Industrial Crops and Products 31 (2010) 534541

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Industrial Crops and Products

journal homepage: www.elsevier.com/locate/indcrop

Measurement of structural carbohydrates, lignins, and micro-components of

straw and shives: Effects of extractives, particle size and crop species
Yukihiro Tamaki, Giuseppe Mazza
Pacic Agri-Food Research Centre, Agriculture and Agri-Food Canada, 4200 Highway 97 Summerland, British Columbia, Canada V0H 1Z0

a r t i c l e

i n f o

Article history:
Received 14 January 2010
Accepted 4 February 2010

Keywords:
Structural carbohydrates
Lignin
Extractives
Particle size
Straw
Shives
Wheat
Triticum aestivum
Durum wheat
Triticum durum
Barley
Hordeum vulgare
Oat
Avena sativa
Flax
Linum usitatissimum

a b s t r a c t
The effects of extractives, particle size, and crop species on the contents of structural carbohydrates,
lignins, and micro-components in Canada Prairie Spring (CPS) wheat, durum wheat, barley, oat, and
triticale straw, as well as ax shives, were determined.
Extraction for 24 h in water followed by 7 h in ethyl alcohol (EtOH) yielded 20.2% extractives in triticale
straw. Acid insoluble lignin decreased from 17.6% in native triticale straw to 13.6% in 24 h water + 7 h EtOH
extracted triticale straw. The sample particle size inuenced the values of glucan, xylan, acid insoluble
lignin, ash, and extractives. Glucan, xylan, and acid insoluble lignin levels increased with increasing
particle size. Protein, ash, and extractive levels decreased with increasing particle size. For ax shives,
glucan, xylan, and acid insoluble lignin levels increased with increasing particle size from 19.1% to 33.2%,
from 6.7% to 18.8%, and from 17.4% to 25.7%, respectively. The protein, ash, and extractive levels in ax
shives decreased with increasing particle size from 6.5% to 1.9%, from 27.7% to 0.9%, and from 15.0% to
3.8%, respectively. Total glycans, lignin and extractive levels for the various straws, from the medium
particle sized fraction, were 56.663.9, 14.719.4, and 6.820.2%, respectively. Total glycans, lignin, and
extractives levels from the medium particle sized ax shives were 51.8, 25.8, and 6.46%, respectively. The
composition of the medium particle size fraction reected the composition of the corresponding original
sample.
2010 Elsevier B.V. All rights reserved.

1. Introduction
In recent years, energy consumption has grown steadily as a
result of the increase in the worlds population and the growing
economies of many developing nations. Almost all the energy to
satisfy this increasing demand comes from fossil fuels. Burning of
fossil fuels has led to environmental problems such as air pollution, climate change, and acid rain and, in addition, fossil fuels are
facing progressive depletion, which has led to a rise in oil prices
and contributed to stagnation of the world economy (Campbell
and Laherrre, 1998; Snchez and Cardona, 2008). Therefore, the
development of clean and alternative energy sources is a global
priority.
Renewable biomass is a promising alternative energy source.
Forest products, agricultural crops and residues, and animal and
municipal wastes are sources of renewable biomass (Reddy and
Yang, 2005). Agricultural crops such as corn and sugar cane have

Corresponding author. Tel.: +1 250 494 6376; fax: +1 250 494 0755.
E-mail address: giuseppe.mazza@agr.gc.ca (G. Mazza).
0926-6690/$ see front matter 2010 Elsevier B.V. All rights reserved.
doi:10.1016/j.indcrop.2010.02.004

been used to produce fuel ethanol on a large scale in the United

States and Brazil (Kim and Dale, 2004; Orts et al., 2008). Worldwide
ethanol production in 2006 was estimated to be approximately 51
billion liters (Snchez and Cardona, 2008). On the other hand, there
has been a signicant effect on the agricultural commodity markets
due to the use of food and feed crops as feedstocks for ethanol production (Orts et al., 2008). Consequently, lignocellulosic biomass,
as an alternative to starch- and sugar-based fuel production, has
attracted considerable interest.
Lignocellulose, produced by vascular plants, is the most abundant source of renewable biomass, with the potential to supply
annually approximately 200 billion metric tons worldwide (Zhang
et al., 2007). Plant cell walls consist of three main biopolymers: cellulose (3050%); hemicellulose (1535%); and lignin (1030%) (da
Costa Sousa et al., 2009).
In a lignocellulosic feedstock biorenery, the three biopolymers are utilized for several applications (Ohara, 2003; Kamm and
Kamm, 2004; Carvalheiro et al., 2008). Cellulose can be used to produce fuels (e.g., ethanol), organic acids (e.g., lactic acid), solvents
(e.g., acetone, butanol), 5-hydroxymethyl furfural, levulinic acid,
and lubricants (Kamm and Kamm, 2004; Reddy and Yang, 2005;

Y. Tamaki, G. Mazza / Industrial Crops and Products 31 (2010) 534541

Orts et al., 2008). Hemicellulose can be used to produce ethanol,

xylite, furfural, nylons, and plant gums such as thickeners, adhesives, and stabilizers (Kamm and Kamm, 2004; Reddy and Yang,
2005; Orts et al., 2008). Lignin can be used to produce binders, adhesives, electricity, sub-bituminous coal, and sulphur-free solid fuel
(Kamm and Kamm, 2004; Reddy and Yang, 2005).
Agricultural residues, such as a cereal straw and sugarcane
bagasse, obtained after harvesting are one of the sources of annually renewable lignocellulosic biomass. They are mainly used as
animal feed and bedding at present; however, these lignocellulosic residues could be principal candidates for the production of
fuel, chemicals, and other industrial products. Wheat, rice, and barley straw, corn stover, sorghum stalks, sugarcane bagasse, coconut
husks, and pineapple and banana leaves are the main lignocellulosic
agricultural residues (Reddy and Yang, 2005). They are available in
considerable quantity, at low cost (Reddy and Yang, 2005) and do
not compete directly with food production (Hallac et al., 2009).
Canada has 27 million ha of cropland and produces 75 million
metric tons of agricultural crops (FAO, 2009). A large amount of
agricultural residues such as straw, stover, and shives are produced
every year. These represent signicant opportunities to utilize
lignocellulose as biorenery feedstocks as well as contribute to
environmental, social, and economic sustainability.
Analysis of components in various agricultural residues is essential to understand their utilization potential because lignocellulose
structures and compositions vary greatly depending on plant
species, plant tissues, and growth conditions (Zhang et al., 2007).
Ruth and Thomas (2003) showed that for corn stover a variation of
1% in carbohydrate content based on dry matter would change the
minimum ethanol selling price by $0.018/gal. In this study, we analyzed the structural carbohydrates, lignins and micro-components
in the straw and shives of selected Canadian crops: CPS wheat,
durum wheat, barley, oat, triticale, and ax.
It has been reported that extractives in the biomass and the particle size of a sample inuence composition (Thammasouk et al.,
1997; Bridgeman et al., 2007). Water and ethanol extractives from
a native sample can include non-structural sugars, organic acids,
inorganic material, nitrogenous material, chlorophyll, waxes, and
other minor components (Sluiter et al., 2005; Chen et al., 2007).
Thammasouk et al. (1997) reported that the extractives from herbaceous biomass (switchgrass, corn stover, and fescue) inuenced the
composition analysis. They concluded that some of the extractives
could be condensed or precipitated under the strong acidic conditions used for the measurement of acid insoluble lignin, leading to
a higher lignin level bias in native biomass. In addition, removal
of the extractives was also needed to obtain a more accurate
cellulose measurement because the extractives contained nonstructural sugars. Bridgeman et al. (2007) reported that fractions
with different particle sizes (<90 and 90600 m) showed different
analytical and chemical properties in reed canary grass and switchgrass. The milling process did not randomly reduce all the different
components of the samples in a uniform manner because a plant
consists of different tissues with different physical and chemical
properties. The large particle sized fraction had higher cellulose,
hemicellulose, and lignin levels, and a lower ash level than the small
particle sized fraction. Therefore, the effect of the extractives and
of the particle size on the quantities of the measured components
was examined.

2. Material and methods

2.1. Materials
Triticale ( Triticosecale W.; cultivars, AC Ultima and Pronghorn)
straw was provided by Agriculture Agri-Food Canada, Lethbridge,

535

Alberta. CPS wheat (Triticum aestivum L.), durum wheat (Triticum

durum L.), barley (Hordeum vulgare L.), and oat (Avena sativa
L.) straws were provided by CBioN ABIP Network, Saskatoon,
Saskatchewan. Flax (Linum usitatissimum L.) shives were provided
by Biolin Research Inc. Saskatoon, Saskatchewan.
2.2. Milling of samples
Raw samples were milled using a Retsch SM 2000 cutting
mill (Retsch GmbH, Haan, Germany) with a 2 mm discharge
screen. Milled samples (CPS wheat, durum wheat, barley, oat,
and triticale straws) were separated into ne (<180 m), medium
(180850 m), and coarse (8502000 m) fractions using a Retsch
AS 200 tap sieve shaker (Retsch GmbH, Haan, Germany) with 20and 80-mesh sieves. Milled ax shives were separated into ne
(<150 m), medium (150850 m), and coarse (8502000 m)
fractions with 20- and 100-mesh sieves. The sieves were shaken
for 5 min. The sieved samples were kept at 20 C until analyzed.
2.3. Extraction
2.3.1. The NREL procedure
Determination of extractives in biomass was carried out by the
NREL procedure (Sluiter et al., 2005). First, a water extraction was
performed for 12 or 24 h using a conventional Soxhlet apparatus
(85 mL extraction tube; 500 mL boiling ask; heating mantle (GlasCol, Terre Haute, IN)). The reux rate of the water was adjusted to
provide four to ve siphon cycles per hour. After the water extraction, an ethanol extraction was performed for an additional 7 h.
The reux rate of the ethanol was adjusted to provide 610 siphon
cycles per hour. The extractives obtained were evaporated at 40 C
using a rotary evaporator to remove most of solvent, and dried in
a vacuum oven at 35 C for 24 h. The dried extractives were kept at
20 C until analyzed.
2.4. Determination of structural carbohydrates, lignins, acetyl
groups, and ash content
Carbohydrates, acid insoluble and acid soluble lignin, acetyl
groups, and ash content in native and extractive-free samples were
determined by the NREL procedure: determination of structural
carbohydrates and lignin in biomass (Sluiter et al., 2008).
The samples were hydrolyzed with a sequential acid hydrolysis procedure utilizing 72% H2 SO4 at 30 C for 1 h and followed by
4% H2 SO4 at 121 C for 1 h. Monosaccharides in the hydrolyzate
were quantitatively measured with HPLC using an Agilent 1100
equipped with a refractive index detector (Agilent Technologies
Inc., Palo Alto, CA). The HPLC analysis was carried out using a Biorad Aminex HPX-87P column (300 7.8 mm, Bio-Rad Laboratories,
Hercules, CA) with a H+ /CO3 De-Ashing Rell Cartridge guard column (30 4.6 mm, Bio-Rad Laboratories, Hercules, CA) operating at
75 C with a MilliQ water mobile-phase at a ow rate of 0.5 mL/min.
Acid insoluble lignin was determined gravimetrically as the
ash-free acid insoluble residue resulting from the hydrolysis. Acid
soluble lignin was calculated from the absorbance at 320 nm of
the liquid phase resulting from the hydrolysis. An absorptivity of
30 L g1 cm1 was used to convert absorbance readings to mass
values (Sluiter et al., 2008).
Acetyl groups in the hydrolyzate were quantitatively measured
with HPLC using an Agilent 1100 equipped with a refractive index
detector (Agilent Technologies Inc., Palo Alto, CA). The HPLC analysis was carried out using a Bio-rad Aminex HPX-87H column
(300 7.8 mm, Bio-Rad Laboratories, Hercules, CA) with a Cation H
Rell Cartridge guard column (30 4.6 mm, Bio-Rad Laboratories,
Hercules, CA) operating at 55 C with a 0.005 M H2 SO4 mobilephase at a ow rate of 0.6 mL/min.

536

Y. Tamaki, G. Mazza / Industrial Crops and Products 31 (2010) 534541

Ash content of the samples was determined by complete

combustion in a mufe furnace (Model F-A1730, Thermolyne Corporation, Dubuque, IA) equipped with a temperature controller
(Furnatrol II series 413, Thermolyne Corporation, Dubuque, IA) running a temperature ramp program: ramp from room temperature to
105 C; hold at 105 C for 12 min; ramp to 250 C at 10 C/min; hold
at 250 C for 30 min; ramp to 575 C at 20 C/min; hold at 575 C
for 180 min; and, drop to and hold at 105 C until removed. The
remaining residue in the crucible was taken as the ash content.
2.5. Protein determination
Protein content was estimated from the nitrogen content of
the samples. Prior to the nitrogen analysis, the samples were all
ball-milled for 3 min at 30 Hz (Mix Mill MM301, Retsch GmbH,
Hann, Germany). The nitrogen content was determined using a
Leco FP-528 nitrogen analyzer (Leco Corporation, St. Joseph, MI)
by combustion of the ball-milled samples at 850 C. Protein content was then estimated by multiplying the nitrogen content (%)
by a factor of 6.25. A standard curve was made using ethylenediaminetetraacetic acid (EDTA) (Leco Corporation, St. Joseph, MI).

Fig. 1. Effect of extractives on the composition of triticale (AC Ultima) straw (results
expressed as a percentage of the native, oven-dried basis (wt/wtnative sample )). Particle
size: medium (180850 m). An error bar represents standard deviation. Different letters above columns in a group indicate a statistically signicant difference
(p < 0.05). NA: not applicable.

2.6. Uronic acids determination

Uronic acids content in the hydrolyzate was determined using
the Scott method (Scott, 2002). An aliquot (0.125 mL) of test solution was added to 0.125 mL of 2% NaCl3% H3 BO3 solution in a
test tube. Concentrated H2 SO4 was added to the test tube in an ice
bath and then mixed. The test tube was heated for 40 min at 70 C.
After the reactant cooled to room temperature, 0.1 mL of 0.1% 3,5dimethylphenol in glacial acetic acid was added to the reactant and
then mixed. After 10 min at room temperature, absorbance at 450
and 400 nm was measured. The A (A450 A400 ) was used to calculate uronic acids concentration. A standard curve was made using
d-glucuronic acid (Sigma-Aldrich Co., St. Louis, MO).
2.7. Prediction of the composition in the original biomass sample
To predict the composition in the original biomass sample we
used the following equation:
%XOriginal =

 %X

Fine

 %X

%Fr.Fine
100

  %X

Medium %Fr.Medium
+

Coarse %Fr.Coarse
100

100
(1)

where X is a percentage of a constituent of the composition of each

fraction, and Fr. is the percentage of each fraction.
2.8. Statistical analysis
The Kyplot v2.0 beta 15 software package (Yoshioka, 2002) was
used for data analysis. The Tukey or TukeyKramer test was used
to compare means. Differences with p < 0.05 were considered signicant.
3. Results and discussion
3.1. Effect of extractives on the composition of biomass
It has been reported that extractives in plants inuence the
structural carbohydrates and lignin analyses (Thammasouk et al.,
1997). Thus, extractives were removed sequentially with water for
12 or 24 h, and then with ethanol for 7 h. Fig. 1 shows the effect of
extraction procedures on the composition of triticale straw (particle size: medium, 180850 m). The 12 h water and 7 h ethanol

extractions (12 h water + 7 h EtOH extraction) yielded a total of

17.7% extractives (water, 15.6%; ethanol, 2.1%) based on dry matter. The 24 h water and 7 h ethanol extractions (24 h water + 7 h
EtOH extraction) yielded a total of 20.2% extractives (water, 18.4%;
ethanol, 1.8%) based on dry matter. This indicates that each water
extraction removed approximately 90% of the extractives, and
that the 24 h water + 7 h EtOH extraction procedure increased the
amount removed. The contents of glucan and xylan, the main carbohydrate components of the cell wall, were not signicantly different
between native and extractive-free samples, and ranged in value
from 34.5% to 35.3% (glucan) and from 18.8% to 19.0% (xylan) for
triticale straw. Galactan and arabinan contents were lower in the
extractive-free samples, indicating that a portion could be watersoluble due to their lower molecular weight and higher branched
structure compared to other glycans. In contrast, the removal of
extractives signicantly reduced acid insoluble lignin and ash contents of the straw. Acid insoluble lignin content decreased from
17.6% in the native sample to 13.7% in the 12 h water + 7 h EtOH
extracted sample and 13.6% in the 24 h water + 7 h EtOH extracted
sample. Ash content decreased from 7.4% in the native sample
to 2.4% in the 12 h water + 7 h EtOH extracted sample and 1.3%
in the 24 h water + 7 h EtOH extracted sample. This indicates that
the extractive-free samples contained approximately 80% of the
acid insoluble lignin and 2030% of the ash in the native sample.
Acid soluble lignin and protein contents were signicantly different between native and two extractive-free samples. This can
be attributed to the removal of compounds having absorbance at
320 nm and water-soluble proteins.
A large decrease in ash and acid insoluble lignin contents of
extractive-free samples was observed for CPS wheat, durum wheat,
barley, and oat straws, and ax shives. The results indicate that
the extractives contained a large amount of inorganic materials
removed from the native samples. In addition, the extractives contained some water- and ethanol-soluble components leading to a
higher lignin value, because lignin is difcult to dissolve in water
and ethanol under the extraction conditions.
The composition of the extractives obtained after 24 h water
extraction from triticale straw was determined using the methods
of analysis used for straws and shives. The results showed that the
ash and acid insoluble lignin accounted for 31.4% and 12.9% of the
extractives content, and corresponded to approximately 5.8% and
2.4% of the native sample, respectively. The ash and acid insoluble
lignin contents of the 7 h ethanol extractives were estimated to be

Y. Tamaki, G. Mazza / Industrial Crops and Products 31 (2010) 534541

0.4% and 1.7% of the native sample, respectively. These values are
in agreement with Thammasouk et al. (1997) and show that the
water and ethanol extractives contained false lignin components
that became insoluble under acidic conditions. To date, the false
lignin in extractives has not been investigated in detail; however, it
is believed represent protein, cutins, and suberin (Theander, 1991;
Christiernin et al., 2005; Rowell et al., 2005). Therefore, removal of
extractives is needed to obtain an accurate lignin measurement.
The glucan content of the extractive-free samples was lower
than the native sample although the difference was not statistically signicant. In addition, the 24 h water extractives obtained
from triticale (cv. AC Ultima) straw contained glucose comprising 12.4% of the extractives. Previous studies (Thammasouk et al.,
1997; Sluiter et al., 2005; Chen et al., 2007) have reported that the
extraction process removes soluble sugars. This indicates that a
native sample contains soluble sugars and the glucan value of an
extractive-free sample can provide a more accurate cellulose measurement. Thus, as for lignin, an extractive-free sample should be
analyzed for cellulose content.
Industry generally utilizes biomass as received, but requires,
for efcient utilization, composition data for the total amount
of carbohydrates that is theoretically available. An analysis of
an extractive-free sample cannot provide the value because the
analysis excludes soluble sugars. Therefore, analyses of both an
extractive-free sample and the extractives, or of the native sample,
may be needed to obtain a more accurate estimate of the available
carbohydrates in a lot of feedstock.
These results indicate that removal of extractives from a native
biomass sample prior to the analysis is needed to obtain more accurate measurements of structural components, especially lignin. The
24 h water + 7 h EtOH extraction was used for subsequent experiments due to its higher extractives yield compared to the 12 h
water + 7 h EtOH extraction.
3.2. Effect of particle size on the composition of biomass
Milled triticale (cv. AC Ultima) and CPS wheat straws were
sieved into three fractions: ne (<180 m), medium (180850 m),
and coarse (8502000 m). The yield of these fractions (ne,
medium, and coarse) was 13.7, 65.6, and 20.8% for triticale straw,
and 11.1, 71.6, and 17.3% for wheat straw, respectively. Milled
ax shives were sieved into three fractions: ne (<150 m),
medium (150850 m), and coarse (8502000 m). The yield (ne,
medium, and coarse) was 6.1, 75.6, and 18.3%, respectively. The
medium-sized fraction was always the largest of the three fractions.
Figs. 2 and 3 show the effect of particle size on the composition of the extractive-free triticale and wheat straw samples. In
extractive-free triticale straw, glucan (30.8%) and xylan (16.5%)
measurements of the ne sample were signicantly lower than the
medium (glucan, 34.5%; xylan, 18.9%) and the coarse (glucan, 34.7%;
xylan, 19.6%) fractions. The measurements for the medium and the
coarse fractions were almost the same. The value of acid insoluble
lignin increased with increasing particle size (ne, 12.0%; medium,
13.6%; coarse, 14.4%), whereas, ash and extractives values of the
triticale straw decreased signicantly with increasing particle size.
The ash measurement (3.8%) of the ne fraction was more than
double that of the other fractions (medium, 1.3%; coarse, 1.6%). The
protein level also signicantly decreased with increasing particle
size.
A similar variation was observed in the CPS wheat straw fractions. In extractive-free wheat straw, glucan and xylan levels
increased signicantly from 34.4% to 39.4% and from 15.2% to
19.5%, respectively, with increasing particle size. The amounts of
acid insoluble lignin were also different depending on the particle size (ne, 15.4%; medium, 17.3%; coarse, 17.0%). The quantities
of ash and extractives decreased signicantly with increasing par-

537

Fig. 2. Effect of particle size on the composition of extractives-free triticale (AC

Ultima) straw (results expressed as a percentage of the native, oven-dried basis
(wt/wtnative sample )). Particle size: ne, <180 m; medium, 180850 m; coarse,
8502000 m. An error bar represents standard deviation. Different letters above
columns in a group indicate a statistically signicant difference (p < 0.05).

ticle size. The ash measurement (6.1%) of the ne fraction was

ve to eight times higher than the other fractions (medium, 1.2%;
coarse, 0.8%). The extractives measurement (16.4%) for the ne fraction was approximately 1.5 times higher than the other fractions
(medium, 11.2%; coarse, 10.0%). Protein levels decreased signicantly with increasing particle size.
A particle size effect was also observed in ax shives fractions
and it was more pronounced (Fig. 4). Overall, the measurements
of glycan, xylan, acid insoluble lignin, protein, ash, and extractives
in the ne fraction were signicantly different from the other two
fractions. The ne fraction contained only approximately 60% of the
glucan (19.1%), 40% of the xylan (6.7%) and 70% of the acid insoluble
lignin (17.4%) in comparison with the other two fractions (glucan,
31.0% and 33.2%; xylan, 17.2% and 18.8%; and, acid insoluble lignin,
25.0% and 25.7% for the medium and the coarse fractions, respectively). In contrast, the ash (27.7%) and extractives (15.0%) levels
in the ne fraction were more than 10 times and two times higher
than in the other fractions (ash, 2.2% and 0.9%; and extractives, 6.5%
and 3.8% for medium and coarse fractions, respectively). The protein (6.5%) level in the ne fraction was approximately three times
higher than the other fractions (medium, 2.5%; coarse, 1.9%).

Fig. 3. Effect of particle size on the composition of extractives-free CPS wheat straw
(results expressed as a percentage of the native, oven-dried basis (wt/wtnative sample )).
Particle size: ne, <180 m; medium, 180850 m; coarse, 8502000 m. An error
bar represents standard deviation. Different letters above columns in a group indicate a statistically signicant difference (p < 0.05).

538

Y. Tamaki, G. Mazza / Industrial Crops and Products 31 (2010) 534541

Fig. 4. Effect of particle size on the composition of extractives-free ax shives

(results expressed as a percentage of the native, oven-dried basis (wt/wtnative sample )).
Particle size: ne, <150 m; medium, 150850 m; coarse, 8502000 m. An error
bar represents standard deviation. Different letters above columns in a group indicate a statistically signicant difference (p < 0.05).

Fig. 6. Effect of particle size on the composition of native CPS wheat straw (results
expressed as a percentage of the native, oven-dried basis (wt/wtnative sample )). Particle size: ne, <180 m; medium, 180850 m; coarse, 8502000 m. An error bar
represents standard deviation. Different letters above columns in a group indicate
a statistically signicant difference (p < 0.05).

The reason for the effect of particle size on composition is not

readily apparent. However, it is well known that surface area affects
extraction efciency (Cacace and Mazza, 2003). Figs. 57 show the
effect of the particle size on native triticale and wheat straws,
and ax shives. In native samples, glucan and xylan levels increased
with increasing particle size, and ash and protein levels decreased
with increasing size, as seen in extractive-free samples. This result
indicates that considerable change or fractionation of the component in samples takes place during milling and sieving. However,
acid insoluble lignin levels in triticale and wheat straws did not
increase with increasing particle size although the level in ax
shives in the ne sized fraction was signicantly lower than the
other larger fractions. Perhaps different false lignin contents in the
straws are also separated into different fractions, indicating that
the ner fraction could contain more false lignin.
Bridgeman et al. (2007) reported that the compositional variation of cellulose, hemicellulose, lignin and ash depended on the
particle size (<90 and 90600 m) in reed canary grass and switchgrass. The reason for separating the two fractions at the 90 m point
was that observable differences occurred with regard to particle
shape and sample color. In our experiment, these differences also

occurred between the ne fraction and the other fractions. Generally, the ne fraction was darker in color and more uniform in
shape than the other fractions. Similar to our results, Bridgeman
et al. (2007) showed that the cellulose, hemicellulose, and lignin
contents of the ne fraction (<90 m) were lower than those of
the coarse fraction (90600 m). Ash content of the ne fraction
was higher than that of the coarse fraction. Bridgeman et al. (2007)
also reported that the process of size reduction did not reduce the
different components of biomass material in a uniform manner.
Tenacious organic materials such as cellulose, hemicellulose and
lignin had a tendency to remain in the larger particle sized fraction.
In addition, inherently small and grindable inorganic constituents,
such as ash, tended to accumulate in the ner sized fraction.
Alvo and Belkacemi (1997) and Chundawat et al. (2007) also
reported similar compositional changes with particle size for timothy, alfalfa, and corn stover. They fractionated a milled sample
into four and seven particle size fractions. The hemicellulose content increased with increasing particle size. There was a tendency
for the glucan content to increase with increasing particle size. The
protein and extractives contents tended to decrease with increasing
particle size. The lignin value, however, did not show clear trends.

Fig. 5. Effect of particle size on the composition of native triticale (AC Ultima) straw
(results expressed as a percentage of the native, oven-dried basis (wt/wtnative sample )).
Particle size: ne, <180 m; medium, 180850 m; coarse, 8502000 m. An error
bar represents standard deviation. Different letters above columns in a group indicate a statistically signicant difference (p < 0.05).

Fig. 7. Effect of particle size on the composition of native ax shives (results

expressed as a percentage of the native, oven-dried basis (wt/wtnative sample )). Particle size: ne, <150 m; medium, 150850 m; coarse, 8502000 m. An error bar
represents standard deviation. Different letters above columns in a group indicate
a statistically signicant difference (p < 0.05).

Y. Tamaki, G. Mazza / Industrial Crops and Products 31 (2010) 534541

539

Table 1
Composition of medium-sized extractive-free CPS wheat, durum wheat, barley, oat, and triticale straws, and ax shives (results expressed as a percentage of the native,
oven-dried basis (wt/wtnative sample )).* .
Component (%: dry weight basis)

Straws

Shives

CPS wheat

Durum wheat

Barley

Oat

AC Ultima

Pronghorn

Total glycans
Glucan
Xylan
Galactan
Arabinan
Mannan

59.48c
37.43d
18.97c
0.77b
1.50b
0.82b

59.72c
38.27c
18.75c
0.80b
1.50b
0.40cd

63.91a
41.65a
19.03c
0.96b
1.90a
0.38cd

61.99b
39.35b
19.68b
0.90b
1.80a
0.25cd

56.62d
34.48e
18.89c
0.85b
1.80a
0.60bc

57.43d
34.33e
20.32a
0.89b
1.76a
0.13d

51.77e
30.95f
17.19d
1.26a
0.46c
1.91a

Total lignin
Acid insoluble lignin
Acid soluble lignin

18.38c
17.34c
1.04b

18.15c
17.07c
1.08b

19.37b
18.21b
1.16a

18.20c
17.15c
1.05b

14.65d
13.59d
1.05b

14.80d
13.77d
1.03b

25.80a
25.02a
0.78c

Protein
Uronic acids
Acetyl groups
Ash
Extractives

3.57a
1.41c
2.01bc
1.21cd
11.18bc

2.21d
1.52c
2.23b
1.34bc
13.23b

2.69c
1.88b
1.92bc
1.11d
6.76c

1.88e
1.54bc
1.74c
1.44b
10.09bc

2.93b
1.38c
1.87bc
1.30bcd
20.16a

2.03e
1.62bc
2.25b
0.88e
19.62a

2.54c
3.43a
3.97a
2.20a
6.46c

Total

97.24abc

98.39ab

97.64abc

96.87bc

98.91a

98.64ab

96.17c

Triticale

Flax

Mean values in a row with different superscript letters are signicantly different (p < 0.05).
Particle size: 180850 m.
Particle size: 150850 m.

Sluiter et al. (2008) reported that deviation to a smaller or larger

particle size may result in a low bias in carbohydrate content and,
consequently, high lignin bias due to excessive carbohydrate degradation or incomplete hydrolysis of polymeric sugars to monomeric
sugars. Thus, it was suggested that the efciency of the hydrolysis
process might inuence the composition. In our study, however, the
measurements of galactan, arabinan and mannan in the ne sized
fraction were equivalent to, or higher than, those of the larger fractions. An absorbance increase at 280 nm in the hydrolyzate solution
caused by degradation byproducts such as furfural and hydroxymethyl furfural was not observed. The mass closure of the ne
fractions ranged between 98.2% and 100.4%. In addition, the total
glycans content of the coarse fraction was the highest of the three
fractions. The peak of polymeric materials by incomplete hydrolysis was not detected in the HPLC prole of the coarse fractions.
The mass closure of the coarse fractions ranged between 95.5% and
97.6%.

Therefore, it was considered that the compositional changes

among the three particle size fractions in our study would be caused
by the constituent fractionation and/or concentration by milling
and sieving, as reported by previous studies (Alvo and Belkacemi,
1997; Bridgeman et al., 2007; Chundawat et al., 2007).
3.3. Comparison of the composition of straws and shives from
different crops
Table 1 shows the composition of various extractive-free
medium-sized biomass types (CPS wheat, durum wheat, barley, oat,
and triticale straws and ax shives) expressed as a percentage of
the oven-dried native basis (%, wt/wtnative sample ).
Of the straws analyzed, barley had the highest content of total
glycans, total lignin, glucan and acid insoluble lignin, as well as
the lowest content of extractives. Triticale (cv. Pronghorn) had the
highest content of xylan, and the lowest content of glucan. Triticale

Table 2
Composition of medium-sized extractive-free CPS wheat, durum wheat, barley, oat, and triticale straws, and ax shives (results expressed as a percentage of the extractivesfree, oven-dried basis (wt/wtextractives-free sample )).* .
Component (%: dry weight basis)

Straws
CPS wheat

Shives
Durum wheat

Barley

Oat

Triticale

Flax

AC Ultima

Pronghorn

Total glycans
Glucan
Xylan
Galactan
Arabinan
Mannan

66.97c
42.14e
21.36c
0.86c
1.69c
0.92b

68.82b
44.11ab
21.60c
0.92bc
1.73c
0.46cd

68.54b
44.67a
20.41d
1.03bc
2.04b
0.40cd

68.94b
43.77bc
21.89c
1.00bc
2.01b
0.28d

70.90a
43.18cd
23.66b
1.06bc
2.25a
0.76bc

71.45a
42.71de
25.28a
1.11b
2.19ab
0.16d

55.35d
33.08f
18.38e
1.35a
0.50d
2.04a

Total lignin
Acid insoluble lignin
Acid soluble lignin

20.69bc
19.52bc
1.17c

20.92b
19.68b
1.24b

20.78b
19.53bc
1.24b

20.24c
19.08c
1.16c

18.34d
17.03d
1.32a

18.41d
17.13d
1.28ab

27.58a
26.75a
0.83d

Protein
Uronic acids
Acetyl groups
Ash
Total
*

4.02a
1.59c
2.27cd
1.36cd
96.89abc

2.54de
1.75bc
2.56bc
1.54bc
98.14ab

2.88c
2.02b
2.06d
1.19de
97.47abc

Mean values in a row with different superscript letters are signicantly different (p < 0.05).
Particle size: 180850 m.
Particle size: 150850 m.

2.09f
1.71bc
1.93d
1.60bc
96.52bc

3.68b
1.73bc
2.34bcd
1.63b
98.62a

2.53e
2.02b
2.80b
1.10e
98.31ab

2.71cd
3.66a
4.24a
2.36a
95.90c

540

Y. Tamaki, G. Mazza / Industrial Crops and Products 31 (2010) 534541

Table 3
Predicted values of the composition of the original triticale (AC Ultima) and wheat
straws, and ax shives. .
Component (%: dry weight basis)

CPS wheat

Triticale (AC Ultima)

Flax

Total glycans
Glucan
Xylan
Galactan
Arabinan
Mannan

58.87
37.10
18.65
0.81
1.52
0.78

55.97
34.03
18.72
0.85
1.80
0.58

51.11
30.62
16.84
1.29
0.46
1.90

Total lignin
Acid insoluble lignin
Acid soluble lignin

18.10
17.06
1.04

14.59
13.54
1.03

25.43
24.67
0.77

Protein
Uronic acids
Acetyl groups
Ash
Extractives

3.77
1.42
2.03
1.68
11.56

2.82
1.40
1.89
1.71
19.72

2.67
3.45
3.90
3.51
6.49

Total

97.42

98.09

96.58

Particle size: 2000 m.

(cv. AC Ultima) contained the highest content of extractives, and

the lowest amounts of total glycans, total lignin and acid insoluble
lignin. The composition of CPS wheat and durum wheat straws, and
triticale straws (AC Ultima and Pronghorn) were comparable. The
glycan (cellulose) and total lignin contents of the wheat, barley and
oat straws were similar to values reported in the literature (wheat,
2949% for cellulose and 521% for total lignin; barley, 3145%
and 1419%; oat, 3137% and 1619%, respectively) (Reddy and
Yang, 2005; Buranov and Mazza, 2008; Snchez, 2009). However,
the hemicellulose content (the sum of xylan, galactan, arabinan,
and mannan) was slightly lower than that reported in the literature (wheat, 2639%; barley, 2438%; oat, 2738%). In the case
of ax shives, the contents of hemicellulose and total lignin were
comparable to the values reported in the literature (hemicellulose,
1326%; total lignin, 2330%) (Sain and Fortier, 2002; Buranov and
Mazza, 2008). The cellulose content was, however, lower than the
values reported in the literature (3453%). It is likely that the degree
of purication of ax shives caused the apparent variance in cellulose content. Total glycans, total lignin, protein, ash and extractives
contents of the cereal crop straws in the medium particle sized fraction ranged from 56.6% to 63.9%, from 14.7% to 19.4%, from 2.0% to
3.6%, from 1.11% to 1.34%, and from 6.8% to 20.2%, respectively.
Flax shives had the highest levels of total lignin, acid insoluble
lignin, uronic acids and acetyl groups, and the lowest level of glucan
compared to the cereal straws.
Table 2 shows the composition of the extractive-free mediumsized biomass types expressed as a percentage of the oven-dried
extractives-free basis (%, wt/wtextractives-free sample ). The data presented in Table 2 are more commonly used in comparative studies
of the chemical composition of different lignocellulosic materials;
meanwhile, the data in Table 1 provide a cumulative measure of
feedstock composition in terms of an actual feedstock that would
be used in biomass-to-ethanol conversion processes. The levels
in triticale straws in Table 2 differed considerably from those in
Table 1 because the triticale straws contain high amounts of extractives (approximately, 20%). Thus, total glycans and xylan contents
of the straws indicated the highest amounts; however, the total
lignin content still remained the lowest amount. Total glycans, total
lignin, protein, and ash contents of lignocellulose in the medium
particle sized cereal straws ranged from 67.0% to 71.5%, from 18.3%
to 20.9%, from 2.1% to 4.0% and from 1.1% to 1.6%, respectively.
Table 3 presents the composition of the original triticale and
wheat straw and ax shives, calculated from the ratio of different
sized fractions and the composition values using Eq. (1). The predicted values of triticale (cv. AC Ultima) and wheat straws and ax

Fig. 8. Correlation between the values of the composition of the medium-sized fractions and predicted values of the composition of the original triticale (AC Ultima)
and CPS wheat straws, and ax shives. Particle size: 2000 m.

shives (Table 3) were comparable with the values of the medium

particle sized fraction of these plants (Table 1). The correlation coefcient between these two sets of data was 0.999, and the slope of
the linear regression was 0.995 (Fig. 8). These results indicate that
the composition of the medium particle sized fraction reects the
composition of the corresponding original sample.
4. Conclusions
The results of this research indicate that extractives and particle size inuence the accuracy of the compositional analysis of
wheat, barley, oat and triticale straws and ax shives. The extractives from native samples contained soluble materials (false lignin)
that become insoluble during acid hydrolysis. Removal of the
extractives is needed prior to the structural composition analysis
of biomass to obtain more accurate measurements of lignin and
other cell wall components. Composition changes related to particle size of samples were also observed. Particle size particularly
inuenced glucan, xylan, acid insoluble lignin, ash content, and
extractives yields. The amount of glucan, xylan, and acid insoluble lignin increased with increasing particle size, whereas, protein,
ash, and extractives content decreased with increasing particle size.
The composition measurements of CPS wheat, durum wheat, barley and oat straws were comparable; however, the composition of
triticale straw differed from the other cereal straws. The triticale
straw contained less glucan and acid insoluble lignin, more extractives, and a relatively higher ratio between total glycans and total
lignin. Flax shives showed a higher total lignin and a lower total
glycans content than the cereal crop straws.
Acknowledgements
The authors are grateful to the ABIP-CTBI Network, ABIPCBioNet and ABIP-NAFGEN for the nancial support. The technical
assistance provided by Lana Fukumoto, David Godfrey, Yasantha
Athukorala, Miao Zhang and Michael Parker is appreciated.
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