Branching of hemicelluloses through an azetidinium salt ring-opening reaction

Article abstract of DOI:10.1016/j.carres.2016.04.005

During the last century there has been a steady increase in the number of publications on practical applications of hemicellulose. Due to the water and moisture sensitivity, poor film-forming ability and lack of thermal processability most of the hemicelluloses need to be chemically modified prior to processing into materials. Within this study we present the results of azetidinium salts as a new functional group for conjugation to polysaccharides. The reactivity of three azetidinium salts on xylan, arabinoxylan and galactoglucomannan was investigated. Carbonyl groups were found to be favorable for the reaction with azetidinium salts and thus the glucuronic acid content in the hemicellulose determines the degree of substitution. TEMPO-oxidation of the hemicelluloses was done which successfully increased the degree of substitution. The highly reactive azetidinium salts are easily synthesized from secondary amines and epichlorohydrin and can be used as a new tool toward functionalization of hemicelluloses into the after sought properties.

Full text of DOI:10.1016/j.carres.2016.04.005

Carbohydrate Research 428 (2016) 23–30
Contents lists available at ScienceDirect
Carbohydrate Research
journal homepage: www.elsevier.com/locate/carres
Branching of hemicelluloses through an azetidinium salt
ring-opening reaction
Mikaela Börjesson a, Gunnar Westman a,b,
*
a Department of Chemistry and Chemical Engineering, Chalmers University of Technology, Kemivägen 10, SE-41296 Gothenburg, Sweden
b Wallenberg Wood Science Center (WWSC), Chalmers University of Technology, Kemivägen 10, SE-41296 Gothenburg, Sweden
A R T I C L E
I N F O
A B S T R A C T
Article history:
Received 19 January 2016
Received in revised form 4 April 2016
Accepted 6 April 2016
Available online 11 April 2016
During the last century there has been a steady increase in the number of publications on practical ap-
plications of hemicellulose. Due to the water and moisture sensitivity, poor film-forming ability and lack
of thermal processability most of the hemicelluloses need to be chemically modified prior to process-
ing into materials. Within this study we present the results of azetidinium salts as a new functional group
for conjugation to polysaccharides. The reactivity of three azetidinium salts on xylan, arabinoxylan and
galactoglucomannan was investigated. Carbonyl groups were found to be favorable for the reaction with
azetidinium salts and thus the glucuronic acid content in the hemicellulose determines the degree of
substitution. TEMPO-oxidation of the hemicelluloses was done which successfully increased the degree
of substitution. The highly reactive azetidinium salts are easily synthesized from secondary amines and
epichlorohydrin and can be used as a new tool toward functionalization of hemicelluloses into the after
sought properties.
Keywords:
Hemicellulose
Branching
Azetidinium
Ring-opening
TEMPO-oxidation
Glucuronic acid
© 2016 Elsevier Ltd. All rights reserved.
1. Introduction
hemicelluloses depend on their carbohydrate composition, the degree
of polymerization (DP), its branching pattern as well as the chem-
Today the society has become increasingly aware of the limited
resource of fossil-fuels and the importance of developing materi-
als from renewable resources with a low environmental impact. The
demand for replacing non-renewable products with bio-based prod-
ucts requires knowledge and tools on how to utilize the resources
in more efficient ways. Key factors for a shift to hemicellulose-
based materials are availability in large quantities at a reasonable
price and that the material manufactured from hemicellulose has
similar or better properties than the fossil fuel-based material that
it will replace. A big difference between fossil fuel-based poly-
mers and biopolymers is the higher content of oxygen in bio-
based materials. A consequence of the higher oxygen content is that
the material has lower heat value, 13.6 MJ/kg; on the other hand
it has the advantage that the reactive oxygen containing groups such
as carboxylic acids and alcohol functions can be functionalized to
tune the properties of the material.1 During the 21st century there
has been a huge increase in the number of publications on practi-
cal applications of hemicelluloses where the main applications have
been toward hydrophobized materials such as gels, films and re-
inforcement in composites.2,3 The overall properties of the
ical structure of added substituents.
Hemicelluloses represent approx. 20–30% of the biomass in nature
and are in general shorter than cellulose polymers. The main hemi-
celluloses found in hardwood are xylans with a DP of 100–400.
Xylans consist of a backbone of D-xylopyranose units and are mostly
branched with L-arabinofuranose and glucuronic acids and the
degree of branching depends on plant species and cell type. The main
hemicelluloses found in softwoods are glucomannans, which consist
of D-glucopyranose and D-mannopyranose units that are often
branched with D-galactopyranose units. The glucomannans are partly
acetylated and is shorter than xylans with a DP of ca 100–200. The
hemicelluloses found in annual plants such as crops are generally
more structurally diverse and complex than the hemicelluloses found
in wood. The hemicelluloses in annual plants often consist of mix-
tures of many different sugar units and can be branched with for
example D-xylopyranose, L-arabinofuranose, D-galactopyranose or
uronic acid side chains.4,5
Hemicellulose can be utilized from sources like low-value waste-
water streams in kraft pulping processes and agricultural crop
residues. As said, the type and composition of hemicelluloses are
highly dependent on the source, but also the isolation process and
chemical reactions during the process can give rise to deacetylation,
debranching and/or degradation. As a consequence the hemicellu-
loses structure depends on its process and fractionation history.
Today, isolation of hemicelluloses such as hardwood xylan from the
black liquor in kraft pulping processes and galactoglucomannans
*
Corresponding author. Department of Chemistry and Chemical Engineering,
Chalmers University of Technology, Kemivägen 10, SE-41296 Gothenburg, Sweden.
Tel: +46 (0) 31 772 3072.
E-mail address: westman@chalmers.se (G. Westman).
http://dx.doi.org/10.1016/j.carres.2016.04.005
0008-6215/© 2016 Elsevier Ltd. All rights reserved.

24
M. Börjesson, G. Westman /Carbohydrate Research 428 (2016) 23–30
Table 1
(GGMs) released and accumulated in process waters in the
production of thermo-mechanical pulp is often done by mem-
brane filtration.6,7 There is also a number of methods available to
obtain hemicellulose from different plant sources like extraction with
alkali,8 dimethyl sulfoxide,9 hot-water treatment,10 microwave
treatment11 or extrusion.12
Relative carbohydrate compositions (Rel %) in the different hemicellulose samples
assuming the samples only consist of detected sugar units. The glucuronic acid (GlcA)
content was determined by NMR analysis
Sample
Araa
Gala
Glca
Xyla
Mana
GlcAb
Xylan
NaOH-xyl
AX
NaOH-AX
TEMPO-AX
GGM
NaOH-GGM
TEMPO-GGM
0.7
1.0
20.9
19.2
9.1
2.1
2.1
2.0
0.3
1.4
1.1
1.7
1.4
14.3
10.9
16.3
3.1
3.3
7.5
10.2
5.8
16.9
16.4
16.9
82.5
81.0
70.5
68.7
83.5
0.1
0.0
0.1
0.0
0.1
0.2
66.6
70.0
64.2
13.4
13.2
–
–
–
–
–
–
Although, being such a close relative to cellulose much less de-
velopment in chemical modification of hemicellulose has been done
compared with cellulose. Traditional modification processes are
etherification, esterification, graft polymerization and oxidation
which are quite often used as an intermediate for further
functionalization. These processes are usually used with the aim to
increase the hydrophobicity of the material. Successful acylation of
hemicelluloses to increase the hydrophobicity has been reported
by several groups.13–19 There are also reports on fluorination of hemi-
celluloses, however thus far; it seems that the water sensitivity of
the trifluoroacetyl group limits the use of these derivatives.20,21
Hemicelluloses can find use within the packaging industry,22,23
food industry,24 biomedical uses25 and for chemical production.26,27
Cross-linked hemicelluloses have also been reported to form hydrogels
and are evaluated for drug delivery systems applications.28,29
We have looked into azetidinium salts as new coupling re-
agents that can be used for functionalization of hemicelluloses.
Azetidines are found in a diverse range of natural products and serve
as a valuable building block for other structural classes. The four
member azetidine-ring can be quarternized to an azetidinium ion
and in combination with the ring strain, it is a suitable reagent due
to its high reactivity.30 Azetidinium ions can easily be synthesized
from secondary amines and epichlorohydrin.31,32 Because of the many
choices of carbon chains or other functional groups that could be
attached to the azetidine-nitrogen, there is a big variety of modi-
fications that can be done. Akzo Nobel Chemicals recently filed a
patent for production of azetidinium compounds with carbon chains
between 1 and 30 carbon atoms.33 A difference compared to tradi-
tional hydrophobization agents from using azetidiniums is that two
substituents in a Y-shaped branching will be formed when the
azetidinium ring is opened and covalently linked to the target.
Hydrophobization and properties tuning in this way will be a com-
plement to the ones formed by ether and ester conjugation to the
polysaccharide. Functionalization with Y-shaped chlorohydrins has
recently been investigated showing that longer alkyl chains at-
tached to CNC increase hydrophobicity and enhance the dispersion
in low-density polyethylene matrices.34
There are only a few reports on azetidinium ions in the field of
polysaccharides. Most of them are found in polyamide amine epi-
chlorohydrin (PAE), which is a water-soluble resins used as wet
strength additive for preparing wet strength papers. In a review
article by Espy35 potential cross-linking reactions of azetidinium
groups are discussed as a possible wet strength agent. In the review
it is stated that alcohol groups may play a part in cross-linking re-
actions as well as carboxylate end groups of cellulosic polymers.
Carboxymetylated cellulose (CMC) or other carboxyl-bearing dry
strength polymers showed an increased effectiveness in reaction with
PAE.36 Also Obokata and Isogai37 found in their study on PAE that
TEMPO-oxidized cellulose under alkaline conditions showed an in-
creased reactivity toward PAE resulting in increased wet strength
in the cellulose sheets.
Branching of hemicelluloses through azetidinium salts opens up
the possibility to attach different functional groups or length of carbon
chains and by that gives new properties to the material depending
on how the branching is designed. In this study, three different and
relatively small azetidinium salts with a 3-hydroxyazetidin-1-ium chlo-
ride structure have been synthesized and used in reaction with the
three different hemicelluloses—xylan, arabinoxylan (AX) and
galactoglucomannan (GGM)—to evaluate the branching effect on the
different hemicellulose structures. AX and GGM were also TEMPO-
0.5
0.7
a
Content detected by ICS.
Content detected by NMR.
b
oxidized to investigate the reactivity of the azetidinium salts. TEMPO-
oxidations are known to oxidize primary alcohol functionalities of
several polysaccharides. Recently, it has been reported that TEMPO-
mediated oxidation is an efficient method for the oxidation of GGM.38,39
2. Experimental procedure
2.1. Materials
Three hemicelluloses were used in the study; a commercial
hardwood-xylan (beech wood, Sigma-Aldrich), arabinoxylan (AX)
extracted from barley husk (Lyckeby Culinar, Sweden) following the
procedure described by Höije et al.40 and galactoglucomannan (GGM)
from spruce kindly received as a gift from Stora Enso (Sweden). The
hemicelluloses have been analyzed with carbohydrate analysis and
NMR to determine the sugar content composition in the samples
(Table 1). All chemicals used for the synthesis of azetidinium salts
(diethylamine, dihexylamine, morpholine and epichlorohydrin) or
treatments of the hemicelluloses with NaOH-activation or TEMPO-
oxidation (2,2,6,6-tetramethyl-1-piperidinyloxy (TEMPO) and
bis(acetoxy)iodobenzene (BAIB) as co-oxidant) were purchased from
Sigma-Aldrich and used as received.
2.2. Preparation of N,N-dialkylazetidinium salts
Preparation of the different azetidinium (Az) salts following pre-
viously known synthesis.32,41–44 A general reaction scheme is shown
in Fig. 1.
The general procedure for preparation of azetidinium salts was
dropwise addition of 0.1 mol epichlorohydrin to a solution of 0.1 mol
secondary amine and solvent. The temperature was kept below 5 °C
during the addition of epichlorohydrin due to an exothermic reac-
tion. The reaction mixture was stirred in an ice bath for 1 h and
additional 47 h at room temperature. The solution was purified with
diethyl ether and water and solvent was removed in vacuum. Fig. 2
shows the molecular structure of the different azetidinium salts used
in this study. For more details see supporting information.
2.3. Pre-treatment of hemicellulose
A pre-treatment of the hemicelluloses was performed before re-
action with the azetidinium salts, where a 17.5 wt% NaOH-solution
Fig. 1. A secondary amine is converted to azetidinium chloride through reaction with
epichlorohydrin.

M. Börjesson, G. Westman /Carbohydrate Research 428 (2016) 23–30
25
Fig. 2. The molecular structure of the azetidinium salts used in the study. I: 1,1-diethyl-3-hydroxyazetidin-1-ium chloride prepared from diethylamine, II: 1,1-dihexyl-3-
hydroxyazetidin-1-ium chloride prepared from dihexylamine and, III: N-morpholino-3-hydroxyazetidin-1-ium chloride prepared from morpholine.
was used to activate the hydroxyl groups in order to increase the
reactivity of the polysaccharides.45–47 Ten grams of hemicellulose
was suspended in 35 mL deionized water and heated under reflux
for 15 min before cooled down to room temperature and 10 mL 17.5
wt% NaOH-solution was added to the mixture. After 60 min di-
methyl ether was added to obtain a heterogeneous mixture and the
NaOH-hemicellulose was filtrated and washed with ethanol.
A TEMPO-oxidation was performed on AX and GGM to intro-
duce carbonyl groups in the polymers following a procedure
described by Nylander.48 Dried hemicellulose (1 g) was dissolved
in 6.45 mL distilled water and 19.35 mL acetonitrile was added giving
a two-phase solution. The solution was cooled on ice and 3.48 g BAIB
(10.8 mmol) and 0.12 g (0.768 mmol) TEMPO was added and the so-
lution was magnetically stirred for 2 h at 0 °C and additional 4 h at
room temperature. The mixture was thereafter precipitated in cold
ethanol followed by washing and filtration with ethanol.
D2O. The glucuronic acid content in hemicellulose was analyzed with
NMR according to Westbye et al.50
Fourier-Transform Infrared (FTIR) spectroscopy was recorded on
a PerkinElmer Spectrum One instrument using the potassium
bromide (KBr) pellet technique with 10 wt% sample. FTIR was ana-
lyzed between 4000–400 cm−1 and 32 scans collected.
Neutral carbohydrate composition was analyzed on acid hydro-
lyzed hemicellulose samples with a high performance anion
exchange chromatography with pulsed amperometric detection
(HPAEC-PAD), using an ion chromatography system ICS 3000
(Dionex) equipped with a gradient pump, isocratic post column
pump, column oven and autosampler. The different sugars de-
tected with the available system were L-arabinose (Ara), D-galactose
(Gal), D-glucose (Glc), D-xylose (Xyl) and D-mannose (Man).
The degrees of substitution (DS) were determined from the
amount of nitrogen in the samples measured by elemental analy-
sis at Mikrolab Kolbe in Germany. The DS according to the number
of nitrogen (DSN) were calculated using Eq. (1)
2.4. Modification of hemicellulose with azetidinium salts
(1)
DS = M
×%N 1401−M
×%N
)
Az-reagent
(
)
(
N
(
Average hemicellulose unit
)
Since the azetidinium salts as well as the hemicelluloses used
in the study have slightly different solubility a general method that
works well for all hemicelluloses and azetidinium salts used in this
study was developed. It was found that a general process as follows
gave the best yields. Five grams of hemicellulose, 2–3 mol eq.
azetidinium salt depending on hemicellulose (1 mol equiv./–OH func-
tionality) and a 50 g solution of DMSO/toluene (90:10) were added
to a round bottom flask. A condenser was attached to the flask and
the solution was stirred and kept at 90 °C for 22 h. The mixture was
allowed to cool to room temperature before the modified hemi-
cellulose dropwise was precipitated in cold ethanol during stirring.
The modified hemicellulose was filtrated, washed with ethanol, re-
dissolved in deionized water and precipitated again in ethanol
followed by filtration and washing several times with ethanol before
drying at room temperature.
where M(Average hemicellulose unit) and MAz-reagent are the molecular weights
of the average anhydrous sugar units in the hemicellulose polymer
and the azetidinium reagents respectively, and %N is the amount
of nitrogen in percent measured by elemental analysis. The
maximum DS for the different hemicelluloses are dependent on the
available reaction sites per sugar unit which were estimated from
the carbohydrate analysis presented in Table 1.
Thermogravimetric analysis (TGA) was used to detect thermal
changes in the hemicellulose samples. A TGA/DSC 3+ Star System
(Mettler Toledo) was used where 10 mg sample was heated from
25 °C to 500 °C under N2 atmosphere with a heating rate of 5 °C/
min. The extrapolated onset temperature and the inflection point
calculated from the 1st derivate were measured using the STARe
Excellence Software.
Fig. 3 shows suggested reaction schemes for the three hemicel-
luloses where the hemicelluloses, AX and GGM have been TEMPO-
oxidized before reaction with azetidinium salts.
3. Results and discussion
3.1. Azetidinium salts
2.5. Hydrolysis of hemicellulose
The azetidinium chlorides prepared from secondary amines and
epichlorohydrin were obtained in yields above 70%. The salts were
analyzed with 1H NMR and 13C NMR in DMSO-d6. The synthesis and
NMR-analysis of the different azetidinium chlorides are presented
in detail in supporting information.
The modified hemicelluloses were hydrolyzed to its monosac-
charides by a sulfuric acid hydrolysis.49 Three milliliters of 72% H2SO4
were added to 200 mg dried sample and put under vacuum for
15 min before placed in a water bath at 30 °C for 1 h. Eighty-four
grams of distilled water was added and the samples were put in
an autoclave for 1 h at 125 °C and 20 bar. To analyze the carbohy-
drate content the hydrolyzed samples were filtered and diluted with
distilled water to a concentration of 200 mg/L.
3.2. Carbohydrate content in hemicellulose
The sugar composition in the hydrolyzed hemicellulose samples
was determined from a neutral carbohydrate analysis and the rel-
ative percent (Rel %) of the sugars was calculated assuming only the
detected sugars were present (Table 1).
2.6. Characterization
1H NMR and 13C NMR were recorded on a Varian MR-400
(400 MHz) spectrometer. DMSO-d6 was used as solvent for the
azetidinium salts and the hemicellulose samples were dissolved in
The glucuronic acid content in beech xylan was measured by 1H
giving approximately 13% glucuronic acids. The glucuronic acid

26
M. Börjesson, G. Westman /Carbohydrate Research 428 (2016) 23–30
Fig. 3. Suggested reaction schemes for the reactions between (a) xylan and Az-salt, (b) AX and Az-salt after a TEMPO-oxidation, and (c) GGM and Az-salt after a TEMPO-oxidation.
content in AX and GGM was too low to be detected by NMR and
corresponds to literature, AX and GGM should not contain any or
just a very small amount of uronic acids.51,52 Different pre-treatments
for hemicelluloses and how it affects the carbohydrate composi-
tion are listed in Table 1. The NaOH-activation did not change the
carbohydrate composition for xylan and AX significantly but de-
graded the α-glycosidic linkage between the galactose side chain
and the glucose–mannose backbone in GGM, which is known to be
degraded during oxidation or alkali treatments of the polymer.53
A TEMPO-oxidation under neutral conditions was performed to
introduce carboxylate groups in the AX and GGM polymers without
degrading the polymers too much. A traditional TEMPO-oxidation
process was also performed following published procedures38,39 but
the oxidation of the hemicelluloses was thwarted by unknown
reasons. However, TEMPO-oxidation with BAIB as co-oxidant gave
successful oxidations of the hemicellulose materials. Table 2 shows
the concentrations (mg/L) of the neutral carbohydrate analysis of
TEMPO-oxidized AX and GGM. For the AX samples mainly the arabi-
nose groups are changed due to the oxidation of the primary
hydroxyl group, present only in the arabinose units. For the GGM
samples the largest reduction in detected sugars is seen in the
glucose and mannose sugars and only a small change in the de-
tected concentrations of galactose. This means that the oxidation
primary occurs in the glucose–mannose backbone of GGM. The arabi-
nose concentration for TEMPO-oxidized AX is reduced by 63%
meaning that 63% of the arabinose groups are either oxidized or de-
graded whereas the number of reduced sugars involved in the
TEMPO-oxidation of GGM is 17%.
3.3. FTIR analysis of hemicelluloses
FTIR analysis on hardwood xylan (Fig. 4a) shows a broad ab-
sorption band at 3600–3100 cm−1 and at 2925 cm−1 showing the
hydroxyl groups and CH2 stretching in the xylose ring respective-
ly. A small absorption band at 1711 cm−1 corresponds to carboxylic
groups, which indicates the presence of glucuronic acids in the
sample. The absorption band at 1044 cm−1 is typical ring vibra-
tions and stretching vibrations of C–OH side groups.54 The band at
1167 cm−1 represents the glycosidic bonds (C–O–C) and the absorp-
tion band at 896 cm−1 is typical for β-glycosidic linkage. Other
Table 2
The concentrations (mg/L) of the detected sugars in the carbohydrate analysis before
and after TEMPO-oxidation of AX and GGM
Sample
Ara
Gal
1.5
1.6
18.8
17.8
Glc
Xyl
99.1
100.4
0.1
Man
Total residue (mg/L)
AX
TEMPO-AX
GGM
29.4
11.0
2.8
10.5
7.0
22.2
18.5
0.0
0.2
87.4
70.1
140.6
120.2
131.3
109.3
absorption bands detected are 1614 (absorbed water) and 984
TEMPO-GGM
2.1
0.8
55
(C–O) cm−1
.
FTIR analysis on azetidinium salt branched xylan

M. Börjesson, G. Westman /Carbohydrate Research 428 (2016) 23–30
27
Fig. 4. FTIR spectrums of (a) xylan, (b) Az-DEA modified xylan, (c) Az-DHA modified xylan, and (d) Az-morph modified xylan.
(Fig. 4b–d) shows a new absorption band at 1745 cm−1 correspond-
ing to carbonyl groups (C O) such as saturated esters. Due to the
around 1740 cm−1 due to the presence of both acetylated groups and
–COO- groups caused by the oxidation. The broadening can also be
observed in the Az-DEA modified oxidized sample in Fig. 6d. Sug-
gested reaction schemes for the reaction between TEMPO-oxidized
GGM and azetidinium containing groups are shown in Fig. 3c.
=
formation of an ester the branching reaction between azetidinium
salts and xylan occurs on the carboxylic acid group present in the
glucuronic acids. According to literature,35–37 the azetidiniums con-
taining wet strength resin, PAE, are more prone to react with –COO-
groups compared with secondary –OH groups, which further support
that the reaction takes place on the glucuronic acids in the xylan
samples. Changes in FTIR spectra can also be seen as a narrowing in
the area between 3600 and 3100 indicating structural changes of
the –OH groups. Suggested reaction schemes for the reaction between
xylan and azetidinium containing groups are shown in Fig. 3a.
AX and GGM do not have any carbonyl groups available and as
a consequence FTIR analysis on modified AX and GGM samples does
not show any changes in the carbonyl group absorption band area
(1760–1665 cm−1). For both AX and GGM, primary alcohols are
present on the side groups additional to this GGM also has primary
hydroxyl groups in the backbone. The primary –OH groups are se-
lective for a TEMPO-oxidation and introduction of carbonyl groups
to AX and GGM can increase the reaction efficiency. Figs. 5 and 6
show the TEMPO-oxidation of AX and GGM respectively and the re-
action between hemicellulose and Az-DEA salt.
In Fig. 5 it is clearly seen that the TEMPO-oxidation has intro-
duced carbonyl groups to the polymer due to the new absorption
band at 1740 cm−1. The azetidinium reaction between TEMPO-
oxidized AX and Az-DEA salt is shown in Fig. 5d and the absorption
band at 1740 cm−1 is shifted to 1744 cm−1 indicating structural
changes of the carbonyl groups due to formation of an ester bonding.
Fig. 5b shows the azetidinium reaction for non-oxidized AX where
no carbonyl groups are present. Suggested reaction schemes for the
reaction between TEMPO-oxidized AX and azetidinium contain-
ing groups are shown in Fig. 3b.
3.4. Degree of substitution
The amounts of nitrogen in the samples were measured with el-
emental analysis and used to calculate the degree of substitutions
(DSN) using Eq. (1). The results are presented in Table 3 and the car-
bohydrate composition has been used to calculate the maximum
DS. Since carboxylate groups are favorable prior to hydroxyl groups
the maximum DS, regarding the number of carboxylic acids avail-
able per sugar unit, is determined to 0.13 for xylan. AX and GGM
did not have any measurable amounts of carboxylic acid groups
present and the reaction efficiency is calculated from the number
of primary alcohols available per sugar unit (for un-oxidized
samples), giving a maximum DS of 0.3 for AX and 1.0 for GGM.
DS is highest for the xylan samples due to the glucuronic acids
present in the polymer chain whereas both AX and GGM samples
have too low glucuronic acid content to be detected. Even though
GGM has a higher amount of primary –OH groups with one primary
–OH group on every sugar unit compared with AX, which only has
one primary –OH on the branched arabinose groups, the DS is lowest
for the GGM samples. Thus, the primary –OH groups do not seem
to have any prominent role in the reaction but as seen in Table 3,
some of the –OH groups are substituted in AX and GGM samples.
Primary –OH groups are on the other hand favorable in oxidation
reactions and the introduction of carbonyl groups could make it pos-
sible to control the AX and GGM modifications.
From Table 3 it is also noted that the smallest azetidinium com-
pound, Az-DEA, gave the highest substitution for all hemicelluloses
and the more bulky ones gave lower DS. For reagents with larger
functional groups sterical effects could influence the reaction
efficiency.
The native GGM is partly acetylated which is shown as a dis-
tinct peak at 1736 cm−1 in Fig. 6a and will be present as long as the
sample is not treated with alkali such as during NaOH-activation.5,56,57
The TEMPO-oxidation introduces a broader absorption band area

28
M. Börjesson, G. Westman /Carbohydrate Research 428 (2016) 23–30
Fig. 5. FTIR spectrums of (a) arabinoxylan (AX), (b) Az-DEA modified AX, (c) TEMPO-oxidized AX and, (d) Az-DEA modified TEMPO-oxidized AX.
3.5. Effects of different pre-treatments of hemicellulose
polymer and for the actual amount of oxidized groups in the poly-
mers, shown as the numbers in brackets in Table 4.
As stated before, the reactions with azetidinium groups are fa-
vorable to carbonyl groups and the introduction of carbonyl groups
through TEMPO-oxidation is therefore expected to increase the re-
action efficiency, compared with the DS values present in Table 3.
Table 4 summarizes the different pre-treatments that were done on
the different hemicelluloses which further were reacted with Az-
DEA salt for comparison of the reaction efficiency. DS was calculated
from the amount of nitrogen found in the elemental analysis using
Eq. (1). The TEMPO-oxidation is selective to primary alcohols and
Table 4 shows both the reaction efficiency for a fully oxidized
The 17.5 wt% NaOH-activation increased the reaction efficien-
cy for xylan and AX but did not have an evident effect on GGM,
probably due to the alkaline degradation of the galactose units. On
the other hand, the introduction of carbonyl groups through TEMPO-
oxidation had a significant influence on the reaction efficiency for
both AX and GGM, which further proves the favorable of carbonyl
groups prior hydroxyl groups.
Xylan samples were almost fully substituted while AX and GGM
had a much lower substitution degree. Reaction efficiencies in Table 4
are calculated for both fully oxidized samples and the actual amount
Fig. 6. FTIR spectrums of (a) partly acetylated GGM, (b) Az-DEA modified GGM, (c) TEMPO-oxidized GGM, and (d) Az-DEA modified TEMPO-oxidized GGM.

M. Börjesson, G. Westman /Carbohydrate Research 428 (2016) 23–30
29
Table 3
The ability to control DS is favorable to understand the reac-
tion and an optimization regarding concentration of azetidinium
salts might have increased DS further for all samples, which was
the case for the etherification with EPTMAC described above, where
the excess of 10 mol equiv. EPTMAC/anhydroxylose unit was nec-
essary for a DS of 1.64. On the other hand, the Y-shaped azetidinium
reagent contributes with two alkyl chains per reaction target and
a lower DS could have a higher impact on the material properties.
Degree of substitution (DS) for the different Az-modification reactions of the three
hemicelluloses: xylan, AX and GGM. The reaction efficiencies are calculated from
the maximum DS of available reaction targets in the hemicelluloses
Sample
Pre-treatment
%N
DSN
Reaction efficiency (%) a
Az-DEA-xyl
Az-DHA-xyl
Az-morph-xyl
Az-DEA-AX
Az-DHA-AX
Az-morph-AX
Az-DEA-GGM
Az-DHA-GGM
Az-morph-GGM
NaOH-act.
NaOH-act.
NaOH-act.
NaOH-act.
NaOH-act.
NaOH-act.
NaOH-act.
NaOH-act.
NaOH-act.
1.10
0.82
0.99
0.58
0.44
0.36
0.46
0.08
0.31
0.13
0.10
0.11
0.06
0.05
0.04
0.06
0.01
0.04
96.85
75.01
86.85
19.47
15.07
11.80
5.66
3.6. Thermal effects of modified hemicelluloses
0.95
3.75
The decomposition temperatures for the three hemicelluloses
and its modified form were evaluated by TGA, as shown in Fig. 7.
All modified samples differ from its native hemicellulose indicat-
ing successful reactions. For the modified xylan samples (Fig. 7a)
all modified samples showed an increase in onset temperature as
well as a more rapid decomposition compared with native xylan.
Only the TEMPO-oxidized samples are shown for the AX and GGM
samples due to the higher DS for those samples compared with the
non-oxidized samples (Fig. 7b and c). The results are varying for the
two hemicelluloses but in both cases the onset temperatures are
decreased; and for the GGM sample the decomposition rate is slower
where most of the material decomposes at a higher temperature
compared with native GGM.
a
The reaction efficiency is dependent on the maximum DS for the hemicellu-
lose that has been determined from the carbohydrate composition to 0.13 for xylan,
0.3 for AX and 1.0 for GGM.
of oxidized groups in the polymers. The oxidation reaction seems
to work better on AX than GGM and the oxidation procedure could
be more investigated. In both cases the oxidation increased the re-
action efficiency. By controlling the number of carbonyl groups it
should also be possible to control the number of substituents.
The DS values presented in both Tables 3 and 4 are relatively low
compared with other reactions on hemicelluloses, for example the
etherification with 2,3-epoxypropyl trimethyl ammonium chlo-
ride (EPTMAC) giving a similar ring opening as for azetidinium ions.
DS on etherified birch xylan has been reported to vary between 0.17
and 1.64 depending on process parameters45 and the etherifica-
tion with EPTMAC on AX from barley husk has reported DS values
ranging from 0.03 to 0.2.58 One significant difference with the etheri-
fication with EPTMAC compared with azetidinium ions is that
EPTMAC reacts with both primary and secondary hydroxyl groups,
giving more available substitution targets.
Thermal stability is often necessary for heat processing of ma-
terials like composite manufacturing and the longer alkyl chains with
six carbons show a larger increase in degradation temperature com-
pared with the shorter 2-carbon chains. Even longer alkyl chains
could influence the thermal stability further.
4. Conclusions
This study shows that it is possible to attach different azetidinium
containing groups on hemicelluloses containing carbonyl groups such
as in glucuronic acids or after a TEMPO-oxidation of polymers con-
taining primary alcohols. Carbohydrate analysis of different pre-
treated hemicelluloses shows that alkali conditions degrade the
α-glycosidic linkage between galactose and the glucose–mannose
backbone in GGM and a TEMPO-oxidation with more neutral con-
ditions, such as the TEMPO/BAIB-oxidation was chosen to avoid
degradation of the polymers.
The three different azetidinium containing reagents that were
synthesized were used in attempt to study the reaction between
different hemicelluloses and to understand the chemistry to control
the reaction. There is a huge variety of azetidinium containing salts
that can be synthesized and used for modification of hemicellu-
lose to increase the value of the so far less explored hemicellulose
polymers. By knowing where on the polymer the reactions occur
it is easier to control the reaction and by studying different
azetidinium groups with different functionalities the properties of
Table 4
Degree of substitution (DS) for different pre-treatments of hemicelluloses; no treat-
ment at all, 17.5 wt% NaOH-activation or TEMPO-oxidation. The reaction efficiencies
are calculated from the maximum DS of available reaction targets in the hemicelluloses
Sample
Pre-treatment
%N
DSN
Reaction efficiency (%)a
Az-DEA-xyl
Az-DEA-xyl
Az-DEA-AX
Az-DEA-AX
Az-DEA-AX
Az-DEA-GGM
Az-DEA-GGM
Az-DEA-GGM
None
NaOH-act.
None
NaOH-act.
TEMPO-ox.
None
NaOH-act.
TEMPO-ox.
1.06
1.10
0.54
0.58
1.49
0.47
0.46
1.00
0.12
0.13
0.06
0.06
0.17
0.06
0.06
0.13
92.62
96.85
18.51
19.47
55.97 (92.39)b
5.74
5.66
13.09 (77.91)b
a
The reaction efficiency is dependent on the maximum DS for the hemicellu-
lose that has been determined to 0.13 for xylan, 0.3 for AX and 1.0 for GGM.
b
The numbers in brackets referring to the reaction efficiency for 63% oxidized
primary hydroxyl groups in AX and 17% oxidized primary hydroxyl groups in GGM.
Fig. 7. TGA curves for the three hemicelluloses and its modifications: (a) xylan, (b) arabinoxylan (AX), and (c) galactoglucomannan (GGM).

30
M. Börjesson, G. Westman /Carbohydrate Research 428 (2016) 23–30
18. Sun R, Fang JM, Tomkinson J. Polym Degrad Stabil 2000;67:345–53.
19. Sun R, Fang JM, Tomkinson J, Geng ZC, Liu JC. Carbohyd Polym 2001;44:29–39.
20. Gröndahl M, Gustafsson A, Gatenholm P. Macromolecules 2006;39:2718–21.
21. Cunha AG, Freire CSR, Silvestre AJD, Neto CP, Gandini A, Orblin E, et al. J Colloid
Interf Sci 2007;316:360–6.
22. Ebringerová A, Heinze T. Macromol Rapid Comm 2000;21:542–56.
23. Saadatmand S, Edlund U, Albertsson A-C, Danielsson S, Dahlman O, Karlström
K. Biomacromolecules 2013;14:2929–36.
24. Ebringerová A, Hromádková Z, Heinze T. Adv Polym Sci 2005;186:1–67.
25. Zhang Y-Q, Xie B-J, Gan X. Carbohyd Polym 2005;60:27–31.
26. Mamman AS, Lee J-M, Kim Y-C, Hwang IT, Park N-J, Hwang YK, et al. Biofuel
Bioprod Bior 2008;2:438–54.
the modified hemicelluloses can be tailored and for example longer
alkyl chains increase hydrophobicity. Based on structure–property
relationship hemicellulose derivatives from azetidinium salts have
an interesting spectrum in possible applications including barri-
ers or films for packaging solutions or carrier of drugs due to the
azetidinium reagents commonly used in drug development. Addi-
tionally, the reaction conditions need to be further optimized to reach
a more energy efficient process.
Acknowledgment
27. Reddy N, Yang Y. Trends Biotechnol 2005;23:22–7.
28. Voepel J, Edlund U, Albertsson A-C. J Polym Sci 2009;47:3595–606.
29. Voepel J, Sjöberg J, Reif M, Albertsson A-C, Hultin U-K, Gasslander U. J Polym
Sci 2009;112:2401–12.
The authors would like to acknowledge all involved persons and
companies in the Smartfoam project and Forskningsrådet Formas
for financial support. Thanks to Stora Enso (Karlstad, Sweden) and
Lyckeby Culinar (Kristianstad, Sweden) for providing the hemicel-
lulose material.
30. Bott TM, West FG. Heterocycles 2012;84:223–64.
31. Ross JH, Baker D, Coscia AT. J Org Chem 1964;29:824–6.
32. Chattopadhyay S, Keul H, Moeller M. Macromol Chem Phys 2012;213:500–12.
33. Restorp PA, Andersson AOR, Ronne NE. European Patent 20150059997 2015.
34. Gårdebjer S, Andersson M, Engström J, Restorp P, Persson M, Larsson A. Polym
Chem 2016;7:1756–64.
35. Espy HH. Tappi J 1995;78:90–9.
36. Siqueira EJ, Salon M-CB, Belgacem MN, Mauret E.
2015;132:42144.
Supplementary material
J Appl Polym Sci
37. Obokata T, Isogai A. Colloids Surf A 2007;302:525–31.
38. Leppänen A-S, Xu C, Eklund PC, Lucenius J, Österberg M, Willför SM. J Appl Polym
Sci 2013;130:3122–9.
Supplementary data to this article can be found online at
doi:10.1016/j.carres.2016.04.005.
39. Parikka K, Leppänen A-S, Xu C, Pitkänen L, Eronen P, Österberg M, et al.
Biomacromolecules 2012;13:2418–28.
40. Höije A, Gröndahl M, Tømmeraas K, Gatenholm P. Carbohyd Polym 2005;61:266–
75.
41. Murray L, O’Farrell A-M, Abrams T. U.S. Patent 2004/0209937A1, 2004.
42. Jain PP, Degani MS, Raju A, Ray M, Rajan MGR. Bioorg Med Chem Lett
2013;23:6097–105.
43. Bakalarz A, Helitiski J, Krawieeka B, Michalski J, Potrzebowski MJ. Tetrahedron
55 1999;55:12211–26.
References
1. Cumpstey I. ISRN Org Chem 2013;2013:1–27.
2. Cunha AG, Gandini A. Cellulose 2010;17:1045–65.
3. Heinze T, Koschella A, Ebringerova A. Chemical functionalization of Xylan: a short
review. In: Gatenholm P, Tenkanen M, editors. Hemicelluloses: science and
technology, Vol. 864. Washington, DC: ACS Symposium Series; 2004. p. 312–25.
4. Buchanan CM, Buchanan NL, Debenham JS, Gatenholm P, Jacobsson M, Shelton
MC, et al. Carbohyd Polym 2003;52:345–57.
5. Sjöström E. editor. Wood polysaccharides. In: Wood chemistry fundamentals and
applications. San Diego: Academic Press Inc.; 1993. p. 51–70.
6. Li Z, Brännvall E. Nord Pulp Pap Res J 2011;26:380–5.
7. Doliška A, Willför S, Strnad S, Ribitsch V, Kleinschek KS, Eklund P, et al.
Holzforschung 2012;66:149–54.
8. Gabrielii I, Gatenholm P, Glasser WG, Jain RK, Kenne L. Carbohyd Polym
2000;43:367–74.
9. Saake B, Kruse T, Puls J. Bioresource Technol 2001;80:195–204.
10. Bengtsson S, Åman P. Carbohyd Polym 1990;12:267–77.
11. Lindblad MS, Albertsson A-C. Chemical modification of hemicelluloses and gums.
In: Dumitriu S, editor. Polysaccharides: structural diversity and functional versatility.
New York: CRC Press; 2004. p. 491–508.
12. Vaidya AA, Gaugler M, Smith DA. Carbohyd Polym 2016;136:1238–50.
13. Chaa L, Joly N, Lequart V, Faugeron C, Mollet J-C, Martin P, et al. Carbohyd Polym
2008;74:597–602.
14. Fang JM, Sun R, Fowler P, Tomkinson J, Hill CAS. J Appl Polym Sci 1999;74:2301–11.
15. Moine C, Gloaguen V, Gloaguen J-M, Granet R, Krausz P. J Environ Sci Heal B
2004;B39:627–40.
16. Peng F, Ren J-L, Peng B, Xu F, Sun R-C, Sun J-X. Carbohyd Res 2008;343:2956–62.
17. Sun R, Fang JM, Tomkinson J, Hill CAS. J Wood Chem Technol 1999;19:287–306.
44. Buriks RS, Lovett EG. U.S Patent 4,341,887, 1982.
45. Schwikal K, Heinze T, Ebringerová A, Petzold K. Macromol Sy 2006;232:49–56.
46. Junli R, Xinwen P, Linxin Z, Feng P, Runcang S. Carbohyd Polym 2012;89:152–7.
47. Ebringerová A, Hromádková Z, Kacˇuráková M, Antal M. Carbohyd Polym
1994;24:301–8.
48. Nylander F. Lignin carbohydrate complexes: model compounds synthesis
[licentiate thesis]. Gothenburg: Chalmers University of Technology; 2015.
49. Theander O, Westerlund EA. J Agric Food Chem 1986;34:330–6.
50. Westbye P, Köhnke T, Gatenholm P. Holzforschung 2008;62:31–7.
51. Höije A, Sandström C, Roubroeks JP, Andersson R, Gohil S, Gatenholm P. Carbohyd
Res 2006;341:2959–66.
52. Albertsson A-C, Edlund U. Synthesis, chemistry and properties of hemicelluloses.
In: Plackett D, editor. Biopolymers—new materials for sustainable films and coatings.
West Sussex: John Wiley and Sons, Ltd.; 2011. p. 133–50.
53. Song X, Hubbe MA. Carbohyd Polym 2014;99:617–23.
54. Kacˇuráková M, Wellner N, Ebringerová A, Hromádková Z, Wilson RH, Belton PS.
Food Hydrocoll 1999;13:35–41.
55. Ren J-L, Sun R-C, Peng F. Polym Degrad Stabil. 2008;93:786–93.
56. Mima S, Miya M, Iwamoto R, Yoshikawa S. J Appl Polym Sci 2003;28:1909–17.
57. Bough WA, Salter WL, Wu ACM, Perkins BE. Biotechnol Bioeng 1978;20:1931–43.
58. Köhnke T, Brelid H, Westman G. Cellulose 2009;16:1109–21.