Hindawi Publishing Corporatione Scientific World Journal Volume 2014, Article ID 298153, 13 pages http://dx.doi.org/10.1155/2014/298153 Review Article Bioethanol from Lignocellulosic Biomass: Current Findings Determine Research Priorities Qian Kang,1,2 Lise Appels,2 Tianwei Tan,1 and Raf Dewil2 1 Key Lab of Bioprocess, College of Life Science and Technology, Beijing University of Chemical Technology, Beijing 100029, China Department of Chemical Engineering, Process and Environmental Technology Lab, KU Leuven, 2860 Sint-Katelijne-Waver, Belgium 2 Correspondence should be addressed to Qian Kang;
[email protected] Received 9 November 2014; Accepted 18 December 2014; Published 31 December 2014 Academic Editor: Reeta R. Singhania Copyright © 2014 Qian Kang et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. “Second generation” bioethanol, with lignocellulose material as feedstock, is a promising alternative for first generation bioethanol. This paper provides an overview of the current status and reveals the bottlenecks that hamper its implementation. The current literature specifies a conversion of biomass to bioethanol of 30 to ∼50% only. Novel processes increase the conversion yield to about 92% of the theoretical yield. New combined processes reduce both the number of operational steps and the production of inhibitors. Recent advances in genetically engineered microorganisms are promising for higher alcohol tolerance and conversion efficiency. By combining advanced systems and by intensive additional research to eliminate current bottlenecks, second generation bioethanol could surpass the traditional first generation processes. 1. Introduction 1.1. Bioethanol as Sustainable Fuel. With the global increasing demand for energy, energy shortage will be a global problem. Bioethanol is considered as an important renewable fuel to partly replace fossil-derived fuels. The world production of bioethanol increased from 50 million m3 in 2007 to over 100 million m3 in 2012 [1]: Brazil and the United States represent approximately 80% of the world supply, mostly using corn or sugarcane. In developing economies, food-related feedstock is preferably replaced by nonfood raw materials, such as sweet sorghum or cassava. The use of common biomass could significantly increase the bioethanol production, and lignocellulose-based bioethanol is therefore the topic of the present review paper. The current technological development and bottlenecks define the short- and medium-term research priorities. Industrial ethanol is mainly produced petrochemically through the acid-catalyzed hydration of ethylene. Ethanol for use in alcoholic beverages, and the vast majority of ethanol for use as biofuel, is produced by fermentation where certain species of yeast (e.g., Saccharomyces cerevisiae) or bacteria (e.g., Zymomonas mobilis) metabolize sugars in oxygen-lean conditions to produce ethanol and carbon dioxide. The main reasons for the enhanced development of bioethanol are its use as a favourable and near carbonneutral renewable fuel, thus reducing CO2 emissions and associated climate change; its use as octane enhancer in unleaded gasoline; and its use as oxygenated fuel-mix for a cleaner combustion of gasoline, hence reducing tailpipe pollutant emissions and improving the ambient air quality. The largest single use of ethanol is as engine fuel and fuel additive, with common types of available fuel-mixes listed in Table 1. Ethanol has appropriate properties for spark ignition IC engines. Its octane numbers, motor octane number (MON) and research octane number (RON), are 90 and 109, respectively, on average 99 compared to 91 for regular gasoline. Due to its low cetane number, ethanol does not burn efficiently by compression ignition and is moreover not easily miscible with diesel fuel. To improve the use of ethanol in compressionignition (CI) engine vehicles, measures can be taken, such as the addition of an emulsifier in order to increase the ethanol-diesel miscibility; the addition of ethylhexyl nitrate Malinen et al. sewage treatment sludge. as such unusable in industrial and fuel applications. as residues produced during and after application end.2. The unit EJ (eta-joule) is equal to 1018 J. such as straw. and so forth [4]. second. 15% petrol Hydrous ethanol (∼5. 1. Europe USA. [8] and Demirbas¸ [9]. Code E5 E10 E15 E25 E85 E100 Composition Max. In general the final availability of organic wastes and residues may fluctuate and is affected by market growth. bagasse. 15% anhydrous ethanol. including cotton stalks.3 wt% water) or diterbutyl peroxide to enhance the cetane number. The overall potential of the organic fraction of municipal waste and its waste wood fraction is strongly reliant on economic growth. produced as either crop or key product. min. as residues from the production processes. 85% petrol Max. and others.1. and the use of biomaterials. The energy potential of the world’s forests is again difficult to estimate. 1. Although these lignocellulosic biomass resources represent a significant energy value. Whether first. jute sticks. whereas the use of lignocellulose raw materials is commonly called “second generation” bioethanol. It is estimated at between 5 and 50 EJ/year [4]. Lignocellulose. although climate and other factors have influences. secondary sources. Agroindustrial biomass residues are byproducts of agriculture or its related industry. The potential of agriculture residues varies from 15 up to 70 EJ/year [4]. wheat and rice straw. as reported by Kang et al. 95% petrol Max. through forest management operations such as precommercial thinning and removal of dead and dying trees. Due to the high transportation cost. fermentation produces an alcohol-lean broth only. The organic fraction of municipal solid waste (MSW) is an inexpensive source of biomass and covers domestic and industrial waste collected in a specific area. Forestry residues include biomass. 75% petrol Max.6 vol% (89. the total land area available. for reasons given below. 90% petrol Max. Sources. and straw.2 The Scientific World Journal Table 1: Common ethanol-petrol mixtures [1. especially when considering the primary sources. sawdust. and rice husks [5]. with the lower estimate due to the current use as fertiliser. or third generation feedstock is used. for example. dried manure is considered as a tertiary source. Lignocellulosic Biomass 1. [1]. and . The specificities of using lignocellulosic raw materials will be dealt with in Section 1. The extraction costs and the required transportation to centralized processing plants make forest fuels expensive. 31]. wood trimmings. Further ethanol enrichment by common distillation is impossible. 7]. the use of a dual fuel operation in which ethanol and diesel are introduced separately into the cylinder. despite the cost of the required enzymes to hydrolyse cellulose. as a function of regional production. for example. hemicellulose (20–40%). [3]. Forestry waste includes wood chips. Within the agroindustrial residues. It can provide 65% of the biomass energy potential [6. and tertiary sources. The agricultural residues are produced decentralised and have a low density. or the modification of diesel engines in order to adapt their characteristics of autoignition [2]. Composition. ∼150 EJ/year. Since the present paper deals with second generation bioethanol. corresponding to the azeotropic composition with a boiling point of 78. for example. and recoverability factors.2∘ C. relevant and recent (>2010) literature is summarized in Table 2. Several studies have focused on their use for energy production at a district level. The first generation processes were discussed in detail by Kang et al.3. Europe Brazil Comments Blends for regular cars Flex-fuel vehicles on the yield. bamboo.3. rice husks. and given as 5 to 55 EJ/year. The possible contribution by 2050 is estimated at 98 EJ/year of excess natural forest growth and at 32–52 EJ/year of processing residues. the organic fraction of municipal solid waste (MSW). A total worldwide estimate is difficult to make. not harvested or removed from sorting regions in commercial hardwood and softwood production. min. 85% anhydrous ethanol. min. The “third generation” of algal bioethanol is at an early stage of investigation. Available biomass can be categorized into primary sources. only part of this resource can be used as feedstock for bioethanol. 1. is mainly composed of cellulose (40– 60% of the total dry weight). harvesting processing factors. the principal component of the plant cell walls. sugar cane. The ethanol must hence be purified. cars >2000 Brazil USA. Development of this technology could deal with a number of cellulosecontaining agricultural byproducts. Bioethanol from Different Feedstocks.3. 10% anhydrous ethanol. but different alternatives have been investigated. it is expensive to apply agricultural residues as the main fuel in power stations.2. Fractional distillation can concentrate ethanol to 95. 5% anhydrous ethanol. coconut shells. for example. The energy potential of biomass residues and organic wastes depends Countries Western Europe USA. while the higher estimate considers the total technical potential [5]. min. The 2nd generation bioethanol processes will use cellulose-released sugars. wood trimmings.3. Fermentation of sugar-based raw materials is referred to as “first generation” bioethanol. maize cobs. and the type of production. applying appropriate designs of decentralized smaller plants. short rotation energy plantations. sawdust. and bark. min. 25% anhydrous ethanol. consumption.5 mol%). and conversion of C5 sugars to ethanol (ii) The residues need to be processed for byproducts through biorefinery to improve the economics of the whole process (i) This enzyme extract promoted the conversion of approximately 32% of the cellulose (ii) C.025% and decreases the hydrolytic enzyme requirements (ii) Obtained ethanol production in excess of 60 L/1000 kg fresh lemon peel biomass (i) The maximum glucose yield obtained was 91.252 m3 ton−1 biomass (i) Maximal bioethanol concentration (18.1 g/L (i) Increase of the sugar yield from 33 to 54% and reduction of the quantity of enzymatic mixture by 40% (i) The highest overall ethanol yield was obtained with the yeast Pachysolen tannophilus: yielded 163 mg ethanol per gram of raw wheat straw (23 and 35% greater) (i) Reuse existing assets to the maximum extent (ii) Keep the process as simple as possible (iii) Match the recalcitrance of the biomass with the severity of the pretreatment (i) Reduces the residual content of essential oils below 0.4 g/L (MTCC 7450) (ii) The hydrolyte obtained was 91.6 ± 1.69% (ii) Bamboo can be used as feedstock for the production of bioethanol (i) An ethanol yield based on total sugar of 480 g kg−1 was obtained (ii) Ethanol produced on marginal land at 0. enzymes used in hydrolysis. Reference Objectives [61] Optimal industrial symbiosis system to improve bioethanol production [62] Bioethanol production from dilute acid pretreated Indian bamboo variety by separate hydrolysis and fermentation [63] Fuel ethanol production from sweet sorghum bagasse using microwave irradiation [64] [65] [66] [67] [68] [69] Ultrasonic-assisted simultaneous SSF of pretreated oil palm fronds for bioethanol production Convert sucrose and homocelluloses in sweet sorghum stalks into ethanol Low-intensity pulsed ultrasound to increase bioethanol production Different process configurations for bioethanol production from pretreated olive pruning biomass Bioethanol production from water hyacinth Eichhornia crassipes Enhanced saccharification of biologically pretreated wheat straw for ethanol production [70] Fermentation of biologically pretreated wheat straw for ethanol production [71] Integration of pulp and paper technology with bioethanol production [72] Production of bioethanol by fermentation of lemon peel wastes pretreated with steam explosion [73] Ultrasonic-assisted enzymatic saccharification of sugarcane bagasse for bioethanol production [74] Status and barriers of advanced biofuel technologies [75] Sugarcane bagasse hydrolysis using yeast cellulolytic enzymes [76] Pretreatment of unwashed water-insoluble solids of reed straw and corn stover pretreated with liquid hot water to obtain high concentrations of bioethanol Main results (i) Reduced bioethanol production and logistic costs (ii) 2nd generation biomass should be used for bioethanol production (i) Bioethanol yield of 1.7 vol% was obtained (i) Yeast Saccharomyces cerevisiae TY2 produced ethanol at 9.1% (reed straw) and 71.2 g/L) and yield (57.28 g/L (reed straw) and 52.28% of the theoretical yield and the maximum amount of glucose obtained was 38.1% (corn stover) .26 g/L (corn stover) was obtained (ii) Ethanol yield reached a maximum of 69.0%) (i) All sugars in sweet sorghum stalk lignocellulose were hydrolysed into fermentable sugars (i) Increase of the production of bioethanol from lignocellulosic biomass to 52 ± 16% (i) Ethanol concentration of 3.The Scientific World Journal 3 Table 2: Recent literature about second generation bioethanol production.22% of the theoretical ethanol yield (MTCC 89) (iii) Decreases the reaction time (iv) The application of low intensity ultrasound enhanced the enzyme release and intensified the enzyme-catalysed reaction (i) The major barriers for the commercialization of 2nd generation ethanol production are the high costs of pretreatment.76% (v/v) with an efficiency of 41. laurentii is a good 𝛽-glucosidase producer (i) A high ethanol concentration of 56. Cellulose consists of long chains of 𝛽glucose monomers gathered into microfibril bundles.4% and 89. Softwood especially can hence be excluded. mostly xyloglucans or xylans. cellulolyticus gave ethanol yield 0. cellulose requires 320∘ C as well as high pressures (up to 25 MPa) to transform the rigid crystalline structure into an amorphous structure in water [12].1 g/L. and D-mannose) as well as pentose (D-xylose and L-arabinose). coniferyl. MG-60 [85] Furfural and xylose production from sugarcane bagasse in ethanol production lignin (10–25%). and Cu2+ ) increased the ethanol production (i) The furfural yield and xylose yield were 6 and 15. the released sugars can be fermented and the downstream process is similar to that of first generation feedstock [1].32% (13. loss of hemicelluloses. and chemical pulps represent the purest sources of cellulose. flax. Fe2+ . Hemicellulose is an amorphous structure formed of different heteropolymers including hexoses (D-glucose. respectively (ii) Ethanol was produced from the residual solid materials obtained from furfural and xylose at 87. Cotton. and removal or modification of lignin (i) Lowest environmental impact for second generation bioethanol production (ii) Highest exergy efficiency (steam explosion pretreatment + SSF + dehydration) reaching 79.. are linked to the microfibrils by hydrogen bonds. Reference Objectives [77] Waste paper sludge as a potential biomass for bioethanol production [78] Assessment of combinations between pretreatment and conversion configurations for bioethanol production [79] Combined use of gamma ray and dilute acid for bioethanol production [80] Ethanol production from lignocellulosic biomass (exergy analysis) [81] Alkaline pretreatment on sugarcane bagasse for bioethanol production [82] Influence of dual salt pretreatment of sugarcane bagasse for bioethanol production [83] Bioethanol production from alkaline pretreated sugarcane bagasse using Phlebia sp. The hemicelluloses. and synapyl alcohols).19 (g ethanol/g Solka floc) (i) The process based on dilute acid pretreatment and enzymatic hydrolysis and cofermentation combination shows the best economic potential (ii) The cellulose hydrolysis based on an enzymatic process showed the best energy efficiency (i) Increasing enzymatic hydrolysis after combined pretreatment is resulting from or decrease in crystallinity of cellulose.208 (g ethanol/g PS organic material) (ii) Consolidated biomass processing (CBP) technology gave ethanol yield 0. Once the lignocellulosic biomass is pretreated and hydrolysed. Lignins are phenolic compounds which are formed by polymerisation of three types of monomers (p-coumaryl. as shown in Table 4. while soft and hardwoods contain less than 50% of cellulose.16%) was obtained (ii) Cellulose content increased after alkaline pretreatment (i) Better performance was observed using H2 O2 with MnSO4 ⋅H2 O and ZnO (ii) The inhibitor formation was limited (iii) The maximum theoretical ethanol yield of 84. Although starches require temperatures of only 60–70∘ C to be converted from crystalline to amorphous texture. Potential lignocellulosic feedstocks and their composition are summarized in Table 3. The . MG-60 [84] Integrated fungal fermentation of sugarcane bagasse for bioethanol production by Phlebia sp. Its backbone chain is primarily composed of xylan linkages including 𝛼-xylose (∼90%) and L-arabinose (∼10%) [14].7% (4. Mn2+ . The extensive hydrogen linkages among Main results (i) SSF using cellulase produced by A.5 g/g of sugarcane bagasse. High lignin and/or high ash concentrations are unfavorable for bioethanol production.5 g/L) was achieved during the fermentation (i) 75% moisture content was suitable for subsequent ethanol production (ii) Some additives improved delignification in integrated fungal fermentation (IFF) (iii) Some inorganic chemicals (e. respectively cellulose molecules lead to a crystalline and strong matrix structure [11].g. It may contain sugar acids (uronic acids) [13]. Dgalactose.184 g/g sugarcane bagasse) was achieved during the fermentation (i) MG-60 produced cellulose and xylanase rapidly during consolidated bioprocessing (CBP) (ii) The maximum theoretical ethanol yield of 65.58% (i) The lowest lignin content (7. 0. Lignin adds compressive strength and stiffness to the cell wall [10].4 The Scientific World Journal Table 2: Continued.3%. 9 7.1 6.9 70.6 .08 53.8 40.4 0.02 51.1 19 18.7 56.7 5.1 70.1 69.5 41.8 70.6 6.4 6 1.1 0.2 6.3 6.8 17.2 5.7 6.22 45.1 12.7 0.7 1 0.7 6.2 14 16.1 12.8 0.8 49.4 0.88 43.6 25 43.4 6.3 8.4 3.4 34.1 14.6 10.9 17.7 0.8 Other biomass sources 60.2 19 11.4 43.6 0.4 0.09 49.5 0.15 49.9 15.80 0.9 0.02 52.7 74.01 42.8 12.3 13 11.4 43 5.2 1.8 6.7 41.4 0.2 3.6 7.26 15.4 16.10 49.1 20 14.8 6 0.4 0.1 43 5.4 7.7 0.4 0.3 2.2 0.7 4.17 51.7 6 0.8 8. ash) 12–16 0.8 43.9 9.08 49.7 0.5 0.6 15.4 71 71.8 43.2 0.9 6 0.10 50.1 0..1 5.3 8.5 73 78.7 44.2 44.7 42.1 0.5 35.16 49.3 45 40.1 13.1 0.2 0.1 1. A: ash.4 1.1 0.4 38.7 43.3 7 16.8 34.6 6.1 16.6 0.3 6.04 50 42.1 1.3 3.1 0.7 3.3 0.2 6 0.8 NA 50.01 51.8 7. Raw material Agricultural residues Hardwood Softwood Grasses Waste papers from chemical pulps Newspaper Switch grass a Hemicelluloses 25–50 25–40 25–29 35–50 12–20 25–40 30–35 Cellulose 37–50 45–47 40–45 25–40 50–70 40–55 40–45 Lignin 5–15 20–25 30–60 —a 6–10 18–30 12 Others (i.1 7.2 6 0. Sample Pine Eucalyptus bark Forest residue Land clearing wood Olive wood Pine chips Pine sawdust Poplar Mixed sawdust Spruce wood Willow Bamboo Miscanthus grass Sweet sorghum Switchgrass Corn straw Rice straw Wheat straw Coconut shell Cotton husks Corn stover Groundnut shell Hazelnut shell Olive husks Rice husks Soya husks Bagasse Sunflower husks Tea wastes Chicken litter Agricultural residue Mixed waste paper Refuse-derived fuel Sewage sludge Wood yard waste Ultimate analysis (wt% on dry basis) C O H N S Wood and woody biomass 54.1 71.1 0.5 1.1 3.50 2–5 2 5–8 4-5 Not present or not available.4 3.5 1.4 7 11.8 3.2 6.2 40.3 VM: volatile matter.4 0.4 59.1 0.1 43.8 70.04 49.6 20.7 43.2 1.9 17.4 7.04 49.1 18.8 2.3 7.4 0.5 5.3 6.3 0.2 1.2 52. 87].3 43.4 7.5 1.42 48. VM Proximate analysis M FC A 46.1 2 0.4 39.1 0.3 22 16.1 48.4 8.5 9.7 0.33 52.08 50.09 51 42.1 4.3 40.9 10.4 3 5.7 76.e.2 37.3 30.1 5.3 5.1 50.5 42.6 76.1 5.8 1.8 67.5 5.8 14.5 73.9 2.1 6.04 52.4 6.11 48.1 0.9 6.4 0.7 6.4 41.3 54.1 75.3 10. M: moisture.5 7.1 68.7 34.08 50.3 66.8 12 56.6 42.5 0.4 9.1 68.9 0.2 0.6 5.7 0.8 0.1 0.9 71.9 5.1 0.47 50.8 4.4 79.03 52.1 2.5 38.6 0.2 4.6 7.7 0.5 6.4 6.4 46.3 41.5 0.4 4.6 69.4 17.2 70.7 0.9 7.1 0.07 49 44. FC: fixed carbon.6 0.06 Herbaceous and agriculture biomass 52 42.9 33.4 0.9 6 0.9 40.5 1.8 36.8 43.4 74.3 17.2 11.6 5 0.4 2.6 6.2 7.7 45.02 49.2 6.2 6.7 55.9 6.05 52.9 19.7 10.4 67.1 0.57 0.8 10.7 6.3 12.5 25.The Scientific World Journal 5 Table 3: Potential lignocellulosic biomass sources and compositions (% dry weight) [86.1 7. Table 4: Ultimate and proximate analyses of different biomasses (wt%).6 41. This certainly eliminates most of the soft woods. Some typical values of the C-. since most of the ash will concentrate in the lignin residue. Pretreatment and First Stage Hydrolysis. physicochemical. The ash content of biomass sources is generally low. The elements in the ash are O. as described in Section 3. which is again catalysed by dilute acid. and O are the key components of biomass and largely determine their calorific value. C. membrane techniques have been investigated [3. Processing of Biomass to Ethanol 2. Al. The phenolic groups. Second Stage Hydrolysis. However. Pretreatment is a costly separation. Its composition should. as illustrated in Table 4. Lignin components are gaining importance because of their dilution effect on the processes of hydrolysis and fermentation [16. much of the sulphuric acid and other inhibitors produced during the degradation of the materials need to be removed. The main steps are summarized in Figure 1. K.2. 2. Steam explosion has a few limitations since the lignin-carbohydrate matrix is not completely broken down. The ash content and its chemical composition are a strong function of the biomass species. The Scientific World Journal 2. The exact composition is largely dependent on the biomass sources. it needs to be carefully stored and conditioned to prevent early fermentation and bacterial contamination. Genetic manipulation could modify the metabolic routes to produce bioethanol or other value-added compounds in an efficient manner. Biomass feedstock with a high lignin content is not readily applicable as raw material for the bioethanol fermentation. the most common methods are steam explosion and dilute acid prehydrolysis. To be totally hydrolysed into free monomers. Cl. which are followed by enzymatic hydrolysis. formed from the degradation of lignin. 2. and a portion of the xylan fraction is destroyed. 20] and own analyses. H.111 kg per kg of glucan and xylan. 23] and some examples are summarized in Table 5. however. Chen et al. using physical. P. that is. either produced in a separate reactor or bought externally from industrial suppliers [27– 30]. An extensive literature survey was provided by several authors [19. Through pretreatment. respectively. The use of dilute sulphuric acid (0. coniferyl. . Cl. mannose.3. and the release of microbial and chemical contamination that possibly reduces the overall yield needs further attention. concentrated acid. substantially deactivate cellulolytic enzymes and hence hamper enzymatic hydrolysis. Al. but also other compounds of the cellulosic matrix can however inhibit the enzymatic hydrolysis and fermentation. and galactose). as a result of an extensive literature survey [9. Lignin is an aromatic and rigid biopolymer. chemical. This could however be problematic as lignin components serve as the major plant defence system to pathogens and insects and its modification could disrupt the plants’ natural protection [18]. Ca. The different composition of biomass feedstock (dry) is also reflected in its elemental composition. 17].136 kg and 1. H-. Na. The lignin fraction in biomass sources varies considerably. Generalities. and neutralisation is performed before fermentation. Hydrolysis of mycoLB (LB after fungal pretreatment) has been recognized as a promising approach to avoid fermentation inhibitors and improve total sugar recovery. as illustrated in Table 5. Fe. 31]. the released cellulose of the biomass is converted into glucose. coumaryl. Mg.1. and O-content. Chandel et al. 433–463 K for 10 minutes) has the preference of the US National Renewable Energy Laboratory [25]: hemicellulose is largely hydrolysed releasing different simple sugars (xylose.5–1%. covalently bonded to hemicellulosic xylans and responsible for the rigidity and high level of compactness of the plant cell wall [15]. accounting for approximately 33% of the total cost [26]: the economy needs to be improved. and biological treatment (Table 6). 22. and Na. and Si as well as lower concentrations of Ca. Biomass contains a significant content of K. or preferably by cellulase enzymes. simple sugars are made available in proportions depending on the type of biomass used and the pretreatment process. thus being a potential energy self-sustaining source of the process. Sulphuric acid or carbon dioxide is often added in order to reduce the production of inhibitors and improve the solubilisation of hemicellulose [15]. To overcome inhibition by hydrolyte components. be taken into consideration. Si. S. Carbonic acid and alkaline extraction have the best performance. [17] however demonstrated that lignin modification via genetic engineering could considerably reduce lignin formation and improve ethanol yield. Mg. In the second stage hydrolysis. providing a general production flow sheet. Once the feedstock is delivered to the ethanol plant. since once recovered. Pretreatment involves delignification of the feedstock [24] in order to make cellulose more accessible in the hydrolysis step. The conversion of cellulose and hemicellulose can be expressed by the reaction of glucan (for hexoses) and xylan (for pentose) with water: (C6 H10 O5 )𝑛 + 𝑛H2 O → 𝑛C6 H12 O6 (1) (C5 H8 O4 )𝑛 + 𝑛H2 O → 𝑛C5 H10 O5 (2) The maximum theoretical yield of hexoses and pentoses is 1. and sinapyl alcohol. and Ti [21]. hemicellulose requires a wide range of enzymes in view of the diversity of its sugars. Retaining the lignin could moreover benefit the energyeconomy of the process. Mn. Fe. Lignin is composed of monomers of phenyl propionic alcohol. degradation products are generated that reduce the efficiency of the hydrolysis and fermentation steps.6 degree of branching and the xylan composition vary with the nature and the source of raw materials. arabinose. as well as other essential data are summarized in Table 4. Part of the acetic acid. it can be applied in a combined heat and power unit (CHP). thus possibly hampering further energy generation by fouling or sintering. Further research is certainly required. [32] investigated the strategies that have been adopted to detoxify lignocellulosic hydrolysates and their effects on the chemical composition of the hydrolysates to improve the fermentability of lignocellulosics. 19. The conversion reaction for hexoses (C6) and pentoses (C5) is as follows: C6 H12 O6 → 2C2 H5 OH + 2CO2 (3) 3C6 H10 O5 → 5C2 H5 OH + 5CO2 (4) Wastes Investment Very low Low Very low Very low Low High High Low High High High Low Low Moderate High Low Low Very high The theoretical maximum yield of broth hexoses and pentoses is 0.47 0.91 15.02 2.03 0.79 23.43 1. S.75 7.01 8.96 4. Pretreatment process (i) Mechanical (ii) Steam explosion (iii) Ammonia fiber explosion (AFEX) (iv) Carbonic acid (i) Dilute acid (ii) Concentrated acid (iii) Alkaline extraction (iv) Wet oxidation (v) Organosolv Yield of fermentable sugars Physical or physicochemical Low High Moderate Very high Chemical Very high Very high Very high High Very high 2.12 4.47 0. Contrarily to the conversion of disaccharides and starch to ethanol. cannot metabolize xylose.17 Fe2 O3 SO3 Na2 O TiO2 1. switchgrass) into fermentable sugars [33.25 1.03 4.21 4. or fungi.35 94..25 50.23 0.49 3.88 12.05 2.38 20.01 4. Sample SiO2 CaO Eucalyptus bark Poplar bark Willow Wood residue 10.54 0.47 0.71 24.74 77.81 13. Additional research tried to find microorganisms which can effectively .g.06 0. 35].47 0.03 4. such as bacteria.28 1.01 0. and ETEK (Sweden) [34] have built pilot plants capable of producing a few hundred thousand litres of ethanol per year. Genetically engineered fungi that produce large volumes of cellulase.45 1.48 46.53 7.6 4.28 60.92 56.91 2.02 0.11 0.77 14.33 Other biomass varieties 12.25 1.41 10.62 2.04 1.92 3.12 0.57 4.57 19.67 6. respectively. Fermentation is the biological process to convert the hexoses and pentoses into ethanol by a variety of microorganisms. modern lignocellulose-to-ethanol processes are at pilot and demonstration stage: NREL (USA) [25].83 4.24 0.4 13.54 0.9 0.511 kg ethanol and 0.74 3 1.16 1.47 0.67 0.61 0.77 7.97 4.98 3.4.01 1.19 15. Fermentation.74 0.66 4.64 3.06 0.31 Chicken litter Mixed waste paper Refuse-derived fuel Sewage sludge Wood yard waste 5.45 0.91 8.52 0.64 3.79 3. Iogen Corporation (Canada) [33].19 6.2 53.95 3.86 4.89 3.68 2.31 0.7 1.74 2.6 15.81 3.32 Table 6: Assessment of selected pretreatment processes [15.22 11. These could convert agricultural residues (e.92 K2 O P2 O5 Al2 O3 MgO Wood and woody biomass 9.49 2.63 26.31 46.54 1.36 0. yeast.8 0.67 33.1 56. the yeast commonly used for first generation ethanol production.88 0.37 1.The Scientific World Journal 7 Table 5: Elemental ash composition of different biomass.54 1.04 23.06 Herbaceous and agriculture biomass 53. which are mature technologies.15 57.13 8.. and sugar cane bagasse) and energy crops (e.85 1.29 0.02 0.73 3.86 6.g.46 0.48 0.93 2.08 2. and hemicellulase enzymes are under investigation.53 2.28 0.75 5.35 3.1 53.74 6.95 0.58 3.46 10.56 28.42 73. The overall theoretical ethanol yield (at 20∘ C) hence becomes 0.85 66.8 0.12 9.09 11.21 0. straw.26 15.66 Bamboo whole Miscanthus Sorghum grass Sweet sorghum Switchgrass Wheat straw Rice husks Sugar cane bagasse Sunflower husks 9. cerevisiae.27 3.47 0.736 liters per kg of glucan (and/or other 6C structures) and xylan (and/or other 5C structures).01 2.15 0.77 28.22 4.84 1.29 2.09 0.37 12.57 0. 88–96].719 and 0.61 4.33 0.489 kg CO2 per kg sugar. Other yeasts and bacteria are under investigation to ferment xylose and other pentoses into ethanol.02 10.59 1. xylanase.16 0.99 1.21 8.1 10.62 38.07 0.54 2.87 14.6 0.24 3.21 66.94 0.82 6.21 0.83 2.11 3.58 1.01 0.98 1.36 2.4 1.98 3.54 6.4 0.97 4.85 7.12 0.92 2. corn stover.12 0.36 23. that need to be dealt with to operate hydrolysis and fermentation under optimum conditions and maximum conversion. different integration methods of hydrolysis and fermentation steps are proposed. such as furfural and 5-hydroxymethylfurfural (5-HMF). the consolidated bioprocessing (CBP) proceeds by producing all required enzymes and ethanol using a single type of microorganisms in a single reactor. 39]. adapted yeasts or bacteria can be used. Unfortunately. and syringaldehyde. Separate hydrolysis and cofermentation (SHCF) and simultaneous saccharification and cofermentation (SSCF) are possible alternatives: cofermenting both C5 and C6 sugars by a single strain of microorganisms in the same reactor significantly improves the process economics and enhances the commercial production of lignocellulosic ethanol in the short term [39–41]. such as 4hydroxybenzaldehyde (4-HB). Although this facilitates both the optimization of each separate reactor and the selection of sugar-appropriate microorganisms to ferment the different sugars. furans. and phenols. since it implies neither capital nor operating costs for dedicated enzyme production together with a reduced consumption of substrate for enzyme production. the liberated cellulose is treated in a different reactor for hydrolysis and subsequent fermentation than the hydrolysed hemicellulose and lignin. CBP is considered as the ultimate evolution of biomassto-bioethanol conversion technology. When using enzymatic hydrolysis. Research has been done on more promising yeasts and bacteria: Zymomonas mobilis succeeds . 37]. for example. chemical inhibition is a more severe problem than encountered in first generation raw materials. With bioethanol production from lignocellulosic biomass. Pretreatment and hydrolysis of lignocellulosics release specific inhibitors. and Zymomonas mobilis as promising candidates [36.8 The Scientific World Journal Lignocellulosic 2nd Harvesting Pretreatment ST Hydrolysis 1st stage ST Hydrolysis 2nd stage CO2 CHP Wastes Fermentation Waste water treatment ST ST Distillation ST Drying to anhydrous ethanol Figure 1: Second generation biomass-to-ethanol production (ST: steam addition). Klebsiella oxytoca. vanillin. A novel development. The most commonly used yeast is Saccharomyces cerevisiae. the higher investment costs for two separate reactors and the inhibition of the high glucose concentration to fermenting organisms are major disadvantages [38. To increase the critical ethanol-inhibition concentration. ferment both types of sugars into ethanol with Escherichia coli. In the separate hydrolysis and fermentation (SHF). it is predicted that it will take several years of research to determine such microorganisms or compatible combinations of microorganisms [41]. with a moderate yield of fermentation. 5 wt%. or adaptive mutation to produce acetic acid resistant strains of Z. and sucrose. To overcome inhibition by hydrolyte components. although further research is certainly required.. This system allows the plant to be self-sufficient in energy supply. simultaneous saccharification and combined fermentation (SSCombF) of the enzymatic hydrolyzate. 9 Pretreatment. mobilis [42. well above the tolerance threshold of Z. in spite of these attractive advantages. thus preventing their industrial application. makes this a very popular yeast for industrial application. but resulted in low ethanol concentrations and produced acetic acid.6. Unlike yeast. [1]. 3. and coculturing [45]. Z. Genetic engineering has succeeded in altering the conventional S. for softening (polyvalent cation removal) and removal of disinfection byproduct precursors such as natural organic matter and synthetic organic matter. 2. were isolated from sugarcane molasses. However. or reverse osmosis dealing with dissolved contaminants. As described before. It was already stated that membrane techniques could help to overcome some of the problems. some groups of bacteria such as Zymomonas mobilis can convert sugars into ethanol [47]. In ultrafiltration (UF). mobilis is indeed that its plasma membrane contains hopanoids. around 16 wt%. the yield and productivity are much lower. Almeida et al. not only capable of cofermenting . as described in Section 3.The Scientific World Journal to survive higher ethanol concentrations in the fermenter up to 16 vol%. fructose. Burning the solid residues for steam and power production is the most beneficial option and meets the energy demand of the plant. Typical ethanol concentrations are in the range of 3–6 vol% only. cerevisiae’s capacity to ferment glucose and pentose sugars simultaneously [48]. gene expression. cerevisiae. Better and cost-efficient pretreatment techniques need further investigation. together with methods to reduce or eliminate microbial and chemical contaminants that can reduce the yields. Ultrafiltration is used in industry and research for purifying and concentrating macromolecular (103 –106 Da) solutions. [3]. and generates additional revenue through sales of excess electricity [39. 2. Biomass to bioethanol will only be a technical and economic viable alternative to first generation bioethanol. Some natural wild yeast species appear capable of replacing S. with microfiltration (suspended solids) and ultrafiltration. Steam and Electricity Generation. mobilis cannot tolerate toxic inhibitors present in lignocellulosic hydrolytes such as acetic acid. while water and low molecular weight solutes pass through the semipermeable membrane in the permeate. The bottom product of the first distillation column (stillage) contains mainly lignin and water next to unconverted cellulose and hemicellulose. mobilis to overcome its inherent deficiencies by metabolic engineering. 44].g. T. Enhancing ethanol production by pretreatment involving fungi (e. typical inhibitors present in biomass hydrolysis. very low in comparison with 12 to 15 vol% obtained from 1st generation feedstock [1]. Nanofiltration and reverse osmosis are high-pressure membrane filtration processes used most often with low total dissolved solids water (surface water and fresh groundwater). but they are more vulnerable to chemical inhibition than S. if appropriate solutions are developed. and Dekkera. reduces solid waste disposal cost. especially protein solutions. and CBP are also considered to be cost-effective whilst reducing end-product inhibition. The fermentation rate is also higher with Zymomonas mobilis in comparison to Saccharomyces cerevisiae [1]. An interesting characteristic of Z. solutes of high molecular weight are retained in the so-called retentate. It cannot ferment C5 sugars like xylose and arabinose which are important components of lignocellulosic hydrolytes. However. [49] investigated a modified S. mobilis. Different process improvements. Removal of suspended solids prior to feeding the membrane is essential to prevent damage to the membrane and minimize the effects of fouling which greatly reduce the separation efficiency. Some genera. Purification. including energy pinch. 43]. several bottlenecks emerge. such as Candida. While fungi act slowly. Current production problems hence determine immediate and future research priorities. The development of genetically modified fermentative and cellulolytic microorganisms is recommended to increase the ethanol yield and productivity under the stress conditions of high production bioethanol processes [47]. membrane techniques have been investigated.5. Several attempts have been made to engineer Z. but their low bioethanol yield and poor survival in the fermenter need further improvement. cerevisiae. an inhibitor of the fermentative yeast [46]. but also a moderate tolerance for acids and sugars. Current Research Priorities in Biomass to Ethanol From the previous process assessment. mutagenesis. nanofiltration. Simultaneous saccharification and fermentation (SSF). accounts for about 33% of the total cost [26]. are described in detail by Kang et al. additional distillation efforts are required. reesei and Basidiomycetes) with appropriate lignocellulolytic properties at low pH and high temperatures is also a promising and added-value step in SSF ethanol bioconversion. and hybrid processes. thus providing an extraordinary tolerance to ethanol in its environment. The possible application of microfiltration to eliminate suspended solids has recently been confirmed by Kang et al. Pichia. its substrate range is limited to glucose. Not only this advantage. These membrane separation technologies have been examined in different stages of the bioethanol production. Concentration of acetic acid in lignocellulosic hydrolytes can be as high as 1. very high gravity fermentation. This insoluble fraction is dewatered by a pressure filter and sent to a fluidized bed combustor system for steam and electricity generation. as the first step. pentacyclic compounds similar to eukaryotic sterols. Due to the higher water content of the broth. when these engineered strains metabolize mixed sugars in the presence of inhibitors. potential lignocellulolytic fungi have been produced by mutagenesis. cerevisiae in second generation bioethanol [47]. Bi.47 g/g ethanol. xylose and arabinose [97]. that is. Reconstitution of the N. especially with respect to the CBP option [59. mobilis and S. 4. Fermentation of C5 and C6 sugars needs adapted microorganisms. R. Acknowledgments This work was supported by the Chinese Science Council in support of the delegation of the principal author to the KU Leuven. This needs to be complemented by a comprehensive systems approach. 60]. is required to design new strategies for increasing the ethanol fermentation performance. Lignin should be considered as a valuable energy source.” Advances in Bioscience and Biotechnology. “Performance and emission characteristics of diesel engine fueled with ethanol-diesel blends in different altitude regions. “Energyefficient production of cassava-based bio-ethanol.365 g/g. New combined processes reduce both the number of operation steps and the production of chemical inhibitors. cerevisiae fermentation has already been proven for first generation large-scale bioethanol production. K. 2014. Conflict of Interests The authors declare that there is no conflict of interests regarding the publication of this paper. Genetic engineering. encompassing the different individual steps and accounting for all inputs and outputs during the entire operation regardless of modifications in any of these individual steps. Kang.g. and T. and proving to be an ideal gene modified organism (GMO) for CBP processes [58]. respectively. It improved ethanol yield when compared to fermentation with either cellobiose or xylose as sole carbon sources. Tan. Industrial operations using antibiotics to control microbial contaminants in fermenters or as strain markers would generate and release antibiotic resistant organisms and offer another potential environmental and public health risk. The final ethanol production of 1. The resulting consortium was demonstrated to utilize phosphoric acid swollen cellulose (PASC) for growth and ethanol production. Ge et al. J. CBP combines hydrolysis and fermentation operations in a single reactor. Its genetic improvement is gaining increasing research. Since fermentative microorganisms must be capable of surviving the high temperatures of SSF/SSCombF/CBP processes. 55]. Second generation bioethanol could surpass the traditional first generation processes. Such high temperature operations do not require cooling and cellulase addition [57]. The ethanol yield for HDY-ZMYWBG1 and HDY-ZMYWBG3 is 0. Appels. 92. namely.” Journal of Biomedicine and Biotechnology. This avoids the costs related to the purchase of cellulolytic enzymes [50]. Conclusions The cellulosic bioethanol production process involves specific processing steps. T. mobilis are promising for higher alcohol tolerance and conversion efficiency. [2] L. especially in the pretreatment and hydrolysis. Baeyens. marxianus was moreover genetically modified to exhibit T. marxianus. [56] engineered yeasts to coferment mixtures of xylose and cellobiose. vol. Ha et al. has been documented as a candidate for its ability to coferment both hexose and pentose sugars and survive temperatures of 42–45∘ C [58]. The thermotolerant yeast. crassa cellodextrin transport system in Saccharomyces cerevisiae promotes efficient growth of this yeast on cellodextrins [54. Shen. [52] obtained three recombinants: HDY-ZMYWBG1. 5. cerevisiae cells.25 g/L corresponded to 87% of the theoretical value and was 3-fold higher than a similar yeast consortium secreting only the three cellulases [53]. provided present processing bottlenecks are removed and the best combination of advanced systems is used. further research is required: high temperature ethanol fermentation is an emerging technology provided appropriate microorganisms can be developed. J. K. where hydrolysis and substrate fermentation are possible in a single step. 10 pages. used as a fuel in a CHP and being capable of supplying the power and heat requirements of the complete conversion process. still to be further investigated. cerevisiae have proven high ethanol yield and adaptability [98]. . Improvement in each of these individual aspects is required to achieve high conversion and cost-effective biomass-to-bioethanol operations. as a powerful biotechnological tool. no. Z. 12. reesei and Basidiomycetes) that exhibit lignocellulolytic properties at low pH levels and high temperature. The use of GMOs is questionable.368 g/g and 0. The engineered yeast strains more rapidly convert cellulose to ethanol when compared with yeast lacking this system..2% of the theoretical yield. 925–939. Dewil. As mentioned before. HDY-ZMYWBG2. The industrial potential for S. Both the gene engineered Z. Article ID 417421. Upregulation of stress tolerance genes by recombinant DNA technology can be a useful approach to overcome inhibitory situations [51]. and HDY-ZMYWBG3 using the lithium acetate transformation method into the S. cerevisiae and Z. and Y. 2011. mobilis remains an attractive candidate due to its high ethanol yield and resistance to temperatures in the range of 40∘ C [36]. Further research is certainly required in optimizing biological pretreatment involving fungi (e. Numerous genes have been introduced and heterologous expression has been incorporated into Z. 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