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Using wheat B starch as raw material, optimize the one-step enzymatic hydrolysis process for preparing fermentable sugars and subsequent fermentation of alcohol by brewing yeast to improve its utilization value. Initially, the elemental composition of Wheat B starch was quantified. A composite enzymatic catalyst containing α-amylase and saccharase were then employed to generate fermentable sugars from wheat B starch through enzymatic hydrolysis efficiently. Various parameters, including composite enzyme concentration, solid-liquid ratio, enzyme temperature, enzymatic duration, and pH were thoroughly investigated to assess their effects on reducing sugar yield. An orthogonal optimization approach was conducted to determine the optimal conditions for enzymatic hydrolysis. Results showed that wheat B starch contained 85.61% starch, with approximately 2% protein and mineral content, and pentosan composition nearing 5%. Following the orthogonal optimization, the most effective parameters for enzymatic saccharification were identified: A composite enzyme dosage of 4 mL, a solid-liquid ratio of 1:25 g/mL, an enzymatic digestion temperature of 60 ℃, an enzymatic digestion duration of 10 h, and a pH of 5.0. Under these specific conditions, the yield of reducing sugar reached 94.6%. Subsequently, wheat B starch was converted into alcohol through yeast-mediated fermentation. This process led to a final alcohol concentration 12.76 g/L, accompanied by an alcohol yield of 31.90%. In conclusion, wheat B starch holded remarkable promise for large-scale bioethanol production, providing valuable insights for maximizing the utilization of this resource on a broad scale.

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Waterlogging stress is one of the major abiotic stresses affecting the productivity and quality of many crops worldwide. However, the mechanisms of waterlogging tolerance are still elusive in barley. In this study, we identify key differentially expressed genes (DEGs) and differential metabolites (DM) that mediate distinct waterlogging tolerance strategies in leaf and root of two barley varieties with contrasting waterlogging tolerance under different waterlogging treatments. Transcriptome profiling revealed that the response of roots was more distinct than that of leaves in both varieties, in which the number of downregulated genes in roots was 7.41-fold higher than that in leaves of waterlogging sensitive variety after 72 h of waterlogging stress. We also found the number of waterlogging stress-induced upregulated DEGs in the waterlogging tolerant variety was higher than that of the waterlogging sensitive variety in both leaves and roots in 1 h and 72 h treatment. This suggested the waterlogging tolerant variety may respond more quickly to waterlogging stress. Meanwhile, phenylpropanoid biosynthesis pathway was identified to play critical roles in waterlogging tolerant variety by improving cell wall biogenesis and peroxidase activity through DEGs such as Peroxidase (PERs) and Cinnamoyl-CoA reductases (CCRs) to improve resistance to waterlogging. Based on metabolomic and transcriptomic analysis, we found the waterlogging tolerant variety can better alleviate the energy deficiency via higher sugar content, reduced lactate accumulation, and improved ethanol fermentation activity compared to the waterlogging sensitive variety. In summary, our results provide waterlogging tolerance strategies in barley to guide the development of elite genetic resources towards waterlogging-tolerant crop varieties. [ABSTRACT FROM AUTHOR]

Despite the significant progress in the research, the problem of finding an efficient method for producing bioethanol from renewable lignocellulosic waste materials remains unresolved. Our investigation aimed to assess the efficacy of ethanol production from sugar beet pulp (SBP) utilising various approaches, including pretreatment variations, enzymatic processes, and microbial hydrolysis. Our research involved using the post-cultivation concentrate of T. viride LOCK 0588 grown in the SBP-based medium as a source of enzymes. The SBP hydrolysis process was carried out for 48 h at 50 °C. The quantity of sugar released, up to 61 g dm−3, through the utilisation of this extract proved to be on par with the outcomes achieved by the application of the commercial Cellic Ctec2 preparation. The final yields of the ethanol production with the use of the coculture of S. cerevisiae (Ethanol Red) and Scheffersomyces stipitis LOCK 0047 strain were in the range 5.1 ± 0.11 kg 100 kg−1 ÷ 5.38 ± 0.11 kg 100 kg−1. These results provide a solid basis for improving larger-scale industrial procedures that involve converting SBP into bioethanol using a cost-efficient approach of microbial hydrolysis with T. viride and a blend of pentose and hexose fermenting yeast. [ABSTRACT FROM AUTHOR]

The use of renewable feedstocks and the variability of lignocellulosic feedstocks generating fermentable sugars also offer the advantages of the most abundant, sustainable and competitive non-food biomass. The aim of this study is to valorize the oilcake obtained from non-conventional Congolese Afzelia bella oilseeds, after extraction of the oil used in biodiesel production. The conversion of A. bella oilseeds cake into ethanol was carried out using the two methods of pretreatment (with the synthesized ionic liquid 1-butyl-3-methyl imidazolium chloride and dilute sulfuric acid), followed by hydrolysis with dilute sulfuric acid and fermentation by Saccharomyces cerevisiae. The lignocellulosic composition of the sample was 47.98% cellulose, 28.15% hemicellulose and 22.3% lignin. Pretreatment with 1-butyl-3-methyl-imidazolium chloride (ionic liquid) showed an increase in cellulose content of around 4.25% and hemicellulose content of around 7.7%, with simultaneous delignification of 12.25%. In contrast, with dilute sulfuric acid, the lignin and cellulose content of the sample fell to 3.88% and 1.88% respectively. Hemicellulose content, on the other hand, increased by only 3.58%. After dilute acid hydrolysis (5% H2SO4), the values for total reducing sugar, glucose and xylose were 66.06±1.34 mg.g-1; 36.25 ± 1.2 mg.g-1 and 29.71 ± 0.6 mg.g-1 respectively. The percentage conversion of sugars was 55.16 ± 0.5% for cellulose to glucose and 53.76 ± 0.75% for hemicellulose to xylose. The hydrolyzed product of the complex polysaccharide was then converted to ethanol using commercial yeast. Results showed an ethanol yield of around 10.3%, and Fourier Transform Infrared (FTIR) spectroscopy results confirmed bioethanol production. A. bella oilseeds cakes are therefore a potential source for ethanol production.

Background Cayratia trifolia has been extensively studied for its bioactive components and medicinal properties. This study was carried out to evaluate the fermentation ability of Saccharomyces cerevisiae 2.1 yeast to determine suitable fermentation conditions. Methods The initial sugar content for three-leaf cayratia fermentation and fermentation efficiency were calculated. The temperature for three-leaf cayratia fermentation, incubation time, and preliminary 1 liter of three-leaf cayratia wine fermentation were determined. All treatments showed that the total sugar content decreased after 9 days of fermentation compared to the initial level. Temperature during fermentation has a direct effect on yeast activity. Temperature increases the growth of yeast and speed of enzyme activity. The fermentation time changes depending on temperature, initial soluble solid contents, and yeast strains, leading to changes in the ethanol content after the end of fermentation. Results The results showed that the S. cerevisiae 2.1 was able to ferment at room temperature with an initial pH of 4.5, 24 °Brix, and a yeast density of 107 cells/mL. The appropriate fermentation conditions determined for S. cerevisiae 2.1 were 24 °Brix and incubation at room temperature (28-33°C) with a fermentation time of 11 days. Concusion In 1-L scale fermentation, three-leaf cayratia wine had 9.46% (v/v) and the fermentation efficiency was 90.34%. The wine produced had a unique color, flavor, and aroma that met the sensory evaluation of the Vietnamese national standard TCVN-3217:79.

Mycotoxins (ochratoxin A (20 ppb), aflatoxin B1 (40 ppb), deoxynivalenol (4 ppm), and zearalenone (800 ppb)) were intentionally added to rice bran raw materials. After fermentation, their contents were determined in the distillate and distillery stillage obtained using single-stage and continuous pilot plant-scale columns. After single-stage distillation, aflatoxin B1, deoxynivalenol, and zearalenone were not detected in the distillate, indicating that even if a certain amount (four times the maximum residue limit (MRL)) was present in the raw material, it would not remain in the distillate after fermentation and distillation. Most mycotoxins remained in the distillery stillage, and their residual rates ranged from 54.0–96.2%. For ochratoxin A, 0.19 ppb was found in the distillate and this migration occurred in three consecutive distillations (0.11–0.22 ppb). Ochratoxin A and aflatoxin B1 were not detected in the distillate (alcohol content 93.9% and 95.4%, respectively) obtained from the contaminated fermented liquid (approximately three times the MRL based on the raw material) using the pilot-plant scale continuous distillation column. Therefore, the migration of mycotoxins is difficult when the distilled spirit is produced using a continuous distillation column, even if the raw material is contaminated with certain amounts of the investigated mycotoxins. [ABSTRACT FROM AUTHOR]

Background: Cayratia trifolia has been extensively studied for its bioactive components and medicinal properties. This study was carried out to evaluate the fermentation ability of Saccharomyces cerevisiae 2.1 yeast to determine suitable fermentation conditions. Methods: The initial sugar content for three-leaf cayratia fermentation and fermentation efficiency were calculated. The temperature for three-leaf cayratia fermentation, incubation time, and preliminary 1 liter of three-leaf cayratia wine fermentation were determined. All treatments showed that the total sugar content decreased after 9 days of fermentation compared to the initial level. Temperature during fermentation has a direct effect on yeast activity. Temperature increases the growth of yeast and speed of enzyme activity. The fermentation time changes depending on temperature, initial soluble solid contents, and yeast strains, leading to changes in the ethanol content after the end of fermentation. Results:The results showed that the S. cerevisiae 2.1 strain was able to ferment at room temperature with an initial pH of 4.5, 24 °Brix, and a yeast density of 107 cells/mL. The appropriate fermentation conditions determined for S. cerevisiae 2.1 yeast were 24 °Brix and incubation at room temperature (28-33°C) with a fermentation time of 11 days. Concusion: In 1 liter scale fermentation, three-leaf cayratia wine had 9.46% (v/v) and the fermentation efficiency was 90.34%. The wine produced had a unique color, flavor, and aroma that met the sensory evaluation of the Vietnamese standard TCVN-3217:79.

In this study, the primary paper-mill sludge characterized as containing 51% glucan was used to optimize the enzymatic saccharification process for the production of bioethanol using a Box–Behnken design (BBD). Polyethylene glycol 4000 (PEG-4000) surfactant-assisted enzymatic saccharification of dried primary sludge (DPS) showed a 12.8% improvement in saccharification efficiency. There was a statistically significant effect of solid enzyme loading and saccharification time on the enzymatic saccharification of DPS at a 95% confidence level (p < 0.05). The optimum levels of 10.4% w/w DPS solid loading, 2.03% enzyme loading (10 FPU g/DPS), and 1% (w/w DPS) PEG-4000 loading for a saccharification efficiency of 57.66% were validated experimentally and found to be non-significant with regard to the lack of fit with the predicted saccharification efficiency of 56.76%. Furthermore, Saccharomyces cerevisiae fermented the saccharified sugars into ethanol (9.35 g/L) with a sugar-to-ethanol conversion yield of 91.6% compared with the theoretical maximum. Therefore, DPS is a more suitable renewable biomass for determining the presence of fermentable sugar and for the production of ethanol. [ABSTRACT FROM AUTHOR]

Despite the significant progress in the research, the problem of finding an efficient method for producing bioethanol from renewable lignocellulosic waste materials remains unresolved. Our investigation aimed to assess the efficacy of ethanol production from sugar beet pulp (SBP) utilising various approaches, including pretreatment variations, enzymatic processes, and microbial hydrolysis. Our research involved using the post-cultivation concentrate of T. viride LOCK 0588 grown in the SBP-based medium as a source of enzymes. The SBP hydrolysis process was carried out for 48 h at 50 °C. The quantity of sugar released, up to 61 g dm−3, through the utilisation of this extract proved to be on par with the outcomes achieved by the application of the commercial Cellic Ctec2 preparation. The final yields of the ethanol production with the use of the coculture of S. cerevisiae (Ethanol Red) and Scheffersomyces stipitis LOCK 0047 strain were in the range 5.1 ± 0.11 kg 100 kg−1 ÷ 5.38 ± 0.11 kg 100 kg−1. These results provide a solid basis for improving larger-scale industrial procedures that involve converting SBP into bioethanol using a cost-efficient approach of microbial hydrolysis with T. viride and a blend of pentose and hexose fermenting yeast.

Ligusticum chuanxiong is a common traditional edible-medicinal herb that has various pharmacological activities. However, its effects on Saccharomyces cerevisiae (S. cerevisiae) remains unknown. In this study, we found that water extract of Ligusticum chuanxiong (abbreviated as WEL) exhibited excellent free radical scavenging ability in-vitro. Moreover, WEL treatment could delay the aging of S. cerevisiae, an important food microorganism sensitive to reactive oxygen species (ROS) stress. Biochemical analyses revealed that WEL significantly increased the activity of antioxidant enzymes in S. cerevisiae, including superoxide dismutase (SOD), catalase (CAT) and glutathione reductase (GR), as well as their gene expression. As a result, ROS level was significantly decreased and accompanied with the decline of malondialdehyde (MDA), which represented a state of low oxidative stress. The reduction of oxidative stress could elevate S. cerevisiae's ethanol fermentation efficiency. Taken together, WEL plays a protective role against S. cerevisiae aging via improving antioxidant activity.

Saccharomyces cerevisiae is the most important industrial microorganism used to fuel ethanol production worldwide. Herein, we obtained a mutant S. cerevisiae strain with improved capacity for ethanol fermentation, from 13.72% (v/v for the wild-type strain) to 16.13% (v/v for the mutant strain), and analyzed its genomic structure and gene expression changes. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment revealed that the changed genes were mainly enriched in the pathways of carbohydrate metabolism, amino acid metabolism, metabolism of cofactors and vitamins, and lipid metabolism. The gene expression trends of the two strains were recorded during fermentation to create a timeline. Venn diagram analysis revealed exclusive genes in the mutant strain. KEGG enrichment of these genes showed upregulation of genes involved in sugar metabolism, mitogen-activated protein kinase pathway, fatty acid and amino acid degradation, and downregulation of genes involved in oxidative phosphorylation, ribosome, fatty acid and amino acid biogenesis. Protein interaction analysis of these genes showed that glucose-6-phosphate isomerase 1, signal peptidase complex subunit 3, 6-phosphofructokinase 2, and trifunctional aldehyde reductase were the major hub genes in the network, linking pathways together. These findings provide new insights into the adaptive metabolism of S. cerevisiae for ethanol production and a framework for the construction of engineered strains of S. cerevisiae with excellent ethanol fermentation capacity. [ABSTRACT FROM AUTHOR]

The Sugar industry faces several sustainability challenges due to dumping of excess global production which cause stiff competition for high cost sugar producers like Kenya. Through diversification into cogeneration and ethanol production, sugar companies in high cost producing countries can remain competitive and survive in the market. In this study, the technical and economic feasibility of ethanol production is done for the case of Nzoia Sugar Company in Kenya whose milling capacity is 3000 tons of cane per day (TCD). Molasses is taken through a series of stages starting with pretreatment, followed by fermentation and distillation to produce ethanol. The ethanol formed is then transferred to the distillation columns for fractionation to remove water and hence have ethanol rich product. Through further distillation and molecular sieve dehydration the purity and quality of the final ethanol product is enhanced. The main equipment for the process includes the fermenter, boilers, condensers, and distillation columns, design to suit the product characteristics and design capacity of the plant. The analysis showed that ethanol production by a 3000 TCD sugar factory is a profitable venture. The study showed that investment is both technically and economically feasible and will improve the financial performance and sustainability of the sugar factory in a competitive market environment. Therefore, diversification into ethanol production is an important strategy in sustaining the sugar industry in a competitive global market.

As a flooding-tolerant tree species, Taxodium distichum has been utilized in afforestation projects and proven to have important value in flooding areas. Alcohol dehydrogenase (ADH), which participates in ethanol fermentation, is essential for tolerance to the anaerobic conditions caused by flooding. In a comprehensive analysis of the ADH gene family in T. distichum, TdADHs were cloned on the basis of whole-genome sequencing, and then bioinformatic analysis, subcellular localization, and gene expression level analysis under flooding were conducted. The results show that the putative protein sequences of 15 cloned genes contained seven TdADHs and eight TdADH-like genes (one Class III ADH included) that were divided into five clades. All the sequences had an ADH_N domain, and except for TdADH-likeE2, all the other genes had an ADH_zinc_N domain. Moreover, the TdADHs in clades A, B, C, and D had a similar motif composition. Additionally, the number of TdADH amino acids ranged from 277 to 403, with an average of 370.13. Subcellular localization showed that, except for TdADH-likeD3, which was not expressed in the nucleus, the other genes were predominantly expressed in both the nucleus and cytosol. TdADH-likeC2 was significantly upregulated in all three organs (roots, stems, and leaves), and TdADHA3 was also highly upregulated under 24 h flooding treatment; the two genes might play key roles in ethanol fermentation and flooding tolerance. These findings offer a comprehensive understanding of TdADHs and could provide a foundation for the molecular breeding of T. distichum and current research on the molecular mechanisms driving flooding tolerance. [ABSTRACT FROM AUTHOR]

Mycotoxins (ochratoxin A (20 ppb), aflatoxin B1 (40 ppb), deoxynivalenol (4 ppm), and zearalenone (800 ppb)) were intentionally added to rice bran raw materials. After fermentation, their contents were determined in the distillate and distillery stillage obtained using single-stage and continuous pilot plant-scale columns. After single-stage distillation, aflatoxin B1, deoxynivalenol, and zearalenone were not detected in the distillate, indicating that even if a certain amount (four times the maximum residue limit (MRL)) was present in the raw material, it would not remain in the distillate after fermentation and distillation. Most mycotoxins remained in the distillery stillage, and their residual rates ranged from 54.0–96.2%. For ochratoxin A, 0.19 ppb was found in the distillate and this migration occurred in three consecutive distillations (0.11–0.22 ppb). Ochratoxin A and aflatoxin B1 were not detected in the distillate (alcohol content 93.9% and 95.4%, respectively) obtained from the contaminated fermented liquid (approximately three times the MRL based on the raw material) using the pilot-plant scale continuous distillation column. Therefore, the migration of mycotoxins is difficult when the distilled spirit is produced using a continuous distillation column, even if the raw material is contaminated with certain amounts of the investigated mycotoxins.

Background: Cayratia trifolia has been extensively studied for its bioactive components and medicinal properties. This study was carried out to evaluate the fermentation ability of Saccharomyces cerevisiae 2.1 yeast to determine suitable fermentation conditions. Methods: The initial sugar content for three-leaf cayratia fermentation and fermentation efficiency were calculated. The temperature for three-leaf cayratia fermentation, incubation time, and preliminary 1 liter of three-leaf cayratia wine fermentation were determined. All treatments showed that the total sugar content decreased after 9 days of fermentation compared to the initial level. Temperature during fermentation has a direct effect on yeast activity. Temperature increases the growth of yeast and speed of enzyme activity. The fermentation time changes depending on temperature, initial soluble solid contents, and yeast strains, leading to changes in the ethanol content after the end of fermentation. Results: The results showed that the S. cerevisiae 2.1 strain was able to ferment at room temperature with an initial pH of 4.5, 24 °Brix, and a yeast density of 107 cells/mL. The appropriate fermentation conditions determined for S. cerevisiae 2.1 yeast were 24 °Brix and incubation at room temperature (28-33°C) with a fermentation time of 11 days. Concusion: In 1 liter scale fermentation, three-leaf cayratia wine had 9.46% (v/v) and the fermentation efficiency was 90.34%. The wine produced had a unique color, flavor, and aroma that met the sensory evaluation of the Vietnamese standard TCVN-3217:79.

Introduction. Recent studies have shown the benefits of phytolytic enzymes to prepare grain wort in ethanol production. However, there is a lack of data on the effect of phytases and their amount on the conversion of grain polymers, the ionic composition of wort and mash, and the efficiency of yeast generation and ethanol fermentation. Study objects and methods. Wheat and corn wort samples were treated with a complex of hydrolases, including phytases. Capillary electrophoresis determined the ionic composition of wort and mash. Gas chromatography measured the content of volatile metabolites. Results and discussion. The key enzymes were phytases and proteases. They improved the conversion of grain polymers and stimulated the growth and metabolism of yeast cells. Their synergism enriched the wort with assimilable nitrogen, phosphorus, and other valuable minerals. In addition, it intensified the growth of the Saccharomyces cerevisiae yeast, increased the rate of carbohydrate consumption, and reduced the formation of side metabolites 1.7–1.9 times, mainly due to higher and aromatic alcohols. The concentration of phosphates remained practically unchanged during the fermentation of grain wort treated with phytases. However, by the end of fermentation, it was 2.4–5.1 times higher than in the mash samples without phytolytic treatment. Finally, we identified a complex of enzymes and optimal amounts of phytases that have a stimulating effect on ethanol fermentation. Conclusion. Phytases, whether used individually or together with proteases, enriched grain wort with soluble macro- and microelements, improved yeast metabolism, directed ethanol synthesis, and decreased the formation of fermentation by-products.

Abstract Background As one of the clean and sustainable energies, lignocellulosic ethanol has achieved much attention around the world. The production of lignocellulosic ethanol does not compete with people for food, while the consumption of ethanol could contribute to the carbon dioxide emission reduction. However, the simultaneous transformation of glucose and xylose to ethanol is one of the key technologies for attaining cost-efficient lignocellulosic ethanol production at an industrial scale. Genetic modification of strains and constructing consortia were two approaches to resolve this issue. Compared with strain improvement, the synergistic interaction of consortia in metabolic pathways should be more useful than using each one separately. Results In this study, the consortia consisting of suspended Scheffersomyces stipitis CICC1960 and Zymomonas mobilis 8b were cultivated to successfully depress carbon catabolite repression (CCR) in artificially simulated 80G40XRM. With this strategy, a 5.52% more xylose consumption and a 6.52% higher ethanol titer were achieved by the consortium, in which the inoculation ratio between S. stipitis and Z. mobilis was 1:3, compared with the Z. mobilis 8b mono-fermentation. Subsequently, one copy of the xylose metabolic genes was inserted into the Z. mobilis 8b genome to construct Z. mobilis FR2, leading to the xylose final-consumption amount and ethanol titer improvement by 15.36% and 6.81%, respectively. Finally, various corn stover hydrolysates with different sugar concentrations (glucose and xylose 60, 90, 120 g/L), were used to evaluate the fermentation performance of the consortium consisting of S. stipitis CICC1960 and Z. mobilis FR2. Fermentation results showed that a 1.56–4.59% higher ethanol titer was achieved by the consortium compared with the Z. mobilis FR2 mono-fermentation, and a 46.12–102.14% higher ethanol titer was observed in the consortium fermentation when compared with the S. stipitis CICC1960 mono-fermentation. Furthermore, qRT-PCR analysis of xylose/glucose transporter and other genes responsible for CCR explained the reason why the initial ratio inoculation of 1:3 in artificially simulated 80G40XRM had the best fermentation performance in the consortium. Conclusions The fermentation strategy used in this study, i.e., using a genetically modified consortium, had a superior performance in ethanol production, as compared with the S. stipitis CICC1960 mono-fermentation and the Z. mobilis FR2 mono-fermentation alone. This result showed that this strategy has potential for future lignocellulosic ethanol production.

Online monitoring of fermentation processes is a necessary task to determine concentrations of key biochemical compounds, diagnose faults in process operations, and implement feedback controllers. However, obtaining the signals of all-important variables in a real process is a task that may be difficult and expensive due to the lack of adequate sensors, or simply because some variables cannot be directly measured. From the above, a model-based approach such as state observers may be a viable alternative to solve the estimation problem. This work shows a comparative analysis of the real-time performance of a family of sliding-mode observers for reconstructing key variables in a batch bioreactor for fermentative ethanol production. These observers were selected for their robust performance under model uncertainties and finite-time estimation convergence. The selected sliding-mode observers were the first-order sliding mode observer, the proportional sliding mode observer, and the high-order sliding mode observer. For estimation purposes, a power law kinetic model for ethanol production by Saccharomyces cerevisiae was performed. A hybrid methodology allows the kinetic parameters to be adjusted, and an approach based on inference diagrams allows the observability of the model to be determined. The experimental results reported here show that the observers under analysis were robust to modeling errors and measurement noise. Moreover, the proportional sliding-mode observer was the algorithm that exhibited the best performance. [ABSTRACT FROM AUTHOR]

Simple Summary: Pyruvate decarboxylase (PDC) is a key enzyme involved in ethanol fermentation, a process for the production of biofuels. Thermostable and oxygen-stable PDC activity is highly desirable for biotechnological applications at high temperatures. The enzymes from the thermoacidophiles Saccharolobus (formerly Sulfolobus) solfataricus (Ss, Topt = 80 °C) and Sulfolobus acidocaldarius (Sa, Topt = 80 °C) were purified and characterized, and their biophysical and biochemical properties were determined comparatively. The purified enzymes were CoA-dependent and thermostable. There was no loss of activity in the presence of oxygen. In conclusion, both thermostable SsPDC and SaPDC catalyze the CoA-dependent production of acetaldehyde from pyruvate in the presence of oxygen. Pyruvate decarboxylase (PDC) is a key enzyme involved in ethanol fermentation, and it catalyzes the decarboxylation of pyruvate to acetaldehyde and CO2. Bifunctional PORs/PDCs that also have additional pyruvate:ferredoxin oxidoreductase (POR) activity are found in hyperthermophiles, and they are mostly oxygen-sensitive and CoA-dependent. Thermostable and oxygen-stable PDC activity is highly desirable for biotechnological applications. The enzymes from the thermoacidophiles Saccharolobus (formerly Sulfolobus) solfataricus (Ss, Topt = 80 °C) and Sulfolobus acidocaldarius (Sa, Topt = 80 °C) were purified and characterized, and their biophysical and biochemical properties were determined comparatively. Both enzymes were shown to be heterodimeric, and their two subunits were determined by SDS-PAGE to be 37 ± 3 kDa and 65 ± 2 kDa, respectively. The purified enzymes from S. solfataricus and S. acidocaldarius showed both PDC and POR activities which were CoA-dependent, and they were thermostable with half-life times of 2.9 ± 1 and 1.1 ± 1 h at 80 °C, respectively. There was no loss of activity in the presence of oxygen. Optimal pH values for their PDC and POR activity were determined to be 7.9 and 8.6, respectively. In conclusion, both thermostable SsPOR/PDC and SaPOR/PDC catalyze the CoA-dependent production of acetaldehyde from pyruvate in the presence of oxygen. [ABSTRACT FROM AUTHOR]

Abstract Background The industrial production of various alcohols from organic carbon compounds may be performed at high rates and with a low risk of contamination using thermophilic microorganisms as whole-cell catalysts. Thermoanaerobacter species that thrive around 50–75 °C not only perform fermentation of sugars to alcohols, but some also utilize different organic acids as electron acceptors, reducing them to their corresponding alcohols. Results We purified AdhE as the major NADH- and AdhB as the major NADPH-dependent alcohol dehydrogenase (ADH) from the cell extract of the organic acid-reducing Thermoanaerobacter sp. strain X514. Both enzymes were present in high amounts during growth on glucose with and without isobutyrate, had broad substrate spectra including different aldehydes, with high affinities (

Municipal solid waste management and its disposal are considered one of the major challenges facing the urban communities around the world. Effective solid waste treatment involves a variety of approaches, treatment technologies and concepts to ensure the protection of public health and the environment. Waste landfill is the method most commonly used worldwide, despite all the significant environmental, health and economic consequences. Thus, alternative methods such as the municipal solid waste pretreatment, fermentation, ethanol fermentation and anaerobic fermentation have been the focus of heightened attention. Using these methods, an alcohol-containing liquid was obtained from 1.5 kg of the organic fraction of municipal solid waste (OFMSW), which had an ethanol percentage of 97.45%. Consequently, if properly managed and used, municipal solid waste can be a viable source of energy rather than a source of pollution.

Saccharomyces cerevisiae is the most important industrial microorganism used to fuel ethanol production worldwide. Herein, we obtained a mutant S. cerevisiae strain with improved capacity for ethanol fermentation, from 13.72% (v/v for the wild-type strain) to 16.13% (v/v for the mutant strain), and analyzed its genomic structure and gene expression changes. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment revealed that the changed genes were mainly enriched in the pathways of carbohydrate metabolism, amino acid metabolism, metabolism of cofactors and vitamins, and lipid metabolism. The gene expression trends of the two strains were recorded during fermentation to create a timeline. Venn diagram analysis revealed exclusive genes in the mutant strain. KEGG enrichment of these genes showed upregulation of genes involved in sugar metabolism, mitogen-activated protein kinase pathway, fatty acid and amino acid degradation, and downregulation of genes involved in oxidative phosphorylation, ribosome, fatty acid and amino acid biogenesis. Protein interaction analysis of these genes showed that glucose-6-phosphate isomerase 1, signal peptidase complex subunit 3, 6-phosphofructokinase 2, and trifunctional aldehyde reductase were the major hub genes in the network, linking pathways together. These findings provide new insights into the adaptive metabolism of S. cerevisiae for ethanol production and a framework for the construction of engineered strains of S. cerevisiae with excellent ethanol fermentation capacity.

Currently, most cotton textile waste is sent to landfill. However, due to the use of synthetic additives and the chemical treatment of cotton fibers, cotton textile waste is difficult to biodegrade. Cotton textile waste can also be subjected to material recycling, or to incineration/gasification to produce energy. Here, we present the optimization of acid hydrolysis of cotton yarn fibers for glucose efficiency. The cotton yarn hydrolysates showed great potential for replacing simple sugar solutions in fermentation media. The highest glucose concentration was obtained in the hydrolysates of cotton yarn hydrolyzed in a 2% solution of sulfuric acid or phosphoric acid at 140–160 °C for 2 h. After 2 h of hydrolysis at 140 °C with 2% H3PO4, the concentration of glucose in the cotton yarn hydrolysate (13.19 g/L) increased fivefold compared with cotton yarn treated under the same conditions with H2SO4 (2.65 g/L). The structural modifications in the solid residues after acid hydrolysis were analyzed using a scanning electron microscope with energy dispersive spectroscopy (SEM-EDS), attenuated total reflectance Fourier-transform infrared spectroscopy (FTIR-ATR), and Raman spectroscopy. The SEM images, IR spectra, and Raman spectra revealed that the most significant changes in the morphology of the fibers occurred when the process was carried out at high temperatures (≥140 °C). Better growth of the yeast strains Saccharomyces cerevisiae Ethanol Red and Saccharomyces cerevisiae Tokay ŁOCK0204 was observed in the medium containing phosphoric acid hydrolysate. The maximum methane yield of 278 dm3/kgVS and the maximum hydrogen yield of 42 dm/kgVS were reported for cotton yarn waste after pretreatment with H3PO4. This might have been linked to the beneficial effect of phosphorus, which is a key nutrient for anaerobic digestion. The proposed hydrolysis method does not generate fermentation inhibitors. [ABSTRACT FROM AUTHOR]

Thermotolerant ethanologenic yeasts have attracted the interest of many scientists due to the current challenges caused by increasing global temperature, the benefits associated with processing at high temperatures, and the potential to reduce cooling costs. The objectives of this study are to characterize the selected thermotolerant yeasts and to evaluate their use in Cayratia trifolia fermentation at high temperatures. A total of 151 yeast strains isolated from 53 samples of Cayratia trifolia were studied for their morphology, physiology, biochemistry, and their phylogenetic relationship. Based on the results of tests for thermotolerance ability (37-45oC) and ethanol tolerance capacity (9-12% v/v), 57 of the 151 yeast isolates were selected to be tested for use in wine fermentation from three-leaf cayratia at 37oC. Thirty isolates that were found to have high fermentation ability and that produced an ethanol concentration of between 6.0 and 9.9% (v/v) were selected for identification using amplified 26S rDNA sequences. The yeasts were identified as follows: Candida glabrata (BL2.1, CT1.1, CT1.3, CT2.3, HG2.1), Candida tropicalis (KG1.1, KG3.2, CM3.3, HG3.3, TG1.1, TG3.1), Candida nivariensis (DT1.2, CM3.2, ST2.1, BT1.2), Pichia kudriavzevii (KG2.1, KG5.1, AG2.1, AG2.3, AG4.2, DT3.2, LA1.3, CM4.4, BT2.1, BT3.3, TV4.2, CT4.2, VL1.1), Clavispora lusitaniae (TG4.2), and Saccharomyces cerevisiae (HG1.3). The phylogenetic tree constructed using MEGA 6 with bootstrap analysis performed by repeating the data 1,000 times revealed that the selected yeast strains were closely related. The newly isolated strain of S. cerevisiae HG1.3 producing the highest ethanol concentration of 9.9% (v/v) in Cayratia trifolia wine fermentation at 370C was selected for further study.

Kiwifruit (Actinidia deliciosa) ‘Bruno’ is prone to accumulate ethanol rapidly after respiratory climacteric during storage at ambient conditions without stresses, which causes quality deterioration of the fruit associated with alcohol off-flavor. For maintaining the postharvest quality of kiwifruit ‘Bruno’, the effects of 1.0 µL • L−1 1-methylcyclopropene (1-MCP) treatment on regulating the ethanol fermentation and γ-aminobutyric acid (GABA) shunt pathway associated with control of alcohol off-flavor were investigated during storage at room temperature (24 ± 1) °C for 27 days. The results showed that 1-MCP treatment significantly reduced the respiration rate, ethylene production, decay rate, ascorbic acid (AsA) loss, and delayed the decline in the firmness and titratable acidity (TA), and the increase in total soluble solid (TSS) in kiwifruit. Furthermore, 1-MCP treatment effectively inhibited the increases in contents of acetaldehyde, ethanol, and GABA along with the suppressed activities of key enzymes involved in ethanol fermentation and GABA shunt pathway, such as pyruvate decarboxylase (PDC), alcohol dehydrogenase (ADH), glutamate decarboxylase (GAD), and GABA-transaminase (GABA-T) in kiwifruit during storage. In conclusion, 1-MCP treatment efficiently regulated the ethanol fermentation and GABA shunt pathway by delaying the ripening process to avoid the alcohol off-flavor development, thereby contributed to maintaining the quality of the kiwifruit.

As a flooding-tolerant tree species, Taxodium distichum has been utilized in afforestation projects and proven to have important value in flooding areas. Alcohol dehydrogenase (ADH), which participates in ethanol fermentation, is essential for tolerance to the anaerobic conditions caused by flooding. In a comprehensive analysis of the ADH gene family in T. distichum, TdADHs were cloned on the basis of whole-genome sequencing, and then bioinformatic analysis, subcellular localization, and gene expression level analysis under flooding were conducted. The results show that the putative protein sequences of 15 cloned genes contained seven TdADHs and eight TdADH-like genes (one Class III ADH included) that were divided into five clades. All the sequences had an ADH_N domain, and except for TdADH-likeE2, all the other genes had an ADH_zinc_N domain. Moreover, the TdADHs in clades A, B, C, and D had a similar motif composition. Additionally, the number of TdADH amino acids ranged from 277 to 403, with an average of 370.13. Subcellular localization showed that, except for TdADH-likeD3, which was not expressed in the nucleus, the other genes were predominantly expressed in both the nucleus and cytosol. TdADH-likeC2 was significantly upregulated in all three organs (roots, stems, and leaves), and TdADHA3 was also highly upregulated under 24 h flooding treatment; the two genes might play key roles in ethanol fermentation and flooding tolerance. These findings offer a comprehensive understanding of TdADHs and could provide a foundation for the molecular breeding of T. distichum and current research on the molecular mechanisms driving flooding tolerance.

In this study, major factors involved in regulating ethanol production from wood by the saprotrophic white-rot fungus Phanerochaete sordida YK-624 were investigated. P. sordida YK-624 produced ethanol from wood meal culture without the addition of any nutrients, and ethanol was produced from wood culture only when the oxygen concentration in headspace was reduced to ≤5%; thereafter, ethanol production ceased within a few days. Analyses of gene expression during aerobic incubation indicated that P. sordida simultaneously upregulates the glycolytic pathway from sugar uptake to pyruvate conversion during ethanol fermentation and suppresses pyruvate influx into the TCA cycle. Upon termination of ethanol fermentation, the expression of all tested genes was repressed, and the fungus ceased to grow. In contrast, the fungus could utilize ethanol for aerobic growth. These results suggest that ethanol fermentation by P. sordida functions as a short-term stress response system under anaerobic conditions during wood decay, enabling the fungus to rapidly resume growing when oxygen is supplied (e.g., following breakdown of plant cell walls or removal of the fungus from water immersion). This is the first report to describe the physiologic significance of ethanol fermentation in saprotrophic white-rot fungi. [Display omitted] • The biological significance of ethanol fermentation by saprotrophic white-rot fungus P. sordida YK-624 was speculated. • P. sordida YK-624 produced ethanol from wood meal without addition of any nutrients under hypoxic condition. • P. sordida YK-624 could survive and ferment for only a few days under hypoxia conditions during wood decay. • Ethanol fermentation in P. sordida YK-624 appears to be a short-term hypoxia tolerance system. [ABSTRACT FROM AUTHOR]

Bioethanol is alternative renewable energy typically obtained from glucose through a fermentation process using Saccharomyces cerevisiae. In the bioethanol fermentation process using yeast, there are several inhibiting factors, such as a high concentration of substrate, ethanol as the product, and nutrients. The present study aimed to investigate the effect of fermentation time (12- 72 hours), immobilized carrier size (sizes of 0.5 cm3, 1 cm3, and 1.5 cm3), and medium pH (3.0, 4.0, and 5.0) on the ethanol fermentation process using immobilized yeast in jackfruit (Artocarpus heterophyllus) seeds and subsequently to compare its performance with a free cell system. The highest ethanol concentration (89.15 g/L) with a yield of 96.92% was obtained by immobilizing yeast in jackfruit seed at a fermentation time of 72 hours, carrier size of 0.5 cm3, and medium pH of 5.0. When compared to the free cell system fermentation under identical operating conditions, immobilized yeast in jackfruit seed obtained 1.41 times higher ethanol concentration. Jackfruit seed also led to a higher ethanol concentration compared to other S. cerevisiae carriers. Altogether, our findings imply that jackfruit seed has great potential as a carrier of S. cerevisiae in the process of fermenting glucose into ethanol. [ABSTRACT FROM AUTHOR]

Banana (M. acuminata x balbisiana) is an abundant lignocellulosic waste material in large plantations all over the Philippines, especially in Mindanao, which can be utilized as substrate in producing high-value products like ethanol. To compensate for the low yield based on total weight of substrate due to the high moisture content of banana pseudostem, there is the primary challenge to make the conversion of this lignocellulosic biomass into monomeric sugar and then into ethanol more efficiently in order to achieve yields that would make it cost-competitive. Hence, this study evaluated the effects of solid loading, incubation time and amount of enzyme on yield of reducing sugars in the enzymatic hydrolysis process and attempted to optimize the significant factors by Response Surface Methodology (RSM), specifically using Box-Behnken design. There was significant improvement on the reducing sugar yield of the pretreated banana pseudostem at 20 h incubation time, 15 g solid loading and 0.55 % enzyme concentration. Ethanol production was observed to be higher in the detoxified substrate although biomass was higher for the non-detoxified substrate. As to our knowledge, the present study is the first attempt to produce second generation ethanol using banana pseudostem waste as feedstock in the Philippines.

In this work, we have tested individual and combination of applications of ozonolysis and liquid hot water (LHW) to pretreat sugarcane bagasse (SCB) for the removal of enzyme and/or microbial inhibitors and generation of potential value-added chemicals. A solid content with 80% cellulose and a liquid phase (liquor) rich in phenolic derived compounds (3 g.L−1) from lignin, sugars (>20 g.L−1), and other compounds, such as furfural and hydroxymethylfurfural (HMF), were generated. Maximal (59%) glucan conversion occurred in the presence of double-pretreated bagasse, which had 32–50% more glucan available than the samples that were individually LHW or ozone-pretreated, resulting in maximal ethanol production (92% after 42 h) from double-pretreated SCB enzyme hydrolyzate. In summary, this work showed that ozone reacts effectively with lignin without the use of any other chemical reagent, and LHW pretreatment, followed by a washing step, was effective in solubilizing and cleaning up the fiber enzyme and microbial inhibitory compounds with ozone being effective against phenolics. Moreover, the generated cellulose-rich substrate is readily fermentable. The acidic liquor fraction removed by sequential washings and containing mainly sugars and phenolic compounds may be evaluated for use in green chemistry bioconversions processes.

Background: As one of the clean and sustainable energies, lignocellulosic ethanol has achieved much attention around the world. The production of lignocellulosic ethanol does not compete with people for food, while the consumption of ethanol could contribute to the carbon dioxide emission reduction. However, the simultaneous transformation of glucose and xylose to ethanol is one of the key technologies for attaining cost-efficient lignocellulosic ethanol production at an industrial scale. Genetic modification of strains and constructing consortia were two approaches to resolve this issue. Compared with strain improvement, the synergistic interaction of consortia in metabolic pathways should be more useful than using each one separately. Results: In this study, the consortia consisting of suspended Scheffersomyces stipitis CICC1960 and Zymomonas mobilis 8b were cultivated to successfully depress carbon catabolite repression (CCR) in artificially simulated 80G40XRM. With this strategy, a 5.52% more xylose consumption and a 6.52% higher ethanol titer were achieved by the consortium, in which the inoculation ratio between S. stipitis and Z. mobilis was 1:3, compared with the Z. mobilis 8b mono-fermentation. Subsequently, one copy of the xylose metabolic genes was inserted into the Z. mobilis 8b genome to construct Z. mobilis FR2, leading to the xylose final-consumption amount and ethanol titer improvement by 15.36% and 6.81%, respectively. Finally, various corn stover hydrolysates with different sugar concentrations (glucose and xylose 60, 90, 120 g/L), were used to evaluate the fermentation performance of the consortium consisting of S. stipitis CICC1960 and Z. mobilis FR2. Fermentation results showed that a 1.56–4.59% higher ethanol titer was achieved by the consortium compared with the Z. mobilis FR2 mono-fermentation, and a 46.12–102.14% higher ethanol titer was observed in the consortium fermentation when compared with the S. stipitis CICC1960 mono-fermentation. Furthermore, qRT-PCR analysis of xylose/glucose transporter and other genes responsible for CCR explained the reason why the initial ratio inoculation of 1:3 in artificially simulated 80G40XRM had the best fermentation performance in the consortium. Conclusions: The fermentation strategy used in this study, i.e., using a genetically modified consortium, had a superior performance in ethanol production, as compared with the S. stipitis CICC1960 mono-fermentation and the Z. mobilis FR2 mono-fermentation alone. This result showed that this strategy has potential for future lignocellulosic ethanol production. [ABSTRACT FROM AUTHOR]

Bioethanol is an environmentally benign renewable energy commonly obtained from glucose fermentation using Saccharomyces cerevisiae. The purposes of this study are to investigate the effects of time, temperature, pH, immobilized yeast cell loading, beads reuse during ethanol production through batch fermentation of glucose derived from oil palm empty fruit bunches by S. cerevisiae immobilized on Na-alginate beads and to compare the performance of fermentation using immobilized yeast cells and that of using a free cell system. The results revealed that time, temperature, pH, yeast mass and beads reuse significantly affected the ethanol and final glucose concentrations. As expected, a maximum ethanol concentration was obtained from fermentation using immobilized yeast cells at 30 °C, pH 5, and immobilized yeast cell loading of 0.75 g for 48 hours. However, fermentation with a free cell system at the same conditions resulted in lower ethanol yield. The highest ethanol concentration of 88.125 g/L with a productivity of 1.84 g/L·h was achieved from the second cycle fermentation using of immobilized cells beads. The results suggest that an immobilized cell system exhibits great potential applications for improved ethanol production due to its ability to sustain the stability of cell activity, reduce contamination tendency, and protect yeast cells from any possible inhibitions. [ABSTRACT FROM AUTHOR]

Municipal solid waste management and its disposal are considered one of the major challenges facing the urban communities around the world. Effective solid waste treatment involves a variety of approaches, treatment technologies and concepts to ensure the protection of public health and the environment. Waste landfill is the method most commonly used worldwide, despite all the significant environmental, health and economic consequences. Thus, alternative methods such as the municipal solid waste pretreatment, fermentation, ethanol fermentation and anaerobic fermentation have been the focus of heightened attention. Using these methods, an alcohol-containing liquid was obtained from 1.5 kg of the organic fraction of municipal solid waste (OFMSW), which had an ethanol percentage of 97.45%. Consequently, if properly managed and used, municipal solid waste can be a viable source of energy rather than a source of pollution. [ABSTRACT FROM AUTHOR]

Background: Ethanol concentration (PE), ethanol productivity (QP) and sugar consumption (SC) are important values in industrial ethanol production. In this study, initial sugar and nitrogen (urea) concentrations in sweet sorghum stem juice (SSJ) were optimized for high PE (≥10%, v/v), QP, (≥2.5 g/L·h) and SC (≥90%) by Saccharomyces cerevisiae SSJKKU01. Then, repeated-batch fermentations under normal gravity (NG) and high gravity (HG) conditions were studied. Results: The initial sugar at 208 g/L and urea at 2.75 g/L were the optimum values to meet the criteria. At the initial yeast cell concentration of ~1 × 108 cells/mL, the PE, QP and SC were 97.06 g/L, 3.24 g/L·h and 95.43%, respectively. Repeated-batch fermentations showed that the ethanol production efficiency of eight successive cycles with and without aeration were not significantly different when the initial sugar of cycles 2 to 8 was under NG conditions (~140 g/L). Positive effects of aeration were observed when the initial sugar from cycle 2 was under HG conditions (180–200 g/L). The PE and QP under no aeration were consecutively lower from cycle 1 to cycle 6. Additionally, aeration affected ergosterol formation in yeast cell membrane at high ethanol concentrations, whereas trehalose content under all conditions was not different. Conclusion: Initial sugar, sufficient nitrogen and appropriated aeration are necessary for promoting yeast growth and ethanol fermentation. The SSJ was successfully used as an ethanol production medium for a high level of ethanol production. Aeration was not essential for repeated-batch fermentation under NG conditions, but it was beneficial under HG conditions.How to cite: Sriputorn B, Laopaiboon P, Phukoetphim N, et al. Enhancement of ethanol production efficiency in repeated-batch fermentation from sweet sorghum stem juice: Effect of initial sugar, nitrogen and aeration. Electron J Biotechnol 2020;46. https://doi.org/10.1016/j.ejbt.2020.06.001

The objective of this study was to assess the suitability of buckwheat grain for agricultural distillate production. The effects of the method of starch liberation on the physicochemical composition of the prepared mashes, fermentation results, the concentration of volatile compounds in the obtained distillates, and their organoleptic features, were investigated. The raw materials used were two cultivars of buckwheat grains, Panda and Kora. Both cultivars were characterized by similar starch content, however Panda cultivar had a significantly higher content of reducing sugars. Fermentation of the Kora cultivar-based mashes resulted in a higher fermentation efficiency (up to approximately 85% of the theoretical yield) compared to the Panda cultivar-based mashes (up to approx. 75%). Of the tested methods of starch liberation, the pressure-thermal treatment was revealed as superior, especially in the case of the Panda cultivar. In the case of the Kora cultivar, both the pressure-thermal method and the pressureless method of starch liberation resulted in a high process efficiency (up to 85% of the theoretical). Supplementation of the buckwheat mashes with (NH 4 ) 2 HPO 4 improved fermentation results. The highest scores in sensory assessment were given to distillates from mashes prepared with the pressure-thermal treatment, which contained relatively low concentrations of undesirable compounds, such as acetaldehyde and methanol, and revealed pleasant organoleptic features.

The nicotine from tobacco stalk showed obvious inhibitory effect on the activity of cellulase and fermentability of microorganisms, which seriously hinders the utilization of tobacco stalk. Dilute sulfuric acid presoak of tobacco stalk was used to enhance the performance of instant catapult steam explosion (ICSE) for tobacco stalk pretreatment. The presoak was beneficial to break the recalcitrant structure of tobacco stalk, reduce nicotine content to relieve the inhibition on the activity of cellulase and metabolism of microorganisms, and promote the performance of enzymatic hydrolysis and ethanol fermentation. The optimized 0.8% sulfuric acid (w/w) presoak–integrated ICSE pretreatment resulted in 85.54% nicotine removal from tobacco stalk; meanwhile, the total sugar concentration from enzymatic hydrolysis of pretreated tobacco stalk increased from 33.40 to 53.81 g/L (the ratio of dry tobacco stalk to water was 1:8, w/w), ethanol concentration increased 103.36% from 5.95 to 12.10 g/L in flask, compared with separate ICSE pretreatment. Finally, the ethanol concentration achieved the highest 23.53 g/L in a 5-L fermenter with the ethanol yield from the glucose of tobacco stalk hydrolysate achieving 71.40% by increasing the solid loading of the tobacco stalk in the enzymatic hydrolysis process (the ratio of dry tobacco stalk to water was 1:4, w/w). These results achieved the expected purpose of efficient utilization of discarded tobacco stalk.

Background: The industrial production of various alcohols from organic carbon compounds may be performed at high rates and with a low risk of contamination using thermophilic microorganisms as whole-cell catalysts. Thermoanaerobacter species that thrive around 50–75 °C not only perform fermentation of sugars to alcohols, but some also utilize different organic acids as electron acceptors, reducing them to their corresponding alcohols. Results: We purified AdhE as the major NADH- and AdhB as the major NADPH-dependent alcohol dehydrogenase (ADH) from the cell extract of the organic acid-reducing Thermoanaerobacter sp. strain X514. Both enzymes were present in high amounts during growth on glucose with and without isobutyrate, had broad substrate spectra including different aldehydes, with high affinities (< 1 mM) for acetaldehyde and for NADH (AdhE) or NADPH (AdhB). Both enzymes were highly thermostable at the physiological temperature of alcohol production. In addition to AdhE and AdhB, we identified two abundant AdhA-type ADHs based on their genes, which were recombinantly produced and biochemically characterized. The other five ADHs encoded in the genome were only expressed at low levels. Conclusions: According to their biochemical and kinetic properties, AdhE and AdhB are most important for ethanol formation from sugar and reduction of organic acids to alcohols, while the role of the two AdhA-type enzymes is less clear. AdhE is the only abundant aldehyde dehydrogenase for the acetyl-CoA reduction to aldehydes, however, acid reduction may also proceed directly by aldehyde:ferredoxin oxidoreductase. The role of the latter in bio-alcohol formation from sugar and in organic acid reduction needs to be elucidated in future studies. [ABSTRACT FROM AUTHOR]

Fuel ethanol has been consolidated in view of the current energy matrix. However, for an efficient process, the yeast and the quality of the substrate used must be taken into account. Thus, this study aims to evaluate sugarcane juice and sweet sorghum and analyze cell viability, ethanol production and the assimilation of amino acids present in these substrates as a function of temperature. The Industrial yeasts Saccharomyces cerevisiae FT858 and Pedra-2 (Pe-2) were activated with pre-inoculum of 2% YPD. The biomass obtained was inoculated on the substrates and incubated at 30 and 40°C at 250 rpm. Aliquots were collected for feasibility analyses with methylene blue, ethanol by gas chromatography and amino acids by high performance liquid chromatography. Yeasts showed better performance in sorghum broth at 30°C. The viability of 89% for Pe-2 and 85% for FT858 and ethanol concentration of 10% (v v-1) for both yeasts. At 40°C there was a reduction in these parameters. Sorghum had the highest amount of amino acids and serine, arginine, alanine and tryptophan were effectively assimilated by yeasts. The temperature of 40°C interfered in the metabolic capacity of the yeasts, causing thermal stress, inducing a greater consumption of amino acids. [ABSTRACT FROM AUTHOR]

ABSTRACT Mbp1p is a component of MBF (MluI cell cycle box binding factor, Mbp1p-Swi6p) and is well known to regulate the G1–S transition of the cell cycle. However, few studies have provided clues regarding its role in fermentation. This work aimed to recognize the function of the MBP1 gene in ethanol fermentation in a wild-type industrial Saccharomyces cerevisiae strain. MBP1 deletion caused an obvious decrease in the final ethanol concentration under oxygen-limited (without agitation), but not under aerobic, conditions (130 rpm). Furthermore, the mbp1Δ strain showed 84% and 35% decreases in respiration intensity under aerobic and oxygen-limited conditions, respectively. These findings indicate that MBP1 plays an important role in responding to variations in oxygen content and is involved in the regulation of respiration and fermentation. Unexpectedly, mbp1Δ also showed pseudohyphal growth, in which cells elongated and remained connected in a multicellular arrangement on yeast extract-peptone-dextrose (YPD) plates. In addition, mbp1Δ showed an increase in cell volume, associated with a decrease in the fraction of budded cells. These results provide more detailed information about the function of MBP1 and suggest some clues to efficiently improve ethanol production by industrially engineered yeast strains. IMPORTANCE Saccharomyces cerevisiae is an especially favorable organism used for ethanol production. However, inhibitors and high osmolarity conferred by fermentation broth, and high concentrations of ethanol as fermentation runs to completion, affect cell growth and ethanol production. Therefore, yeast strains with high performance, such as rapid growth, high tolerance, and high ethanol productivity, are highly desirable. Great efforts have been made to improve their performance by evolutionary engineering, and industrial strains may be a better start than laboratory ones for industrial-scale ethanol production. The significance of our research is uncovering the function of MBP1 in ethanol fermentation in a wild-type industrial S. cerevisiae strain, which may provide clues to engineer better-performance yeast in producing ethanol. Furthermore, the results that lacking MBP1 caused pseudohyphal growth on YPD plates could shed light on the development of xylose-fermenting S. cerevisiae, as using xylose as the sole carbon source also caused pseudohyphal growth.

Online monitoring of fermentation processes is a necessary task to determine concentrations of key biochemical compounds, diagnose faults in process operations, and implement feedback controllers. However, obtaining the signals of all-important variables in a real process is a task that may be difficult and expensive due to the lack of adequate sensors, or simply because some variables cannot be directly measured. From the above, a model-based approach such as state observers may be a viable alternative to solve the estimation problem. This work shows a comparative analysis of the real-time performance of a family of sliding-mode observers for reconstructing key variables in a batch bioreactor for fermentative ethanol production. These observers were selected for their robust performance under model uncertainties and finite-time estimation convergence. The selected sliding-mode observers were the first-order sliding mode observer, the proportional sliding mode observer, and the high-order sliding mode observer. For estimation purposes, a power law kinetic model for ethanol production by Saccharomyces cerevisiae was performed. A hybrid methodology allows the kinetic parameters to be adjusted, and an approach based on inference diagrams allows the observability of the model to be determined. The experimental results reported here show that the observers under analysis were robust to modeling errors and measurement noise. Moreover, the proportional sliding-mode observer was the algorithm that exhibited the best performance.

Pyruvate decarboxylase (PDC) is a key enzyme involved in ethanol fermentation, and it catalyzes the decarboxylation of pyruvate to acetaldehyde and CO2. Bifunctional PORs/PDCs that also have additional pyruvate:ferredoxin oxidoreductase (POR) activity are found in hyperthermophiles, and they are mostly oxygen-sensitive and CoA-dependent. Thermostable and oxygen-stable PDC activity is highly desirable for biotechnological applications. The enzymes from the thermoacidophiles Saccharolobus (formerly Sulfolobus) solfataricus (Ss, Topt = 80 °C) and Sulfolobus acidocaldarius (Sa, Topt = 80 °C) were purified and characterized, and their biophysical and biochemical properties were determined comparatively. Both enzymes were shown to be heterodimeric, and their two subunits were determined by SDS-PAGE to be 37 ± 3 kDa and 65 ± 2 kDa, respectively. The purified enzymes from S. solfataricus and S. acidocaldarius showed both PDC and POR activities which were CoA-dependent, and they were thermostable with half-life times of 2.9 ± 1 and 1.1 ± 1 h at 80 °C, respectively. There was no loss of activity in the presence of oxygen. Optimal pH values for their PDC and POR activity were determined to be 7.9 and 8.6, respectively. In conclusion, both thermostable SsPOR/PDC and SaPOR/PDC catalyze the CoA-dependent production of acetaldehyde from pyruvate in the presence of oxygen.

This study aimed to utilize the enzymatic hydrolysis of leftover cooked rice (LCR) for fermentative ethanol production. Effect of glucoamylase volumes (V1: 5 U/g, V2: 25 U/g, and V3: 50 U/g) on the performance of LCR hydrolysis was also evaluated. It was found that the highest chemical oxygen demand (COD) of 77.5 g/L and reducing sugar (RS) of 34.6 g/L were achieved at V3. The LCR hydrolyzate obtained from enzymatic hydrolysis was then used as feedstock for ethanol fermentation. Higher ethanol production was obtained when RS increased from 18.7 g/L (V1) to 23.2 g/L (V2). However, lower ethanol production was found when RS further increased to 34.6 g/L (V3) probably because too high RS concentration led to the inhibition on the yeast. The maximum ethanol production and yield were 21.1 g/L and 0.3 g ethanol/g LCR, respectively. The LCR could be a promising substrate for fermentative ethanol production for industrial application.

Increasing ethanol demand and public concerns about environmental protection promote the production of lignocellulosic bioethanol. Compared to that of starch- and sugar-based bioethanol production, the production of lignocellulosic bioethanol is water-intensive. A large amount of water is consumed during pretreatment, detoxification, saccharification, and fermentation. Water is a limited resource, and very high water consumption limits the industrial production of lignocellulosic bioethanol and decreases its environmental feasibility. In this review, we focused on the potential for reducing water consumption during the production of lignocellulosic bioethanol by performing pretreatment and fermentation at high solid loading, omitting water washing after pretreatment, and recycling wastewater by integrating bioethanol production and anaerobic digestion. In addition, the feasibility of these approaches and their research progress were discussed. This comprehensive review is expected to draw attention to water competition between bioethanol production and human use. [ABSTRACT FROM AUTHOR]

Mathematical models of ethanol fermentation in continuous conventional bioreactor (CCBR) and continuous membrane bioreactor (CMBR) are discussed theoretically. Each of these models is a system of nonlinear equations containing nonlinear terms related to Monod model for each retention time at non-steady condition. Analytical expressions of cell growth, glucose consumption, and ethanol production are obtained using a new approach of the homotopy perturbation method. A detailed study of the parameters effects on each of the governing systems is also presented. The obtained analytical solution is shown to be in strong agreement with most reliable numerical methods. [ABSTRACT FROM AUTHOR]

Currently, most cotton textile waste is sent to landfill. However, due to the use of synthetic additives and the chemical treatment of cotton fibers, cotton textile waste is difficult to biodegrade. Cotton textile waste can also be subjected to material recycling, or to incineration/gasification to produce energy. Here, we present the optimization of acid hydrolysis of cotton yarn fibers for glucose efficiency. The cotton yarn hydrolysates showed great potential for replacing simple sugar solutions in fermentation media. The highest glucose concentration was obtained in the hydrolysates of cotton yarn hydrolyzed in a 2% solution of sulfuric acid or phosphoric acid at 140–160 °C for 2 h. After 2 h of hydrolysis at 140 °C with 2% H3PO4, the concentration of glucose in the cotton yarn hydrolysate (13.19 g/L) increased fivefold compared with cotton yarn treated under the same conditions with H2SO4 (2.65 g/L). The structural modifications in the solid residues after acid hydrolysis were analyzed using a scanning electron microscope with energy dispersive spectroscopy (SEM-EDS), attenuated total reflectance Fourier-transform infrared spectroscopy (FTIR-ATR), and Raman spectroscopy. The SEM images, IR spectra, and Raman spectra revealed that the most significant changes in the morphology of the fibers occurred when the process was carried out at high temperatures (≥140 °C). Better growth of the yeast strains Saccharomyces cerevisiae Ethanol Red and Saccharomyces cerevisiae Tokay ŁOCK0204 was observed in the medium containing phosphoric acid hydrolysate. The maximum methane yield of 278 dm3/kgVS and the maximum hydrogen yield of 42 dm/kgVS were reported for cotton yarn waste after pretreatment with H3PO4. This might have been linked to the beneficial effect of phosphorus, which is a key nutrient for anaerobic digestion. The proposed hydrolysis method does not generate fermentation inhibitors.

Abstract Background Ensiling cannot be utilized as a stand-alone pretreatment for sugar-based biorefinery processes but, in combination with hydrothermal processing, it can enhance pretreatment while ensuring a stable long-term storage option for abundant but moist biomass. The effectiveness of combining ensiling with hydrothermal pretreatment depends on biomass nature, pretreatment, and silage conditions. Results In the present study, the efficiency of the combined pretreatment was assessed by enzymatic hydrolysis and ethanol fermentation, and it was demonstrated that ensiling of sugarcane bagasse produces organic acids that can partly degrade biomass structure when in combination with hydrothermal treatment, with the consequent improvement of the enzymatic hydrolysis of cellulose and of the overall 2G bioethanol process efficiency. The optimal pretreatment conditions found in this study were those using ensiling and/or hydrothermal pretreatment at 190 °C for 10 min as this yielded the highest overall glucose recovery yield and ethanol yield from the raw material (0.28–0.30 g/g and 0.14 g/g, respectively). Conclusion Ensiling prior to hydrothermal pretreatment offers a controlled solution for wet storage and long-term preservation for sugarcane bagasse, thus avoiding the need for drying. This preservation method combined with long-term storage practice can be an attractive option for integrated 1G/2G bioethanol plants, as it does not require large capital investments or energy inputs and leads to comparable or higher overall sugar recovery and ethanol yields.

We previously observed that as glucose is completely exhausted during ethanol fermentation, the dissolved carbon dioxide (DCO2) level in the fermenter will suddenly decline. This observation was implemented to design and develop a DCO2‐driven‐and‐controlled repeated batch fermentation process for ethanol production. The process was tested at four different glucose concentrations (~150 g/L, ~200 g/L, ~250 g/L, and ~300 g/L), and each glucose concentration was controlled under three respective DCO2 control levels (without DCO2 control, and DCO2 controlled at either 1000 mg/L or 750 mg/L). The results show that reported process features complete glucose utilization and is self‐driven. For glucose concentration less than 200 g/L, ~41%‐50% of fermentation time per batch was saved during the repeated batch operation. It took 12.1 ± 1.1 hours‐14.9 ± 1.9 hours to complete a batch with glucose feed at ~250 g/L and 21.7 ± 6 hours‐31.5 ± 7 hours to complete a batch with glucose feed at ~300 g/L. The reported process is time saving and stable, but the ethanol yield is ~20% lower than the operation without DCO2 control. Dissolved CO2 control became essential for glucose concentrations greater than 250 g/L if zero glucose discharge in each batch during the operation is desired. [ABSTRACT FROM AUTHOR]

Zymomonas mobilis is a model ethanologenic bacterium for diverse biochemical production. Rich medium (RM) is a complex medium that is routinely used to cultivate Z. mobilis, which contains carbon sources such as glucose, nitrogen sources such as yeast extract (YE), and KH2PO4. Glucose consumption and cell growth of Z. mobilis is usually coupled during ethanol fermentation. However, sometimes glucose was not consumed during the exponential growth phase, and it took extended time for cells to consume glucose and produce ethanol, which eventually reduced the ethanol productivity. In this study, the effects of different nitrogen sources, as well as the supplementation of an additional nitrogen source into RM and minimal medium (MM), on cell growth and glucose consumption of Z. mobilis were investigated to understand the uncoupled cell growth and glucose consumption. Our results indicated that nitrogen sources such as YE from different companies affected cell growth, glucose utilization, and ethanol production. We also quantified the concentrations of major ion elements in different nitrogen sources using the quantitative analytic approach of Inductively Coupled Plasma Optical Emission Spectroscopy (ICP-OES), and demonstrated that magnesium ion in the media affected cell growth, glucose consumption, and ethanol production. The effect of magnesium on gene expression was further investigated using RNA-Seq transcriptomics. Our results indicated that the lack of Mg2+ triggered stress responses, and the expression of genes involved in energy metabolism was reduced. Our work thus demonstrated that Mg2+concentration in nitrogen sources is essential for vigorous cell growth and ethanol fermentation, and the difference of Mg2+concentration in different YE is one of the major factors affecting the coupled cell growth, glucose consumption and ethanol fermentation in Z. mobilis. We also revealed that genes responsive for Mg2+ deficiency in the medium were majorly related to stress responses and energy conservation. The importance of magnesium on cell growth and ethanol fermentation suggests that metal ions should become one of the parameters for monitoring the quality of commercial nitrogen sources and optimizing microbial culture medium.