Comparison of alkaline pulping with steam explosion for glucose production from rice straw

Article abstract of DOI:10.1016/j.carbpol.2010.08.046

Agricultural residues, such as rice straw, are renewable, largely unused, and abundantly available resources. They contain cellulose and hemicellulose, which could be used to produce ethanol and many other value-added products. The current research invest

Full text of DOI:10.1016/j.carbpol.2010.08.046

Carbohydrate Polymers 83 (2011) 720–726
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Carbohydrate Polymers
journal homepage: www.elsevier.com/locate/carbpol
Comparison of alkaline pulping with steam explosion for glucose production
from rice straw
Maha M. Ibrahima,∗, Waleed K. El-Zawawya, Yasser R. Abdel-Fattahb, Nadia A. Solimanb,
Foster A. Agblevorc
a National Research Center, Cellulose and Paper Department, El-Tahrir St., Dokki, Cairo, Egypt
b Genetic Engineering and Biotechnology Research Institute, Mubarak City for Scientific Research and Technology Applications,
Universities and Research Institutes Zone, 21934 New Burg El-Arab City, Alexandria, Egypt
c Virginia Polytechnic Institute and State University, Department of Biological Systems Engineering, 213 Seitz Hall, Blacksburg, VA 24061, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
Agricultural residues, such as rice straw, are renewable, largely unused, and abundantly available
resources. They contain cellulose and hemicellulose, which could be used to produce ethanol and many
other value-added products. The current research investigates the utilization of rice straw as a lignocel-
lulosic biomass feedstock to produce a value-added product. Investigation was carried to convert the rice
straw into glucose which can be further fermented to produce ethanol. Different pretreatment methods,
such as chemical pretreatment process using alkaline pulping and steam explosion were applied in this
study to pretreat the lignocellulosic biomass. A Spezyme CP® cellulase enzyme was used in the exper-
iment to hydrolyze the pretreated material into glucose. The total reducing sugars produced from the
enzymatic hydrolysis of cellulose was measured by the dinitrosalicylic acid (DNS) method. The data from
the enzyme hydrolysis time study were analyzed to provide information on enzyme hydrolysis rates.
The results indicated that 28.9–58.4 g/L of glucose can be produced from rice straw depending on the
pretreatment method.
Received 5 August 2010
Accepted 17 August 2010
Available online 24 August 2010
Keywords:
Agricultural residues
Rice straw
Pretreatment process
Alkaline pulping
Steam explosion
Glucose production
© 2010 Elsevier Ltd. All rights reserved.
1. Introduction
and subsequently fermented to ethanol by microorganisms (Millati,
Niklasson, & Taherzadeh, 2002; Palmqvist & Hahn-Hägerdal, 2000).
Research in utilization of lignocellulosic plant materials for
bioethanol production has grown significantly over the last few
decades as the depletion of non-renewable fuels and increasing
greenhouse gas emissions continue to create an increasing need for
an alternative non-fossil transportation fuel. Agricultural residues,
such as rice straw, are natural lignocellulosic materials. Lignocel-
lulosic materials are renewable, largely unused, and abundantly
available sources of raw materials for the production of fuel ethanol.
They can be obtained at low cost from a variety of resources, e.g. for-
est residues, municipal solid waste, waste paper, and crop residue
resources (Cheng & Zhu, 2008). These materials predominantly
contain a mixture of carbohydrate polymers (cellulose and hemi-
cellulose), lignin, extractives, and ashes. The term “holocellulose” is
often used to describe the total carbohydrate contained in a plant or
microbial cell. Holocellulose is therefore comprised of cellulose and
hemicellulose in lignocellulosic materials. Sugars polymerized in
form of cellulose and hemicellulose can be liberated by hydrolysis
Pretreatment of lignocelluloses is intended to disorganize the
crystalline structure of macro- and microfibrils, in order to release
the polymer chains of cellulose and hemicellulose, and/or modify
the pores in the material to allow the enzymes to penetrate into
the fibers to render them amenable to enzymatic hydrolysis (Galbe
& Zacchi, 2002). Pretreatment should be effective to achieve this
goal, avoid degradation or loss of carbohydrate, and avoid forma-
tion of inhibitory by-products for the subsequent hydrolysis and
fermentation; obviously, it must be cost-effective (Sun & Cheng,
2002).
Steaming with or without explosion (autohydrolysis) is one
of the popular pretreatment methods of lignocellulosic materials.
Steam pretreatment removes the major part of the hemicellulose
from the solid material and makes the cellulose more susceptible
to enzymatic digestion. In this method the biomass is treated with
high pressure steam. The pressure is then swiftly reduced, in steam
explosion, which makes the materials undergo an explosive decom-
pression. Steam explosion is typically initiated at a temperature of
160–260 ◦C for several seconds to a few minutes before the mate-
rial is exposed to atmospheric pressure (Kurabi et al., 2005; Sun &
Cheng, 2002; Varga, Reczey, & Zacchi, 2004).
∗
Corresponding author. Tel.: +20 124436805.
E-mail address: mwakleed@hotmail.com (M.M. Ibrahim).
0144-8617/$ – see front matter © 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.carbpol.2010.08.046

M.M. Ibrahim et al. / Carbohydrate Polymers 83 (2011) 720–726
721
On the other hand, chemical pulping methods attempt to dis-
solve the lignin with a minimum of dissolution or degradation of
the cellulose and hemicellulose components of the fibers under
high temperature and pressure. Historically, the most successful
approaches to achieve these goals have employed NaOH (the soda
2.2.2. Water and alkali extraction
The steam-exploded fiber (10 g, oven dry fiber, odf) was
extracted with water at 80 ◦C for 1 h using fiber to water ratio of
1:10. The mixture was vacuum filtered in a Buchner funnel and
then washed with 500 mL water. The water-extracted fiber was
further extracted with 20 wt.% sodium hydroxide (NaOH) at 80 ◦C
for 1 h using a fiber to liquor ratio of 1:10. The mixture was filtered
and the alkali-extracted water was collected and the residue was
washed with water until the pH was neutral. The alkali-extracted
fiber was air-dried at ambient laboratory conditions and stored in
a cold room.
(the sulfite process), and alkaline S2− (the kraft
2−
process), SO3
process) (Rojas & Hubbe, 2004).
The present study was conducted to compare alkaline pulp-
ing (soda process), as the one used in Egyptian pulp mills for rice
straw, and steam explosion as alternative pretreatment methods
for ethanol production from starchy materials. The agricultural
residue was subjected to the pretreatment then followed by enzy-
matic hydrolysis. The results of the glucose concentration in the
culture sample were determined using enzymatic colorimetric
method.
2.3. Compositional analysis of the steam exploded and pulped
material
The holocellulose (TAPPI T257 om-85), Klason lignin (TAPPI
T222 om-88), and ash content (TAPPI T211 om-85) of the steam-
exploded and pulped fiber samples were determined and collected
in Table 1.
2. Experimental
2.1. Materials and chemicals
Rice straw, as an Egyptian agricultural residue, was used as a
source for the lignocellulosic material. A Spezyme CP® cellulase
enzyme preparation (Lot #301-03195-109, Genencor, Interna-
tional, Rochester, NY) was used for the enzymatic hydrolysis for
glucose production.
2.4. Fiber characterization
2.4.1. FTIR spectra
Fourier transform infrared (FTIR) spectroscopy was used to eval-
uate the fiber resulting from alkaline pulping and steam explosion.
Also, it was used to assess the crystallinity index (C.I.) of the fibers
according to Hulleman, Vanhazendonk, and Vandam (1994) and
Wistara, Zhang, and Young (1999), which is based on the ratio of
the absorbance of the bands at 1429 (CH2 scissoring) and 893 cm−1
(C(1) group vibration).
The FTIR spectra of steam exploded and alkaline pulping rice
straw were performed using a Thermo-Nicolet Model 670 Instru-
ment (Thermo Electron, Inc., Madison, WI).
2.2. Pretreatment
2.2.1. Steam explosion and pulping process
The steam explosion and alkaline pulping of the rice straw were
carried out as a pretreatment process. Steam explosion of the agri-
cultural residue sample was carried out in a 25-L batch reactor
located at the Thomas M. Books Forest Products Center, Blacks-
burg, VA. The procedure reported by Jeoh and Agbelvor (2001) was
used in this study. About 1 kg of agricultural waste sample were
weighed and loaded into a 25-L batch steam explosion chamber
and the ball valve was closed. Saturated steam was admitted into
the chamber, and the biomass temperature was raised to 220 ◦C.
When the material attained the reaction temperature, timing of
the reaction was started. For this experiment, 240 s was used for
the reaction. At the end of the allotted steaming time, the valve
was opened for the “explosive depressurization” to occur. The
steam-exploded material shot through the connecting piping and
collected in the collection bin. The product came out in a sludge
form and the fibers were bagged, weighed and stored in a cold
room.
The pulping was done to the same waste, i.e. rice straw, on
an electrically heated rotatory autoclave located at the National
Research Center, Egypt, and the conditions were as follows: total
chemical as NaOH was 10% (wt/wt), and liquor to fiber ratio was
6:1. The fibers were cooked at 170 ◦C for 2 h. After cooking, the pres-
sure was released to atmosphere. The pulped fiber was washed with
water until neutrality, then air-dried.
2.4.2. Scanning electron microscopy
Scanning electron microscopy (SEM) was used to investigate the
morphology of the different types of fibers, i.e. steam exploded and
alkaline pulped rice straw using a JEOL JXA-840 A electron micro-
probe analyzer (JEOL USA Inc., Peabody, MA). The specimens for
the fibers were coated with gold/palladium and observed using an
applied tension of 30 kV.
2.5. Hydrolysis
Enzymatic hydrolysis is the most promising alternative to the
use of dilute acid, but it is certainly not a replacement process. We
aimed to develop an optimum condition of using the enzyme for
the pretreated agricultural residues to convert the resulted cellu-
lose into glucose. The condition is to adjust the temperature, pH,
and time of incubation. The total reducing sugars produced from
the enzymatic hydrolysis of cellulose was measured by the dini-
trosalicylic acid (DNS) method.
Table 1
Chemical analysis for raw material and both steam-exploded and pulped rice straw fibers.
Raw material and unbleached
steam exploded and pulped
fibers
Severity
Temperature (◦C)
Retention
time (min)
Holocellulose (%)
␣-Cellulose (%)
Lignin (%)
Ash (%)
C.I.
parametera
Rice straw raw material
Steam rice straw
Alkali-extracted
–
4.15
–
–
220
80
–
4
120
68.09
50.74
86.90
38.83
44.95
78.37
14.55
23.84
4.20
16.34
22.75
12.00
–
2.33
–
steam-exploded rice straw
Pulped rice straw
–
170
120
71.61
58.37
10.32
13.70
2.00
a
The severity parameter is a relation between the temperature and the retention time.

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M.M. Ibrahim et al. / Carbohydrate Polymers 83 (2011) 720–726
2.5.1. Enzyme hydrolysis time study
rice straw was higher than those of the steam-exploded rice straw.
The steam explosion technology involves short time cooking of
biomass feedstock at high temperature and pressure in saturated
steam. Following the cooking, the digester is discharged by a fast
decompression. This process can be characterized as a thermo-
mechanochemical process in which hydrolytic reactions as well
as mechanical forces acting in the decompression step lead to the
massive liberation of fibers. The main chemical reactions that occur
during the process are the cleavage of easily hydrolyzed glyco-
sidic bonds, cleavage of some ether bonds in lignin, and cleavage of
lignin–carbohydrate linkages. In addition, the steam explosion pro-
cess induces, as mentioned above, a re-condensation of the lignin
onto the fibers, as shown from the higher lignin content for the
steam-exploded rice straw (Table 1), and hence a higher ash con-
tent. During the alkaline pulping, the lignin dissolved and separated
in the black liquor.
Samples of rice straw, at different pretreatments, i.e. steam
explosion, alkaline pulping, and steam explosion-alkali extraction,
were selected for an initial study of enzyme hydrolysis. The mate-
rials were either steam-exploded or pulped according to the same
experiment as described above.
A Spezyme CP® cellulase enzyme preparation (Lot #301-03195-
109, Genencor, International, Rochester, NY) was used for the
hydrolysis. The pretreated fibers (10 g odf) in 100 mL sodium
acetate buffer at pH 4.7 were sterilized first at 121 ◦C for 20 min.
2.0 mL of Spezyme® (64 FPU/mL) was added after cooling to the
above samples. The mixture was incubated in a shaker bath at 50 ◦C
and 75 rpm for 4 days. After a reaction time of 2, 3 and 4 days, the
reaction was stopped by placing the sample in a boiling beaker
of water for 5 min. The mixture was vacuum filtered and the glu-
cose concentration was analyzed using the enzymatic colorimetric
method. A reference for pure cellulose was used to compare for the
waste material.
This means that, for steam explosion, the treatment depoly-
merizes and degrades hemicellulose while most of the crystalline
structure of the cellulose is preserved. After delignification of the
pretreated material, less Klason lignin can be detected.
2.5.2. Enzyme hydrolysis calculations
Steam-exploded rice straw was also investigated microscopi-
cally. Steam explosion is considered one of the most promising
pretreatment technologies. Fig. 1A and B shows the SEM images
of steam-exploded rice straw under various magnifications. Fig. 1A
clearly shows the shape and size distribution of the fibers in
steam-exploded rice straw at severity of 4.15. Well-separated
fibers can be seen with an average diameter of 5.114 ␮m. The
SEM image (Fig. 1B) of one individual fiber at larger magnifica-
tion shows many holes on the surface of steam-exploded rice
straw. This is probably due to the removal of some fibrous lay-
ers. In a previous work (Ibrahim, Agblevor, & El-Zawawy, 2010) we
proposed that these changes to the steam-exploded fibers result
from the removal of very reactive amorphous cellulose on the sur-
face.
Data from the enzyme hydrolysis time study were analyzed to
provide information on enzyme hydrolysis rates. Enzyme hydrol-
ysis rates were computed as concentration of glucose released per
hydrolysis time:
dS
dt
Glut − Glu0
t − t0
v =
=
(1)
where v= enzyme hydrolysis rate (mg/mL glucose per hour),
Glut = concentration of glucose at time t (mg/mL), Glu0 = initial glu-
cose concentration at time = 0 h (mg/mL), t = hydrolysis time (h),
and t0 = time = 0 h.
3. Results and discussion
3.1. Fiber composition and SEM
Moreover, the most apparent effect of the alkaline pulp-
ing pretreatment apart from a color change from yellow into
dark brown is the defibration, or separation of individual fibers
and cell types of rice straw. The pretreated material is quite
heterogeneous and contains larger pieces, which are easily rec-
ognized (Fig. 1C). When looking more closely at the pretreated
fibers it becomes apparent that the surface appeared smooth and
was covered with deposits (Fig. 1D). No holes or cracks were
seen in the fibers, but semispherical formations appear to be
emerging from the cell wall (Fig. 1D). Selig et al. (2007) hypoth-
esized that the observed droplet formations evolve from the
lignocellulosic matrix during pretreatment, potentially depositing
back onto the cell wall surface. In addition, Simola, Malkavaara,
Alen, and Peltonen (2000) and Simola-Gustafsson, Hortling, and
Peltonen (2001) mentioned that on the basis of previous guid-
ance from the pulp and paper industry it can be hypothesized
that these droplets originate from lignin present in the untreated
biomass.
Generally, pretreatment had an effect on the overall structure
of the pretreated material apart from a change in color; the fibers
were partially defibrated in the alkaline pulping (Fig. 1C), pre-
sumably due to the hemicellulose content of the middle lamella
(Donaldson, Hague, & Snell, 2001), and well separated in the steam
explosion (Fig. 1A). However, upon closer observation, the surface
of the individual fibers had changed drastically. The uneven surface
now appeared smooth after alkaline pulping, while it showed holes
and cracks upon steam explosion.
A common feature of the enzymatic hydrolysis step is the need
for pretreatment of the lignocellulosic material resulting in a more
efficient reaction despite the recalcitrant nature of the plant cell
wall (Himmel et al., 2007). The effect of the pretreatment has been
described as a disruption of the cell wall matrix including the
connection between carbohydrates and lignin, as well as depoly-
merizing and solubilising hemicellulose polymers (Ramos, 2003).
Pretreatment is also able to change the degree of cellulose crys-
tallinity (Chang & Holtzapple, 2000).
Steam explosion was selected as the processing technology
because it requires little or no chemical input and thus is envi-
ronmentally benign relative to other technologies. Both the steam
exploded and the pulped rice straw materials were coarse, pow-
dery, and had dark-brown color. These color changes during the
steam and pulping pretreatment could have resulted from the
degradation of the cell wall components and extractives (Sun, Xu,
Sun, Fowler, & Baird, 2005).
The chemical characterization revealed the proportion of each
component of the fibers from agricultural residues. As seen in
Table 1, the alkaline pulping treatment results in a higher holo-
cellulose and lower lignin compared to the raw material, while for
steam explosion the holocellulose is lower compared to the raw
material and this can be explained due to the higher lignin content
after steam explosion which may be due to the re-condensation
of the lignin during the steam explosion process. On the other
hand, the comparison between the two pretreatment methods
indicated that the main effect of the hydrothermal pretreatment
on the composition of the biomass is the partial but substantial
removal of hemicelluloses, where the results seen in Table 1 indi-
cated that the holocellulose for the unbleached alkaline pulping
3.2. FTIR spectroscopic analysis
FTIR spectroscopy was used as an analytical tool to qualitatively
determine the chemical changes in the surface of pretreated agri-

M.M. Ibrahim et al. / Carbohydrate Polymers 83 (2011) 720–726
723
Fig. 1. Microscopic images. SEM images of steam exploded (A and B) and alkaline pulped rice straw (C and D). Steam explosion causes fiber separation (A) and a surface layer
with holes (B). Holes are indicated with arrows. In alkaline pulping pretreated rice straw, the defibrating effect of the pretreatment causes the individual fibers to partially
separate, as can be seen in (C). The pretreatment leaves a surface layer of droplets on the individual fibers (D). Droplets are indicated with arrows.
ture (Ibrahim, 2000), the peaks at 1170–1040 cm−1 arise from the
ether linkage. The absorbances at 1431, 1372, 1319, 1165, 1059 and
896 cm−1 which are typical of pure cellulose can be seen in the FTIR
spectra.
cultural residue to complement and understand the microscopic
investigations. The FTIR spectra of delignified alkaline pulping
pretreated and steam-exploded rice straw samples are shown in
Fig. 2A. Excerpts of the spectra are presented in Fig. 2B.
The infrared spectra in the 400–4000 cm−1 region for steam-
exploded and alkaline pulped rice straw are shown in Fig. 2A.
Kristensen, Thygesen, Felby, Jørgensen, and Elder (2008) have men-
tioned that one of the effects of the pretreatment is the removal of
wax from the straw, where the CH2– stretching bands at approx-
imately 2850 and 2920 cm−1 reduced for the pretreated sample,
signifying a reduction in the amount of the aliphatic fractions of
waxes. The same observation can be noticed for the raw material of
rice straw, Fig. 2A, where a reduction in the CH2 band can be inves-
tigated after alkaline pulping and steam explosion. On the other
hand, absorption bands for different pretreatments appeared sim-
Two interesting features are revealed in Fig. 2B. It can be seen
that the carbonyl band at 1735 cm−1, which has been ascribed to
hemicelluloses and celluloses, is changed according to the treat-
ment used. This is expected, as the pretreatment is known to affect
the hemicelluloses and the celluloses in this region. Second, lignin
bands at approximately 1595 and, in particular, 1510 cm−1 (aro-
matic ring stretch) (Stewart, Wilson, Hendra, & Morrison, 1995)
are strongly enhanced in the alkaline pulping pretreated samples
compared with steam-exploded samples, where these peaks are
reduced (Fig. 2B). One explanation for this could be a relative
increase in the amount of lignin in the steam explosion pretreat-
ment due to the removal of hemicelluloses. Another reason could be
that lignin is released and re-deposited on the surface. The increase
in lignin is believed to be too significant to be only due to the
hemicellulose removal.
One of the strategies employed in increasing convertibility is
to change the cellulose crystallinity. Differences between samples
with regard to the relative amounts of amorphous and crystalline
cellulose have earlier been described through infrared peak ratios.
At least four different peak pairs have been proposed (Hulleman
et al., 1994; Wistara et al., 1999). Of these, only the peak pair
1429 cm−1 (crystalline) and 893 cm−1 (amorphous) is seen for
the samples of the present study. The peak ratios for the steam
ilarly. The absorption band at the region of 3422 and 3345 cm−1
,
may be due to various hydroxyl (OH) stretching vibrations. The
OH compounds may include absorbed water, aliphatic primary and
secondary alcohols found in cellulose, hemicellulose, lignin, extrac-
tives; and carboxylic acids in extractives (Khan, Idriss Ali, & Basu,
1993; Kolboe & Ellefsen, 1962). The absorption band near the OH
stretching vibrations, at 2918 and 2904 cm−1 may be associated
with CH stretching vibrations. Kolboe and Ellefsen (1962) sug-
gested that the bands in the 1644 cm−1 region for cellulose may be
attributed to C O stretching vibration of the alpha-keto carbonyl.
The presence of the bands at the region of 1323 and 1318 cm−1 may
be attributed to the lignin (syringic group). According to the litera-

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M.M. Ibrahim et al. / Carbohydrate Polymers 83 (2011) 720–726
Fig. 2. FTIR spectra for steam exploded and alkaline pulping pretreatment rice straw (the arrows in (A) indicate the peaks at 1429 and 893 cm−1 for the pretreated fibers and
at 2850 cm−1 for the raw material).
exploded and pulped raw materials were gathered in Table 1. The
peak ratio for the alkaline pulped rice straw was 2.0, while it was
2.33 for the steam-exploded one. When comparing the results, it
appears that the crystallinity index showed higher value for the
steam-exploded fibers compared to the alkaline pulped fibers. This
means that, the steam explosion raises the crystallinity of the cel-
lulose component because the main reactions that occur during
the process are the cleavage of the amorphous fractions of the cel-
lulose, while the crystalline fraction is more resistant to cleavage
under these conditions.
3.3. Glucose production
3.3.1. Effect of pretreatment on sugar content
The data presented in Table 2 show that the glucose concen-
tration resulting from the enzyme hydrolysis of the pretreated
rice straw depended on the pretreatment method. Glucose data in
Table 2 is graphically represented in Fig. 3. The graph clearly shows
that the glucose concentration was higher for alkaline pulped rice
straw compared to steam-exploded material. On the other hand,
the further treatment for the steam-exploded fibers with alkali

M.M. Ibrahim et al. / Carbohydrate Polymers 83 (2011) 720–726
725
Table 2
Glucose and theoretical ethanol concentrations (g/L) for the pretreated samples.
Table 3
Rate of enzyme hydrolysis (mg/mL glucose per hour) for pretreated materials.
Samples
Glucose
concentration (g/L)
Theoretical ethanol
concentration (g/L)
Samples
Rate of enzyme hydrolysis
2nd day
3rd day
4th day
Pure cellulose
45.8
47.8
28.9
58.4
23.41
14.77
24.43
29.84
Pure cellulose
0.575
0.592
0.491
0.589
0.581
0.649
0.387
0.753
0.477
0.498
0.301
0.608
Alkaline pulped rice straw
Steam-exploded rice straw
Alkali-extracted-steam-
exploded rice
Alkaline pulped rice straw
Steam-exploded rice straw
Alkali-extracted-steam-exploded rice straw
straw
reaction is as follows:
extraction resulted in higher glucose concentration after enzyme
hydrolysis for four days compared to both steam explosion and
alkaline pulped pretreatments.
C6H12O6
−→
Fermentation2C2H5OH +
2CO2
carbon dioxide
glucose
ethanol
46 g/mol
44 g/mol
180 g/mol
From the above observations the product of steam explosion
pretreatment has been shown to be easily digestible by enzymes
after alkali extraction. This is may be due to the fact that lignin
is re-localized on the fiber by steam explosion, resulting in higher
lignin content, as can be seen from Table 1, and by further alkali
treatment it can be removed. It is known that lignin is responsible
for unproductive adsorption of cellulases (Eriksson, Börjesson, &
Tjerneld, 2002; Kristensen, Börjesson, Bruun, Tjerneld, & Jørgensen,
2007), where it encases the cellulose in the cell wall matrix, hin-
dering cellulases from reaching cellulose fibrils. We hypothesize
that the migration of lignin to the outer surface exposes internal
cellulose surfaces. Selig et al. (2007) have also observed the for-
mation and migration of spherical lignin deposits onto the surface
of fibers as a result of pretreatment. They also suggest that the
deposited lignin can have a negative impact on the enzymatic cellu-
lose hydrolysis. It is possible, however, that the surface lignin layer
is easily removed by mild conditions of alkali extraction for the
steam-exploded fibers, due to lignin being less strongly bound to
carbohydrate polymers, compared with its native linkages. More-
over, we theorize that the re-located lignin has exposed cellulose
inside the cell wall, thus increasing the enzyme accessibility.
Based on these observations, we therefore propose that the re-
localization of lignin as well as partial hemicellulose removal are
likely to be important factors in increasing the enzymatic digestibil-
ity through pretreatment. It seems that exposing cellulose through
manipulation of hemicelluloses and lignin are equally as important
as altering the crystallinity and rupture of the skeletal structure of
the cell wall.
Based on the balanced equation, the alkali-extracted steam-
exploded rice straw should produce 29.84 g/L ethanol compared
to 23.41 g/L for pure cellulose, 24.43 g/L for steam-exploded rice
straw and 14.77 g/L for alkaline pulped rice straw (Table 2). From
the theoretical ethanol concentration, one can notice that the pro-
duction of ethanol depends on the pretreatment method according
to the glucose formation. From the data given here, it is evident that
steam explosion treatment, after alkali extraction, can improve the
potential for pretreated rice straw to glucose conversion.
Furthermore, the calculated rates of enzyme hydrolysis are
gathered together in Table 3, where one can notice that the higher
rate of enzyme hydrolysis was obtained for the alkali-extracted
steam-exploded rice straw. The rate of the enzyme hydrolysis
was rapid for the first days, where it depends on the action of
the enzyme on converting the cellulosic products into glucose.
It is known that, enzymatic degradation of cellulose to glucose
is generally accomplished by synergistic action of at least three
major classes of enzymes: endoglucanases, exoglucanases, and
␤-glucosidases. These enzymes are usually called together cellu-
lase or cellulolytic enzymes (Wyman, 1996, chap. 1). According
to that the enzyme plays an action where the endoglucanases
attack the low-crystallinity regions of the cellulose fiber and cre-
ate free chain-ends. The exoglucanases further degrade the sugar
chain by removing cellobiose units (dimers of glucose) from the
free chain-ends. The produced cellobiose is then cleaved to glu-
cose by ␤-glucosidase. This action is very important to complete
depolymerization of cellulose to glucose.
From the glucose concentration, one can calculate the theoreti-
cal yield for ethanol production. For the theoretical calculation, it is
known that each mole of glucose will produce two moles of ethanol.
Glucose has a molar mass of 180 g/mol. The molar mass of ethanol
is 46 g/mol. The other product of fermentation is carbon dioxide,
which is not considered in the calculations. The equation for this
4. Conclusions
The results indicated that the higher glucose concentration
can be obtained from the alkali-extracted steam-exploded rice
straw. This was noticed compared to the results obtained from
alkaline pulping and steam explosion of the waste material as a
pretreatment process. This can be attributed to the fact that the
steam explosion affects the cellulosic region and the further alkali
extraction facilitates removing of the re-localized lignin and hence
facilitates the enzyme treatment. Furthermore, the results indi-
cated that the rate of hydrolysis differs according to the different
pretreatments applied to the raw material. Also, the results indicate
that via enzymatic hydrolysis, rice straw can be used as a cellu-
losic source after pretreatment for glucose production, where it can
produces glucose higher than that resulted from pure cellulose.
Acknowledgments
The authors acknowledge the USA-Egypt Science and Tech-
nology Program, USDA, and NSF for funding the project
(ENV8-004-002). Also, the authors acknowledge Bob Wright for
conducting the steam explosion experiments.
Fig. 3. Glucose concentration upon enzyme hydrolysis of pure cellulose and pre-
treated rice straw.

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M.M. Ibrahim et al. / Carbohydrate Polymers 83 (2011) 720–726
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