Bioethanol from Agricultural
Waste Residues
Pascale Champagne
Department of Civil Engineering, Queen’s University, Kingston, ON K7L 3N6, Canada;
champagne@civil.queensu.ca (for correspondence)
Published online 3 December 2007 in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/ep.10242
Under the Kyoto Protocol, the Government of Can-
ada has committed to reducing its greenhouse gas
emissions by 6% from 1990 levels between 2008 and
2012. Ethanol-blended gasolines have the potential to
contribute significantly to these emission reductions.
Ethanol is derived from biologically renewable resour-
ces and can be employed to replace octane enhancers
and aromatic hydrocarbons or oxygenates. To date,
the ethanol production industry in Canada is com-
prised mainly of small-scale plants producing ethanol
primarily from agricultural crops as feedstock.
Research interests in the area of bioethanol produc-
tion from organic waste materials emerged in the late
1980. Significant advances in lignocellulosic material
extraction and enzymatic hydrolysis have been
reported in the last decade, however, continued
research efforts are essential for the development of
technically feasible and economically viable large-
scale enzyme-based biomass-to-ethanol conversion
processes. This research aims to develop and test an
enzyme-based biomass-to-ethanol conversion process,
which employs organic waste materials, such as live-
stock manures, as alternative sources of cellulosic
material feedstock. The source of the livestock ma-
nure, manure management practices and cellulose
extraction procedures have a significant impact on
the quantity and quality of the cellulosic materials
derived. As such, raw feedstock materials must be
carefully characterized to assess the impact of these
factors on the yield of bioethanol and residual end
products. The success of cellulose-to-ethanol conver-
sion processes for cellulose extracted from these waste
materials as feedstock is generally a function of cellu-
lose fiber pretreatment, enzyme selection and operat-
ing conditions. These will differ depending on the
source of the waste material feedstock. The long-term
benefits of this research will be to introduce a sus-
Ó 2007 American Institute of Chemical Engineers
Environmental Progress (Vol.27, No.1) DOI 10.1002/ep
tainable solid waste management strategy for a num-
ber of livestock manure and other lignocellulosic
waste materials; contribute to the mitigation in green-
house gases through sustained carbon and nutrient
recycling; reduce the potential for water, air, and soil
contamination associated with land disposal of or-
ganic waste materials; and to broaden the feedstock
source of raw materials for the ethanol production
industry. Ó 2007 American Institute of Chemical Engineers
Environ Prog, 27: 51–57, 2008
Keywords: bioethanol, manure, enzymatic hydroly-
sis, agricultural waste
INTRODUCTION
Under the Kyoto Protocol, the Government of Can-
ada has committed to reducing its greenhouse gas
emissions by 6% from 1990 levels between 2008 and
2012. Ethanol-blended gasolines have the potential to
contribute significantly to these emissions reductions.
Ethanol is an alternative fuel derived from biologically
renewable resources and can be employed to replace
octane enhancers such as methylcyclopentadienyl
manganese tricarbonyl (MMT) and aromatic hydrocar-
bons such as benzene or oxygenates such as methyl
tertiary butyl ether (MTBE). A potential source for
low-cost ethanol production is to utilize lignocellulosic
materials (crop residues, grass, sawdust, woodchips,
sludges, livestock manure), but the feasibility of using
these materials as a feedstock is often limited by the
cost of bioethanol production, which is relatively high
based on current technologies. The challenges are
generally associated with the low yield and the high
cost of the hydrolysis process. Research involving bio-
ethanol production from lignocellulosic waste materi-
als have included crop residues [1–4], municipal solid
wastes [5–8], forest products wastes [9–11], leaf and
yard wastes [12], municipal sludges [13], as well as a
few studies involving dairy and cattle manures [14–16].
The current bioethanol production industry in Canada
April 2008 51
Figure 1. Potential value-added products recovery from lignocellulosic waste materials.
from lignocellulosic materials is comprised of only a
few small-scale plants, producing ethanol mainly from
agricultural crops as feedstock materials. Iogen Corpo-
ration has developed a demonstration plant for the
conversion of wood, hay, straw and other agricultural
crop residuals to ethanol. Other ethanol production
plants currently in operation in Canada include:
Mohawk Oil Canada (wheat-based), Pound-Maker
Agventures (wheat-based), Commercial Alcohols
(corn-based), API Grain Processors (wheat-based),
and Tembec (forestry product-based).
In 1995, Dinel and Dumontet [17] introduced a
concept of sustainable waste management termed Bi-
ological and Chemical Integrated Recycling System.
They proposed a series of possible extraction and
fractionation procedures which would allow for the
recovery of a number of value-added products,
including cellulose, from various lignocellulosic waste
streams (see Figure 1). Livestock manures and
sludges contain large quantities of lignocelluloses,
polysaccharides, proteins and other organic materials.
The conversion of these materials to value-added
products has been recognized as an attractive waste
management approach. In addition, the recovery of
raw materials from organic wastes such as manures
and sludges, and their conversion to value-added
products has the potential to reduce the microbial
production of CO2 and CH4, thereby minimizing
potential environmental impacts and the strain on
nonrenewable resource reserves. However as feed-
stocks, organic wastes often have a complex physical
and chemical composition which has made it difficult
to utilize, and the limited research efforts to convert
these materials to higher-value chemicals and energy
on a commercial scale have been unsuccessful [14].
Bioethanol production utilizing lignocellulosic
materials generally takes place in three phases. First,
52 April 2008
delignification to liberate cellulose and hemicellulose
from lignin. Second, depolymerization of the carbo-
hydrate polymers (cellulose and hemicellulose) to
produce free sugars. Third, fermentation of mixed
hexose and pentose sugars to produce ethanol. In
addition, a preliminary step is often required when
employing livestock manure and sludges as feedstock
to obtain a better solids/liquid separation and to solu-
bilize and remove metals. Factors affecting the acid
and enzymatic hydrolysis of cellulose include poros-
ity of the waste materials, cellulose fiber crystallinity,
and lignin and hemicellulose content. Hydrolysis can
be significantly enhanced by introducing physical and
chemical fiber pretreatments including: grinding, py-
rolysis, steam explosion, ammonia fiber explosion
(AFEX), CO2 explosion, ozonolysis, acid hydrolysis,
alkaline hydrolysis, oxidative delignification, Organo-
solv process involving extraction with hot aqueous
ethanol and biological pretreatment of the lignocellu-
losic materials to remove lignin and hemicellulose
and reduce fiber crystallinity. Enzymatic hydrolysis
can be improved through the optimization of sub-
strate concentration, cellulase dosing, use of surfac-
tants, enzyme mixtures, and enzyme recycling. Simul-
taneous saccharification and fermentation (SSF) is an
effective process configuration which removes glu-
cose, an inhibitor to cellulase activity, thereby increas-
ing the yield and rate of cellulose hydrolysis [18].
DELIGNIFICATION OF AGRICULTURAL WASTE MATERIALS
Rivers and Emert [3] studied the effects of substrate
composition, cellulose crystallinity, and particle size
on the yields of enzymatic hydrolysis using bagasse
and rice straw. They concluded that each type of
lignocellulosic feedstock requires a specific pretreat-
ment in order to optimize enzymatic hydrolysis. The
Environmental Progress (Vol.27, No.1) DOI 10.1002/ep
Table 1. Percent metal reduction in hog manure
after HCl pretreatment.
Metal removal efficiency (%)
Metal 0.1 N HCl 1.0 N HCl
Al 4.9 6 2.7 39.3 6 22.2
Ca 19.4 6 5.8 69.4 6 6.5
Cu 88.0 6 4.3 81.3 6 12.0
Fe 70.9 6 2.8 91.9 6 4.2
Mg 82.3 6 1.5 92.5 6 2.3
Zn 90.3 6 2.3 82.1 6 12.4
Na 82.1 6 10.3 94.5 6 1.2
P 37.9 6 3.4 87.6 6 4.2
change in the crystallinity of rice straw following pre-
treatment appeared to correlate well with conversions
to either glucose or ethanol. Modification of the
lignocellulose matrix from its native state by alkaline
pretreatment resulted in the most significant increases
in enzymatic hydrolysis reinforcing the concept that
the nature of the lignocellulose matrix is a significant
limiting factor in the enzymatic hydrolysis of bagasse
and rice straw. In the research conducted at Carleton
University, (Ottawa, Canada) general laboratory-scale
chemical extraction and fractionation procedures and
protocols to be applied to agricultural crop residues,
municipal sludges and livestock manures were devel-
oped. Henderson et al. [19] elaborated an alkaline
delignification technique and reported that the alka-
line delignification technique was effective in separat-
ing cellulose as the nonhydrolyzable product (NHP)
from the lignin and hemicellulose as the hydrolysable
product (HP) of organic crop residues. Experimental
variables included the type and strength of hydrolyz-
ing base, temperature, and type of acid used for final
neutralization of the HP. The efficiency of separation
was assessed based on material recovery, and the car-
bon, nitrogen, phosphorous and ash recovered in
each of the two fractions. Using bagasse and corn
stovers as feedstock, the most efficient separations
from the bagasse (70% NHP) and corn stovers (50%
NHP) were obtained by using two alkaline hydrolysis
cycles of 0.5 N KOH at a temperature of 708C.
The alkaline delignification technique was then
adapted to extract and fractionate hog manure, a
more complex waste material, into NHP and HP. Levy
et al. [20, 21] and Champagne et al. [22] also demon-
strated 0.1–1.0 N acid pretreatments were effective in
separating metals from the hog manure. Although a
1.0 N solution showed a higher metal removal effi-
ciency (Table 1), it also had a negative impact on
NHP and HP recovery compared to the dilute 0.1 N
HCl pretreatment, which implied that organic matter
and nitrogen recovery could also be impacted (Table
2). Aside from Al (5%), more than 71% of metals
were separated from the hog manure using the 0.1 N
HCl pretreatment. It was suggested that better metal
separation could be achieved through coagulation
with the removal the colloidal matter. Upon further
investigation it was noted that nonionic polymers
Environmental Progress (Vol.27, No.1) DOI 10.1002/ep
Table 2. Total solids recovery in the HP and NHP
after pretreatment and alkaline hydrolysis.
HCl/KOH NHP (g TSNHP/ HP (g TSHP/
strength (N) 100g TSOM) 100g TSOM)
0.0/0.25 64.1 6 2.0 10.9 6 0.7
0.1/0.25 72.5 6 4.3 2.5 6 1.3
0.0/0.5 62.0 6 0.4 6.3 6 0.2
0.1/0.5 61.0 6 1.6 11.5 6 1.8
0.0/0.75 64.2 6 1.8 10.8 6 0.4
0.1/0.75 60.9 6 2.5 6.0 6 0.5
Figure 2. 13C NMR scan of HP resulting from 0.1 N
HCl pretreatment and 0.5N alkaline hydrolysis.
were most effective in removing the colloidal matter
from suspension. A separation of 61% NHP and
11.5% HP were observed using a 0.1 N HCL pretreat-
ment solution and a 0.5 N KOH alkaline hydrolysis
solution. The NHP accounted for the largest volatile
solids (62.4%) and carbon (60.1%) recovery, while
the HP accounted for the largest nitrogen (45.5%) re-
covery. FTIR and 13C NMR [22] spectra of the prod-
ucts suggested that the HP had a more complex
structure than the NHP. It was shown that the HP
possessed humic-like characteristics with the pres-
ence of alkenes and alkanes, proteinaceous carbon
and nitrogen, as well as aromatic carboxylic and phe-
nolic structures. The spectra for the NHP showed
mainly aliphatic chemical structures with the presence
of carbohydrate carbon as would be observed for cel-
lulose (Figures 2 and 3).
BIOETHANOL PRODUCTION FROM AGRICULTURAL WASTES
A research group at Washington State University
conducting research on the extraction of value-added
products from dairy and cattle manures [14–16],
developed a process for hydrolyzing lignocellulosic
materials from manure into fermentable sugars. They
reported that when raw dairy manure was pretreated
with 3% sulfuric acid at 1108C for 1 h, hemicellulose
was completely degraded to arabinose, galactose and
xylose. The pretreated materials were then treated
with cellulolytic enzymes to hydrolyze the cellulose.
The optimal enzyme loadings were identified as 13
FPU cellulose/g substrate and 5 IU b-glucosidase/g
April 2008 53
 13
Figure 3. C NMR Scan of NHP resulting from 0.1 N
HCl pre treatment and 0.5 N alkaline hydrolysis.

Figure 4. Pretreatment of poultry manures cellulose

substrate. The optimal temperature and pH were material fibers.
determined to be 468C and 4.8, respectively. A sub-
strate concentration of 50 g/L favored both glucose
concentration and glucose yield. It was also found
that a reduced particle size of 590 lm resulted in a
high glucose yield and further decreases in particle
size did not increase the yield which supports find-
ings by our research group. For each particle size
tested, the addition of a surfactant resulted in at least
20% improvement in glucose yield. The optimized
hydrolysis process achieved a glucose yield of 11.3 g/
100 g manure, which corresponded to 40% cellu-
lose.
The feasibility of producing ethanol from waste
materials comparing crop residues, poultry manure
and municipal sludges as low-cost lignocellulosic
feedstock was examined by Li and Champagne [23,
24]. The enzymatic hydrolysis of crop residues Figure 5. Glucose yields from poultry manure using
achieved relatively high glucose yields. At 408C, with different feedstock and fiber pretreatments.
an enzyme loading of 800 units/g substrate, the per-
centages of conversion in 24 h were 65.4% and 51.1%
for KOH-treated corn stovers and bagasse respec-
tively. The study showed that physical and/or chemi- ing, also exhibited an impact on the hydrolysis (see
cal NHP fiber pretreatments (grinding, drying and Figure 4). The glucose yields for poultry manure with
phosphorylation) had a great impact on the glucose various pretreatment are shown in Figure 5. At 408C,
yields from the saccharification process, primarily with an enzyme loading 400 units/g substrate, the
because it affected the enzyme access to the cellulose glucose yield for raw poultry manure was 7.1, and
material. It was concluded that higher glucose yields this increased to 10.2 when KOH pretreatment was
might be achieved by tailoring the sequence of fiber employed. These results were quite different to that
pretreatment processes depending on the feedstock observed with crop residues, where a 10-fold
from which the NHP fibers were derived. The increase in the glucose yield was noted as a result of
sequence of fiber pretreatments affected the enzy- KOH treatment. This would suggest that in the poul-
matic hydrolysis rate, where crop residues ground try manure, most of the cellulose was not as easily
before alkaline treatment and the use of a wet sub- extracted from other organic matters. Although the
strate yielded higher conversion rates, compared to integrated structure of cellulosic material might be
trials using dry substrate which was first treated with broken up during the KOH treatment, the broken
an alkaline solution followed by grinding. The hydro- structure could still not provide enough access for
lysis rate was also found to increase when half of the the enzymes to act on the cellulose molecules. In the
enzyme dose was added each at the beginning and case of KOH-treated manure, dried before enzymatic
midpoint of hydrolysis rather than entirely at the be- hydrolysis, the glucose yield increased to 13.9. These
ginning. results demonstrated that drying the KOH-treated ma-
In a study using poultry manure as a lignocellulo- nure did improve the glucose yield from enzymatic
sic feedstock [23, 24] it was found that the alkaline hydrolysis. It could be inferred that some of the or-
KOH delignification technique was effective in ganic matter might be degraded at high temperature
improving enzymatic hydroloysis to glucose. In addi- (up to 708C), thus more cellulose could be uncovered
tion, physical pretreatments such as drying and grind- during the drying process; or that the physical struc-

54 April 2008 Environmental Progress (Vol.27, No.1) DOI 10.1002/ep

Figure 6. Pretreatment of primary sludge cellulosic
material fibers.
ture of substrate became more accessible to the
enzyme due to the drying. The highest glucose yield
was noted for KOH-treated and ground poultry ma-
nure. With a conversion of 27.6, this represented a
twofold increase in yield compared to that of
unground manure. These results suggested that by
grinding the poultry manure, the substances that cov-
ered the cellulose were broken up, thus, exposing
the surface of the cellulose. In addition, reducing the
particle size of the substrate also created a larger sur-
face area for reaction. Thus, the substrate became
much more accessible to the enzyme. As a result, the
glucose yield increased significantly.
The cellulose content in primary sludge is mainly
from waste paper, however, the cellulosis fraction
must be separated from the noncellulosic compo-
nents prior to the enzymatic hydrolysis to maximize
glucose yield [23, 24]. Pretreatment approaches for
primary sludge include KOH, HCl, and HCl followed
by KOH alkaline delignification (see Figure 6). Figure
7 presents the glucose yields from primary sludge fol-
lowing various feedstock material and fiber pretreat-
ments. Both wet and dry substrates were tested for
each of them. All the trials were conducted at 408C
for a period of 24 h. Results indicated that when
KOH pretreatment was performed on primary sludge,
the conversion for wet substrate increased from
31.1% to 35.4%, which represented a 4.3% increase in
glucose yield. However, compared to that of crop
residues, which achieved an almost ten-fold increase
in the conversion percentages, the KOH pretreatment
was not as effective on the primary sludge. Similarly,
when the primary sludge was treated with HCl, the
glucose yield increased to 42.6% for wet substrate,
which was 11.5% higher than that observed without
acid and alkaline treatment. This suggested that
through HCl pretreatment, which would remove met-
als, enzyme inhibition due to metals may have been
reduced, resulting in a higher glucose yield from the
enzymatic hydrolysis. When both acid and alkaline
pretreatments were performed on the primary sludge,
Environmental Progress (Vol.27, No.1) DOI 10.1002/ep
Figure 7. Glucose yields from primary sludge using
different feedstock and fiber pretreatments.
the glucose yield reached 54.2% and 37.0% for wet
and dry substrates, respectively, which supported the
value of acid and alkaline pretreatments for effective
glucose conversion from enzymatic hydrolysis. More-
over, the results implied that considerable cellulose is
contained in the primary sludge on a dry mass basis,
and cellulosic material recovery and conversion to
ethanol might present a valuable waste management
alternative when employed as a wet feedstock. This
is contrary to what was noted with poultry manure,
which demonstrated a higher glucose conversion
when dry substrate was used. This might indicate that
the cellulose in primary sludge was more readily ac-
cessible to the enzymes and that drying and grinding
might not be required to improve the enzymatic hy-
drolysis as it was using with poultry manure.
CONCLUSIONS
Preliminary research using organic waste materials
as lignocellulosic feedstock for ethanol production
has shown great promise to date, however, further
research is essential to investigate its application
beyond the laboratory-scale. For instance, cellulose
extraction procedures and fiber pretreatments should
be optimized as a function of feedstock to maximize
the enzymatic hydrolysis and fermentation processes.
Enzyme recycling and the use of simultaneous sac-
charification and fermentation to integrate the ethanol
production process and achieve higher reaction rates
at lower costs would also be beneficial to the devel-
opment of this waste management strategy.
FUTURE RESEARCH NEEDS AND ANTICIPATED
SIGNIFICANCE OF RESEARCH
The research will result in the development of an
innovative waste management approach that uses ag-
ricultural waste materials as a renewable resource for
the extraction of a value-added product, cellulose,
and its conversion to bioethanol. This alternative
strategy would be of great benefit to the agricultural
industry where traditional manure management prac-
April 2008 55
tices have begun to inhibit rural development and
limit the establishment of livestock facilities due to
potential health risks for the general public. Munici-
palities and industries that rely heavily on landfilling
as a disposal strategy for their biosolids and process
sludge may be benefited as well. The long-term ben-
efits will be to reduce the potential for water, air, and
soil contamination associated with land disposal of
organic residuals; to broaden the feedstock source of
raw materials for the ethanol production industry;
and to contribute to the mitigation of greenhouse
gases through sustained carbon and nutrient recy-
cling.
In order to develop efficient and economically via-
ble bioethanol conversion processes for waste materi-
als, future research needs to include the elaboration
of existing laboratory procedures and protocols,
design recommendations, as well as economic and
environmental assessments generated from ongoing
research on lignocellulosic materials. Processes could
then be adapted and applied to other waste materials
which could be employed as lignocellulosic feedstock
for the production of bioethanol. These could include
municipal and industrial sludge, biosolids, leaf and
yard wastes, as well as other organic residues. As a
secondary research objective, the extraction of other
value-added products from various organic residues
could be investigated to broaden the biomass feed-
stock pool relevant to the emerging bioproducts
industry. For instance, the NHP could be used as a
substrate for microbial synthesis to produce ethanol,
as well as organic acids or complex proteins in the
form of live biomass. Products useful for agriculture
could be generated from the nitrogen and carbon-
rich HP, which would involve using the HP and the
residual fraction of the NHP that contribute effectively
in developing a sustainable waste management
approach.
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