Energy 189 (2019) 116190

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Energy
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Methane production performances of different compositions in

lignocellulosic biomass through anaerobic digestion
Shuaishuai Ma, Hongliang Wang*, Jingxue Li, Yu Fu, Wanbin Zhu**
College of Biomass Sciences and Engineering /College of Agronomy and Biotechnology, China Agricultural University, Beijing, 100193, China

a r t i c l e i n f o a b s t r a c t

Article history: Lignocellulosic biomass holds great potential in the production of renewable energy such as biomethane
Received 20 May 2019 via anaerobic digestion. However, the digestion performance of the three main compositions of ligno-
Received in revised form cellulose during biogas production is not well understood so far, and this has greatly hindered the
17 August 2019
efficient use of biomass by anaerobic fermentation. In this study, the anaerobic digestion performances of
Accepted 22 September 2019
glucose, cellulose, hemicellulose, lignin, and their combinations in biomethane production were inves-
Available online 24 September 2019
tigated. The biomethane production potential of cellulose was higher than that of hemicellulose and
lignin, and high crystallinity cellulose shows adverse impact on methane production. Hemicellulose had
Keywords:
Biogas
a fast hydrolysis rate and was easier to be digested than cellulose, but it also easily led to excessive acids
Biomethane production potential accumulation. Lignin showed a very limited potential in biomethane production. The co-fermentation of
Lignocellulose cellulose and hemicellulose could promote cellulose conversion and eliminate the over-acidification of
Anaerobic digestion hemicellulose, resulting in efficient and stable biogas production. The co-fermentation of lignin with
Co-fermentation cellulose and hemicellulose showed an inhibition in biomethane production, and lignin with different
structure and sulfur content had different degree of inhibition. This study provides insights into the
mechanisms of anaerobic fermentation of lignocellulosic biomass, and paves the way for the better use of
biomass.
© 2019 Elsevier Ltd. All rights reserved.

1. Introduction composed of three major components named cellulose, hemicel-

Lignocellulosic biomass, as a kind of renewable organic
resource, is readily available and has a great potential in the pro-
duction of clean energy. Global lignocellulosic biomass production
is approximately 200 billion tons per year [1]. Such a huge amount
of biomass resource can provide sufficient feedstock for the pro-
duction of sustainable energy. With the energy crisis increasing, the
rational use of biomass resources is particularly important. Anaer-
obic digestion (AD) is a promising technology that could be used to
convert a variety of lignocellulosic biomass into biogas [2e4]. AD is
identified as the least energy-intensive process among all bio-
energy production technologies with greenhouse gas abatement
benefits [5]. Lignocellulosic biomass such as agricultural waste and
forestry residue is promising in biogas production via AD process
[6].
Lignocellulose is the main constituent of plant cell wall, and
* Corresponding author.
** Corresponding author.
E-mail addresses: hlwang@cau.edu.cn (H. Wang), wanbin@cau.edu.cn (W. Zhu).
lulose, and lignin [7,8]. From the chemical composition and struc-
tural point of view, cellulose is a linear polymer formed by D-
glucose subunits through b-(1 / 4) glycosidic bonds. Hemicellu-
lose is relatively complex, which is composed of a variety of poly-
saccharides including xylan, glucuronoxylan, arabinoxylan,
glucomannan, and xyloglucan [9]. The most complicated compo-
sition in lignocellulose is lignin which is a three-dimensional
amorphous biopolymer composed of methoxy or hydroxy-
substituted phenylpropane monomers through disordered poly-
merization. Lignin encapsulates cellulose and hemicellulose [10,11]
by crosslinks through hydrogen bonds and covalent bonds, forming
a hydrophobic three-dimensional structure - “Lignin-Carbohydrate
Complexes” (LCC) [12,13]. In addition, about two-thirds of the cel-
lulose arranges in an ordered structure to form stable crystalliza-
tion zones, and the hydrogen bonds with high density in the
crystallization zones make cellulose less soluble and recalcitrant in
its conversion [14,15]. This complex structure of lignocellulose
prevents plant cell wall from rapid degradation by microorganisms,
thereby rendering biomass hard to be efficiently converted during
AD [16]. The complex structure of lignocellulose also causes trouble

https://doi.org/10.1016/j.energy.2019.116190
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2 S. Ma et al. / Energy 189 (2019) 116190
Abbreviations
AD Anaerobic digestion
AL Alkali lignin
APHA American Public Health Association
BD Biodegradablity
CMY Cumulative methane yield
DC Decrystallization cellulose
DMY Daily methane yield
ECF Experimental chemical formula
EMY Experimental methane yield
GLU Glucose
LCC Lignin-Carbohydrate Complexes
for understanding fermentation performance of cellulose, hemi-
cellulose, and lignin in biogas production, and hinders biomass
efficient utilization. Therefore, the investigation of digestion char-
acteristics of the three compositions of lignocellulose during their
separated or mixed fermentations is necessary, which will help to
better understand and utilize of the AD process of lignocellulose.
At present, there are tremendous researches on biogas pro-
duction by using lignocellulosic biomass through various pre-
treatments. However, most of these studies only consider the
overall conversion of lignocellulose, and there is only a rough es-
timate of the biogas production potential, utilization rate, and
digestion mechanism of each components in lignocellulose
[17e20]. The digestion rate and the biogas production potential of
the individual component in lignocellulose, as well as the interac-
tion of the three components in AD process have not been clearly
studied. In this study, the AD performance of cellulose, hemicel-
lulose, lignin, and their mixtures were carefully studied for biogas
production. The cumulative methane yield (CMY), hydrogen ion
concentration (pH), volatile fatty acids (VFAs), daily methane yield
(DMY), total alkalinity (TA), soluble chemical oxygen demand
(sCOD) of AD with different ratios of lignocellulosic compositions
were investigated. Besides, the modified Gompertz model was used
to evaluate the kinetic parameters during biogas production. On the
basis of this research, reliable supports were given for the better
utilization of lignocellulosic biomass resources to produce bio-
methane through AD processes.
2. Materials and methods
2.1. Materials
Microcrystalline cellulose (MC) was bought from Shanghai test
Co., Ltd., China. Decrystallization cellulose (DC) was made by MC via
the following method: 20 g MC was dry-milled for 10 min, and then
added to a high speed blender containing 1 L 4 wt% NaOH solution,
shearing for 30 min [21,22]. The pretreated cellulose was separated
by centrifugation and dried in a freezer dryer. Xylan (XY), extracted
from corncob, was bought from Macklin, China, and it was used as
the representative of hemicellulose. Glucose (GLU) was bought
from Shanghai test Co., Ltd., China. Lignin (LI) was prepared from
beech wood powder by milling extraction. Alkali lignin (AL) was
bought from Macklin, China, and sodium lignosulfonate (SL) was
bought from Aladdin, China. Besides, the inoculum was taken from
a continuous biogas fermenter in Zhuozhou City, Hebei, China,
which was fed by corn stalk and pig manure.
LI Lignin
MC Microcrystalline cellulose
pH Hydrogen ion concentration
sCOD Soluble chemical oxygen demand
SL Sodium lignosulfonate
SUM Sum of C, H, O, N and S
TA Total alkalinity
TMMY Theoretical maximum methane yield
TS Total solid
VFAs Volatile fatty acids
VS Volatile solid
XY Xylan
2.2. Experimental design
To investigate the biogas production rate and potential in AD of
the individual components of lignocellulose and the interactions of
them, several combinations were set. The total initial organic
loading was 15 g-VS/L in all combinations. To compare the AD
features of cellulose, hemicellulose, glucose, and lignin during their
separated fermentation, they were digested in separated reactors.
The substrate-to-inoculum ratio in each digester was 1, which was
based on VS content. Hemicellulose with cellulose, glucose with
cellulose, hemicellulose with cellulose and different type of lignin
were mixed to co-digestions at a ratio of 1:1 or 1:1:1 which were
also based on VS content to explore their interactions during AD.
The total volume of each reactor was 500 mL with a 200 mL
working volume, and the initial pH of each digester was adjusted to
7.0 ± 0.1 by hydrochloric acid or sodium hydroxide solution. The
inoculum without any feedstock addition was used as a control. All
experiments were conducted in triplicate. The headspace of all
digesters was filled with pure nitrogen to produce anaerobic con-
ditions before fermentation. Batch fermentations were kept in a
36 ± 1
C constant temperature chamber for 50 days. 2 mL samples
were collected at 0, 2, 4, 8, 13, 18, 23, 30, 40 and 50 days for each
parallel, and mixed into a 10 mL centrifuge tube before use. During
analysis, solids were separated from liquids by centrifugation at
5000 rpm for 10 min.
2.3. Analysis of solid samples
The elemental compositions of each substrate were measured
by an elemental analyser (EA3000, EuroVector, China). The volatile
solid (VS), total solid (TS) and ash content were measured according
to American Public Health Association (APHA) Standard Methods
[23].
2.4. Analysis of liquid samples
The total alkalinity (TA) of samples was titrated by bromocresol
green-methyl red titration standard solution. The soluble chemical
oxygen demand (sCOD) was determined by a COD rapid detector
(Model ET99731, Lovibond, Germany, 131 program). The sample
was centrifuged and mixed with the digestion solution with a ratio
of 2:3 to digest for 2 h. The hydrogen ion concentration (pH) of
liquids was determined by a portable pH meter (Shanghai Sanxin
Instrument Factory, model SX-620). The concentration of volatile
fatty acids (VFAs) including formic acid, acetic acid, propionic acid,
butyric acid and lactic acid were measured by high-performance
liquid chromatography (HPLC; Model LCMS-2020, Shimadzu,
Tokyo, Japan) with a HPX-87H column (300  7.8 mm; Bio-Rad
 S. Ma et al. / Energy 189 (2019) 116190 3

Laboratories, Hercules, CA, USA).

BD ¼ ðEMY = TMMYÞ  100% (5)
2.5. Analysis of biogas where EMY is experimental methane yield.

In the batch anaerobic fermentation reactor, the daily biogas

2.7. Kinetic model fitting
production was measured using a pressure measurement system
(Mess-und Regelsysteme GmbH, WAL, Germany) with a measure-
The biogas production process was fitted by using the modified
ment accuracy of 0.01%. The measured pressure was converted to
Gompertz model and analyzed using Origin 9.1 software. The for-
the yield of biogas through Eq. (1) [24]:
mula was Eq. (6):

.
Vbiogas ¼ DP  Vheadspace  M ðG  TÞ (1) P ¼ Pmax  expf  exp½b  e  ðl  xÞ = Pmax þ 1g (6)

where Vbiogas is the volume of Nissan biogas (mL), DP is the absolute
pressure difference between pressure and atmospheric pressure in
the reactor (kPa), Vheadspace is the volume of the head space of the
reactor (mL), M is the molar volume (22.41 at 273.15 K L/mol,
101.325 kPa), G is the general gas constant (8.314 L kPa/K/mol), T is
the absolute temperature (K).
The methane content was determined by a gas chromatography
(GC-2014, Shimadzu, Japan). The formula for calculating methane
content was shown in Eq. (2):
where P refers to the cumulative methane yield (mL/g-VS); Pmax
represents the predicted potential methane yield (mL/g-VS); b
represents the maximum methane production rate (mL/g-VS/day);
e is a constant with a value of 2.7183; l represents the fermentation
period of delay, and x represents the fermentation time (days) [26].
3. Results and discussion
3.1. Characteristics of substrates and inoculum

YCH4 ¼ 3229865XCH4 þ 29545 (2) The characteristics of substrates and inoculum is listed in
Table 1. MC, DC, XY and GLU had high VS contents (more than
where Y is the peak area, X is the methane content (%), and
98.12%). SL and AL showed relatively low VS contents and high ash
R2 ¼ 0.9931. Eq. (2) was derived from a standard curve of CH4 peak
contents. These results confirmed that cellulose, hemicellulose, and
area (Y) correlated with CH4 content (X) in mixed gases of N2, CH4,
glucose had high content of organic matter. Besides, LI contained
and CO2. Gas content was analyzed by a gas chromatograph.
only 0.09% sulfur, which was lower than AL (3.22%) and SL (3.82%).
All the components of lignocellulose had very high C/N ratios,
2.6. Theoretical maximum methane yield and biodegradability indicating they are not suitable for AD alone. Therefore, in actual
calculations industrial production, lignocellulosic biomass was always co-
fermented with nitrogen-rich raw materials.
The theoretical maximum methane yield (TMMY) was deter-
mined by the formula which was based on the elemental compo- 3.2. Cumulative methane yield
sitions of the raw materials [25], as shown in Eqs. (3) and (4):
    3.2.1. Cumulative methane yield (CMY) of cellulose, hemicellulose
b c 3d a b c 3d
Ca Hb Oc Nd þ a   þ H2 O/ þ   CH4 and glucose
4 2 4 2 8 4 8
  As shown in Fig. 1, the CMYs of MC and DC were 211.8 mL/g-VS
a b c 3d and 298.2 mL/g-VS, respectively, which were higher than XY (133.9
þ  þ þ CO2 þ dNH3 (3)
2 8 4 8 mL/g-VS) and GLU (100.2 mL/g-VS). After pretreatment, the crys-
tallinity of MC decreased from 95.39% to 59.91% (Fig. 2). Meanwhile,
22:4  1000  ða=2 þ b=8  c=4  3d=8Þ the CMY of MC improved from 211.8 mL/g-VS to 298.2 mL/g-VS,
TMMY ¼ (4) clearly indicating that a reduction of the crystallinity in cellulose
12a þ b þ 16c þ 14d
was beneficial for the methane production in AD. This result is
where CaHbOcNd is experimental chemical formula. consistent with previous findings that the crystallinity of cellulose
The formula for calculating biodegradability (BD) was deter- decreased with the increase in the rate of enzymatic hydrolysis
mined by Eq. (5): [27,28]. A high efficiency of cellulose hydrolysis will result in an

Table 1
Characteristics of substrates and inoculum.

Parameter MC DC XY GLU LI AL SL Inoculum

TS (%)a 97.71 ± 0.15 97.89 ± 0.38 96.36 ± 0.22 91.63 ± 0.1 94.09 ± 0.05 95.53 ± 0.17 95.76 ± 0.14 11.16 ± 1.3
VS (%)a 97.70 ± 0.12 97.87 ± 0.77 96.31 ± 0.11 91.62 ± 0.12 92.70 ± 0.11 30.6 ± 0.26 64.94 ± 1.53 4.16 ± 0.28
Ash (%)a 0.01 ± 0.00 0.02 ± 0.01 0.04 ± 0.00 0.01 ± 0.01 0.00 ± 0.00 67.97 ± 0.26 29.06 ± 4.73 62.59 ± 1.75
C (%)b 44.72 ± 0.03 44.78 ± 0.01 42.89 ± 0.05 42.61 ± 0.12 46.93 ± 0.04 55.04 ± 0.08 44.70 ± 0.10 12.31 ± 0.13
H (%)b 6.23 ± 0.02 5.01 ± 0.02 5.59 ± 0.03 6.41 ± 0.34 5.06 ± 0.12 3.98 ± 0.28 4.05 ± 0.32 1.61 ± 0.05
O (%)b 48.21 ± 0.12 48.24 ± 0.16 50.52 ± 0.22 50.13 ± 0.31 46.81 ± 0.13 31.34 ± 0.25 37.81 ± 0.27 17.17 ± 0.63
N (%)b 0.14 ± 0.00 0.01 ± 0.00 0.01 ± 0.00 0.01 ± 0.00 0.02 ± 0.00 0.05 ± 0.00 0.03 ± 0.00 1.22 ± 0.02
S (%)b 0.06 ± 0.01 0.08 ± 0.01 0.09 ± 0.01 0.08 ± 0.00 0.09 ± 0.00 3.22 ± 0.01 3.82 ± 0.02 0.46 ± 0.04
SUM (%)b 99.37 ± 0.12 98.12 ± 0.18 99.10 ± 0.20 99.23 ± 0.53 98.91 ± 0.05 93.63 ± 0.10 90.41 ± 0.47 32.77 ± 0.61
ECF C27H45O22N C365H501O302N C357H559O316N C322H641O313N C196H253O146N C92H80O39N C124H135O79N C8H13O9N

SUM: Sum of C, H, O, N and S; ECF: Experimental chemical formula; MC: Microcrystalline cellulose; DC: Decrystallization cellulose; XY: Xylan; GLU: Glucose; LI: Lignin; AL:
Alkali lignin; SL: Sodium lignosulfonate.
a
Calculated as total weight (%).
b
Calculated as TS (%).
4 S. Ma et al. / Energy 189 (2019) 116190
Fig. 1. Cumulative biomethane yield of cellulose (MC and DC), hemicellulose (XY) and
glucose (GLU).
Fig. 2. Comparison of crystallinity between microcrystalline cellulose (MC) and
decrystallization cellulose (DC).
increase in methane production. This result also demonstrates that
appropriate pretreatments to reduce the crystallinity of cellulose in
raw materials can obviously increase the yield of methane in AD.
Compared to MC and DC, the CMYs of XY and GLU were only
133.9 mL/g-VS and 100.2 mL/g-VS, respectively. XY and GLU did not
produce biogas at the beginning of the fermentation. They began to
produce a small amount of biogas till digested for 24e26 days. This
only lasted for a short time, and biomethane production stopped
after about 35 days of digestion. The duration of biogas production
by using XY and GLU as feedstocks was also much shorter than that
by using cellulose. Biogas production stopped after digestion for
about 35 days. This was probably due to the generation of large
amount of acids volatile fatty acids (VFAs) during AD, which caused
rapidly decrease in pH as measured to below 6 during AD. The rapid
reduction of pH inhibited the activity of methanogenic microor-
ganisms. After 24 days, XY resumed biogas production, while GLU
was delayed for a few days. This result is mainly attributed to that
VFAs is gradually converted to methane leading to an increase in pH
and the activity of methanogenesis. Hemicellulose is usually the
first used lignocellulosic composition by microorganisms during
AD. Many studies had also shown that hemicellulose has the most
obvious change before and after the fermentation among all the
three lignocellulosic compositions [29e31]. Compared to hemi-
cellulose, glucose was more susceptible to hydrolysis in the early
stage of anaerobic fermentation, and the acidification stage was
earlier than hemicellulose, which led a lower CMY.
As shown in Fig. 2, XY and GLU were too easy to be decomposed,
leading to fermentation stagnation during AD. To avoid this unde-
sirable feature, the co-fermentation of MC with XY or GLU was
carried out. As shown in Fig. 3, biogas from co-fermentations of MC
with GLU or XY could be generated at the early stage of the
fermentation, and did not stop until day 40. This result was
completely different from that of the AD process by using XY and
GLU alone. Moreover, the CMYs of MC þ XY and MC þ GLU were
38.24% and 62.18%, respectively, which were higher than those of
XY and GLU mono-fermented. The addition of MC to XY or GLU
could alleviate the acidification phenomenon during fermentation,
which was consistent with the result of co-digestion of hemicel-
lulose and cellulose reported by Li et al. [32]. This result is due to
that complexes formed from MC with XY or GLU had a low hy-
drolysis rate, and then prevented the rapid generation of VFAs and
eliminated the inhibition of methane production. All these results
indicate that the addition of cellulose could be a good way to
improve the AD processes of xylan and glucose, resulting in a
higher biomethane production.
3.2.2. Cumulative methane yield of LI, AL and SL in MC þ XY
As an important composition of lignocellulose, the digestion
behavior of lignin was studied. Biogas production was negligible
when lignin was digested alone. Thus, lignin with different char-
acters was co-fermented with MC and XY to investigate their in-
fluences on biogas production during AD. It was observed that CMY
decreased after the addition of lignin (Fig. 4), indicating lignin has
an inhibition effect in carbohydrate fermentation. Lignin acted as a
shield to protect cellulose and hemicellulose from enzymatic
decomposition in lignocellulose. Besides, during enzymatic hy-
drolysis process, some enzymes might combine with lignin irre-
versibly through hydrophobic interactions, which led to an
inactivation of them [33]. The inhibition caused by AL and SL was
more serious than that caused by LI. LI, AL and SL contained 0.09%,
3.22%, and 3.82% sulfur, respectively, indicating that sulfur was also
an adverse factor on AD. Sürmeli et al. investigated the effect of
sulfides on anaerobic fermentation of chicken manure and found
Fig. 3. Cumulative biomethane yield of microcrystalline cellulose (MC) co-fermented
with xylan (XY) or glucose (GLU).
 S. Ma et al. / Energy 189 (2019) 116190 5

Fig. 4. Cumulative biomethane yield of three kinds of lignin co-fermented with

microcrystalline cellulose (MC) and xylan (XY).

that both sulfur and H2S gas generated during the fermentation
could inhibit the activity of methanogens, and resulted in a
decrease in methane production [34]. Besides, the CMY of SL was
lower than AL, probably because the sulfonic acid group in SL
inhibited AD more seriously than other sulfur containing functional
groups. Moreover, SL is an anionic surfactant which is believed to
inhibit anaerobic fermentation because of its ability to cause loss of
biological function by reacting with components of cells and
various macromolecules such as proteins and DNA [35].

3.3. Changes of VFAs, pH and daily methane yield during AD

To get insights into AD process, the changes of VFAs, pH and

daily methane yield (DMY) during the AD of different feedstocks
were carefully studied. From Fig. 5, it could find that in the first 30
days, the total VFAs concentration of each AD system was high, and
Fig. 5. Changes in pH, VFAs and daily biomethane yield during anaerobic digestion.
then the VFAs gradually decreased and the biogas production
increased, indicating that VFAs were the important intermediates
in biogas production. The total VFAs of XY and GLU in mono-
was below 1800 mg/L all the time during co-fermentation (Fig. 5b).
fermentation were higher than that of other groups (Fig. 5a). In
All the phenomena indicated that the co-fermentation of hemi-
the early stage of AD, XY and GLU were rapidly hydrolyzed to form
cellulose or glucose materials with high-cellulose-content mate-
lots of VFAs, including acetic acid, butyric acid and propionic acid.
rials would be a viable strategy to eliminate excessive VFAs
The pH decreased to below 6 which could inhibit methanogen
cumulation. In the co-fermentation of carbohydrates with different
growth in the fermentation broth. However, this environment was
lignin, the total VFAs concentration was low, which indirectly
beneficial for acid-production bacteria, since the optimal pH range
indicated that lignin could inhibit carbohydrate hydrolysis to acids.
for them is 5.5e6.5 [36]. So, at the beginning of AD, biomethane
The addition of lignin might interact with carbohydrates or en-
was produced in a small amount, and stopped shortly because XY
zymes, resulting in preventing enzymes from well contacting with
and GLU were easily hydrolyzed to various acids. The concentration
carbohydrates.
of butyric acid in XY (1.506e0.944 g/L) and GLU (2.102e1.069 g/L)
digestions was higher than that in MC þ XY (0.254e0.163 g/L) and
MC þ GLU (0.174e0.147 g/L) digestions. Li et al. studied the 3.4. Changes of VFAs/TA, pH and sCOD during AD
fermentation of xylan, arabinogalactan, and glucomannan, and
found the same phenomenon [32]. Studies had shown that the Changes of VFAs/TA, pH and sCOD during the AD of different
activity of methanogens would not be significantly inhibited when feedstocks were also studied. The larger the sCOD value, the more
the concentration of butyric acid and acetic acid was lower than content of easy degradable substances in AD. In the co-
1800 and 2400 mg/L, respectively [37]. However, more than fermentation system containing AL or SL, sCOD was over
1800 mg/L butyric acid was produced during the fermentation of 13000 mg/L, obviously higher than other fermentations (Fig. 6c). AL
XY and GLU, and that could be the reason for the inhibition of AD and SL were by-products of pulp and paper. The pulp black liquor
processes. With acetic acid and butyric acid being gradually itself contained high sCOD [38]. During the fermentation of XY or
consumed, biogas production was gradually recovered. The VFAs of GLU, the sCOD gradually decreased from high, but VFAs/TA was
MC þ XY and MC þ GLU were relative lower as compared with XY above 0.4 for a long time (Fig. 6a). The VFAs/TA value was generally
and GLU mono-fermentation, and the butyric acid concentration used to evaluate the stability of AD system. Once VFAs/TA was
6 S. Ma et al. / Energy 189 (2019) 116190

MC, which indicated that cellulose with low crystallinity benefited

for fermentation conversion. The BD of XY and GLU were 35% and
26.3%, respectively, indicating that it was not a good choice for
fermenting hemicellulose or glucose separately, since they were
easily acidified and inhibited the activity of methanogen. After co-
digestion with MC, the BD of XY and GLU increased to 46.9% and
41.2%, respectively. This might be due to the addition of MC in XY or
GLU caused a nutritional balance in AD, which was beneficial for
the growth of various microorganisms [32]. Thus, the co-
fermentation avoided the inhibition of methane production by
high VFAs concentrations [39,40]. After the addition of LI, AL, or SL
in MC and XY co-fermentation system, the co-digestion BD changed
from 46.9% to 56.1%, 42.9% and 35.6%, respectively. The reason for
the increase in BD after adding LI was that LI contained a small
amount of hemicellulose. The addition of AL and SL to the
fermentation system decreased BD due to the inhibition effect of
lignin itself and sulfur in it. These results once more demonstrate
that lignin, especially those with high content of sulfur, is a sig-
nificant obstacle for the production of biomethane through ligno-
cellulose by AD.

3.6. Kinetic analysis of biogas production by modified Gompertz

model

As an important model for the study of methane production

Fig. 6. Changes of VFAs/TA, pH and sCOD during anaerobic digestion.
higher than 0.4, the AD system would become unstable and
adversely affect biogas production 32. The pH of fermentation
broth by using XY or GLU as feedstock was also below 6.5, which
was bad for biogas production. MC and DC had a short period of
VFAs/TA greater than 0.4 during the fermentation process, but the
system could return to stability soon.
3.5. Biodegradability
As shown in Table 2, DC had a higher biodegradability (BD) than
kinetics, the modified Gompertz model can be used to predict/
describe the relationship between CMY and time (Table 2). All R2
values were above 0.956, which means the modified Gompertz
model can well reflect the methanogenesis of AD. The l value can
mirror the reaction rate of the substrate in the AD system, which
reflects the degree of adaptation of microorganisms to raw mate-
rials in the AD system. A low l values reflects that microorganisms
can adapt quickly to the new environment with no significant
delay, which means that the fermentation system will produce
methane without delay. Both XY and GLU had a large l, indicating
that the microbes had been greatly affected during the fermenta-
tion process, causing methane production to stagnate for a while.
When MC was added to XY or GLU fermentation systems, the value
of l decreased significantly, indicating that the addition of cellulose
could improve the fermentation of glucose and hemicellulose. Be-
sides, XY and GLU had almost the same TMMY (382.5 mL/g-VS and
381.4 mL/g-VS), but the Pmax and EMY of XY were higher than GLU.
This result indicates that the fermentation potential of XY and GLU
was close, but GLU was more susceptible to acidification during AD.
MC had higher Pmax and TMMY values than XY and GLU, indicating
that the fermentation potential of cellulose was the highest as
compared with other lignocellulose components.

Table 2
Methane production performance of cellulose, hemicellulose, glucose and their combinations.

Substrates TMMY (mL/g-VS) EMY (mL/g-VS) BD (%) Modified Gompertz model

Pmax b l R2

MC 407.8 211.8 51.9% 240.2 6.7 3.8 0.992

DC 390.3 298.2 76.4% 367.7 8.5 5.2 0.993
XY 382.5 133.9 35% 136.2 15.5 23.3 0.991
GLU 381.4 100.2 26.3% 102.8 13.1 27.4 0.984
MC þ XY 394.6 185.1 46.9% 191.2 10.3 12.2 0.989
MC þ GLU 394.1 162.5 41.2% 171.1 7.1 7.7 0.993
MC þ XY þ LI 402.4 225.8 56.1% 226.6 15.6 5.8 0.995
MC þ XY þ AL 414.8 178.1 42.9% 196.1 4.8 2.6 0.956
MC þ XY þ SL 450.2 160.1 35.6% 159.0 13.1 3.3 0.994

Pmax: Simulated maximum cumulative methane yield (mL/g-VS); b: Maximum methane production rate (mL/g-VS/day); l: Lag phase time (day); R2: Correlation coefficient.
 S. Ma et al. / Energy 189 (2019) 116190
4. Conclusions
The anaerobic digestion characteristics of cellulose, hemicellu-
lose, lignin, glucose and their mixtures during AD were investi-
gated. It was found that the biomethane potential of cellulose was
higher than that of hemicellulose and lignin. Cellulose with high
crystallinity was negative for methane production. Hemicellulose
had a fast hydrolysis rate, which was easier to be used than cellu-
lose, but also easily led to over acidified the fermentation system.
The co-fermentation of cellulose and hemicellulose could eliminate
the rancidity of hemicellulose, and benefit to biogas production.
Lignin shown very limited biogas production potential. The addi-
tion of lignin to the cellulose and hemicellulose co-fermentation
system resulted in an inhibition of AD. Lignin containing sulfur
had a more serious inhibition on AD than that without sulfur. Re-
sults obtained in this study showed biomass pretreatment to
remove or reduce lignin content in the fermentation feedstock,
meanwhile reduce the crystallinity of cellulose, would benefit for
biogas production.
Acknowledgements
This work was supported by the National Key R&D Program of
China (2018YFB1501500) and the National Natural Science Foun-
dation of China (No. 21706277).
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