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The study aims to enhance ethanol production by Wickerhamomyces subpelliculosus ZE75 isolated from marine sediment. In addition, analyzing the kinetic parameters of ethanol production and optimization of the fermentation conditions was performed. The marine yeast isolate ZE75 was selected as the front runner ethanol-producer, with an ethanol yield of 89.77 gL -1 . ZE75 was identified relying on the phenotypic and genotypic characteristics of W. subpelliculosus. The genotypic characterization based on the Internal Transcribed Spacer (ITS) sequence was deposited in the GenBank database with the accession number OP715873. The maximum specific ethanol production rate (v max ) was 0.482 gg -1  h -1 at 175 gL -1 glucose concentration, with a high accuracy of R 2 0.95. The maximum growth specific rates (μ max ) were 0.141 h -1 obtained at 150 gL -1 glucose concentration with R 2 0.91. Optimization of the fermentation parameters such as pH and salinity has been achieved. The highest ethanol yield 0.5637 gg -1 was achieved in a 100% natural seawater-based medium. The maximum ethanol production of 104.04 gL -1 was achieved at pH 4.5 with a specific ethanol rate of 0.1669 gg -1  h -1 . The findings of the present study recommend the possibility of ethanol production from a seawater-based medium on a large scale using W. subpelliculosus ZE75., (© 2024. The Author(s).)

The soluble and fermentable carbohydrate contents was detected over 47% of glucose and fructose in Chinese Elaeagnus angustifolia fruit powder (EAF), being over 47 wt% sugar content more than that of grape. Ethanol was therefore fermented directly from EAF, and different submerged fermentation modes were comparatively employed to optimize ethanol harvest. The results indicated that glucose has certain competitive inhibition on fructose bio-utilization, as well as the EAF solid residue involved fermentation mode also hindered the fermented-ethanol titer. Pectinase addition and in situ hydrolysis seemed to assist somewhat the fermentation. The water-solute fermentation mode is preferable, and glucose and fructose components were completely consumed and converted to 80.96 g/L ethanol at 87.6% ethanol yield even under tannin and pectin inhibition. The fermentation result could provide some experimental data and an approach to not only new biomass resource explores of bioethanol and alcohol beverage production, but also the technological development on valorization commercials of EAF in global draught areas., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

NADH-dependent reductase enzyme catalyzes the phenolic aldehyde conversion and correspondingly improves the ethanol fermentability of the ethanologenic Zymomonas mobilis. This study constructed the transcriptional landscape of mono/dioxygenase genes in Z. mobilis ZM4 under the stress of the toxic phenolic aldehyde inhibitors of 4-hydroxybenzaldehyde, syringaldehyde, and vanillin. One specific dioxygenase encoding gene ZMO1721 was differentially expressed by 3.07-folds under the stress of 4-hydroxybenzaldehyde among the eleven mono/dioxygenase genes. The purified ZMO1721 shared 99.9% confidence and 48.0% identity with the oxidoreductase in Rhodoferax ferrireducens T118 was assayed and the NADH-dependent reduction activity was confirmed for phenolic aldehyde vanillin conversion. The ZMO1721 gene was then overexpressed in Z. mobilis ZM4 and the 4-hydroxybenzaldehyde conversion rate was accelerated. The cell growth, glucose consumption, and ethanol productivity of Z. mobilis ZM4 were also improved by ZMO1721 overexpression. The genes identified on improving phenolic aldehyde tolerance and ethanol fermentability in this study could be used as the synthetic biology tools for modification of ethanologenic strains., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

In this study, the effect of sonication on the fermentation process of a single-celled fungus was examined. During the experiment, Saccharomyces cerevisiae (S. cerevisiae) was used as the starting strain for ethanol fermentation (batch fermentation) in a 7.5 L automated fermentation tank. The fermentation tank connected with a six-frequency ultrasonic equipment. Non-sonication treatment was set up as the control. Sonication treatment with power density of 280 W/L and 48 h of treatment time were set up as trial groups for investigating the influence of different ultrasound frequency including 20, 23, 25, 28, 33 and 40 kHz on the changes in dry cell-weight, glucose consumption rate, and ethanol yield. The results showed that the dry cell-weight, glucose consumption rate, and ethanol content reached the best results under the ultrasonic condition of 28 kHz ultrasound frequency in comparison with other ultrasound frequency. The dry cell-weight and ethanol content of the 28 kHz ultrasonic treatment group increased by 17.30% and 30.79%, respectively in comparison with the control group The residual sugar content dropped to a lower level within 24 h, which was consistent with the change in ethanol production. Besides, the results found that the glucose consumption rate increased compared to the control. It indicated that ultrasound accelerated glucose consumption contributed to increase the rate of ethanol output. In order to explore the mechanism of sonication enhanced the content of ethanol output by S. cerevisiae, the morphology, permeability of S. cerevisiae and key enzyme activities of ethanol synthesis were investigated before and after sonication treatment. The results showed that after sonication treatment, the extracellular nucleic acid protein content and intracellular Ca 2+ concentration increased significantly. The morphology of S. cerevisiae was observed by SEM and found that the surface of the strain had wrinkles and depressions after ultrasonic treatment. furthermore after sonication treatment, the activities of three key enzymes which catalyze three irreversible reactions in glycolysis metabolism, namely, hexokinase, phosphofructokinase and pyruvate kinase increased by 59.02%, 109.05% and 87.27%, respectively. In a word, low-intensity ultrasound enhance the rate of ethanol output by S. cerevisiae might due to enhancing the growth and cell permeability of strains, and increasing the activities of three key enzymes of ethanol biosynthesis., (Copyright © 2021. Published by Elsevier B.V.)

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., (Copyright © 2020 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.)

In this study, the effect of soluble polysaccharides (SPs) derived from agricultural waste, rice straw, on fermentation-associated stresses (temperature and concentrations of glucose and ethanol) was investigated to achieve high-performance ethanol production. The increase in temperature and concentrations of glucose and ethanol significantly inhibited Saccharomyces cerevisiae growth and lowered ethanol fermentation efficiency. Flow cytometric assays indicated that SPs could alleviate membrane permeability damage caused by fermentation-associated stresses. Atomic force microscopy and transmission electron microscopy revealed that fermentation-associated stresses induced cell surface shrinkage, causing a decrease in the cell size, whereas SPs stimulated the formation of extracellular matrices (EMs), which made the cell surface smooth and the cell morphology regular. Cells with EMs induced by SPs could efficiently produce ethanol under severe stresses. As a result, the titer of ethanol in the fermentation with SPs was 1.40-fold (from 26.40 to 36.98 g/L) higher than that in the fermentation without SPs, suggesting the stress-alleviating effect of SPs on ethanol production.

The budding yeast Saccharomyces cerevisiae remains a central position among biofuel-producing organisms. However, the gene expression regulatory networks behind the ethanol fermentation is still not fully understood. Using a static fermentation model, we have examined the ethanol yields on biomass of deletion mutants for all yeast nonessential genes encoding transcription factors and their related proteins in the yeast genome. A total of 20 (about 10%) transcription factors are identified to be regulators of ethanol production during fermentation. These transcription factors are mainly involved in cell cycling, chromatin remodeling, transcription, stress response, protein synthesis and lipid synthesis. Our data provides a basis for further understanding mechanisms regulating ethanol production in budding yeast., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Trehalase catalyzes the hydrolysis of the non-reducing disaccharide trehalose. The highly active trehalase MthT from Myceliophthora thermophila was screened from the trehalase genes of six species of filamentous fungi. An ingenious multi-copy knock-in expression strategy mediated by the CRISPR/Cas9 tool and medium optimization were used to improve MthT production in Aspergillus niger, up to 1698.83 U/mL. The protein background was dramatically abated due to insertion. The recombinant MthT showed optimal activity at pH 5.5 and 60 °C, and exhibited prominent thermal stability between 50 and 60 °C under acid conditions (pH 4.5-6.5). The ethanol conversion rate (ethanol yield/total glucose) was significantly improved by addition of MthT (51.88%) compared with MthT absence (34.38%), using 30% starch saccharification liquid. The results of this study provided an effective strategy, established a convenient platform for heterologous expression in A. niger and showed a potential strategy to decrease production costs in industrial ethanol production.

A previously published genome-scale metabolic model namely iFF708 was modified to depict the metabolic flux distribution within Saccharomyces cerevisiae grown under a redox potential-controlled very high-gravity condition. The following modifications were made: electron transport chain (ETC) and oxidative phosphorylation, proton gradient and ATP transportation, and malate-aspartate shuttle. With these modifications, this model could describe the experimental data collected from the above-mentioned ethanol fermentation. As a result, the simulation unveiled that the P/O ratio is critical under microaerobic conditions and the malate-aspartate shuttle is inactivated due to the shortage of electron transport across mitochondria. In other words, the limited supply of oxygen suppresses the functionality of oxidative phosphorylation, tricarboxylic acid (TCA) cycle, and ETC. In terms of the glycolytic pathway, fluxes coming from glucose-6-phosphate and pyruvate nodes are insensitive to the changes of fermentation redox potential. As the initial glucose concentration is greater than 250 g/L, the interactive effect between the initial glucose concentration and redox potential level becomes noticeable., (© 2019 International Union of Biochemistry and Molecular Biology, Inc.)

Herbicides inhibiting either aromatic or branched-chain amino acid biosynthesis trigger similar physiological responses in plants, despite their different mechanism of action. Both types of herbicides are known to activate ethanol fermentation by inducing the expression of fermentative genes; however, the mechanism of such transcriptional regulation has not been investigated so far. In plants exposed to low-oxygen conditions, ethanol fermentation is transcriptionally controlled by the ethylene response factors-VII (ERF-VIIs), whose stability is controlled in an oxygen-dependent manner by the Cys-Arg branch of the N-degron pathway. In this study, we investigated the role of ERF-VIIs in the regulation of the ethanol fermentation pathway in herbicide-treated Arabidopsis plants grown under aerobic conditions. Our results demonstrate that these transcriptional regulators are stabilized in response to herbicide treatment and are required for ethanol fermentation in these conditions. We also observed that mutants with reduced fermentative potential exhibit higher sensitivity to herbicide treatments, thus revealing the existence of a mechanism that mimics oxygen deprivation to activate metabolic pathways that enhance herbicide tolerance. We speculate that this signaling pathway may represent a potential target in agriculture to affect tolerance to herbicides that inhibit amino acid biosynthesis., (© The Author(s) 2019. Published by Oxford University Press on behalf of the Society for Experimental Biology.)

Raman spectroscopy as a process analytical technology tool was implemented for the monitoring and control of ethanol fermentation carried out with Saccharomyces cerevisiae. The need for the optimization of bioprocesses such as ethanol production, to increase product yield, enhanced the development of control strategies. The control system developed by the authors utilized noninvasive Raman measurements to avoid possible sterilization problems. Real-time data analysis was applied using partial least squares regression (PLS) method. With the aid of spectral pretreatment and multivariate data analysis, the monitoring of glucose and ethanol concentration was successful during yeast fermentation with the prediction error of 4.42 g/L for glucose and 2.40 g/L for ethanol. By Raman spectroscopy-based feedback control, the glucose concentration was maintained at 100 g/L by the automatic feeding of concentrated glucose solution. The control of glucose concentration during fed-batch fermentation resulted in increased ethanol production. Ethanol yield of 86% was achieved compared to the batch fermentation when 75% yield was obtained. The results show that the use of Raman spectroscopy for the monitoring and control of yeast fermentation is a promising way to enhance process understanding and achieve consistently high production yield., (© 2019 The Authors. Biotechnology Progress published by Wiley Periodicals, Inc. on behalf of American Institute of Chemical Engineers.)

A thermotolerant ethanol fermenting yeast strain is a key requirement for effective ethanol production at high temperature. This work aimed to select a thermotolerant yeast producing a high ethanol concentration from molasses and increasing its ethanol production by mutagenesis. Saccharomyces cerevisiae DMKU 3-S087 was selected from 168 ethanol producing strains because it produced the highest ethanol concentration from molasses at 40 °C. Optimization of molasses broth composition was performed by the response surface method using Box-Behnken design. In molasses broth containing optimal total fermentable sugars (TFS) of 200 g/L and optimal (NH 4 ) 2 SO 4 of 1 g/L, with an initial pH of 5.5 by shaking flask cultivation at 40 °C ethanol, productivity and yield were 58.4 ± 0.24 g/L, 1.39 g/L/h and 0.29 g/g, respectively. Batch fermentation in a 5 L stirred-tank fermenter with 3 L optimized molasses broth adjusted to an initial pH of 5.5 and fermentation controlled at 40 °C and 300 rpm agitation resulted in 72.4 g/L ethanol, 1.21 g/L/h productivity and 0.36 g/g yield at 60 h. Strain DMKU 3-S087 improvement was performed by mutagenesis using ultraviolet radiation and ethyl methane sulfonate (EMS). Six EMS mutants produced higher ethanol (65.2 ± 0.48-73.0 ± 0.54 g/L) in molasses broth containing 200 g/L TFS and 1 g/L (NH 4 ) 2 SO 4 by shake flask fermentation at 37 °C than the wild type (59.8 ± 0.25 g/L). Among these mutants, only mutant S087E100-265 produced higher ethanol (62.5 ± 0.26 g/L) than the wild type (59.5 ± 0.02 g/L) at 40 °C. In addition, mutant S087E100-265 showed better tolerance to high sugar concentration, furfural, hydroxymethylfurfural and acetic acid than the wild type.

N-Acetyltransferase Mpr1 was originally discovered as an enzyme that detoxifies L-azetidine-2-carboxylate through its N-acetylation in the yeast Saccharomyces cerevisiae Σ1278b. Mpr1 protects yeast cells from oxidative stresses possibly by activating a novel L-arginine biosynthesis. We recently constructed a stable variant of Mpr1 (N203K) by a rational design based on the structure of the wild-type Mpr1 (WT). Here, we examined the effects of N203K on ethanol fermentation of the sake yeast S. cerevisiae strain lacking the MPR1 gene. When N203K was expressed in the diploid Japanese sake strain, its fermentation performance was improved compared to WT. In a laboratory-scale brewing, a sake strain expressing N203K produced more ethanol than WT. N203K also affected the contents of flavor compounds and organic acids. These results suggest that the stable Mpr1 variant contributes to the construction of new industrial yeast strains with improved fermentation ability and diversity of taste and flavor.

The present study investigated cold hydrolysis of cassava pulp (CP) and the use of cold hydrolysis with simultaneous saccharification and fermentation (SSF) for ethanol production. Cold hydrolysis of 100 g-CP/L at 50 °C for 2 h, followed by at 30 °C for 72 h resulted in the production of 71.5 ± 1.8 g/L of reducing sugar, with a yield of 0.72 g/g-CP. A mathematical model describing the cold hydrolysis process was subsequently developed. The model proved to be applicable for other cold hydrolysis systems with satisfactory results. The sequential process of cold hydrolysis at 50 °C for 2 h, followed by SSF at 30 °C for 72 h gave 27.4 g-ethanol/L, with a productivity of 0.37 g/(L h) and a fermentation efficiency of 57.58%. Based on the results, a bioconversion process for CP to ethanol was proposed. In this process, 1 kg of ethanol could be produced from 3.65 kg of CP without any nutrient supplementation., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Dekkera bruxellensis is continuously changing its status in fermentation processes, ranging from a contaminant or spoiling yeast to a microorganism with potential to produce metabolites of biotechnological interest. In spite of that, several major aspects of its physiology are still poorly understood. As an acetogenic yeast, minimal oxygen concentrations are able to drive glucose assimilation to oxidative metabolism, in order to produce biomass and acetate, with consequent low yield in ethanol. In the present study, we used disulfiram to inhibit acetaldehyde dehydrogenase activity to evaluate the influence of cytosolic acetate on cell metabolism. D. bruxellensis was more tolerant to disulfiram than Saccharomyces cerevisiae and the use of different carbon sources revealed that the former yeast might be able to export acetate (or acetyl-CoA) from mitochondria to cytoplasm. Fermentation assays showed that acetaldehyde dehydrogenase inhibition re-oriented yeast central metabolism to increase ethanol production and decrease biomass formation. However, glucose uptake was reduced, which ultimately represents economical loss to the fermentation process. This might be the major challenge for future metabolic engineering enterprises on this yeast., (© 2018 John Wiley & Sons, Ltd.)

The development of sustainable bioethanol fuel production from food waste has increasingly become an attractive topic. Food waste is recognized as the most available and costless feedstock. Therefore, ethanol production has been adopted as cost-efficient and an ecological way for FW disposal. This paper reviewed the microorganisms utilized for ethanol fermentation, the effect of enzymatic hydrolysis on ethanol concentration, optimization of accurate process parameters, and recycling of huge volumes of stillage for ethanol production towards reducing any incurred environmental burdens and minimizing the cost. The statistical tools which may enhance the process efficiency had been presented. Also, the perspective and the future development were introduced. All these aimed to fully utilize the food waste and also reduce the cost for side-product in this process; proper operation conditions and the control methods for stillage recycling were considered as the methods to improve ethanol fermentation from food waste.

Miscanthus×giganteus is a perennial rhizomatous grass which is used as a biofuel crop. Due to its high yields, low production costs, resistance to low temperatures, low soil requirements and, above all, high cellulose content, miscanthus can be a useful resource for ethanol production. The aim of this study was to determine the effect of two fertilization regimes (sewage sludge/mineral NPK) during miscanthus cultivation on the chemical composition of biomass, the content of major lignocellulosic factions and the effectiveness of miscanthus conversion to bioethanol. The results indicate that fertilization treatments influenced biomass yield and the content of major lignocellulosic fractions. Bioethanol production was higher when hydrolysis and fermentation processes were conducted separately than when saccharification and fermentation were conducted simultaneously. Ethanol production increased by 30% and 40% in response to sewage sludge and NPK (equivalent nitrogen content=160kgN/ha) fertilization, respectively, in comparison with unfertilized crops., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

In this study, it was found that the type of phenolic acids derived from rice straw was the major factor affecting ethanol fermentation by Pichia stipitis. The aim of this study was to investigate the inhibitory effect of phenolic acids on ethanol fermentation with rice straw. Different cellulases produced different ratios of free phenolic acids to soluble conjugated phenolic acids, resulting in different fermentation efficiencies. Free phenolic acids exhibited much higher inhibitory effect than conjugated phenolic acids. The flow cytometry results indicated that the damage to cell membranes was the primary mechanism of inhibition of ethanol fermentation by phenolic acids. The removal of free phenolic acids from the hydrolysates increased ethanol productivity by 2.0-fold, indicating that the free phenolic acids would be the major inhibitors formed during saccharification. The integrated process for ethanol and phenolic acids may constitute a new strategy for the production of low-cost ethanol., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

The aim of this study was to evaluate the influence of different starch liberation and saccharification methods on microbiological contamination of distillery mashes. Moreover, the effect of hop α-acid preparation for protection against microbial infections was assessed. The quality of agricultural distillates was also evaluated. When applying the pressureless liberation of starch (PLS) and malt as a source of amylolytic enzymes, the lactic acid bacteria count in the mashes increased several times during fermentation. The mashes obtained using the pressure-thermal method and malt enzymes revealed a similar pattern. Samples prepared using cereal malt exhibited higher concentrations of lactic and acetic acids, as compared to mashes prepared using enzymes of microbial origin. The use of hop α-acids led to the reduction of bacterial contamination in all tested mashes. As a result, fermentation of both mashes prepared with microbial origin enzyme preparations and with barley malt resulted in satisfactory efficiency and distillates with low concentrations of aldehydes., Competing Interests: The authors declare no conflict of interest.

High enzyme loading and low productivity are two major issues impeding low cost ethanol production from lignocellulosic biomass. This work applied rapid bioconversion with integrated recycle technology (RaBIT) and extractive ammonia (EA) pretreatment for conversion of corn stover (CS) to ethanol at high solids loading. Enzymes were recycled via recycling unhydrolyzed solids. Enzymatic hydrolysis with recycled enzymes and fermentation with recycled yeast cells were studied. Both enzymatic hydrolysis time and fermentation time were shortened to 24 h. Ethanol productivity was enhanced by two times and enzyme loading was reduced by 30%. Glucan and xylan conversions reached as high as 98% with an enzyme loading of as low as 8.4 mg protein per g glucan. The overall ethanol yield was 227 g ethanol/kg EA-CS (191 g ethanol/kg untreated CS). Biotechnol. Bioeng. 2017;114: 1713-1720. © 2017 Wiley Periodicals, Inc., (© 2017 Wiley Periodicals, Inc.)

Since both ethanol and butanol fermentations are urgently developed processes with the biofuel-demand increasing, performance comparison of aerobic ethanol fermentation and anerobic butanol fermentation in a continuous and closed-circulating fermentation (CCCF) system was necessary to achieve their fermentation characteristics and further optimize the fermentation process. Fermentation and pervaporation parameters including the average cell concentration, glucose consumption rate, cumulated production concentration, product flux, and separation factor of ethanol fermentation were 11.45 g/L, 3.70 g/L/h, 655.83 g/L, 378.5 g/m 2 /h, and 4.83, respectively, the corresponding parameters of butanol fermentation were 2.19 g/L, 0.61 g/L/h, 28.03 g/L, 58.56 g/m 2 /h, and 10.62, respectively. Profiles of fermentation and pervaporation parameters indicated that the intensity and efficiency of ethanol fermentation was higher than butanol fermentation, but the stability of butanol fermentation was superior to ethanol fermentation. Although the two fermentation processes had different features, the performance indicated the application prospect of both ethanol and butanol production by the CCCF system.

Lignocellulosic materials have been considered low-cost effective substrates for bioethanol production. However, lignocellulosic pretreatment releases toxic compounds such as 5-hydroxymethylfurfural (HMF) that is known to inhibit the yeast growth and ethanol production. In this work, flow cytometry was used to monitor the physiological response of the yeast Saccharomyces carlsbergensis ATCC 6269 in the presence of different initial HMF concentrations within the range of 0-15 g/L, in terms of cell membrane integrity, potential, and intracellular lipids. It was observed that the HMF presence affected more significantly the yeast growth than the ethanol production. At 15 g/L HMF, the yeast growth and fermentation ability were completely inhibited. The cell membrane integrity and potential decreased as the initial HMF concentration increased. At the end of the fermentation process with 10 g/L HMF, the yeast culture contained 45 % of cells with depolarized plasma membrane, 52 % of cells with permeabilized plasma membrane, and 53 % of cells with increasing reactive oxygen species (ROS) levels. Using the Nile Red stain, it was observed that intracellular polar lipids were more affected by the initial HMF concentration than the neutral lipids, probably due to the extensive membrane damage.

A total monosaccharide concentration of 37.8 g/L and 85.9% conversion from total fermentable monosaccharides of 44.0 g/L from 110 g dw/L Eucheuma denticulatum slurry were obtained by thermal acid hydrolysis and enzymatic saccharification. Subsequent adsorption treatment to remove 5-hydroxymethylfurfural (5-HMF) using 5% activated carbon and an adsorption time of 10 min were used to prevent a prolonged lag phase, reduced cell growth, and low ethanol production. The equilibrium adsorption capacity (q e ) of HMF (58.183 mg/g) showed high affinity to activated carbon comparing to those of galactose (2.466 mg/g) and glucose (2.474 mg/g). The efficiency of cell growth and ethanol production with activated carbon treatment was higher than that without activated carbon treatment. Fermentation using S. stipitis KCTC7228 produced a cell concentration of 3.58 g dw/L with Y X/S of 0.107, and an ethanol concentration of 15.8 g/L with Y P/S of 0.48 in 96 h.

Two different plants namely Phoenix canariensis and Opuntia ficus-indica were used as substrate for reducing sugar generation and ethanol production. Dilute acid, alkaline and steam explosion were used as pretreatment methods in order to depolymerize lignin and/or hemicellulose and recover cellulose. By using alkaline pretreatment with 2.5% NaOH 71.08% for P. canariensis and 74.61% for O. ficus-indica lignin removal and 81.84% for P. canariensis and 72.66% for O. ficus-indica cellulose recovery yields were obtained. Pretreated materials were hydrolyzed by cellulase with high efficiency (87.0% and 84.5% cellulose conversion yields for P. canariensis and O. ficus-indica) and used as substrate for fermentation. Maximum ethanol production of 15.75g/L and 14.71g/L were achieved from P. canariensis and O. ficus-indica respectively. Structural differences were observed by XRD, FTIR and SEM for untreated, pretreated, hydrolyzed and fermented samples and were highly correlated with compositional analysis results., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

An oxidative production process of xylonic acid using xylose in distillation stillage of cellulosic ethanol fermentation broth was designed, experimentally investigated, and evaluated. Dry dilute acid pretreated and biodetoxified corn stover was simultaneously saccharified and fermented into 59.80g/L of ethanol (no xylose utilization). 65.39g/L of xylose was obtained in the distillation stillage without any concentrating step after ethanol was distillated. Then the xylose was completely converted into 66.42g/L of xylonic acid by Gluconobacter oxydans. The rigorous Aspen Plus modeling shows that the wastewater generation and energy consumption was significantly reduced comparing to the previous xylonic acid production process using xylose in pretreatment liquid. This study provided a practical process option for xylonic acid production from lignocellulose feedstock with significant reduction of wastewater and energy consumption., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

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., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

One of the key processes in making beer is fermentation. In the fermentation process, brewer's yeast plays an essential role in both the production of ethanol and the flavor profile of beer. Therefore, the mechanism of ethanol fermentation by of brewer's yeast is attracting much attention. The high ethanol productivity of sake yeast has provided a good basis from which to investigate the factors that regulate the fermentation rates of brewer's yeast. Recent studies found that the elevated fermentation rate of sake Saccharomyces cerevisiae species is closely related to a defective transition from vegetative growth to the quiescent (G 0 ) state. In the present study, to clarify the relationship between the fermentation rate of brewer's yeast and entry into G 0 , we constructed two types of mutant of the bottom-fermenting brewer's yeast Saccharomyces pastorianus Weihenstephan 34/70: a RIM15 gene disruptant that was defective in entry into G 0 ; and a CLN3ΔPEST mutant, in which the G 1 cyclin Cln3p accumulated at high levels. Both strains exhibited higher fermentation rates under high-maltose medium or high-gravity wort conditions (20° Plato) as compared with the wild-type strain. Furthermore, G 1 arrest and/or G 0 entry were defective in both the RIM15 disruptant and the CLN3ΔPEST mutant as compared with the wild-type strain. Taken together, these results indicate that regulation of the G 0 /G 1 transition might govern the fermentation rate of bottom-fermenting brewer's yeast in high-gravity wort., (Copyright © 2016 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.)

Stillage reflux fermentation in food waste ethanol fermentation could reduce sewage discharge but exert a harmful effect because of side-product accumulation. In this study, regulation methods based on metabolic regulation and side-product alleviation were conducted. Result demonstrated that controlling the proper oxidation-reduction potential value (-150mV to -250mV) could reduce the harmful effect, improve ethanol yield by 21%, and reduce fermentation time by 20%. The methods of adding calcium carbonate to adjust the accumulated lactic acid showed that ethanol yield increased by 17.3%, and fermentation time decreased by 20%. The accumulated glyceal also shows that these two methods can reduce the harmful effect. Fermentation time lasted for seven times without effect, and metabolic regulation had a better effect than side-product regulation., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

In this study, a Saccharomyces cerevisiae recombinant strain 14 was constructed through genome shuffling method by transferring the whole genomic DNA of Candida intermedia strain 23 into a thermo-tolerant S. cerevisiae strain. The recombinant strain 14 combined the good natures of both parent strains that efficiently produced ethanol from glucose and single cell protein from xylose with 54.6% crude protein and all essential amino acids except cysteine at 35°C. Importantly, the recombinant strain 14 produced 64.07g/L ethanol from 25%(w/v) NaOH-pretreated and washed corn stover with the ethanol yield of 0.26g/g total stover by fed-batch simultaneous saccharification and fermentation and produced 66.50g/L dry cell mass subsequently from the residual hydrolysate and ethanol. Therefore, this study represents a feasible method to comprehensively utilize hexose and pentose in lignocellulosic materials., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

The use of field beans, a non-food leguminous crop, was studied for ethanol, feed components and fungal biomass production. The seeds were hydrolyzed using enzymes or with combination of acid (H3PO4) and alkaline (Ca(OH)2) pretreatment and enzymatic hydrolysis. Fermentation by Saccharomyces cerevisiae, with or without removal of suspended solids, yielded 38.3-42.5gL(-1) ethanol (71.3-79.2% efficiency). The filtration residues contained ca. 247-326gkg(-1) crude protein, 10.6-15.5% acid detergent fiber and 19.9-29.1% neutral detergent fiber. They were enriched in phenolics (by up to 93.4%) and depleted in condensed tannin (by up to 59.3%) in comparison to the raw material. The thin stillages were used for cultivation of edible fungus Neurospora intermedia which produced 8.5-15.9gL(-1) ethanol and 4.8-16.2gL(-1) biomass containing over 62% protein. The mass balances showed that fermentation of unfiltered mashes was more efficient yielding up to 195.9gkg(-1) ethanol and 84.4% of protein recovery., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Corn stover (CS) was hydrothermally pretreated using CH3COOH (0.3 %, v/v), and subsequently its ability to be utilized for conversion to ethanol at high-solids content was investigated. Pretreatment conditions were optimized employing a response surface methodology (RSM) with temperature and duration as independent variables. Pretreated CS underwent a liquefaction/saccharification step at a custom designed free-fall mixer at 50 °C for either 12 or 24 h using an enzyme loading of 9 mg/g dry matter (DM) at 24 % (w/w) DM. Simultaneous enzymatic saccharification and fermentation (SSF) of liquefacted corn stover resulted in high ethanol concentration (up to 36.8 g/L), with liquefaction duration having a negligible effect. The threshold of ethanol concentration of 4 % (w/w), which is required to reduce the cost of ethanol distillation, was surpassed by the addition of extra enzymes at the start up of SSF achieving this way ethanol titer of 41.5 g/L.

In this paper, genetic algorithm was used to investigate mathematical modeling of ethanol fermentation in a continuous conventional bioreactor (CCBR) and a continuous membrane bioreactor (CMBR) by ethanol permselective polydimethylsiloxane (PDMS) membrane. A lab scale CMBR with medium glucose concentration of 100gL(-1) and Saccharomyces cerevisiae microorganism was designed and fabricated. At dilution rate of 0.14h(-1), maximum specific cell growth rate and productivity of 0.27h(-1) and 6.49gL(-1)h(-1) were respectively found in CMBR. However, at very high dilution rate, the performance of CMBR was quite similar to conventional fermentation on account of insufficient incubation time. In both systems, genetic algorithm modeling of cell growth, ethanol production and glucose concentration were conducted based on Monod and Moser kinetic models during each retention time at unsteady condition. The results showed that Moser kinetic model was more satisfactory and desirable than Monod model., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Hyper thermal (HT) acid hydrolysis of Kappaphycus alvarezii, a red seaweed, was optimized to 12% (w/v) seaweed slurry content, 180mM H2SO4 at 140°C for 5min. The maximum monosaccharide concentration of 38.3g/L and 66.7% conversion from total fermentable monosaccharides of 57.6g/L with 120gdw/L K. alvarezii slurry were obtained from HT acid hydrolysis and enzymatic saccharification. HT acid hydrolysis at a severity factor of 0.78 efficiently converted the carbohydrates of seaweed to monosaccharides and produced a low concentration of inhibitory compounds. The levels of ethanol production by separate hydrolysis and fermentation with non-adapted and adapted Kluyveromyces marxianus to high concentration of galactose were 6.1g/L with ethanol yield (YEtOH) of 0.19 at 84h and 16.0g/L with YEtOH of 0.42 at 72h, respectively. Development of the HT acid hydrolysis process and adapted yeast could enhance the overall ethanol fermentation yields of K. alvarezii seaweed., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Raw materials and pollution control are key issues for the ethanol fermentation industry. To address these concerns, food waste was selected as fermentation substrate, and stillage reflux was carried out in this study. Reflux was used seven times during fermentation. Corresponding ethanol and reducing sugar were detected. Accumulation of by-products, such as organic acid, sodium chloride, and glycerol, was investigated. Lactic acid was observed to accumulate up to 120g/L, and sodium chloride reached 0.14mol/L. Other by-products did not accumulate. The first five cycles of reflux increased ethanol concentration, which prolonged fermentation time. Further increases in reflux time negatively influenced ethanol fermentation. Single-factor analysis with lactic acid and sodium chloride demonstrated that both factors affected ethanol fermentation, but lactic acid induced more effects., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Saccharomyces cerevisiae is the most widely used fermentation organism for ethanol production. However, the gene expression regulatory networks behind the ethanol fermentation are still not fully understood. Using a static fermentation model, we examined the ethanol yields on biomass of deletion mutants for 77 yeast genes encoding nonessential transcription factors, and found that deletion mutants for ACE2 and SWI5 showed dramatically increased ethanol yields. Overexpression of ACE2 or SWI5 in wild type cells reduced their ethanol yields. Furthermore, among the 34 target genes regulated by Ace2 and Swi5, deletion of CTS1,RPS4a,SIC1,EGT2,DSE2, or SCP160 led to increased ethanol yields, with the former two showing higher effects. Overexpression of CTS1 or RPS4a in both ace2/ace2 and swi5/swi5 mutants reduced their ethanol yields. In contrast, deletion of MCR1 or HO significantly decreased ethanol yields, with the former one showing the highest effect. Therefore, Ace2 and Swi5 are two negative regulators of ethanol yield during static fermentation of yeast cells, and both CTS1 and RPS4a are major effectors mediating these two transcription factors in regulating ethanol production., (© FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Pretreatment with ozone combined with aqueous ammonia was used to recover residual organic carbon from recalcitrant solid digestate for ethanol production after anaerobic digestion (AD) of rice straw. Methane yield of AD at mesophilic and thermophilic conditions, and ethanol production of solid digestate were investigated. The results showed that the methane yield at thermophilic temperature was 72.2% higher than that at mesophilic temperature under the same conditions of 24days and 17% solid concentration. And also the ethanol production efficiency of solid digestate after thermophilic process was 24.3% higher than that of solid digestate after mesophilic process. In this study, the optimal conditions for integrated methane and ethanol processes were determined as 55°C, 17% solid concentration and 24days. 58.6% of glucose conversion, 142.8g/kg of methane yield and 65.2g/kg of ethanol yield were achieved, and the highest net energy balance was calculated as 6416kJ/kg., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Dried distillers grains with solubles (DDGS), a co-product of corn ethanol production in the dry-grind process, was pretreated by soaking in aqueous ammonia (SAA) using a 15 % w/w NH4OH solution at a solid/liquid ratio of 1:10. The effect of pretreatment on subsequent enzymatic hydrolysis was studied at two temperatures (40 and 60 °C) and four reaction times (6, 12, 24, and 48 h). Highest glucose yield of 91 % theoretical was obtained for the DDGS pretreated at 60 °C and 24 h. The solubilized hemicellulose in the liquid fraction was further hydrolyzed with dilute H2SO4 to generate fermentable monomeric sugars. The conditions of acid hydrolysis included 1 and 4 wt% acid, 60 and 120 °C, and 0.5 and 1 h. Highest yields of xylose and arabinose were obtained at 4 wt% acid, 120 °C, and 1 h. The fermentability of the hydrolysate obtained by enzymatic hydrolysis of the SAA-pretreated DDGS was demonstrated in ethanol fermentation by Saccharomyces cerevisiae. The fermentability of the hydrolysate obtained by consecutive enzymatic and dilute acid hydrolysis was demonstrated using a succinic acid-producing microorganism, strain Escherichia coli AFP184. Under the fermentation conditions, complete utilization of glucose and arabinose was observed, whereas only 47 % of xylose was used. The succinic acid yield was 0.60 g/g total sugar consumed.

In this study, the effects of different parts of corn stalk, including stem, leaf, flower, cob and husk on second generation ethanol production were evaluated. FTIR, XRD and SEM were performed to investigate the effect of dilute acid pretreatment. The bagasse obtained after pretreatment were further hydrolyzed by cellulase and used as the substrate for ethanol fermentation. As results, hemicelluloses fractions in different parts of corn stalk were dramatically removed and the solid fractions showed vivid compositions and crystallinities. Compared with other parts of corn stalk, the cob had higher sugar content and better enzymatic digestibility. The highest glucose yield of 94.2% and ethanol production of 24.0 g L(-1) were achieved when the cob was used as feedstock, while the glucose yield and the ethanol production were only 86.0% and 17.1 g L(-1) in the case of flower., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

This study proposes a novel alternative for utilization of whey permeate, a by-product stream from the dairy industry, in wheat fermentation for ethanol production using Saccharomyces cerevisiae. Whey permeates were hydrolyzed using enzymes to release fermentable sugars. Hydrolyzed whey permeates were integrated into wheat fermentation as a co-substrate or to partially replace process water. Cold starch hydrolysis-based simultaneous saccharification and fermentation was done as per the current industrial protocol for commercial wheat-to-ethanol production. Ethanol production was not affected; ethanol yield efficiency did not change when up to 10% of process water was replaced. Lactic acid bacteria in whey permeate did not negatively affect the co-fermentation or reduce ethanol yield. Whey permeate could be effectively stored for up to 4 wk at 4 °C with little change in lactose and lactic acid content. Considering the global abundance and nutrient value of whey permeate, the proposed strategy could improve economics of the dairy and biofuel sectors, and reduce environmental pollution. Furthermore, our research may be applied to fermentation strategies designed to produce value-added products other than ethanol., (Copyright © 2016 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.)

Genomic structural variation (GSV) is a ubiquitous phenomenon observed in the genomes of Saccharomyces cerevisiae strains with different genetic backgrounds; however, the physiological and phenotypic effects of GSV are not well understood. Here, we first revealed the genetic characteristics of a widely used industrial S. cerevisiae strain, ZTW1, by whole genome sequencing. ZTW1 was identified as an aneuploidy strain and a large-scale GSV was observed in the ZTW1 genome compared with the genome of a diploid strain YJS329. These GSV events led to copy number variations (CNVs) in many chromosomal segments as well as one whole chromosome in the ZTW1 genome. Changes in the DNA dosage of certain functional genes directly affected their expression levels and the resultant ZTW1 phenotypes. Moreover, CNVs of large chromosomal regions triggered an aneuploidy stress in ZTW1. This stress decreased the proliferation ability and tolerance of ZTW1 to various stresses, while aneuploidy response stress may also provide some benefits to the fermentation performance of the yeast, including increased fermentation rates and decreased byproduct generation. This work reveals genomic characters of the bioethanol S. cerevisiae strain ZTW1 and suggests that GSV is an important kind of mutation that changes the traits of industrial S. cerevisiae strains., (© FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

This study reported a new solution of lignocellulose feedstock storage based on the distributed pretreatment concept. The dry dilute sulfuric acid pretreatment (DDAP) was conducted on corn stover feedstock, instead of ammonia fiber explosion pretreatment. Then the dry dilute acid pretreated corn stover was stored for three months during summer season with high temperature and humidity. No negative aspects were found on the physical property, composition, hydrolysis yield and ethanol fermentability of the long term stored pretreated corn stover, plus the additional merits including no chemicals recovery operation, anti-microbial contaminant environment from stronger acid and inhibitor contents, as well as the mild and slow hydrolysis in the storage. The new pretreatment method expanded the distributed pretreatment concept of feedstock storage with potential for practical application., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

This study evaluated ethanol fermentation and its correlation with glutathione (GSH) synthesis under various cadmium-conditions with different metal ions and nitrogen sources. We found that corn steep liquor (CSL) and yeast extract have differential roles to play in GSH accumulation in cell even though both of them could alleviate the inhibition by cadmium. The different GSH accumulation in cell resulted from the different contents of metal ions in CSL and yeast extract. Intracellular GSH decreased with increasing calcium concentrations, and high calcium concentrations rendered the yeast more tolerant to cadmium stress than the nitrogen sources did. When the mole ratio of calcium to cadmium was 100:1, yeast tolerated 1000 µmol/L cadmium with no decrease in efficiency in ethanol production. As a result, the use of calcium allowed a significant saving of high-cost nutrient yeast extract with an efficient ethanol production, making the bioconversion of cadmium-containing biomass into ethanol possible., (© 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

The polydimethylsiloxane (PDMS) membrane, prepared in water phase, was investigated in separation ethanol from model ethanol/water mixture and fermentation-pervaporation integrated process. Results showed that the PDMS membrane could effectively separate ethanol from model solution. When integrated with batch ethanol fermentation, the ethanol productivity was enhanced compared with conventional process. Fed-batch and continuous ethanol fermentation with pervaporation were also performed and studied. 396.2-663.7g/m(2)h and 332.4-548.1g/m(2)h of total flux with separation factor of 8.6-11.7 and 8-11.6, were generated in the fed-batch and continuous fermentation with pervaporation scenario, respectively. At the same time, high titre ethanol production of ∼417.2g/L and ∼446.3g/L were also achieved on the permeate side of membrane in the two scenarios, respectively. The integrated process was environmental friendly and energy saving, and has a promising perspective in long-terms operation., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Corncob residue as the lignocellulosic biomass accumulated phenolic compounds generated from xylitol production industry. For utilization of this biomass, Zymomonas mobilis ZM4 was tested as the ethanol fermenting strain and presented a better performance of cell growth (2.8 × 10(8)  CFU/mL) and ethanol fermentability (54.42 g/L) in the simultaneous saccharification and fermentation (SSF) than the typical robust strain Saccharomyces cerevisiae DQ1 (cell growth of 2.9 × 10(7)  CFU/mL, ethanol titer of 48.6 g/L). The physiological response of Z. mobilis ZM4 to the twelve typical phenolic compounds derived from lignocellulose was assayed and compared with that of S. cerevisiae DQ1. Z. mobilis ZM4 showed nearly the same tolerance to the phenolic aldehydes with S. cerevisiae DQ1, but the stronger tolerance to the phenolic acids existing in corncob residue (2-furoic acid, p-hydroxybenzoic acid, p-coumaric acid, vanillic acid, ferulic acid, and syringic acid). The tolerance mechanism of Z. mobilis was investigated in terms of inhibitor degradation, cell morphology and membrane permeability under the stress of phenolics using GC-MS, scanning and transmission electron microscopies (SEM and TEM), as well as fluorescent probes. The results reveal that Z. mobilis ZM4 has the capability for in situ detoxification of phenolic aldehydes, and the lipopolysaccharide aggregation on the cell outer membrane of Z. mobilis ZM4 provided the permeable barrier to the attack of phenolic acids., (© 2015 Wiley Periodicals, Inc.)

This paper reports on processing options for the conversion of feedlot cattle manures into composite sugars for ethanol fermentation. Small-scale anaerobic digestion trials revealed that the process significantly reduces the content of glucan and xylan (ca. 70%) without effecting lignin. Moreover, anaerobic digestate (AD) fibres were poor substrates for cellulase (Cellic® CTec 2) saccharification, generating a maximum combined sugar yield of ca. 12% per original dry weight. Dilute acid pretreatment and enzyme saccharification of raw manures significantly improved total sugar recoveries, totalling 264 mg/g (79% theoretical). This was attained when manures were pretreated with 2.5% H2SO4 for 90 min at 121°C and saccharified with 50 FPU CTec 2/g glucan. Saccharomyces cerevisiae efficiently fermented crude hydrolysates within 6 h, yielding 7.3 g/L ethanol, representing glucose to ethanol conversion rate of 70%. With further developments (i.e., fermentation of xylose), this process could deliver greater yields, reinforcing its potential as a biofuel feedstock., (Crown Copyright © 2015. Published by Elsevier Ltd. All rights reserved.)

While softwoods represent a potential feedstock for second generation ethanol production, compounds present in their hydrolysates can inhibit fermentation. In this study, a novel Design of Experiments (DoE) approach was used to identify significant inhibitory effects on Saccharomyces cerevisiae D5A for the purpose of guiding kinetic model development. Although acetic acid, furfural and 5-hydroxymethyl furfural (HMF) were present at potentially inhibitory levels, initial factorial experiments only identified ethanol as a significant rate inhibitor. It was hypothesized that high ethanol levels masked the effects of other inhibitors, and a subsequent factorial design without ethanol found significant effects for all other compounds. When these non-ethanol effects were accounted for in the kinetic model, R¯(2) was significantly improved over an ethanol-inhibition only model (R¯(2)=0.80 vs. 0.76). In conclusion, when ethanol masking effects are removed, DoE is a valuable tool to identify significant non-ethanol inhibitors and guide kinetic model development., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Unstructured kinetic models were proposed to describe the principal kinetics involved in ethanol fermentation in a continuous and closed-circulating fermentation (CCCF) process with a pervaporation membrane bioreactor. After ethanol was removed in situ from the broth by the membrane pervaporation, the secondary metabolites accumulated in the broth became the inhibitors to cell growth. The cell death rate related to the deterioration of the culture environment was described as a function of the cell concentration and fermentation time. In CCCF process, 609.8 g L(-1) and 750.1 g L(-1) of ethanol production were obtained in the first run and second run, respectively. The modified Gompertz model, correlating the ethanol production with the fermentation period, could be used to describe the ethanol production during CCCF process. The fitting results by the models showed good agreement with the experimental data. These models could be employed for the CCCF process technology development for ethanol fermentation., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Commercialization of lignocellulosic ethanol fermentation requires its high titer, but the reactive oxygen species (ROS) accumulation during the bioprocess damaged the cells and compromised this goal. To improve the cellular anti-oxidative activity during non-detoxified corncob residue hydrolysate fermentation, seed cells were prepared to possess a higher level of intracellular biotin pool (IBP), which facilitated the biosyntheses of catalase and porphyrin. As a result, the catalase activity increased by 1.3-folds compared to control while the ROS level reduced by 50%. Cell viability in high-IBP cells was 1.7-folds of control and the final ethanol titer increased from 31.2 to 41.8 g L(-1) in batch fermentation. The high-IBP cells were further used for repeated-batch fermentation in the non-detoxified lignocellulosic hydrolysate, and the highest titer and average productivity of ethanol reached 63.7 g L(-1) and 1.2 g L(-1)h(-1). The results were favorable to future industrial application of this lignocellulosic bioethanol process., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

The production of raw starch-degrading amylases by recombinant Saccharomyces cerevisiae provides opportunities for the direct hydrolysis and fermentation of raw starch to ethanol without cooking or exogenous enzyme addition. Such a consolidated bioprocess (CBP) for raw starch fermentation will substantially reduce costs associated with energy usage and commercial granular starch hydrolyzing (GSH) enzymes. The core purpose of this review is to provide comprehensive insight into the physiological impact of recombinant amylase production on the ethanol-producing yeast. Key production parameters, based on outcomes from modifications to the yeast genome and levels of amylase production, were compared to key benchmark data. In turn, these outcomes are of significance from a process point of view to highlight shortcomings in the current state of the art of raw starch fermentation yeast compared to a set of industrial standards. Therefore, this study provides an integrated critical assessment of physiology, genetics and process aspects of recombinant raw starch fermenting yeast in relation to presently used technology. Various approaches to strain development were compared on a common basis of quantitative performance measures, including the extent of hydrolysis, fermentation-hydrolysis yield and productivity. Key findings showed that levels of α-amylase required for raw starch hydrolysis far exceeded enzyme levels for soluble starch hydrolysis, pointing to a pre-requisite for excess α-amylase compared to glucoamylase for efficient raw starch hydrolysis. However, the physiological limitations of amylase production by yeast, requiring high biomass concentrations and long cultivation periods for sufficient enzyme accumulation under anaerobic conditions, remained a substantial challenge. Accordingly, the fermentation performance of the recombinant S. cerevisiae strains reviewed in this study could not match the performance of conventional starch fermentation processes, based either on starch cooking and/or exogenous amylase enzyme addition. As an alternative strategy, the addition of exogenous GSH enzymes during early stages of raw starch fermentation may prove to be a viable approach for industrial application of recombinant S. cerevisiae, with the process still benefitting from amylase production by CBP yeast during later stages of cultivation.

Ethanol fermentation was investigated at the high solids content of the dry dilute sulfuric acid pretreated corn stover feedstock using an evolutionary adapted Saccharomyces cerevisiae DQ1 strain. The evolutionary adaptation was conducted by successively transferring the S. cerevisiae DQ1 cells into the inhibitors containing corn stover hydrolysate every 12h and finally a stable yeast strain was obtained after 65 days' continuous adaptation. The ethanol fermentation performance using the adapted strain was significantly improved with the high ethanol titer of 71.40 g/L and the high yield of 80.34% in the simultaneous saccharification and fermentation (SSF) at 30% solids content. No wastewater was generated from pretreatment to fermentation steps. The results were compared with the published cellulosic ethanol fermentation cases, and the obvious advantages of the present work were demonstrated not only at the high ethanol titer and yield, but also the significant reduction of wastewater generation and potential cost reduction., (Copyright © 2015 Elsevier Ltd. All rights reserved.)