Reduced Gelatinization, Hydrolysis, and Digestibility in Whole Wheat Bread
in Comparison to White Bread
Kristin Whitney and Senay Simsek†
ABSTRACT
Wheat, an important crop in North Dakota and the United States, is
often used for bread. Health concerns related to chronic diseases have
caused a shift toward consumption of whole wheat bread. There has been
some indication that the rate and amount of starch digestibility of whole
wheat breads may be lower than for their refined flour counterparts. This
research investigated the components of whole wheat bread that may
reduce starch digestibility and impact nutritional quality. Six formulations
of flour were used, which included two refined flours, two whole wheat
flours, and two whole wheat flours with added starch. The starch was
added to whole wheat flours to increase the starch level to that of the
refined flour so that we can determine whether or not the dilution of
the starch in whole wheat bread was a factor in lowering the estimated
Health concerns related to chronic diseases such as diabetes
mellitus, cardiovascular disease, cancer, and obesity have caused a
shift away from consumption of white bread toward whole grain and
whole wheat products (Slavin 2004). Whole wheat bread has in-
creased dietary fiber (arabinoxylan and cellulose), B vitamins,
mineral content, and phytochemicals, such as phenolic compounds,
phytates, and avenanthramides (Slavin 2004). These additional
components may have some interaction with starch or other com-
ponents, which result in changes to the digestibility of starch in
whole wheat bread.
Digestion and uptake of carbohydrates are typically slowed when
whole foods (e.g., whole wheat) are consumed. The rate of digestion
and absorption of glucose affects the glycemic response to a food.
Glycemic index (GI) is one indicator that can be used to compare
the in vivo glycemic response to foods (Slavin 2004). When in vitro
assay methods are employed, the term is referred to as estimated GI
(eGI) and is measured based on the glucose released from the test
food compared with the glucose released by the reference food
(Ovando-Martı́nez et al. 2011a). Foods can be classified as high
(>70), medium (56–69), or low (<55) GI foods (Venn and Green
2007; American Diabetes Association 2013). There have been
many studies showing that the glycemic response and starch di-
gestion are lower in whole wheat and whole grain products com-
pared with products produced from refined flour (Bravo et al.
1998; Slavin 2004; Tucker et al. 2014).
One component affecting the GI in bread is the amount of re-
sistant starch (RS), which is classified as dietary fiber because it
resists hydrolysis in the small intestine and is fermented by gut
microbiota in the large intestine (Holm and Björck 1992; Liljeberg
et al. 1996). RS can be classified into four groups based on the
mechanism of resistance. The types of RS are as follows: type I
(RS-I, physically inaccessible), type II (RS-II, granular morphol-
ogy), type III (RS-III, retrograded starch), and type IV (RS-IV,
modified starch). RS-III is composed of retrograded starch that is
formed during the cooking and cooling of starch-based food products
† Corresponding author. Phone: +1.701.231.7737. E-mail: senay.simsek@ndsu.edu
North Dakota State University, Department of Plant Sciences, P.O. Box 6050, Dept.
7670 Fargo, ND 58108-6050, U.S.A.
https://doi.org/10.1094/CCHEM-05-17-0116-R
© 2017 AACC International, Inc.
Cereal Chem. 94(6):991–1000
glycemic index (eGI) of whole wheat bread. White and whole wheat
flours and breads were evaluated for chemical composition, baking
quality by AACC International Approved Method 10-09.01, and eGI
by the Englyst assay. Whole wheat breads had significantly (P < 0.05)
higher mineral, protein, arabinoxylan, and phenolic acid contents, as
well as significantly (P < 0.05) lower eGI. The starch molecular weight
was also significantly (P < 0.05) higher for whole wheat and whole
wheat + starch breads compared with white breads. The eGIs of refined
flour breads were 93.1 and 92.7, whereas the eGIs of whole wheat and
whole wheat + starch breads ranged from 83.5 to 85.1. Overall, several
factors in the whole wheat bread composition can be found to affect the
quality and starch hydrolysis.
and makes up the majority of the RS found in bread (Sajilata et al.
2006).
Previous research has found that the process of baking bread
causes the formation of retrograded starch (RS-III) (Johansson et al.
1984; Holm and Björck 1992). When starches are cooled, retro-
gradation of amylose occurs, which reduces the digestibility of that
portion of the starch. Owing to a slower rate of retrogradation,
amylopectin will remain digestible for a longer period of time. The
cooking and cooling methods of starchy foods will influence starch
structure and effect digestibility (Lovegrove et al. 2017). Along with
the formation of RS-III, there may be other physicochemical causes
of lower GI in whole wheat breads. These include the amylose-
to-amylopectin ratio (Björck et al. 1994), presence of nonstarch
polysaccharides (arabinoxylans) (Choct and Annison 1992), anti-
nutrients (lectins, phytates, and enzyme inhibitors) (Thompson and
Yoon 1984), Maillard reaction products (Chung et al. 2011), and
starch–protein interaction (Jenkins et al. 1987).
Limiting water reduces starch gelatinization, which may reduce
starch digestibility. Arabinoxylans bind water and may make it less
available for starch gelatinization. Partially intact birefringent starch
granules have been observed in white bread containing about 60%
moisture. Also, studies have shown that the presence of hydrating
components, such as nonstarch polysaccharides, in cell wall com-
ponents reduce the water available for gelatinization and result in
the inhibition of starch digestibility by allowing the native starch
structure to persist (Mishra et al. 2012). The arabinoxylan in whole
wheat bread may also form an indigestible film around the starch
granules, acting as a physical barrier to starch digestion.
As seen in previous research, there are many possible causes of
the reduced GI of whole wheat bread (Thompson and Yoon 1984;
Jenkins et al. 1987; Bravo et al. 1998; Chung et al. 2011); however,
these interactions need to be studied in more detail. In this study,
starch digestibility in white and whole wheat breads will be in-
vestigated in relationship to the changes occurring to the starch
during baking.
MATERIALS AND METHODS
Materials. Four wheat flour samples were tested; these included
one commercially milled sample and three varieties (Barlow, Glenn,
and Prosper) of hard red spring wheat samples that were milled on a
laboratory mill (MLU-202, Buhler, Plymouth, MN, U.S.A.). Data
from analysis of the samples (refined, whole wheat, and whole
wheat + starch) made from these three varieties was combined for
Vol. 94, No. 6, 2017 991

statistical analysis and presentation of data. Commercially milled
white patent flour (nonenriched, nonbleached) and whole wheat
flour were obtained from North Dakota State Mill (Grand Forks,
ND, U.S.A.). After blending bran and shorts fractions in a cross-
flow blender, the particle size of the bran and shorts was reduced
by grinding in a hammer mill with a 0.8 mm screen (Perten In-
struments, Springfield, IL, U.S.A.).
Table I shows the formulation of the flour blends used in this
study. White flour and ground bran plus shorts from the three wheat
varieties were blended at the ratio of material obtained from mill-
ing to produce whole wheat flours. Starch was extracted with a
Glutomatic gluten washing machine (Perten Instruments) from each
white flour sample. Starch extracted from white flour was used to
prepare the whole wheat + starch samples. Starch was added to the
whole wheat flours from each of the three varieties to increase their
starch content to the same level as their corresponding white
straight-grade flour.
All bread samples were freeze dried and ground to a uniform
particle size in a mortar and pestle before chemical analysis. The
amyloglucosidase was obtained from Megazyme International
(Bray, Ireland), and the invertase, pancreatin, and other reagents
were purchased from Sigma-Aldrich (Saint Louis, MO, U.S.A.).
Flour Composition and Quality. Proximate analysis of all
flour blends was conducted to determine the quality of flour used for
these experiments. Determination of moisture, ash, and protein
contents was conducted according to AACC International Approved
Methods 44-15.02, 08-01.01, and 46-30.01, respectively. Starch
damage of the flour blends and bread samples was measured fol-
lowing AACCI Approved Method 76-30.02. The water absorption
and dough strength of the flours were determined with a farino-
graph, according to AACCI Approved Method 54-21.02.
Baking and Bread Evaluation. Bread formulations were
baked according to AACCI Approved Method 10-09.01, with some
modifications. Fungal a-amylase was used instead of malt powder,
and instant dry yeast was used instead of compressed yeast to im-
prove the constancy of these ingredients used in the baking formula.
Ammonium phosphate (5 ppm) was added to improve yeast func-
tion. The bread was prepared using a 2 h fermentation schedule,
rather than a 3 h fermentation, to avoid overfermentation (Gonzalez-
Gracia et al. 2012). The dough was punched once during fermen-
tation. After baking, the bread was evaluated for loaf volume by
rapeseed displacement (AACCI Approved Method 10-05.01). The
bread was lyophilized and ground to a fine and homogenous powder
in a food processor before additional analysis.
Bread Crumb Microstructure. The bread was also evaluated
with scanning electron microscopy (SEM) to determine starch granule
morphology and if there may be a physical barrier to the starch di-
gestion. Thin slices of bread were dried at room temperature and
mounted on aluminum mounts using colloidal silver or carbon adhe-
sive tabs and coated with gold using a Balzers SCD 030 sputter coater
(BAL-TEC RMC, Tucson, AZ, U.S.A.). Images were obtained with a
JEOL JSM-6300 SEM (JEOL USA, Peabody, MA, U.S.A.) using an
accelerating voltage of 10 kV (Ovando-Martı́nez et al. 2011b).
Arabinoxylan Content. Arabinoxylan content of the flour and
bread was measured according to the method of Blakeney et al.
TABLE I
Formulation of Flour Blends
Flour Flour Type
Flour 1 Commercial refined
Flour 2 Refined
Flour 3 Commercial whole wheat
Flour 4 Whole wheat
Flour 5 Commercial whole wheat + starch
Flour 6 Whole wheat + starch
z Base flours: formula 1, commercially milled straight grade flour; formulas 2, 4, and 6, laboratory-milled straight-grade flour; and formulas 3 and 5, commercially
milled whole wheat flour.
992 CEREAL CHEMISTRY
19433638, 2017, 6, Downloaded from https://onlinelibrary.wiley.com/doi/10.1094/CCHEM-05-17-0116-R by Cnr Roma, Wiley Online Library on [16/01/2024]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
(1983). Samples (6–7 mg) were hydrolyzed with trifluoroacetic
acid (250 µl, 2M) at 121°C for 1 h, and m-inositol was added as
the internal standard. The samples were then dried and reduced
with ammonium hydroxide (100 µl, 1M) and sodium borohydride
(20 mg/mL) in dimethyl sulfoxide. The reaction was stopped with
glacial acetic acid, and 1-methylimidazol (100 µl) and acetic anhydride
(0.5 mL) were added to acetylate the samples. The samples were par-
titioned twice with 1 mL of methylene chloride, which was transferred
to a second tube. The methylene chloride fractions were pooled and dried
at 45°C under nitrogen and the samples redissolved in 1 mL of acetone.
The derivitized sugars were quantified by an Agilent (Santa
Clara, CA, U.S.A.) 7890 gas chromatograph with a flame ionization
detector. An SPTM-2380 column (30 m × 0.25 mm × 0.2 µm, Supelco,
Bellefonte, PA, U.S.A.) was used for separation. The flow rate was
kept constant at 0.8 mL/min, and the carrier gas was helium. The
injector, oven, and detector temperatures were set to 230, 100, and
250°C, respectively. The total arabinoxylan content was calculated
according to this formula: total arabinoxylan = (% arabinose +
% xylose) × 0.88 (Henry 1986).
Polyphenol and Phytic Acid Contents. Samples were
extracted by shaking at room temperature with methanol/water
acidified with HCl (50:50 v/v, pH 2, 50 mL/g of sample, 60 min,
room temperature, constant shaking) and acetone/water (70:30 v/v,
50 mL/g of sample, 60 min, room temperature, constant shaking).
After centrifugation (15 min, 25°C, 3,000 × g), supernatants were
combined and used to determine extractable polyphenols. Ferulic
acid was used to prepare a standard curve. Extractable polyphenols
were determined by the Folin–Ciocalteu procedure (Singleton et al.
1999). The results were expressed as ferulic acid equivalents.
Hydrolyzable polyphenols comprise hydrolyzable tannins, phe-
nolic acids, and hydroxycinnamic acids that are released from the
food matrix by strong acidic hydrolysis. These compounds were
extracted by a methanol/18.4M H2SO4 90:10 (v/v) hydrolysis at
85°C for 20 h from the residues of methanol/acetone/water ex-
traction that was done for determination of soluble polyphenols
(Hartzfeld et al. 2002); after centrifugation (15 min, 25°C, 3,000 ×
g), supernatants were combined and used to determine the hydro-
lyzable polyphenols by the Folin–Ciocalteu method with a ferulic
acid standard curve (Singleton et al. 1999). The results were ex-
pressed as ferulic acid equivalents. Although the Folin–Ciocalteu
method has some limitations in regard to overestimation of phenolic
compounds owing to reaction with other components (nitrogenous
compounds, nonphenolic antioxidants, etc.) (Everette et al. 2010;
Sánchez-Rangel et al. 2013), the aim was to get an overall picture of
differences in phenolic content in the samples.
Phytic acid content of bread was determined according to the
method of Haug and Lantzsch (1983) with modifications by Guttieri
et al. (2006). Phytic acid was extracted with 0.2M hydrochloric acid
overnight. The extract was diluted, and the sample extracts and
standard solutions were boiled before the addition of ferric am-
monium chloride. After cooling on ice, the samples were added to
microplates along with 2,2-bipyridine-thioglcolic acid, and the ab-
sorbance was read at 530 nm (Guttieri et al. 2006). The phytic acid
content was determined by plotting the absorbance of the standard
curve against concentration.
Base Flour (%)z Bran (%) Starch (%)
100 0 0
100 0 0
100 0 0
74 26 0
86 0 14
65 22 13

Starch Digestibility. The Englyst in vitro assay was conducted
to determine the starch hydrolysis curves (Englyst et al. 1992). The
moisture of bread samples and the reference sample was measured
according to AACCI Approved Method 44-15.02. The samples
were weighed (0.3 g) into 50 mL plastic centrifuge tubes, and the
sample weight was adjusted for moisture content to a dry weight
basis. The bread samples were incubated at 37°C with an enzyme
mix (amyloglucosidase, invertase, and pancreatin) for 180 min. Ali-
quots of the digest were taken every 20 min to determine the amount
of glucose released by reaction with glucose oxidase/peroxidase
(GOPOD). A sample of commercial white bread (purchased from a
local grocery store and air dried at room temperature) was analyzed as
a reference. The hydrolysis index (HI) was obtained by dividing the
area under the hydrolysis curve of the sample by the area obtained for
commercial white bread (hydrolysis curve 0–180 min). The eGI of
the samples was calculated using the equation described by Ovando-
Martı́nez et al. (2011b): eGI = 8.198 + 0.862 × HI.
Owing to the high level of retrogradation of starch occurring in
baked bread, the amount of RS-III (retrograded starch) was de-
termined. The RS-III was measured using a modified enzymatic-
gravimetric determination of dietary fiber. After incubation at
100°C with thermostable a-amylase for 35 min, protease was added
to the samples for further incubation at 60°C for 35 min. Then, the
samples were treated with amyloglucosidase to hydrolyze the dex-
trans and physically inaccessible starch. The samples were centri-
fuged and the remaining pellets washed with distilled water, 96%
ethanol, and acetone. The pellets were dissolved in potassium hy-
droxide and hydrolyzed with amyloglucosidase. The glucose was
measured with GOPOD reagent (Saura-Calixto et al. 1993).
Amylose Content and Starch Molecular Weight. For de-
termination of apparent amylose content and starch molecular mass,
the starch was extracted from bread and flour blends following the
method of Simsek et al. (2013). The extracted starch was dissolved
in potassium hydroxide/urea solution and heated for 90 min at
100°C. The samples were then neutralized with hydrochloric acid
and filtered before analysis by high-performance size-exclusion
chromatography (HPSEC) with multiangle light scattering. The re-
fractive index increment (dn/dc value) for calculation of the starch
molecular mass was 0.146 (You and Lim 2000; Simsek et al. 2013).
The Debye model with a fit degree of one was used for calculation
of the molar mass. The results were fitted to a first-order polynomial
model.
Statistical Analysis. The results were analyzed with SAS 9.3
statistical analysis software package. Data were averaged across all
three varieties used for laboratory-milled flour for each treatment
(white, whole wheat, and whole wheat + starch), owing to few
significant differences between the three wheat variety flour sample
types. Analysis of variance was conducted using a completely
random design. The mean separation was conducted by Tukey’s
studentized range test with a = 0.05. Correlation coefficients were
calculated with Excel.
TABLE II
Dough and Bread Quality of White and Whole Wheat Breadsz
Farinograph
Absorption Peak Time Stability
Flour Type (%, 14% MB) (min) (min)
Commercial refined 61.35e 7.85ab 11.70b
Refined 64.52d 8.62a 14.50a
Commercial whole 65.60c 5.65c 10.20c
wheat
Whole wheat 71.50a 6.23bc 8.80d
Commercial whole 66.16b 5.80c 11.05bc
wheat + starch
Whole wheat + starch 71.64a 6.92abc 9.00d
z Values in the same column with the same letter are not significantly different (P > 0.05). MB = moisture basis; MTI = mixing tolerance index; and BU =
Brabender units.
19433638, 2017, 6, Downloaded from https://onlinelibrary.wiley.com/doi/10.1094/CCHEM-05-17-0116-R by Cnr Roma, Wiley Online Library on [16/01/2024]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
RESULTS AND DISCUSSION
Quality of White and Whole Wheat Dough and Breads.
Dough and bread quality parameters added are reported in Table II.
The white flours had significantly (P < 0.05) lower water absorption
(61.35 and 64.52%) than the whole wheat and whole wheat + starch.
The whole wheat and whole wheat + starch samples had absorp-
tions of 71.50 and 71.64%, respectively. The bran component
in the whole wheat flours absorbs water at a higher rate, owing to
the presence of arabinoxylans and other nonstarch polysaccha-
rides (D’Appolonia and Kunerth 1984). The whole wheat and
whole wheat + starch flours had significantly (P < 0.05) lower
dough strength than the white flour dough. The lower stability in
whole wheat is in part owing to the physical interference of the
protein matrix by the bran particles (Gonzalez-Gracia et al. 2012).
However, there is likely an interaction between the gluten matrix
and the components of bran, such as arabinoxylans, phenolic
compounds, and lipids (Khalid et al. in press). These interactions
during mixing have an effect on dough quality that carry over into
the end-product quality of the flour (Khalid et al. in press).
The bake absorption ranged from 62.65 to 73.05% and followed
the same trend as the farinograph absorption. As was expected
based on previous studies (Gan et al. 1992; Gonzalez-Gracia et al.
2012), the loaf volume of both whole wheat breads was significantly
(P < 0.05) lower than white bread. The addition of starch to the
whole wheat did not significantly (P > 0.05) affect the loaf volume
compared with the other whole wheat bread. These results show the
typical challenges with the physical quality of bread when adding
bran and germ to prepare whole wheat bread. However, along with
changes in the physical quality, the chemical composition and nu-
tritional quality of whole wheat breads are different from white
breads.
Microstructure of White and Whole Wheat Bread Crumb.
The crumb structure of bread is an important factor in evaluating
end-product quality of bread. In this study, SEM was used to take
images of the bread microstructure. The microstructure of the bread
may have had some influence on the digestibility of the starch in the
bread. The SEM images showing the bread microstructure are
shown in Figure 1. The most obvious difference between the mi-
crostructure of the white breads and the whole wheat breads is the
visibility of the starch granules. The starch granules in the white
bread can be seen far more clearly than in the whole wheat and
whole wheat + starch breads. The starch granules in the white bread
samples have become deformed and pitted by gelatinization and the
action of amylolytic enzymes. The degradation of the starch gran-
ules will allow for more rapid hydrolysis by human digestive en-
zymes when the bread is eaten.
The starch granules in all the bread samples were coated with a
matrix of protein and leached starch (Rojas et al. 2000). This matrix
could be seen in all of the bread samples in this study (Fig. 1). The
whole wheat and whole wheat + starch samples seemed to have a
Baking
MTI Quality Bake Absorption Mix Time Loaf Volume
(BU) Number (BU) (%, 14% MB) (min) (cm3)
28.50a 132.50b 62.65c 3.00b 1,072.50a
21.00c 171.17a 67.25b 3.29ab 1,028.33a
19.50c 135.50b 71.55a 3.00b 717.50b
24.17b 129.67b 72.98a 3.38a 731.67b
20.50c 140.50b 67.60b 3.13ab 707.50b
25.17b 134.33b 73.05a 3.33a 735.83b
Vol. 94, No. 6, 2017 993

thicker matrix coating the starch granules. The thick matrix in these
samples blocked the starch granules from view in the SEM im-
ages of the bread. The matrix in the whole wheat samples might be
thicker because of arabinoxylans that have leached from the bran
during the mixing and fermentation processes of baking. A thicker
matrix will act more strongly as a physical barrier to digestive
enzymes. There will also be a chemical barrier to starch diges-
tion if arabinoxylans are forming a portion of the matrix covering
the starch granules, because arabinoxylans are not hydrolyzed by
Fig. 1. Scanning electron microscopic images of white and whole wheat
bread crumb. Images taken at ×1,000 magnification. Boxes indicate starch
granules, and arrows indicate protein matrix.
994 CEREAL CHEMISTRY
19433638, 2017, 6, Downloaded from https://onlinelibrary.wiley.com/doi/10.1094/CCHEM-05-17-0116-R by Cnr Roma, Wiley Online Library on [16/01/2024]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
human digestive enzymes. Studies done by Bravo et al. (1998) and
Choct and Annison (1992) determined that wheat products con-
taining higher levels of arabinoxylans had slower starch diges-
tibility. Based on previous research and the SEM images in this
study, it is possible that the arabinoxylans formed a barrier to pre-
vent starch hydrolysis. The microstructure of the bread and possible
components making up these structures led us to investigate the
chemical composition of the bread in more detail to determine any
link between the chemical composition of the bread and its end-use
quality and starch digestibility.
Composition of White and Whole Wheat Flours and
Breads. Certain components in the flour may affect starch hy-
drolysis and digestibility. The composition of white and whole
wheat flours and breads is given in Table III. One minor but im-
portant component of wheat flour is the mineral content. Whole
wheat flours will have higher mineral content, because the majority
of the minerals in wheat are concentrated in the bran and germ
fraction (Anson 2010). The white flours had significantly (P < 0.05)
less ash (0.74 and 0.77%) than both the whole wheat flours (1.91
and 1.94%) and whole wheat + starch flours (1.90 and 1.92%).
Addition of other ingredients during baking resulted in higher ash
content in the bread samples. The ash contents of the bread samples
are significantly (P < 0.05) higher than the flours they were made
from but follow a similar trend. The white bread sample (1.51%)
had significantly (P < 0.05) lower ash content than the whole wheat
(2.53%) and whole wheat + starch breads (2.51%).
During milling and processing of wheat, a portion of the starch
can become damaged by mechanical action or gelatinization
(Prabhasankar and Rao 2001). Flours with high starch damage will
have higher water absorption, but the dough may overferment more
rapidly or the bread crumb may be sticky (Boyacı et al. 2004). The
damaged starch may also be more easily digested and affect the eGI
of the bread. The commercial whole wheat and commercial whole
wheat + starch samples had significantly (P < 0.05) lower starch
damage than the other flours. The difference in starch damage
between commercial whole wheat and the other flours was likely
owing to the method of flour milling. The starch damage increased
significantly (P < 0.05) in all samples after baking. This was caused
by the starch hydrolysis and gelatinization that occurred in the
baking process. The white breads had significantly (P < 0.05)
higher starch damage (20.80 and 18.81%) than the whole wheat and
whole wheat + starch breads. The starch damage ranged from 12.57
to 14.92% for whole wheat and whole wheat + starch breads. The
large increase in starch damage after baking of the white bread
samples may indicate that the starch in the white breads was more
susceptible to gelatinization and hydrolysis than the starch in the
whole wheat and whole wheat + starch breads. These main com-
ponents of wheat flour greatly affect the end-product and nutritional
quality. However, there are other components that may be minor but
can also have strong effects on the quality.
Arabinoxylans are considered a minor component of wheat, but
they have important effects on the bread. The majority of the dietary
fiber in wheat is comprised of arabinoxylans (Anson 2010). The
arabinoxylan content and arabinose-to-xylose ratio (A/X) of the
white and whole wheat flours and breads are shown in Table III.
The arabinoxylan content of the flour samples ranged from 2.56 to
11.04% and from 2.53 to 10.27% for the breads. The additional
arabinoxylan content in the whole wheat samples may affect the
sample viscosity during digestion and access to starch by enzymes,
which, in turn, could affect the eGI of the whole wheat breads
(Bravo et al. 1998; Choct and Annison 1992).
It is important to evaluate how the starch functional and chemical
properties are affecting end-product and nutritional quality. To fully
investigate the effects of starch on the bread quality and eGI, the
components and properties of the starch must be determined. The
proportion of amylose and other starch properties, such as molec-
ular mass, can have an influence on the end-product and nutritional
quality. The amylopectin and amylose contents of the starches

extracted from the flour and bread samples are presented in Table
III. The amylopectin and amylose contents of the white, whole
wheat, and whole wheat + starch flour samples are similar to each
other. All of the flours have amylose contents that are in the normal
range for native wheat starch. Native cereal starches typically have
around 25% amylose content (Eliasson 2004; Simsek et al. 2013).
After baking, the amylose content significantly (P < 0.05) de-
creased for all samples; however, a larger decrease in amylose
content was seen for the white bread sample. Because, during
starch gelatinization, amylose leaches from the starch granule first
(Eliasson 2004), the amylose will be more readily available for
hydrolysis than the amylopectin. The hydrolysis of amylose during
baking causes the bread samples to have a lower percentage of
amylose content. The white breads (19.34–20.34%) had signi-
ficantly (P < 0.05) lower amylose content than the whole wheat
breads (23.10–25.39%) and whole wheat + starch breads
(22.85–23.26%) (Table III). The reduction of amylose in the white
bread samples may indicate that the starch is more readily hy-
drolyzed and may have more rapid digestion in the human gut than
the starch in the whole wheat breads.
Whole grains, such as wheat, are a good source of antioxidants,
although the antioxidant capacity of whole wheat has typically been
underestimated (Slavin 2004; Pérez-Jiménez and Saura-Calixto 2005).
Phytic acid is one antioxidant compound found in wheat that acts
as a chelating agent that binds various metals and suppresses iron-
catalyzed redox reactions (Slavin 2004). The phytic acid content of
the white and whole wheat flour and bread samples is reported in
Table IV. The phytic acid contents of the white, whole wheat, and
whole wheat + starch flours ranged from 12.28 to 20.92 mg/g. The
white flour had significantly (P < 0.05) lower phytic acid content
than the whole wheat and whole wheat + starch flours.
The phytic acid content of all the bread samples was significantly
(P < 0.05) lower than the original flour samples they were produced
from. The most drastic reduction in phytic acid content occurred in
the white bread samples. There was an approximately 98% re-
duction of phytic acid in white bread (0.18–0.24 mg/g) compared
with the white flour (12.28–12.51 mg/g). Although the whole wheat
and whole wheat + starch breads had significantly (P < 0.05) lower
phytic acid than the whole wheat and whole wheat + starch flours,
the decrease after baking was about 2–3 mg/g. The higher phytic
acid content of the whole wheat breads may reduce starch hydro-
lysis and digestibility by binding Ca2+, which would inhibit amylase
activity (Thompson and Yoon 1984; Haros et al. 2001).
Table IV also presents the levels of extractable and hydrolyzable
phenolic compounds in the flour and bread samples. There were
19433638, 2017, 6, Downloaded from https://onlinelibrary.wiley.com/doi/10.1094/CCHEM-05-17-0116-R by Cnr Roma, Wiley Online Library on [16/01/2024]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
significant (P < 0.05) differences between the hydrolyzable phe-
nolic contents of white and whole wheat flours. The hydrolyzable
phenolic content of the white flours was 6.78 and 6.77 mg/g.
The whole wheat (12.03–12.33 mg/g) and whole wheat + starch
(12.29–12.61 mg/g) flours had about twice as much hydrolyzable
phenolic compounds as the white flours. One of the most common
phenolic compounds in wheat, ferulic acid, is found bound to the
arabinoxylan in wheat, making it insoluble until hydrolysis by gut
microflora (Slavin 2004).
The extractable phenolic compound content was significantly
(P < 0.05) reduced after baking bread for all samples. The largest
reduction in extractable phenolic compounds was for the white
bread. The whole wheat and whole wheat + starch breads had sig-
nificantly (P < 0.05) higher extractable phenolic content than the
white bread. The higher level of extractable phenolic compounds
in the whole wheat breads may be owing to hydrolysis of these
compounds from the arabinoxylans during the fermentation process
of baking. Conversely, the hydrolyzable phenolic compound con-
tent was significantly (P < 0.05) higher in the bread samples than
the flours they were prepared from. In the case of the hydrolyzable
phenolic compounds, the white bread had a larger increase than the
whole wheat and whole wheat + starch breads. A study done by
Pérez-Jiménez and Saura-Calixto (2005) showed a substantial
increase in the phenolic content of wheat flour and other cereal
products after treatment with digestive enzymes. The enzyme hy-
drolysis that occurs during baking could be increasing the detect-
able levels of the hydrolyzable phenolic compounds in the bread.
The levels of phytic acid and phenolic compounds found in bread
are important not only for their antioxidant activity but also for their
effect in slowing starch digestibility (Thompson and Yoon 1984).
Phenolic compounds that are found in wheat, such as ferulic acid,
p-coumaric acid, vanillic acid, and hydroxybenzoic acid, were
found to reduce hydrolysis of wheat starch by a-amylase and amy-
loglucosidase. These phenolic compounds can bind with the starch
and the amylolytic enzymes to reduce starch hydrolysis (Kandil et al.
2012).
Starch Molecular Weight. The molecular mass is another
important characteristic of starch that could impact end-product and
nutritional quality. Starch molecular mass can vary based on genetic
and environmental differences in a sample (Simsek et al. 2013). The
variation in starch molecular mass can alter wheat starch swell-
ing and pasting characteristics (Shibanuma et al. 1996; Sasaki
and Matsuki 1998), which may influence end-product quality.
The weight averaged molecular masses (Mw) of amylopectin and
amylose in the flour and bread samples are shown in Figure 2.

TABLE III
Composition of White and Whole Wheat Flours and Breadsz
Ash Protein Starch Damage AX A/X Amylopectin Amylose
Sample Type (% DWB) (% DWB) (%) (% DWB) Ratio (%) (%)
Flours
Commercial refined 0.77f 15.12f 7.15e 2.84f 0.95a 74.60f 25.40a
Refined 0.74f 16.18d 7.88e 2.56f 0.88abc 74.48f 25.52a
Commercial whole wheat 1.94b 16.41c 3.38f 11.04a 0.89abc 75.04def 24.96abc
Whole wheat 1.91bc 17.00a 7.28e 9.01cd 0.93ab 75.37de 24.63bc
Commercial whole wheat + 1.92bc 16.14d 3.35f 10.39ab 0.89abc 75.73d 24.27c
starch
Whole wheat + starch 1.90c 16.78b 6.88e 9.71bc 0.92abc 74.87ef 25.13ab
Breads
Commercial refined 1.56d 14.93g 20.80a 2.96f 0.83c 80.66a 19.34f
Refined 1.51e 15.85e 18.81b 2.53f 0.83bc 79.66b 20.34e
Commercial whole wheat 2.52a 16.16d 13.24d 10.27ab 0.87abc 74.61f 25.39a
Whole wheat 2.53a 16.76b 14.92c 9.92b 0.90abc 76.90c 23.10d
Commercial whole wheat + 2.51a 16.12d 12.57d 7.97e 0.94a 77.15c 22.85d
starch
Whole wheat + starch 2.51a 16.50c 13.40d 8.74de 0.89abc 76.74c 23.26d
z Values in the same column with the same letter are not significantly different (P > 0.05). DWB = dry weight basis; AX = arabinoxylan; and A/X = arabinose to
xylose.

Vol. 94, No. 6, 2017 995


There were no significant differences (P > 0.05) in the Mw of
amylopectin for any of the flours that were from the same source.
The Mw of the amylopectin from the commercially milled flours
was significantly (P < 0.05) lower than the amylopectin from the
experimentally milled flours. The Mw of the amylopectin in the flour
samples was about 14–15 million. Amylopectin is one of the largest
known biopolymers, and as such the large molecular mass of the
amylopectin in this study is within the acceptable range for amy-
lopectin (Eliasson 2004; Gidley et al. 2010).
Although there was no difference in Mw of the amylopectin for
white or whole wheat flours, a significant (P < 0.05) reduction in
Mw was seen for the amylopectin in the bread samples. The amy-
lopectin from the white breads (0.92 × 107 and 1.04 × 107) had
significantly (P < 0.05) lower Mw than the amylopectin in the whole
wheat breads (1.23 × 107 and 1.27 × 107) and whole wheat + starch
breads (1.25 × 107 and 1.27 × 107). The reduction in molecular mass
of the amylopectin in the bread samples is related to the level of
starch hydrolysis that occurred during fermentation and baking of
the bread. These results indicate that the amylopectin in white bread
is more easily hydrolyzed than the amylopectin in whole wheat
breads. There must be some component present in the whole wheat
flour that is lacking in the white flour that prevents the starch hy-
drolysis. This may be affecting the end-product quality by lowering
the amount of starch hydrolyzed for yeast metabolism and gas
production. However, it may have some benefit to the nutritional
quality of the bread. The prevention of starch hydrolysis in the
whole wheat breads may lead to slower starch digestion in the
human digestive tract.
Amylose has a smaller molecular mass than amylopectin, but it
has significant functionality in wheat flour. The amylose molecule
provides for a significant amount of the structure and texture of
bread. Bread made from wheat flour that has low or no amylose
will have a poor texture and may collapse (Hung et al. 2007). The
molecular mass of amylose in wheat flour may also affect the end-
product and nutritional quality. The Mw of amylose in white and
whole wheat flour and bread samples is shown in Figure 2. As seen
in the molecular mass of amylopectin, there were fewer significant
(P < 0.05) differences in the molecular mass of amylose in the flour
samples. The amylose Mw of the commercial refined flour and re-
fined flour samples was 7.40 × 106 and 7.20 × 106, respectively,
whereas the Mw of the amylose in the commercial whole wheat
flour, whole wheat flour, commercial whole wheat flour + starch,
and whole wheat flour + starch flours was 5.73 × 106, 7.21 × 106,
6.53 × 106, and 7.11 × 106, respectively.
The Mw of amylose in the bread samples was all significantly
(P < 0.05) lower than the amylose in the flours from which they
were prepared. As was the case with amylopectin, the white breads
TABLE IV
Phytic Acid and Phenolic Contents of Flours and Breadsz
Sample Type Phytic Acid (mg/g)
Flours
Commercial refined 12.51e
Refined 12.28e
Commercial whole wheat 19.70b
Whole wheat 20.92a
Commercial whole wheat + starch 19.79b
Whole wheat + starch 20.90a
Breads
Commercial refined 0.18f
Refined 0.24f
Commercial whole wheat 18.61c
Whole wheat 17.89d
Commercial whole wheat + starch 18.26cd
Whole wheat + starch 18.32cd
z Values in the same column with the same letter are not significantly different (P > 0.05).
996 CEREAL CHEMISTRY
19433638, 2017, 6, Downloaded from https://onlinelibrary.wiley.com/doi/10.1094/CCHEM-05-17-0116-R by Cnr Roma, Wiley Online Library on [16/01/2024]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
also had significantly (P < 0.05) lower molecular mass of amylose
than the whole wheat and whole wheat + starch breads from the
same source. However, there was less change in the molecular mass
of amylose in all samples between the flours and the breads than
was seen with the molecular mass of the amylopectin. The reason
for this is the manner in which and rate at which each of these
molecules undergoes hydrolysis. When hydrolysis of amylose oc-
curs at the ends, the reduction in Mw will be less drastic than when
amylose is hydrolyzed toward the center of the molecule. If the
amylopectin has few molecules of glucose hydrolyzed from
the ends of the branches it will retain most of its molecular mass.
The branching of amylopectin can inhibit complete hydrolysis of
the molecule. However, if the amylopectin becomes debranched,
the molecular mass will decrease rapidly (Eliasson 2004). Also,
portions of the hydrolyzed amylopectin may be retained in the
HPSEC system along with amylose. This could affect the Mw of the
starch molecules eluting in the amylose fraction (Simsek et al.
2013). Starch gelatinization and degradation were not complete in
white or whole wheat breads. Whole wheat breads, which contain
higher levels of nonstarch polysaccharides (arabinoxylan) from cell
wall components, may have even lower rates of starch gelatiniza-
tion and degradation, which could limit starch digestibility by
allowing the native starch structure to persist (Mishra et al. 2012).
Starch Digestibility. Classically, starch is separated into three
classes based on the rate of digestibility. These are rapidly digestible
starch (RDS), slowly digestible starch (SDS), and RS. Also, there
are several types of RS, which are classified based on their mech-
anism of resistance (Sajilata et al. 2006). In this study we focused on
the total RS content and RS-III. RS-III is classified as starch that
is composed of retrograded starch, most of which is retrograded
amylose (Sajilata et al. 2006) and is very thermostable (Eerlingen
and Delcour 1995). Table V shows the RDS, SDS, RS, and RS-III
contents of the bread samples. The RDS content of the commercial
refined and refined breads was 40.81 and 40.59%, which was sig-
nificantly (P < 0.05) higher than the RDS content of the whole
wheat (36.12 and 35.70%) and whole wheat + starch (36.26 and
35.58%) breads. The white breads also had significantly (P < 0.05)
higher SDS than the whole wheat and whole wheat + starch breads.
The high RDS content may indicate that the starch in the white
bread is more easily digestible than the starch in whole wheat bread.
Overall, the RS and RS-III contents of the breads were quite low.
This was expected, because bread generally has a high glycemic
response. Typical levels of RS in bread range from 0.5 to 2%. The
RS content of the breads in this study ranged from 1.68 to 3.00%.
The RS content of white breads was significantly (P < 0.05) lower
than the RS content of the whole wheat and whole wheat + starch
breads. By determination of RS-III, it can be seen that the RS
Phenolic Compounds (mg/g)
Extractable Hydrolyzable
3.82a 6.78d
4.16a 6.77d
4.26a 12.33b
4.06a 12.03b
4.24a 12.61b
4.14a 12.29b
1.45c 10.16c
1.45c 9.77c
2.23b 13.75a
2.10b 13.83a
2.16b 14.05a
2.13b 13.81a

content of the bread samples was composed mostly (if not com-
pletely) of RS-III. The RS-III content of the bread samples was
opposite of the RS content. The white breads had 1.47 and 1.23%
RS-III, which was significantly (P < 0.05) higher than the RS-III
contents of the whole wheat (0.90 and 0.97%) and whole wheat +
starch (0.91 and 0.99%) breads. The RS-III, which is retrograded
starch, was formed during the storage of the bread prior to lyoph-
ilization as part of the staling process.
GI and starch digestibility are important nutritional factors in
high-starch foods such as bread. The GI refers to the postprandial
glycemic response of a test product compared with that of a ref-
erence food (glucose or white bread) (Björck et al. 1994; Augustin
et al. 2002; Slavin 2004). Wheat breads are typically high-GI
foods, but whole wheat breads often have lower GI than white
flour bread (McKevith 2004; Anson 2010). The starch hydrolysis
properties of the white and whole wheat bread samples are shown
in Table V. The HI is determined by the rate of starch hydrolysis in
the target food compared with the rate of starch hydrolysis in a
reference food (Goñi et al. 1997). The HI of the breads in this
study ranged from 87.23 to 98.51. It is well known that starch in
white bread is easily hydrolyzed and may give a spike in glucose
response (Slavin 2004). The HI of the whole wheat and whole
wheat + starch breads was significantly (P < 0.05) lower than for
the white bread. Starch was added to whole wheat flour to coun-
teract the effect of the difference in starch content between white
and whole wheat bread. The results of this study show that there
were no significant differences (P > 0.05) between the HI of the
19433638, 2017, 6, Downloaded from https://onlinelibrary.wiley.com/doi/10.1094/CCHEM-05-17-0116-R by Cnr Roma, Wiley Online Library on [16/01/2024]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
whole wheat and whole wheat + starch breads. Because of this, we
can assume that the lower starch content of whole wheat bread was
not the factor that results in lower HI. When conducting in vitro
starch hydrolysis assays, the HI can be used to calculate the eGI.
The eGI of the bread samples in this study followed a similar
trend as the HI (Table V). The white bread had significantly (P <
0.05) higher eGI than the whole wheat and whole wheat + starch
breads. The eGI ranged from 83.46 to 93.11 for the breads in
this study. The difference in eGI of the whole wheat and whole
wheat + starch breads was not significant (P > 0.05). As stated
previously, starch was added to whole wheat breads to eliminate the
effect of starch content on starch hydrolysis in the bread samples.
Bravo et al. (1998) determined a starch digestion rate index of about
90 in a whole wheat bread sample. This was slightly higher than the
HI and eGI of the whole wheat breads in this study. The results of
this study show that although the whole wheat and whole wheat +
starch breads were still in the high-GI category, their eGI was still
significantly lower than in the white breads. This might be beneficial
to consumers who want to continue consuming bread products but
desire a more health-conscious option.
Because it was determined that the lower starch content in whole
wheat bread does not decrease the eGI, there must be some other
component of the whole wheat bread resulting in the lower eGI.
From the results of the chemical analysis of the white and whole
wheat flours and breads, the components that may be responsible for
lowering the eGI in whole wheat bread may be determined. When
examining the bread crumb structure by SEM (Fig. 1) a thicker

Fig. 2. Molecular mass (A) and change in molecular mass (B) of amylopectin and amylose in white and whole wheat flour and bread samples. Mw =
weight averaged molecular mass; DMw = change in weight averaged molecular mass between flour and baked bread; CRF = commercial refined flour;
RF = refined flour; CWWF = commercial whole wheat flour; WWF = whole wheat flour; CWWF+S = commercial whole wheat flour + starch; WWF+S =
whole wheat flour + starch. Error bars represent ±standard deviation. Columns with the same pattern with the same letter are not significantly different (P > 0.05).

Vol. 94, No. 6, 2017 997


matrix can be seen coating the starch granules in the whole wheat
bread. This matrix will act more strongly as a physical barrier to
digestive enzymes and could reduce the eGI. The matrix covering
the starch granules is composed of the gluten matrix and possibly
arabinoxylan. Both of these components have been shown to result
in a reduction in starch hydrolysis. Studies by Bravo et al. (1998)
and Choct and Annison (1992) determined that wheat products
containing higher levels of arabinoxylans had slower starch di-
gestibility. There was also a strong highly significant (P < 0.001)
negative correlation (Table VI) between the eGI and arabinoxylan
content of the breads in our study, which shows a strong relationship
between arabinoxylan content and starch digestibility. However, the
arabinoxylan is a marker for the presence of the bran in the whole
wheat breads, and the reduction of eGI is likely owing to a com-
bination of more complex factors. Jenkins et al. (1987) determined
that bread that had the native protein–starch interaction remaining
intact had lower glycemic response and in vitro starch digestion than
bread made from starch and protein that had been extracted from
each other. Based on their previous research and the SEM images in
this study, it is possible that the arabinoxylans and the gluten protein
matrix are forming a barrier to prevent starch hydrolysis.
Phytic acid and phenolic compounds are other relatively minor
components of wheat flour and bread, but they may influence the
starch hydrolysis and eGI of the whole wheat bread samples (Table
VI). The eGI had strong highly significant (P < 0.001) correlations
(–0.977, –0.830, and –0.924) with phytic acid, extractable pheno-
lics, and hydrolyzable phenolics, respectively. In the case of phytic
acid, there was an inverse relationship with the eGI of the bread
19433638, 2017, 6, Downloaded from https://onlinelibrary.wiley.com/doi/10.1094/CCHEM-05-17-0116-R by Cnr Roma, Wiley Online Library on [16/01/2024]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
samples. The whole wheat bread samples, which had significantly
(P < 0.05) higher phytic acid levels (Table IV), had significantly
(P < 0.05) lower eGI than the white breads. The phenolic com-
pounds measured in the bread samples (Table IV) had the same
inverse trend as the phytic acid. Thompson and Yoon (1984) found
that starch hydrolysis was inhibited by the presence of phytic acid
and some phenolic compounds. It is reasonable to presume that the
higher levels of phytic acid and phenolic compounds in the whole
wheat samples are affecting the eGI of the whole wheat breads.
The differences in eGI between the white and whole wheat breads
can also be examined by comparing the properties of the starch in
these samples. The amount of amylose in the bread starches was
measured, and the white breads had significantly (P < 0.05) less
amylose than the whole wheat breads (Table III). The reduction of
amylose in the white bread samples may indicate that the starch is
more readily hydrolyzed and may have more rapid digestion in the
human gut than the starch in the whole wheat breads. Changes in
molecular mass of both amylopectin and amylose in the bread
samples also allude to differences in starch hydrolysis between the
white and whole wheat breads. There are strong highly significant
(P < 0.001) correlations between eGI and the change in molecular
weight of amylopectin (0.697) and amylose (0.929). The molecular
masses of the amylopectin and amylose were significantly (P <
0.05) lower in the white bread samples than in the whole wheat
bread samples (Fig. 2). The bread samples with higher eGI also
tended to have starch with lower molecular mass. The prevention of
starch hydrolysis in the whole wheat breads may lead to slower
starch digestion in the human digestive tract.

TABLE V
Starch Hydrolysis Properties of White and Whole Wheat Breadsz
Sample Type RDS (% DWB) SDS (% DWB) RS (% DWB) RS-III (% DWB) HI eGI
Commercial refined 40.81a 26.42a 1.92ab 1.47a 98.51a 93.11a
Refined 40.59a 25.97a 1.68b 1.23b 98.01a 92.68a
Commercial whole wheat 36.12b 17.33c 2.79ab 0.90c 88.74bc 84.69bc
Whole wheat 35.70b 17.62bc 2.32ab 0.97c 87.32d 83.46d
Commercial whole wheat + starch 36.26b 19.27b 3.00a 0.91c 89.23b 85.11b
Whole wheat + starch 35.58b 19.11b 2.24ab 0.99c 87.97cd 84.02cd
z Values in the same column with the same letter are not significantly different (P > 0.05). DWB = dry weight basis; RDS = rapidly digestible starch; SDS = slowly
digestible starch; RS = resistant starch; RS-III = type III resistant starch; HI = hydrolysis index; and eGI = estimated glycemic index.

TABLE VI
Correlation Between Composition of White and Whole Wheat Breads and Starch Digestibilityz
Parameter Amylopectin DMw Amylose DMw RDS SDS RS RS-III HI eGI
Ash –0.630*** –0.953*** –0.975*** –0.902*** 0.609** –0.445* –0.978*** –0.978***
Protein 0.371 –0.262 –0.330 –0.603** 0.011 –0.332 –0.272 –0.272
Starch damage 0.688*** 0.885*** 0.880*** 0.794*** –0.614** 0.317 0.884*** 0.884***
Starch 0.428* 0.897*** 0.932*** 0.977*** –0.547** 0.363 0.914*** 0.914***
Arabinoxylan –0.558** –0.913*** –0.948*** –0.929*** 0.554** –0.480* –0.946*** –0.946***
A/X –0.650*** –0.555** –0.591** –0.388 0.369 –0.226 –0.597** –0.597**
Phytic acid –0.636*** –0.947*** –0.982*** –0.887*** 0.599** –0.505* –0.977*** –0.977***
Extractable phenolics –0.766*** –0.784*** –0.811*** –0.570** 0.612** –0.395 –0.830*** –0.830***
Hydrolyzable phenolics –0.506* –0.902*** –0.941*** –0.914*** 0.572** –0.518** –0.924*** –0.924***
Amylopectin % –0.158 0.122 0.145 0.371 –0.150 0.056 0.133 0.133
Amylose % 0.158 –0.122 –0.145 –0.371 0.150 –0.056 –0.133 –0.133
Amylopectin Mw –0.229 –0.371 –0.409 –0.565** 0.406* –0.353 –0.436* –0.436*
Amylose Mw –0.515** –0.327 –0.284 –0.329 0.491* –0.221 –0.376 –0.376
Amylopectin DMw 1.000 0.664*** 0.621** 0.338 –0.565** 0.373 0.697*** 0.697***
Amylose DMw 0.664*** 1.000 0.924*** 0.846*** –0.620** 0.364 0.929*** 0.929***
RDS 0.621** 0.924*** 1.000 0.849*** –0.555** 0.494 0.979*** 0.979***
SDS 0.338 0.846*** 0.849*** 1.000 –0.585** 0.262 0.840*** 0.840***
RS –0.565** –0.620** –0.555** –0.585** 1.000 –0.156 –0.588** –0.588**
RS-III 0.373 0.364 0.494* 0.262 –0.156 1.000 0.489* 0.489*
HI 0.697*** 0.929*** 0.979*** 0.840*** –0.588** 0.489* 1.000 1.000
eGI 0.697*** 0.929*** 0.979*** 0.840*** –0.588** 0.489* 1.000 1.000
z RDS = rapidly digestible starch; SDS = slowly digestible starch; RS = resistant starch; RS-III = type III resistant starch; HI = hydrolysis index; eGI = estimated
glycemic index; A/X = arabinose-to-xylose ratio; Mw = weight averaged molecular weight; and DMw = change in weight average molecular weight. *, ** and ***
represent significance at 0.05, 0.01, and 0.001, respectively.

998 CEREAL CHEMISTRY


CONCLUSIONS
When examining the starch hydrolysis properties and eGI of these
bread samples along with the results of the physicochemical anal-
ysis of the flour and bread samples, several interesting conclusions
can be made. This study was focused on the chemical and physical
differences between white and whole wheat breads that may affect
the eGI of the breads. The results of this research show that the
lower starch content in whole wheat bread does not decrease the
eGI, so there must be some other component of the whole wheat
bread resulting in the lower eGI. Because the dilution of starch in
the whole wheat bread was determined not to be the factor causing
lower eGI in whole wheat breads, the relationships between eGI and
the composition of the bread samples was investigated. Several
connections between the eGI and the composition of the bread
samples were found. There was a larger increase in starch damage
of white bread after baking compared with whole wheat bread.
Starch molecular weight also decreased after baking to a greater
degree in white bread compared with whole wheat breads. These
changes in starch indicate that the starch in white bread undergoes
more hydrolysis during fermentation and baking and may be more
easily hydrolyzed during digestion. The lower amount of starch
hydrolysis may also be affecting the end-product quality by re-
ducing the glucose available for yeast metabolism and gas pro-
duction. However, prevention of starch hydrolysis in the whole
wheat breads may lead to slower starch digestion in the human
digestive tract.
On the whole, the end-product quality of whole wheat bread may
be somewhat lower than white bread; however, the nutritional
quality of whole wheat bread may outweigh these faults in end-
product appearance. The results of this study show that although the
whole wheat breads are still in the high-GI category, their eGI is still
significantly (P < 0.05) lower than in white bread. Also, there are
several components of the whole wheat contributing to the reduction
of the eGI in whole wheat bread. This might be beneficial to con-
sumers who want to continue consuming bread products but desire a
more health-conscious option.
LITERATURE CITED
AACC International. Approved Methods of Analysis, 11th Ed. Method 08-
01.01. Ash—Basic method. Approved April 13, 1961. Method 10-05.01.
Guidelines for measurement of volume by rapeseed displacement. Ap-
proved October 17, 2001. Method 10-09.01. Basic straight-dough bread-
baking method—Long fermentation. Approved November 8, 1995. Method
44-15.02. Moisture—Air-oven methods. Approved October 30, 1975.
Method 46-30.01. Crude protein—Combustion method. Approved No-
vember 8, 1995. Method 54-21.02. Rheological behavior of flour by far-
inograph: Constant flour weight procedure. Approved November 8, 1995;
revised and approved January 6, 2011. Method 76-30.02. Determination of
damaged starch. Approved May 1, 1969. Available online only. AACCI:
St. Paul, MN.
American Diabetes Association. 2013. Glycemic index and diabetes.
www.diabetes.org/food-and-fitness/food/what-can-i-eat/understanding-
carbohydrates/glycemic-index-and-diabetes.html?loc=ff-slabnav
Anson, N. M. 2010. Bioactive Compounds in Whole Grain Wheat.
Doctoral thesis. Maastricht University: Maastricht, the Netherlands.
Augustin, L. S., Franceschi, S., Jenkins, D. J., Kendall, C. W., and
La Vecchia, C. 2002. Glycemic index in chronic disease: A review. Eur.
J. Clin. Nutr. 56:1049-1071.
Björck, I., Granfeldt, Y., Liljeberg, H., Tovar, J., and Asp, N. G. 1994.
Food properties affecting the digestion and absorption of carbohy-
drates. Am. J. Clin. Nutr. 59:699S-705S.
Blakeney, A. B., Harris, P. J., Henry, R. J., and Stone, B. A. 1983. A
simple and rapid preparation of alditol acetates for monosaccharide
analysis. Carbohydr. Res. 113:291-299.
Boyacı, I. H., Williams, P. C., and Köksel, H. 2004. A rapid method for
the estimation of damaged starch in wheat flours. J. Cereal Sci. 39:
139-145.
Bravo, L., Englyst, N., and Hudson, J. 1998. Nutritional evaluation of
carbohydrates in the Spanish diet: Non-starch polysaccharides and in
19433638, 2017, 6, Downloaded from https://onlinelibrary.wiley.com/doi/10.1094/CCHEM-05-17-0116-R by Cnr Roma, Wiley Online Library on [16/01/2024]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
vitro starch digestibility of breads and breakfast products. Food Res.
Int. 31:129-135.
Choct, M., and Annison, G. 1992. The inhibition of nutrient digestion by
wheat pentosans. Br. J. Nutr. 67:123-132.
Chung, S. Y., Lee, S. W., and Rhee, C. 2011. Effects of various Maillard
reaction products on in vitro starch hydrolysis and blood glucose re-
sponses in mice. Starch/Stärke 63:443-449.
D’Appolonia, B. L., and Kunerth, W. H. 1984. The farinograph handbook.
American Association of Cereal Chemists: St. Paul MN.
Eerlingen, R. C., and Delcour, J. A. 1995. Formation, analysis, structure
and properties of type III enzyme resistant starch. J. Cereal Sci. 22:
129-138.
Eliasson, A. C., ed. 2004. Starch in Food: Structure, Function and Ap-
plications. CRC Press: Boca Raton, FL.
Englyst, H. N., Kingman, S. M., and Cummings, J. H. 1992. Classifica-
tion and measurement of nutritional important starch fractions. Eur. J.
Clin. Nutr. 46:S33-S50.
Everette, J. D., Bryant, Q. M., Green, A. M., Abbey, Y. A., Wangila,
G. W., and Walker, R. B. 2010. Thorough study of reactivity of various
compound classes toward the Folin–Ciocalteu reagent. J. Agric. Food
Chem. 58:8139-8144.
Gan, Z., Galliard, T., Ellis, P. R., Angold, R. E., and Vaughan, J. G. 1992.
Effect of the outer bran layers on the loaf volume of wheat bread.
J. Cereal Sci. 15:151-163.
Gidley, M. J., Hanashiro, I., Hani, N. M., Hill, S. E., Huber, A., Jane, J. L.,
Liu, Q., Morris, G. A., Rolland-Sabate, A., Striegel, A. M., and Gilbert,
R. G. 2010. Reliable measurements of the size distributions of starch
molecules in solution: Current dilemmas and recommendations. Car-
bohydr. Polym. 79:255-261.
Goñi, I., Garcia-Alonso, A., and Saura-Calixto, F. 1997. A starch hy-
drolysis procedure to estimate glycemic index. Nutr. Res. 17:427-437.
Gonzalez-Gracia, G., Schlepp, E., and Simsek, S. 2012. Effects of wheat
bran color and particle size one whole wheat bread baking quality.
AACC International Annual Meeting, Hollywood, FL.
Guttieri, M. J., Peterson, K. M., and Souza, E. J. 2006. Agronomic per-
formance of low phytic acid wheat. Crop Sci. 46:2623-2629.
Haros, M., Rosell, C. M., and Benedito, C. 2001. Use of fungal phytase to
improve breadmaking performance of whole wheat bread. J. Agric.
Food Chem. 49:5450-5454.
Hartzfeld, P. W., Forkner, R., Hunter, M. D., and Hagerman, A. E. 2002.
Determination of hydrolyzable tannins (gallotannins and ellagitannins)
after reaction with potassium iodate. J. Agric. Food Chem. 50:1785-1790.
Haug, W., and Lantzsch, H. 1983. Sensitive method for the rapid de-
termination of phytate in cereals and cereal products. J. Sci. Food
Agric. 34:1423-1426.
Henry, R. J. 1986. Genetic and environmental variation in the pentosan
and b-glucan contents of barley, and their relation to malting quality.
J. Cereal Sci. 4:269-277.
Holm, J., and Björck, I. 1992. Bioavailability of starch in various wheat-
based bread products: Evaluation of metabolic responses in healthy
subjects and rate and extent of in vitro starch digestion. Am. J. Clin.
Nutr. 55:420-429.
Hung, P. V., Maeda, T., and Morita, N. 2007. Dough and bread qualities of
flours with whole waxy wheat flour substitution. Food Res. Int. 40:
273-279.
Jenkins, D. J., Thorne, M. J., Wolever, T. M., Jenkins, A. L., Rao, A. V.,
and Thompson, L. U. 1987. The effect of starch-protein interaction in
wheat on the glycemic response and rate of in vitro digestion. Am.
J. Clin. Nutr. 45:946-951.
Johansson, C. G., Siljeström, M., and Asp, N. G. 1984. Dietary fibre in
bread and corresponding flours—Formation of resistant starch during
baking. Z. Lebensm. Unters. Forsch. 179:24-28.
Kandil, A., Li, J., Vasanthan, T., and Bressler, D. C. 2012. Phenolic acids
in some cereal grains and their inhibitory effect on starch liquefaction
and saccharification. J. Agric. Food Chem. 60:8444-8449.
Khalid, K. H., Ohm, J.-B., and Simsek, S. In press. Whole wheat bread:
Effect of bran fractions on dough and end-product quality. J. Cereal
Sci. doi:10.1016/j.jcs.2017.03.011
Liljeberg, H., Åkerberg, A., and Björck, I. 1996. Resistant starch for-
mation in bread as influenced by choice of ingredients or baking
conditions. Food Chem. 56:389-394.
Lovegrove, A., Edwards, C., De Noni, I., Patel, H., El, S., Grassby, T.,
Zielke, C., Ulmius, M., Nilsson, L., Butterworth, P., Ellis, P., and
Shewry, P. 2017. Role of polysaccharides in food, digestion and health.
Crit. Rev. Food Sci. Nutr. 57:237-253.
Vol. 94, No. 6, 2017 999

McKevith, B. 2004. Nutritional aspects of cereals. Nutr. Bull. 29:111-142.
Mishra, S., Hardacre, A., and Monro, J. 2012. Food structure and car-
bohydrate digestibility. In: Carbohydrates—Comprehensive Studies on
Glycobiology and Glycotechnology. C.-F. Chang, ed. InTech: Rijeka,
Croatia.
Ovando-Martı́nez, M., Bello-Pérez, L. A., Whitney, K., Osorio-Dı́az, P.,
and Simsek, S. 2011a. Starch characteristics of bean (Phaseolus vul-
garis L.) grown in different localities. Carbohydr. Polym. 85:54-64.
Ovando-Martı́nez, M., Osorio-Dı́az, P., Whitney, K., Bello-Pérez, L. A.,
and Simsek, S. 2011b. Effect of the cooking on physicochemical and
starch digestibility properties of two varieties of common bean (Pha-
seolus vulgaris L.) grown under different water regimes. Food Chem.
129:358-365.
Pérez-Jiménez, J., and Saura-Calixto, F. 2005. Literature data may un-
derestimate the actual antioxidant capacity of cereals. J. Agric. Food
Chem. 53:5036-5040.
Prabhasankar, P., and Rao, P. H. 2001. Effect of different milling methods
on chemical composition of whole wheat flour. Eur. Food Res. Tech-
nol. 213:465-469.
Rojas, J. A., Rosell, C. M., Benedito de Barber, C., Pérez-Munuera, I.,
and Lluch, M. A. 2000. The baking process of wheat rolls followed by
cryo scanning electron microscopy. Eur. Food Res. Technol. 212:
57-63.
Sajilata, M. G., Singhal, R. S., and Kulkarni, P. R. 2006. Resistant
starch—A review. Compr. Rev. Food Sci. Food Saf. 5:1-17.
Sánchez-Rangel, J. C., Benavides, J., Heredia, J. B., Cisneros-Zevallos,
L., and Jacobo-Velázquez, D. A. 2013. The Folin–Ciocalteu assay
revisited: Improvement of its specificity for total phenolic content
determination. Anal. Methods 5:5990-5999.
[Received May 30, 2017. Accepted August 31, 2017.]
1000 CEREAL CHEMISTRY
19433638, 2017, 6, Downloaded from https://onlinelibrary.wiley.com/doi/10.1094/CCHEM-05-17-0116-R by Cnr Roma, Wiley Online Library on [16/01/2024]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
Sasaki, T., and Matsuki, J. 1998. Effect of wheat starch structure on
swelling power. Cereal Chem. 75:525-529.
Saura-Calixto, F., Goñi, I., Bravo, L., and Mañas, E. 1993. Resistant
starch in foods: Modified method for dietary fiber residues. J. Food Sci.
58:642-643.
Shibanuma, Y., Takeda, Y., and Hizukuri, S. 1996. Molecular and pasting
properties of some wheat starches. Carbohydr. Polym. 29:253-261.
Simsek, S., Whitney, K., and Ohm, J. B. 2013. Analysis of cereal starches
by high-performance size exclusion chromatography. Food Anal.
Methods 6:184-190.
Singleton, V. L., Orthofer, R., and Lamuela-Raventos, R. M. 1999.
Analysis of total phenols and other oxidation substrates and antioxi-
dants by means of Folin-Ciocalteu reagent. Methods Enzymol. 299:
152-178.
Slavin, J. 2004. Whole grains and human health. Nutr. Res. Rev. 17:
99-110.
Thompson, L. U., and Yoon, J. H. 1984. Starch digestibility as affected by
polyphenols and phytic acid. J. Food Sci. 49:1228-1229.
Tucker, A. J., Vandermey, J. S., Robinson, L. E., Graham, T. E., Bakovic,
M., and Duncan, A. M. 2014. Effects of breads of varying carbohydrate
quality on postprandial glycaemic, incretin and lipidaemic response
after first and second meals in adults with diet-controlled type 2 di-
abetes. J. Funct. Foods 6:116-125.
Venn, B. J., and Green, T. J. 2007. Glycemic index and glycemic load:
Measurement issues and their effect on diet–disease relationships. Eur.
J. Clin. Nutr. 61:S122-S131.
You, S., and Lim, S. T. 2000. Molecular characterization of corn starch
using an aqueous HPSEC-MALLS-RI system under various dissolution
and analytical conditions. Cereal Chem. 77:303-308.