Food Chemistry 187 (2015) 280–289

Contents lists available at ScienceDirect

Food Chemistry
journal homepage: www.elsevier.com/locate/foodchem

Wheat milling by-products and their impact on bread making

Sami Hemdane, Sofie Leys, Pieter J. Jacobs, Emmie Dornez, Jan A. Delcour, Christophe M. Courtin ⇑
Laboratory of Food Chemistry and Biochemistry and Leuven Food Science and Nutrition Research Centre (LFoRCe), KU Leuven, Kasteelpark Arenberg 20 – box 2463, 3001
Leuven, Belgium

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

Article history: This study investigates the relationship between the properties of dietary fiber (DF) rich wheat milling
Received 19 October 2014 by-products and their impact on bread making. From coarse bran over coarse and fine weatings to low
Received in revised form 10 February 2015 grade flour, the content of starch and lipids increased, while that of DF and ash decreased. Enzyme activ-
Accepted 13 April 2015
ity levels differed strongly and were not related to other by-product properties. Average particle size of
Available online 22 April 2015
the by-products was positively correlated with DF and ash contents and their hydration properties. When
meals from flour and by-products were composed on the same overall starch level to compensate for dif-
Keywords:
ferences in endosperm contamination in the by-products, bread specific volume was more strongly
Wheat
Coarse bran
depressed with fine weatings and low grade flour than with coarse bran and weatings. This suggests that
Coarse weatings the properties of the former were intrinsically more detrimental to bread making than those of the latter.
Fine weatings Ó 2015 Elsevier Ltd. All rights reserved.
Low grade flour
Bread making

1. Introduction
Wheat (Triticum aestivum L.) bran, recovered in a number of by-
products of the wheat milling industry, is a good source of dietary
fiber (DF), antioxidants, B vitamins and minerals (Delcour &
Hoseney, 2010; Shewry, 2009). Epidemiologic evidence exists that
the daily consumption of bran-enriched cereal products may
reduce the risk of welfare diseases like obesity (de Munter, Hu,
Spiegelman, Franz, & van Dam, 2007), cardiovascular disease
(Jacobs & Gallaher, 2004) and colorectal cancer (Jacobs, Marquart,
Slavin, & Kushi, 1998). Furthermore, the European Food Safety
Authority recently recognizes two health claims concerning wheat
bran, allowing to state that wheat bran increases fecal bulk and
accelerates the intestinal transit (EFSA Panel on Dietetic Products
Nutrition and Allergies (NDA), 2010). Therefore, incorporation of
wheat bran in cereal-based products is an interesting and low-cost
strategy to improve their nutritional properties and physiological
effects. However, incorporation of bran-containing milling by-
products in cereal-based foods generally results in inferior end-
product quality. In bread, such incorporation results in a decreased
bread loaf volume, an increased crumb firmness and a decreased
sensory acceptance (Moder, Finney, Bruinsma, Ponte, & Bolte,
1984; Pomeranz, Shogren, Finney, & Bechtel, 1977).
Nowadays, the detrimental effect of wheat bran in bread mak-
ing is counteracted by use of strong wheat flour and addition of
⇑ Corresponding author. Tel.: +32 16 321917; fax: +32 16 321997.
E-mail address: christophe.courtin@biw.kuleuven.be (C.M. Courtin).
bread improvers, rather than working on bran itself. Although sev-
eral studies investigated the detrimental effect of wheat bran in
bread making, a clear view on the contribution of different aspects
of bran on this effect is lacking. Nevertheless, several hypotheses
on the underlying causes of quality loss exist. Firstly, it is generally
accepted that the replacement of flour by wheat bran causes a dilu-
tion of wheat gluten proteins. Pomeranz et al. (1977), for instance,
reported that the addition of up to 5% wheat bran decreased bread
volume to an extent expected from dilution of gluten proteins.
However, when 7% bran or more was added, the volume decrease
could no longer be explained by the dilution of gluten proteins
only. Also the competition for water between meal constituents
might explain the detrimental effect of wheat bran on bread prop-
erties. More recently, Bock, Connelly, and Damodaran (2013)
observed a redistribution of water among dough components
when wheat bran is added, resulting into partial dehydration and
structural changes of the gluten proteins. A third possible explana-
tion of the impact of bran on bread properties is its steric hin-
drance during dough development, disturbing the formation of
an optimal gluten network (Gan, Ellis, Vaughan, & Galliard,
1989). Wheat bran might furthermore not only hinder the forma-
tion of a gluten network, but may also destabilize gas cells, result-
ing into a decrease in gas retention (Gan, Galliard, Ellis, Angold, &
Vaughan, 1992; Pomeranz et al., 1977). Finally, Wootton and
Shams-Ud-Din (1986) noticed that bran extract is more detrimen-
tal to bread volume than the washed residue, suggesting the
involvement of deleterious bran-related enzymes or reactive
chemical components like glutathione or ferulic acid, which might

http://dx.doi.org/10.1016/j.foodchem.2015.04.048
0308-8146/Ó 2015 Elsevier Ltd. All rights reserved.
 S. Hemdane et al. / Food Chemistry 187 (2015) 280–289
react with sulfhydryl groups of gluten proteins (Noort, van Haaster,
Hemery, Schols, & Hamer, 2010).
When studying the impact of bran addition on bread making, it
should be recognized that the use of the term ‘‘bran’’ can lead to
confusion. From a botanical point of view, bran is defined as the
outer layers of the wheat kernel, consisting of the pericarp, seed
coat and nucellar epidermis. Bran defined by the miller contains
besides the botanical bran also aleurone and starchy endosperm
remnants which were not removed during the milling process
(Delcour & Hoseney, 2010). It should also be noted that botanical
bran is distributed over a range of different by-products formed
during milling, namely coarse bran, coarse weatings, fine weatings
and low grade flour. Coarse bran (which is usually defined as reg-
ular bran) and coarse weatings (or fine bran) are separated in the
early breaks of the milling process, while fine weatings (also
referred to as middlings or shorts) are smaller particles of endo-
sperm and bran, which were not separated after grinding
(Delcour & Hoseney, 2010). This latter by-product has also been
reported to contain more aleurone and germ than (coarse) bran
(Lai, Hoseney, & Davis, 1989b). Low grade flour (or red dog) is
the flour like material remaining after the last grinding steps and
contains a high level of non-endosperm material (Delcour &
Hoseney, 2010).
Although all of the above mentioned milling by-products can be
reconstituted with their flour complement to form whole meal, lit-
tle information can be found on their functionality relative to each
other in bread making. This is unfortunate from a scientific point of
view as comparison of their properties with their impact on bread
making might yield insight in the detrimental impact of bran in
general and the different by-products in particular on bread mak-
ing. Also from a practical point of view this is unfortunate, as each
of the fractions is a possible candidate for addition to bread for-
mula to enhance dietary fiber levels in bread.
Therefore, in this study, we try to gain more insight in the phe-
nomena underlying the detrimental effect of wheat bran on bread
making by studying the properties and functionality of different
wheat milling by-products (coarse bran, coarse weatings, fine
weatings and low grade flour) in bread making. To this end, differ-
ences in bran chemical composition, enzyme activity levels and
physical properties were investigated and related to dough and
bread properties.
2. Materials and methods
2.1. Materials
Ten wheat milling by-products were kindly provided by differ-
ent industrial milling companies. In addition, four milling by-prod-
ucts were produced on laboratory scale with a Bühler MLU-202
laboratory mill (Uzwil, Switzerland) by milling wheat cultivars
Akteur and Apache (harvest 2012) obtained from AVEVE (Landen,
Belgium), resulting into coarse bran 5 and fine weatings 3, and
coarse bran 6 and fine weatings 4, respectively. Akteur is a cultivar
with excellent baking properties and a flour protein content of
14.6 ± 0.1%, while Apache gives flour of inferior quality (flour pro-
tein content of 10.3 ± 0.2%). Milling yields were 76.3% and 75.8%,
respectively. The reason to produce the latter by-products was
twofold. First, it allowed to assess the difference in the properties
and functionality of milling by-products obtained from wheat cul-
tivars with superior and inferior baking properties. Secondly, the
production of these samples on laboratory scale resulted in better
insight in how the by-products obtained by laboratory milling dif-
fer from those obtained by industrial milling. The flour used in this
study was Crousti (Dossche Mills, Deinze, Belgium) with following
composition (dry matter basis): 2.3 ± 0.1% DF, 83.2 ± 1.1%
281
polymeric glucose, 13.6 ± 0.2% proteins, 0.63 ± 0.01% ash and
1.5 ± 0.1% lipids. Vital wheat gluten was from Tereos Syral (Aalst,
Belgium) and fresh baker’s yeast from AB Mauri (Merelbeke,
Belgium). Also commercial sugar and salt were used. All chemicals,
solvents and reagents were of analytical grade and purchased from
Sigma–Aldrich (Bornem, Belgium), unless specified otherwise.
2.2. Chemical composition of the milling by-products
DF content was analyzed according to the AOAC Official Method
991-43 (AOAC International., 1995). Total starch (measured as
polymeric glucose), total arabinoxylan (TOT-AX) and water-ex-
tractable arabinoxylan (WE-AX) contents were analyzed in tripli-
cate using a gas chromatography based method as described by
Courtin, Van den Broeck, and Delcour (2000). Saccharides were
hydrolyzed to their constituting monosaccharides with 2.0 M tri-
fluoroacetic acid, reduced to alditols with NaBH4 and derivatized
to alditol acetates using acetic acid anhydride. Polymeric glucose
was calculated as the level of glucose multiplied by 0.90, arabi-
noxylan content was calculated as the sum of the arabinose and
xylose levels multiplied by 0.88 (Courtin et al., 2000). Protein con-
tent was measured in duplicate by the Dumas combustion method,
an adaptation of the AOAC Official Method 990-03 (AOAC
International, 1995) to an automated Dumas protein analysis sys-
tem (EAS VarioMax N/CN, Elt, Gouda, The Netherlands). The
detected nitrogen was converted to protein content using 6.25 as
conversion factor. Total lipid content was determined gravimetri-
cally after extraction with water-saturated butanol as described
by Gerits, Pareyt, and Delcour (2013). Moisture, ash and damaged
starch contents were determined in triplicate following AACC-
methods 44-19.01, 08-01.01 and 76-31.01, respectively (AACC,
2000).
2.3. Enzyme activity levels of the milling by-products
a-Amylase and endoxylanase activity levels in bran extracts
were determined with the Amylazyme and Xylazyme AX methods,
respectively (Megazyme, Bray, Ireland). Enzymes were extracted
by suspending 1.0 g of sample in 10.0 mL maleate buffer
(100 mM, pH 6.0, 5 mM CaCl2) for the Amylazyme method and in
10.0 mL sodium acetate buffer (25 mM, pH 5.0) for the Xylazyme
AX method and shaking (Laboshake, VWR International, Leuven,
Belgium) during 30 min at room temperature. An azurine cross-
linked amylose or AX tablet was added to 1.0 mL pre-equilibrated
sample at 40 °C. After an appropriate incubation time (generally
15 min for amylase activity levels and 6 h for xylanase activity
levels), the reaction was stopped by adding 10.0 mL of
Tris(hydroxymethyl)aminomethane (Tris) base solution (2.0 w/v%
for the Amylazyme method, 1.0 w/v% for the Xylazyme AX
method). After filtration of the suspensions, the filtrates were
cooled to room temperature and the extinction values at 590 nm
(E590) [Ultraspec III UV/vis spectrophotometer (GE Healthcare,
Uppsala, Sweden)] were measured against a control, prepared by
incubating the extracts without the tablet. Correction was made
for non-enzymic color release by the substrate tablets. Activities
were expressed in a-amylase and endoxylanase units (AU and
XU, respectively) per gram dry matter (dm). One unit is defined
as the increase of E590 per hour of incubation, under the conditions
of the assay.
Endopeptidase activity levels were determined using azocasein
as substrate (Feller & Erismann, 1978). Enzymes were extracted by
suspending 1.0 g sample in 10.0 mL maleate buffer (100 mM, pH
6.0, 5 mM CaCl2) and shaking (Laboshake, VWR International,
Leuven, Belgium) during 30 min at room temperature. Azocasein
solution (350 lL, 1.4 w/v%) was added to 250 lL enzyme extract
and incubated for 4 h at 40 °C. After incubation, 500 lL of cooled
282 S. Hemdane et al. / Food Chemistry 187 (2015) 280–289
trichloroacetic acid (7 °C) was added and precipitated proteins
were removed by centrifugation (10,000g, 10 min). Finally,
500 lL of NaOH (500 mM) was added to 500 lL supernatants and
the extinction at 440 nm (E440) was measured. Endopeptidase
activity levels were expressed as endopeptidase units (PU) per
gram dm, with one PU defined as the increase of E440 per hour of
incubation.
2.4. Particle size distribution of the milling by-products
Particle size distribution of samples was determined in tripli-
cate by sieving 20.0 g of sample on a Vibratory Sieve Shaker AS
200 control (Retsch, Aartselaar, Belgium) for 30 min, with intervals
every 30 s and a shaking amplitude of 0.75 mm. Sizes of the sieve
pores were 2000 lm, 1000 lm, 710 lm, 500 lm, 250 lm, 125 lm
and 50 lm. The weight retained on the different sieves was
expressed as w/w%. Mean particle size (MPS) was also calculated,
using the formula of Ensor, Olson, and Colenbrander (1970):
Pn pffiffiffiffiffiffiffiffiffi
d d
ðW i log Þ
1 Pn i iþ1
Wi
MPS ¼ 10 1 ð1Þ
With Wi defined as the mass of sample retained on sieve i (g), di as
the pore diameter of sieve i (lm) and di+1 as the pore diameter of
the upper sieve following sieve i (lm).
2.5. Hydration properties of the milling by-products
The procedure for water retention capacity (WRC) determina-
tion was optimized based on the method described by Mongeau
and Brassard (1982). Wheat bran (1.0 g) was weighed in a 50 mL
tarred centrifuge tube and 10.0 mL deionized water was added.
After vortex stirring, the sample was left for 60 min at room tem-
perature, followed by 10 min of centrifugation at 4,000g. The
supernatant was carefully removed and a draining step of 15 min
was performed in order to remove residual non-retained water.
The centrifuge tube was weighed and the WRC was expressed as
g water retained per g dm bran and as g water per g by-product
DF (WRCDF).
Swelling capacity (SC) was determined by the bed volume tech-
nique as described by Kuniak and Marchessault (1972) with slight
adaptations. Approximately 500 mg of sample was weighed in a
10.0 mL graduated glass cylinder and steeped for 60 min at room
temperature in 5.0 mL deionized water. SC was expressed as vol-
ume occupied by the swollen sample (mL) per g dm bran.
2.6. Composing meals
For the investigation of the impact on wheat milling by-prod-
ucts on dough and bread making properties, meals were enriched
in different bran types by following two different approaches.
In a first approach, meals were composed on a 15% bran basis by
mixing 80% Crousti flour, 15% milling by-product and 5% vital
wheat gluten. In a second approach, meals were composed on an
overall 70% starch basis as starch was considered to be a good mea-
sure for the endosperm contamination of the bran. Therefore,
depending on the starch content of the milling by-products, differ-
ent levels of sample were mixed with Crousti flour. The reason for
the latter approach was that the different milling by-products ana-
lyzed in this study significantly differ in starch content, and thus
consequently in botanical bran content. In order to investigate
the functionality of the botanical bran component in the milling
by-products, we decided to also compose meals on a same overall
starch level of 70%. As a result of this approach, the endosperm
content was equal for all meals. Also in the latter approach, 5% vital
wheat gluten was mixed. In both approaches, a control flour com-
posed of 95% Crousti flour and 5% gluten was used as reference.
2.7. Dough properties
For the determination of dough properties, doughs of the com-
posite meals were made in triplicate using a Brabender E330
Farinograph (Duisburg, Germany) with a 10 g stainless steel mix-
ing bowl, according to AACC method 54-21.02 (AACC, 2000).
Farinograph water absorption (FWA) and dough development time
(DDT) were determined. FWA was expressed on a 14% meal mois-
ture basis and corrected to 500 Farinograph Units.
2.8. Bread making
Breads enriched in the different bran types were prepared in
triplicate according to the straight dough procedure of Shogren
and Finney (1984). The composite meal (10.00 g, moisture content
14.0%), 0.53 g compressed fresh yeast, 0.60 g sucrose, 0.15 g salt
and 6.8 mL water were mixed with a 10 g pin mixer (National
Manufacturing, Lincoln, NE, USA) for 4 min 15 s. For the breads
with low grade flour and composed on a 15% bran basis, a water
volume of 6.6 mL was used. For the white control bread (95%
Crousti flour, 5% gluten), 6.2 mL of water and a mixing time of
4 min were used. Preliminary bread making trials led to the use
of the amount of water and mixing times described here.
Fermentation and final proofing were performed in a fermentation
cabinet (National Manufacturing) at 30 °C and 90% relative humid-
ity for 90 and 36 min, respectively. Doughs were punched after 52
and 77 min fermentation and punched and molded after 90 min.
Baking was performed for 13 min at 232 °C in a rotary oven
(National Manufacturing). Bread volume was measured by rape-
seed displacement, according to AACC method 10-05.01 (AACC,
2000). Breads were weighed and their specific volumes calculated.
2.9. Statistical analysis
Statistical analysis of the results was performed using the
Statistical Analysis Software 9.3 (SAS Institute Inc., Cary, NC,
USA). One-way analysis of variation (ANOVA) was performed to
analyze significant differences between mean values of several
variables. After a positive omnibus test, post hoc analyses were
conducted to detect differences among experimental settings. A
Tukey multiple comparison procedure was used with a 5% family
significance level. Pearson’s correlation coefficients (P < 0.05) for
linear correlations between mean values were also calculated with
SAS 9.3.
Principal Component Analysis (PCA), a multivariate data analy-
sis method, was conducted with the Unscrambler software 9.1.2
(CAMO Technologies, Woodbridge, NJ, USA). In PCA, the high num-
ber of original variables is reduced to a smaller number of new
variables called principal components (PCs), which are linear com-
binations of the original variables. These PCs are independent vari-
ables and describe in decreasing order the variability of the data.
All variables were centered and scaled to unit variance prior to
the multivariate analyses.
3. Results
3.1. Chemical composition of the milling by-products
Table 1 shows the chemical composition of the milling by-prod-
ucts under study. On average, coarse bran contained 51.1% DF, of
which 25.5% TOT-AX, 22.0% starch, 18.9% proteins, 6.4% ash and
2.8% lipids. These values are comparable to data reported in
 S. Hemdane et al. / Food Chemistry 187 (2015) 280–289 283

Table 1
Dietary fiber, total arabinoxylan (TOT-AX), total and damaged starch, protein, ash and lipid contents of wheat milling by-products (coarse bran, coarse weatings, fine weatings and
low grade flour).a

a
Values in grey represent averages of the by-products from a different origin but of a same type. The individual samples are compared to each other for significant
differences and are grouped using small letters. The averages are compared to each other as well and are grouped using capital letters. Values within the same column not
sharing a same letter are significantly different (P < 0.05).

previous studies (Seyer & Gélinas, 2009; Zhang & Moore, 1997).
The average composition of coarse weatings was 43.2% DF, of
which 20.5% TOT-AX, 28.8% starch, 18.8% proteins, 4.6% ash and
3.7% lipids. Fine weatings consisted of 31.9% DF, of which 16.1%
TOT-AX, 40.5% starch, 19.5% proteins, 3.8% ash and 5.0% lipids
and low grade flour contained 14.5% DF, of which 9.1% TOT-AX,
54.4% starch, 18.8% proteins, 2.8% ash and 5.7% lipids. WE-AX-
levels were relatively low in all samples, ranging from 0.53% to
1.37% (results not shown), indicating that arabinoxylan in bran is
mainly water-insoluble. The damaged starch content increased
from 2.5% for coarse bran to 12.8% for low grade flour (P < 0.05,
Table 1). This fivefold increase in damaged starch content in the
finer bran types can only be partly explained by the increase in
total starch content, as the ratio of damaged starch over total
starch only increased twofold, from 11.4% for coarse bran up to
23.5% for low grade flour (P < 0.05).
Overall, when comparing the chemical composition of milling
by-products of the same type, differences in composition were
rather small. For protein contents, there was even little or no vari-
ability over the different milling by-product types (Table 1).

Table 2
a-Amylase, endoxylanase and endopeptidase activity levels of wheat milling by-products (coarse bran, coarse weatings, fine weatings and low grade flour).a
α-Amylase activity Endoxylanase activity Endopeptidase activity
Milling by-product
AU/g dm XU/g dm PU/g dm
Coarse bran 1 6.3 ± < 0.1b 2.4 ± < 0.1 h 3.8 ± 0.1 j
Coarse bran 2 70.5 ± 0.3 a 4.6 ± < 0.1 d 6.3 ± 0.1 d
Coarse bran 3 14.4 ± 0.2 i 4.7 ± < 0.1 c 4.7 ± 0.1 h
Coarse bran 4 4.5 ± 0.1 k 1.2 ± < 0.1 k 3.2 ± 0.2 k
Coarse bran 5 6.2 ± 0.1 j 1.4 ± < 0.1 j 5.1 ± 0.1 g
e a
Coarse bran 6 24.8 ± 0.1 5.6 ± < 0.1 6.4 ± 0.1 d
A A
Average 26.1 ± 24.8 3.3 ± 1.9 4.9 ± 1.3 A
c m
Coarse weatings 1 34.3 ± 0.2 0.9 ± < 0.1 5.4 ± 0.1 f
f e
Coarse weatings 2 21.8 ± 0.1 3.2 ± < 0.1 6.2 ± < 0.1d
d b
Coarse weatings 3 28.2 ± 0.1 5.4 ± < 0.1 7.5 ± 0.1 b
Average 28.1 ± 6.2 A 3.2 ± 2.3 A
6.4 ± 1.0 A
Fine weatings 1 17.7 ± 0.1 h 0.7 ± < 0.1 n
4.1 ± < 0.1i
Fine weatings 2 28.1 ± 0.1 d 2.0 ± < 0.1 i
9.6 ± 0.1 a
Fine weatings 3 9.1 ± 0.3 i 1.0 ± < 0.1 l
5.9 ± 0.1 e
g f c
Fine weatings 4 20.0 ± 0.3 2.6 ± < 0.1 6.6 ± 0.1
A A A
Average 18.7 ± 7.8 1.6 ± 0.9 6.5 ± 2.3
d g c
Low grade flour 28.0 ± 0.2 2.5 ± < 0.1 6.9 ± 0.1
Average 28.0 ± 2 / A 2.5 ± 1/9 A
6.9 ± 1/3 A

a
Values in grey represent averages of the by-products from a different origin but of a same type. The individual samples are compared to each other for significant
differences and are grouped using small letters. The averages are compared to each other as well and are grouped using capital letters. Values within the same column not
sharing a same letter are significantly different (P < 0.05).
284 S. Hemdane et al. / Food Chemistry 187 (2015) 280–289

A high variability in a-amylase activity levels was noticed, varying

80%
from 4.5 AU/g up to 70.5 AU/g, but no relation was found between
60% the type of bran and the a-amylase activity level. Indeed, the aver-
age a-amylase activity levels of the milling by-products were 26.1,
wt% 40% 28.1, 18.7 and 28.0 AU/g for coarse bran, coarse weatings, fine
weatings and low grade flour, respectively. Endoxylanase activity
20% levels ranged from 0.7 XU/g to 5.6 XU/g and endopeptidase activity
levels varied between 3.2 PU/g and 9.6 PU/g.
0%
CB 1 CB 2 CB 3 CB 4 CB 5 CB 6
3.3. Particle size distribution of the milling by-products
B 80%

Fig. 1 shows the particle size distribution of the milling by-

60%
products, with their corresponding MPS shown in Table 3. Coarse
wt% 40% bran samples (Fig. 1A) had a MPS varying between 936 lm and
1344 lm (Table 3). The distribution patterns of coarse and fine
20% weatings shifted to smaller sizes (Fig. 1B and C, respectively) com-
pared to the coarse bran samples (P < 0.05), with MPS-values vary-
0% ing between 348 lm and 523 lm for coarse weatings and between
CW 1 CW 2 CW 3 168 lm and 304 lm for fine weatings (Table 3). For low grade
C flour, the highest proportion of particles had a particle size
80%
between 125 lm and 250 lm (Fig. 1C), resulting in a MPS of
60% 173 lm. Some variations in particle size distribution were also
observed between the industrial and the laboratory samples
wt% 40% within one type of milling by-product. The laboratory samples
had a wider size distribution than the industrial samples, resulting
20% in a lower MPS for coarse bran compared to industrial coarse bran
and a higher MPS for fine weatings compared to industrial fine
0%
weatings.
FW 1 FW 2 FW 3 FW 4 LGF
<50 µm 50-125 µm 125-250 µm 250-500 µm
500-710 µm 710-1000 µm 1000-2000 µm >2000 µm 3.4. Hydration properties of the milling by-products
Fig. 1. Particle size distribution of coarse bran (CB, A), coarse weatings (CW, B), fine
weatings (FW, C) and low grade flour (LGF, C) as determined by the sieving method. The variability in WRC and SC of the different milling by-prod-
The Y-axis shows the weight percentage of the particles between the corresponding ucts was investigated (Table 3). Coarse bran generally had a high
sieve pore diameter. Error bars are shown for each sieve. WRC and SC (average values of 5.3 g H2O/g and 10.6 mL/g, respec-
tively), followed by coarse weatings (3.7 g H2O/g and 7.1 mL/g),
fine weatings (2.8 g H2O/g and 6.7 mL/g) and low grade flour
Table 3
(1.8 g H2O/g and 6.0 mL/g). Assuming that DF was the main chem-
Mean particle size (MPS), water retention capacity (WRC) and swelling capacity (SC)
of wheat milling by-products (coarse bran, coarse weatings, fine weatings and low ical component influencing the WRC, WRCDF-values were calcu-
grade flour).a lated: coarse bran had an average WRCDF of 10.2 ± 1.4 g H2O/g
DF, coarse weatings had a WRCDF of 8.7 ± 1.2 g H2O/g DF, fine weat-
ings 8.9 ± 1.2 g H2O/g DF and low grade flour 12.2 ± 0.7 g H2O/g DF.
Here, no trend could be observed anymore among the different
milling by-products

3.5. Relationships between chemical composition, enzyme activity

levels and physical properties of the milling by-products

To establish the relationship between chemical constituents,

a
Values in grey represent averages of the by-products from a different origin but
of a same type. The individual samples are compared to each other for significant
differences and are grouped using small letters. The averages are compared to each
other as well and are grouped using capital letters. Values within the same column
not sharing a same letter are significantly different (P < 0.05).
3.2. Enzyme activity levels of the milling by-products
a-Amylase, endoxylanase and endopeptidase activity levels of
the milling by-products were also determined (Table 2). A very
enzyme activity levels and physical properties of wheat milling
by-products, PCA was performed (Fig. 2). The first principal compo-
nent (PC 1) explained 62% of the variability in the dataset, while PC
2 explained 16% of the variability. On the left side of the correlation
loading plot (Fig. 2A), total starch, damaged starch and lipids clus-
ter together, while on the right side of the plot, DF and ash are
pooled. When observing the corresponding score plot, the different
milling by-products could clearly be distinguished (Fig. 2B), with
low grade flour on the left side of the score plot and the coarse bran
samples on the other side of the plot. Next, MPS and the hydration
properties (WRC and SC) are located close to the DF and ash con-
tents in the correlation loading plot (Fig. 2A), indicating that they
are positively correlated with these parameters. The second PC
seemed to be a measure for the level of a-amylase and
endoxylanase activity in the bran, though not of endopeptidase
activity.
 S. Hemdane et al. / Food Chemistry 187 (2015) 280–289
Fig. 2. PCA correlation loading plot (A) and score plot (B) of chemical composition,
enzyme activity levels and physical properties of different wheat milling by-
products. PC 1 (horizontal axis) explained 62% of the variability. PC 2 (vertical axis)
explained 14% of the variability. Abbreviations used: dietary fiber (DF), total
arabinoxylan (TOT-AX), water-extractable arabinoxylan (WE-AX), damaged starch
(DStarch), a-amylase activity level (Amy), endoxylanase activity level (Xyl),
endopeptidase activity level (Pep), mean particle size (MPS), water retention
capacity (WRC) and swelling capacity (SC), coarse bran (CB), coarse weatings (CW),
fine weatings (FW) and low grade flour (LGF).
3.6. Functionality of wheat milling by-products in dough and bread
making
Usually, when wheat bran is added to white flour, a specific
percentage of flour is replaced by bran. However, in the present
study, significant differences in starch content, and thus in endo-
sperm contamination, were observed between the analyzed
milling by-products. When using a fixed replacement level, this
implies that relatively less flour is replaced by botanical bran
when the milling by-product contains more endosperm. This is
for example the case for low grade flour compared to coarse bran.
Consequently, with this approach, meals containing equivalent
levels of coarse bran and low grade flour contain the same
amounts of milling by-product, but differ in endosperm content.
In order to investigate the functionality of the botanical bran
component in the milling by-products, we also composed meals
285
on a same overall starch level of 70%. Results of the addition of
the milling by-products under study on dough and bread proper-
ties considering these two approaches are here presented.
3.6.1. Dough and bread made with meals containing 15% milling by-
product
Results on FWA and DDT of the doughs prepared from meals in
which 15% of the flour was replaced by milling by-products are
summarized in Table 4A. FWA and DDT increased from 62.2%
and 2.6 min for white flour up to a maximum average value of
68.7% and 8.6 min for coarse bran. Compared to this latter value,
a slight decrease in FWA was noticed when flour was replaced by
finer bran types (P < 0.05). FWA values went from 68.7% for coarse
bran down to 66.8% for low grade flour. Finally, a shorter DDT was
observed when finer bran types were added, going from 8 to 9 min
for coarse bran down to 5 min for low grade flour.
Table 4A equally shows the results of the bread specific volumes
obtained with the standard flour (5.9 cm3/g) and meal in which
15% of flour was replaced by different milling by-products.
Besides the expected decrease in specific volume when wheat flour
was replaced by bran containing milling fractions, some differ-
ences between the breads with different milling by-products could
also be noticed. Indeed, although no significant differences were
observed between the breads with coarse bran, coarse weatings
and fine weatings (3.6, 3.9 and 3.7 cm3/g, respectively), replace-
ment of flour by low grade flour resulted in bread with a signifi-
cantly higher specific volume compared to breads containing
other milling by-products (4.4 cm3/g).
3.6.2. Dough and bread made with meals having an overall starch
content of 70%
In a second part, meals were composed of flour and milling by-
product to obtain an overall starch level of 70% (Supplementary
Table 1). With this approach, all of the composed meals had by
approximation the same quantity of endosperm and non-en-
dosperm material.
The meals composed using this approach were analyzed for
their dough properties (Table 4B). The FWA for by product contain-
ing meals varied between 69.5% and 73.8%. The highest values
were observed for the dough containing fine weatings and low
grade flour. Next, when the DDTs were investigated, some clear dif-
ferences in development times could be observed (Table 4B). A sig-
nificantly lower DDT was noticed for doughs containing wheat
bran with a smaller average particle size. Indeed, the highest aver-
age DDTs were observed for doughs containing coarse bran
(8.7 min), followed by doughs with coarse weatings (6.5 min), fine
weatings (5.4 min) and finally doughs containing low grade flour
(3.9 min). One exception was the coarse bran 5 sample, where a
DDT comparable to the doughs with coarse weatings was observed
(6.6 min).
Table 4B also shows the differences in bread specific volume. A
different trend was observed compared to breads composed on an
equivalent bran sample basis (Table 4A). Indeed, when considering
bread made with a recipe on a same starch basis (Table 4B), the
specific volume of breads composed with coarse bran was compa-
rable to the breads composed with coarse weatings (4.2 cm3/g and
4.0 cm3/g, respectively), but significantly higher compared to
specific volumes of breads containing fine weatings and low grade
flour (3.4 cm3/g and 2.8 cm3/g, respectively).
4. Discussion
First, relations in the chemical composition, enzyme activity
levels and physical properties between the milling by-products
286 S. Hemdane et al. / Food Chemistry 187 (2015) 280–289
Table 4
Farinograph water absorption (FWA), dough development time (DDT) and bread specific volume for dough and bread made from meals containing 15% by-product (A) and
composed on a 70% overall starch basis (B). The first column shows the milling by-product used in the meal (coarse bran, coarse weatings, fine weatings, low grade flour)a.
a
Values in grey represent averages of the by-products from a different origin but of a same type. The individual samples are compared to each other for significant
differences and are grouped using small letters. The averages are compared to each other as well and are grouped using capital letters. Values within the same column not
sharing a same letter are significantly different (P < 0.05).
were investigated. When comparing the chemical composition of
the different milling by-products, an increase in total starch, dam-
aged starch and lipid content and a decrease in DF and ash content
was observed when the by-product type was finer (P < 0.05). This is
caused by a less efficient separation of starchy endosperm and bran
in the finer fractions. Indeed, as mentioned earlier, it is common
knowledge that low-grade flour and fine weatings consist of endo-
sperm particles that still have bran attached after all the grinding
steps (Delcour & Hoseney, 2010). The higher lipid content in the
finer bran types are probably due to germ remnants or a higher
amount of aleurone in these milling by-products. When relating
these observations to the corresponding correlation loading plot
(Fig. 2A), it can be noticed that constituents which are typically
high in endosperm and/or germ are found to the left of the plot,
while constituents that are typically high in the outer wheat kernel
layers are found to the right of the plot. PC 1 hence represents the
level of endosperm contamination in the bran. It is also clear that
the protein content was not related to any other chemical compo-
nent or bran property. Nevertheless, the protein composition
would probably be different for the samples with a higher or a
lower endosperm contamination, as previous research showed that
the amino acid distribution differs between the different bran lay-
ers and the endosperm (Jensen & Martens, 1983).
Among the different samples in one by-product category only
relatively small variations in DF and starch contents were
observed. These can easily be attributed to the efficiency of the
milling process in different milling facilities. The largest difference
in composition was observed between the fine weatings derived
from industrial scale milling and laboratory scale milling. This
results from the fact that the fine weatings obtained on laboratory
scale also contains material that ends up in the low grade flour
fraction in an industrial mill.
Besides the strong correlation between the chemical compo-
nents, it was clear that DF was also highly positively correlated
to the by-product MPS, WRC and SC (Fig. 2A). The interaction
between bran and water can be explained by molecular interac-
tions like hydrogen bounds, ionic bounds or hydrophilic interac-
tions and by the entrapment of water in the bran matrix
(Chaplin, 2003). It is known that water strongly binds to DF com-
ponents such as water-insoluble arabinoxylan (Chaplin, 2003;
Courtin & Delcour, 2002), which is abundantly present in wheat
bran (Table 1). The comparable calculated WRCDF-values for the
different by-products (shown in the Results section) support this.
Indeed, from the WRCDF-values, we can deduce (i) that the amount
of water bound per unit of DF is virtually the same for all milling
by-product and hence (ii) that DF content is the main factor that
determines water binding capacity. Indeed, no clear correlation
could be found between the WRCDF and the type of milling by-pro-
duct, while WRC was clearly depended on the type of by-product.
The higher the overall level of DF in the by-product, the higher its
water binding capacity. However, it should be taken into account
that the by-products with higher DF-levels also had a higher
MPS. Indeed, also the microporous structure and imperfect packing
of bran particles are said to be important parameters for their
capacity to retain water in the tests used (Jacobs, Hemdane,
Dornez, Delcour, & Courtin, 2015; Mongeau & Brassard, 1982).
Also damaged starch can absorb more water than for instance
granular starch (Berton, Scher, Villieras, & Hardy, 2002).
Nevertheless, damaged starch and WRC are placed on opposite
sides of the PCA plots, indicating that damaged starch does not
noticeably contribute to a high WRC in the presence of high botan-
ical bran levels or DF levels as a proxy for such bran.
A last set of bran properties analyzed are the enzyme activity
levels. Although enzymes are highly concentrated in the outer lay-
ers of the wheat kernel (Poutanen, 1997), little is known about the
contribution of bran related enzymes to its detrimental effects in
bread making. Therefore, it seemed interesting to assess their vari-
ability in milling by-products and their relationship with either
other by-product properties or with their functionality in bread
making. None of the three analyzed enzyme activity levels (a-amy-
lase, endoxylanase, endopeptidase) seemed to be related to the
type of milling by-products. Previous studies reported for a-amy-
lase that milling by-products were relatively high in a-amylase
activity levels compared to the flour streams, but no clear relation-
ship between the activity levels and the different milling by-pro-
duct was noticed (Dornez, Gebruers, Wiame, Delcour, & Courtin,
2006). Furthermore, concerning endoxylanase, Dornez, Gebruers,
et al. (2006) observed high endoxylanase activity levels in different
milling by-products compared to the flour streams. But also in this
study, it was difficult to notice clear relationships between the
endoxylanase activity level and the type of bran. Finally, although
a-amylase and endoxylanase activity levels cluster together in the
PCA correlation loading plot (Fig. 2A), no strong correlations could
be found between the different enzyme activity levels (P > 0.05).
 S. Hemdane et al. / Food Chemistry 187 (2015) 280–289 287

Table 5
Pearson’s correlation coefficients between the average Farinograph water absorption (FWA), dough development time (DDT), bread specific volume (Sp. Vol.) and dietary fiber
(DF), total arabinoxylan (TOT-AX), water-extractable arabinoxylan (WE-AX), starch, damaged starch (D. Starch), proteins (Prot), ash, lipids, a-amylase activity (Amy),
endoxylanase activity (Xyl), endopeptidase activity (Pep), bran mean particle size (MPS), bran water retention capacity (WRC) and bran swelling capacity (SC) in the doughs and
breads containing 15% by-product (A) and composed on a 70% overall starch basis (B). Significant values (P < 0.05) are highlighted in bold.

DF TOT- WE- Starch D. Prot Ash Lipids Amy Xyl Pep MPS WRC SC FWA DDT Sp.
AX AX Starch Vol.
A FWA 0.69 0.75 0.26 0.73 0.53 0.16 0.74 0.56 0.05 0.55 0.12 0.56 0.69 0.64 1.00 – –
DDT 0.80 0.83 0.50 0.85 0.79 0.29 0.93 0.90 0.12 0.36 0.59 0.95 0.86 0.92 0.68 1.00 –
Volume 0.44 0.37 0.31 0.35 0.42 0.33 0.38 0.14 0.22 0.06 0.02 0.18 0.39 0.31 0.25 0.13 1.00
B FWA 0.19 0.09 0.52 – 0.35 0.53 0.19 0.56 0.23 0.28 0.56 0.58 0.40 0.44 1.00 – –
DDT 0.68 0.70 0.58 – 0.78 0.84 0.65 0.86 0.75 0.25 0.81 0.89 0.88 0.88 0.46 1.00 –
Volume 0.60 0.71 0.66 – 0.80 0.68 0.50 0.79 0.59 0.31 0.80 0.72 0.67 0.67 0.59 0.71 1.00

This suggests that higher or lower enzyme activities in the milling
by-products could depend on the wheat varieties or the level of
bran- or endosperm-related enzyme inhibitors or result from
pre-harvest sprouting (Lunn, Kettlewell, Major, & Scott, 2001) or
microbial contamination (Dornez, Joye, Gebruers, Delcour, &
Courtin, 2006). For apparent endoxylanase activity levels, for
instance, it is known that they strongly depend on the wheat vari-
ety and weather conditions, whereas the endoxylanase inhibitor
levels are mainly genetically determined (Dornez, Joye, Gebruers,
Lenartz, et al., 2006).
In a second part of this study, the bread making functionality of
the different milling by-products was investigated and related to
their properties (Table 5 and Supplementary Fig. 2). To this end,
meals were composed both on a constant by-product addition level
and on a same starch level basis. Composing meals on a same
starch level basis is not common. Nevertheless, this way, all breads
contain the same level of non-endosperm compounds, which is not
the case if a specific amount of flour is replaced by a milling by-
product. The impact of the milling by-products on dough and bread
properties using both approaches is discussed here.
In accordance with previous studies (Seyer & Gélinas, 2009;
Shenoy & Prakash, 2002), FWA increased when flour was replaced
by any milling by-product. This increase was expected as milling
by-products are relatively high in DF, which can strongly bind
water (Chaplin, 2003; Courtin & Delcour, 2002). However, it should
also be noticed that doughs containing milling by-products are
higher in damaged starch, which is reported to be highly positively
correlated to the hydration capacities of regular flour (Berton et al.,
2002). Nevertheless, no clear correlations could be found between
FWA and either DF or damaged starch levels in the doughs as
shown in Table 5.
Also DDT increased when flour was replaced by milling by-
products. This increase was dependent on the type of by-product
and highly correlated with bran particle size (Table 5, r = 0.95 for
meals containing 15% milling by-product, r = 0.89 for meals having
an overall starch content of 70%). Previous studies already
observed a decreased DDT with decreasing bran particle size
(Noort et al., 2010; Zhang & Moore, 1997), which Noort et al.
(2010) attributed to a slower water uptake rate for coarser bran
particles than for finer particles. This latter hypothesis would
imply that prehydration of wheat bran decreases the DDT com-
pared to doughs with untreated bran. This has been investigated
by Nelles, Randall, and Taylor (1998), who indeed reported a
decreased DDT when prehydrated bran was added. However, when
observing the Farinograms in that study in more detail, we could
not observe any clear DDT decrease, which makes it difficult to rely
on their conclusions on the one hand and on the hypothesis of a
slower water uptake for coarser particles on the other hand. It is
thus more likely that the larger bran particles hinder the develop-
ment of a proper gluten network more strongly than finer particles
due to their more hindering physical structure. Therefore, more
time is needed to develop an optimal dough if large bran particles
are added compared to doughs containing smaller particles.
Finally, the relationship between milling by-products proper-
ties and bread specific volume can be considered (Table 5 and
Supplementary Fig. 2). Besides the expected decrease in specific
volume when wheat flour was replaced by bran, differences
between the breads with different milling by-products were also
noticed. When flour was replaced by 15% of by-products, low grade
flour resulted in bread with a higher specific volume compared to
breads containing other milling by-products (P < 0.05). This obser-
vation could be explained by less dilution of gluten proteins in
bread containing 15% of low grade flour compared to the other
bran-enriched breads, due to the high endosperm contamination
of low grade flour. This hypothesis is quite plausible, as bread vol-
ume is positively affected by gluten quantity (Delcour & Hoseney,
2010). However, based on this hypothesis alone, one could expect
that the specific volume is higher when recipes containing by-
products with higher levels of endosperm remnants are consid-
ered. This is not entirely the case in present study. Breads contain-
ing fine weatings had a comparable specific volume to that of
breads containing coarse bran or coarse weatings. The botanical
bran of fine weatings seems hence inherently more detrimental
to bread quality than the botanical bran of coarse bran and coarse
weatings. This is confirmed by the results obtained with the breads
composed with a same starch level. From these results, it can be
concluded that some properties more associated with fine weat-
ings are more deleterious to bread quality compared to those more
associated with coarse bran and coarse weatings. They even show
that some properties more associated with low grade flour have
the most detrimental impact on bread quality. These properties
are discussed in more detail later in this paper.
One exception of the deleterious effect of fine weatings and low
grade flour was the bread with the fine weatings 3 sample, which
had a specific volume comparable to that of bread with coarse bran
and coarse weatings. This is probably due to the excellent bread
quality wheat cultivar from which the weatings originated and
thus, to the high quality gluten proteins in its attached endosperm.
Indeed, the proportion of gluten proteins originating from fine
weatings 3 in the meal was estimated to be around 14% of the total
gluten content in the meal, which was not negligible. The propor-
tion of gluten proteins originating from coarse bran was, for
instance, much lower (2.5% to 4% of the total gluten content in
the meals composed on a 70% starch basis).
The deleterious effect of bran in bread making might be either
of a physical, chemical or biochemical nature. Regarding the differ-
ences in physical properties between fine weatings and low grade
flour on the one hand and coarse bran and coarse weatings on the
other hand, a first obvious dissimilarity was the MPS, which was
smaller for the first two by-products than for the latter two. A con-
siderable number of studies tried to investigate the importance of
bran particle size in bread making. For instance, Noort et al. (2010)
288 S. Hemdane et al. / Food Chemistry 187 (2015) 280–289
reported a larger decrease in bread quality when flour was
replaced by finely ground bran compared to coarse bran.
However, it has also been reported that particle size reduction of
coarse bran slightly improved bread volume (Lai, Hoseney, &
Davis, 1989a; Moder et al., 1984), which indicates that the impact
of bran particle size on bread quality is still not very well under-
stood. In this study, no correlation between MPS and bread specific
volume was observed for breads with 15% by-product (Table 5). A
slightly positive correlation was found for the breads made with
meals formulated on a 70% starch (Table 5, r = 0.72). The latter cor-
relation suggests that, if all breads contain the same level of non-
endosperm compounds, smaller bran particles are more detrimen-
tal to bread volume than larger ones. This might be due to the
higher contact surface of smaller bran particles, resulting into a
more pronounced interaction between bran and flour components
(Noort et al., 2010). Besides, it is also possible that the more detri-
mental effect of smaller particles is due to the fact that they have a
higher abrasive effect on the gluten network during mixing, as
more fine bran particles were added than coarse bran, to obtain
breads with a same level of botanical bran. Nevertheless, the rela-
tive small correlation between MPS and the specific volume of
breads made with meal containing 70% starch (Table 5, r = 0.72)
suggests that the particle size of the by-products is not the only
factor that affects bread volume.
Next, a second difference in physical properties between the
milling by-products are the hydration properties. However, no
good correlations could be found between the hydration properties
of the by-products and the bread specific volume (Table 5).
Another explanation for the detrimental effect of fine weatings
and low grade flour might be that these milling by-products con-
tain higher levels of lipids or damaged starch. The specific volume
of breads made with meal containing 70% starch was negatively
correlated with lipids (Table 5, r = 0.79), which are known to neg-
atively influence bread volume, especially non-polar lipids
(Goesaert et al., 2005). Besides, a similar correlation was found
between the specific volume of breads containing 70% starch and
damaged starch levels (Table 5, r = 0.80). Although damaged
starch is not a bran-related component, the abundant presence of
this specific component is important to mention. Indeed, according
to Barrera, Pérez, Ribotta, and León (2007), a highly negative corre-
lation exist between the damaged starch level in flour and bread
volume.
Finally, a last possible reason for the decrease in bread quality
upon bran addition are bran-related enzymes. Here, no good corre-
lation could be found between the measured enzyme activity
levels in the meals and the specific volume of either breads con-
taining 15% by-product or 70% starch (Table 5), except between
endopeptidase and the specific volume of breads containing 70%
starch (Table 5, r = 0.80). Moreover, no clear relationship was
observed between the type of by-product and a-amylase-, endoxy-
lanase- and endopeptidase activity levels as shown in Table 2 and
Fig. 2A. This does not imply that these bran-related enzymes are
not functionally relevant in bread making, but implies that they
do probably not contribute to a net volume increase or decrease.
It can be concluded that depending on the type (coarse bran,
coarse weatings, fine weatings or low grade flour), wheat milling
by-products can strongly differ in chemical composition, enzyme
activity levels and physical properties. They induce clear changes
in dough and bread properties when flour is replaced by these dif-
ferent by-products. Bread specific volumes were more significantly
decreased when fine weatings or low grade flour were added on an
equal overall starch level, indicating that some properties more
associated with these by-products were more detrimental to bread
quality than properties more associated with coarse bran and
coarse weatings. These properties can be related to particle size,
lipid content or damaged starch content.
Acknowledgements
This work was performed within the framework of a Flanders’
FOOD (Brussels, Belgium) project. P.J. Jacobs acknowledges the
Institute for the Promotion of Innovation through Science and
Technology in Flanders (IWT-Vlaanderen, Brussels, Belgium) for
the financial support. E. Dornez acknowledges the ‘‘Fonds voor
Wetenschappelijk Onderzoek – Vlaanderen’’ (FWO, Brussels,
Belgium) for her postdoctoral fellowship. J.A. Delcour is W.K.
Kellogg Chair in Cereal Science and Nutrition at KU Leuven. This
study was also part of the KU Leuven Methusalem program
‘‘Food for the future’’ (20072021).
Appendix A. Supplementary data
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.foodchem.2015.
04.048.
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