Cellulosic ethanol production: Progress,challenges and strategies for solutions

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

Designing the deconstruction of plant cell walls

Cell wall architecture plays a key role in the regulation of plant cell growth and differentiation into specific cell types. Gaining genetic control of the amount, composition, and structure of cell walls in different cell types will impact both the quantity and yield of fermentable sugars from biomass for biofuels production. The recalcitrance of plant biomass to degradation is a function of how polymers crosslink and aggregate within walls. Novel imaging technologies provide an opportunity to probe these higher order structures in their native state. If cell walls are to be efficiently deconstructed enzymatically to release fermentable sugars, then we require a detailed understanding of their structural organization in future bioenergy crops.     

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.     

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.     

基因工程改善植物生物质能利用的研究进展

利用植物木质纤维资源发酵产乙醇越来越受到人们的重视,但是要达到工业生产仍然存在很多难题。最近在利用植物基因工程技术改善植物自身性状,以利于能源植物的研究方面取得了一定的进展,这些研究包括减少植物自身细胞壁中的木质素含量、细胞中积累表达纤维素酶和木聚耱酶等的方法,使产生的生物质更利于降解利用。     

植物基因工程改善生物质能利用的研究进展

利用植物木质纤维资源发酵生产乙醇越来越受到人们的重视,但是要实现工业化生产仍然存在很多难题。最近,利用植物基因工程技术,改善植物自身性状,包括减少植物自身细胞壁中木质素含量、细胞中积累表达纤维素酶和木聚糖酶等方法,使自生产生的生物质更利于降解利用。目前,对这种新的能源转基因植物的研究取得了一定进展。     

The economics of current and future biofuels

This work presents detailed comparative analysis on the production economics of both current and future biofuels, including ethanol, biodiesel, and butanol. Our objectives include demonstrating the impact of key parameters on the overall process economics (e.g., plant capacity, raw material pricing, and yield) and comparing how next-generation technologies and fuels will differ from today’s technologies. The commercialized processes and corresponding economics presented here include corn-based ethanol, sugarcane-based ethanol, and soy-based biodiesel. While actual full-scale economic data are available for these processes, they have also been modeled using detailed process simulation. For future biofuel technologies, detailed techno-economic data exist for cellulosic ethanol from both biochemical and thermochemical conversion. In addition, similar techno-economic models have been created for n-butanol production based on publicly available literature data. Key technical and economic challenges facing all of these biofuels are discussed.     

The greenhouse gas intensity and potential biofuel production capacity of maize stover harvest in the US Midwest

Agricultural residues are important sources of feedstock for a cellulosic biofuels industry that is being developed to reduce greenhouse gas emissions and improve energy independence. While the US Midwest has been recognized as key to providing maize stover for meeting near‐term cellulosic biofuel production goals, there is uncertainty that such feedstocks can produce biofuels that meet federal cellulosic standards. Here, we conducted extensive site‐level calibration of the Environmental Policy Integrated Climate (EPIC) terrestrial ecosystems model and applied the model at high spatial resolution across the US Midwest to improve estimates of the maximum production potential and greenhouse gas emissions expected from continuous maize residue‐derived biofuels. A comparison of methodologies for calculating the soil carbon impacts of residue harvesting demonstrates the large impact of study duration, depth of soil considered, and inclusion of litter carbon in soil carbon change calculations on the estimated greenhouse gas intensity of maize stover‐derived biofuels. Using the most representative methodology for assessing long‐term residue harvesting impacts, we estimate that only 5.3 billion liters per year (bly) of ethanol, or 8.7% of the near‐term US cellulosic biofuel demand, could be met under common no‐till farming practices. However, appreciably more feedstock becomes available at modestly higher emissions levels, with potential for 89.0 bly of ethanol production meeting US advanced biofuel standards. Adjustments to management practices, such as adding cover crops to no‐till management, will be required to produce sufficient quantities of residue meeting the greenhouse gas emission reduction standard for cellulosic biofuels. Considering the rapid increase in residue availability with modest relaxations in GHG reduction level, it is expected that management practices with modest benefits to soil carbon would allow considerable expansion of potential cellulosic biofuel production.     

Metabolic engineering of microbial pathways for advanced biofuels production

Production of biofuels from renewable resources such as cellulosic biomass provides a source of liquid transportation fuel to replace petroleum-based fuels. This endeavor requires the conversion of cellulosic biomass into simple sugars, and the conversion of simple sugars into biofuels. Recently, microorganisms have been engineered to convert simple sugars into several types of biofuels, such as alcohols, fatty acid alkyl esters, alkanes, and terpenes, with high titers and yields. Here, we review recently engineered biosynthetic pathways from the well-characterized microorganisms Escherichia coli and Saccharomyces cerevisiae for the production of several advanced biofuels.     

Biodelignification of lignocellulose substrates: An intrinsic and sustainable pretreatment strategy for clean energy production

Lignocellulosic biomass (LB) is a promising sugar feedstock for biofuels and other high-value chemical commodities. The recalcitrance of LB, however, impedes carbohydrate accessibility and its conversion into commercially significant products. Two important factors for the overall economization of biofuel production is LB pretreatment to liberate fermentable sugars followed by conversion into ethanol. Sustainable biofuel production must overcome issues such as minimizing water and energy usage, reducing chemical usage and process intensification. Amongst available pretreatment methods, microorganism-mediated pretreatments are the safest, green, and sustainable. Native biodelignifying agents such as Phanerochaete chrysosporium, Pycnoporous cinnabarinus, Ceriporiopsis subvermispora and Cyathus stercoreus can remove lignin, making the remaining substrates amenable for saccharification. The development of a robust, integrated bioprocessing (IBP) approach for economic ethanol production would incorporate all essential steps including pretreatment, cellulase production, enzyme hydrolysis and fermentation of the released sugars into ethanol. IBP represents an inexpensive, environmentally friendly, low energy and low capital approach for second-generation ethanol production. This paper reviews the advancements in microbial-assisted pretreatment for the delignification of lignocellulosic substrates, system metabolic engineering for biorefineries and highlights the possibilities of process integration for sustainable and economic ethanol production.     

Two-stage pretreatment of rice straw using aqueous ammonia and dilute acid

Liberation of fermentable sugars from recalcitrant lignocellulosic biomass is one of the key challenges in production of cellulosic ethanol. Here we developed a two-stage pretreatment process using aqueous ammonia and dilute sulfuric acid in a percolation mode to improve production of fermentable sugars from rice straw. Aqueous NH? was used in the first stage which removed lignin selectively but left most of cellulose (97%) and hemicellulose (77%). Dilute acid was applied in the second stage which removed most of hemicellulose, partially disrupted the crystalline structure of cellulose, and thus enhanced enzymatic digestibility of cellulose in the solids remaining. Under the optimal pretreatment conditions, the enzymatic hydrolysis yields of the two-stage treated samples were 96.9% and 90.8% with enzyme loadings of 60 and 15FPU/g of glucan, respectively. The overall sugar conversions of cellulose and hemicellulose into glucose and xylose by enzymatic and acid hydrolysis reached 89.0% and 71.7%, respectively.     

High-level expression of a suite of thermostable cell wall-degrading enzymes from the chloroplast genome

The biological conversion of plant biomass into fermentable sugars is key to the efficient production of biofuels and other renewable chemicals from plants. As up to more than 90% of the dry weight of higher plants is fixed in the cell wall, this will require the low-cost production of large amounts of cell wall-degrading enzymes. Transgenic plants can potentially provide an unbeatably cheap production platform for industrial enzymes. Transgene expression from the plastid genome is particularly attractive, due to high-level foreign protein accumulation in chloroplasts, absence of epigenetic gene silencing and improved transgene containment. Here, we have explored the potential of transplastomic plants to produce large amounts of thermostable cell wall-degrading enzymes from the bacterium Thermobifida fusca. We show that a set of four enzymes that are required for efficient degradation of cellulose (and the hemicellulose xyloglucan) could be expressed successfully in transplastomic tobacco plants. However, overexpression of the enzymes (to between approximately 5 and 40% of the plant's total soluble protein) resulted in pigment-deficient mutant phenotypes. We demonstrate that the chloroplast-produced cellulolytic enzymes are highly active. Although further optimization is needed, our data indicate that transgenic plastids offer great potential for the production of enzyme cocktails for the bioconversion of cellulosic biomass.     

Process and technoeconomic analysis of leading pretreatment technologies for lignocellulosic ethanol production using switchgrass

Six biomass pretreatment processes to convert switchgrass to fermentable sugars and ultimately to cellulosic ethanol are compared on a consistent basis in this technoeconomic analysis. The six pretreatment processes are ammonia fiber expansion (AFEX), dilute acid (DA), lime, liquid hot water (LHW), soaking in aqueous ammonia (SAA), and sulfur dioxide-impregnated steam explosion (SO(2)). Each pretreatment process is modeled in the framework of an existing biochemical design model so that systematic variations of process-related changes are consistently captured. The pretreatment area process design and simulation are based on the research data generated within the Biomass Refining Consortium for Applied Fundamentals and Innovation (CAFI) 3 project. Overall ethanol production, total capital investment, and minimum ethanol selling price (MESP) are reported along with selected sensitivity analysis. The results show limited differentiation between the projected economic performances of the pretreatment options, except for processes that exhibit significantly lower monomer sugar and resulting ethanol yields.     

Effect of surfactant and particle size reduction on hydrolysis of deinking sludge and nonrecyclable newsprint

Disposal of sludge from deinking mills represents a significant proportion of operating costs. Bioconversion of the cellulosic fraction of deinking sludge (DIS) to ethanol greatly reduces disposal costs while producing an environmentally friendly fuel. In this study, the cellulosic fraction of newsprint and deinking sludge was hydrolysed to produce fermentable sugars. For newsprint, a particle size of 1 to 1.5 mm provided optimal reaction rates in batch reactors over practical hydrolysis times, and reducing sugar concentrations as high as 35 g/L could be achieved using a fed-batch reactor configuration. For both newsprint and DIS, the hydrolysis rate increased nonlinearly with enzyme loading. Tween-80 only marginally improved sugar production but was able to release sugars from cellulosic substrates in the absence of lytic enzymes, in an amount proportional to the surfactant concentration and the substrate particle size. DIS was relatively recalcitrant to enzymatic hydrolysis, possibly due in part to inhibition by hydrophobic constituents. (c) 1995 John Wiley & Sons, Inc.     

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.     

Fungal pretreatment of lignocellulosic biomass

Pretreatment is a crucial step in the conversion of lignocellulosic biomass to fermentable sugars and biofuels. Compared to thermal/chemical pretreatment, fungal pretreatment reduces the recalcitrance of lignocellulosic biomass by lignin-degrading microorganisms and thus potentially provides an environmentally-friendly and energy-efficient pretreatment technology for biofuel production. This paper provides an overview of the current state of fungal pretreatment by white rot fungi for biofuel production. The specific topics discussed are: 1) enzymes involved in biodegradation during the fungal pretreatment; 2) operating parameters governing performance of the fungal pretreatment; 3) the effect of fungal pretreatment on enzymatic hydrolysis and ethanol production; 4) efforts for improving enzymatic hydrolysis and ethanol production through combinations of fungal pretreatment and physical/chemical pretreatment; 5) the treatment of lignocellulosic biomass with lignin-degrading enzymes isolated from fungal pretreatment, with a comparison to fungal pretreatment; 6) modeling, reactor design, and scale-up of solid state fungal pretreatment; and 7) the limitations and future perspective of this technology.     

Composition of corn dry-grind ethanol by-products: DDGS, wet cake, and thin stillage

DDGS and wet distillers' grains are the major co-products of the dry grind ethanol facilities. As they are mainly used as animal feed, a typical compositional analysis of the DDGS and wet distillers' grains mainly focuses on defining the feedstock's nutritional characteristics. With an increasing demand for fuel ethanol, the DDGS and wet distillers' grains are viewed as a potential bridge feedstock for ethanol production from other cellulosic biomass. The introduction of DDGS or wet distillers' grains as an additional feed to the existing dry grind plants for increased ethanol yield requires a different approach to the compositional analysis of the material. Rather than focusing on its nutritional value, this new approach aims at determining more detailed chemical composition, especially on polymeric sugars such as cellulose, starch and xylan, which release fermentable sugars upon enzymatic hydrolysis. In this paper we present a detailed and complete compositional analysis procedure suggested for DDGS and wet distillers' grains, as well as the resulting compositions completed by three different research groups. Polymeric sugars, crude protein, crude oil and ash contents of DDGS and wet distillers' grains were accurately and reproducibly determined by the compositional analysis procedure described in this paper.     

Plant genetic engineering to improve biomass characteristics for biofuels

Currently, most ethanol produced in the United States is derived from maize kernel, at levels in excess of four billion gallons per year. Plant lignocellulosic biomass is renewable, cheap and globally available at 10-50 billion tons per year. At present, plant biomass is converted to fermentable sugars for the production of biofuels using pretreatment processes that disrupt the lignocellulose and remove the lignin, thus allowing the access of microbial enzymes for cellulose deconstruction. Both the pretreatments and the production of enzymes in microbial tanks are expensive. Recent advances in plant genetic engineering could reduce biomass conversion costs by developing crop varieties with less lignin, crops that self-produce cellulase enzymes for cellulose degradation and ligninase enzymes for lignin degradation, or plants that have increased cellulose or an overall biomass yield.     

Ethanol from sugarcane in Brazil: a ‘midway’ strategy for increasing ethanol production while maximizing environmental benefits

This article reviews the history and current state of ethanol production from sugarcane in Brazil and presents a strategy for improving ecosystem services and production. We propose that it is possible to produce ethanol from sugarcane while maintaining or even recovering some of Brazil's unique neotropical biodiversity and ecosystem climate services. This approach to the future of sustainable and responsible ethanol production is termed the ‘midway’ strategy. The ‘midway’ strategy involves producing the necessary biotechnology to increase productivity while synergistically protecting and regenerating rainforest. Three main areas of scientific and technological advance that are key to realizing the ‘midway’ strategy are: (i) improving the quality of scientific data on sugarcane biology as pertains to its use as a bioenergy crop; (ii) developing technologies for the use of bagasse for cellulosic ethanol; and (iii) developing policies to improve the ecosystem services associated with sugarcane landscapes. This article discusses these three issues in the general context of biofuels production and highlights examples of scientific achievements that are already leading towards the ‘midway’ strategy.     

Engineering cellulolytic ability into bioprocessing organisms

Lignocellulosic biomass is an abundant renewable feedstock for sustainable production of commodities such as biofuels. The main technological barrier that prevents widespread utilization of this resource for production of commodity products is the lack of low-cost technologies to overcome the recalcitrance of lignocellulose. Organisms that hydrolyse the cellulose and hemicelluloses in biomass and produce a valuable product such as ethanol at a high rate and titre would significantly reduce the costs of current biomass conversion technologies. This would allow steps that are currently accomplished in different reactors, often by different organisms, to be combined in a consolidated bioprocess (CBP). The development of such organisms has focused on engineering naturally cellulolytic microorganisms to improve product-related properties or engineering non-cellulolytic organisms with high product yields to become cellulolytic. The latter is the focus of this review. While there is still no ideal organism to use in one-step biomass conversion, several candidates have been identified. These candidates are in various stages of development for establishment of a cellulolytic system or improvement of product-forming attributes. This review assesses the current state of the art for enabling non-cellulolytic organisms to grow on cellulosic substrates.