Research Article

Received: 17 July 2014 Revised: 23 August 2014 Accepted article published: 29 August 2014 Published online in Wiley Online Library:

(wileyonlinelibrary.com) DOI 10.1002/jctb.4530

Enhanced saccharification of lignocellulosic

biomass by pretreatment with quaternary
ammonium hydroxide
Chao Zhong,a Chunming Wang,a Fengxue Wang,a Honghua Jia,a* Ping Weia
and Yin Zhaob

Abstract
BACKGROUND: As one of the most abundant natural resources, lignocellulosic biomass is usually utilized at low eciencies
because of its complicated structure and poor degradability, and pretreatments prior to utilization are therefore considered
necessary. To overcome the disadvantages of traditional pretreatments for lignocellulosic biomass, a method based on the
application of quaternary ammonium hydroxide was investigated.

RESULTS: Remarkable structural transformation and compositional changes were detected in pretreated biomass, presenting
significant removal of lignin and decreased cellulose crystallinity, which could contribute to enhanced degradability. Significant
factors in this process were investigated by single-factor experiment and response surface methodology, and optimum
conditions were calculated as: reaction time 7.0 h, temperature 51.6 C, and concentration of solvent 16.8%. Biomass pretreated
under these conditions presented a maximum saccharification yield of 73.71%, which was a 3-fold increase compared with the
untreated sample. In addition, the solvent quaternary ammonium hydroxide was recycled five times for biomass pretreatment
with high activity retained.

CONCLUSIONS: Enhanced saccharification eciency of lignocellulosic biomass was achieved by pretreatment with quaternary
ammonium hydroxide.
2014 Society of Chemical Industry

Keywords: lignocellulosic biomass; pretreatment; saccharification; quaternary ammonium hydroxide; recycling

INTRODUCTION and hemicelluloses.8 Hence, in order to eectively utilize these

As a major component of biomass, lignocelluloses make up about
half the materials produced via photosynthesis.1 It is known
that lignocelluloses consist primarily of biopolymers stored in
the form of cellulose, hemicelluloses and lignin, which strongly
interact and are chemically connected by non-covalent forces
and covalent cross-linkages, resulting in low eciency for ligno-
cellulosic biomass utilization in view of its poor degradability.1,2
Currently, only a minority of lignocellulosic biomass has been
utilized for such fields as the production of animal feeds and
organic fertilizers, paper-making, and energy utilization,3,4 with
the rest being directly burnt every year, which is a substan-
tial waste of natural resources and also a contamination of the
environment.
With environmental problems being more exacerbated, recently
there has been an increasing interest in more ecient utiliza-
tion of agro-industrial residues for the production of renewable
energy.5,6 It is believed that most renewable energy is origi-
nated from carbohydrates stored in agro-industrial residues.
However, conversion from residues to carbohydrates is always
hindered by their complicated structures.7 Lignocellulosic
materials are generally resistant to be degraded, which is
attributed to the tight association between lignin, cellulose,
materials, pretreatment before utilization are considered to be
necessary.
Currently, extensive eorts have been made to develop methods
for lignocellulosic biomass pretreatment, such as biological, physi-
cal, physicochemical and chemical methods, and among which the
method based in the use of chemicals has drawn increasing atten-
tion in view of its low energy consumption and high eciency.9 11
So far, reported biomass pretreatments based on chemicals have
mostly been carried out with inorganic acid or base.12,13 However,
these methods involved great consumption of solvent and strict
operating conditions may dramatically increase the pretreatment
cost. In addition, application of inorganic acid/base is strictly con-
trolled because of serious pollution risk, which to some extent
Correspondence to: Honghua Jia, College of Biotechnology and Pharmaceuti-
cal Engineering, Nanjing Tech University, Nanjing 211800, P. R. China. E-mail:
hhjia@njtech.edu.cn
a College of Biotechnology and Pharmaceutical Engineering, Nanjing Tech Uni-
versity, Nanjing 211800, P. R. China
b Henan Tianguan Group Co. Ltd, Nanyang 473000, P.R. China

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 www.soci.org
prevents it being an ideal method for lignocellulosic biomass
pretreatment.
In comparison, recent utilization of polar solvents for lignocellu-
losic biomass pretreatment has been considered as an economic
promising approach, and it is found that these reports are mostly
based on ionic liquids (ILs),14 such as N-methylmorpholine N-oxide
monohydrate (NMMO), LiCl/N,N-dimethylacetamide (DMAC) and
1-butyl-3-methylimidazolium chloride (BMIMCl). Compared with
the traditional process, this method based on the use of ionic
liquids is considered to be a more ecient lignocelluloses pre-
treatment. There are still some limitations with this process, such
as high viscosity and high cost, which inhibit its application to
biomass pretreatment at large scale. Besides, degradation of cellu-
lose or hemicelluloses is also detected during the pretreatment,15
which is not beneficial for further application of pretreated
biomass.
To overcome these disadvantages, quaternary ammonium
hydroxide containing polar hydroxide anion is considered as an
ideal substitute solvent.16 18 Compared with sodium hydroxide,
which requires strict operational conditions and causes serious
pollution,19 tetra-n-methylammonium hydroxide (TMAH) with
its advantages of mild operation conditions, high eciency, and
good recyclability was considered to be appropriate for biomass
pretreatment. The feasibility of biomass pretreatment with TMAH,
and its recyclability were investigated based on serial biological
analyses. In addition, eects of significant factors on biomass
pretreatment were investigated by single-factor experiment
and response surface methodology. All research work in this
paper was done to investigate the feasibility of TMAH-based
biomass pretreatment for enhanced biomass utilization
eciency.
EXPERIMENTAL
Materials
Lignocellulosic biomass (wheat straw) is annually harvested from
Siyang, Jiangsu Province in Southeast China, and then stored
in piles on the ground near the biogas pilot plant in Nanjing
Tech University. Straw samples are dried in sunlight and cut
into appropriate dimension of 5 cm. Tetra-n-methylammonium
hydroxide (40% aqueous solution), acetic anhydride and sodium
hydroxide were purchased from Alfa Company. Cellulase (cel-
lulase from Trichoderma viride), hemicellulase (hemicellulase
from Aspergillus niger), and alkaline lignin were purchased from
Sigma-Aldrich Company. Sulfuric acid, ethanol, were provided by
Aladdin Reagent Company.
Pretreatment of wheat straw with tetra-n-
methylammonium hydroxide
Investigations of wheat straw pretreatment with tetra-n-
methylammonium hydroxide were performed as follows:
Tetra-n-methylammonium hydroxide aqueous solution (40%)
was initially diluted to lower concentration (1030%) as the sol-
vent for pretreatment. Wheat straw sample (1.00 g) was added
and mixed with 10 mL solvent in a triangular flask incubated in
a water bath at 50 C. After the required time, the system was
cooled down by incubating the flask in an ice-water bath, and
10 mL of distilled water was then added with stirring to stop
the reaction. The mixtures were then filtered through gauze,
with the pretreated samples being washed and dried in a vac-
uum oven for further analyses. Filtrates were collected for lignin
recovery.
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C Zhong et al.
Characterizations of pretreated wheat straw
Series of tests for untreated and pretreated straw samples includ-
ing compositional and structural analyses were performed.
Compositions of straw (cellulose and hemicelluloses) were
analyzed according to the method described by Goering and
Van Soest,20 and lignin content was determined by the Klason
method.21
The crystal structure was detected by X-ray diraction (XRD),
recorded on XTRA/3KW X-ray diractometer (ARL). It was per-
formed with CuK radiation ( = 0.1541 nm) and the diraction
scans were recorded with 2 ranging from 3 to 50 . The Crystallo-
graphic Index (CI) was calculated according to the Segal Equation
as follows: 22
I I
CI = hkl am 100%
Ihkl
where Ihkl was the diraction intensity of crystallization area
diraction peak; Iam was the diraction intensity of non-
crystallization area diraction peak (2 = 18 ).
In addition, scanning electronic microscopy (SEM) was per-
formed to investigate outer structural changes in straw samples
during pretreatment.
Design of experiment for wheat straw pretreatment
According to the previous result, it was found that several fac-
tors might have great influence on pretreatment, among which
reaction temperature, reaction time, and concentration of solvent
were assumed to be the three primary factors. Hence, a single fac-
tor experiment was conducted to investigate each factors influ-
ence on the process.
The investigations were carried out by initially measuring 1.00 g
wheat straw sample and then mixing with 10 mL of solvent diluted
to concentrations 5%, 10%, 15%, 20%, 25%, and 30%, with the
mixtures being heated to temperatures of 30, 40, 50, 60, and 70 C
for 2, 4, 6, 8, 10, and 12 h, respectively. After the required times,
the system was incubated in an ice-water bath to cool it down,
and 10 mL of distilled water was added with stirring to stop the
reaction. The mixtures were then filtrated, with the pretreated
samples being washed and dried in vacuum oven for further
analysis.
In this study, enzymatic saccharification yield was proposed for
evaluating the biodegradability of biomass samples. Untreated
or pretreated dried samples (0.5 g) were employed per sacchari-
fication experiment, and each test was performed in duplicate to
ensure the accuracy of results. Enzymes used for the saccharifica-
tion were cellulase (cellulase from Trichoderma viride) and hemi-
cellulase (hemicellulase from Aspergillus niger), which were both
provided by Sigma and employed according to instruction as: cel-
lulase (12 mg) and hemicellulase (270 mg) dissolved in 20 mL of
phosphate buer solution (pH 4.8) designed for 1 g biomass sac-
charification test, and the systems were fixed in static state at 50 C
for 48 h, with the pulps being filtrated. Corresponding sugars in
the filtrate were quantitatively analyzed by gas chromatography
as follows: filtrate was initially made neutral. Alditol acetates of
neutral sugars in filtrates were prepared according to the method
described by Blakeney.23 The alditol acetate derivatives of sugars
were quantified by HP 5890 GC using an Alltech DB-225 capillary
column (30 m 0.25 mm 0.25 m film) and the temperature pro-
gram initial temperature 190 C with a hold of 4 min, followed by a
temperature rise of 2 C min1 to 230 C with a final hold of 25 min.
Helium was used as the carrier gas, and the signals were detected
by flame ionization detector (FID). Saccharification yield was
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precipitate washed with a specified amount of distilled water and

Table 1. Design of experiments and response for saccharification
yield of straw sample dried under vacuum at 50 C. The precipitate yield was calculated
based on the lignin content of untreated straw sample as:
Factor 1 Factor 2 Factor 3 Response:
MRe
X3 : Lignin Yield = 100%
MK
X1 : X2 : Reaction Concentration Saccharification
Reaction temperature of solvent yield where MRe was the mass of precipitate, and MK the lignin content
Index time (h) ( C) (%) of straw (%) a of untreated straw sample determined by the Klason method.
The precipitate was characterized by elemental analysis
1 8 40 5 45.23
and Fourier transform infrared (FTIR) spectroscopy, which was
2 4 50 25 59.26
recorded in the range from 4000 to 400 cm1 with a resolution of
3 4 50 15 72.65
4 cm1 .
4 12 60 15 50.23
5 12 50 25 54.24
6 8 50 15 71.98 Recycling of TMAH
7 4 60 15 63.24 After pretreatment, the filtrate was initially processed with isola-
8 8 50 15 72.46 tion of lignin and some cellulosic monomers, with the residual
9 8 50 15 72.78 TMAH in the filtrate recovered by simply reducing the pressure
10 8 60 25 64.23 and subsequently distilling to remove water under the conditions
11 4 40 15 60.67 0.1 MPa and 45 C.14 The recycled TMAH was diluted and applied
12 12 50 5 52.23 as solvent for wheat straw pretreatment again. Repeated cycles
13 12 40 15 46.23 of pretreatment followed by sample isolation and TMAH purifi-
14 8 50 15 72.11 cation were performed to investigate the recyclability of TMAH
15 4 50 5 57.26 for biomass pretreatment. All cyclic experiments were carried out
16 8 40 25 59.23 under the optimal conditions determined above.
17 8 60 5 58.03
a Saccharification time: 48 h. RESULTS AND DISCUSSION
Pretreatment of lignocellulosic biomass
In this study, scheme for wheat straw pretreatment with
calculated based on the content of glucose and hemicelluloses in
tetra-n-methylammonium hydroxide was designed and is pre-
an untreated straw sample.
sented in Fig. 1. It could be found that wheat straw samples with
stable lignocellulosic structure could be readily disrupted within
Optimization of pretreatment by response surface several minutes, and the possible mechanism was speculated
methodology and described as: ion pairs in solvent were initially dissociated
Response surface methodology is an empirical modeling into [(n-t-Me)4 -N]+ cation and hydroxide anion, and the free
technique used to evaluate the relationship between a set of cations and anions might immediately interact with lignocellu-
experimental factors and results.24 In order to optimize the factors loses, resulting in the rupture of tight bonding existed between
determined for pretreatment, a Box-Behnken designed experi- cellulose, hemicelluloses and lignin.25
ment with the three variables reaction time (X1 ), temperature (X2 ), According to previous results, solvents with concentration 40%
and concentration of solvent (X3 ) at three levels was performed or higher might be suitable for lignocelluloses dissolution, leading
(Table 1). to the distribution of all components in solvent. In this study, a
A two-way interaction quadratic model equation was employed diluted solvent with lower concentration was considered to be
to predict the optimal parameters as follows: suitable for lignocellulosic biomass pretreatment. It was found that
the inner structure of lignocelluloses was dramatically disrupted
y = k0 + k1 X1 + k2 X2 + k3 X3 + k11 X12 + k22 X22 + k33 X32 + k12 X1 X2 after pretreatment, which could improve cellulose accessibility
+ k13 X1 X3 + k23 X2 X3 in lignocelluloses and lead to enhanced degradability of straw
samples.
where y was the response; k0 a constant; k1 , k2 and k3 the linear
coecients; k11 , k22 and k33 the quadratic coecients; k12 , k13 and Eects of pretreatment on lignocellulosic biomass
k23 the cross-coecients. In order to investigate the eects of pretreatment on straw sam-
A total of 17 experiments were carried out to estimate the coe- ples, serial analyses including structural/compositional detections
cients of the equation, with experimental data being subsequently and saccharification experiments were performed.
analyzed using Design-Expert 8.0 to determine the optimal value Structural analyses of straw samples including scanning elec-
of each factor. All tests were performed in duplicate to guaran- tronic microscopy (SEM) and X-ray diraction (XRD) were per-
tee the accuracy of results. Finally, verifying experiment conducted formed. According to SEM analyses (Fig. 2), obvious structural
under the optimal conditions was repeated several times to con- destruction was detected after pretreatment, achieving a looser
firm the results. structure and enhanced cellulose accessibility in pretreated sam-
ple, which was beneficial for further enzymatic degradability.
Lignin recovery Crystallinity of lignocellulosic biomass represents the relative
The filtrates obtained from pretreatment were fixed in vacuum at amount of crystalline cellulose in material. X-ray diraction anal-
4 C for 60 min, with the suspension being centrifuged and the yses of untreated and pretreated straw samples were performed,

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 www.soci.org C Zhong et al.

Figure 1. Scheme of lignocellulosic biomass pretreatment with TMAH.

Figure 2. Scanning electronic microscopy (SEM) of straw samples before and after pretreatment: (a, b) untreated straw sample; (c, d) pretreated straw
sample.

with samples exhibiting similar diraction peaks to cellulose II
(2 = 16.0 and 22.5 ).26 Results indicated that the untreated sam-
ple showed a lower CI value (46.43 0.56%) compared with the
pretreated samples (48.76 0.44%). It is believed that an increased
CI value is normal when biomass is subjected to pretreatment,
which is probably explained by the removal of amorphous frac-
tions (lignin and hemicelluloses). A reduction of cellulose crys-
tallinity was also detected in the pretreated sample, indicating that
conversion to an amorphous form in the crystalline structure of cel-
lulose was achieved, which contributed to enhanced degradability
of straw samples.5
Compositional analyses of straw sample are shown in Fig. 3,
and it is interesting to find that the compositional variation in
straw samples pretreated under dierent conditions all presented
the similar trend of distinct removal of lignin and slight loss
of cellulose or hemicelluloses. This result might be attributed
to the dissolution and isolation of alkali-soluble lignin from
lignocelluloses during the pretreatment process. It is known
that lignin is a major obstacle to cellulase, because it prevents
enzyme accessibility and also irreversibly adsorbs cellulase.27 Our
results confirmed the existence of strong correlation between
the fraction of hydrolyzed cellulose and lignin removed from
straw samples (Fig. 4).28 A promising result was concluded as it
was not necessary to remove the majority of lignin to achieve
greater cellulose degradability, which meant that a degree of
lignin removal (6070%) might be enough for greater cellulose
degradability.
After pretreatment, disrupted structures accompanied with
lignin removal and lower degree of crystallinity were achieved
in lignocelluloses, which improved the accessibility of cellulose
and contributed to enhanced degradability. In this study, enzy-
matic saccharification experiments were performed to evaluate
the eects of pretreatment on biomass, and result show that
compared with the untreated sample, a 3-fold increase of sac-
charification yield to about 73% was achieved for the pretreated
samples (Fig. 5).

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Enhanced saccharification of lignocellulosic biomass
Figure 3. (a) Eect of reaction time on straw composition (temperature 50 C, concentration 15%); (b) eect of reaction temperature on straw composition
(time 8 h, concentration 15%); (c) eect of solvent concentration on straw composition (time 8 h, temperature 50 C).
Figure 4. Relationship between lignin content removed from straw sam-
ples and biodegradability of pretreated samples.
It is also known that traditional chemicals for biomass pretreat-
ment such as sodium hydroxide, sulfuric acid, etc. always require
higher temperature (>100 C) and more time, and sometimes even
generate serious pollution in the environment.10 In comparison,
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pretreatment based on TMAH was completely achieved and pre-
sented higher eciency with less consumption at lower tem-
perature (around 50 C) and in less time (about 7 h) (Fig. 5),
which showed its advantages over traditional methods and fur-
ther proved it to be a potential pretreatment for lignocellulosic
biomass.
Factors aecting pretreatment process
In present study, pretreatment of lignocellulosic biomass for
dierent reaction times was investigated as seen in Fig. 5(a). Pre-
treatment of straw for 4 h resulted in a saccharification yield of
64.30%. A slight increase of yield less than 10% was obtained with
further extending reaction times to 8 h, followed by a decreased
yield with longer pretreatment times.
At room temperature, pretreatment could not be accomplished
because of the weak activity at low temperature. As temperature
was increased, pretreatment could be readily completed. As seen
in Fig. 5(b), samples pretreated at temperatures from 3050 C
presented obvious increase of saccharification yield from 43.12%
to 72.68%, followed by a decreased yield at higher tempera-
ture. This result might be explained by the enhanced activity
and improved cellulose/hemicelluloses content in pretreated
samples. With increasing temperature, solvent with enhanced
activity might rapidly interact with lignocelluloses and chemically
disrupt its inner interactions, leading to a looser structure and
enhanced degradability of pretreated samples. However, a slight
loss of hemicelluloses and cellulose was detected in samples
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Figure 5. (a) Eect of reaction time on saccharification yield of pretreated sample (temperature 50 C, concentration 15%); (b) eect of reaction
temperature on saccharification yield of pretreated sample (time 8 h, concentration 15%); (c) eect of solvent concentration on saccharification yield
of pretreated sample (time 8 h, temperature 50 C).

Table 2. ANOVA for response surface quadratic model

Source Sum of Squares Degree of Freedom Mean Square F-value P-value Prob > F

Model 1335.97 9 148.44 11.74 0.0019 significant

X1 -Time 175.78 1 175.78 13.90 0.0074a
X2 -Temperature 74.24 1 74.24 5.87 0.0459a
X3 -Concentration 73.27 1 73.27 5.79 0.0470a
X1 X2 0.51 1 0.51 0.04 0.8464
X1 X3 2.5 105 1 2.5 105 1.9 106 0.9989
X2 X 3 15.21 1 15.21 1.20 0.3090
X1 2 358.09 1 358.09 28.32 0.0011a
X2 2 289.33 1 289.33 22.88 0.0020a
X3 2 245.41 1 245.41 19.41 0.0031a
Residual 88.51 7 12.64
Lack of Fit 86.37 3 28.79 53.94 0.0011 significant
Pure Error 2.13 4 0.53
Cor Total 1424.48 16
a Values of Prob > F less than 0.0500 indicate model terms are significant.
Std.dev 3.56; R2 : 0.9879.

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Figure 6. (a) Response surface plot for interaction eect of time and temperature on biomass saccharification yield; (b) response surface plot for interaction
eect of time and solvent concentration on biomass saccharification yield; (c) response surface plot for interaction eect of temperature and solvent
concentration on biomass saccharification yield.

The eect of solvent concentration on saccharification yield is

shown in Fig. 5(c), indicating that saccharification yield increase
from 56.21% to 72.68% was achieved with a concentration
increase from 5% to 15%, which was explained by the enhanced

Table 3. Results of enzymatic tests with recycling of TMAH

Saccharification
Cycle Volume of Loss of yield of
indexa solvent (mL) TMAH b(%) straw (%) c

1 10 72.56
2 10 2.3 67.56
3 10 3.4 62.67
4 10 2.1 57.23
5 10 3.7 51.34
Figure 7. FTIR spectra of reference lignin and recovered lignin. a Samples for cyclic experiment were pretreated under the optimal
conditions obtaiend by RSM .
b Mass of TMAH would slightly decrease during the reuse experiments
pretreated at higher temperature, resulting in a decreased saccha-
due to operational loss.
rification yield. Therefore, pretreatment of straw samples within a c Saccharification time: 48 h.
prescribed temperature range was considered to be necessary.

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alkalinity with increasing amounts of hydroxide anions. A gentle
decrease in yield was detected with further increase of con-
centration of solvent, which might be caused by the loss of
cellulose/hemicelluloses in samples pretreated by solvents with
higher concentration.
Optimization of pretreatment process
In recent publications, response surface methodology had been
successfully applied to optimize factors that might have a strong
influence on experimental results.27 For this purpose, according
to the results of single-factor experiments, a Box-Behnken design
was employed, with levels of three determined variables: reaction
time (X1 ); temperature (X2 ); and concentration of solvent (X3 ), and
their corresponding experimental designs being shown in Table 1.
The experimental results were analyzed to establish an empirical
model for optimizing the response, based on which a quadratic
model in terms of the coded factors for saccharification yield
was obtained to reveal the mathematical relationship between
the independent variables and the response, with the polynomial
model being calculated by Design-Expert 8.0 as follows:
y = 72.60 4.69X1 + 3.05X2 + 3.03X3 + 0.36X1 X2
0.0025X1 X3 1.95X2 X3 9.22X12 8.29X22 7.63X32
Results for analysis of variance (ANOVA) are listed in Table 2, and
it is found that the predicted response fitted the experimentally
obtained response well. The coecient of determination (R2 ) for
checking the goodness-of-fit of the model was 0.9897, implying
that 98.97% of the variability in response could be explained by
this model. Besides, the F-value was much greater than the tabular
F 0.01 (9, 7) value of 6.72, suggesting that the treatment dierences
were highly significant.29 Significance of coecient was supposed
to be directly proportional to P value, and the three selected
variables in this study were all analyzed with P values less than 0.05,
indicating that a highly statistically significant confidence level of
about 95% was obtained.
In addition, the interaction eects of reaction time, tempera-
ture and concentration of solvent on biomass pretreatment were
investigated by response surface methodology, with shape of con-
tour curves implying that these variables were identified to have
greater eects on this process (Fig. 6).
All experimental data were analyzed for determining the opti-
mized value for each factor. As results, the optimal conditions
of reaction time, temperature and solvent concentration were,
respectively, calculated as 7.0 h, 51.6 C and 16.8%, with a maxi-
mum response (saccharification yield) as 73.71% obtained from
biomass pretreated under these conditions. To verify the results,
repeated tests were performed under these conditions, with
average saccharification yield of 73.12% obtained for pretreated
samples, which was consistent with the predicted value and
confirmed its significance.
Lignin recovery
We attempted to recover lignin from the filtrate, as this had been
successfully fulfilled in other studies.30 It was found that diluting
the filtrate with water could precipitate fine powders, and more
powder could be generated in weak acidic conditions. The powder
was characterized by elemental analysis and FTIR spectroscopy
detection (Fig. 7). Comparison with a reference lignin showed that
the precipitate was most likely to be lignin. In addition, when
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C Zhong et al.
ethanol was used for washing the pulp, increased precipitates
recovery was observed. Hence, the final protocol was an initial
washing of the pulp with ethanol, drying the combined filtrate by
evaporating the ethanol, and precipitating the lignin by diluting
the dried TMAH with water, with results showing that an average
yield of precipitate (lignin) as about 45% could be achieved.
Recycling of TMAH for pretreatment
According to the above method, residual TMAH could be simply
recovered and then applied for wheat straw pretreatment, with
serial tests being carried out to check its recyclability. Its note-
worthy that cycles 25 employed recycled TMAH as solvent for
straw pretreatment and exhibited saccharification yields ranging
from 67.56% to 51.34% (Table 3), which basically met the require-
ment for its utilization, indicating that no significant loss of activ-
ity occurred during the recycling process. It is apparent that this
advantage will promote the industrial application of TMAH for
biomass pretreatment in the future.
ACKNOWLEDGEMENT
The research was supported financially by the State Key Basic
Research and Development Plan of China (2009CB724700),
National Key Technology R&D Program (2011BAD15B02), National
Hi-Tech R&D Program (2012AA021405), Jiangsu Key Technol-
ogy R&D Program (BE2010359), PAPD, Jiangsu Province Natural
Science Foundation Innovation Project (SBK200910195), Open
Project of State Key Laboratory of Motor Vehicle Oriented Biofuels
Technology (2013024), and PCSIRT (IRT1066).
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