Review TRENDS in Biotechnology
12
Bio-ethanol – the fuel of tomorrow
from the residues of today
B. Hahn-Hägerdal, M. Galbe, M.F. Gorwa-Grauslund, G. Lidén and G. Zacchi
Lund University, PO Box 124, S-221 00 Lund, Sweden, Getingevägen 60
The increased concern for the security of the oil supply
and the negative impact of fossil fuels on the environ-
ment, particularly greenhouse gas emissions, has put
pressure on society to find renewable fuel alternatives.
The most common renewable fuel today is ethanol
produced from sugar or grain (starch); however, this
raw material base will not be sufficient. Consequently,
future large-scale use of ethanol will most certainly have
to be based on production from lignocellulosic materi-
als. This review gives an overview of the new technol-
ogies required and the advances achieved in recent years
to bring lignocellulosic ethanol towards industrial pro-
duction. One of the major challenges is to optimize the
integration of process engineering, fermentation tech-
nology, enzyme engineering and metabolic engineering.
Introduction
One of the greatest challenges for society in the 21st
century is to meet the growing demand for energy for
transportation, heating and industrial processes, and to
provide raw material for the industry in a sustainable way.
An increasing concern for the security of the oil supply has
been evidenced by increasing oil prices, which during 2006
approached US$80 per barrel. More importantly, the
future energy supply must be met with a simultaneous
substantial reduction of green house gas emissions.
Actions towards this aim have been initiated. The Eur-
opean Commission plans to substitute progressively 20% of
conventional fossil fuels with alternative fuels in the trans-
port sector by 2020, with an intermittent goal set at 5.75%
in 2010. In the USA, the Energy Policy Act of 2005 requires
blending of 7.5 billion gallons of alternative fuels by 2012
[1], and recently the US President, in his state of the union
address, set the goal to replace more than 75% of imported
oil with alternative fuels by the year 2025 [2]. Liquid
biofuels from renewable resources, particularly from lig-
nocellulose materials, will have a substantial role in meet-
ing these goals (Box 1).
Ethanol has already been introduced on a large scale in
Brazil, the US and some European countries, and we
expect it to be one of the dominating renewable biofuels
in the transport sector within the coming 20 years. Ethanol
can be blended with petrol or used as neat alcohol in
dedicated engines, taking advantage of the higher octane
number and higher heat of vaporization; furthermore, it is
an excellent fuel for future advanced flexi-fuel hybrid
Corresponding author: Zacchi, G. (Guido.Zacchi@chemeng.lth.se).
Available online 16 October 2006.
www.sciencedirect.com 0167-7799/$ – see front matter ß 2006 Elsevier Ltd. All rights reserved. doi:10.1016/j.tibtech.2006.10.004
Vol.24 No.
vehicles. Currently, ethanol for the fuel market is produced
from sugar (Brazil) or starch (USA) at competitive prices.
However, this raw material base, which also has to be used
for animal feed and human needs, will not be sufficient to
meet the increasing demand for fuel ethanol; and the
reduction of greenhouse gases resulting from use of sugar-
or starch-based ethanol is not as high as desirable [3]. Both
these factors call for the exploitation of lignocellulose feed-
stocks, such as agricultural and forest residues as well as
dedicated crops, for the production of ethanol. This review
summarizes recent developments in the bioconversion
processes aimed at fuel ethanol production, with emphasis
on process integration. In particular, the concept that each
individual unit operation has to be developed and
optimized in relation to the preceding and subsequent
process steps will be discussed.
Overview of the conversion process
With so many advantages, why are there still no
production facilities using lignocellulosic materials?
Ethanol is currently produced from sugar cane and
starch-containing materials, where the conversion of
starch to ethanol includes a liquefaction step (to make
the starch soluble) and a hydrolysis step (to produce glu-
cose). The resulting glucose is then readily fermented.
Although there are similarities between the lignocellulosic
and the starch process, the techno-economic challenges
facing the former are large. There are several options for
a lignocellulose-to-ethanol process but, regardless of which
is chosen, the following features must be assessed in
comparison with established sugar- or starch-based
ethanol production.
(i) Efficient de-polymerization of cellulose and hemi-
cellulose to soluble sugars.
(ii) Efficient fermentation of a mixed-sugar hydrolysate
containing six-carbon (hexoses) and five-carbon (pen-
toses) sugars as well as fermentation inhibitory
compounds.
(iii) Advanced process integration to minimize process
energy demand.
(iv) Cost-efficient use of lignin.
The first step in the conversion of biomass to ethanol is
size reduction and pretreatment. In this review, only the
enzymatic process (Figure 1) will be discussed because it is
considered to be the most promising technology [4–6]. The
hemicellulose and cellulose polymers are hydrolyzed with
enzymes or acids to release monomeric sugars. The sugars
from the pretreatment and enzymatic hydrolysis steps
are fermented by bacteria, yeast or filamentous fungi,
550 Review TRENDS in Biotechnology Vol.24 No.12
Box 1. Advantages of lignocellulose-based liquid biofuels
 Biofuel sources are geographically more evenly distributed than
the fossil fuels; thus, the sources of energy will, to a larger extent,
be domestic and provide security of supply.
 Lignocellulosic raw materials minimize the potential conflict
between land use for food (and feed) production and energy
feedstock production. The raw material is less expensive than
conventional agricultural feedstock and can be produced with
lower input of fertilizers, pesticides, and energy.
 Biofuels from lignocellulose generate low net greenhouse gas
emissions, reducing environmental impacts, particularly climate
change.
 Biofuels might also provide employment in rural areas
although the enzymatic hydrolysis and fermentation can
also be performed in a combined step – a so-called simul-
taneous saccharification and fermentation (SSF). After
final purification (by distillation and molecular sieves or
other separation techniques), the ethanol is ready to be
used as a fuel, either neat or blended with petrol. A part of
the lignin, the principal solid part of the biomass remain-
ing, can be burnt to provide heat and electricity for the
process, whereas the rest is retained as a valuable co-
product. The most probable use today would be as an
ash-free solid fuel, but various technologies are under
development to convert it to a higher-value product, which
could form the basis for a new branch of industrial chem-
istry [7].
First, the biomass is converted to sugars. . .
It is only 40 years ago that the biodegradation of
lignocellulosics was first discussed [8]. Enzyme conversion
is substrate-specific without by-product formation, which
reduces inhibition of the following process steps. However,
enzyme-catalysed conversion of cellulose to glucose is slow
unless the biomass has been subjected to pretreatment,
which is also required to reach high yields and to make the
process commercially successful [9]: the pretreatment aims
Figure 1. Schematic flowsheet for the conversion of biomass to ethanol.
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to increase pore size and reduce cellulose crystallinity. In
acid-catalyzed pre-treatment, the hemicellulose layer is
hydrolyzed, whereas in alkali-catalyzed pretreatment,
mainly, a part of the lignin is removed and hemicellulose
has to be hydrolysed by the use of hemicellulases. Hence,
pretreatment is necessary to expose the cellulose fibres to
the enzymes or to at least make the cellulose more acces-
sible to the enzymes. An efficient pretreatment can sub-
stantially reduce the enzyme requirements, which make
up a large part of the production cost.
Pretreatment is usually assessed in a number of
ways: by enzymatic hydrolysis (EH) of the solid material
to determine the digestibility; by fermentation of the liquid
to assess the effect of potential inhibitors towards the
fermenting microorganism; and/or by simultaneous sac-
charification and fermentation (SSF) of the pretreated
material. The conditions for the assessment can vary,
particularly in terms of washed or non-washed solids
and the concentration of solids and enzymes in the EH
and SSF, making comparison of results from different
investigations difficult.
In an extensive study undertaken in the USA, where the
same batch of corn stover was pretreated using various
methods (e.g. dilute acid, AFEX, hot water treatment) and
then subjected to standard evaluation techniques, the
yields of sugars were found to be more or less the same
[10]. Total sugar yields – after pretreatment followed by
enzymatic hydrolysis – of around 90% or more were
reached, demonstrating that corn stover is an easily
degradable material. When corn stover was steam pre-
treated with small amounts of SO2, overall sugar yields
close to the theoretical value were obtained; steam pre-
treatment without a catalyst also resulted in 90% glucose
yield [11].
In countries such as Sweden, Canada and the USA,
much of the available biomass is softwood, which is more
difficult to hydrolyse than corn stover. For softwood, steam
pretreatment with the addition of an acid catalyst such as
 Review TRENDS in Biotechnology
H2SO4 or SO2 is a prerequisite to reach high sugar yields.
Acid increases the recovery of hemicellulose sugars and it
improves the enzymatic hydrolysis of the solid fraction; the
acid catalyst in steam pretreatment functions similar to
acid pulp cooking but with less liquid.
Steam pretreatment of SO2 impregnated spruce chips
yields a material that is relatively easy to hydrolyze and
ferment, whereas dilute acid impregnation results in a
material harder to ferment because of the generation of
inhibitory compounds [12]. With present bench-scale tech-
nology, it is possible to obtain around 300 litres of ethanol
per metric ton spruce, which is 70% of the theoretical
overall yield based on hexose sugars [12,13].
Steam pretreatment with the addition of a catalyst for
hydrolysis and improved enzymatic digestibility is the
closest to commercialisation. It has been widely tested
in pilot-scale equipment, for example, in the Iogen pilot
plant (Canada) [14], the Souston pilot plant (France) [15]
and in the pilot plant in Örnsköldsvik (Sweden), and is
used in a demonstration-scale ethanol plant at Iogen
(Canada). It is also to be used in Salamanca (Spain), in
a plant constructed by the company Abengoa.
Most probably, there will not be one general method
because different types of raw material require different
pretreatments. So far, methods such as ammonia fiber
explosion (AFEX), wet oxidation and liquid hot water
(LHW) treatment seem to be more successful for agricul-
tural residues [16–18], whereas steam pretreatment has
resulted in high sugar yields for both forestry and agri-
cultural residues. Glucose yields >90% and xylose yields
>80% were obtained after enzymatic hydrolysis, both with
and without the addition of an acid catalyst [12,19–21].
Acid-catalyzed pretreatment primarily solubilizes the
hemicellulose fraction into the liquid phase. For softwood,
the liquid mainly contains solubilized mannose in addition
to small amounts of xylose, arabinose, galactose and glucose.
The solid phase comprises lignin and cellulose, the latter of
which is subjected to enzymatic hydrolysis. The maximum
cellulase activity of most fungal-derived cellulases and b-
glucosidases is observed at 50 8C and at a pH of 4.0–5.0;
however, the optimal conditions vary with the hydrolysis
time and are dependent on the source of the enzymes [22].
Cellulases belong to two groups of enzymes known as endo-
glucanases (EG) and cellobiohydrolases (CBH), respec-
tively. EG randomly attack the cellulose chain, creating
free ends for CBH to cleave dimers of glucose (cellobiose)
off [23]. A third type of enzyme, b-glucosidase, which hydro-
lyzes cellobiose into two glucose molecules, is also necessary:
in the absence of b-glucosidase, end-product inhibition from
cellobiose will occur. Furthermore, compounds generated
during pretreatment might have an adverse effect on enzy-
matic hydrolysis. The enzymatic hydrolysis of spruce was
greatly improved when the liquid fraction from the pretreat-
ment step was replaced with a buffer solution [24]. This
could not be entirely ascribed to the reduction in end-pro-
duct inhibition, suggesting that inhibitory compounds had
also been removed.
In an initiative of the US Department of Energy, two
companies, Genencor International (http://www.genencor.
com/) and Novozymes Inc. (http://www.novozymes.com/),
were awarded $17 million each, with the goal to reduce
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Vol.24 No.12 551
the enzyme cost 10-fold (http://www.eere.energy.gov/).
However, the production cost of enzymes is still too high
and requires further reduction. One way to achieve this
is to use a fraction of the feedstock and/or the hydrolysate
for in situ enzyme production by fungi or other microorgan-
isms [25].
. . . and then the sugars are fermented to ethanol
Contrary to sucrose- and starch-based ethanol production,
lignocellulose-based production is a mixed-sugar fermen-
tation in the presence of inhibiting compounds – low
molecular weight organic acids, furan derivatives, pheno-
lics and inorganic compounds – released and formed during
pretreatment and/or hydrolysis of the raw material [26].
Lignocellulosic raw materials, in particular hardwood and
agricultural raw materials, can contain 5–20% (or more) of
the pentose sugars xylose and arabinose, which are not
fermented to ethanol by the most commonly used indus-
trial fermentation microorganism, the yeast Saccharo-
myces cerevisiae. Xylose is by far the most abundant
pentose sugar, whereas arabinose can constitute as much
as 14–15% in corncob hulls and wheat bran, respectively.
Consequently, most research efforts have been devoted to
the development of efficient xylose-fermenting microorgan-
isms [27,28].
Xylose-fermenting microorganisms are found among
bacteria, yeast and filamentous fungi [29]. Anaerobic
bacteria ferment pentoses, but are inhibited already at
low sugar and ethanol concentrations. In addition,
the ethanolic fermentation occurs with considerable
by-product formation, which reduces the ethanol yield
[30]. Natural xylose-fermenting yeast, notably Pichia
stipitis CBS 6054, ferment xylose to ethanol with
reasonable yield and productivity; however, these yeast
strains are inhibited by compounds generated during
pretreatment and hydrolysis of the lignocellulose mate-
rial [31]. Filamentous fungi tolerate inhibitors but are
too slow for a competitive industrial process. Therefore,
efforts have predominantly been made to obtain recom-
binant strains of bacteria and yeast able to meet the
requirements of industrial lignocellulose fermentation
(Figure 2).
Figure 2. Strains that are metabolically engineered for ethanol production
from pentoses. In essence, either the tail end, as in Escherichia coli and
Klebsiella oxytoca, or the front end of metabolism, as for Saccharomyces
cerevisiae and Zymomonas mobilis, have been engineered. Abbreviation: rec,
recombinant.
552 Review TRENDS in Biotechnology Vol.24 No.12

Pentose-fermenting Escherichia coli [32] and Klebsiella
oxytoca [33] have been generated by introducing ethanolo-
genic genes from Zymomonas mobilis (Figure 2). At the
same time, the first xylose-fermenting S. cerevisiae strain
was generated through the introduction of genes for xylose-
metabolizing enzymes from P. stipitis [34] (Figure 2). Later
xylose-fermenting strains of S. cerevisiae were constructed
by introducing the genes encoding xylose isomerase from
the bacterium Thermus thermophilus [35] and the anae-
robic fungus Piromyces sp. [36], respectively. For xylose-
using S. cerevisiae, high ethanol yields from xylose also
require metabolic engineering strategies to enhance the
xylose flux. This was independently demonstrated in an
SSF set-up for corn stover, where glucose and xylose were
fermented simultaneously [37], and in a recombinant
strain with increased carbon flux [38]. Z. mobilis also
efficiently produces ethanol from the hexose sugars glucose
and fructose but not from pentose sugars, although a xylose
fermenting Z. mobilis was generated by introducing a
xylose-metabolizing pathway from E. coli [39]. More
recently, the obligatory anaerobic bacterium Thermoa-
naerobacterium saccharolyticum has been genetically engi-
neered for improved ethanolic fermentation (Joe Shaw
et al., oral presentation, Nashville, 2006).
E. coli and K. oxytoca naturally metabolize arabinose,
such that the ethanologenic strains ferment all lignocellu-
lose-derived sugars [40]; furthermore, xylose- and arabi-
nose-fermenting strains of Z. mobilis have been
constructed [41]. Because yeast only ferment arabinose
to ethanol in rich media [42,43], S. cerevisiae has been
engineered for arabinose use by introducing both bacterial
[44] and fungal genes [45] encoding arabinose-metaboliz-
ing enzymes, where the fungal approach did not result
in appreciable arabinose fermentation. The functional
arabinose-metabolizing pathway has recently been
integrated into the diploid xylose-fermenting S. cerevisiae
strain TMB 3400, and co-usage of xylose and arabinose has
been demonstrated [46].
In addition to being able to ferment both hexose and
pentose sugars, the fermenting microorganisms must do
this in the presence of inhibiting compounds such as weak
acids, furan derivatives and phenolics [47]. Detoxification
by chemical or physical methods before fermentation
reduces the concentration of inhibitors and improves the
performance in the fermentation step. Treatment with
Ca(OH)2 (so-called ‘overliming’) and ion-exchange treat-
ment increased the fermentability significantly [48]; how-
ever, this was at the expense of increased process cost and
sugar loss [49]. Furthermore, Ca(OH)2 is unlikely to be
accepted in a full-scale ethanol plant, where precipitation
of calcium salts might foul distillation columns, evapora-
tors and heat-exchanger surfaces [50,51].
In ethanolic yeast fermentation, in-situ biological detox-
ification occurs when carbonyl compounds – furans and
phenolics – are reduced to the corresponding alcohols
[52,53], which are less inhibitory to yeast [54]. This phe-
nomenon has been successfully exploited in online probing
process control in fed-batch fermentation, by adjusting the
feed rate of hydrolysate to the intrinsic capacity for inhi-
bitor conversion of the yeast [55]. In such cases, the fer-
mentation productivity will be a function of the inhibitor
concentration, the conversion capacity of the yeast and the
quality of the process control. Only recently it was demon-
strated that the overexpression of a gene encoding alcohol
dehydrogenase activity improved the fermentation of a
5-hydroxymethyl furfural-containing medium [56].
The competitiveness of fuel ethanol research makes
strain comparisons difficult. Literature data are hampered
by incomplete reporting, differences in fermentation
conditions (including media composition and oxygenation),

Table 1. Latest fermentation results on hydrolysate from various recombinant and native xylose-fermenting organisms.
Organism Hydrolysate Detoxification Fermentation mode Yield Fermentation time Sp. Refs
g/g initial sugar (hours) productivity
g/g cells.h
Escherichia coli KO11 Bagasse + n/a 0.49 n/a 0.37 [66]
hemicellulose 1
Corn fiber 2 + Batch 0.39–0.41 3 93–102 n/a [67]
Batch 0.30–0.38 4 29–68
Fed-batch 0.35–0.39 5 118
E. coli FBR5 Corn stover + n/a 0.46 n/a 0.214 [66]
Rice hull Batch 0.43 64 n/a [68]
Rice hull +3 Batch 0.40 39 n/a [68]
Zymomonas Corn stover 7 +3 Batch 0.42 n/a n/a [66] [69]
mobilis 8b 6
Pichia stipitis 8 Wheat straw +3 n/a 0.41 n/a 0.44 [66,70] 9
P. stipitis CBS 5773 Spent sulfite liquor + 10 Continuous 0.35 - 0.05 [66]
Saccharomyces Corn fiber +3 Batch 12 0.36 48 0.072 13 [71]
cerevisiae 424A (LNF- Corn stover +3 Batch 12 0.41 24 0.044 13 [71]
11
ST) Corn stover Batch 12 0.45 55 0.048 13 [9]
S. cerevisiae TMB3006 Spruce Fed batch 0.37 n/a 0.66 [66]
S. cerevisiae TMB3400 Spruce Fed batch 0.43 n/a 0.25 [66]
Corn stover 14 SSF, Batch 15 0.33 16 96 0.036 [37]
SSF, Fed batch 15 0.30 16 96 0.076
1
Supplemented with 2.5% (w/v) corn steep liquor; 2 supplemented with 5% (w/v) corn steep liquor; 3 overliming; 4 neutralization by anion exchange; 5 membrane
pervaporation; 6 acid-tolerant derivative of ZM4/AcR (pZB5); 7 80% hydrolysate supplemented at 100 g/l glucose; 8 adapted strain of NRRL Y-7124; 9 calculated from Nigam 2001
[70]; 10 rotoevaporated spent sulfite liquor; 11 optimized pH-controlled liquid hot water pretreatment; 12 Semi-anaerobic fermentations with 10 g/l yeast extract added; 13
Calculated with the given initial cell dry weight of 8.5 g/l; 14 0.05% water insoluble solid; 15 1 g/l yeast extract added; 16 % of theoretical based on the glucose and xylose content
in the raw material, (i.e. taking into account both hydrolysis and fermentation yields).
Abbreviations. n/a, not available; SHF, separate hydrolysis and fermentation; SSF, simultaneous saccharification and fermentation.

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 Review TRENDS in Biotechnology
vastly different raw materials, and pretreatment and
hydrolysis conditions [29]. Table 1 summarizes the most
recently published results on the fermentation of lignocel-
lulose hydrolysate with natural and recombinant bacteria
and yeast. In essence, so far only recombinant S. cerevisiae
strains have been able to ferment xylose in non-detoxified
hydrolysates, where the fed-batch technology also permits
the fermentation of extremely inhibitory softwood hydro-
lysates.
Moving on from research to a commercial process
The estimated cost of producing ethanol from cellulosic
materials varies widely between investigations, as shown
both in some earlier review papers [57,58], with production
costs in the range of 0.28 to 1.0 US$/l ethanol, and in more
recent techno-economic evaluations [13,59,60]. However,
most cost estimations are based on laboratory-scale and, to
some extent, pilot-scale data for individual process steps
and should be treated with caution. The cost of raw mate-
rial, which varies considerably between different studies
(US$22–US$61 per metric ton dry matter), and the capital
costs, which makes the total cost dependent on plant
capacity, contribute most to the total production cost.
The cost for hydrolysis, particularly for the enzymatic
process, is also a major cost contributor.
Raw material cost is reduced by using the whole crop for
products and co-products. High ethanol yield requires
complete hydrolysis of both cellulose and hemicellulose
with a minimum of sugar degradation, followed by efficient
fermentation of all sugars in the biomass. In the short-
term, co-products are likely to be used for the production of
fuel, heat and electricity; however, in the long term,
bioethanol technology will form the basis for the sustain-
able production of commodity chemicals and materials in
future biorefineries. High co-product yield requires
reduced energy demand for ethanol production. This is
achieved when high solids concentrations (Figure 3) are
combined with integration of energy-intensive process
steps (e.g. pretreatment, distillation, evaporation and
Figure 3. Ethanol production cost as a function of water-insoluble solids (WIS) in
the SSF step (expressed as weight-% of total material) for production of ethanol
from spruce, based on a production capacity of 200 000 DM raw material per year.
Broken line: use of meachanical vapour recompression in evaporation (1
SEK = $0.13). This figure clearly shows the importance of working at high WIS.
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Vol.24 No.12 553
drying). In SSF, 12 % WIS (water-insoluble solids) result
in an ethanol concentration >4 wt-% (weight-%; kg ethanol
per 100 kg solution), which is necessary to reduce the
energy demand in the distillation steps (A. Wingren,
PhD thesis, Lund University, 2005). Further reductions
in the energy demand can be obtained by recycling certain
process streams, to minimise the amount of fresh water
used [61]. However, high solids concentrations and recy-
cling of process streams increase the concentration of
compounds that are inhibitory to enzymatic hydrolysis
and fermentation, necessitating detoxification or fed-batch
technology, as described previously. It also results in high
viscosity, which limits mixing and pumping.
Process integration reduces the capital costs. In the
separate hydrolysis and fermentation (SHF) process, cel-
lulose is first hydrolyzed to glucose and then glucose is
fermented to ethanol. The primary advantage of SHF is
that hydrolysis and fermentation occur at optimum condi-
tions; the disadvantage is that cellulolytic enzymes are
end-product inhibited so that the rate of hydrolysis is
progressively reduced when glucose and cellobiose accu-
mulate [24]. Product inhibition was the rationale for the
first report on simultaneous saccharification and fermen-
tation (SSF) of cellulose [62]: in SSF, hydrolysis and fer-
mentation occur simultaneously in the same vessel, and
the end-product inhibition of the enzymes is relieved
because the fermenting organism immediately consumes
the released sugars. Furthermore, the fermentation seems
to decrease the inhibition of the enzymes by converting
some of the toxic compounds present in the hydrolysate
[24]. This increases the overall ethanol productivity, the
ethanol concentration and the final ethanol yield [12,63]
(Figure 4).
More recently, the SSF technology has proved advanta-
geous for the simultaneous fermentation of hexose and
pentose sugars (so called SSCF). In SSCF, the enzymatic
hydrolysis continuously releases hexose sugars, which
increases the rate of glycolysis such that the pentose
sugars are fermented faster and with higher yield [37].
Figure 4. Overall yields of fermentable sugars and ethanol from steam-pretreated
spruce. Abbreviations: EH, sugar yield after separate enzymatic hydrolysis; SSF,
ethanol yield after SSF; SO2, after impregnation with sulfur dioxide; H2SO4, after
impregnation with sulfuric acid. Adapted from data presented in [12].
554 Review TRENDS in Biotechnology Vol.24 No.12
Figure 5. Biorefinery – integration of a combined heat and power plant with an
ethanol production plant.
Further process integration can be achieved by performing
both hydrolysis and fermentation in a single reactor, using
one or a mixture of microorganisms that produce all the
required enzymes and ferment all sugars – so-called con-
solidated bioprocessing (CBP) [64]. However, no such
microorganisms are currently available, and the concept
is subject to further research.
The economic analysis [60] of the cellulosic bioethanol
process shows that reliable cost estimations require
laboratory results are verified in pilot and demonstration
plants, where all steps are integrated into a continuous
process. This also provides the possibility to explore the
benefits of process integration to reduce the number of
process steps and the energy demand, and to recirculate
process streams to eliminate the use of fresh water and to
reduce the amount of waste streams. Currently, the Iogen
Corp. (http://www.iogen.ca/) demo-plant is the only operat-
ing plant for the production of bioethanol from lignocellu-
lose using the enzymatic hydrolysis process. The plant can
handle up to 40 tonnes per day of wheat, oat and barley
straw and is designed to produce up to 3 million litres of
cellulose ethanol per year. Abengoa Bioenergy (http://
www.abengoabioenergy.com) is also constructing a pilot
plant in York, USA, to convert residual starch, cellulose
and hemicellulose – mainly corn stover – to bioethanol and
high-protein feed. In Salamanca, the same company con-
structed a demonstration plant integrated with a fuel-
ethanol-from-grain plant, producing 195 million liters. In
the demonstration plant, an additional 5 million liters of
ethanol per year will be produced from cellulose, mainly
Box 2. Major research challenges
 Improving the enzymatic hydrolysis with efficient enzymes,
reduced enzyme production cost and novel technology for high
solids handling.
 Developing robust fermenting organisms, which are more
tolerant to inhibitors and ferment all sugars in the raw material
in concentrated hydrolysates at high productivity and with high
ethanol concentration.
 Extending process integration to reduce the number of process
steps and the energy demand and to re-use process streams to
eliminate the use of fresh water and to reduce the amount of
waste streams.
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from wheat staw. In Sweden, a fully integrated pilot plant
for ethanol production from softwood, comprising both two-
stage dilute acid hydrolysis and the enzymatic process, was
taken into operation in mid 2004. The pilot has a maximum
capacity of 2 ton (DM) wood per day (http://www.etek.se).
The step from pilot- and demo-scale production of lig-
nocellulosic ethanol to competitive full-scale production
requires further reduction of the production cost. One
approach to this is the integration of ethanol production
with a combined heat and power plant (Figure 5) or with a
pulp and paper mill. This has been estimated to reduce the
ethanol production cost by up to 20 percent for conditions
prevailing in Sweden [65] and it is the main strategy
pursued in the Swedish cellulosic ethanol effort. Similar
conclusions were reached in a study on co-production of
ethanol and electricity from softwood, based on conditions
in California. Another option is to integrate cellulosic
ethanol production with starch-based ethanol production
to use the whole agricultural crop. For the immediate
future, we believe that these integrated plant concepts
will be used in the first successful industrial scale produc-
tion of lignocellulosic fuel ethanol. The transition of lig-
nocellulosic fuel ethanol production into a mature
industrial technology requires research and development
efforts in the areas summarized in Box 2.
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Progress in Biophysics and Molecular Biology

January 2007: Vol. 93
Themed issue on
Effects of ultrasound and infrasound relevant to human health
Guest edited by Alastair McKinlay

26 articles, including:

* Medical diagnostic applications and sources – Tony A. Whittingham

* Therapeutic applications of ultrasound – Gail ter Haar
* Medical ultrasound imaging – Jørgen Arendt Jensen
* Medical and non-medical protection standards for ultrasound and infrasound – Francis A. Duck
* Rapporteur report: Mechanisms and interactions – Timothy G. Leighton
* Quantification of risk from fetal exposure to diagnostic ultrasound – Charles C. Church
* Ultrasound, microbubbles and the blood-brain barrier – Stephen Meairs
* Shear stress in cells generated by ultrasound – Junru Wu
* The enhancement of bone regeneration by ultrasound – Lutz Claes
* Cardiac imaging: The biological effects of diagnostic cardiac ultrasound – Maria Grazia Andreassi
http://www.sciencedirect.com/science/journal/00796107

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