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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.

Solid-state fermentation, featured by water-saving, eco-friendly and high concentration product, is a promising technology in lignocellulosic ethanol industry. However, in solid-state fermentation system, large gas content inside the substrate directly leads to high oxygen partial pressure and inhibits ethanol fermentation. Z. mobilis can produce ethanol from glucose near the theoretical maximum value, but this ethanol yield would be greatly decreased by high oxygen partial pressure during solid-state fermentation. In this study, we applied N2 periodic pulsation process intensification (NPPPI) to ethanol solid-state fermentation, which displaced air with N2 and provided a proper anaerobic environment for Z. mobilis. Based on the water state distribution, the promotion effects of NPPPI on low solid loading and solid-state fermentation were analyzed to confirm the different degrees of oxygen inhibition in ethanol solid-state fermentation. During the simultaneous saccharification solid-state fermentation, the NPPPI group achieved 45.29% ethanol yield improvement and 30.38% concentration improvement compared with the control group. NPPPI also effectively decreased 58.47% of glycerol and 84.24% of acetic acid production and increased the biomass of Z. mobilis. By coupling the peristaltic enzymatic hydrolysis and fed-batch culture, NPPPI made the ethanol yield and concentration reach 80.11% and 55.06 g/L, respectively, in solid-state fermentation.

Assessment of viable biomass is challenging in bioprocesses involving complex media with distinct biomass and media particle populations. Biomass monitoring in these circumstances usually requires elaborate offline methods or sophisticated inline sensors. Reliable monitoring tools in an at-line capacity represent a promising alternative but are still scarce to date. In this study, a flow cytometry-based method for biomass monitoring in spent sulfite liquor medium as feedstock for second generation bioethanol production with yeast was developed. The method is capable of (i) yeast cell quantification against medium background, (ii) determination of yeast viability, and (iii) assessment of yeast physiology though morphological analysis of the budding division process. Thus, enhanced insight into physiology and morphology is provided which is not accessible through common online and offline biomass monitoring methods. To demonstrate the capabilities of this method, firstly, a continuous ethanol fermentation process of Saccharomyces cerevisiae with filtered and unfiltered spent sulfite liquor media was analyzed. Subsequently, at-line process monitoring of viability in a retentostat cultivation was conducted. The obtained information was used for a simple control based on addition of essential nutrients in relation to viability. Thereby, inter-dependencies between nutrient supply, physiology, and specific ethanol productivity that are essential for process design could be illuminated.

Ethanol is one of the most important platform chemicals that can be produced in a continuous biofilm reactor. The continuous system can be easily adapted to a biofilm reactor, which is a useful tool for ethanol production by microorganisms. In this study, two different media (the first medium: acid-pretreated/detoxified and glucose-enriched rice husk hydrolysate; the second medium: acid-pretreated/detoxified and xylose-enriched rice husk hydrolysate) were used for ethanol production in a continuous biofilm reactor. Both medium (1.5 L) were supplemented with 1% (w/v) yeast extract and 2% (w/v) peptone. The dilution rate for the first medium was between 0.02 and 0.12 h−1, while for the second medium, it was between 0.01 and 0.05 h−1. When the first medium was used for ethanol fermentation in a continuous system, maximum ethanol productivity of 0.418 g/L/h and maximum biomass productivity of 0.196 g/L/h were yielded at dilution rates of 0.08 and 0.10 h−1, respectively. As for the second medium for ethanol fermentation in a continuous system, their values were 0.083 and 0.079 g/L/h at dilution rates of 0.03 and 0.04 h−1, respectively. Additionally, the yield factors for biomass and ethanol (Y0X/S and Y0P/S) were also found to be 0.642 g X/g S and 0.49 g P/g S for the first medium and 0.254 g X/g S and 0.27 g P/g S for the second medium, respectively. In addition, although cost-effective ethanol production regarding energy cost and recovery time is desired, the use of the non-enriched sterile and enriched non-sterile media in a repeated-batch biofilm reactor caused low fermentation kinetics. Consequently, ethanol production was successfully performed by using Scheffersomyces stipitis in a continuous PCS-biofilm reactor including acid-pretreated/detoxified and glucose- or xylose-enriched rice husk hydrolysate, which gave higher ethanol concentration compared with subsequent ethanol fermentation in a repeated-batch biofilm reactor.

Background The growth of the cellulosic ethanol industry is currently impeded by high production costs. One possible solution is to improve the performance of fermentation itself, which has great potential to improve the economics of the entire production process. Here, we demonstrated significantly improved productivity through application of an advanced fermentation approach, named self-cycling fermentation (SCF), for cellulosic ethanol production. Results The flow rate of outlet gas from the fermenter was used as a real-time monitoring parameter to drive the cycling of the ethanol fermentation process. Then, long-term operation of SCF under anaerobic conditions was improved by the addition of ergosterol and fatty acids, which stabilized operation and reduced fermentation time. Finally, an automated SCF system was successfully operated for 21 cycles, with robust behavior and stable ethanol production. SCF maintained similar ethanol titers to batch operation while significantly reducing fermentation and down times. This led to significant improvements in ethanol volumetric productivity (the amount of ethanol produced by a cycle per working volume per cycle time)—ranging from 37.5 to 75.3%, depending on the cycle number, and in annual ethanol productivity (the amount of ethanol that can be produced each year at large scale)—reaching 75.8 ± 2.9%. Improved flocculation, with potential advantages for biomass removal and reduction in downstream costs, was also observed. Conclusion Our successful demonstration of SCF could help reduce production costs for the cellulosic ethanol industry through improved productivity and automated operation.

Thiswork aimed to describe fermentation characteristics of ethanol productivity by the hybrids of baker's yeast. In this study five parental isolates and 15 genotypic hybrids of Saccharomyces cerevisiae derived from three crosses were used for estimating fermentation characteristics during bioethanol production. At 0.02 concentration of sugarcane sub- products the hybrids of P1 x P4 increased turbidity and pH value above the mid–parent. In contrast, biomass formation showed decline production. However, at 0.04 and 0.06 sugarcane sub- products some of the hybrids increased significantly pH value above the mid–parent indicating that fermentation profile differs from one concentration of dissolved solids to another. However, ethanol productivity was affected mainly by low pH, high acidity, high alcohol content, high temperature and osmotic pressure. The low pH was the main stress factor on yeast during fermentation. Though, the hybrids derived from the cross between P2 x P5 significantly increased pH value above the mid – parent at 0.02 and 0.04 concentration of sugarcane sub–products. The yeast hybrids showed better turbidity and biomass formation at 0.02 concentration if compared with the mid–parents and the higher concentrations of sugarcane sub–products. Most hybrids derived from the cross between P3 x P5 showed significant pH values above the mid – parents at all concentrations of sugarcane sub- products. In addition, most hybrids showed significant turbidity above the mid – parents at 0.02 sub–products. However, insignificant differences were shown between yeast genotypes for turbidity and biomass formation at 0.04 and 0.06 concentration of dissolved solids.

Construction of acid-tolerant strains of Saccharomyces cerevisiae is required for various bioproduction processes. We previously isolated the gene IoGAS1 from multiple stress-tolerant Issatchenkia orientalis as a gene conferring sulfuric acid resistance in S. cerevisiae, but its acid tolerance was only investigated using sulfuric acid. Here, we evaluated the growth and ethanol fermentation ability of the IoGAS1-expressing S. cerevisiae strain, B4-IoGAS1, by using various acidic reagents. B4-IoGAS1 exhibited faster growth than the control strain, B4-CON, when cultured aerobically with sulfuric, hydrochloric, formic, acetic, and lactic acids at pH below 2.4. However, the growth of B4-IoGAS1 was suppressed at pH above 2.48, irrespective of the type of acid reagents. Furthermore, B4-IoGAS1 exhibited higher performance of ethanol fermentation than B4-CON under 250 mM lactic acid condition at pH 2.37. These results demonstrate that IoGAS1 could facilitate the aerobic growth and anaerobic ethanol production under different acidic stressed conditions.

In this paper, the effect of a variable temperature fermentation model was studied. We identified a model which considers substrate, product, and biomass inhibition, as well as an active and inactive cell phase of the yeast Saccharomyces cerevisiae. The model was evaluated using MATLAB at five different temperatures (28, 31, 34, 37, and 40°C). In the proposed model, we optimized the results through parameter fitting and compared it to the initial model which failed to simulate cell death. Through this work, we were able to improve the fit of the model considerably by 33% decrease, in the residual standard deviation. We also found that a smaller range of temperature performs better by maintaining the ethanol concentration and decreasing the temperature through the fermentation. Our model can show fermentation over a change in temperature over time and can show how fermentations will perform with temperature change.

In this study, introduction of a viable cell sensor and electronic nose into ethanol fermentation was investigated, which could be used in real-time and on-line monitoring of the amount of living cells and product content, respectively. Compared to the conventional off-line biomass determination, the capacitance value exhibited a completely consistent trend with colony forming units, indicating that the capacitance value could reflect the living cells in the fermentation broth. On the other hand, in comparison to the results of off-line determination by high-performance liquid chromatography, the ethanol concentration measured by electronic nose presented an excellent consistency, so as to realize the on-line monitoring during the whole process. On this basis, a dynamic feeding strategy of glucose guided by the changes of living cells and ethanol content was developed. And consequently, the ethanol concentration, productivity and yield were enhanced by 15.4%, 15.9% and 9.0%, respectively. The advanced sensors adopted herein to monitor the key parameters of ethanol fermentation process could be readily extended to an industrial scale and other similar fermentation processes.

Este trabajo tiene como objetivo principal obtener etanol mediante fermentación alcohólica aplicando procesos biotecnológicos; debido a su alto contenido en azucares se emplea el mosto de uva como sustrato y la levadura Saccharomyces cerevisiae como agente biológico para completar la fermentación, debido a que, este microorganismo ha sido probado en investigaciones anteriores mostrando una elevada tolerancia al etanol. Se tuvo en cuenta variables como el pH, tiempo de fermentación, los grados Brix, formación de biomasa, entre otros. Los rendimientos de etanol y biomasa fueron de 0,483 gg-1 y 0,069 gg-1, respectivamente, además se establecieron las dimensiones del reactor en la escala industrial asumiendo una producción de 30L de etanol. Palabras clave: etanol, fermentación, mosto de uva.

KOH (1%) and glycerol (20–40%) pretreatment of bamboo, rubber, elephant grass and nSiam weed were investigated with respect to biomass composition, enzymatic hydrolysis and ethanol fermentation. Delignification was higher under alkaline environment and increased considerably with temperature (25-121 °C). Lignin removal had a positive effect on biomass digestibility, sugars production and ethanol yield. The highest sugar and ethanol yields were observed for KOH pretreatment (121 °C, 1 h) on elephant grass which also recorded the highest delignification (67.5%) and consequently the lowest solids recovery (59.5%). The grass gave the highest yield on glucose (19 g/100 g raw material), xylose (8.2 g/100 g raw material) and ethanol (9.8 g/100 g raw material) under KOH pretreatment (121 °C, 1 h). The elephant grass has great potential to be used as a major energy crop in Africa in a biorefinery where production of sugars, ethanol and other valuable biochemicals are optimised.

The search for a balance between the energy-related challenges of the future and providing nutritional security has resulted in the development of a market for biofuels of successive generations. The larger their portion in biofuel production, the less the prices of agricultural products will increase. The use of algae, cyanobacteria and aquatic plants in the production of liquid fuels is an alternative. The aim of this study was to determine the effect of thermal hydrolysis on degradation of polysaccharides contained in biomass of cyanobacteria Arthrospira platensis and to assess the effectiveness of ethanol production from preconditioned biomass. The study is aimed at the selection of the most advantageous parameters of thermochemical hydrolysis to reach the experiment variant with the best effects, degree of polysaccharide degradation and effectiveness of alcohol fermentation. The experiment was divided into two stages; in stage I, the possibility of obtaining fermentable sugars by hydrothermal and chemical treatment of the substrate was tested. Stage II involved an assessment of the effectiveness of the pretreatment methods to produce bioethanol in alcohol fermentation. Yeast used in industrial ethanol production—Saccharomyces cerevisiae As4—was used in the alcohol fermentation. The results have shown that the temperature of 150 °C was the most beneficial for the process of thermohydrolysis, and the mash in the microwave-heated sample contained the highest concentration of alcohol (0.97 g/l), which is 98% more than in the control mash and 37% more than in the conventionally heated sample.

Lignocellulosic biomass is a sustainable feedstock for fuel ethanol production, but it is characterized by low mass and energy densities, and distributed production with relatively small scales is more suitable for cellulosic ethanol, which can better balance cost for the feedstock logistics. Lignocellulosic biomass is recalcitrant to degradation, and pretreatment is needed, but more efficient pretreatment technologies should be developed based on an in-depth understanding of its biosynthesis and regulation for engineering plant cell walls with less recalcitrance. Simultaneous saccharification and co-fermentation has been developed for cellulosic ethanol production, but the concept has been mistakenly defined, since the saccharification and co-fermentation are by no means simultaneous. Lignin is unreactive, which not only occupies reactor spaces during the enzymatic hydrolysis of the cellulose component and ethanol fermentation thereafter, but also requires extra mixing, making high solid loading difficult for lignocellulosic biomass and ethanol titers substantially compromised, which consequently increases energy consumption for ethanol distillation and stillage discharge, presenting another challenge for cellulosic ethanol production. Pentose sugars released from the hydrolysis of hemicelluloses are not fermentable with Saccharomyces cerevisiae used for ethanol production from sugar- and starch-based feedstocks, and engineering the brewing yeast and other ethanologenic species such as Zymomonas mobilis with pentose metabolism has been performed within the past decades. However strategies for the simultaneous co-fermentation of pentose and hexose sugars that have been pursued overwhelmingly for strain development might be modified for robust ethanol production. Finally, unit integration and system optimization are needed to maximize economic and environmental benefits for cellulosic ethanol production. In this article, we critically reviewed updated progress, and highlighted challenges and strategies for solutions.

Production of fuel ethanol is one of the possible ways to utilize crude glycerol, substantial amounts of which are produced by biodiesel industry. Earlier, we have described construction of the recombinant strains of methylotrophic thermotolerant yeast Ogataea polymorpha with simultaneous overexpression of the genes PDC1 and ADH1, which produced increased amounts of ethanol from glycerol. In this work, we have further improved these strains by overexpression of genes involved either in oxidative (through dihydroxyacetone) or phosphorylative (through glycerol-3-phosphate) pathway of glycerol catabolism, as well as heterologous gene coding for glycerol transporter FPS1 from Komagataella phaffii (formerly, Pichia pastoris). Obtained recombinant strains produced up to 10.7 g/L of ethanol (with ethanol productivity 30 mg/g of biomass/hr and yield 132 mg/g of consumed glycerol) from pure glycerol and up to 3.55 g/L of ethanol (with ethanol productivity 11.6 mg/g of biomass/hr and yield 72.3 mg/g of consumed glycerol) from crude glycerol as a carbon source, which is approximately 15 times more relative to that of the O. polymorpha wild-type strain and 2.2 more relative to the earlier constructed strain.

International audience; Yeast strains were isolated from sugar cane molasses (S1), dates (S2) and figs (S3) and the ethanol production was evaluated in batch condition. A comparison was made with the yeast Saccharomyces cerevisiae. The strains showed tolerant characteristics to stressful conditions like salinity and ethanol. The isolated strains produced ethanol; at 20 h of fermentation ethanol yields were 0.38-0.39 g.g-1 , and the productivities were almost 0.58 g.L-1. S. cerevisiae and S1 tolerated up to 14% (v/v) of ethanol; while interestingly the isolates S2 and S3 were highly tolerant, up to 20% (v/v) ethanol. Thus, S2 and S3 could serve as potential strains for ethanol fermentation, with 0.27 and 0.29 g.g-1 yield of ethanol in the presence of 1.37 mol.L-1 NaCl. These values were higher than the value obtained using the yeast of reference and S1 (0.16 g.g-1). Co-cultures of S2 and S3 enhanced the ethanol production, increasing the yield of ethanol by 12.5% compared with the single culture. The strains were identified as species S.cerevisiae, and S2 and S3 were very similar. For an application in the valorization of biomass such as green macro-algae, some assays were done on a synthetic model medium of hydrolysate of macro-algae and the strains S2 and S3 demonstrated excellent fermentative performances.

The aim of this work was to study ethanol fermentation properties of the robust mutant Candida glycerinogenes UG21 from non-detoxified lignocellulose hydrolysate. C. glycerinogenes UG21 with high tolerance to elevated temperature, acetic acid, and furfural was obtained and applied in lignocellulose-based ethanol production. C. glycerinogenes UG21 exhibited highly-efficient degradation ability to furfural. High levels of acetic acid and furfural inhibited cell growths and ethanol production of Saccharomyces cerevisiae ZWA46 and industrial Angel yeast but had a slight impact on biomass and ethanol titer of C. glycerinogenes UG21. Using non-detoxified sugarcane bagasse hydrolysate, C. glycerinogenes UG21 reached 1.24 g/L/h of ethanol productivity at 40 °C but ethanol production of S. cerevisiae ZWA46 and Angel yeast was inhibited. Further, C. glycerinogenes UG-21 exhibited 2.42-fold and 1.58-fold higher productivity than S. cerevisiae ZWA46 and Angel yeast under low-toxicity hydrolysate. Therefore, C. glycerinogenes UG-21 could be an excellent candidate for low-cost lignocelluloses ethanol production.

White-rot fungi are microorganisms capable of ethanol fermentation; however, the specific conditions activating ethanol fermentation are unclear in contrast to fermentation by yeasts. In this study, we investigated the conditions favoring ethanol fermentation by the white-rot fungus Phanerochaete sordida YK-624, which is able to produce ethanol from woody material. In aerobic stationary cultivation with various concentrations of glucose (0.8-33 g/l), the fungus produced ethanol in media containing an initial glucose concentration of 2.8 g/l or higher. The amount of glucose consumption, mycelial weight, and ethanol production on the second day of culture increased in a concentration-dependent manner at low glucose concentrations; however, these were saturated at high concentrations. Biomass yields (growth/glucose consumption) were decreased until the initial glucose concentration increased to 6.0 g/l, after which the biomass yields showed constant values at higher concentrations (12-33 g/l). On the other hand, ethanol yields increased with decreasing biomass yields. In short shaking cultivation using mycelial suspension, trace amounts of instantaneous aerobic ethanol production were observed with 1.1 and 2.1 g/l glucose, but the relative gene expression levels of key enzymes at the pyruvate branch point showed no significant differences between ethanol production and non-production conditions. From these experimental results, it appears that the white-rot fungus P. sordida YK-624 produces ethanol due to overflow in sugar metabolism under aerobic conditions, although P. sordida YK-624 prioritizes glucose utilization for respiratory growth.

A novel biomass pretreatment method, i.e. pressurized microwave hydrothermal treatment with inorganic salts, and the simultaneous saccharification and fermentation (SSF) using salt-tolerant cellulase, i.e. Meicelase, and salt-tolerant yeast, i.e. Saccharomyces cerevisiae BA11, were studied for the efficient ethanol production from sugarcane bagasse. The component analysis, enzymatic saccharification, and alcohol fermentation were carried out using pressurized microwave treated sugarcane bagasse with various inorganic salts. The pressurized microwave treatment using 2 wt% MgCl2 at a treatment temperature of 200 °C for a treatment time of 5 min followed by SSF without washing treatment of inorganic salts provided the maximum glucose and ethanol yields, i.e. 0.392 and 0.18 g/g-raw material, those corresponded to 99 and 90% of saccharification and ethanol conversion ratios, respectively.

Background Large-scale processing of lignocellulosics for glucose production generally relies on high temperature and acidic or alkaline conditions. However, extreme conditions produce chemical contaminants that complicate downstream processing. A method that mainly rely on mechanical and enzymatic reaction completely averts such problem and generates unmodified lignin. Products from this process could find novel applications in the chemicals, feed and food industry. But a large-scale system suitable for this purpose is yet to be developed. In this study we applied simultaneous enzymatic saccharification and communition (SESC) for the pre-treatment of a representative lignocellulosic biomass, cedar softwood, under both laboratory and large-scale conditions. Results Laboratory-scale comminution achieved a maximum saccharification efficiency of 80% at the optimum pH of 6. It was possible to recycle the supernatant to concentrate the glucose without affecting the efficiency. During the direct alcohol fermentation of SESC slurry, a high yield of ethanol was attained. The mild reaction conditions prevented the generation of undesired chemical inhibitors. Large-scale SESC treatment using a commercial beads mill system achieved a saccharification efficiency of 60% at an energy consumption of 50 MJ/kg biomass. Conclusion SESC is very promising for the mild and clean processing of lignocellulose to generate glucose and unmodified lignin in a large scale. Economic feasibility is highly dependent on its potential to generate high value natural products for energy, specialty chemicals, feed and food application.

Understanding the specific response of yeast cells to environmental stress factors is the starting point for selecting the conditions of adaptive culture in order to obtain a yeast line with increased resistance to a given stress factor. The aim of the study was to evaluate the specific cellular response of Saccharomyces cerevisiae strain Ethanol Red to stress caused by toxic by-products generated during the pretreatment of lignocellulose, such as levulinic acid, 5-hydroxymethylfurfural, furfural, ferulic acid, syringaldehyde and vanillin. The presence of 5-hydroxymethylfurfural at the highest analyzed concentration (5704.8 ± 249.3 mg/L) under aerobic conditions induced the overproduction of ergosterol and trehalose. On the other hand, under anaerobic conditions (during the alcoholic fermentation), a decrease in the biosynthesis of these environmental stress indicators was observed. The tested yeast strain was able to completely metabolize 5-hydroxymethylfurfural, furfural, syringaldehyde and vanillin, both under aerobic and anaerobic conditions. Yeast cells reacted to the presence of furan aldehydes by overproducing Hsp60 involved in the control of intracellular protein folding. The results may be helpful in optimizing the process parameters of second-generation ethanol production, in order to reduce the formation and toxic effects of fermentation inhibitors.

Made available in DSpace on 2021-06-25T11:10:24Z (GMT). No. of bitstreams: 0 Previous issue date: 2021-02-01 Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) The efficiency of unpressurized ozonolysis (O) and acid washing (A) was investigated as a simple pretreatment procedure to depolymerize sugarcane biomass for posterior enzymatic hydrolysis and alcoholic fermentation to improve cellulosic ethanol production. The fiber chemical compositional analyses showed that the ozonolysis reduces the lignin content by 25%. High-performance anion-exchange chromatography (HPAEC-PAD) analysis showed that the loss of glucose during treatment was less than 1.5%. After enzymatic hydrolysis, the maximum glucose and xylose yield (310 ± 2 and 122 ± 3 mg g−1, respectively) were achieved from the OA treatment sequence and the fermentation yield was three times more ethanol than the yield from untreated bagasse. This simple treatment may be an alternative for improving cellulosic ethanol production using a mild-operational strategy. São Paulo State University (UNESP) Institute of Biosciences Humanities and Exact Sciences São Paulo State University (UNESP) Institute of Biosciences Humanities and Exact Sciences FAPESP: 2013/06234-3 FAPESP: 2015/11588-4

O processo fermentativo para a produção de etanol brasileiro vem despertando interesse mundial, pois é realizado a partir da cana-de-açúcar, uma biomassa renovável de fermentação direta e, por meio do metabolismo das leveduras. Assim, este estudo visa comparar as características das biomassas cana-de-açucar e sorgo sacarino e avaliar o acúmulo de glycerol e a produção de etanol da levedura FT-858. Inicialmente, foi realizado um estudo bibliográfico qualitativo-exploratório acerca das características tecnológicas dos substratos sacarinos e, para a parte laboratorial um pré-inoculo com o meio líquido YPD 2%, esterilizado, no qual foi inoculada 0,10 g da levedura liofilizada FT-858 e incubada a 30 °C por 10h a 250rpm, que foi prontamente centrifugação e a biomassa inoculada no meio fermentativo a base de cana e de sorgo na concentração de 22 °Brix, que foi conduzido por reciclo celular a 30 e 40 °C a 250rpm e alíquotas de 4ml foram retiradas para as análises de glicerol intracelular utilizando o kit enzimático de triglicerídios e a concentração de etanol por cromatografia gasosa. Os substratos sacarinos apresentaram composição semelhante de açúcares, contudo, a sacarose foi superior ao encontrado no sorgo. O acúmulo de glicerol intracelular foi maior em caldo de cana em ambas as temperaturas e a concentração de etanol sofreu queda ao longo dos reciclos fermentativos. O acúmulo de glicerol e etanol foi inversamente proporcional, independente do substrato da sacarina. No entanto, os fatores de estresse associados causaram alterações no perfil de produção do metabólito da levedura, o que influenciou um maior acúmulo de glicerol. Assim, o sorgo sacarina tem grande potencial para ser utilizado na produção de etanol, uma vez que pode ser processado na mesma instalação industrial. Made available in DSpace on 2020-08-27T04:10:58Z (GMT). No. of bitstreams: 1 37068.pdf: 989234 bytes, checksum: 7c15d71101299d1412d45ee712418d48 (MD5) Previous issue date: 2020

Intelligent biorefinery design that addresses both the composition of the biomass feedstock as well as fermentation microorganisms could benefit from dedicated tools for computational simulation and computer-assisted optimization. Here we present the BioLego Vn2.0 framework, based on Microsoft Azure Cloud, which supports large-scale simulations of biomass serial fermentation processes by two different organisms. BioLego enables the simultaneous analysis of multiple fermentation scenarios and the comparison of fermentation potential of multiple feedstock compositions. Thanks to the effective use of cloud computing it further allows resource intensive analysis and exploration of media and organism modifications. We use BioLego to obtain biological and validation results, including (1) exploratory search for the optimal utilization of corn biomasses-corn cobs, corn fiber and corn stover-in fermentation biorefineries; (2) analysis of the possible effects of changes in the composition of K. alvarezi biomass on the ethanol production yield in an anaerobic two-step process (S. cerevisiae followed by E. coli); (3) analysis of the impact, on the estimated ethanol production yield, of knocking out single organism reactions either in one or in both organisms in an anaerobic two-step fermentation process of Ulva sp. into ethanol (S. cerevisiae followed by E. coli); and (4) comparison of several experimentally measured ethanol fermentation rates with the predictions of BioLego.

The objective of this study was to obtain two types of fuels, i.e., bioethanol and biogas, in a sequential combination of biochemical processes from lignocellulosic biomass (corn straw). Waste from the agricultural sector containing lignocellulose structures was used to obtain bioethanol, while the post-fermentation (cellulose stillage) residue obtained from ethanol fermentation was a raw material for the production of high-power biogas in the methane fermentation process. The studies on obtaining ethanol from lignocellulosic substrate were based on the simultaneous saccharification and fermentation (SSF) method, which is a simultaneous hydrolysis of enzymatic cellulose and fermentation of the obtained sugars. Saccharomyces cerevisiae (D-2) in the form of yeast cream was used for bioethanol production. The yeast strain D-2 originated from the collection of the Institute of Agricultural and Food Biotechnology. Volatile compounds identified in the distillates were measured using gas chromatography with flame ionization detector (GC-FID). CH4 and CO2 contained in the biogas were analyzed using a gas chromatograph in isothermal conditions, equipped with thermal conductivity detector (katharometer) with incandescent fiber. Our results show that simultaneous saccharification and fermentation enables production of bioethanol from agricultural residues with management of cellulose stillage in the methane fermentation process.

Modeling is gain importance to knowledge about kinetic-metabolic nature of fermentations. In this study, Modified Richards Model (MRM) was applied to model ethanol fermentation in a biofilm reactor...

The budding yeast Saccharomyces cerevisiae is an important microorganism for fermentation and the food industry. However, during production, S. cerevisiae commonly uses the ethanol fermentation pathway for glucose utilization if excess sugar is present, even in the presence of sufficient oxygen levels. This aerobic ethanol fermentation, referred to as the Crabtree effect, is one of the most significant reasons for low cell yield. To weaken the Crabtree effect in fed-batch and continuous culture, sugar flow should be limited. In addition, in continuous culture, the dilution rate must be reduced to avoid washing out cells. However, under such conditions, production speed might be sacrificed. It is difficult to solve this problem with the tradeoff between cell yield and production speed by using conventional tactics. However, a metabolomics approach may be an effective way to search for clues regarding metabolic modulation. Therefore, the purpose of this study was to reduce ethanol production in continuous culture of S. cerevisiae at a higher dilution rate through a metabolomics approach. We used a metabolomics analysis to identify metabolites that were drastically increased or decreased in continuous culture when the dilution rate shifted from biomass formation to ethanol fermentation. The individual addition of two of the selected metabolites, fumaric acid and malic acid, reduced ethanol production at a higher dilution rate. This result demonstrates the potential for using metabolomics approaches to identify metabolites that reduce ethanol production in continuous culture at high dilution rates.

In recent decades, there has been a tendency in the world to increase ethanol production significantly in order to solve energy problems, that is, to use it as a biofuel. The factors determining the production cost of targeted biotechnological products include the output of these products from the raw materials used. One of the modern and effective ways to intensify alcoholic fermentation and reduce the cost of fuel ethanol is yeast recirculation. The research objects were: raw material (sugarbeet molasses), molasses wort, yeast Saccharomyces cerevisiae of the strain M-5, fermented wash and its distillates. In the raw materials, intermediate products, and fermented wash, the techno-chemical parameters recommended by the current technology regulations for obtaining spirit from molasses have been determined. Acoholic fermentation was carried out in an industrial environment, in a battery of series-connected fermentors. Recirculation of yeast was carried out by separating it from the final stage of fermentation, concentrating it on the separator, and introducing it into the first fermentor. The experimental data obtained prove that for wort fermentation, it is effective to use yeast that recirculates in the anaerobic stage. It has been established that the alcohol-forming power of recycled yeast increases as the yeast adapts to the environment in which it has been staying for a long time. The yeast becomes more active biochemically, with more efficient metabolism. Its need for continuously cultured biomass is reduced, the share of aerobically assimiliated sugars decreases, and, consequently, the losses during yeast generation are fewer. At the same time, accelerating the initial period of anaerobic fermentation helps inhibit the biosynthesis of glycerol, the formation of which consumes the largest amount of sugar among all the secondary products. The parameters of molasses wort fermentation, with yeast biomass recirculating, have been determined in an industrial environment. It has been established that the alcohol output from the raw materials increases as the synthesis of secondary metabolic products weakens. The advantages of this fermentation method will be used in further studies, namely when fermenting molasses wort, with an increased concentration of dry matter, in order to reduce the specific heat energy consumption in production and to make it cheaper. The developed biotechnology of alcohol can be usefully employed to produce fuel ethanol, and increasing its production will contribute to Ukraine’s energy self-sufficience.

Xylose is a second-most abounded sugar after glucose in lignocellulosic hydrolysates and should be efficiently fermented for economically viable second-generation ethanol production. Despite significant progress in metabolic and evolutionary engineering, xylose fermentation rate of recombinant Saccharomyces cerevisiae remains lower than that for glucose. Our recent study demonstrated that peroxisome-deficient cells of yeast Ogataea polymorpha showed a decrease in ethanol production from xylose. In this work, we have studied the role of peroxisomes in xylose alcoholic fermentation in the engineered xylose-utilizing strain of S. cerevisiae. It was shown that peroxisome-less pex3Δ mutant possessed 1.5-fold decrease of ethanol production from xylose. We hypothesized that peroxisomal catalase Cta1 may have importance for hydrogen peroxide, the important component of reactive oxygen species, detoxification during xylose alcoholic fermentation. It was clearly shown that CTA1 deletion impaired ethanol production from xylose. It was found that enhancing the peroxisome population by modulation the peroxisomal biogenesis by overexpression of PEX34 activates xylose alcoholic fermentation.

Conversion of lignocellulosic biomass, such as corn stover (CS), to ethanol has encountered issues of inhibition from degradation products, low ethanol titer and low ethanol productivity. This work integrated CS into corn ethanol process for effective conversion. CS was pretreated using either dilute alkali or dilute acid pretreatment. The pretreated CS was enzymatically hydrolyzed and then mixed with liquefied corn for ethanol fermentation. Fermentation strains, substrate mixing ratios and fed-batch strategy were investigated. The mixture of alkali pretreated CS and corn at solids loadings of 10% and 20%, respectively, resulted in 92.30 g/L ethanol. Ethanol titer was further improved to 96.43 g/L with a fed-batch strategy. The mixture of dilute acid pretreated CS and corn achieved a better performance, leading to 104.9 g/L ethanol with 80.47% ethanol yield and a productivity as high as 2.19 g/L/h. This work demonstrated effective conversion of CS and corn together to ethanol.

To decrease alkaline dosage and increase the enzymatic saccharification of sugarcane bagasse (SB), dilute mixed alkali salts soaking (0.5% K2CO3, 0.068% K2SO3) in an autoclave at 100–125 °C for 10–60 min was attempted to pretreat SB. The increased pretreatment severity at elevated temperatures resulted in the increase of delignification and enzymatic saccharification. It was found the optimum pretreatment temperature and pretreatment time were 120 °C for 40 min. Furthermore, FTIR, XRD and SEM characterized the porosity, morphology, and crystallinity changes of the pretreated SB samples. After the enzymatic in situ saccharification of 50 g/L SB in dilute K2CO3/K2SO3 (0.35% K2CO3, 0.048% K2SO3) media for 72 h, the reducing sugars and glucose could be obtained at 32.94 and 20.18 g/L, respectively. Finally, the SB-hydrolysates containing 20 g/L glucose were used for ethanol fermentation in the presence of dilute alkali salts. After 48 h, the ethanol yield was 0.394 g ethanol/g glucose, which represents 77.25% of the theoretical yield. In conclusion, an effective process about the biomass pretreatment, enzymatic saccharification and ethanol fermentation was successfully demonstrated in “one-pot” media containing low loading of alkali salts K2CO3/K2SO3.

The waste seaweed from Gwangalli beach, Busan, Korea was utilized as biomass for ethanol production. Sagassum fulvellum (brown seaweed, Mojaban in Korean name) comprised 72% of the biomass. The optimal hyper thermal acid hydrolysis conditions were obtained as 8% slurry contents, 138 mM sulfuric acid, and 160oC of treatment temperature for 10 min with a low content of inhibitory compounds. To obtain more monosaccharides, enzymatic saccharification was carried out with Viscozyme L for 48 h. After pretreatment, 34 g/l of monosaccharides were obtained. Pichia stipitis and Pichia angophorae were selected as optimal co-fermentation yeasts to convert all of the monosaccharides in the hydrolysate to ethanol. Co-fermentation was carried out with various inoculum ratios of P. stipitis and P. angophorae. The maximum ethanol concentration of 16.0 g/l was produced using P. stipitis and P. angophorae in a 3:1 inoculum ratio, with an ethanol yield of 0.47 in 72 h. Ethanol fermentation using yeast co-culture may offer an efficient disposal method for waste seaweed while enhancing the utilization of monosaccharides and production of ethanol.

In the present investigation, an collagen fibre (CF), abundant natural biomass, was successfully grafted by polyethyleneimine (PEI). The resultant PEI grafted collagen fibre (CF-PEI) was employed as biocompatible and high cell loading support matrix for the immobilization of Saccharomyces cerevisiae cells. The as-prepared CF-PEI immobilized cells (CF-PEI-cell) exhibited high activity and stability for both batch and continuous fermentation. In batch fermentation, CF-PEI-cells showed enhanced stability as compared with other matrices supported cells, and produced an average ethanol concentration of 45.04 g/L with ethanol yield (YP/S) of 0.46 g/g and glucose conversion efficiency (η) of 90.4%. Continuous fermentation was operated stably in a down-flow trickling bed reactor charged with CF-PEI-cell for a total of 2 months. When the dilution rate was 0.16 1/h, the average ethanol productivity reached 7.18 g/(L h) with η of 88.94%. Further scanning electron microscopy observations confirmed that yeast cells can proliferate on the surface of CF-PEI during ethanol fermentation, which demonstrates that CF-PEI is indeed an ideal matrix for the immobilization of yeast cells.

The aim of the present study was to develop an inhibitor-tolerant strain of Scheffersomyces stipitis and establish an efficient ethanol fermentation process for cost-effective ethanol production from lignocellulosic biomass. By a strategy of three successive rounds of UV mutagenesis following adaptation, we isolated a S. stipitis mutant with improved tolerance against ethanol and inhibitors in the form of acetic acid, furfural and vanillin. The mutant strain exhibited excellent ethanol fermentation performance; both the xylose and glucose consumption rate and ethanol productivity were almost two times higher than the parental strain in batch fermentation. To overcome the issue of product inhibition and carbon catabolite repression (CCR) effect, the membrane integrated continuous fermentation system was employed. The maximum ethanol titer of 43.2 g l−1 and productivity of 2.16 g l−1 h−1 was achieved at a dilution rate of 0.05 h−1, higher than the relevant studies ever reported. These results suggested the novel process of cell recycling continuous fermentation using S. stipitis mutant has great potential for commercial ethanol production from lignocelluloses-based biomass.

Bioethanol as a substitute for traditional sources of energy, especially oil transport, is currently one of the most researched alternative motor fuels. Normally, bioethanol is produced from agricultural crops such as sugar cane or corn. However, this is counter-productive, because agriculture is primarily serving to ensure enough food for the people. It is therefore necessary to look for new production of appropriate non-food crops or find an added value to this process. Utilisation of contaminated water from metal industry could be one of them. Based on the hypothesis of reduction of some toxic metals with higher oxidation number is opening the possibility of using this wastewater in alcohol fermentation of any kind of biomass. In this study, hexavalent chromium Cr(VI) was used as a model contaminant in the process of aerobic fermentation of corn to bioethanol. To determine the reduction potential of glucose to Cr(VI), and to quantitatively determinate the glucose content after saccharification, UV/VIS spectrophotometry was used. As a method of qualitative determination of fermentation product, gas chromatography with mass detection was used. Infrared spectrometry was used for qualitative analyses of produced ethanol. Based on the established results shown in this paper, we can conclude that the presence of hexavalent chromium in the fermentation process does not have a significant negative impact, while offering the opportunity of using the industrial wastewaters for the production of bioethanol fuel.

The aim of this work was to develop a novel strategy for improving the vanillin tolerance and ethanol fermentation performances of Saccharomyces cerevisiae strains. Isogeneic diploid, triploid, and tetraploid S. cerevisiae strains were generated by genome duplication of haploid strain CEN.PK2-1C. Ploidy increments improved vanillin tolerance and diminished proliferation capability. Antimitotic drug methyl benzimidazol-2-ylcarbamate (MBC) was used to introduce chromosomal aberrations into the tetraploid S. cerevisiae strain. Interestingly, aneuploid mutants with DNA contents between triploid and tetraploid were more resistant to vanillin and showed faster ethanol fermentation rates than all euploid strains. The physiological characteristics of these mutants suggest that higher bioconversion capacities of vanillin and ergosterol contents might contribute to improved vanillin tolerance. This study demonstrates that genome duplication and MBC treatment is a powerful strategy to improve the vanillin tolerance of yeast strains.

A strategy for monitoring fermentation processes, specifically, simultaneous saccharification and fermentation (SSF) of corn mash, was developed. The strategy covered the development and use of first principles, semimechanistic and unstructured process model based on major kinetic phenomena, along with mass and energy balances. The model was then used as a reference model within an identification procedure capable of running on-line. The on-line identification procedure consists on updating the reference model through the estimation of corrective parameters for certain reaction rates using the most recent process measurements. The strategy makes use of standard laboratory measurements for sugars quantification and in situ temperature and liquid level data. The model, along with the on-line identification procedure, has been tested against real industrial data and have been able to accurately predict the main variables of operational interest, i.e., state variables and its dynamics, and key process indicators. The results demonstrate that the strategy is capable of monitoring, in real time, this complex industrial biomass fermentation. This new tool provides a great support for decision-making and opens a new range of opportunities for industrial optimization.

For practical applications of bioethanol, the uses of both highly concentrated biomass materials and their effective fermentation by yeasts are indispensable in order to produce ethanol at low costs. However, as the saccharified products of those biomass generally contain abundant sugars, the yeasts are affected by the compounds and are inclined to decrease their physiological activities. In the process of fermentation, ethanol is gradually produced by the yeasts in the culture; the concentrated metabolic product also damages itself, and inhibition of the fermentation frequently occurs. The application of yeasts with high fermentative activities under stress pressures such as sugars and ethanol is thus desired for bioethanol production. In this study, various types of high-fermentative yeasts under stress pressures were isolated mainly from coastal waters in Japan and characterized. All yeast strains with high fermentative activities under 20% v/v ethanol were found to be Saccharomyces cerevisiae. The HK21 strain isolated from Tokyo Bay and identified as S. cerevisiae had the highest fermentation activity under 30% w/v sorbitol and under 20% v/v ethanol, and it produced approx. 70 g/l (9% v/v) ethanol from the 15% w/v glucose solution at 25 oC within 5 days.

This paper opens up a new issue dealing with Luffa cylindrica (LC) lignocellulosic biomass recovery in order to produce 2G bioethanol. LC fibers are composed of three principal fractions, namely, α-cellulose (45.80% ± 1.3), hemicelluloses (20.76% ± 0.3), and lignins (13.15% ± 0.6). The optimization of LC fibers hydrothermal and diluted acid pretreatments duration and temperature were achieved through the cubic central composite experimental design CCD. The pretreatments optimization was monitored via the determination of reducing sugars. Then, the 2G bioethanol process feasibility was tested by means of three successive steps, namely, LC fibers hydrothermal pretreatment performed at 96°C during 54 minutes, enzymatic saccharification carried out by means of a commercial enzyme AP2, and the alcoholic fermentation fulfilled with Saccharomyces cerevisiae. LC fibers hydrothermal pretreatment liberated 33.55 g/kg of reducing sugars. Enzymatic hydrolysis allowed achieving 59.4 g/kg of reducing sugars. The conversion yield of reducing sugar to ethanol was 88.66%. After the distillation step, concentration of ethanol was 1.58% with a volumetric yield about 70%.

This work focuses on the performance of ten commercial Saccharomyces yeast strains in the batch alcoholic fermentation of sugars contained in selected industrial wastewaters from the soft drink industry. Fermentation has been applied successfully to treat these effluents prior to their disposal. Although many strains were investigated, similar behaviour was observed between all of the Saccharomyces strains tested. When media were inoculated with 2 g L−1 of yeast, all strains were able to completely consume the available sugars in less than 14 h. Thus, any of the strains studied in this work could be used in non-conventional wastewater treatment processes based on alcoholic fermentation. However, ethanol production varied between strains, and these differences could be significant from a production point of view. Saccharomyces bayanus produced the most ethanol, with a mean yield of 0.44 gethanol gsugarconsumed−1 and an ethanol specific production rate of 5.96 gethanol (L h)−1. As the assayed soft drinks wastewaters contain about 105 gsugar/L of fermentable sugars, the concentration of ethanol achieved after the fermentations process was 46.2 gethanol/L. A rigorous kinetic modelling methodology was used to model the Saccharomyces bayanus fermentation process. The kinetic model included coupled mass balances and a minimal number of parameters. A simple unstructured model based on the Andrews equation (substrate inhibition) was developed. This model satisfactorily described biomass growth, sugar consumption and bioethanol production. In addition to providing insights into the fermentative performance of potentially relevant strains, this work can facilitate the design of large-scale ethanol production processes that use wastewaters from the sugar-sweetened beverage industry as feedstock. Fil: Comelli, Raul Nicolas. Universidad Nacional del Litoral. Facultad de Ingeniería y Ciencias Hídricas; Argentina Fil: Seluy, Lisandro Gabriel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Desarrollo Tecnológico para la Industria Química. Universidad Nacional del Litoral. Instituto de Desarrollo Tecnológico para la Industria Química; Argentina Fil: Isla, Miguel Angel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Desarrollo Tecnológico para la Industria Química. Universidad Nacional del Litoral. Instituto de Desarrollo Tecnológico para la Industria Química; Argentina

Jerusalem artichoke (JA) is a potential energy crop for biorefinery due to its unique agronomic traits such as resistance to environmental stresses and high biomass yield in marginal lands. Although JA tubers have been explored for inulin extraction and biofuels production, there is little concern on its stalk (JAS). In this article, the pretreatment of JAS by alkaline hydrogen peroxide was optimized using the response surface methodology to improve sugars yield and reduce chemicals usage. Scanning electron microscopy, X-ray diffraction, and thermogravimetric analysis were applied to characterize the structures of the pretreated JAS to evaluate the effectiveness of the pretreatment. Furthermore, the feeding of the pretreated JAS and cellulase was performed for high solid uploading (up to 30%) to increase ethanol titer, and simultaneous saccharification and fermentation with 55.6g/L ethanol produced, 36.5% more than that produced through separate hydrolysis and fermentation, was validated to be more efficient.

Among the many types of lignocellulosic biomass pretreatment methods, the use of ionic liquids (ILs) is regarded as one of the most promising strategies. In this study, the effects of four kinds of ILs for pretreatment of lignocellulosic biomass such as bagasse, eucalyptus, and cedar were evaluated. In direct ethanol fermentation from biomass incorporated with ILs by cellulase-displaying yeast, 1-butyl-3-methylimidazolium acetate ([Bmim][OAc]) was the most effective IL. The ethanol production and yield from [Bmim][OAc]-pretreated bagasse reached 0.81 g/L and 73.4% of the theoretical yield after fermentation for 96 h. The results prove the initial concept, in which the direct fermentation from lignocellulosic biomass effectively promoted by the pretreatment with IL.

The cell growth and ethanol production from hydrolysates of various types were estimated from the volatile composition of lignocellulosic biomass by deep neural network (DNN) and the significant compositions estimated by asymmetric autoencoder-decoder (AAE). A six-layer DNN achieved good accuracy with learning and validation losses—0.033 and 0.507, respectively—and estimated overall time courses of yeast growth and ethanol fermentation. The AAE decoded the volatile compositions and represented the features of significant inhibitors via nonlinear dimensionality reduction, which was partly different from those using partial least squares regression reported previously. It revealed the significant features of hydrolysates for bioethanol production, which are lost in conventional approaches. The approach using DNN and AAE is, therefore, useful for bioethanol fermentation and other bioproductions using raw materials.