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Research Paper

Comparing the hydrolysis and biogas production

performance of alkali and acid pretreatments of
rice straw using two-stage anaerobic fermentation
Dianlong Wang a, Ping Ai a,*, Liang Yu b, Zhongxin Tan c, Yanlin Zhang a

College of Engineering, Huazhong Agricultural University, Wuhan 430070, China

Department of Biological Systems Engineering, Washington State University, Pullman, WA 99164, USA
College of Resources and Environment, Huazhong Agricultural University, Wuhan 430070, China

article info

To enhance rice straw degradation and improve biogas production in a two-stage anaer-

Article history:

obic system, rice straw was treated with either dilute alkali or acid. Firstly, the effects of

Received 23 August 2014

NaOH or HCl concentration, temperature and time on glucose conversion, degradation of

Received in revised form

cellulose and hemi-cellulose were investigated. It was found that glucose conversion rate

1 February 2015

and the organic matter content were greater with alkali rather than acid pretreatment.

Accepted 16 February 2015

Suitable conditions for hydrolysis using NaOH pretreatment were 2% (w/w), 60 h and 60  C.

Published online 4 March 2015

With these conditions the glucose conversion rate was 55.5%. Secondly, the hydrolysate
was used as substrate for batch anaerobic fermentation tests at different organic loadings.


The maximum methane yield of hydrolysate from NaOH pretreatment was 193.2 ml g1

Rice straw

[COD]. For the HCl pretreatment, the maximum methane production rate was 287.0 ml g1


[COD] which was 43.89% higher than with NaOH. Although the NaOH solution was more


efficient in improving rice straw hydrolysis, the hydrolysate from the HCl pretreatment had

Anaerobic fermentation

greater methane production at a similar organic loading.




Research into renewable energy has received more attention

in recent years to help solve energy supply and global warming problems (Kemppainen et al., 2012; Oleskowicz-Popiel
et al., 2012; Talebnia, Karakashev, & Angelidaki, 2010). Biogas
is one of the most popular forms of renewable energy able to
be produced from organic wastes. To ensure adequate sources
of feedstock for biogas production, lignocellulosic materials

2015 IAgrE. Published by Elsevier Ltd. All rights reserved.

such as manure fibre and crop residues have widely been

studied to enhance digestibility and improve the productivity
of anaerobic digestion (AD) (Hendriks & Zeeman, 2009; Yu
et al., 2014).
Rice straw is one of the most abundant, low-cost and
renewable lignocellulosic materials in the world. The lignocellulose in rice straw contains about 32e47% cellulose,
19e27% hemicellulose and 5e24% lignin (Zhao et al., 2010).
Cellulose is a crystalline material while hemicellulose is
amorphous (Guerra-Rodrguez, Portilla-Rivera, Jarqun-

* Corresponding author. Room B306, College of Engineering, Huazhong Agricultural University, No.1, Shizishan Street, Hongshan District,
Wuhan 430070, PR China. Tel./fax: 86 27 87288723.
E-mail address: (P. Ai).
1537-5110/ 2015 IAgrE. Published by Elsevier Ltd. All rights reserved.


b i o s y s t e m s e n g i n e e r i n g 1 3 2 ( 2 0 1 5 ) 4 7 e5 5

 zquez, 2012). Several researchers reEnrquez, Ramrez, & Va

ported that crystalline structure of cellulose limits the utilisation of lignocellulose (Sun et al., 2007). Therefore,
pretreatment of lignocellulose is an important step in
breaking down the chemical structure of the straw and
enhancing the conversion efficiency of cellulose and hemicellulose to increase sugars. Various pretreatment methods
have been investigated, such as using ammonia, alkali, acid,
and steam explosion (Bauer et al., 2014; McIntosh & Vancov,
2011; Nguyen et al., 2010; Satari Baboukani, Vossoughi, &
Alemzadeh, 2012; Zhao, Wang, Zhu, Ragauskas, & Deng,
2008). The hydrolysate obtained following the pretreatment
of lignocellulosic materials contains abundant reducing
sugars. These reducing sugars are easily degraded and utilised
in anaerobic fermentation.
Acid and alkali hydrolyses are commonly used in pretreatment methods that have been successfully used to treat
lignocellulosic materials (Hendriks & Zeeman, 2009;
Teramura et al., 2013). Acid pretreatment serves to break
down the hemicellulose and make the cellulose more
accessible. The xylose is mainly obtained when the hydrolysis of hemicellulose is performed under acid condition. The
cellulose in pretreated solid residue can easily be utilised to
produce ethanol. Also, many byproducts are generated, such
as furfural, hydroxymethylfurfural (HMF) and acetic acid
which have been reported as inhibitors for AD (Palmqvist &
gerdal, 2000). To avoid the formation of these inHahn-Ha
hibitors, moderate temperatures may be used in the acid
hydrolysis. Compared with that of acid pretreatment, the
reaction pathways of alkali pretreatments are more complex. The delignification reaction is very significant in alkaline hydrolysis, resulting in an increase of internal surface
area, a decrease in the degree of polymerisation, a decrease
in crystallinity, separation of structural linkages between
lignin and carbohydrates, and disruption of the lignin
structure (Sun & Cheng, 2002). This peeling reaction produces monomers of hemicellulose that are easily degraded
into other inhibitors at high temperature. Sodium hydroxide,
ammonia and lime pretreatments have received widespread
attention due to their high efficiency. However, many studies
treating straw at high temperatures and pressures has lead
to inhibition and high energy consumption (McIntosh &
Vancov, 2011; Satari Baboukani et al., 2012) moderate pretreatment temperatures still needs further research to
improve the economics of alkali pretreatment (Auxenfans
et al., 2012; Cabrera et al., 2014; Xu, Cheng, SharmaShivappa, & Burns, 2010). Furthermore, comparisons between pretreatments using alkaline and acid giving methane
yields and lignocellulose degradation on methane yield and
lignocellulose degradation are seldom reported (Zheng,
Zhao, Xu, & Li, 2014). The effect of hydrolysate concentration on methane productivity in rice straw has also not been
In this study, alkaline and acid concentration, moderate
temperatures and retention time were considered in the pretreatment processes of wheat straw, and their effects on the
lignocellulose removal and glucose yield were investigated.
The methane productivity from hydrolysate obtained in
alkaline pretreatment was compared with that of acid pretreatment. Moreover, the effect of hydrolysate concentration

on methane productivity was also tested in batch fermentation experiments.


Materials and methods


Preparation of rice straw

Rice straw was collected from fields in a suburb of Wuhan,

China. After collection, the straw was air dried, ground using a
grinder and passed through a 2 mm aperture standard screen.
The processed straw was then sealed in plastic bags and
stored at room temperature for further use and analysis. The
characteristics of the rice straw samples were shown in Table


Alkali pretreatment

Alkali treatment was carried out using a single factor experiment to study the effects of NaOH concentration (0.5%, 1%, 2%,
3%, 4%, w/w), moderate temperatures (15  C, 30  C, 45  C, 60  C,
75  C) and retention time (12 h, 24 h, 36 h, 48 h, 60 h) on the
degradation of rice straw. While studying the effects of NaOH
concentration and retention time, temperature was fixed at
45  C. While studying the effects of temperature and retention
time, NaOH concentration was fixed at 2%. The dry matter of
37.5 g was used in each batch, and fibre concentration was
fixed at 7% based on dry matter content. Treated mixture solutions were filtered with filter paper, the contents watered
using 1000 ml deionised water, and the wet solid residues
prepared analysis of dry matter, fibre composition and structure. A small amount of filtrate was taken for the analysis of
glucose and chemical oxygen demand (COD). The remaining
filtrates were stored at 4  C for further use. Each run was
duplicated and the average data was reported.


Acid pretreatment

Similar with alkali treatment, single factor experiment design

was used to study the effects of HCl concentration (0.5%, 1.0%,
1.5%, 2.0%, 2.5%, w/w), moderate temperatures (15  C, 30  C,
45  C, 60  C, 75  C) and retention time (12 h, 24 h, 36 h, 48 h,
60 h) on the degradation of rice straw. While studying the effects of HCl concentration and retention time, temperature
was fixed at 60  C. While studying the effects of temperature
and retention time, HCl concentration was fixed at 1%. The dry
matter of 37.5 g was used in each batch, and fibre concentration was fixed at 7% dry matter. Treated mixture solutions
were filtered with filter paper and the contents watered using

Table 1 e Characteristics of rice straw.



Total solids (TS, %)

Volatile solids (VS, %)
Neutral detergent fibre (NDF, %)
Acid detergent fibre (ADF, %)
Cellulose (%)
Hemicellulose (%)
Lignin (%)


b i o s y s t e m s e n g i n e e r i n g 1 3 2 ( 2 0 1 5 ) 4 7 e5 5

1000 ml deionised water, and the wet solid residues prepared

for the analysis of dry matter, fibre composition and structure.
A small amount of filtrate was taken for the analysis of
glucose and chemical oxygen demand. The remaining filtrates
were stored at 4  C for further use. Each run was duplicated
and the average data was reported.

Glucose conversion%


Hydrolysate from NaOH and HCl pretreatment was used as

substrate for batch anaerobic fermentation experiment. The
hydrolysates were neutralised to pH value of 7.0 using 20%
sulphuric acid or 20% sodium hydroxide prior to feeding the
reactors. The 1.5 l distillation flasks were used as batch
fermentation reactors. Each run included 0.8 l hydrolysate of
different concentrations (25%, 50%, 75% and 100% (v/v)) and
inoculated with sludge 0.4 l. All reactors were capped with
rubber stoppers and put into a water bath at 37  C. Prior to the
fermentation test, the reactors were flushed with nitrogen to
remove oxygen from the headspace and maintain an anaerobic environment. To minimise the effect of random errors,
each run was duplicated and the average data was reported.
The biogas volume was determined by drainage method,
and the biogas was collected by a 1 ml plastic syringe for the
gas component analysis. The biogas volume and component
were measured at 24 h interval and initial and final liquid
samples were taken.


The glucose conversion of rice straw after pretreatment

was used as an indicator of the extent of hydrolysis. The
glucose conversion was calculated based on following equation (MacLellan et al., 2013).

glucose concentration  volume of hydrolysate

total solids  cellulose content  1:11

Anaerobic fermentation

Analytical methods

Total solids (TS), volatile solids (VS) and COD were measured
using standard methods (APHA, 1998). The samples were
heated overnight at 105  C to determine TS and ashed at 550  C
to determine VS. The pH was measured using pH meter (FE20
LAB). Fibre composition was measured using the Laboratory
Analytical Procedure (LAP) developed by the National
Renewable Energy Laboratory (NREL) (Sluiter et al., 2008). The
solid samples were dried and observed with a scanning electron microscope (SEM) (JSM-6390LV, NTC, and Japan). Glucose
in the hydrolysate was analysed using an Agilent1220 chromatography system equipped with a Zorbax carbonhydrate
analytical column, 4.6  150 mm, (Agilent, Palo Alto, CA, USA)
and a refractive index detector. The mobile phase was 75%
acetonitrile with a flow rate of 0.8 ml min1. Samples were
filtered through a 0.22 mm membrane before injection.
The biogas component was detected by a gas chromatography, GC9790II, (FULI, Zhejiang, China) equipped with a
thermal conductivity detector (TCD), a 1.5 m stainless steel
packed column with 5A molecular sieve and a Hayesep Q
packed column (ATEO, Lanzhou, China). The temperatures of
the injector, detector and oven were maintained at 55  C,
100  C and 50  C, respectively. Argon was used as the carrier
gas at a flow rate of 30 ml min1.



A modified Gompertz equation (Eq. (2)) was used to fit cumulative methane production in the batch experiments
(Chang, Tu, Huang, Lay, & Lin, 2011).

Rm e
l  t 1
Ht P exp  exp


where H (t) is the cumulative methane volume at time t (h), P

(ml [CH4]) is the methane potential obtained during stationary
phase, Rm (ml [CH4] h1) is the maximum methane production
rate and obtained during exponential phase, and l (h) is lag
time. Those parameters were estimated by nonlinear curve
fitting in Origin 9.0 (OriginLab, Northampton, MA, USA). The
specific methane potential, Ps (ml [CH4] g1 [CODadded] h1) and
the specific maximum methane production rate, Rs
(ml [CH4] g1 [CODadded] h1) were obtained by dividing P and
Rm by COD added. Batch fermentation from different hydrolysate was compared based on Ps, Rs and l.


Results and discussions

Effect of pretreatment on the glucose conversion and
COD concentration
Dilute alkaline and acid pretreatment methods were adopted
to enhance the biogas production of hydrolysate and degradation of lignocellulose. The effects of concentration and
temperature on glucose conversion with retention time from
12 to 60 h were shown in Fig. 1.
For alkaline pretreatment, it could be seen that the glucose
conversion rate increased sharply with the increase of NaOH
concentration (Fig. 1a). It reached 42.8% at 2% [NaOH], 60 h of
retention time and 45  C of retention time, and improved by
7% at 3% [NaOH] compared with 2% [NaOH]. As the NaOH
concentration was 4%, the glucose conversion decreased
dramatically. A possible reason is the conversion of reducing
sugars into other byproducts because of high alkaline concentration. The xylose and glucose respectively converted into
the furfural and HMF through dehydration. Palmqvist and
gerdal (2000) reported similar findings. It was
observed that COD also decreased when NaOH concentration
reached 4% (Fig. 2). This is consistent with the phenomenon of
glucose conversion. A least squares analysis was conducted to
assess the interaction between time and concentration. It
showed no significant (p > 0.05) difference between NaOH
concentration of 2% and 3% at 60 h. The NaOH concentration


b i o s y s t e m s e n g i n e e r i n g 1 3 2 ( 2 0 1 5 ) 4 7 e5 5

Fig. 1 e Glucose conversion from rice straw under different treatment conditions.

Fig. 2 e COD concentration from hydrolysates under different treatment conditions.

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of 2% was beneficial by considering the economic cost. The

temperature of pretreatment had an obvious effect on the
glucose conversion. Higher temperatures result in more
intense hydrolysis and enhance glucose conversion. Many
studies have been conducted at high temperatures, such as
120e160  C (Satari Baboukani et al., 2012; Yue, Teater,
MacLellan, Liu, & Liao, 2011). To reduce energy consumption,
this study considered moderate temperatures (15e75  C). It
was observed that as temperature increased from 15  C to
75  C, glucose conversion gradually improved (Fig. 1b). Relative to the reference value of 60  C glucose conversion at 75  C
was not significantly improved at 60 h. Thus a temperature of
60  C was suitable for NaOH pretreatment. In addition, to
achieve higher sugar yield and COD, the pretreated time was
increased and the hydrolysis of rice straw was sufficient. It is
likely that long reaction times will result in complex reactions.
Moreover, glucose conversion at 60 h was greater than at 48 h.
So to avoid further degradation reaction of sugars, 60 h was
chosen as a suitable time. In summary, the suitable conditions
determined in this study were NaOH concentration of 2%, a
retention time of 60 h, and a temperature of 60  C. Under these
conditions, in the experiments, glucose conversion reached
Compared with alkaline pretreatment, it was interesting
that glucose conversion decreased at 48 h and 60 h with an
increase in HCl concentration (Fig. 1c). The main reason was
that the hemicellulose was easily degraded in the acid solution (Hendriks & Zeeman, 2009). Satari Baboukani et al. (2012)
also reported that the maximum xylose yield of 91% was
produced from conversion of hemicelluloses via acid pretreatment. Glucose conversion was greatest when the acid
concentration was 0.5%. But in order to achieve a higher COD
concentration (Fig. 2b), a 1% concentration of HCl was
considered as being a better condition. The effect of temperature on glucose conversion was clear (Fig. 1d). Increasing
temperature dramatically enhanced glucose conversion. At a
retention time of 48 h, the glucose conversion at 75  C
improved nearly 4 fold compared with the conversion at 15  C.
In addition, increasing retention time also increased glucose
conversion. A retention time of 48 h was considered suitable
because long retention times result in increased concentrations of degradation products, such as acetic acid, furfural and
HMF (Hendriks & Zeeman, 2009; Rajan & Carrier, 2014). Thus,
analysis of the results concluded that the suitable conditions
for HCl pretreatment were 1.0% [HCl], 48 h of retention time
and a temperature of 75  C.
As is shown in Fig. 2a, the COD concentration of hydrolysate increased rapidly when the NaOH concentration was
raised. After 60 h, the COD concentration ranged from 9.4 g l1
at 0.5% [NaOH] to 14.8 g l1 at 2.0% [NaOH]. Compared with
2.0% [NaOH], 3.0% [NaOH] gave a slight improvement in COD.
However, when the NaOH concentrations were 4.0%, the COD
concentration gradually decreased. This phenomenon agreed
with the trend of glucose conversion. This indicated that the
use of strong alkaline resulted in a dehydration reaction that
reduced sugars (Hendriks & Zeeman, 2009). The average COD
concentration at 2% and 3% [NaOH] from 12 to 60 h were
14.22 g l1 and 14.50 g l1, respectively, which had no significant difference. Thus, this also showed that the selected alkali
treatment conditions were reasonable. Effect of acid


concentration on COD concentration was shown in Fig. 2b.

The COD concentration at different retention times showed a
similar trend as the increase of HCl concentration reaching its
highest value at 1.0% [HCl] and 48 h. This indicated that high
HCl concentrations could result in a decrease in COD concentration. However, the COD concentration from acid hydrolysate was lower than alkaline hydrolysate. Thus, the
moderate alkaline pretreatment was more efficient with
respect to promoting hydrolysis and increasing COD
The effect of temperature on the COD concentration is
shown in Fig. 2c and d. For alkaline pretreatment, the COD
concentration increased significantly when temperature and
retention time was raised. For the 48 h tests, the COD concentration ranged from 6.3 at 15  C to 16.5 g l1 at 60  C, and
slowly decreased to 14.3 g l1 at 75  C. The overall trend was
similar for 48 h and 60 h. To obtain high concentrations of
hydrolysates and low energy consumption, the temperature
of 60  C was judged suitable for alkaline pretreatment. This
was consistent with the previous conclusions. In addition, for
acid pretreatment, the effect of temperature on COD concentration was significant. At 48 h and 60 h, the COD concentration increased as the temperature ranged from 15  C to
75  C. At 48 h, the COD concentration ranged from 4.1 at 15  C
to 9.3 g l1 at 75  C. The COD concentration of 48 h and 60 h
was 9.3 g l1 at 75  C. So the most suitable temperature for acid
pretreatment was 75  C.


Degradation of rice straw via different conditions

To evaluate the degradation of lignocellulose, compositions of

pretreated straw are shown in the Tables 2 and 3. Compared
with the results from non-treated straw, it is evident that
concentration, temperature and retention time had important
effects on cellulose, hemicellulose and lignin contents.
Under the experimental conditions, the hemicellulose and
lignin from alkaline hydrolysis were highly degraded
compared to the cellulose. It can be inferred that the crystallinity of cellulose produced less cellulose degradation (Iroba,
Tabil, Sokhansanj, & Dumonceaux, 2014). The loss of hemicellulose and lignin were 89.45% and 88.92%, respectively, at
4.0% [NaOH]. It can be seen that as the concentration of NaOH
increased, cellulose was gradually hydrolysed. This was due to
the removal of hemicellulose and the lignin to enable the
hydrolysis of cellulose to occur more readily. This could
explain the higher glucose conversion at alkaline pretreatment compared with acid pretreatment. Teater, Yue,
MacLellan, Liu, and Liao (2011) reported a similar finding
when examining the effects of NaOH concentration on corn
stover hydrolysis. When NaOH concentration increased from
0.5% to 2.0% lignin concentration decreased from 14.1% to
10.0%. The delignification reaction was also obvious during
NaOH pretreatment. It was essential to enhance the degradation of lignocellulose which was influenced by NaOH concentration. At 45  C, lignin content ranged from 6.5% (0.5%
[NaOH] for 60 h) to a minimum of 0.27% (3% [NaOH] for 60 h).
Many researchers have reported that lignin is a physical barrier, restricting cellulase access to cellulose and reducing
enzyme activity (McIntosh & Vancov, 2011; Yoon, Han, & Shin,
2014). Moreover, the increase of temperature could improve


b i o s y s t e m s e n g i n e e r i n g 1 3 2 ( 2 0 1 5 ) 4 7 e5 5

Table 2 e Cellulose and hemicellulose content of alkaline pretreated straw.

Concentration (%)

Temperature (oC)

Retention time (h)

Cellulose (%)

Hemicellulose (%)

Lignin (%)






degradation of lignocellulose. At 2% [NaOH], cellulose and

hemicellulose ranged from 33.67% to 12.72% (at 15  C) to
26.47% and 5.95% (at 75  C), respectively.
As regards to acid pretreatment (Table 3), cellulose and
lignin contents were obviously higher in the straw. This
resulted from the decrease in hemicellulose. The hemicellulose contents of different HCl pretreatments showed nearly no
difference. At HCl concentration of 2.5%, the hemicellulose
content (7.29%) was reduced by 75.58% compared with nontreated straw. The results indicated that dilute HCl pretreatments are an efficient method to remove hemicellulose.
Thus, much xylose was contained in the HCl hydrolysate. This
illustrated the phenomenon shown later in the anaerobic
fermentation that gas production of HCl hydrolysate was
higher than in the alkaline hydrolysate. In addition, it can be
seen that as temperature increased, the hemicellulose content gradually reduced. At 0.5% [HCl], hemicellulose was
reduced and ranged from 11.19% (15  C for 60 h) to 7.08% (75  C
for 60 h) in Table 3. However, the lignin content of the straw in
every test was increased. This showed that dilute HCl hardly
 ez, & Castro, 2008).
dissolves lignin (Cara, Ruiz, Oliva, Sa


Scanning electron microscopy of solid residues

To study the effect of alkaline and acid concentration on the

damaged extent of rice straw, solid residues after pretreatment were examined using scanning electron microscopy
(SEM). The results are shown in Fig. 3. It was found that the
fibres were greatly affected by alkaline concentration. At
NaOH concentration of 0.5%, surface of solid residues were
relatively smooth and regular. With the increase of NaOH
concentration, the protective layer included hemicellulose
and lignin was degraded. The cellulose was obviously exposed
in image of 2% [NaOH]. The extent of damage is serious

enough to expand and loosen the fibres. The possible reason

for this could be that high alkaline concentrations degrade the
lignin giving access to the cellulose. Nguyen et al. (2010) reported similar results and made similar inferences. Compared
with alkaline pretreatments, the extent of damage from acid
pretreatments was slight. This could be explained by the fact
that acid degraded the hemicellulose but did not break down
the lignin. Because there was only slight damage to the lignin,
the surface of acid pretreatment samples remained smooth.

Batch methane production at different hydrolysate
Batch anaerobic fermentation was conducted to study the
biogas potential of alkaline and acid hydrolysate. To explore
the best feeding load, hydrolysates were set up four different
concentrations by diluting using de-ionized water: 100%, 75%,
50%, and 25% (v/v). Methane production at different hydrolysate concentrations and the theoretical methane production
calculated using the Gompertz equation (Eq. (2)) is shown in
Fig. 4.
For both alkali and acid hydrolysate, methane production
was improved with increasing hydrolysate concentration.
After 72 h fermentation was basically complete since the cumulative gas production at 72 h reached over 85% of total gas
production. After 120 h, no biogas was produced in fermented
reactors. Methane yields from alkaline hydrolysate were
193.2, 162.6, 135.8, and 89.4 ml g1 [COD] at different concentrations of 100%, 75%, 50%, and 25%, respectively. Similarly,
the methane yields from acid hydrolysate were 275.5, 279.2,
248.7, and 287.0 ml g1 [COD] at the different concentrations of
100%, 75%, 50%, and 25%, respectively. Obviously, the
methane yield from acid hydrolysate was higher compared
with the alkali hydrolysate. This was because the anaerobic

Table 3 e Cellulose and hemicellulose content of acid pretreated straw.

Concentration (%)

Temperature (oC)

Retention time (h)

Cellulose (%)

Hemicellulose (%)

Lignin (%)






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Fig. 3 e SEM images of the solid residues from different pretreatment.

Fig. 4 e Methane productions from hydrolysate at different concentration during batch fermentation (a. NaOH pretreatment,
b. HCl pretreatment).


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Table 4 e Gompertz equation kinetic parameters, final pH, and COD consumption during batch experiments at different
hydrolysate concentrations.
Alkali 75%
Alkali 50%
Alkali 25%
Acid 100%
Acid 75%
Acid 50%
Acid 25%

COD conc.
(g l1)

Ps (ml [CH4] g1


Rs (ml [CH4] g1


l (h)

CH4 conc.


COD consumption








fermentation was conducted under similar COD of acid and

alkali hydrolysates conditions. Although the alkali pretreatment gave hydrolysate with higher COD and better cellulose
degradation, the gas production rate of the acid hydrolysates
was high. Another possible reason was that alkali pretreatment resulted in lignin solubilisation and formation of
gerdal, 2000).
inhibitory substances (Palmqvist & Hahn-Ha
The methane content, final pH and COD consumption are
shown in Table 4. For alkaline or acid hydrolysate, the
methane contents of biogas were higher at 75% and 100% (v/
v) hydrolysate. It reached 68 1% at 75% and 100% experiments. In addition, it was found that the COD consumption
from the acid hydrolysate was higher than the alkali hydrolysate. This phenomenon was consistent with the change in
gas yield. The results were the opposite of those obtained by
Kongjan, Kotay, Min, and Angelidaki (2010) where hydrogen
yield decreased as the hydrolysate concentration increased.
This difference could be attributed to the influence of high
organic loading, which inhibited hydrogen producing bacteria by changing the acid-producing rate. This study indicated
that high organic loading resulted in more rapid acid production and improved methanogenic bacterial activity producing higher gas yields. The highest methane potential was
287.0 ml g1 [COD] obtained from fermentation with 25% acid
hydrolysate. We conclude that acid hydrolysate obtains
higher methane production than alkali hydrolysate at similar
COD conditions.
The parameters of the Gompertz model were shown in
Table 4. The specific methane yields (Ps) from the simulated
results are slightly different from the experimental methane
potential values. Gas production can be assessed based on
these parameters and l can be easily estimated. The current
study suggests that high COD loading results in high methane
production. For the same concentration hydrolysate from alkali and acid pretreatment, l from alkali hydrolysate was
longer than for the acid hydrolysate. This indicates that under
similar conditions, alkali hydrolysate required a longer time to
begin fermentation.



In this study, it was found glucose conversion and COD of

alkali pretreatment were higher than acid pretreatment.
And suitable conditions for hydrolysis were 2% (w/w), 60 h
and 60  C with NaOH pretreatment. Under these conditions

glucose conversion was 55.5%. In batch anaerobic fermentation, the results showed acid pretreatment was an efficient method to improve biogas production. The maximum
methane production of alkaline hydrolysate was
193.2 ml g1 [COD]. For HCl hydrolysate, the maximum
methane production of 287.0 ml g1 [COD] was 43.89% higher
than alkali hydrolysate. Although the NaOH solution was
more efficient in improving straw hydrolysis, hydrolysate
from the HCl pretreatment had the greater methane

The project was financed by the National Natural Science
Foundation Program of China for (No. 51406064) and Central
Universities Fundamental Research Funds (No. 2013PY056).
The authors gratefully acknowledge their support.



Cumulative methane volume, ml

Methane potential, ml
Specific methane potential, ml g1
Maximum methane production rate, ml h1
Specific maximum methane production rate,
ml g1 h1
Lag time, h
Time, h

Chemical oxygen demand, g l1
Anaerobic digestion
Total solids, %
Volatile solids, %
Laboratory Analytical Procedure
National Renewable Energy Laboratory
Scanning electron microscope
thermal conductivity detector
Neutral detergent fibre, %
Acid detergent fibre, %

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