotopes with arid and semi-arid environmental conditions as a basic substrate to isolate thermo- and ethanol-tolerant AAB strains. The isolation and morphological, biochemical and metabolic characterization of these bacteria allowed us to isolate a total number of 400 strains with characters similar to AAB, of which six strains (FAGD1, FAGD10, FAGD18 and GCM2, GCM4, GCM15) were found to be mobile and immobile Gram-negative bacteria with ellipsoidal rod-shaped colonies that clustered in pairs and in isolated chains. These strains are capable of producing acetic acid from ethanol, growing on peptone and oxidizing acetate to CO2 and H2O. Strains FAGD1, FAGD10 and FAGD18 show negative growth on YPG medium containing D-glucose > 30%, while strains GCM2, GCM4 and GCM15 show positive growth. These six strains stand out on CARR indicator medium as isolates of the genus Acetobacter ssp. Analysis of 16S rDNA gene sequencing allowed us to differentiate these strains as Acetobacter fabarum and Acetobacter pasteurianus. The study of the tolerance of these six isolates towards pH showed that most of the six strains are unable to grow at pH 3 and pH 9, with an ideal pH of 5. The behavior of the six strains at different concentrations of ethanol shows an optimal production of acetic acid after incubation at concentrations between 6% and 8% (v/v) of ethanol. All six strains tolerated an ethanol concentration of 16% (v/v). The resistance of the strains to acetic acid differs between the species of AAB. The optimum acetic acid production is obtained at a concentration of 1% (v/v) for the strains of FAGD1, FAGD10 and FAGD18, and 3% (v/v) for GCM2, GCM4 and GCM15. These strains are able to tolerate an acetic acid concentration of up to 6% (v/v). The production kinetics of the six strains show the highest levels of growth and acetic acid production at 30 °C. This rate of growth and acetic acid production is high at 35 °C, 37 °C and 40 °C. Above 40 °C, the production of acid is reduced. All six strains continue to produce acetic acid, even at high temperatures up to 48 °C. These strains can be used in the vinegar production industry to minimize the load on cooling systems, especially in countries with high summer temperatures.