review article bioethanol from lignocellulosic biomass...

Review ArticleBioethanol from Lignocellulosic BiomassCurrent Findings Determine Research Priorities

Qian Kang12 Lise Appels2 Tianwei Tan1 and Raf Dewil2

1Key Lab of Bioprocess College of Life Science and Technology Beijing University of Chemical Technology Beijing 100029 China2Department of Chemical Engineering Process and Environmental Technology Lab KU Leuven 2860 Sint-Katelijne-Waver Belgium

Correspondence should be addressed to Qian Kang kangqian1985gmailcom

Received 9 November 2014 Accepted 18 December 2014 Published 31 December 2014

Academic Editor Reeta R Singhania

Copyright copy 2014 Qian Kang et al This is an open access article distributed under the Creative Commons Attribution Licensewhich permits unrestricted use distribution and reproduction in any medium provided the original work is properly cited

ldquoSecond generationrdquo bioethanol with lignocellulose material as feedstock is a promising alternative for first generation bioethanolThis paper provides an overview of the current status and reveals the bottlenecks that hamper its implementation The currentliterature specifies a conversion of biomass to bioethanol of 30 to sim50 only Novel processes increase the conversion yield to about92 of the theoretical yield New combined processes reduce both the number of operational steps and the production of inhibitorsRecent advances in genetically engineeredmicroorganisms are promising for higher alcohol tolerance and conversion efficiency Bycombining advanced systems and by intensive additional research to eliminate current bottlenecks second generation bioethanolcould surpass the traditional first generation processes

1 Introduction

11 Bioethanol as Sustainable Fuel With the global increasingdemand for energy energy shortage will be a global problemBioethanol is considered as an important renewable fuel topartly replace fossil-derived fuels The world production ofbioethanol increased from 50 million m3 in 2007 to over 100million m3 in 2012 [1] Brazil and the United States representapproximately 80 of the world supply mostly using corn orsugarcane In developing economies food-related feedstockis preferably replaced by nonfood raw materials such assweet sorghum or cassava The use of common biomasscould significantly increase the bioethanol production andlignocellulose-based bioethanol is therefore the topic of thepresent review paperThe current technological developmentand bottlenecks define the short- and medium-term researchpriorities

Industrial ethanol is mainly produced petrochemicallythrough the acid-catalyzed hydration of ethylene Ethanol foruse in alcoholic beverages and the vast majority of ethanolfor use as biofuel is produced by fermentation where certainspecies of yeast (eg Saccharomyces cerevisiae) or bacteria

(eg Zymomonas mobilis) metabolize sugars in oxygen-leanconditions to produce ethanol and carbon dioxide

The main reasons for the enhanced development ofbioethanol are its use as a favourable and near carbon-neutral renewable fuel thus reducing CO

2emissions and

associated climate change its use as octane enhancer inunleaded gasoline and its use as oxygenated fuel-mix fora cleaner combustion of gasoline hence reducing tailpipepollutant emissions and improving the ambient air qualityThe largest single use of ethanol is as engine fuel and fueladditive with common types of available fuel-mixes listed inTable 1

Ethanol has appropriate properties for spark ignition ICengines Its octane numbers motor octane number (MON)and research octane number (RON) are 90 and 109 respec-tively on average 99 compared to 91 for regular gasoline Dueto its low cetane number ethanol does not burn efficiently bycompression ignition and ismoreover not easilymisciblewithdiesel fuel To improve the use of ethanol in compression-ignition (CI) engine vehicles measures can be taken suchas the addition of an emulsifier in order to increase theethanol-diesel miscibility the addition of ethylhexyl nitrate

Hindawi Publishing Corporatione Scientific World JournalVolume 2014 Article ID 298153 13 pageshttpdxdoiorg1011552014298153

2 The Scientific World Journal

Table 1 Common ethanol-petrol mixtures [1 31]

Code Composition Countries CommentsE5 Max 5 anhydrous ethanol min 95 petrol Western Europe

Blends for regular carsE10 Max 10 anhydrous ethanol min 90 petrol USA EuropeE15 Max 15 anhydrous ethanol min 85 petrol USA cars gt2000E25 Max 25 anhydrous ethanol min 75 petrol BrazilE85 Max 85 anhydrous ethanol min 15 petrol USA Europe Flex-fuel vehiclesE100 Hydrous ethanol (sim53 wt water) Brazil

or diterbutyl peroxide to enhance the cetane number theuse of a dual fuel operation in which ethanol and diesel areintroduced separately into the cylinder or the modificationof diesel engines in order to adapt their characteristics ofautoignition [2]

12 Bioethanol from Different Feedstocks Fermentation ofsugar-based raw materials is referred to as ldquofirst generationrdquobioethanol whereas the use of lignocellulose raw materials iscommonly called ldquosecond generationrdquo bioethanol The ldquothirdgenerationrdquo of algal bioethanol is at an early stage of investi-gationThe first generation processes were discussed in detailby Kang et al [1] Since the present paper deals with secondgeneration bioethanol relevant and recent (gt2010) literatureis summarized in Table 2 The 2nd generation bioethanolprocesses will use cellulose-released sugars despite the costof the required enzymes to hydrolyse cellulose Developmentof this technology could deal with a number of cellulose-containing agricultural byproducts such as straw woodtrimmings sawdust bamboo and others

The specificities of using lignocellulosic raw materialswill be dealt with in Section 13 Whether first second orthird generation feedstock is used fermentation producesan alcohol-lean broth only as such unusable in industrialand fuel applications The ethanol must hence be purifiedFractional distillation can concentrate ethanol to 956 vol(895mol) corresponding to the azeotropic compositionwith a boiling point of 782∘C Further ethanol enrichment bycommon distillation is impossible but different alternativeshave been investigated as reported by Kang et al [3]

13 Lignocellulosic Biomass

131 Sources Available biomass can be categorized intoprimary sources produced as either crop or key product forexample sugar cane short rotation energy plantations sec-ondary sources as residues from the production processesfor example bagasse rice husks and straw and tertiarysources as residues produced during and after applicationend for example the organic fraction of municipal solidwaste (MSW) sewage treatment sludge wood trimmingsand so forth [4] In general the final availability of organicwastes and residues may fluctuate and is affected by marketgrowth although climate and other factors have influencesespecially when considering the primary sourcesThe energypotential of biomass residues and organic wastes depends

on the yield the total land area available and the type ofproduction

The organic fraction of municipal solid waste (MSW)is an inexpensive source of biomass and covers domesticand industrial waste collected in a specific area The overallpotential of the organic fraction of municipal waste and itswaste wood fraction is strongly reliant on economic growthconsumption and the use of biomaterials It is estimated atbetween 5 and 50 EJyear [4] The unit EJ (eta-joule) is equalto 1018 J

Agroindustrial biomass residues are byproducts of agri-culture or its related industry including cotton stalks wheatand rice straw coconut shells maize cobs jute sticks andrice husks [5] The agricultural residues are produced decen-tralised andhave a lowdensityDue to the high transportationcost it is expensive to apply agricultural residues as the mainfuel in power stations The potential of agriculture residuesvaries from 15 up to 70 EJyear [4] as a function of regionalproduction harvesting processing factors and recoverabilityfactors Within the agroindustrial residues dried manure isconsidered as a tertiary source A total worldwide estimate isdifficult to make and given as 5 to 55 EJyear with the lowerestimate due to the current use as fertiliser while the higherestimate considers the total technical potential [5]

Forestry residues include biomass not harvested orremoved from sorting regions in commercial hardwood andsoftwood production through forestmanagement operationssuch as precommercial thinning and removal of dead anddying trees Forestry waste includes wood chips sawdustand bark It can provide 65 of the biomass energy potential[6 7] The extraction costs and the required transportationto centralized processing plants make forest fuels expen-sive Several studies have focused on their use for energyproduction at a district level applying appropriate designsof decentralized smaller plants for example Malinen et al[8] and Demirbas [9] The energy potential of the worldrsquosforests is again difficult to estimateThe possible contributionby 2050 is estimated at 98 EJyear of excess natural forestgrowth and at 32ndash52 EJyear of processing residues Althoughthese lignocellulosic biomass resources represent a signifi-cant energy value sim150 EJyear only part of this resourcecan be used as feedstock for bioethanol for reasons givenbelow

132 Composition Lignocellulose the principal componentof the plant cell walls is mainly composed of cellulose (40ndash60 of the total dry weight) hemicellulose (20ndash40) and

The Scientific World Journal 3

Table 2 Recent literature about second generation bioethanol production

Reference Objectives Main results

[61] Optimal industrial symbiosis system to improve bioethanolproduction

(i) Reduced bioethanol production and logistic costs(ii) 2nd generation biomass should be used for bioethanolproduction

[62] Bioethanol production from dilute acid pretreated Indianbamboo variety by separate hydrolysis and fermentation

(i) Bioethanol yield of 176 (vv) with an efficiency of 4169(ii) Bamboo can be used as feedstock for the production ofbioethanol

[63] Fuel ethanol production from sweet sorghum bagasse usingmicrowave irradiation

(i) An ethanol yield based on total sugar of 480 g kgminus1 wasobtained(ii) Ethanol produced on marginal land at 0252m3 tonminus1biomass

[64] Ultrasonic-assisted simultaneous SSF of pretreated oil palmfronds for bioethanol production

(i) Maximal bioethanol concentration (182 gL) and yield(570)

[65] Convert sucrose and homocelluloses in sweet sorghumstalks into ethanol

(i) All sugars in sweet sorghum stalk lignocellulose werehydrolysed into fermentable sugars

[66] Low-intensity pulsed ultrasound to increase bioethanolproduction

(i) Increase of the production of bioethanol fromlignocellulosic biomass to 52 plusmn 16

[67] Different process configurations for bioethanol productionfrom pretreated olive pruning biomass (i) Ethanol concentration of 37 vol was obtained

[68] Bioethanol production from water hyacinth Eichhorniacrassipes

(i) Yeast Saccharomyces cerevisiae TY2 produced ethanol at96 plusmn 11 gL

[69] Enhanced saccharification of biologically pretreated wheatstraw for ethanol production

(i) Increase of the sugar yield from 33 to 54 and reductionof the quantity of enzymatic mixture by 40

[70] Fermentation of biologically pretreated wheat straw forethanol production

(i) The highest overall ethanol yield was obtained with theyeast Pachysolen tannophilus yielded 163mg ethanol pergram of raw wheat straw (23 and 35 greater)

[71] Integration of pulp and paper technology with bioethanolproduction

(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 severityof the pretreatment

[72] Production of bioethanol by fermentation of lemon peelwastes pretreated with steam explosion

(i) Reduces the residual content of essential oils below0025 and decreases the hydrolytic enzyme requirements(ii) Obtained ethanol production in excess of 60 L1000 kgfresh lemon peel biomass

[73] Ultrasonic-assisted enzymatic saccharification of sugarcanebagasse for bioethanol production

(i) The maximum glucose yield obtained was 9128 of thetheoretical yield and the maximum amount of glucoseobtained was 384 gL (MTCC 7450)(ii) The hydrolyte obtained was 9122 of the theoreticalethanol yield (MTCC 89)(iii) Decreases the reaction time(iv) The application of low intensity ultrasound enhanced theenzyme release and intensified the enzyme-catalysed reaction

[74] Status and barriers of advanced biofuel technologies

(i) The major barriers for the commercialization of 2ndgeneration ethanol production are the high costs ofpretreatment enzymes used in hydrolysis and conversion ofC5 sugars to ethanol(ii) The residues need to be processed for byproducts throughbiorefinery to improve the economics of the whole process

[75] Sugarcane bagasse hydrolysis using yeast cellulolyticenzymes

(i) This enzyme extract promoted the conversion ofapproximately 32 of the cellulose(ii) C laurentii is a good 120573-glucosidase producer

[76]Pretreatment of unwashed water-insoluble solids of reedstraw and corn stover pretreated with liquid hot water toobtain high concentrations of bioethanol

(i) A high ethanol concentration of 5628 gL (reed straw)and 5226 gL (corn stover) was obtained(ii) Ethanol yield reached a maximum of 691 (reed straw)and 711 (corn stover)


Table 2 Continued


[77] Waste paper sludge as a potential biomass for bioethanolproduction

(i) SSF using cellulase produced by A cellulolyticus gaveethanol yield 0208 (g ethanolg PS organic material)(ii) Consolidated biomass processing (CBP) technology gaveethanol yield 019 (g ethanolg Solka floc)

[78] Assessment of combinations between pretreatment andconversion configurations for bioethanol production

(i) The process based on dilute acid pretreatment andenzymatic hydrolysis and cofermentation combination showsthe best economic potential(ii) The cellulose hydrolysis based on an enzymatic processshowed the best energy efficiency

[79] Combined use of gamma ray and dilute acid for bioethanolproduction

(i) Increasing enzymatic hydrolysis after combinedpretreatment is resulting from or decrease in crystallinity ofcellulose loss of hemicelluloses and removal or modificationof lignin

[80] Ethanol production from lignocellulosic biomass (exergyanalysis)

(i) Lowest environmental impact for second generationbioethanol production(ii) Highest exergy efficiency (steam explosion pretreatment+ SSF + dehydration) reaching 7958

[81] Alkaline pretreatment on sugarcane bagasse for bioethanolproduction

(i) The lowest lignin content (716) was obtained(ii) Cellulose content increased after alkaline pretreatment

[82] Influence of dual salt pretreatment of sugarcane bagasse forbioethanol production

(i) Better performance was observed using H2O2 withMnSO4sdotH2O and ZnO(ii) The inhibitor formation was limited(iii) The maximum theoretical ethanol yield of 8432(131 gL 0184 gg sugarcane bagasse) was achieved duringthe fermentation

[83] Bioethanol production from alkaline pretreated sugarcanebagasse using Phlebia sp MG-60

(i) MG-60 produced cellulose and xylanase rapidly duringconsolidated bioprocessing (CBP)(ii) The maximum theoretical ethanol yield of 657 (45 gL)was achieved during the fermentation

[84] Integrated fungal fermentation of sugarcane bagasse forbioethanol production by Phlebia sp MG-60

(i) 75 moisture content was suitable for subsequent ethanolproduction(ii) Some additives improved delignification in integratedfungal fermentation (IFF)(iii) Some inorganic chemicals (eg Fe2+ Mn2+ and Cu2+)increased the ethanol production

[85] Furfural and xylose production from sugarcane bagasse inethanol production

(i) The furfural yield and xylose yield were 6 and 155 gg ofsugarcane bagasse respectively(ii) Ethanol was produced from the residual solid materialsobtained from furfural and xylose at 874 and 893respectively

lignin (10ndash25) Cellulose consists of long chains of 120573-glucose monomers gathered into microfibril bundles Thehemicelluloses mostly xyloglucans or xylans are linked tothe microfibrils by hydrogen bonds Lignins are phenoliccompounds which are formed by polymerisation of threetypes of monomers (p-coumaryl coniferyl and synapyl alco-hols) Lignin adds compressive strength and stiffness to thecell wall [10] Once the lignocellulosic biomass is pretreatedand hydrolysed the released sugars can be fermented andthe downstream process is similar to that of first generationfeedstock [1] Potential lignocellulosic feedstocks and theircomposition are summarized in Table 3

High lignin andor high ash concentrations are unfa-vorable for bioethanol production Softwood especially canhence be excluded The extensive hydrogen linkages among

cellulose molecules lead to a crystalline and strong matrixstructure [11] Although starches require temperatures of only60ndash70∘C to be converted from crystalline to amorphoustexture cellulose requires 320∘C as well as high pressures (upto 25MPa) to transform the rigid crystalline structure into anamorphous structure in water [12] Cotton flax and chemicalpulps represent the purest sources of cellulose while soft andhardwoods contain less than 50 of cellulose as shown inTable 4

Hemicellulose is an amorphous structure formed ofdifferent heteropolymers including hexoses (D-glucose D-galactose and D-mannose) as well as pentose (D-xylose andL-arabinose) It may contain sugar acids (uronic acids) [13]Its backbone chain is primarily composed of xylan linkagesincluding 120572-xylose (sim90) and L-arabinose (sim10) [14] The


Table 3 Potential lignocellulosic biomass sources and compositions ( dry weight) [86 87]

Raw material Hemicelluloses Cellulose Lignin Others (ie ash)Agricultural residues 25ndash50 37ndash50 5ndash15 12ndash16Hardwood 25ndash40 45ndash47 20ndash25 080Softwood 25ndash29 40ndash45 30ndash60 050Grasses 35ndash50 25ndash40 mdasha 2ndash5Waste papers from chemical pulps 12ndash20 50ndash70 6ndash10 2Newspaper 25ndash40 40ndash55 18ndash30 5ndash8Switch grass 30ndash35 40ndash45 12 4-5aNot present or not available

Table 4 Ultimate and proximate analyses of different biomasses (wt)

Ultimate analysis (wt on dry basis) Proximate analysisSample C O H N S VM M FC A

Wood and woody biomassPine 545 387 59 05 042 461 378 129 32Eucalyptus bark 487 453 57 03 005 687 12 151 42Forest residue 527 411 54 07 010 345 568 73 14Land clearing wood 507 428 6 04 007 354 492 7 84Olive wood 49 449 54 07 003 743 66 161 3Pine chips 528 405 61 05 009 669 76 20 55Pine sawdust 51 429 6 01 001 704 153 142 01Poplar 516 417 61 06 002 797 68 115 2Mixed sawdust 498 437 6 05 002 551 349 93 07Spruce wood 523 412 61 03 010 757 67 171 05Willow 498 434 61 06 006 742 101 143 14

Herbaceous and agriculture biomassBamboo 52 425 51 04 004 71 13 152 08Miscanthus grass 492 442 6 04 015 719 114 14 27Sweet sorghum 497 437 61 04 009 718 7 168 44Switchgrass 497 434 61 07 011 708 119 128 45Corn straw 487 441 64 07 008 674 74 178 71Rice straw 501 43 57 1 016 594 76 144 186Wheat straw 494 436 61 07 017 672 101 163 64Coconut shell 511 431 56 01 01 705 44 22 31Cotton husks 504 398 84 14 001 73 69 169 32Corn stover 425 426 5 08 NA 781 106 176 37Groundnut shell 509 404 75 12 002 681 79 209 31Hazelnut shell 515 416 55 14 004 715 72 199 14Olive husks 50 421 62 16 004 737 68 174 21Rice husks 493 437 61 08 022 561 106 172 161Soya husks 454 469 67 09 008 696 63 19 51Bagasse 498 439 6 02 008 766 104 111 19Sunflower husks 504 43 55 11 01 691 91 19 28Tea wastes 486 422 54 38 703 726 1857 388

Other biomass sourcesChicken litter 605 253 68 62 12 433 93 131 343Agricultural residue 524 412 6 04 004 547 303 127 23Mixed waste paper 523 402 72 02 008 768 88 68 76Refuse-derived fuel 538 368 78 11 047 703 42 05 25Sewage sludge 509 334 73 61 233 45 64 53 433Wood yard waste 522 404 6 11 03 409 381 84 126VM volatile matter M moisture FC fixed carbon A ash


degree of branching and the xylan composition vary withthe nature and the source of raw materials To be totallyhydrolysed into free monomers hemicellulose requires awide range of enzymes in view of the diversity of its sugars

Lignin is an aromatic and rigid biopolymer covalentlybonded to hemicellulosic xylans and responsible for therigidity and high level of compactness of the plant cell wall[15] Lignin is composed of monomers of phenyl propionicalcohol that is coumaryl coniferyl and sinapyl alcoholThe lignin fraction in biomass sources varies considerablyas illustrated in Table 4 Lignin components are gainingimportance because of their dilution effect on the processesof hydrolysis and fermentation [16 17] The phenolic groupsformed from the degradation of lignin substantially deac-tivate cellulolytic enzymes and hence hamper enzymatichydrolysis Chen et al [17] however demonstrated that ligninmodification via genetic engineering could considerablyreduce lignin formation and improve ethanol yield Thiscould however be problematic as lignin components serve asthe major plant defence system to pathogens and insects andits modification could disrupt the plantsrsquo natural protection[18] Retaining the lignin could moreover benefit the energy-economy of the process since once recovered it can beapplied in a combined heat and power unit (CHP) thusbeing a potential energy self-sustaining source of the processBiomass feedstock with a high lignin content is not readilyapplicable as raw material for the bioethanol fermentationThis certainly eliminates most of the soft woods

The different composition of biomass feedstock (dry)is also reflected in its elemental composition The exactcomposition is largely dependent on the biomass sources CH and O are the key components of biomass and largelydetermine their calorific value Some typical values of theC- H- and O-content as well as other essential data aresummarized in Table 4 as a result of an extensive literaturesurvey [9 19 20] and own analyses

The ash content of biomass sources is generally lowas illustrated in Table 5 Its composition should howeverbe taken into consideration since most of the ash willconcentrate in the lignin residue thus possibly hamperingfurther energy generation by fouling or sintering Biomasscontains a significant content of K Cl and Si as well as lowerconcentrations of CaMg Al Fe andNaThe ash content andits chemical composition are a strong function of the biomassspeciesThe elements in the ash are O Ca K Si Mg Al S FeP Cl Na Mn and Ti [21] An extensive literature survey wasprovided by several authors [19 22 23] and some examplesare summarized in Table 5

2 Processing of Biomass to Ethanol

21 Generalities Once the feedstock is delivered to theethanol plant it needs to be carefully stored and conditionedto prevent early fermentation and bacterial contaminationThrough pretreatment simple sugars are made available inproportions depending on the type of biomass used and thepretreatment process The main steps are summarized inFigure 1 providing a general production flow sheet

22 Pretreatment and First Stage Hydrolysis Pretreatmentinvolves delignification of the feedstock [24] in order tomakecellulose more accessible in the hydrolysis step using phys-ical physicochemical chemical and biological treatment(Table 6) Carbonic acid and alkaline extraction have the bestperformanceHowever themost commonmethods are steamexplosion and dilute acid prehydrolysis which are followedby enzymatic hydrolysis Sulphuric acid or carbon dioxide isoften added in order to reduce the production of inhibitorsand improve the solubilisation of hemicellulose [15] Steamexplosion has a few limitations since the lignin-carbohydratematrix is not completely broken down degradation productsare generated that reduce the efficiency of the hydrolysis andfermentation steps and a portion of the xylan fraction isdestroyed

The use of dilute sulphuric acid (05ndash1 433ndash463K for10 minutes) has the preference of the US National RenewableEnergy Laboratory [25] hemicellulose is largely hydrolysedreleasing different simple sugars (xylose arabinosemannoseand galactose) but also other compounds of the cellulosicmatrix can however inhibit the enzymatic hydrolysis andfermentation Part of the acetic acid much of the sulphuricacid and other inhibitors produced during the degradationof the materials need to be removed and neutralisationis performed before fermentation Pretreatment is a costlyseparation accounting for approximately 33of the total cost[26] the economy needs to be improved and the release ofmicrobial and chemical contamination that possibly reducesthe overall yield needs further attention

23 Second Stage Hydrolysis In the second stage hydrolysisthe released cellulose of the biomass is converted into glucosewhich is again catalysed by dilute acid concentrated acid orpreferably by cellulase enzymes either produced in a separatereactor or bought externally from industrial suppliers [27ndash30]

The conversion of cellulose and hemicellulose can beexpressed by the reaction of glucan (for hexoses) and xylan(for pentose) with water

(C6H10O5)

119899+ 119899H2O 997888rarr 119899C

6H12O6 (1)

(C5H8O4)

119899+ 119899H2O 997888rarr 119899C

5H10O5 (2)

The maximum theoretical yield of hexoses and pentoses is1136 kg and 1111 kg per kg of glucan and xylan respectively

To overcome inhibition by hydrolyte components mem-brane techniques have been investigated [3 31] Chandel etal [32] investigated the strategies that have been adoptedto detoxify lignocellulosic hydrolysates and their effects onthe chemical composition of the hydrolysates to improvethe fermentability of lignocellulosics Hydrolysis of myco-LB (LB after fungal pretreatment) has been recognized asa promising approach to avoid fermentation inhibitors andimprove total sugar recovery Genetic manipulation couldmodify the metabolic routes to produce bioethanol or othervalue-added compounds in an efficient manner Furtherresearch is certainly required as described in Section 3


Table 5 Elemental ash composition of different biomass

Sample SiO2 CaO K2O P2O5 Al2O3 MgO Fe2O3 SO3 Na2O TiO2

Wood and woody biomassEucalyptus bark 1004 5774 929 235 31 1091 112 347 186 012Poplar bark 186 7731 893 248 062 236 074 074 484 012Willow 61 4609 234 1301 196 403 074 3 161 006Wood residue 5315 1166 485 137 1264 306 624 199 447 057

Herbaceous and agriculture biomassBamboo whole 992 446 5338 2033 067 657 067 368 031 001Miscanthus 5642 1077 1975 554 079 301 094 228 047 003Sorghum grass 7321 702 897 443 183 221 095 111 025 002Sweet sorghum 6685 1041 449 347 081 312 058 347 147 006Switchgrass 6625 1021 964 392 222 471 136 083 058 028Wheat straw 5035 821 2489 354 154 274 088 424 352 009Rice husks 9448 097 229 054 021 019 022 092 016 002Sugar cane bagasse 4679 491 695 387 146 456 1112 357 161 202Sunflower husks 2366 1531 2853 713 875 733 427 407 08 015

Other biomass varietiesChicken litter 577 5685 1219 154 101 411 045 359 06 003Mixed waste paper 2862 763 016 02 5353 24 082 173 054 437Refuse-derived fuel 3867 2681 023 077 1454 645 626 301 136 19Sewage sludge 3328 1304 16 1588 1291 249 157 205 225 08Wood yard waste 601 2392 298 198 308 217 198 246 101 032

Table 6 Assessment of selected pretreatment processes [15 88ndash96]

Pretreatment process Yield of fermentable sugars Wastes InvestmentPhysical or physicochemical

(i) Mechanical Low Very low Low(ii) Steam explosion High Low High(iii) Ammonia fiber explosion (AFEX) Moderate Very low High(iv) Carbonic acid Very high Very low Low

Chemical(i) Dilute acid Very high High Moderate(ii) Concentrated acid Very high High High(iii) Alkaline extraction Very high High Low(iv) Wet oxidation High Low Low(v) Organosolv Very high Low Very high

24 Fermentation Contrarily to the conversion of disaccha-rides and starch to ethanol which are mature technologiesmodern lignocellulose-to-ethanol processes are at pilot anddemonstration stage NREL (USA) [25] Iogen Corporation(Canada) [33] and ETEK (Sweden) [34] have built pilotplants capable of producing a few hundred thousand litres ofethanol per year

Fermentation is the biological process to convert thehexoses and pentoses into ethanol by a variety ofmicroorgan-isms such as bacteria yeast or fungiThe conversion reactionfor hexoses (C6) and pentoses (C5) is as follows

C6H12O6997888rarr 2C

2H5OH + 2CO

2 (3)

3C6H10O5997888rarr 5C

2H5OH + 5CO

2 (4)

The theoreticalmaximumyield of broth hexoses andpentosesis 0511 kg ethanol and 0489 kg CO

2per kg sugarThe overall

theoretical ethanol yield (at 20∘C) hence becomes 0719 and0736 liters per kg of glucan (andor other 6C structures) andxylan (andor other 5C structures) respectively

S cerevisiae the yeast commonly used for first generationethanol production cannot metabolize xylose Other yeastsand bacteria are under investigation to ferment xylose andother pentoses into ethanol

Genetically engineered fungi that produce large volumesof cellulase xylanase and hemicellulase enzymes are underinvestigation These could convert agricultural residues (egcorn stover straw and sugar cane bagasse) and energy crops(eg switchgrass) into fermentable sugars [33 35] Additionalresearch tried to find microorganisms which can effectively


Lignocellulosic

2nd

Harvesting

Pretreatment

ST

ST

ST

ST

ST

Hydrolysis1st stage

Hydrolysis2nd stage

CO2

CHP Wastes Fermentation

Waste watertreatment

Distillation

Drying toanhydrous ethanol

Figure 1 Second generation biomass-to-ethanol production (ST steam addition)

ferment both types of sugars into ethanol with Escherichiacoli Klebsiella oxytoca and Zymomonas mobilis as promisingcandidates [36 37]

When using enzymatic hydrolysis different integrationmethods of hydrolysis and fermentation steps are pro-posed In the separate hydrolysis and fermentation (SHF)the liberated cellulose is treated in a different reactor forhydrolysis and subsequent fermentation than the hydrolysedhemicellulose and lignin Although this facilitates both theoptimization of each separate reactor and the selection ofsugar-appropriate microorganisms to ferment the differ-ent sugars the higher investment costs for two separatereactors and the inhibition of the high glucose concentra-tion to fermenting organisms are major disadvantages [3839] Separate hydrolysis and cofermentation (SHCF) andsimultaneous saccharification and cofermentation (SSCF) arepossible alternatives cofermenting both C5 and C6 sugarsby a single strain of microorganisms in the same reactorsignificantly improves the process economics and enhancesthe commercial production of lignocellulosic ethanol in theshort term [39ndash41]

A novel development the consolidated bioprocessing(CBP) proceeds by producing all required enzymes and

ethanol using a single type ofmicroorganisms in a single reac-tor CBP is considered as the ultimate evolution of biomass-to-bioethanol conversion technology since it implies neithercapital nor operating costs for dedicated enzyme productiontogether with a reduced consumption of substrate for enzymeproduction Unfortunately it is predicted that it will takeseveral years of research to determine such microorganismsor compatible combinations of microorganisms [41]

With bioethanol production from lignocellulosicbiomass chemical inhibition is a more severe problemthan encountered in first generation raw materialsPretreatment and hydrolysis of lignocellulosics releasespecific inhibitors for example furans such as furfural and5-hydroxymethylfurfural (5-HMF) and phenols such as 4-hydroxybenzaldehyde (4-HB) vanillin and syringaldehydethat need to be dealt with to operate hydrolysis andfermentation under optimum conditions and maximumconversion

To increase the critical ethanol-inhibition concentrationadapted yeasts or bacteria can be used The most commonlyused yeast is Saccharomyces cerevisiae with a moderateyield of fermentation Research has been done on morepromising yeasts and bacteria Zymomonas mobilis succeeds


to survive higher ethanol concentrations in the fermenterup to 16 vol Not only this advantage but also a moderatetolerance for acids and sugars typical inhibitors present inbiomass hydrolysis makes this a very popular yeast for indus-trial application The fermentation rate is also higher withZymomonas mobilis in comparison to Saccharomyces cere-visiae [1] An interesting characteristic of Z mobilis is indeedthat its plasma membrane contains hopanoids pentacycliccompounds similar to eukaryotic sterols thus providing anextraordinary tolerance to ethanol in its environment around16wt However in spite of these attractive advantages itssubstrate range is limited to glucose fructose and sucrose Itcannot ferment C5 sugars like xylose and arabinose which areimportant components of lignocellulosic hydrolytes Unlikeyeast Z mobilis cannot tolerate toxic inhibitors present inlignocellulosic hydrolytes such as acetic acid Concentrationof acetic acid in lignocellulosic hydrolytes can be as highas 15 wt well above the tolerance threshold of Z mobilisSeveral attempts have been made to engineer Z mobilis toovercome its inherent deficiencies by metabolic engineeringmutagenesis or adaptive mutation to produce acetic acidresistant strains of Z mobilis [42 43] However when theseengineered strains metabolize mixed sugars in the presenceof inhibitors the yield and productivity are much lower thuspreventing their industrial application

To overcome inhibition by hydrolyte components mem-brane techniques have been investigated although furtherresearch is certainly required as described in Section 3

25 Purification Typical ethanol concentrations are in therange of 3ndash6 vol only very low in comparison with 12 to15 vol obtained from 1st generation feedstock [1] Due tothe higher water content of the broth additional distillationefforts are required Different process improvements includ-ing energy pinch very high gravity fermentation and hybridprocesses are described in detail by Kang et al [1]

26 Steam and Electricity Generation The bottom product ofthe first distillation column (stillage) contains mainly ligninand water next to unconverted cellulose and hemicelluloseThis insoluble fraction is dewatered by a pressure filter andsent to a fluidized bed combustor system for steam andelectricity generation This system allows the plant to beself-sufficient in energy supply reduces solid waste disposalcost and generates additional revenue through sales of excesselectricity [39 44] Burning the solid residues for steam andpower production is themost beneficial option andmeets theenergy demand of the plant

3 Current Research Priorities inBiomass to Ethanol

From the previous process assessment several bottlenecksemerge Biomass to bioethanol will only be a technical andeconomic viable alternative to first generation bioethanolif appropriate solutions are developed Current productionproblems hence determine immediate and future researchpriorities

Pretreatment as the first step accounts for about 33of the total cost [26] Better and cost-efficient pretreat-ment techniques need further investigation together withmethods to reduce or eliminate microbial and chemicalcontaminants that can reduce the yields It was alreadystated that membrane techniques could help to overcomesomeof the problemswithmicrofiltration (suspended solids)and ultrafiltration nanofiltration or reverse osmosis dealingwith dissolved contaminants The possible application ofmicrofiltration to eliminate suspended solids has recentlybeen confirmed by Kang et al [3]

In ultrafiltration (UF) solutes of high molecular weightare retained in the so-called retentate while water and lowmolecular weight solutes pass through the semipermeablemembrane in the permeate Ultrafiltration is used inindustry and research for purifying and concentratingmacromolecular (103ndash106Da) solutions especially proteinsolutions Removal of suspended solids prior to feeding themembrane is essential to prevent damage to the membraneand minimize the effects of fouling which greatly reduce theseparation efficiency

Nanofiltration and reverse osmosis are high-pressuremembrane filtration processes used most often with low totaldissolved solids water (surface water and fresh groundwater)for softening (polyvalent cation removal) and removal ofdisinfection byproduct precursors such as natural organicmatter and synthetic organic matter These membrane sep-aration technologies have been examined in different stagesof the bioethanol production

Enhancing ethanol production by pretreatment involvingfungi (eg T reesei and Basidiomycetes) with appropriatelignocellulolytic properties at low pH and high temperaturesis also a promising and added-value step in SSF ethanol bio-conversionWhile fungi act slowly potential lignocellulolyticfungi have been produced by mutagenesis gene expressionand coculturing [45] Some genera such as Candida Pichiaand Dekkera were isolated from sugarcane molasses butresulted in low ethanol concentrations and produced aceticacid an inhibitor of the fermentative yeast [46] Somenatural wild yeast species appear capable of replacing Scerevisiae in second generation bioethanol [47] but theirlow bioethanol yield and poor survival in the fermenterneed further improvement As described before some groupsof bacteria such as Zymomonas mobilis can convert sugarsinto ethanol [47] but they are more vulnerable to chemicalinhibition than S cerevisiae

The development of genetically modified fermentativeand cellulolytic microorganisms is recommended to increasethe ethanol yield and productivity under the stress conditionsof high production bioethanol processes [47] Simultane-ous saccharification and fermentation (SSF) simultaneoussaccharification and combined fermentation (SSCombF) ofthe enzymatic hydrolyzate and CBP are also consideredto be cost-effective whilst reducing end-product inhibitionGenetic engineering has succeeded in altering the conven-tional S cerevisiaersquos capacity to ferment glucose and pentosesugars simultaneously [48] Almeida et al [49] investigateda modified S cerevisiae not only capable of cofermenting


saccharides but also of generating less furfural inhibitorsAs mentioned before CBP combines hydrolysis and fermen-tation operations in a single reactor by using geneticallymodified microorganisms that produce cellulase enzyme toferment sugars in a single step This avoids the costs relatedto the purchase of cellulolytic enzymes [50] Lignin shouldbe considered as a valuable energy source used as a fuel ina CHP and being capable of supplying the power and heatrequirements of the complete conversion process

Genetic engineering as a powerful biotechnological toolis required to design new strategies for increasing theethanol fermentation performance Upregulation of stresstolerance genes by recombinant DNA technology can be auseful approach to overcome inhibitory situations [51] Geet al [52] obtained three recombinants HDY-ZMYWBG1HDY-ZMYWBG2 and HDY-ZMYWBG3 using the lithiumacetate transformationmethod into the S cerevisiae cellsTheethanol yield for HDY-ZMYWBG1 and HDY-ZMYWBG3 is0368 gg and 0365 gg respectively The resulting consor-tium was demonstrated to utilize phosphoric acid swollencellulose (PASC) for growth and ethanol production Thefinal ethanol production of 125 gL corresponded to 87 ofthe theoretical value and was 3-fold higher than a similaryeast consortium secreting only the three cellulases [53]Reconstitution of the N crassa cellodextrin transport systemin Saccharomyces cerevisiae promotes efficient growth of thisyeast on cellodextrins [54 55] The engineered yeast strainsmore rapidly convert cellulose to ethanol when comparedwith yeast lacking this systemHa et al [56] engineered yeaststo coferment mixtures of xylose and cellobiose It improvedethanol yield when compared to fermentation with eithercellobiose or xylose as sole carbon sources This is a criticalstep towards enabling economic biofuel production

Since fermentative microorganisms must be capableof surviving the high temperatures of SSFSSCombFCBPprocesses further research is required high temperatureethanol fermentation is an emerging technology providedappropriate microorganisms can be developed Such hightemperature operations do not require cooling and cellulaseaddition [57] The thermotolerant yeast K marxianus hasbeen documented as a candidate for its ability to cofermentboth hexose and pentose sugars and survive temperaturesof 42ndash45∘C [58] K marxianus was moreover geneticallymodified to exhibit T reesei and Aspergillus aculeatus cellu-lolytic activities allowing direct and continuous conversionof cellulosic 120573-glucan into ethanol at 48∘C yielding 047 ggethanol that is 922 of the theoretical yield and provingto be an ideal gene modified organism (GMO) for CBPprocesses [58]

The industrial potential for S cerevisiae fermentationhas already been proven for first generation large-scalebioethanol production Its genetic improvement is gainingincreasing research especially with respect to the CBP option[59 60] where hydrolysis and substrate fermentation arepossible in a single step

Z mobilis remains an attractive candidate due to itshigh ethanol yield and resistance to temperatures in therange of 40∘C [36] Numerous genes have been introducedand heterologous expression has been incorporated into Z

mobilis to extend its effectiveness toward other substratesnamely xylose and arabinose [97] Both the gene engineeredZmobilis and S cerevisiae have proven high ethanol yield andadaptability [98]

Further research is certainly required in optimizingbiological pretreatment involving fungi (eg T reesei andBasidiomycetes) that exhibit lignocellulolytic properties atlow pH levels and high temperature

The use of GMOs is questionable since their introductioninto large-scale fermentation operations can pose risks ofenvironmental dissemination and potential exposure risksto public health Industrial operations using antibiotics tocontrol microbial contaminants in fermenters or as strainmarkers would generate and release antibiotic resistantorganisms and offer another potential environmental andpublic health risk

Improvement in each of these individual aspects isrequired to achieve high conversion and cost-effectivebiomass-to-bioethanol operations This needs to be comple-mented by a comprehensive systems approach encompassingthe different individual steps and accounting for all inputsand outputs during the entire operation regardless of mod-ifications in any of these individual steps

4 Conclusions

The cellulosic bioethanol production process involves spe-cific processing steps especially in the pretreatment andhydrolysis Fermentation of C5 and C6 sugars needs adaptedmicroorganisms still to be further investigated

New combined processes reduce both the number ofoperation steps and the production of chemical inhibitorsRecent advances in genetically engineered S cerevisiae andZ mobilis are promising for higher alcohol tolerance andconversion efficiency Second generation bioethanol couldsurpass the traditional first generation processes providedpresent processing bottlenecks are removed and the bestcombination of advanced systems is used

Conflict of Interests

The authors declare that there is no conflict of interestsregarding the publication of this paper

Acknowledgments

This work was supported by the Chinese Science Council insupport of the delegation of the principal author to the KULeuven

References

[1] Q Kang L Appels J Baeyens R Dewil and T Tan ldquoEnergy-efficient production of cassava-based bio-ethanolrdquo Advances inBioscience and Biotechnology vol 5 no 12 pp 925ndash939 2014

[2] L Shen J Lei and Y Bi ldquoPerformance and emission character-istics of diesel engine fueledwith ethanol-diesel blends in differ-ent altitude regionsrdquo Journal of Biomedicine and Biotechnologyvol 2011 Article ID 417421 10 pages 2011


[3] Q Kang J Huybrechts B van der Bruggen J Baeyens T WTan and R Dewil ldquoHydrophilic membranes to replace molec-ular sieves in dewatering the bio-ethanolwater azeotropicmixturerdquo Separation and Purification Technology vol 136 pp144ndash149 2014

[4] G Fischer and L Schrattenholzer ldquoGlobal bioenergy potentialsthrough 2050rdquo Biomass amp Bioenergy vol 20 no 3 pp 151ndash1592001

[5] M F Demirbas M Balat and H Balat ldquoPotential contributionof biomass to the sustainable energy developmentrdquo EnergyConversion andManagement vol 50 no 7 pp 1746ndash1760 2009

[6] J Werther M Saenger E-U Hartge T Ogada and Z SiagildquoCombustion of agricultural residuesrdquo Progress in Energy andCombustion Science vol 26 no 1 pp 1ndash27 2000

[7] H Baath A Gallerspang G Hallsby et al ldquoRemote sensingfield survey and long-term forecasting an efficient combinationfor local assessments of forest fuelsrdquoBiomass and Bioenergy vol22 no 3 pp 145ndash157 2002

[8] J Malinen M Pesonen T Maatta and M Kajanus ldquoPotentialharvest for wood fuels (energywood) from logging residues andfirst thinnings in Southern FinlandrdquoBiomass andBioenergy vol20 no 3 pp 189ndash196 2001

[9] A Demirbas ldquoEnergy balance energy sources energy policyfuture developments and energy investments in Turkeyrdquo EnergyConversion and Management vol 42 no 10 pp 1239ndash12582001

[10] P H Raven R F Evert and S E Eichhorn Biology of PlantsFreeman andCompanyWorth Publishers NewYork NYUSA6th edition 1999

[11] A Ebringerova Z Hromadkova and T Heinze ldquoHemicellu-loserdquo Advances in Polymer Science vol 186 pp 1ndash67 2005

[12] S Deguchi S-A Mukai M Tsudome and K HorikoshildquoFacile generation of fullerene nanoparticles by hand-grindingrdquoAdvanced Materials vol 18 no 6 pp 729ndash732 2006

[13] B C Saha ldquoHemicellulose bioconversionrdquo Journal of IndustrialMicrobiology and Biotechnology vol 30 no 5 pp 279ndash2912003

[14] FMGirio C Fonseca F Carvalheiro L C Duarte SMarquesand R Bogel-Lukasic ldquoHemicelluloserdquo Bioresource Technologyvol 101 pp 4775ndash4800 2010

[15] A T W M Hendriks and G Zeeman ldquoPretreatments toenhance the digestibility of lignocellulosic biomassrdquoBioresourceTechnology vol 100 no 1 pp 10ndash18 2009

[16] M R Ladisch N SMosier Y Kim E Ximenes andDHogsettldquoConverting cellulose to biofuels SBE special supplement bio-fuelsrdquo Chemical Engineering Progress vol 106 no 3 pp 56ndash632010

[17] F Chen R M S Srinivasa S Temple L Jackson G Shadleand R A Dixon ldquoMulti-site genetic modulation of monolignolbiosynthesis suggests new routes for formation of syringyllignin andwallbound ferulic acid in alfalfa (Medicago sativaL)rdquoPlant Journal vol 48 no 1 pp 113ndash124 2006

[18] X Li J-K Weng and C Chapple ldquoImprovement of biomassthrough lignin modificationrdquo Plant Journal vol 54 no 4 pp569ndash581 2008

[19] A Demirbas ldquoCombustion characteristics of different biomassfuelsrdquo Progress in Energy and Combustion Science vol 30 no 2pp 219ndash230 2004

[20] S Mahmoudi J Baeyens and J P K Seville ldquoNO119909formation

and selective non-catalytic reduction (SNCR) in a fluidized bedcombustor of biomassrdquo Biomass and Bioenergy vol 34 no 9pp 1393ndash1409 2010

[21] J Shen S Zhu X Liu H Zhang and J Tan ldquoThe prediction ofelemental composition of biomass based onproximate analysisrdquoEnergy Conversion and Management vol 51 no 5 pp 983ndash9872010

[22] S van Loo and J Koppenjan The Handbook of BiomassCombustion and Cofiring Earthscan London UK 2008

[23] C Telmo J Lousada and N Moreira ldquoReview proximateanalysis backwards stepwise regression between gross calorificvalue ultimate and chemical analysis of woodrdquo BioresourceTechnology vol 101 no 11 pp 3808ndash3815 2010

[24] Y Sun and J Cheng ldquoHydrolysis of lignocellulosic materials forethanol production a reviewrdquo Bioresource Technology vol 83no 1 pp 1ndash11 2002

[25] US Department of Energy (DOE) ldquoThe DOE bioethanol pilotplantrdquo Tech Rep DOE leaflet GO-10200-1114 2000

[26] E Tomas-Pejo J M Oliva and M Ballesteros ldquoRealisticapproach for full-scale bioethanol production from lignocellu-lose a reviewrdquo Journal of Scientific and Industrial Research vol67 no 11 pp 874ndash884 2008

[27] Y J Zhang Q Li J M Su et al ldquoA green and efficienttechnology for the degradation of cellulosic materials structurechanges and enhanced enzymatic hydrolysis of natural cellulosepretreated by synergistic interaction of mechanical activationand metal saltrdquo Bioresource Technology vol 177 pp 176ndash1812015

[28] A Singh S Bajar and N R Bishnoi ldquoEnzymatic hydrolysis ofmicrowave alkali pretreated rice husk for ethanol production bySaccharomyces cerevisiae Scheffersomyces stipitis and their co-culturerdquo Fuel vol 116 pp 699ndash702 2014

[29] G P Maitan-Alfenas E M Visser and V M GuimaraesldquoEnzymatic hydrolysis of lignocellulosic biomass convertingfood waste in valuable productsrdquo Current Opinion in FoodScience vol 1 pp 44ndash49 2015

[30] X H Cui X B Zhao J Zeng S K Loh Y M Choo and DH Liu ldquoRobust enzymatic hydrolysis of formiline-pretreatedoil palm empty fruit bunches (EFB) for efficient conversion ofpolysaccharide to sugars and ethanolrdquo Bioresource Technologyvol 166 pp 584ndash591 2014

[31] J Baeyens Q Kang L Appels R Dewil Y Lv and T TanldquoChallenges and opportunities in improving the production ofbio-ethanolrdquo Progress in Energy and Combustion Science vol 47pp 60ndash88 2015

[32] A K Chandel S S da Silva and O V Singh ldquoDetoxificationof lignocellulose hydrolysates biochemical andmetabolic engi-neering toward white biotechnologyrdquo Bioenergy Research vol6 no 1 pp 388ndash401 2013

[33] D M Mousdale ldquoBiofuels biotechnology chemistry and sus-tainable developmentrdquo in The Iogen Corporation Process asa Template and Paradigm chapter 41 CRC Press Taylor ampFrancis New York NY USA 2008

[34] J Lindstedt ldquoAlcohol production from lignicellulosic feed-stockrdquo in FVS Fachtagung pp 228ndash237 2003

[35] D Deswal R Gupta P Nandal and R C Kuhad ldquoFungalpretreatment improves amenability of lignocellulosic materialfor its saccharification to sugarsrdquo Carbohydrate Polymers vol99 pp 264ndash269 2014

[36] B S Dien M A Cotta and T W Jeffries ldquoBacteria engineeredfor fuel ethanol production current statusrdquo Applied Microbiol-ogy and Biotechnology vol 63 no 3 pp 258ndash266 2003

[37] B Hahn-Hagerdal H B K Karhumaa C Fonseca I Spencer-Martins and M F Gorwa-Grauslund ldquoTowards industrial


pentose-fermenting yeast strainsrdquo Applied Microbiology andBiotechnology vol 74 no 5 pp 937ndash953 2007

[38] A Aden and T Foust ldquoTechnoeconomic analysis of the dilutesulfuric acid and enzymatic hydrolysis process for the conver-sion of corn stover to ethanolrdquo Cellulose vol 16 no 4 pp 535ndash545 2009

[39] F K Kazi J A Fortman R P Anex et al ldquoTechno-economiccomparison of process technologies for biochemical ethanolproduction from corn stoverrdquo Fuel vol 89 supplement 1 ppS20ndashS28 2010

[40] D Humbird and A Aden ldquoBiochemical production of ethanolfrom Corn Stover 2008 State of Technology modelrdquo NRELReport No TP-510-46214 National Renewable Energy Labora-tory (NREL) Golden Colo USA 2009

[41] D Klein-Marcuschamer P Oleskowicz-Popiel B A SimmonsandHWBlanch ldquoTechnoeconomic analysis of biofuels awiki-based platform for lignocellulosic biorefineriesrdquo Biomass andBioenergy vol 34 no 12 pp 1914ndash1921 2010

[42] E L Joachimsthal and P L Rogers ldquoCharacterization of ahigh-productivity recombinant strain of Zymomonas mobilisfor ethanol production from glucosexylose mixturesrdquo AppliedBiochemistry and Biotechnology vol 84ndash86 pp 343ndash356 2000

[43] R Chen Y Wang H D Shin M Agrawal and Z C MaoldquoStrains ofZymomonasmobilis for fermentation of biomassrdquo USPatent no US20090269797 A1 2009

[44] D Humbird R Davis L Tao et al ldquoProcess design and eco-nomics for biochemical conversion of lignocellulosic biomass toethanolrdquo Tech RepNRELTP-5100-47764National RenewableEnergy Laboratory Golden Colo USA 2011

[45] M Dashtban H Schraft andW Qin ldquoFungal bioconversion oflignocellulosic residues opportunities amp perspectivesrdquo Interna-tional Journal of Biological Sciences vol 5 no 6 pp 578ndash5952009

[46] A C M Basılio P R L de Araujo J O F de Morais E A daSilva Filho M A de Morais Jr and D A Simoes ldquoDetectionand identification of wild yeast contaminants of the industrialfuel ethanol fermentation processrdquo Current Microbiology vol56 no 4 pp 322ndash326 2008

[47] Y C B Chen Initial investigation of xylose fermentation forlignocellulosic bioethanol production [PhD thesis] AuburnUniversity Auburn Ala USA 2009

[48] N W Y Ho Z D Chen and A P Brainard ldquoGenetically engi-neered Saccharomyces yeast capable of effective cofermentationof glucose and xyloserdquoApplied andEnvironmentalMicrobiologyvol 64 no 5 pp 1852ndash1859 1998

[49] J R M Almeida T Modig A Roder G Liden and M-F Gorwa-Grauslund ldquoPichia stipitis xylose reductasehelps detoxifying lignocellulosic hydrolysate by reducing5-hydroxymethyl-furfural (HMF)rdquo Biotechnology for Biofuelsvol 1 article 12 2008

[50] L R LyndWH van Zyl J EMcBride andM Laser ldquoConsoli-dated bioprocessing of cellulosic biomass an updaterdquo CurrentOpinion in Biotechnology vol 16 no 5 pp 577ndash583 2005

[51] A Dogan S Demirci A O Aytekin and F Sahin ldquoImprove-ments of tolerance to stress conditions by genetic engineeringin Saccharomyces cerevisiae during ethanol productionrdquoAppliedBiochemistry and Biotechnology vol 174 no 1 pp 28ndash42 2014

[52] J P Ge L Y Zhang W X Ping M Y Zhang Y Shen and GSong ldquoGenetically engineered Saccharomyces cerevisiae strainthat can ultilize both xylose and glucose for fermentationrdquoApplied Mechanics and Materials vol 448ndash453 pp 1637ndash16432014

[53] G Goyal S-L Tsai B Madan N A DaSilva and W ChenldquoSimultaneous cell growth and ethanol production from cellu-lose by an engineered yeast consortium displaying a functionalmini-cellulosomerdquo Microbial Cell Factories vol 10 article 892011

[54] J M Galazka C Tian W T Beeson B Martinez N L Glassand J H D Cate ldquoCellodextrin transport in yeast for improvedbiofuel productionrdquo Science vol 330 no 6000 pp 84ndash86 2010

[55] S-J Ha Q Wei S R Kim J M Galazka J Cate and Y-S JinldquoCofermentation of cellobiose and galactose by an engineeredSaccharomyces cerevisiae Strainrdquo Applied and EnvironmentalMicrobiology vol 77 no 16 pp 5822ndash5825 2011

[56] S-J Ha J M Galazka S R Kim et al ldquoEngineered Saccha-romyces cerevisiae capable of simultaneous cellobiose and xylosefermentationrdquo Proceedings of the National Academy of Sciencesof the United States of America vol 108 no 2 pp 504ndash509 2011

[57] G G Fonseca E Heinzle C Wittmann and A K GombertldquoThe yeast Kluyveromyces marxianus and its biotechnologicalpotentialrdquo Applied Microbiology and Biotechnology vol 79 no3 pp 339ndash354 2008

[58] S Yanase T Hasunuma R Yamada et al ldquoDirect ethanolproduction from cellulosic materials at high temperature usingthe thermotolerant yeast Kluyveromyces marxianus displayingcellulolytic enzymesrdquo Applied Microbiology and Biotechnologyvol 88 no 1 pp 381ndash388 2010

[59] W H van Zyl L R Lynd R den Haan and J E McBrideldquoConsolidated bioprocessing for bioethanol production usingSaccharomyces cerevisiaerdquo Advances in Biochemical Engineer-ingBiotechnology vol 108 pp 205ndash235 2007

[60] M Lilly H-P Fierobe W H van Zyl and H VolschenkldquoHeterologous expression of a Clostridium minicellulosome inSaccharomyces cerevisiaerdquo FEMSYeast Research vol 9 no 8 pp1236ndash1249 2009

[61] V Gonela and J Zhang ldquoDesign of the optimal industrialsymbiosis system to improve bioethanol productionrdquo Journal ofCleaner Production vol 64 pp 513ndash534 2014

[62] R Sindhu M Kuttiraja P Binod R K Sukumaran and APandey ldquoBioethanol production from dilute acid pretreatedIndian bamboo variety (Dendrocalamus sp) by separate hydrol-ysis and fermentationrdquo Industrial Crops and Products vol 52pp 169ndash176 2014

[63] S Marx B Ndaba I Chiyanzu and C Schabort ldquoFuel ethanolproduction from sweet sorghum bagasse using microwaveirradiationrdquo Biomass and Bioenergy vol 65 pp 145ndash150 2014

[64] C Ofori-Boateng and K T Lee ldquoUltrasonic-assisted simulta-neous saccharification and fermentation of pretreated oil palmfronds for sustainable bioethanol productionrdquo Fuel vol 119 pp285ndash291 2014

[65] J H Li S Z Li B HanMH Yu GM Li andY Jiang ldquoA novelcost-effective technology to convert sucrose and homocel-luloses in sweet sorghum stalks into ethanolrdquo Biotechnology forBiofuels vol 6 no 1 article 174 2013

[66] M Shaheen M Choi W Ang et al ldquoApplication of low-intensity pulsed ultrasound to increase bio-ethanol produc-tionrdquo Renewable Energy vol 57 pp 462ndash468 2013

[67] P Manzanares M J Negro J M Oliva et al ldquoDifferent processconfigurations for bioethanol production from pretreated olivepruning biomassrdquo Journal of Chemical TechnologyampBiotechnol-ogy vol 86 no 6 pp 881ndash887 2011

[68] T Takagi M Uchida R Matsushima M Ishida and NUrano ldquoEfficient bioethanol production from water hyacinth


Eichhornia crassipes by both preparation of the saccharifiedsolution and selection of fermenting yeastsrdquo Fisheries Sciencevol 78 no 4 pp 905ndash910 2012

[69] M Lopez-Abelairas T A Lu-Chau and J M Lema ldquoEnhancedsaccharification of biologically pretreated wheat straw forethanol productionrdquo Applied Biochemistry and Biotechnologyvol 169 no 4 pp 1147ndash1159 2013

[70] M Lopez-Abelairas T A Lu-Chau and JM Lema ldquoFermenta-tion of biologically pretreated wheat straw for ethanol produc-tion comparison of fermentative microorganisms and processconfigurationsrdquo Applied Biochemistry and Biotechnology vol170 no 8 pp 1838ndash1852 2013

[71] R B Phillips H Jameel and H M Chang ldquoIntegrationof pulp and paper technology with bioethanol productionrdquoBiotechnology for Biofuels vol 6 no 1 article 13 2013

[72] M Boluda-Aguilar and A Lopez-Gomez ldquoProduction of bio-ethanol by fermentation of lemon (Citrus limon L) peel wastespretreatedwith steam explosionrdquo Industrial Crops and Productsvol 41 no 1 pp 188ndash197 2013

[73] R Velmurugan and K Muthukumar ldquoSono-assisted enzymaticsaccharification of sugarcane bagasse for bioethanol produc-tionrdquo Biochemical Engineering Journal vol 63 pp 1ndash9 2012

[74] J J Cheng andG R Timilsina ldquoStatus and barriers of advancedbiofuel technologies a reviewrdquoRenewable Energy vol 36 no 12pp 3541ndash3549 2011

[75] A C de Souza F P Carvalho C F S e Batista R F Schwanand D R Dias ldquoSugarcane bagasse hydrolysis using yeast cellu-lolytic enzymesrdquo Journal ofMicrobiology and Biotechnology vol23 no 10 pp 1403ndash1412 2013

[76] J Lu X Li R F Yang J Zhao and Y B Qu ldquoTween 40pretreatment of unwashed water-insoluble solids of reed strawand corn stover pretreated with liquid hot water to obtain highconcentrations of bioethanolrdquo Biotechnology for Biofuels vol 6no 1 article 159 2013

[77] J Prasetyo and E Y Park ldquoWaste paper sludge as a potentialbiomass for bio-ethanol productionrdquo Korean Journal of Chemi-cal Engineering vol 30 no 2 pp 253ndash261 2013

[78] C Conde-Mejıa A Jimenez-Gutierrez and M M El-HalwagildquoAssessment of combinations between pretreatment and con-version configurations for bioethanol productionrdquoACS Sustain-able Chemistry and Engineering vol 1 no 8 pp 956ndash965 2013

[79] S H Hong J T Lee S Lee et al ldquoImproved enzymatichydrolysis of wheat straw by combined use of gamma ray anddilute acid for bioethanol productionrdquo Radiation Physics andChemistry vol 94 no 1 pp 231ndash235 2014

[80] K Ojeda E Sanchez and V Kafarov ldquoSustainable ethanolproduction from lignocellulosic biomass-application of exergyanalysisrdquo Energy vol 36 no 4 pp 2119ndash2128 2011

[81] R Maryana D Marsquorifatun A I Wheni K W Satriyo and WAngga Rizal ldquoAlkaline pretreatment on sugarcane bagasse forbioethanol productionrdquo Energy Procedia vol 47 pp 250ndash2542014

[82] G Ramadoss andKMuthukumar ldquoInfluence of dual salt on thepretreatment of sugarcane bagasse with hydrogen peroxide forbioethanol productionrdquo Chemical Engineering Journal vol 260pp 178ndash187 2015

[83] L D Khuong R Kondo R de Leon T K Anh K Shimizuand I Kamei ldquoBioethanol production from alkaline-pretreatedsugarcane bagasse by consolidated bioprocessing using Phlebiasp MG-60rdquo International Biodeterioration and Biodegradationvol 88 pp 62ndash68 2014

[84] L Duy Khuong R Kondo R de Leon et al ldquoEffect of chemicalfactors on integrated fungal fermentation of sugarcane bagassefor ethanol production by a white-rot fungus Phlebia sp MG-60rdquo Bioresource Technology vol 167 pp 33ndash40 2014

[85] L Mesa M Morales E Gonzalez et al ldquoRestructuring theprocesses for furfural and xylose production from sugarcanebagasse in a biorefinery concept for ethanol productionrdquoChem-ical Engineering and Processing Process Intensification vol 85pp 196ndash202 2014

[86] J A A Swart J Jiang and P Ho ldquoRisk perceptions and GMcrops the case of Chinardquo Tailoring Biotechnologies The Social-ization of Science and Technology vol 3 no 3 pp 11ndash28 2008

[87] A Harel Noritech Seaweed Biotechnologies Ltd Algae WorldConference Rotterdam The Netherlands 2009

[88] L V AGurgelM T B Pimenta andAA S Curvelo ldquoEnhanc-ing liquid hot water (LHW) pretreatment of sugarcane bagasseby high pressure carbon dioxide (HP-CO

2)rdquo Industrial Crops

and Products vol 57 pp 141ndash149 2014[89] S Behera R Arora N Nandhagopal and S Kumar ldquoImpor-

tance of chemical pretreatment for bioconversion of lignocel-lulosic biomassrdquo Renewable amp Sustainable Energy Reviews vol36 pp 91ndash106 2014

[90] J J Zeng Z H Tong L T Wang J Y Zhu and L IngramldquoIsolation and structural characterization of sugarcane bagasselignin after dilute phosphoric acid plus steam explosion pre-treatment and its effect on cellulose hydrolysisrdquo BioresourceTechnology vol 154 pp 274ndash281 2014

[91] D T Phan and C S Tan ldquoInnovative pretreatment of sugarcanebagasse using supercritical CO

2followed by alkaline hydrogen

peroxiderdquo Bioresource Technology vol 167 pp 192ndash197 2014[92] J M Prado L A Follegatti-Romero T Forster-Carneiro

M A Rostagno F Maugeri Filho and M A A MeirelesldquoHydrolysis of sugarcane bagasse in subcritical waterrdquo Journalof Supercritical Fluids vol 86 pp 15ndash22 2014

[93] J Singh M Suhag and A Dhaka ldquoAugmented digestion of lig-nocellulose by steam explosion acid and alkaline pretreatmentmethods a reviewrdquoCarbohydrate Polymers vol 117 pp 624ndash6312015

[94] R Singh A Shukla S Tiwari and M Srivastava ldquoA reviewon delignification of lignocellulosic biomass for enhancementof ethanol production potentialrdquo Renewable and SustainableEnergy Reviews vol 32 pp 713ndash728 2014

[95] R Biswas H Uellendahl and B K Ahring ldquoWet explosionpretreatment of sugarcane bagasse for enhanced enzymatichydrolysisrdquo Biomass amp Bioenergy vol 61 pp 104ndash113 2014

[96] S Sharma R Kumar R Gaur et al ldquoPilot scale study on steamexplosion andmass balance for higher sugar recovery from ricestrawrdquo Bioresource Technology vol 175 pp 350ndash357 2014

[97] K Deanda M Zhang C Eddy and S Picataggio ldquoDevelop-ment of an arabinose-fermenting Zymomonas mobilis strainby metabolic pathway engineeringrdquoApplied and EnvironmentalMicrobiology vol 62 no 12 pp 4465ndash4470 1996

[98] C Weber A Farwick F Benisch et al ldquoTrends and challengesin the microbial production of lignocellulosic bioalcohol fuelsrdquoApplied Microbiology and Biotechnology vol 87 no 4 pp 1303ndash1315 2010

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Table 1 Common ethanol-petrol mixtures [1 31]

Code Composition Countries CommentsE5 Max 5 anhydrous ethanol min 95 petrol Western Europe

Blends for regular carsE10 Max 10 anhydrous ethanol min 90 petrol USA EuropeE15 Max 15 anhydrous ethanol min 85 petrol USA cars gt2000E25 Max 25 anhydrous ethanol min 75 petrol BrazilE85 Max 85 anhydrous ethanol min 15 petrol USA Europe Flex-fuel vehiclesE100 Hydrous ethanol (sim53 wt water) Brazil

or diterbutyl peroxide to enhance the cetane number theuse of a dual fuel operation in which ethanol and diesel areintroduced separately into the cylinder or the modificationof diesel engines in order to adapt their characteristics ofautoignition [2]

12 Bioethanol from Different Feedstocks Fermentation ofsugar-based raw materials is referred to as ldquofirst generationrdquobioethanol whereas the use of lignocellulose raw materials iscommonly called ldquosecond generationrdquo bioethanol The ldquothirdgenerationrdquo of algal bioethanol is at an early stage of investi-gationThe first generation processes were discussed in detailby Kang et al [1] Since the present paper deals with secondgeneration bioethanol relevant and recent (gt2010) literatureis summarized in Table 2 The 2nd generation bioethanolprocesses will use cellulose-released sugars despite the costof the required enzymes to hydrolyse cellulose Developmentof this technology could deal with a number of cellulose-containing agricultural byproducts such as straw woodtrimmings sawdust bamboo and others

The specificities of using lignocellulosic raw materialswill be dealt with in Section 13 Whether first second orthird generation feedstock is used fermentation producesan alcohol-lean broth only as such unusable in industrialand fuel applications The ethanol must hence be purifiedFractional distillation can concentrate ethanol to 956 vol(895mol) corresponding to the azeotropic compositionwith a boiling point of 782∘C Further ethanol enrichment bycommon distillation is impossible but different alternativeshave been investigated as reported by Kang et al [3]

13 Lignocellulosic Biomass

131 Sources Available biomass can be categorized intoprimary sources produced as either crop or key product forexample sugar cane short rotation energy plantations sec-ondary sources as residues from the production processesfor example bagasse rice husks and straw and tertiarysources as residues produced during and after applicationend for example the organic fraction of municipal solidwaste (MSW) sewage treatment sludge wood trimmingsand so forth [4] In general the final availability of organicwastes and residues may fluctuate and is affected by marketgrowth although climate and other factors have influencesespecially when considering the primary sourcesThe energypotential of biomass residues and organic wastes depends

on the yield the total land area available and the type ofproduction

The organic fraction of municipal solid waste (MSW)is an inexpensive source of biomass and covers domesticand industrial waste collected in a specific area The overallpotential of the organic fraction of municipal waste and itswaste wood fraction is strongly reliant on economic growthconsumption and the use of biomaterials It is estimated atbetween 5 and 50 EJyear [4] The unit EJ (eta-joule) is equalto 1018 J

Agroindustrial biomass residues are byproducts of agri-culture or its related industry including cotton stalks wheatand rice straw coconut shells maize cobs jute sticks andrice husks [5] The agricultural residues are produced decen-tralised andhave a lowdensityDue to the high transportationcost it is expensive to apply agricultural residues as the mainfuel in power stations The potential of agriculture residuesvaries from 15 up to 70 EJyear [4] as a function of regionalproduction harvesting processing factors and recoverabilityfactors Within the agroindustrial residues dried manure isconsidered as a tertiary source A total worldwide estimate isdifficult to make and given as 5 to 55 EJyear with the lowerestimate due to the current use as fertiliser while the higherestimate considers the total technical potential [5]

Forestry residues include biomass not harvested orremoved from sorting regions in commercial hardwood andsoftwood production through forestmanagement operationssuch as precommercial thinning and removal of dead anddying trees Forestry waste includes wood chips sawdustand bark It can provide 65 of the biomass energy potential[6 7] The extraction costs and the required transportationto centralized processing plants make forest fuels expen-sive Several studies have focused on their use for energyproduction at a district level applying appropriate designsof decentralized smaller plants for example Malinen et al[8] and Demirbas [9] The energy potential of the worldrsquosforests is again difficult to estimateThe possible contributionby 2050 is estimated at 98 EJyear of excess natural forestgrowth and at 32ndash52 EJyear of processing residues Althoughthese lignocellulosic biomass resources represent a signifi-cant energy value sim150 EJyear only part of this resourcecan be used as feedstock for bioethanol for reasons givenbelow

132 Composition Lignocellulose the principal componentof the plant cell walls is mainly composed of cellulose (40ndash60 of the total dry weight) hemicellulose (20ndash40) and




































Table 2 Continued











































(C6H10O5)

119899+ 119899H2O 997888rarr 119899C

6H12O6 (1)

(C5H8O4)

119899+ 119899H2O 997888rarr 119899C

5H10O5 (2)
















2H5OH + 2CO

2 (3)

3C6H10O5997888rarr 5C

2H5OH + 5CO

2 (4)







Lignocellulosic

2nd

Harvesting

Pretreatment

ST

ST

ST

ST

ST

Hydrolysis1st stage

Hydrolysis2nd stage

CO2



Distillation































4 Conclusions





Acknowledgments


References

















































































































FuelsJournal of







Advances in



Journal of


Renewable Energy





RotatingMachinery


EnergyJournal of




















































Table 2 Continued











































(C6H10O5)

119899+ 119899H2O 997888rarr 119899C

6H12O6 (1)

(C5H8O4)

119899+ 119899H2O 997888rarr 119899C

5H10O5 (2)
















2H5OH + 2CO

2 (3)

3C6H10O5997888rarr 5C

2H5OH + 5CO

2 (4)







Lignocellulosic

2nd

Harvesting

Pretreatment

ST

ST

ST

ST

ST

Hydrolysis1st stage

Hydrolysis2nd stage

CO2



Distillation































4 Conclusions





Acknowledgments


References

















































































































FuelsJournal of







Advances in



Journal of


Renewable Energy





RotatingMachinery


EnergyJournal of




































(C6H10O5)

119899+ 119899H2O 997888rarr 119899C

6H12O6 (1)

(C5H8O4)

119899+ 119899H2O 997888rarr 119899C

5H10O5 (2)
















2H5OH + 2CO

2 (3)

3C6H10O5997888rarr 5C

2H5OH + 5CO

2 (4)







Lignocellulosic

2nd

Harvesting

Pretreatment

ST

ST

ST

ST

ST

Hydrolysis1st stage

Hydrolysis2nd stage

CO2



Distillation































4 Conclusions





Acknowledgments


References

















































































































FuelsJournal of







Advances in



Journal of


Renewable Energy





RotatingMachinery


EnergyJournal of






























2H5OH + 2CO

2 (3)

3C6H10O5997888rarr 5C

2H5OH + 5CO

2 (4)







Lignocellulosic

2nd

Harvesting

Pretreatment

ST

ST

ST

ST

ST

Hydrolysis1st stage

Hydrolysis2nd stage

CO2



Distillation































4 Conclusions





Acknowledgments


References

















































































































FuelsJournal of







Advances in



Journal of


Renewable Energy





RotatingMachinery


EnergyJournal of


















Lignocellulosic

2nd

Harvesting

Pretreatment

ST

ST

ST

ST

ST

Hydrolysis1st stage

Hydrolysis2nd stage

CO2



Distillation































4 Conclusions





Acknowledgments


References

















































































































FuelsJournal of







Advances in



Journal of


Renewable Energy





RotatingMachinery


EnergyJournal of







































4 Conclusions





Acknowledgments


References

















































































































FuelsJournal of







Advances in



Journal of


Renewable Energy





RotatingMachinery


EnergyJournal of































































































































FuelsJournal of







Advances in



Journal of


Renewable Energy





RotatingMachinery


EnergyJournal of

















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