Natural Science, 2009, 1, 17-22 NS
http://dx.doi.org/10.4236/ns.2009.11004
Research on the Graft Copolymerization of EH-lignin
with acrylamide
Run Fang1,2, Xian-Su Cheng*1, Jian Fu1, Zuan-Bin Zheng1
1College of Material Science and Engineering, Fuzhou University, Fuzhou 350002, China; 2Department of Chemistry and Chemical
Engineering, Minjiang University, Fuzhou 350011, China.
*Corresponding author:
Received 13 April 2009; revised 14 May 2009; accepted 23 May 2009.
ABSTRACT
Lignin isolated from enzymatic hydrolyzed corn-
stalks (EH-lignin) is a renewable natural polymer
noted for its versatility and applicability in a vari-
ety of uses. Graft copolymerization of EH-lignin
with acrylamide (AM) and the application of this
copolymer as a flocculant in dye wastewater
treatment were studied in this article. The influ-
ences of some factors on yield of copolymer and
the grafting ratio were investigated and the
structure of EH-lignin/AM graft copolymer was
characterized by FT-IR. According to the yield and
the grafting ratio, the optimum conditions for
graft copolymerization were as follows: initiator
K2S2O8-Na2S2O3 with a quantity 3 wt% of EH-lignin,
mass ratio of AM to EH-lignin was 2~3, reaction
time 4h and temperature at 50℃. It was found that
the absorption capacity of graft copolymer to two
azo-dyes was enhanced with the increase of
grafting ratio. Furthermore, the residue concen-
tration of EH-lignin/AM graft copolymer remained
in the supernatant after flocculation was much
lower than that of pure EH-lignin.
Keywords: Lignin; Acrylamide; Graft Copolymeriza-
tion; Dye Wastewater; Decoloration
1. INTRODUCTION
At present, the fossil resources are rapidly running out
and the environmental pollutions are getting even more
serious throughout the world. Great attention has been
paid to the development of sustainable technologies
based on renewable raw materials [1,2,3,4]. As a natural
polymer, lignin is a renewable and biodegradable re-
source and noted for its versatility and applicability in a
variety of uses. Making use of these biomaterials will
not only enhance the economic benefit of bioengineering
but also diminish environmental pollutions [5,6].
EH-lignin is a novel ornanosolv lignin isolated from
the residue of enzymatically hydrolyzed cornstalks as a
by-product of fuel ethanol industry [7]. Compared with
traditional lignosulfonate or alkali lignin, EH-lignin pos-
sesses some valuable characteristics: lower content of
sugar, less impurities and narrow molecular weight dis-
tribution. Furthermore, since the enzymatic hydrolysis
process of the cornstalks is carried out under relatively
mild conditions, many functional groups such as pheno-
lic hydroxyl, alcoholic hydroxyl and methoxyl are well
preserved in EH-lignin [8,9,10].
Due to its abundant functional groups, EH-lignin can
be used in dye wastewater treatment by adsorbing dyes
through hydrogen bonding under acidic conditions.
However, the concentration of residue lignin remains in
supernatant after flocculation is very high, which may
leads to a secondary pollution. In order to minimize the
potential secondary pollution, graft copolymerization of
EH-lignin with acrylamide (AM) was studied in this
paper. The effects of some factors on the copolymeriza-
tion were investigated. The structure of EH-lignin/AM
copolymer was analyzed by FT-IR and its application in
the dye wastewater treatment was evaluated. The residue
concentration of this flocculant remains in the super-
natant after flocculation was measured.
2. EXPERIMENTAL
2.1. Materials
EH-lignin was supplied by Tianguang fuel ethanol com-
pany (He’nan, China) in powder form and purified in
laboratory according to procedures described in our pre-
vious article [4]. Characteristics of purified EH-lignin
are shown in Table 1. The details of two azo-dyes, acid
red 274 (AR 274) and reactive red X-3B (RR X-3B),
were shown in Fig. 1 and Table 2. Acrylamide were
purchased from Guanghua chemical reagent Co., Ltd,
China. All other reagents were of analytical grade.
Table 1. Characteristics of EH-lignin.
Residual
sugar/%
Ash
/%
Phenolic hy-
droxyl/mmol.g-1 Mw Mw/Mn
EHLignin
0.22 0.394.25 20621.22

18 R. Fang et al. / Natural Science 1 (2009) 17-22
SO3Na
NaO3S
NH
O
OH
NN O
NN
OH
NaO 3SSO3Na
NHC
N
N
C
C
N
Cl
Cl
(a) (b)
Figure 1. Molecular structure of (a) Acid red 274 and (b) Reactive red X-3B.
Table 2. Details of the dyes.
Dyes Abbreviation Molecular formula CAS number λmax(nm)
Acid red 274 AR 274 C35H31N3Na2O9S2 72828-83-2 527
Reactive red X-3B RR X-3B C19H10Cl2N6Na2O7S2 12226-03-8 538
2.2. Synthesis and Characteristics of EH-
lignin/ AM Graft Copolymer
Graft copolymerization reactions were carried out in a
jacketed reactor flask equipped with a stirrer and a reflex
condenser under N2 protection. Appropriate amount of
EH-lignin, AM and initiators were dissolved in NaOH
aqueous solution and then reacted at different tempera-
tures for a period of time. When a reaction was finished,
copolymer product was precipitated by acidification and
isolated in a centrifuge. In order to remove monomers,
EH-lignin/AM graft copolymers were washed by dis-
tilled water and then vacuum dried.
Viscosity measurement of lignin/AM copolymer in
water solution was conducted by an Ubbelohde type
viscometer at 30.0±0.1. Extrapolation procedure from ℃
data obtained for 5 concentrations of solutions was used
to calculate [η] from Huggins equation, ηsp/c=[η]+k
[η]2c. The intrinsic viscosity was then used to evaluate
the molecular weight of graft copolymers prepared with
different initiators.
The chemical structure of graft copolymer was ana-
lyzed using FT-IR2000 spectrometer (Perkinelmer, U. S.)
and the spectra were recorded in the range of 500-4000
cm-1.
Yield of EH-lignin/AM copolymer and the grafting
ration were determined by Eq.1 and Eq.2 respectively.
Yield
2
01
(%) 100%
W
YWW 
 (1)
Grafting ratio
203
03
(%) 100%
WWW
GR WW


 (2)
where W0 is the weight of EH-lignin; W1 is the weight of
AM monomer; W2 is the weight of the graft copolymer;
W3 is the weight of lignin remained in the supernatant.
2.3. Adsorption and Decoloration of AR 274
and RR X-3B Dye Wastewater
The adsorption and decoloration of AR 274 and RR
X-3B dye wastewater by EH-lignin and EH-lignin/AM
grafted copolymer was investigated by static adsorption
method. Firstly, a certain amount of flocculant was
weighed and dissolved in 2ml 1%NaOH aqueous solu-
tion. Afterward, 200ml dye wastewater with a concentra-
tion of 500mg/L was added into aforesaid copolymer
solution, stirring 3 min to make sure the mixture well-
mixed and then kept undisturbed for 1h. Finally, the so-
lution was acidified with HCl to pH=4 and then filtered
after another 10 min standing. The concentrations of dye
wastewater before and after treatment were measured at
λmax mentioned in Table 2 by UV-2450 spectropho-
tometer. The total organic carbon (TOC) content of the
supernatant was measured by TOC-V analyzer (Shima-
dzu, Japan) to evaluate the residue amount of the floc-
culant and the dyes remained in the supernatant after
flocculation. The absorption amount, decoloration rate
and TOC removal can be calculated by Eq.3, Eq.4 and
Eq.5 respectively.
Adsorption amount
W
VCC
Q

)( 0 (3)
Decoloration rate
%100
0
0


A
AA
E (4)
TOC removal rate

R. Fang et al. / Natural Science 1 (2009) 17-22 19
%100
0
0


B
BB
R (5)
where Q is the adsorption amount, mg/g; C0 and C are
the concentrations of the dye solution before and after
treatment, mg/L; V is the volume of dye solution, L; W
is the amount of graft copolymer, g; E is decoloration
rate; A0 and A are the absorbance of the dye solution
before and after treatment; R is the TOC removal rate;
B0 and B are the TOC values of the dye solution before
and after treatment.
3. RESULTS AND DISCUSSION
The effects of various factors on the yield of copolymer
and the grafting ratio were investigated to determine the
optimum conditions for graft copolymerization. The
basic reaction conditions of these experiments were as
follows: the dosage of EH-lignin was 2.0g, acrylamide
was 4.0g, 100g 1% NaOH aqueous solution was used as
solvent, the amount of K2S2O8-Na2S2O3 was 3% of the
weight of EH-lignin, the reaction temperature was 50 ℃
and the reaction time was 4h.
3.1. Effects of Some Factors on Graft
Copolymerization
3.1.1. Effects of Different Kinds of Initiators on
Graft Copolymerization
The graft copolymerization of EH-lignin with AM was
carried out in aqueous solution, therefore six water-
soluble radical initiators were chosen and their effect on
grafted copolymerization was studied. The results were
showed in Table 1 and the synthesis conditions were as
follows: weight of lignin was 2.0g, acrylamide 4.0g,
reaction temperature was 50, r℃eaction time was 4h and
the dosage of initiator was 3% of the weight of EH-lig-
nin.
Since the raw materials and the reaction procedures
are identical, the chemical structure of lignin/AM co-
polymers initiated by different initiators is quite similar
to each other. Therefore, higher [η] of a lignin/AM co-
polymer’s aqueous solution may indicates a larger mo-
lecular weight of this copolymer. It can be seen in Table 1
Table 3. Effects of different initiators on yield and intrinsic
viscosity of the graft copolymers.
Initiator Yield/% [η]/mL.g -1
Fe2+-H2O2 34.30 35.76
(NH4)2Ce(NO3)6 30.86 34.51
K2S2O8 37.83 35.37
(NH4)S2O8 34.55 37.15
K2S2O8-NaHSO3 37.40 36.93
K2S2O8-Na2S2O3 39.60 37.86
that the graft copolymer initiated by K2S2O8-Na2S2O3
has the largest yield and highest intrinsic viscosity,
which means this binary-initiating system is more effec-
tive in grafting AM onto EH-lignin. For this reason,
K2S2O8-Na2S2O3 was employed in our further research
on the graft copolymerization of EH-lignin with AM.
3.1.2. Effect of Initiator Dosage on Graft
Copolymerization
The effect of initiator dosage on graft copolymerization
was evaluated considering the yield of copolymer and
the grafting ratio. The results were shown in Fig. 2. It
was found that the yield of the copolymer and the graft-
ing ratio increased with increasing initiator dosage at
first. However, when the initiator dosage was more than
3% of the weight of lignin, Y(%) and GR(%) increased
slowly and then decreased when the dosage of initiator
reach 5%.
In the process of the copolymerization, the binary-
initiating system of K2S2O8-Na2S2O3 generated free
radicals to initiate the polymerization of PAM and the
graft copolymerization of EH-lignin with AM or with
PAM chains. On one hand, high free radical concentra-
tion may enhance the possibility of graft copolymeriza-
tion and increase the yield and molecular weight of co-
polymer; on the other hand, it will lower the polymeriza-
tion degree of PAM that may graft onto EH-lignin and
decrease the molecular size of copolymer. Therefore,
Y(%) and GR(%) reach their maximum when the con-
tradiction reaches a equilibrium and the optimum dosage
of initiator is 3% of the weight of EH-lignin.
3.1.3. Effect of Acrylamide Dosage on Graft
Copolymerization
The mass ratio of acrylamide to EH-lignin is another
important factor that may affect the yield of copolymer
and the grafting ratio. It can be found in Fig. 3 that when
the dosage of AM was no more than 6g, the grafting
ratio of EH-lignin/AM copolymer increased quickly
12345
10
15
20
25
30
35
40
10
15
20
25
30
35
40
Grafting ratio (%)
Yield (%)
Mass fraction of initiator/EH-lignin (%)
Y(%)
GR(%)
Figure 2. Effect of initiator dosage on the yield of copoly-
mer and the grafting ratio.

20 R. Fang et al. / Natural Science 1 (2009) 17-22
024681012
10
20
30
40
50
60
70
80
10
20
30
40
50
60
70
80
Grafting ratio (%)
Yield (%)
Y(%)
GR(%)
Dosage of AM (g)
Figure 3. Effect of AM dosage on the yield of copolymer and
the grafting ratio (the dosage of EH-lignin is 2g).
as the mass ratio of AM to EH-lignin increased. This is
because the higher AM concentration can make it easier
for EH-lignin to come into contact with monomer and
then speed up graft copolymerization and improve
grafting ratio. However, when the concentration of the
AM is higher than a certain level, the probability of the
homopolymerization of AM will increase rapidly. This
reaction, which leads to the formation of PAM, will
compete with graft copolymerization and diminish the
grafting ratio. The yield of copolymer, on the contrary,
decreased gradually as the dosage of AM increased from
1g to 12g. This phenomenon can be ascribed to the rising
water-solubility of the copolymer. It has been mentioned
above that higher monomer concentration will enhance
the probability of homopolymerization of AM and extend
the length of some PAM chains that have been grafted
onto EH-lignin. Thus, the water-solubility of EH-lignin/
AM copolymer increases with the rise of AM dosage and
the quantity of copolymer that can be separated from
aqueous solution declines simultaneously. We can see
from the above analysis that the ideal dosage of acryla-
mide is 4~6g or the mass ratio of AM to EH-lignin is 2~3.
3.1.4. Effects of Reaction Time and Reaction
Temperature on Graft Copolymerization
The effects of reaction time and reaction temperature on
the yield of copolymer and the grafting ratio were shown
in Fig. 4 and Fig. 5 respectively. From Fig. 4, we could
see the yield and grafting ratio improved rapidly as the
reaction time increased from 2h to 4h. When the reaction
time was further prolonged, the growth of yield and
grafting ratio became unremarkable. This phenomenon
is similar to the regular pattern of radical polymerization.
The graft copolymerization took place mostly in the pe-
riod of initiation and the speed of the copolymerization
was high at first. However, as the concentration of
acrylamide and initiators declined with elapsed time, the
graft copolymerization slowed down and the yield and
grafting ratio stopped growing.
23456
15
20
25
30
35
40
15
20
25
30
35
40
Yield (%)
Grafting ratio (%)
Y(%)
GR(%)
reaction time (h)
Figure 4. Effect of reaction time on graft copolymerization.
30 40 50 60 70
15
20
25
30
35
40
15
20
25
30
35
40
Grafting ratio (%)
Yield (%)
Y(%)
GR(%)
Reaction temperature (℃)
Figure 5. The influence of reaction temperature on graft co-
polymerization.
The influence of different reaction temperature on
graft copolymerization was shown in Fig. 5. The highest
yield and grafting ratio can both be reached at 50 and ℃
this is the ideal reaction temperature for the bi-
nary-initiating system of K2S2O8-Na2S2O3. It is well
known that low reaction temperature will postpone the
decomposition of initiator and restrain the copolymeri-
zation. On the contrary, when the reaction temperature
was higher than 50, the decomposition of initiator ℃
would be too fast and the possibility of radical transfer
would be greatly enhanced, both of which will lead to
the decrease of yield and grafting ratio.
3.2. FT-IR Spectral Analysis
The FT-IR spectra of EH-lignin and EH-lignin/AM co-
polymer were shown in Fig. 6. It can be seen from the
FT-IR spectrum of EH-lignin/AM copolymer, compared
with pure EH-lignin, the relative intensity of the band at
about 1700 cm-1 increased significantly. This adsorption
peak is assigned to the vibration absorbance of C=O and
the rise of its intensity implies that AM have been
grafted onto EH-lignin. Furthermore, the intensity of the

R. Fang et al. / Natural Science 1 (2009) 17-22 21
4000 3500 3000 2500 2000 15001000
Wavenu m ber / cm -1
a
b
Figure 6. FT-IR spectra of EH-lignin and EH-lignin/ AM co-
polymer (a EH-lignin; b EH-lignin/AM copolymer).
band at 1020 cm-1, which is assigned to the absorbance of
N-H, also increased and this is another proof of the suc-
cessful graft copolymerization. All the information pro-
vided by FT-IR analysis had indicated that the product is an
EH-lignin/AM copolymer with numerous functional groups,
such as phenolic hydroxyl (3400cm-1), carbonyl (1700cm-1)
and amide (1550 and 1020cm-1) groups [11,12].
3.3. Adsorption and Decoloration of Dye-
Wastewater by EH-lignin/AM Copolymer
The adsorption and decoloration of AR 274 and RR X-
3B dye wastewater by EH-lignin/AM grafted copolymer
and pure EH-lignin was investigated according to the
procedures described in 2.3. The effect of grafting ratio
on the adsorption capacity of graft copolymer was shown
in Fig. 7. It was found that the absorption capacity of
EH-lignin/AM copolymer to both dye wastewaters in-
creased remarkably with the rising of grafting ratio. When
the grafting ratio was 43.6 %, the maximum adsorption
amount of AR 274 and RR X-3B by graft copolymer may
reach 751mg/g and 512mg/g respectively.
0 10203040
3
4
5
6
7
8
Adsorption amount (102mg/g)
Grafting ratio (%)
AR 274
RR X-3B
Figure 7. Effect of grafting ratio on the adsorption capacity of
EH-lignin/AM copolymer.
The flocculation of dye colloids result from various
mechanisms, including electrostatic attraction, sorption
(related to protonated amine groups and phenolic hy-
droxyl), and bridging (related to the high molecular
weight of the polymer) [13]. There are sulfonic, carbonyl
and amino groups in AR 274 and RR X-3B. The sulfonic
groups can be electrostatic attracted by protonated amide
groups of the EH-lignin/AM graft copolymer and the
amino groups can from hydrogen bonding with phenolic
hydroxyl groups of the copolymer and EH-lignin. Thus,
higher grafting ratio will help to strengthen the su-
pramolecular interaction between dyes and graft co-
polymers, which can bind copolymer molecules closer,
trap and flocculate dyes more effectively.
The relationship between the dosage of coagulants
and the decoloration rate of dye wastewater was shown
in Fig. 8. The concentration of AR 274 dye wastewater
was 500mg/L and the grafting ratio of EH-lignin/AM
copolymer was 30.8%. It can be seen in Fig. 8 that the
decoloration rate of dye wastewater increased rapidly as
the dosage of both flocculants increased from 50mg/L to
200mg/L. When the dosage of flocculant reached
800mg/L, the AR 274 dye had almost been removed
completely.
Results also showed that both pure EH-lignin and EH-
lignin/AM graft copolymer performed well in the re-
moval of AR 274 from dye wastewater when the dosage
of flocculant is higher than 200mg/L. As has been men-
tioned in 3.2, EH-lignin and lignin/AM copolymer pos-
sess lots of functional groups, such as phenolic hydroxyl,
carbonyl and amide groups. Meanwhile, AR 274 also
contains alcoholic hydroxyl and carbonyl groups, which
leads to the adsorption of dye molecules on EH-lignin
and EH-lignin/AM copolymer through hydrogen bond-
ing. Subsequently, the flocculant molecules bridge to
each other under acidic environment and form large
flocs, which will trap the dyes dissolved in wastewater
and then precipitate simultaneously.
0100 200 300400 500 600 700 800 900
0
10
20
30
40
50
60
70
80
90
100
Decoloration rate (%)
Dosage of coagulant (mg/L)
EH-lignin
Graft copolymer
Figure 8. Effect of coagulant dosage on the decoloration rate
of AR 274 wastewater.

22 R. Fang et al. / Natural Science 1 (2009) 17-22
0100200 300 400 500 600700 800 900
10
20
30
40
50
60
70
80
90
100
Dosage of coagulant (mg/L)
EH-lignin
Graft copolymer
TOC removal rate of dye wastewater(%)
Figure 9. TOC removal rate of dye wastewater after flocculation.
The residue of EH-lignin and EH-lignin/AM copoly-
mer in the supernatant after flocculation was measured
by TOC test to evaluate the potential secondary pollution
of this flocculant. These experiments were carried out
with various dosage of flocculant at pH=4. It can be
found in Fig. 9 that, compared with the decoloration rate
of AR 274 wastewater flocculated by both flocculants,
the TOC removal rate of these samples are much lower.
This phenomenon implies that part of EH-lignin and
EH-lignin/AM copolymer remains in the supernatant
after flocculation. Furthermore, TOC removal rate of the
dye wastewater treated by EH-lignin/AM graft copoly-
mer is much higher than that of pure EH-lignin, which
means the graft copolymerization of EH-lignin with AM
will help to minimize the residue amount of this co-
polymer in wastewater and diminish the potential sec-
ondary pollution.
4. CONCLUSIONS
The preparation and the application of EH-lignin/AM
graft copolymer were presented in this article. The op-
timum synthesis conditions were discussed and the func-
tional groups of the copolymer were characterized by
FT-IR The adsorption and decoloration of two azo-dye
wastewater by EH-lignin/AM grafted copolymer and
pure EH-lignin was investigated.
1) The optimum conditions for the graft copolymeri-
zation of EH-lignin with acrylamide were as follows:
initiator K2S2O8-Na2S2O3 with a quantity 3 wt% of EH-
lignin, mass ratio of AM to EH-lignin was 2~3, reaction
time 4h and temperature at 50.℃
2) FT-IR spectrum of EH-lignin/AM copolymer indi-
cated that acrylamide had been grafted onto EH-lignin
successfully and the copolymer had numerous functional
groups, such as phenolic hydroxyl, carbonyl and amide
groups.
3) The absorption capacity of EH-lignin/AM copoly-
mer to dye wastewater was enhanced with the increase
of grafting ratio. Both pure EH-lignin and EH-lignin/AM
graft copolymer performed well in the removal of AR
274 from dye wastewater when the dosage of flocculant
is higher than 200mg/L. However, the residue concen-
tration of EH-lignin/AM graft copolymer remained in
the supernatant after flocculation was much lower than
that of pure EH-lignin.
ACKNOWLEDGEMENTS
The author thanks Key Laboratory of Cellulose and Lignocellulosics
Chemistry, Guangzhou Institute of Chemistry, Chinese Academy of
Sciences for financial support (LCLC-2004-158).
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