Review Article

Biofuels: Engineering and Biological ChallengesPURNENDU GHOSH*Birla Institute of Scientific Research, Statue Circle, Jaipur 302 001, India

(Received on 30 March 2014; Accepted on 02 August 2015)

The second generation biofuel technologies are evolving rapidly to provide solutions for the partial replacement of fossil

fuels. Both bioethanol and biodiesel have great potential in India. Both the technologies, however, have to overcome various

bottlenecks before they become commercial technologies. In this regard, several critical questions, besides science and

technology, need to be resolved. This will require new ways of thinking about agriculture, energy infrastructure and rural

economic development.

Keywords: Biofuels Technology; Bioethanol; Biomass; Algal Biofuel; Bioenergy

*Author for Correspondence: E-mail: ghoshiitbisr@gmail.com; Phone: 0141-2385283

Proc Indian Natn Sci Acad 81 No. 4 September 2015 pp. 765-773 Printed in India. DOI: 10.16943/ptinsa/2015/v81i4/48295

Introduction

In recent times, a great concern about fossil fuelssupplies, their non-renewable nature andenvironmental consequences of their use has driveninterest in biofuel programmes all over the world.There is no doubt that the “best substitute forpetroleum is petroleum” and, as one analyst puts it,replacement of fossil fuel by biofuel is not possible,butaugmentation of fuel supply probably is. As Churchand Regis (2012) write in their book Regenesis,“We’re now in a transitional period, caught betweenthe age of fossil fuels and the age of biofuels.”

It is believed that a partial transition from oil tobiofuels can stabilize the energy market significantly.To be a viable alternative, a biofuel should provide anet energy gain, have environmental benefits, beeconomically competitive and be producible in largequantities without affecting food security of thecountry.

The National Policy on Biofuels of India (2009)proposes a target of 20% blending of bioethanol by2017. A target of 10% petrol blending seem morerealistic for 2017. Even this seems a difficult

proposition keeping in view the present supply anddemand situation. The intermediate target of 5% and10% blending by 2007-2008 has not been achieved.The government is unable to implement compulsoryblending of 5% ethanol in petrol.

First and Second Generation Biofuels

The basic routes for converting biomass to biofuelare biochemical and thermochemical. The two classicthermochemical options, namely, gasification andpyrolysis produce different intermediates. Gasificationinvolves rapid heating and partial oxidation to producesyngas, which is largely carbon monoxide andhydrogen. The high oxygen content of biomass resultsin the production of significant quantities of carbondioxide, which reduces carbon efficiency. Also thesulphur, nitrogen, phosphorous, potassium, and mineralcontent of biomass complicates matters further. Inpyrolysis, lower temperatures are used to break downbiomass into smaller molecules such as oxygenatedaromatics, ketones, organic acids, and otheroxygenates, as well as light hydrocarbon gases. Inaddition to the lower energy input to achieve biomassdeconstruction, pyrolysis has a high theoretical yieldfor liquid products.

Published Online on 13 October 2015

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In biochemical processing, biomass is typicallyprocessed to yield monosacchrides, which are thenconverted by microbes to produce fuels. Thoughbioethanol is an established biofuel, there is an alternateview that it would be a good idea to look at theconversion of lignocellulose into organic acids ratherthan sugars.

Another biochemical route uses anaerobicdigestion to produce biogas, a mix of methane andcarbon dioxide. Here, natural consortia of bacteriadecompose organic matter into methane in theabsence of oxygen. Although much of the biomassresource might be dedicated to biofuel production (thusdiminishing its role in electricity generation), biogastechnologies could provide a small but nontrivial partof a renewable electricity portfolio, particularly giventheir flexibility and potential for distributed generation.

The feedstock for first generation biofuelsproduced through biochemical routes are primarily foodcrops, such as sugar cane, grain (corn), oil seeds andvegetable oils. Their limited contribution to meet theenergy demands of the future has raised questionsabout their role in the transport fuel mix of the future.This makes the need for second generation biofueltechnologies inevitable and desirable.

The feedstock for second generation biofuels isnon-food biomass, such as lignocellulosic materials(bagasse, cereal straw, forest residues, and short-rotation energy crops). The second-generation biofuelproduction has the potential to provide benefits suchas consuming waste residues and making use ofabandoned land. Job creation and regional growthare probably the most important drivers for theimplementation of second-generation biofuel projectsin major economies and developing countries.

According to the estimates of InternationalEnergy Agency (2010) biofuels are expected toprovide 9% (11.7 EJ) of the total transport fuel demand(126 EJ) in 2030, 26% (29 EJ) of total transportationfuel (112 EJ) in 2050, with second-generation biofuelsaccounting for roughly 90% of all biofuels .

Biofuels derived from lignocellulosic biomass andalgae are promising additional sources to meet energy

demands of the country. Both can play a significantpart to solve energy supply picture in the futureprovided key obstacles are overcome. Both, however,are future technologies as there are no commercialplants, but a considerable number of pilot anddemonstration plants have been planned or set up inrecent years, mainly in North America, Europe, Brazil,China, India and Thailand.

In India, the commercial viability of both theoptions is highly dependent on the future price of oiland the government policy. There is thus a promiseas well as an uncertainty. The promise is tosignificantly reduce our dependence on imported oil,create new jobs, improve rural economies, reducegreenhouse gas emissions, and enhance national fuelsecurity. The major uncertainties are feedstockavailability and cost, conversion technologies and cost,and the impact of technologies on the environment.The milestones (USDOE, 2006) that are suggestedfor the development of biofuels are provided in Fig.1.

Fig. 1: Biofuels development milestones (USNAS, 2012)

A brief overview on the future of cellulose andalgae-based biofuels is given here.

Cellulose-Based Biofuel

Production of ethanol from biomass follows variousconversion routes (Fig. 2). Ethanol is produced in India

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from cane molasses. Efforts to produce ethanol fromother sugar-based feedstock such as sweet sorghum,sugar beets, and sweet potatoes are at present in theexperimental stage. Lower molasses availability andconsequent higher prices impact ethanol’s cost ofproduction, thereby causing a disruption in the supplyof ethanol at pre-negotiated fixed ethanol prices.

All the countries in the world are looking forsolutions for their growing energy needs usingsustainable and renewable resources. The first-generation technologies for bioethanol productionbased on sugars and starches cannot provide long-term solution. They compete for land with food crops,resulting in misleading cost-benefit analysis. What weneed is a cheap, abundant and renewable raw materialthat does not interfere with food production.Lignocellulosic biomass (LCB) is such a feedstockfor the production of second-generation bioethanol.

Supply of Biomass

The global supply of cellulosic biomass is estimatedto contain energy that is equivalent to much morethan the world’s current annual consumption oftransportation fuel. The sources of cellulosic biomassinclude crop wastes, forest residues, and dedicatedenergy crops. Lignocellulosic biomass (LCB) is lessexpensive than sugar or starch-based feedstock, butits conversion to ethanol at present is more costly.The commercialization of this technology thus has toovercome various bottlenecks. These includefeedstock availability, scale of operation, cheaper andeffective pretreatment technologies, efficienthydrolytic agents, availability of recombinantorganisms capable of co-fermenting the whole rangeof sugars at a temperature compatible to optimum

hydrolysis, and better co-product value (Ghosh andGhose, 2003).

Supply of biomass is one of the most criticalfactors for the development of a viable bioethanoltechnology. Three distinct goals need to be met forthe development of biomass-based biofuels, namely,maximizing the total amount of biomass produced perhectare per year, maintaining sustainability whileminimizing inputs, and maximizing the amount of fuelthat can be produced per unit of biomass. Exact valuesfor each of these parameters would vary, dependingupon the type of energy crop and the growing zone.Logistics of raw material supply (availability,collection, storage and handling) to meet largedemands of biofuels is a major issue of concern. Inaddition, the availability of the feedstock on asustainable basis would need either large storagefacilities or availability of plants to operate on multiplefeedstock for their continued operation throughout theyear.

Ideal Pretreatment Technology

Pretreatment of LCB continues to be a major barrierfor the development of a viable technology. In theLCB-based bioethanol technology, cellulose andhemicellulose present in the lignocellulose arehydrolysed to sugars (hexoses and pentoses) usingacids or enzymes. Lignin is the major interference inthe hydrolysis of native lignocellulose. In the enzymaticprocess, the LCB is pretreated in order to increasethe accessibility of cellulolytic enzymes (cellulases)to the substrate. Typically, hydrolysis yields in theabsence of pretreatment are less than 20% oftheoretical yields, whereas yields after pretreatmentoften exceed 90%. The rationale for pretreatmenthas thus been to separate individual components ofLCB with minimum component losses, concomitantwith an increase in surface area and a decrease incrystallinity.

An ideal technology is expected to produce areactive fibre that will require little or no size reduction,and can be operated at a high solid/liquid ratio. Oneneeds to ascertain what is more important forenzymatic hydrolysis – the extent of delignificationthat requires harsher conditions for complete lignin

Fig. 2: Biomass-based bioethanol conversion routes

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separation or loosening of cellulose – hemicellulose-lignin bonds under milder conditions. The benefits oflignin solubilization need to be weighed against thepotential for fermentation inhibition by soluble ligninderivatives.

Various mechanical, physical, chemical, andbiological approaches, either singly or in combinationhave been attempted to meet these objectives, butnone has shown the promise expected from an idealpretreatment technology. Development of LCB-basedenergy plants with traits such as increased celluloseand hemicellulose and less lignin not only has thepotential to improve ethanol yields, but also thepossibility of application of much simpler pretreatmenttechnologies. Metabolic engineering of the ligninbiosynthetic pathway has been suggested as a methodfor modifying lignin content in the feedstock.

Enzyme and Enzymatic Hydrolysis

The important parameters of enzymatic hydrolysis aresugar yield, duration of hydrolysis, enzyme loading,characteristics of substrate cellulose, and enzymecellulases.

The most desirable attributes of the enzymecellulases include the ability to produce a completecellulase system with high catalytic activity againstcrystalline cellulose, thermal stability, decreasedsusceptibility to enzyme inhibition by the products ofhydrolysis (glucose, and cellobiose), selectiveadsorption of the enzyme on cellulose, and the abilityto withstand shear forces. Strategies to improvecellulases include discovering new enzymes throughbioprospecting, creating new/better mixtures ofenzymes, and developing improved expression systemsthrough protein engineering. De-novo and in-silicodesigning of improved cellulases are also beingattempted. Creating a more effective cellulose bindingdomain in the enzyme molecule is another approachto increase enzyme efficiency.

A critical element for the success of bioethanoltechnology is the availability of cheap cellulases.Industrial enzyme producers are trying to achievereduction in enzyme cost in order to support aneconomical and robust cellulose biorefinery. Cellulase

enzymes are too expensive for bioethanol. Forexample, costs of amylase enzymes for convertinggrain starch to ethanol are about ten times cheaperthan the most optimistic cost estimates for cellulasepreparations. There is, however, a good possibility ofproducing effective cellulases at a much reduced cost.For the hydrolysis of pretreated biomass, extremelycomplex cellulases may not be required; simplercellulase systems may serve the purpose. The majormarket for cellulase enzymes is the textile industry,and the enzymes produced are tailored to meet therequirement of this industry. It is important to recognizethat biofuels application needs are significantlydifferent from textile applications.

An Ideal Ethanol Producing Organism

The bioethanol process needs an efficient organismwith capability to convert sugars (both hexoses andpentoses) to ethanol. An ideal ethanol producingorganism should have characteristics such as highethanol tolerance, capacity to withstand high osmoticpressure, high temperature, and low pH, high cellviability, appropriate flocculation and sedimentationcharacteristics, capability to ferment broad range ofsugars mainly to ethanol and possibly negligible levelsof by-products (such as acids and glycerol), andresistance to inhibitory compounds present in thepretreatment/hydrolysis stream. A strategy forincreasing ethanol tolerance or other traits usesevolutionary engineering concepts and methods. Thisstrategy allows the microbial process to evolve underproper selective pressure (in this case, higher ethanolconcentrations) to increasingly higher ethanoltolerances.

Conversion of cellulose and hemicellulose toethanol comprises hydrolysis followed by fermentationof hexoses and pentoses by ethanol producingorganisms. Simultaneous saccharification andfermentation (SSF) integrates the processes ofhydrolysis with fermentation. The development ofthermophilic ethanol-producing organisms for use inSSF could allow the consolidated process to run athigher temperatures, thus realizing significant savingsby reducing cellulase enzyme requirements.Combining cellulase production, cellulose hydrolysis,

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and co-fermentation of hexose and pentose sugars ina single step, called “consolidated bioprocessing”, isconsidered the ultimate low-cost configuration forcellulose hydrolysis and fermentation.

Companies are engaged in creating syntheticmicrobes to accelerate the conversion of agriculturalwaste to ethanol. Diverse microbial strains collectedfrom the seawater is being used to create newsynthetic microbes. The hunt is on for a better microbethat will cheaply and efficiently break down celluloseto sugars and then ferment those sugars into ethanol.Such designer organisms are yet not available.

Commercialization of Technology

The projected cost of ethanol from LCB has declinedsignificantly in the last ten years. Further cost reductionis needed. This is possible by employing a celluloseand hemicellulose rich, but lignin lean feedstock.

Relatively large investments are required toinstall LCB-based bioethanol plants. In India ethanolplants are comparatively small in capacity. This bringsto the fore another related issue, namely, scale ofoperation vis-à-vis feedstock availability. Keeping inview the logistics of feedstock procurement, it isneeded to decide if it is advisable to build very largeplants as increased feedstock cost (due to collectionand transport of large amounts of feedstock) mayoffset savings due to economies of scale.

Commercialization of ethanol needs the attentionof researchers, entrepreneurs, and more importantly,the policy makers. India has the capacity to produce4000 million litres of ethanol from molasses, but itproduces around 2800 million litres. A sizable utilizablecapacity needs to be utilized. It will depend upon howpricing issues are addressed.

A Global Market Survey (Ethanol 2020), reportsthat it is possible to replace 20% of gasolineconsumption in the US, China, and India by 2020, ifthe promises of competitive, large-scale cellulosicethanol production are realized, and if national import/export policies for biofuels are further liberalized.

These are big ‘ifs’. There are many questionsthat need to be resolved, both at the researchers and

planners stage. Large-scale lignocellulose-basedbioethanol technology will require major changes insupply chain infrastructure. It will require new waysof thinking about agriculture, energy infrastructure,and rural economic development.

Algal Biofuel

The National Bio-diesel Mission (NBM) has identifiedJatropha curcas as the most suitable tree-borneoilseed for bio-diesel production on wastelands.Biodiesel production in India is very small due toinadequate supplies of Jatropha. NBM had set anambitious target of covering 11.2 to 13.4 millionhectares of land under Jatropha cultivation by the endof 2011-12.

The Government of India’s ambitious plan ofproducing sufficient bio-diesel to meet the mandateof 20% blending with diesel by 2011-12 has proceededslowly. According to trade and industry estimates,Jatropha has been planted across 500,000 hectaresof wasteland, of which 65-70% is estimated to benew plantation, and would take three to four years tomature. As a result, there are insufficient Jatrophaseeds available for biodiesel production.

Lack of high-yielding, drought-tolerant Jatrophaseeds, smaller land holdings, ownership issues withgovernment or community owned wastelands, littleprogress made by state governments to meet largescale Jatropha plantations, and negligible commercialproduction of biodiesel have hindered the efforts andinvestments made by both private and public sectorcompanies in this sector.

According to the report on Biofuels forTransportation Programme for 12th Five Year Planprepared by the sub-group constituted by the Ministryof New and Renewable Energy, the absence ofguaranteed national market due to the absence ofminimum support price is bound to deter theinvestment especially in long duration crops with littlehistory of cultivation such as Jatropha curcas orPongamia pinnata.

There are about 20 large capacity biodiesel plants(10,000 to 100,000 tons per year) in India that producebiodiesel from edible oil waste (unusable oil fractions),

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animal fat and non-edible oils. Commercial productionof biodiesel from Jatropha and non-edible oilseeds issmall, with estimates varying from 140 to 300 millionlitres per year. The biodiesel produced is sold to theunorganized sector (irrigation pumps, agriculturalusage, diesel generators, etc.), and to experimentalprojects carried out by automobile and transportcompanies. Biodiesel production cost is higher thanthe government notified purchase price. This is mainlydue to the lack of supply of Jatropha seeds.

An Ideal Algae

Algae are an attractive way to harvest solar energy,and turn carbon dioxide into biofuel. Much money isthus being poured into the idea of turning algae intomini oil wells.

The algae-derived biofuel is projected to reducefossil fuel consumption equivalent to 6% of roadtransport diesel by 2030. An efficient algae-basedbiofuel process promises around 40000 litres of oilper hectare of land. The worldwide microalgalmanufacturing infrastructure is devoted to extractionof high value products such as carotenoids and omega3 fatty acids used for food and feed ingredients.Although microalgae are not yet produced at largescale for bulk applications, recent advances,particularly in the methods of systems biology, geneticengineering, and biorefining present opportunities todevelop this process in a sustainable and economicalway, within the next twenty years. But the challengesare many.

Algae use carbon dioxide to produce oilmolecules via photosynthesis. In non-stressed growingalgae, lipids are mostly present in the form ofphospholipids in the cell membranes. Some microalgae,when exposed to stress conditions (e.g., nutrientdeprivation or high light intensities), accumulate lipidsin the form of triacylglycerols in so-called oil bodies.This accumulation occurs at the expense of energyused for growth, leading to a decrease in growth rateand a consequent decrease in productivity.The carbondioxide discharged from power plants and oil refineriescan be captured by algae and used to produce biofuel,and thereby reducing carbon dioxide build up in theatmosphere. A schematic of algae-based biodiesel

processing is provided in Fig. 3.

An ideal algae that can produce biofuel shouldhave high yield on high light intensity, large cells with

Fig. 3: Algae-based biodiesel processing

thin membranes, stable and resistant to infections, andinsensitive to high oxygen concentration. Algae shouldbe able to grow and produce lipids at the same time,should form flocs, and preferably should excrete oilsoutside the cells. Such a magic bug has not beendiscovered yet. Efforts, nevertheless, are being madeto design such magic bugs.

Knowledge of the biosynthesis mechanism oftriacylglycerols and their accumulation in oil bodies islimited and often based on analogies with higher plants.If the mechanisms are known, it could open thepossibility of inducing lipid accumulation in oil bodieswithout having to apply a stress factor. A detailedinsight into metabolic pathways may lead to strategiesto induce lipid accumulation based on processconditions, defined nutrient regimens, and/or the useof metabolic engineering techniques.

Mass Cultivation of Algae

Both open and closed mass cultivation systems canbe used for growing algae. Table 1 summarizes thepros and cons of open and closed system. The obviousproblems with open systems are low biomass growth

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and biomass loss due to lesser control on growthparameters, including disrupted availability of sunlight.Prevention of algae from the predators andcontamination by natural strains are other obviousdifficulties (USNAS, 2012).

Closed and controlled photobioreactors are moreefficient. The design and fabrication of efficientphotobioreactors equipped with optimal lightingtechniques and configurations with emphasis on lightefficiency, less shear damage and low cell adherenceat the surface of the bioreactor is quiteexpensive.Advantages and disadvantages of typicalclosed bioreactors for algal biofuel production arementioned in Table 2 (Kunjapur and Eldridge, 2010).

In recent years, much effort is put into increasingphotosynthetic efficiency of microalgae underoversaturating light (the normal condition on a sunnyday). Certain strains of microalgae can harness 3%of the incoming sunlight to make plant matter, asopposed to roughly 1% for corn or sugar.

The photosynthetic efficiency can be increasedby developing new microalgae strains with smallerantenna sizes, and by decreasing the light path ofphotobioreactors, while increasing mixing in high celldensity cultures. Researchers obtained highphotosynthetic efficiencies under bright sunlight in

systems with lower energy requirements by reducingthe light intensity at the reactor surface. To reducethe cost of manufacturing these systems, verticalpanels made from thin plastic films such aspolyethylene have been be used.

Arranging for carbon dioxide that could beutilized for commercial algae production is challenging;a total of 1.8 kg of carbon dioxide is needed to produce1 kg of algal biomass. While carbon dioxide could besourced from power plants for sequestration,arranging large quantities of fresh or saline water isextremely difficult. In addition, algae like any plantwould require nutrients such as NPK and othermicronutrients for optimal growth.

For sustainable production of biofuel frommicroalgae, it will be important to make use of residualnutrient sources, and to recycle nutrients as much aspossible. Utilization of wastewater will also achievetwin objectives of algal biomass production andwastewater treatment.Waste-water may offer auseful point source, which can be either municipal,organic industrial (e.g., food processing), organicagricultural (e.g., confined animal facilities), oreutrophic waters with low organic content but highnutrient content (e.g., agricultural drainage, lakes andrivers). Microalgae can also grow in seawater. Evendeserts would be suitable if there is access to saltaquifers.

Table 1: Algal Biofuel. Open and closed systems (USDOE,2006)

Parameter Open system Closed system

Cost Lower Higher

Pumping energy Lower Higher

Ease and scale up Greater Lower

Evaporative water loss Higher Negligible

Land area required Higher Lower

Contamination risks Higher Lower

Productivity Lower Higher

Productivity stability More variable Less variable

Sparged CO2 loss Higher Lower

Table 2: Closed Bioreactors: Advantages and disadvantages(Kunjapur and Eldridge, 2010)

Reactor typeAdvantages Disadvantages

Flat plate Shortest oxygen path Low photosyntheticLow power consumptionefficiency

Tubular High volumetric biomassOxygen accumulationdensity Photoinhibition

Most land use

Vertical Greatest gas exchange Support costsBest exposer to light/ Scalabilitydark cyclesLeast land useHigh photosyntheticefficiency

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Algae Harvesting and Processing

After the growth, the algal biomass needs to beharvested, the lipids extracted, and the remaining cellcomponents recovered. Harvesting of microalgae isexpensive because of the high energy requirementsand capital costs involved.

Since most microalgae are small individual cells,centrifugation is often used as a preferred harvestingmethod. However, as the biomass concentration isgenerally low (<3 g/L), centrifugation of dilutedstreams requires a large capacity of the centrifuge,which makes the process energy-demanding andexpensive. Flocculation, followed by sedimentation andflotation, before centrifugation or filtration willsubstantially reduce harvesting costs and energyrequirements. Ideally, algae should flocculatespontaneously at a certain stage of the process.

The process involves extraction of stored oil fromthe algae (by breaking oil-rich algae). This adds todownstream processing cost. The algal oil is extractedfrom the algal cells and then converted into biodieselby transesterification with short-chain alcohols or byhydrogenation of fatty acids into linear hydrocarbons.

After harvesting, the cells are disrupted and theoil extracted with solvents. Most microalgae strainsare, in general, relatively small and have a thick cellwall. For this reason, very harsh conditions (e.g.,mechanical, chemical, and physical stress) are neededto break the cells for extraction of the products.Excretion of the oils, in a manner similar to whatnaturally occurs in the microalgae Botryococcusbraunii, will lead to a simplified biorefinery andimprove downstream economics. However, it will notprovide a complete solution because the remainingcell components still need to be recovered from thecells.

Thin cell membranes, such as those present inDunaliella, strong enough to prevent shear damageduring production, would facilitate cell disruption.Research is needed to explore mild cell disruption,extraction, and separation technologies that retain thefunctionality of the different cell components.

A genetically engineered bacteria (E. coli) hasshown promise to convert sunlight, carbon dioxide andwater into different hydrocarbons, including biodiesel.The bacterium grows happily (three times faster thanthe yeast) at tropical temperature. The designedorganism secretes oil, instead of storing it inside theorganism, so as to reduce downstream costs.

Concluding Remarks

It is now a well-established fact that fossil fuels arein short supply and have limited reserves. They needto be replaced. It must, however, be recognized thattheir full replacement is neither desirable, nor possible.Its partial replacement seems to be a reality. Amongthe alternatives available, biofuels have great potentialin India, because of the availability of feedstock,environmental benefits, and the possibility of improvedrural economies.

The potential of a technology is one thing andits availability at the desirable ‘çost’ is another. Indiahas to overcome various bottlenecks before it becomesa commercial technology. The country has to answerseveral critical questions. The answer to thesequestions will decide the future of biofuels technologyin India. The questions, related to technology and itsdissemination, include: What kind of support, otherthan science, will be needed for its viability? Who willbe the major promoter of policy – agricultural sector,sugar industry or petroleum industry? What kind ofgovernment support is needed to make this technologyviable? Who will be the major promoter of large scalebiofuel technology? Other than science, what kind ofsupport is needed to make this a viable technology?Under what circumstances or conditions can refinersconsider participation in the ethanol industry? Thesequestions continue to bother researchers and planners.Large-scale lignocellulose based bioethanol technologywill require major changes in supply chaininfrastructure. It will require new ways of thinkingabout agriculture, energy infrastructure, and ruraleconomic development.

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second-generation biofuel – potential and perspectives in

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