Ethanol production from non-grain feedstocks

A general view of the possibilities of producing ethanol from sugar, starch and cellulose feedstocks is given. For the 3 variants net energy analysis of ethanol production and evaluation of costs are presented. With the exception of the case using molasses as feedstock the net energy balances are positive. The greatest possible net energy yield can be expected with sugar cane followed by sugar beets, wood and paper waste. Based on feedstock availability, net energy utilization and production costs, the most promising processes for producing ethanol from non-grain feedstocks over the next 20 years will be those processes using fermentable sugars available from nongrain starchy materials, cellulosics and whey. The feedstock prices for cellulosics are low and if the developments in cellulose hydrolysis will lead to improve the ethanol yields from cellulose fermentation to nearer 90 percent of the theoretical value, cellulosic materials can become a good feedstock for ethanol production.     

Yeasts in sustainable bioethanol production: A review

Bioethanol has been identified as the mostly used biofuel worldwide since it significantly contributes to the reduction of crude oil consumption and environmental pollution. It can be produced from various types of feedstocks such as sucrose, starch, lignocellulosic and algal biomass through fermentation process by microorganisms. Compared to other types of microoganisms, yeasts especially Saccharomyces cerevisiae is the common microbes employed in ethanol production due to its high ethanol productivity, high ethanol tolerance and ability of fermenting wide range of sugars. However, there are some challenges in yeast fermentation which inhibit ethanol production such as high temperature, high ethanol concentration and the ability to ferment pentose sugars. Various types of yeast strains have been used in fermentation for ethanol production including hybrid, recombinant and wild-type yeasts. Yeasts can directly ferment simple sugars into ethanol while other type of feedstocks must be converted to fermentable sugars before it can be fermented to ethanol. The common processes involves in ethanol production are pretreatment, hydrolysis and fermentation. Production of bioethanol during fermentation depends on several factors such as temperature, sugar concentration, pH, fermentation time, agitation rate, and inoculum size. The efficiency and productivity of ethanol can be enhanced by immobilizing the yeast cells. This review highlights the different types of yeast strains, fermentation process, factors affecting bioethanol production and immobilization of yeasts for better bioethanol production.     

Evaluation of waste mushroom logs as a potential biomass resource for the production of bioethanol

In order to investigate the possibility of using waste mushroom logs as a biomass resource for alternative energy production, the chemical and physical characteristics of normal wood and waste mushroom logs were examined. Size reduction of normal wood (145 kW h/tone) required significantly higher energy consumption than waste mushroom logs (70 kW h/tone). The crystallinity value of waste mushroom logs was dramatically lower (33%) than normal wood (49%) after cultivation by Lentinus edodes as spawn. Lignin, an enzymatic hydrolysis inhibitor in sugar production, decreased from 21.07% to 18.78% after inoculation of L. edodes. Total sugar yields obtained by enzyme and acid hydrolysis were higher in waste mushroom logs than in normal wood. After 24h fermentation, 12 g/L ethanol was produced on waste mushroom logs, while normal wood produced 8 g/L ethanol. These results indicate that waste mushroom logs are economically suitable lignocellulosic material for the production of fermentable sugars related to bioethanol production.     

Trends in biotechnological production of fuel ethanol from different feedstocks

Present work deals with the biotechnological production of fuel ethanol from different raw materials. The different technologies for producing fuel ethanol from sucrose-containing feedstocks (mainly sugar cane), starchy materials and lignocellulosic biomass are described along with the major research trends for improving them. The complexity of the biomass processing is recognized through the analysis of the different stages involved in the conversion of lignocellulosic complex into fermentable sugars. The features of fermentation processes for the three groups of studied feedstocks are discussed. Comparative indexes for the three major types of feedstocks for fuel ethanol production are presented. Finally, some concluding considerations on current research and future tendencies in the production of fuel ethanol regarding the pretreatment and biological conversion of the feedstocks are presented.     

Process simulation of modified dry grind ethanol plant with recycle of pretreated and enzymatically hydrolyzed distillers' grains

Distillers' grains (DG), a co-product of a dry grind ethanol process, is an excellent source of supplemental proteins in livestock feed. Studies have shown that, due to its high polymeric sugar contents and ease of hydrolysis, the distillers' grains have potential as an additional source of fermentable sugars for ethanol fermentation. The benefit of processing the distillers' grains to extract fermentable sugars lies in an increased ethanol yield without significant modification in the current dry grind technology. Three different potential configurations of process alternatives in which pretreated and hydrolyzed distillers' grains are recycled for an enhanced overall ethanol yield are proposed and discussed in this paper based on the liquid hot water (LHW) pretreatment of distillers' grains. Possible limitations of each proposed process are also discussed. This paper presents a compositional analysis of distillers' grains, as well as a simulation of the modified dry grind processes with recycle of distillers' grains. Simulated material balances for the modified dry grind processes are established based on the base case assumptions. These balances are compared to the conventional dry grind process in terms of ethanol yield, compositions of its co-products, and accumulation of fermentation inhibitors. Results show that 14% higher ethanol yield is achievable by processing and hydrolyzing the distillers' grains for additional fermentable sugars, as compared to the conventional dry grind process. Accumulation of fermentation by-products and inhibitory components in the proposed process is predicted to be 2-5 times higher than in the conventional dry grind process. The impact of fermentation inhibitors is reviewed and discussed. The final eDDGS (enhanced dried distillers' grains) from the modified processes has 30-40% greater protein content per mass than DDGS, and its potential as a value-added process is also analyzed. While the case studies used to illustrate the process simulation are based on LHW pretreated DG, the process simulation itself provides a framework for evaluation of the impact of other pretreatments.     

Improving the efficiency of enzyme utilization for sugar beet pulp hydrolysis

Sugar beet pulp (SBP) is a carbohydrate-rich residue of table sugar processing. It shows promise as a feedstock for fermentable sugar and biofuel production via enzymatic hydrolysis and microbial fermentation. This research focused on the enzymatic hydrolysis of SBP and examined the effects of solid loading (2–10?%, dry basis), enzyme preparation, and enzyme recycle on the production of fermentable sugars. The enzyme partitioning to the solid and liquid phases during SBP enzymatic hydrolysis and loss during recycling were investigated using SDS-PAGE and Zymogram analysis. Without considering product inhibition, the cellulase added initially to the SBP hydrolysis lost only 6?% filter paper activity and negligible carboxymethyl cellulose activity upon multiple cycles of SBP hydrolysis. It was found that enzyme dosage can be reduced by 50?% while maintaining similar, and in some cases higher fermentable sugar yield. The removal of hydrolysis products will further improve enzymatic hydrolysis of SBP for biofuel production.     

Efficient conversion of sugarcane stalks into ethanol employing low temperature alkali pretreatment method

An alternative route for bio-ethanol production from sugarcane stalks (juice and bagasse) featuring a previously reported low temperature alkali pretreatment method was evaluated. Test-tube scale pretreatment-saccharification experiments were carried out to determine optimal LTA pretreatment conditions for sugarcane bagasse with regard to the efficiency of enzymatic hydrolysis of the cellulose. Free fermentable sugars and bagasse recovered from 2 kg of sugarcane stalks were jointly converted into ethanol via separate enzymatic hydrolysis and fermentation (SHF). Results showed that 98% of the cellulose present in the optimally pretreated bagasse was hydrolyzed into glucose after 72-h enzymatic saccharification using commercially available cellulase and β-glucosidase preparations at relatively low enzyme loading. The fermentable sugars in the mixture of the sugar juice and the bagasse hydrolysate were readily converted into 193.5 mL of ethanol by Saccharomyces cerevisiae within 12h, achieving 88% of the theoretical yield from the sugars and cellulose.     

The use of tamarind waste to improve ethanol production from cane molasses

Tamarind wastes such as tamarind husk, pulp, seeds, fruit and the effluent generated during tartaric acid extraction were used as supplements to evaluate their effects on alcohol production from cane molasses using yeast cultures. Small amounts of these additives enhanced the rate of ethanol production in batch fermentations. Tamarind fruit increased ethanol production (9.7%, w/v) from 22.5% reducing sugars of molasses as compared to 6.5% (w/v) in control experiments lacking supplements after 72 h of fermentation. In general, the addition of tamarind supplements to the fermentation medium showed more than 40% improvement in ethanol production using higher cane molasses sugar concentrations. The direct fermentation of aqueous tamarind effluent also yielded 3.25% (w/v) ethanol, suggesting its possible use as a diluent in molasses fermentations. This is the first report, to our knowledge, in which tamarind-based waste products were used in ethanol production. Received 2 April 1998/ Accepted in revised form 13 November 1998     

Kinetics of sugars consumption and ethanol inhibition in carob pulp fermentation by <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis> in batch and fed-batch cultures

The waste materials from the carob processing industry are a potential resource for second-generation bioethanol production. These by-products are small carob kibbles with a high content of soluble sugars (45–50%). Batch and fed-batch Saccharomyces cerevisiae fermentations of high density sugar from carob pods were analyzed in terms of the kinetics of sugars consumption and ethanol inhibition. In all the batch runs, 90–95% of the total sugar was consumed and transformed into ethanol with a yield close to the theoretical maximum (0.47–0.50 g/g), and a final ethanol concentration of 100–110 g/l. In fed-batch runs, fresh carob extract was added when glucose had been consumed. This addition and the subsequent decrease of ethanol concentrations by dilution increased the final ethanol production up to 130 g/l. It seems that invertase activity and yeast tolerance to ethanol are the main factors to be controlled in carob fermentations. The efficiency of highly concentrated carob fermentation makes it a very promising process for use in a second-generation ethanol biorefinery.     

Effective production of fermentable sugars from brown macroalgae biomass

Brown macroalgae are renewable and sustainable biomass resources for the production of biofuels and chemicals, owing to their high levels of carbohydrates and low levels of lignin. To increase the biological usage of brown macroalgae, it is necessary to depolymerize the polysaccharides that generate macroalgal monomeric sugars or sugar derivatives and to convert them into fermentable sugars for the production of biofuels and chemicals. In this review, we discuss the chemical and enzymatic saccharification of the major carbohydrates found in brown macroalgae and the use of the resulting constituents in the production of biofuels and chemicals, as well as high-value health-benefiting functional oligosaccharides and sugars. We also discuss recently reported experimental results, novel enzymes, and technological breakthroughs that are related to polysaccharide depolymerization, fermentable sugar production, and the biological conversion of non-favorable sugars for fermentation using industrial microorganisms. This review provides a comprehensive perspective of the efficient utilization of brown macroalgae as renewable resources for the production of biofuels and chemicals.     

Solid-state fermentation of grape pomace for ethanol production

Summary Grape pomace was used as a substrate for the production of ethanol under solid-state fermentation conditions. The yield of ethanol amounted to greater than 80% of the theoretical, based on the fermentable sugar consumed.     

Trends and challenges in the microbial production of lignocellulosic bioalcohol fuels

Bioalcohols produced by microorganisms from renewable materials are promising substitutes for traditional fuels derived from fossil sources. For several years already ethanol is produced in large amounts from feedstocks such as cereals or sugar cane and used as a blend for gasoline or even as a pure biofuel. However, alcohols with longer carbon chains like butanol have even more suitable properties and would better fit with the current fuel distribution infrastructure. Moreover, ethical concerns contradict the use of food and feed products as a biofuel source. Lignocellulosic biomass, especially when considered as a waste material offers an attractive alternative. However, the recalcitrance of these materials and the inability of microorganisms to efficiently ferment lignocellulosic hydrolysates still prevent the production of bioalcohols from these plentiful sources. Obviously, no known organism exist which combines all the properties necessary to be a sustainable bioalcohol producer. Therefore, breeding technologies, genetic engineering and the search for undiscovered species are promising means to provide a microorganism exhibiting high alcohol productivities and yields, converting all lignocellulosic sugars or are even able to use carbon dioxide or monoxide, and thereby being highly resistant to inhibitors and fermentation products, and easy to cultivate in huge bioreactors. In this review, we compare the properties of various microorganisms, bacteria and yeasts, as well as current research efforts to develop a reliable lignocellulosic bioalcohol producing organism.     

On-line estimation of sugar concentration for control of fed-batch fermentation of lignocellulosic hydrolyzates by Saccharomyces cerevisiae

A feed control strategy, based on estimated sugar concentrations, was developed with the purpose of avoiding severe inhibition of the yeast Saccharomyces cerevisiae during fermentation of spruce hydrolyzate. The sum of the fermentable hexose sugars, glucose and mannose, was estimated from on-line measurements of carbon dioxide evolution rate and biomass concentration by use of a simple stoichiometric model. The feed rate of the hydrolyzate was controlled to maintain constant sugar concentration during fed-batch fermentation, and the effect of different set-point concentrations was investigated using both untreated and detoxified hydrolyzates. The fed-batch cultivations were evaluated with respect to cellular physiology in terms of the specific ethanol productivities, ethanol yields, and viability of the yeast. The simple stoichiometric model used resulted in a good agreement between estimated sugar concentrations and off-line determinations of sugar concentrations. Furthermore, the control strategy used made it possible to maintain a constant sugar concentration without major oscillations in the feed rate or the sugar concentration. For untreated hydrolyzates the average ethanol productivity could be increased by more than 130% compared to batch fermentation. The average ethanol productivity was increased from 0.12 to 0.28 g/g h. The productivity also increased for detoxified hydrolyzates, where an increase of 16% was found (from 0.50 to 0.58 g/g h).     

Ethanol production from eucalyptus wood hemicellulose hydrolysate by Pichia stipitis

Ethanol production was evaluated from eucalyptus wood hemicellulose acid hydrolysate using Pichia stipitis NRRL Y-7124. An initial lag phase characterized by flocculation and viability loss of the yeast inoculated was observed. Subsequently, cell regrowth occurred with sequential consumption of sugars and production of ethanol. Polyol formation was detected. Acetic acid present in the hydrolysate was an important inhibitor of the fermentation, reducing the rate and the yield. Its toxic effect was due essentially to its undissociated form. The fermentation was more effective at an oxygen transfer rate between 1.2 and 2.4 mmol/L h and an initial pH of 6.5. The hydrolysate used in the experiences had the following composition (expressed in grams per liter): xylose 30, arabinose 2.8, glucose 1.5, galactose 3.7, mannose 1.0, cellobiose 0.5, acetic acid 10, glucuronic acid 1.5, and galacturonic acid 1.0. The best values obtained were maximum ethanol concentration 12.6 g/L, fermentation time 75 h, fermentable sugar consumption 99% ethanol yield 0.35 g/g sugars consumed, and volumetric ethanol productivity 4 g/L day. (c) 1992 John Wiley & Sons, Inc.     

Molecular Adaptation Mechanisms Employed by Ethanologenic Bacteria in Response to Lignocellulose-derived Inhibitory Compounds

Current international interest in finding alternative sources of energy to the diminishing supplies of fossil fuels has encouraged research efforts in improving biofuel production technologies. In countries which lack sufficient food, the use of sustainable lignocellulosic feedstocks, for the production of bioethanol, is an attractive option. In the pre-treatment of lignocellulosic feedstocks for ethanol production, various chemicals and/or enzymatic processes are employed. These methods generally result in a range of fermentable sugars, which are subjected to microbial fermentation and distillation to produce bioethanol. However, these methods also produce compounds that are inhibitory to the microbial fermentation process. These compounds include products of sugar dehydration and lignin depolymerisation, such as organic acids, derivatised furaldehydes and phenolic acids. These compounds are known to have a severe negative impact on the ethanologenic microorganisms involved in the fermentation process by compromising the integrity of their cell membranes, inhibiting essential enzymes and negatively interact with their DNA/RNA. It is therefore important to understand the molecular mechanisms of these inhibitions, and the mechanisms by which these microorganisms show increased adaptation to such inhibitors. Presented here is a concise overview of the molecular adaptation mechanisms of ethanologenic bacteria in response to lignocellulose-derived inhibitory compounds. These include general stress response and tolerance mechanisms, which are typically those that maintain intracellular pH homeostasis and cell membrane integrity, activation/regulation of global stress responses and inhibitor substrate-specific degradation pathways. We anticipate that understanding these adaptation responses will be essential in the design of ''intelligent'' metabolic engineering strategies for the generation of hyper-tolerant fermentation bacteria strains.     

Production of biobutanol from acid-pretreated corncob using Clostridium beijerinckii TISTR 1461: Process optimization studies

Corncob is a potential feedstock in Thailand that can be used for fermentable sugar production through dilute sulfuric acid pretreatment and enzymatic hydrolysis. To recover high amounts of monomeric sugars from corncob, the sulfuric pretreatment conditions were optimized by using response surface methodology with three independent variables: sulfuric acid concentration, temperature, and time. The highest response of total sugars, 48.84 g/L, was found at 122.78°C, 4.65 min, and 2.82% (v/v) H₂SO₄. With these conditions, total sugars from the confirmation experiment were 46.29 g/L, with 5.51% error from the predicted value. The hydrolysate was used as a substrate for acetone–butanol–ethanol fermentation to evaluate its potential for microbial growth. The simultaneous saccharification and fermentation (SSF) showed that C. beijerinckii TISTR 1461 can generate acetone–butanol–ethanol products at 11.64 g/L (5.29 g/L acetone, 6.26 g/L butanol, and 0.09 g/L ethanol) instantly using sugars from the hydrolysed corncob with Novozymes 50013 cellulase enzyme without an overliming process.     

可发酵糖为底物不同菌种丁醇生产能力的研究

随着新一代生物质能源的研发,利用梭菌的发酵生产丁醇已成为热点。选用能生产丁醇的Clostridium acetobutylicum AS1.7,Clostridium acetobutylicum AS1.132,Clostridium acetobutylicumAS1.134和Clostridium beijerinckii NCMIB 8052,在多种糖源下进行发酵培养,通过比较其在不同糖源条件下的生长情况、糖利用率、丁醇及副产物产量、对丁醇、木糖耐受能力等,综合筛选出了最适用于发酵生产丁醇的备选菌种。NCMIB8052因具有最高产量、相对优良的耐受性及可利用多种糖源的特点,而被确定为发酵能力最强的菌种。     

A novel cost-effective technology to convert sucrose and homocelluloses in sweet sorghum stalks into ethanol

Background

Sweet sorghum is regarded as a very promising energy crop for ethanol production because it not only supplies grain and sugar, but also offers lignocellulosic resource. Cost-competitive ethanol production requires bioconversion of all carbohydrates in stalks including of both sucrose and lignocellulose hydrolyzed into fermentable sugars. However, it is still a main challenge to reduce ethanol production cost and improve feasibility of industrial application. An integration of the different operations within the whole process is a potential solution.

Results

An integrated process combined advanced solid-state fermentation technology (ASSF) and alkaline pretreatment was presented in this work. Soluble sugars in sweet sorghum stalks were firstly converted into ethanol by ASSF using crushed stalks directly. Then, the operation combining ethanol distillation and alkaline pretreatment was performed in one distillation-reactor simultaneously. The corresponding investigation indicated that the addition of alkali did not affect the ethanol recovery. The effect of three alkalis, NaOH, KOH and Ca(OH)₂ on pretreatment were investigated. The results indicated the delignification of lignocellulose by NaOH and KOH was more significant than that by Ca(OH)₂, and the highest removal of xylan was caused by NaOH. Moreover, an optimized alkali loading of 10% (w/w DM) NaOH was determined. Under this favorable pretreatment condition, enzymatic hydrolysis of sweet sorghum bagasse following pretreatment was investigated. 92.0% of glucan and 53.3% of xylan conversion were obtained at enzyme loading of 10 FPU/g glucan. The fermentation of hydrolyzed slurry was performed using an engineered stain, Zymomonas mobilis TSH-01. A mass balance of the overall process was calculated, and 91.9 kg was achieved from one tonne of fresh sweet sorghum stalk.

Conclusions

A low energy-consumption integrated technology for ethanol production from sweet sorghum stalks was presented in this work. Energy consumption for raw materials preparation and pretreatment were reduced or avoided in our process. Based on this technology, the recalcitrance of lignocellulose was destructed via a cost-efficient process and all sugars in sweet sorghum stalks lignocellulose were hydrolysed into fermentable sugars. Bioconversion of fermentable sugars released from sweet sorghum bagasse into different products except ethanol, such as butanol, biogas, and chemicals was feasible to operate under low energy-consumption conditions.

     

Liberation of fermentable sugars from soybean hull biomass using ionic liquid 1‐butyl‐3‐methylimidazolium acetate and their bioconversion to ethanol

Optimized hydrolysis of lignocellulosic waste biomass is essential to achieve the liberation of sugars to be used in fermentation process. Ionic liquids (ILs), a new class of solvents, have been tested in the pretreatment of cellulosic materials to improve the subsequent enzymatic hydrolysis of the biomass. Optimized application of ILs on biomass is important to advance the use of this technology. In this research, we investigated the effects of using 1‐butyl‐3‐methylimidazolium acetate ([bmim][Ac]) on the decomposition of soybean hull, an abundant cellulosic industrial waste. Reaction aspects of temperature, incubation time, IL concentration, and solid load were optimized before carrying out the enzymatic hydrolysis of this residue to liberate fermentable glucose. Optimal conditions were found to be 75°C, 165 min incubation time, 57% (mass fraction) of [bmim][Ac], and 12.5% solid loading. Pretreated soybean hull lost its crystallinity, which eased enzymatic hydrolysis, confirmed by Fourier Transform Infrared analysis. The enzymatic hydrolysis of the biomass using an enzyme complex from Penicillium echinulatum liberated 92% of glucose from the cellulose matrix. The hydrolysate was free of any toxic compounds, such as hydroxymethylfurfural and furfural. The obtained hydrolysate was tested for fermentation using Candida shehatae HM 52.2, which was able to convert glucose to ethanol at yields of 0.31. These results suggest the possible use of ILs for the pretreatment of some lignocellulosic waste materials, avoiding the formation of toxic compounds, to be used in second‐generation ethanol production and other fermentation processes. © 2015 American Institute of Chemical Engineers Biotechnol. Prog., 32:312–320, 2016     

Fermentations with new recombinant organisms.

United States fuel ethanol production in 1998 exceeded the record production of 1.4 billion gallons set in 1995. Most of this ethanol was produced from over 550 million bushels of corn. Expanding fuel ethanol production will require developing lower-cost feedstocks, and only lignocellulosic feedstocks are available in sufficient quantities to substitute for corn starch. Major technical hurdles to converting lignocellulose to ethanol include the lack of low-cost efficient enzymes for saccharification of biomass to fermentable sugars and the development of microorganisms for the fermentation of these mixed sugars. To date, the most successful research approaches to develop novel biocatalysts that will efficiently ferment mixed sugar syrups include isolation of novel yeasts that ferment xylose, genetic engineering of Escherichia coli and other gram negative bacteria for ethanol production, and genetic engineering of Saccharoymces cerevisiae and Zymomonas mobilis for pentose utilization. We have evaluated the fermentation of corn fiber hydrolyzates by the various strains developed. E. coli K011, E. coli SL40, E. coli FBR3, Zymomonas CP4 (pZB5), and Saccharomyces 1400 (pLNH32) fermented corn fiber hydrolyzates to ethanol in the range of 21-34 g/L with yields ranging from 0.41 to 0.50 g of ethanol per gram of sugar consumed. Progress with new recombinant microorganisms has been rapid and will continue with the eventual development of organisms suitable for commercial ethanol production. Each research approach holds considerable promise, with the possibility existing that different "industrially hardened" strains may find separate applications in the fermentation of specific feedstocks.