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.     

木质纤维生产燃料乙醇工艺的研究进展

利用丰富而廉价的木质纤维原料代替粮食生产燃料乙醇,对经济和社会的可持续发展有着重要的意义。以木质纤维为原料发酵生产燃料乙醇可分为4种工艺:分步糖水解化发酵法、同步糖化发酵法、同步糖化共发酵法和直接微生物转化法。介绍了以上4种工艺的研究进展,并对今后进一步研究提出了建议。     

微生物木糖发酵产乙醇的代谢工程

利用木质纤维素发酵生产乙醇具有广泛的应用前景。而自然界中缺少有效转化木糖为乙醇的微生物是充分利用纤维素水解产物、提高乙醇产率、降低生产成本的关键因素。多年来研究者利用分子生物学技术对微生物菌株进行了代谢工程改造,使其能更有效地利用木糖生产乙醇。以下主要对运动发酵单胞菌、大肠杆菌和酵母等候选产乙醇微生物的木糖代谢工程研究进展进行了概述。     

褐藻生物乙醇的研究进展

利用大型褐藻转化生产的第三代燃料乙醇已受到研究者的广泛关注。我国拥有丰富的褐藻资源,具备了褐藻生物乙醇转化的有利条件。为了实现工业化生产,还需要通过筛选分离和基因工程手段获得高效发酵褐藻的优良菌株及优化预处理、发酵条件等。主要介绍了我国褐藻资源概况、预处理方法和微生物发酵褐藻不同组分生产乙醇的研究进展,提出了当前褐藻乙醇转化中存在的问题,展望了褐藻乙醇的发展方向。     

Bermuda grass as feedstock for biofuel production: a review

Bermuda grass is a promising feedstock for the production of fuel ethanol in the Southern United States. This paper presents a review of the significant amount of research on the conversion of Bermuda grass to ethanol and a brief discussion on the factors affecting the biomass production in the field. The biggest challenge of biomass conversion comes from the recalcitrance of lignocellulose. A variety of chemical, physico-chemical, and biological pretreatment methods have been investigated to improve the digestibility of Bermuda grass with encouraging results reported. The subsequent enzymatic hydrolysis and fermentation steps have also been extensively studied and effectively optimized. It is expected that the development of genetic engineering technologies for the grass and fermenting organisms has the potential to greatly improve the economic viability of Bermuda grass-based fuel ethanol production systems. Other energy applications of Bermuda grass include anaerobic digestion for biogas generation and pyrolysis for syngas production.     

燃料乙醇发酵技术研究进展

在分析木质纤维素类生物质制备燃料乙醇原理基础上,重点对燃料乙醇转化过程的发酵工艺进行了论述。目前乙醇发酵工艺主要包括直接发酵、分步糖化发酵、同步糖化发酵、同步糖化共发酵和联合生物加工技术等,对这几种技术的研究现状进行了分析并对其发展趋势进行了展望,通过基因工程构建高效发酵菌种的联合生物加工技术将是未来高效发酵工艺的发展趋势,旨在为有效提高发酵菌株的底物代谢能力,获得高的乙醇产量提供重要参考。     

Hemicellulose bioconversion

Various agricultural residues, such as corn fiber, corn stover, wheat straw, rice straw, and sugarcane bagasse, contain about 20–40% hemicellulose, the second most abundant polysaccharide in nature. The conversion of hemicellulose to fuels and chemicals is problematic. In this paper, various pretreatment options as well as enzymatic saccharification of lignocellulosic biomass to fermentable sugars is reviewed. Our research dealing with the pretreatment and enzymatic saccharification of corn fiber and development of novel and improved enzymes such as endo-xylanase, β-xylosidase, and α-l-arabinofuranosidase for hemicellulose bioconversion is described. The barriers, progress, and prospects of developing an environmentally benign bioprocess for large-scale conversion of hemicellulose to fuel ethanol, xylitol, 2,3-butanediol, and other value-added fermentation products are highlighted.     

纤维素乙醇统合加工过程中的酵母多酶共展示体系研究进展

利用统合生物加工过程(Consolidated bioprocessing,CBP)生产纤维素乙醇是目前国内外的研究热点。CBP需要一种“集成化”微生物,既能生产水解木质纤维素的多种酶类又能利用水解木质纤维素产生的糖类发酵产乙醇。以酿酒酵母表面展示技术为依托,建立CBP菌株多酶共展示体系的研究主要分为以下两个方向：一是直接将纤维素酶展示在细胞表面,即非复合型纤维素酶体系;另一种是通过表面展示纤维小体(Cellulosome)将纤维素酶间接地锚定在细胞表面,即复合型纤维素酶体系,本文主要从以上两个方向阐述了近几年对于纤维素乙醇生物统合加工过程的研究进展。因纤维小体对纤维素的降解能力比非复合型纤维素酶体系更强,所以其在酿酒酵母细胞表面的组装研究受到越来越多的关注,为了更深入透彻地了解纤维小体的酵母展示技术,文中对纤维小体的结构与功能及其在纤维素乙醇发酵中的应用研究进行重点论述,并对该领域的发展方向进行展望。     

Quantifying the metabolic capabilities of engineered <Emphasis Type="Italic">Zymomonas mobilis</Emphasis> using linear programming analysis

Background  

The need for discovery of alternative, renewable, environmentally friendly energy sources and the development of cost-efficient, "clean" methods for their conversion into higher fuels becomes imperative. Ethanol, whose significance as fuel has dramatically increased in the last decade, can be produced from hexoses and pentoses through microbial fermentation. Importantly, plant biomass, if appropriately and effectively decomposed, is a potential inexpensive and highly renewable source of the hexose and pentose mixture. Recently, the engineered (to also catabolize pentoses) anaerobic bacterium Zymomonas mobilis has been widely discussed among the most promising microorganisms for the microbial production of ethanol fuel. However, Z. mobilis genome having been fully sequenced in 2005, there is still a small number of published studies of its in vivo physiology and limited use of the metabolic engineering experimental and computational toolboxes to understand its metabolic pathway interconnectivity and regulation towards the optimization of its hexose and pentose fermentation into ethanol.     

A Survey of Bioenergy Research in Forest Service Research and Development

Forest biomass represents 25–30 % of the annual biomass available in the USA for conversion into bio-based fuels, bio-based chemicals, and bioproducts in general. The USDA Forest Service Research and Development (R&D) has been focused on producing products from forest biomass since its inception in 1905, with direct combustion, solid sawn lumber, pulp and paper, ethanol as fuel, and silvichemicals all among the mission areas of product research and development. The renewed national interest in biomass conversion to fuels and chemicals is supportive of the most critical need of USDA Forest Service R&D, uses for small-diameter trees and other forest biomass that needs to be removed in the fuel mitigation–fire suppression and forest restoration work of the USDA Forest Service. This paper will summarize the recent USDA Forest Service research on direct combustion, fuel pellets, and conversion of forest biomass to ethanol, both as stand-alone biorefinery processes and as an addition to the traditional wood pulping process.     

Microbial‐based motor fuels: science and technology

The production of biofuels via microbial biotechnology is a very active field of research. A range of fuel molecule types are currently under consideration: alcohols, ethers, esters, isoprenes, alkenes and alkanes. At the present, the major alcohol biofuel is ethanol. The ethanol fermentation is an old technology. Ongoing efforts aim to increase yield and energy efficiency of ethanol production from biomass. n‐Butanol, another microbial fermentation product, is potentially superior to ethanol as a fuel but suffers from low yield and unwanted side‐products currently. In general, biodiesel fuels consist of fatty acid methyl esters in which the carbon derives from plants, not microbes. A new biodiesel product, called microdiesel, can be generated in engineered bacterial cells that condense ethanol with fatty acids. Perhaps the best fuel type to generate from biomass would be biohydrocarbons. Microbes are known to produce hydrocarbons such as isoprenes, long‐chain alkenes and alkanes. The biochemical mechanisms of microbial hydrocarbon biosynthesis are currently under study. Hydrocarbons and minimally oxygenated molecules may also be produced by hybrid chemical and biological processes. A broad interest in novel fuel molecules is also driving the development of new bioinformatics tools to facilitate biofuels research.     

Scientific challenges of bioethanol production in Brazil

Bioethanol (fuel alcohol) has been produced by industrial alcoholic fermentation processes in Brazil since the beginning of the twentieth century. Currently, 432 mills and distilleries crush about 625 million tons of sugarcane per crop, producing about 27 billion liters of ethanol and 38.7 million tons of sugar. The production of bioethanol from sugarcane represents a major large-scale technology capable of producing biofuel efficiently and economically, providing viable substitutes to gasoline. The combination of immobilization of CO₂ by sugarcane crops by photosynthesis into biomass together with alcoholic fermentation of this biomass has allowed production of a clean and high-quality liquid fuel that contains 93% of the original energy found in sugar. Over the last 30 years, several innovations have been introduced to Brazilian alcohol distilleries resulting in the improvement of plant efficiency and economic competitiveness. Currently, the main scientific challenges are to develop new technologies for bioethanol production from first and second generation feedstocks that exhibit positive energy balances and appropriately meet environmental sustainability criteria. This review focuses on these aspects and provides special emphasis on the selection of new yeast strains, genetic breeding, and recombinant DNA technology, as applied to bioethanol production processes.     

Study of the enzymatic hydrolysis of cellulose for production of fuel ethanol by the simultaneous saccharification and fermentation process

The biochemical conversion of cellulosic biomass to ethanol, a promising alternative fuel, can be carried out efficiently and economically using the simultaneous saccharification and fermentation (SSF) process. The SSF integrates the enzymatic hydrolysis of cellulose to glucose, catalyzed by the synergistic action of cellulase and beta-glucosidase, with the fermentative synthesis of ethanol. Because the enzymatic step determines the ethanol. Because the enzymatic step determines the availability of glucose to the ethanologenic fermentation, the kinetic of cellulose hydrolysis by cellulase and beta-glucosidase and the susceptibility of the two enzymes to inhibition by hydrolysis and fermentation products are of significant importance to the SSF performance and were investigated under realistic SSF conditions. A previously developed SSF mathematical model was used to conceptualize the depolymerization of cellulose. The model was regressed to the collected data to determine the values of the enzyme parameters and was found to satisfactorily predict the kinetics of cellulose hydrolysis. Cellobiose and glucose were identified as the strongest inhibitors of cellulase and beta-glucosidase, respectively. Experimental and modeling results are presented in light of the impact of enzymatic hydrolysis on fuel ethanol production. (c) 1993 Wiley & Sons, Inc.     

Thermophilic,lignocellulolytic bacteria for ethanol production: current state and perspectives

Lignocellulosic biomass contains a variety of carbohydrates, and their conversion into ethanol by fermentation requires an efficient microbial platform to achieve high yield, productivity, and final titer of ethanol. In recent years, growing attention has been devoted to the development of cellulolytic and saccharolytic thermophilic bacteria for lignocellulosic ethanol production because of their unique properties. First of all, thermophilic bacteria possess unique cellulolytic and hemicellulolytic systems and are considered as potential sources of highly active and thermostable enzymes for efficient biomass hydrolysis. Secondly, thermophilic bacteria ferment a broad range of carbohydrates into ethanol, and some of them display potential for ethanologenic fermentation at high yield. Thirdly, the establishment of the genetic tools for thermophilic bacteria has allowed metabolic engineering, in particular with emphasis on improving ethanol yield, and this facilitates their employment for ethanol production. Finally, different processes for second-generation ethanol production based on thermophilic bacteria have been proposed with the aim to achieve cost-competitive processes. However, thermophilic bacteria exhibit an inherent low tolerance to ethanol and inhibitors in the pretreated biomass, and this is at present the greatest barrier to their industrial application. Further improvement of the properties of thermophilic bacteria, together with the optimization production processes, is equally important for achieving a realistic industrial ethanol production.     

Consolidated Bioprocessing of Lignocellulosic Feedstocks for Ethanol Fuel Production

Ethanol fuel can be produced renewably from numerous plant and waste materials, but harnessing the energy of lignocellulosic feedstocks has been particularly challenging in the development of this alternative fuel as a substitute for petroleum-based fuels. Consolidated bioprocessing has the potential to make the conversion of biomass to fuel an economical process by combining enzyme production, polysaccharide hydrolysis, and sugar fermentation into a single unit operation. This consolidation of steps takes advantage of the synergistic nature of enzyme systems but requires the use of one or a few organisms capable of producing highly efficient cellulolytic enzymes and fermenting most of the resulting sugars to ethanol with minimal byproduct formation while tolerating high levels of ethanol. In this review, conventional ethanol production, consolidated bioprocessing, and simultaneous saccharification and fermentation are described and compared. Several wild-type and genetically engineered microorganisms, including strains of Clostridium thermocellum, Saccharomyces cerevisiae, Klebsiella oxytoca, Escherichia coli, Flammulina velutipes, and Zymomonas mobilis, among others, are highlighted for their potential in consolidated bioprocessing. This review examines the favorable and undesirable qualities of these microorganisms and their enzyme systems, process engineering considerations for particular organisms, characteristics of cellulosomes, enzyme engineering strategies, progress in commercial development, and the impact of these topics on current and future research.     

Enteric bacterial catalysts for fuel ethanol production.

The technology is available to produce fuel ethanol from renewable lignocellulosic biomass. The current challenge is to assemble the various process options into a commercial venture and begin the task of incremental improvement. Current process designs for lignocellulose are far more complex than grain to ethanol processes. This complexity results in part from the complexity of the substrate and the biological limitations of the catalyst. Our work at the University of Florida has focused primarily on the genetic engineering of Enteric bacteria using genes encoding Zymomonas mobilis pyruvate decarboxylase and alcohol dehydrogenase. These two genes have been assembled into a portable ethanol production cassette, the PET operon, and integrated into the chromosome of Escherichia coli B for use with hemicellulose-derived syrups. The resulting strain, KO11, produces ethanol efficiently from all hexose and pentose sugars present in the polymers of hemicellulose. By using the same approach, we integrated the PET operon into the chromosome of Klebsiella oxytoca to produce strain P2 for use in the simultaneous saccharification and fermentation (SSF) process for cellulose. Strain P2 has the native ability to ferment cellobiose and cellotriose, eliminating the need for one class of cellulase enzymes. Recently, the ability to produce and secrete high levels of endoglucanase has also been added to strain P2, further reducing the requirement for fungal cellulase. The general approach for the genetic engineering of new biocatalysts using the PET operon has been most successful with Enteric bacteria but was also extended to Gram positive bacteria, which have other useful traits for lignocellulose conversion. Many opportunities remain for further improvements in these biocatalysts as we proceed toward the development of single organisms that can be used for the efficient fermentation of both hemicellulosic and cellulosic substrates.     

Metabolic engineering of bacteria for ethanol production

Technologies are available which will allow the conversion of lignocellulose into fuel ethanol using genetically engineered bacteria. Assembling these into a cost-effective process remains a challenge. Our work has focused primarily on the genetic engineering of enteric bacteria using a portable ethanol production pathway. Genes encoding Zymomonas mobilis pyruvate decarboxylase and alcohol dehydrogenase have been integrated into the chromosome of Escherichia coli B to produce strain KO11 for the fermentation of hemicellulose-derived syrups. This organism can efficiently ferment all hexose and pentose sugars present in the polymers of hemicellulose. Klebsiella oxytoca M5A1 has been genetically engineered in a similar manner to produce strain P2 for ethanol production from cellulose. This organism has the native ability to ferment cellobiose and cellotriose, eliminating the need for one class of cellulase enzymes. The optimal pH for cellulose fermentation with this organism (pH 5.0-5.5) is near that of fungal cellulases. The general approach for the genetic engineering of new biocatalysts has been most successful with enteric bacteria thus far. However, this approach may also prove useful with Gram-positive bacteria which have other important traits for lignocellulose conversion. Many opportunities remain for further improvements in the biomass to ethanol processes. These include the development of enzyme-based systems which eliminate the need for dilute acid hydrolysis or other pretreatments, improvements in existing pretreatments for enzymatic hydrolysis, process improvements to increase the effective use of cellulase and hemicellulase enzymes, improvements in rates of ethanol production, decreased nutrient costs, increases in ethanol concentrations achieved in biomass beers, increased resistance of the biocatalysts to lignocellulosic-derived toxins, etc. To be useful, each of these improvements must result in a decrease in the cost for ethanol production. Copyright 1998 John Wiley & Sons, Inc.     

Emerging technology for fermenting d-xylose

In the past four years, numerous yeasts which convert d-xylose to ethanol have been reported. The conversion occurs most readily under aerobic conditions. Various aspects of this conversion have provided new insight into the mechanisms and metabolic regulation of ethanol fermentation in yeasts. Although specific fermentation rates, product yields and product concentrations are significantly lower with d-xylose than with d-glucose, technology is emerging which may prove to be feasible for commercial fermentation of d-xylose-containing waste streams.     

氧限制下粗糙脉孢菌乙醇发酵的数学模型及过程模拟

粗糙脉孢菌(Mzcrospora crassan)具有直接转化植物纤维性物质生产乙醇的能力。研究了不同氧限制条件对粗糙脉孢菌发酵葡萄糖生产乙醇的影响,构建了该过程的数学模型,并利用数学模型进行了模拟和预测研究。结果表明,数学模型能够很好地预测氧限制条件下乙醇的发酵过程,即使微量氧对乙醇发酵也有较大的负面影响。