Food Hydrocolloids 87 (2019) 287–296

Contents lists available at ScienceDirect

Food Hydrocolloids
journal homepage: www.elsevier.com/locate/foodhyd

Effect of egg white solids on the rheological properties and bread making T
performance of gluten-free batter
Aiyun Hana,d, Hollman Motta Romeroa, Noriaki Nishijimab,c, Tsukasa Ichimurab, Akihiro Handab,
Changmou Xua, Yue Zhanga,∗
a
Department of Food Science and Technology, University of Nebraska-Lincoln, Lincoln, NE, 68588, United States
b
Institute of Technology Solution, R&D Division, Kewpie Corporation, 2-5-7 Sengawa, Chofu, Tokyo, 1820002 Japan
c
Henningsen Foods, Inc., Omaha, NE, 68144, United States
d
Department of Food Science and Engineering, Shijiazhuang University, Shijiazhuang, Hebei, 050035, PR China

A R T I C LE I N FO A B S T R A C T

Keywords: Developing baked gluten-free (GF) products is difficult since gluten is essential for many organoleptic properties
Gluten-free bread like texture and taste. Egg white proteins (EW) show strong cohesive behavior with excellent foaming capacity
Egg white and stability, which may improve both organoleptic quality and nutritional value of GF bread. This work aimed
Rheology to study the impact of two EW powders prepared by different methods (e.g. P110 and M200) on bread volume,
Texture
shape, texture and their potential use in retarding the staling process. The two EW samples were used to sub-
Interfacial properties
stitute 5–15% of GF flour in a control GF bread formulation. Compared with control, breads with EW had larger
specific volumes and more homogeneous texture. The rheological properties of GF bread batter were evaluated
through temperature and frequency sweep tests. In general, addition of EW increased the elasticity of GF batter
and improved the texture properties of resultant bread during storage. M200 with more water-soluble protein
aggregates produced a more significant improvement in bread quality than general standardization sample
P110. To elaborate the stabilizing mechanism of EW on bread network, the surface properties of the two EW
samples including surface tension, zeta-potential as well as their conformational changes during thermal
treatments were studied and compared.

1. Introduction
The production of high quality baked gluten-free (GF) foods remains
a technological challenge and the degree of challenge is closely asso-
ciated with how functional gluten is in a particular food product. For
bread, gluten plays a crucial role in the gas retention and structure
formation (Hager & Arendt, 2013). And the lack of gluten results in a
significantly reduced retention of carbon dioxide produced by yeast and
hence a coarse and rigid texture and short shelf-life of products made
with GF flours (Skendi, Mouselemidou, Papageorgiou, & Papastergiadis,
2018). In addition, GF flours and starches are not generally enriched or
fortified, which may lead to nutritional deficiencies in protein and
micronutrients of resulting GF breads (Capriles, dos Santos, & Arêas,
2016). A GF bread with good organoleptic properties and high nutri-
tional value is still the most desired product by individuals with gluten-
related disorders (Sandri, Santos, Fratelli, & Capriles, 2017).
Many studies have been focused on quality improvements of GF
bread by the addition of hydrocolloids including both polysaccharides
and proteins (Anton & Artfield, 2008; Bize, Smith, Aramouni, & Bean,
2017). It is suggested that they can mimic viscoelastic properties of
gluten and increase the gas retention properties of the dough, and thus
enhance the loaf volume and retard the hardness of the resultant bread
(Morreale, Garzón, & Rosell, 2018). Polysaccharides such as xanthan
gum, hydroxypropyl methylcellulose (HPMC) and guar gum have fre-
quently been incorporated into GF formulations to help form a network
structure and then improve the bread quality (Foschia, Horstmann,
Arendt, & Zannini, 2016; Morreale et al., 2018). However, since the
production background of these polysaccharides may vary (e.g. by
biosynthesis pathway) and some production information do not appeal
to consumers, there are some concerns regarding the economic pro-
duction of polysaccharides with stable and standardized quality and the
appropriate incorporated doses (Giavasis, 2014; Horstmann, Axel, &
Arendt, 2018). And the product with addition of polysaccharide alone
may still not resemble traditional bread in texture and sensory
(Crockett, Ie, & Vodovotz, 2011). Alternatively, proteins are in-
corporated to GF bread, in combination of polysaccharides such as

∗
Corresponding author. Department of Food Science and Technology, University of Nebraska-Lincoln, 1901 21st Street, Lincoln, NE 68588, United States.
E-mail address: yue.zhang@unl.edu (Y. Zhang).

https://doi.org/10.1016/j.foodhyd.2018.08.022
Received 24 June 2018; Received in revised form 27 July 2018; Accepted 13 August 2018
Available online 14 August 2018
0268-005X/ © 2018 Elsevier Ltd. All rights reserved.
A. Han et al. Food Hydrocolloids 87 (2019) 287–296

HPMC, to improve both the perceived quality and texture by enhancing Table 1
Maillard browning and flavor, increasing elastic modulus and gas re- The formulas of gluten-free batters with/without substitution of egg white so-
taining capacity (Crockett et al., 2011; Phongthai, D'Amico, lids.
Schoenlechner, & Rawdkuen, 2016). For example, egg white solids at Formulas % flour basis Control M5 M10 M15 P5 P10 P15
15% improved the loaf volume of GF bread (Crockett et al., 2011). The
nutritional value of GF breads can also be improved by protein en- Ingredients (g)
richment. Egg white protein, as the most widely used surface active
GF flour 100–85 200 190 180 170 190 180 170
agent, can form strong cohesive viscoelastic films which are essential Egg white M200 0–15 – 10 20 30 – – –
for gas retention in GF bread (Bize et al., 2017). An improvement of the Egg white P110 0–15 – – – – 10 20 30
overall quality and storage stability of GF breads has been observed by Rice fiber 4 8 8 8 8 8 8 8
Tapioca starch 3 6 6 6 6 6 6 6
the addition of liquid whole egg at 20–30% (flour basis) while increased
Sugar 10 20 20 20 20 20 20 20
height and specific volume of GF muffin were achieved by adding egg Salt 1.6 3.2 3.2 3.2 3.2 3.2 3.2 3.2
white powder at 17.3% (flour basis) with a combination of HPMC or Trehalose 5 10 10 10 10 10 10 10
xanthan gum (Bize et al., 2017; Matos, Sanz, & Rosell, 2014). In these Soybean oil 12 24 24 24 24 24 24 24
previous studies, it was obvious that, when egg white applied, espe- Yeast 3 6 6 6 6 6 6 6
Water 100 200 200 200 200 200 200 200
cially at a low concentration, a combination of other hydrocolloids such
as HPMC, pectin and guar gum was needed to achieve the improvement Note: M5 represents using 5% of Egg white solids M200, P5 represents using 5%
on GF bread quality (Ziobro, Juszczak, Witczak, & Korus, 2016; Bize of Egg white solids P110. Same rules were applied to M10, M15, P10, and P15.
et al., 2017). As stated by Ziobro et al. (2016), adding egg white in the
absence of guar gum and pectin showed negative influence on bread with two egg white solids at different levels are stated in Table 1.
staling. Meanwhile, a reduced dough stability was observed on HPMC- The dry yeast was prehydrated in water (38–43 °C) for 8 min. The
treated GF formulation supplemented with egg white at 5% and 10%, other dry ingredients were blended to homogeneity. The yeast mixture
while the antagonistic interaction with HPMC network was overcome was added, and the batter was mixed for 5 min using a Kitchen-Aid
by 15% egg white (Crockett et al., 2011). There is still much room for Professional Stand Mixer (KitchenAid, St. Joseph, MI) with a dough hook
improvement in developing GF bread formulation supplemented with at speed 4 (scale = 1–10). For each bread loaf, 350 g batter was placed
egg white at a low concentration. In addition, the role of egg white in into non-stick baking pan (L = 21.6 cm × W = 11.4 cm × H = 7.35 cm)
the dough network formation and stabilization, and the resultant im- and proofed in a proofing chamber (Model 6030, CARON Products &
provement of GF dough/batter and baked bread remains somewhat Services, Inc., Marietta, OH) at 38 °C and 75% relative humidity for
unclear, which retards its further development as a regular ingredient 45 min. The GF batter samples were then baked in an electric oven for
in GF formulations. 25 min at 176 °C. After baking, loaves were removed from pans and
Therefore, two spray-dried egg white solids with same chemical cooled for 5 h at room temperature (21 °C) before measurements. To
compositions but manufactured by different industrial processes were study the effect on staling, bread loaves were sealed in polyethylene bags
incorporated into GF formulations and the resultant batters were baked and stored in ambient conditions for up to 4 days. Three breads were
in this study. The first objective was to investigate the impact of egg made for each recipe.
white solids at different levels on the rheological properties of GF bread
batters and the quality of resultant breads, and hence to enunciate the
function of egg white in the structure of bread. The second objective 2.3. Bread quality
was to develop a GF bread with better quality.
2.3.1. Specific volume
2. Materials and methods Bread loaves were weighted 5 h postbaking and their volumes were
determined following AACC method 10–14.01 by using a laser sensor
2.1. Materials with a BVM-L 370 vol analyzer (Texvol Instruments, Viken, Sweden).
The specific volume for each loaf was calculated by dividing the sample
Two spray-dried egg white solids M200 and P110 were provided by volume (cm3) by the sample weight (g). All measurements were done in
Henningsen Foods, Inc. (Omaha, NE). The physical and chemical at- triplicate for each recipe.
tributes of these two samples are shown in Supplementary Table 1.
Commercial all-purpose gluten-free flour (a mixture of garbanzo bean
flour, potato starch, tapioca flour, whole grain sorghum flour and fava 2.3.2. Moisture measurement
bean flour) were purchased from Bob's Red Mill Natural Foods, Inc. The bread moisture content was determined following AACC
(Milwaukie, OR). Rice fiber came from J. Rettenmaier (Schoolcraft, method 44–11.01 by using HB 43-S Halogen moisture analyzer (Mettler
MI), while the tapioca starch was obtained from Tate &Lyle Ingredients, Toledo Inc., Greifensee, Switzerland). Weight loss during baking was
Inc. (Decatur, IL). The rest of ingredients including Fleischmann's Ac- assessed by weighing the pans before and after baking. All breads were
tive dry yeast, trehalose (100% pure), sugar (sucrose), salt and soybean analyzed in triplicate.
oil were acquired in the local market. The extrinsic fluorescence probe
ANS (8- anilinonaphthalene-1-sulfonic acid) was purchased from
Sigma-Aldrich (St. Louis, MO) while other reagents were purchased 2.3.3. Texture analysis
from either Fisher Scientific Inc. (Pittsburgh, PA) or Bio-Rad Labora- The crumb texture was determined using a TA-XT2 texture analyzer
tories, Inc. (Hercules, CA). (Stable Microsystems, Surrey, UK) with the ‘‘Texture Expert’’ software.
A 25 mm diameter cylindrical probe was used in a ‘Texture Profile
2.2. Preparation of gluten-free batter and bread Analysis’ (TPA) double compression test to penetrate to 30% depth,
with a test speed of 1 mm/s, and a 10 s delay between the first and
The GF batters were prepared following the recipe on a 100 g GF second compressions. Hardness (gf), springiness, cohesiveness, resi-
flour basis with 0%–15% substitution of egg white solids. Other in- lience and chewiness were calculated from the TPA plot based on the
gredients were constant in all the formula: 4% rice fiber, 3% tapioca methods stated by Gómez, Ronda, Caballero, Blanco, and Rosell (2007).
starch, 10% sucrose, 1.6% salt, 5% trehalose, 12% vegetable oil, 3% dry For each recipe, texture determinations were made in triplicate using
yeast and 100% water. The formulations of GF batters incorporated bread slices with 25 mm thickness.

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2.4. Rheological properties of batter
Rheological properties of GF batter samples (without yeast) were
measured using a MCR-301 rotational rheometer (Anton Parr, Graz,
Austria) equipped with parallel plate spindle (25 mm diameter). The
gap size, strain and frequency were set at 1 mm, 0.5% and 1 Hz re-
spectively (within the viscoelastic region). To perform temperature
sweep tests, the samples were heated from 30 °C to 95 °C and then
cooled down to 30 °C at a rate of 2 °C/min. After temperature sweep
tests, samples were held at 30 °C for 5 min and a frequency sweep test
from 1 to 100 rad/s was performed to study the viscoelastic properties
of the gels formed. Each test was performed in triplicate.
2.5. Characterization of egg white solids
2.5.1. Surface tension analysis
The dried egg white samples M200 and P110 were dissolved in
water at a concentration of 0.1% or 1% (w/v). The protein solutions
were loaded onto a DSA25 droplet shape analyzer (Kruss, Hamburg,
Germany) to create a 10 μL pendant drop to measure the protein ad-
sorption at the water-air interface (A. Wouters, Fierens, Rombouts,
Brijs, Blecker, 2017). The dynamic surface tension was conducted by
monitoring the shape of pendant drop for 10 min. The surface tension
was plotted against t−1/2 to further estimate the surface tension (ST)
data as a function of time. The steady state surface tension values (γ∞)
was determined from the intercept of the plot.
2.5.2. Surface hydrophobicity analysis
Four mL of protein solution (0.01–0.1%) was mixed with 20 μL of
8 mM ANS (8- anilinonaphthalene-1-sulfonic acid). Subsequently, the
fluorescence of the supernatant was measured with a LS55 fluorescence
spectrophotometer (PerkinElmer, Waltham, MA) equipped with a
quartz cell of 1.0 cm pathlength. The emission wavelength was set at
350 nm and the emission spectra was recorded from 400 to 600 nm.
Protein concentration was plotted against the fluorescence intensity at
470 nm and surface hydrophobicity was calculated as the slope of the
linear regression S0.
2.5.3. Zeta-potential analysis
The zeta-potential of egg white samples was performed on a nano ZS
Zetasizer (Malvern Instruments, Malvern, UK) using a universal dip cell
at room temperature (21 °C). The protein stock solutions were prepared
in distilled water at 1 mg/mL, and adjusted to pH 7 or pH 10, respec-
tively. After 10 times dilution, the zeta-potential values of samples at
the original or adjusted pH conditions were measured.
2.5.4. Fluorescence and circular dichroism analysis
Egg white samples at 1 mg/mL in PBS buffer (pH 7.4) were heated
at 80 °C for 5–10 min, followed by a cooling down at 21 °C for 20 min
before measurements. The fluorescence intensity of egg white samples
with and without heat treatment was measured by LS55 fluorometer (as
described above). The excitation wavelength was 285 nm, while the
emission spectra were recorded in the range of 300 nm–450 nm. The
excitation and emission slit widths were set at 10 nm and 3 nm, re-
spectively, while the scan speed was set at 1200 nm min−1. The circular
dichroism (CD) spectra were obtained by using a J815 CD spectrometer
(JASCO, Tokyo, Japan) within a 0.1 cm cylindrical cuvette at 20 °C in
nitrogen atmosphere. Results were expressed as millidegrees (mdeg).
All measurements were carried out in triplicate.
2.5.5. SDS-PAGE
The egg white samples were analyzed by sodium dodecyl sulfate
polyacrylamide gel electrophoresis (SDS-PAGE) under reducing condi-
tions. The egg white samples were first dissolved in 1 × Laemmli
sample buffer with presence of 5% β-mercaptoethanol. And a total of
40 μg of the sample was loaded onto a mini protean TGX Pre-cast (12%)
gel (Bio-Rad Laboratories, Hercules, CA) for electrophoresis under a
constant voltage of 200 V. After fixing and staining with Coomassie
brilliant blue, the gel was destained and scanned by an Odyssey CLx
imager (LI-COR, Nebraska, USA). Apparent molecular weights were
determined by using a broad range molecular weight standard marker
(Bio-Rad Laboratories, catalog nr 161–0317).
2.6. Statistical analysis
Data were analyzed using SPSS version 19.0 software (SPSS Inc.,
Chicago, IL). Independent-sample t-test was used to test the statistical
significance by a comparison among the 7 groups of bread. A one-way
ANOVA was carried out for analyzing the texture parameters of breads
individually. Fisher's least significant difference (LSD) test was used to
describe means with 95% confidence. The data were expressed as
means ± SEM.
3. Results and discussion
3.1. Effect of egg white solids on the specific volume and texture of the bread
crumb
Texture, specific volume and moisture were selected as the in-
dicators of the bread quality. As shown in Table 2, specific volume
value was increased by 16–44% when the two egg white samples were
added to the breads at three levels. Generally, bread supplemented with
M200 showed a greater mean volume than that with P110 especially at
10% level. M200 showed a better improvement than P110 since bread
supplemented with 10% M200 had a significantly higher volume
(4.28 cm3/g) than control (3.08 cm3/g) while the significant difference
could only be observed with 15% P110 incorporated (p < 0.05). The
impact of two egg white samples on the specific volume is also illu-
strated in Fig. 1.
The moisture content and other texture properties of bread crumbs
are summarized in Table 2. The moisture content of bread crumb was
not significantly influenced by the addition of egg white. This ob-
servation agrees with some previous works in which the addition of
hydrocolloids did not affect the moisture values (Lazaridou, Duta,
Papageorgiou, Belc, & Biliaderis, 2007).
The increase of crumb hardness was observed by other studies when

Table 2
Specific volume, moisture content and texture properties of breads with/without egg white solids.a
Sample Specific volume (cm3/g) Moisture content (%) Hardness (gf) Springiness Cohesiveness Chewiness Resilience

Control 3.08 ± 0.24a 36.48 ± 1.94a 454.55 ± 30.31ab 0.92 ± 0.04a 0.50 ± 0.08a 210.71 ± 24.94a 0.25 ± 0.06a
M5 3.63 ± 0.49a 38.79 ± 3.22a 527.97 ± 39.65b 0.93 ± 0.02a 0.56 ± 0.08ab 274.82 ± 52.87ab 0.28 ± 0.08a
M10 4.28 ± 0.17bc 37.63 ± 2.27a 347.94 ± 17.90a 0.94 ± 0.02a 0.73 ± 0.03b 237.61 ± 16.49a 0.42 ± 0.04b
M15 4.45 ± 0.17c 36.33 ± 1.74a 511.18 ± 91.15b 0.95 ± 0.02a 0.78 ± 0.02b 377.14 ± 62.46c 0.46 ± 0.03b
P5 3.56 ± 0.40ab 37.11 ± 1.59a 494.48 ± 18.15b 0.93 ± 0.01a 0.54 ± 0.06a 248.53 ± 24.02a 0.26 ± 0.05a
P10 3.49 ± 0.64a 38.32 ± 1.03a 528.39 ± 48.18b 0.94 ± 0.02a 0.70 ± 0.04b 348.39 ± 48.34bc 0.40 ± 0.03b
P15 4.30 ± 0.20bc 36.19 ± 2.80a 550.78 ± 116.95b 0.95 ± 0.02a 0.74 ± 0.06b 383.41 ± 79.98c 0.43 ± 0.06b

a
Values are expressed as the mean and SD of at least three measurements. Letters (a–c) mean significant (p < 0.05) difference within the same column.

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Fig. 1. Appearance of crumb structure of gluten-free breads depending on egg

white levels and types.

hydrocolloids such as soy protein isolate, xanthan gum were in-

corporated (Lazaridou et al., 2007; Marco & Rosell, 2008). In this study,
the inclusion of egg white has resulted in an insignificant increase
(p > 0.05) of crumb hardness for all samples but a decrease for M10.
The insignificant increase in the hardness could be attributed to the
excellent water binding and gelling properties of egg white, which built
a network to decrease the swelling of the starch granules and the re-
duced amylose leaching from the granules, which was similar to other
hydrocolloids as network builder (Lazaridou et al., 2007). However, the
crumb hardness was also correlated with bread volume that a higher
bread volume results in higher amounts of air retained in the bread
structure that justifies a lower crumb hardness (Marco & Rosell, 2008).
The low hardness of M10 could be a combined result of high specific
volume but relatively not strong enough network built by 10% egg
white. Springiness of all egg white enriched bread crumbs was similar
to that of the control bread. The two parameters cohesiveness and re-
silience are related to the bread crumb instant and retarded recovery
capacity after a compression cycle. The incorporation of egg white in-
creased both parameters and the increase appeared to be dose-depen-
dent. This observation agrees with previous studies on GF bread and
dough with hydrocolloids such as β-glucan, xanthan gum, etc.
(Lazaridou et al., 2007; Ronda, Perez-Quirce, Lazaridou, & Biliaderis,
2015). A slightly higher cohesiveness and resilience may be favorable
for consumer acceptance, because the product is less susceptible to
crumbling. However, opposite results were reported by others since a
decrease in the cohesiveness was observed when soy protein con-
centrate, pea protein isolate, and individual albumin ingredient were Fig. 2. Effect of egg white solids on crumb texture properties of gluten-free
added to breads (Ziobro et al., 2016). The chewiness was increased bread during storage at 21 °C for up to 4 days. ■ = day 0; □ = day 1; = day
significantly with increasing egg white except M10, probably because 2; = day 4; Error bars represent standard deviation; * Statistically significant
this parameter is mainly affected by hardness (Ronda et al., 2015). difference (P < 0.05) as compared with the same sample on day 0.

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Fig. 3. Storage modulus (G′) values of different gluten-free batters as a function of (A) increasing temperature, (B) decreasing temperature (insert: control for-
mulation), (C) increasing angular frequency at 30 °C, respectively. (D) Values of loss factor (tanδ) as a function of increasing angular frequency at 30 °C. Filled
symbols correspond to batters with egg white M200 and empty symbols correspond to batters with P110 at three different levels (grey squares 5%; red circles 10%;
blue triangles 15%). Solid black line represents the control batter without egg white. (For interpretation of the references to colour in this figure legend, the reader is
referred to the Web version of this article.)

The crumb structure of GF breads with and without egg white solids
is presented in Fig. 1. Good quality GF breads are characterized by
smaller gas cells since they have been found to produce loaves with
higher specific volumes (Bize et al., 2017). Crumb structure was im-
proved by addition of the two egg white samples, as cell size was
smaller and specific volume was larger with increasing amount of egg
white in the formulation (Fig. 1 and Table 2). It was suggested that a
weak network did not succeed in maintaining integrity of gas bubbles
and the coalescence of gas bubble resulted in large cells and a collapse
of the GF batter during proofing and baking (Bize et al., 2017). The
control GF bread had a collapsed appearance, confirming a weak
structure. M200 showed a better improvement on the crumb structure
than P110 that is supported by the smaller cell size and more homo-
geneous appearance at all three levels. This phenomenon is more pro-
nounced in formulation with higher levels of egg white.
3.2. Effect of egg white solids on the texture of the bread crumb during
storage
The textural properties of the bread crumbs during storage were
also evaluated (Fig. 2). The hardness of control bread was significantly
increased from 454.6 gf to 1053.9 gf after 4-day storage, showing a very
quick staling rate of bread. As shown in Fig. 2A, the hardness of bread
crumb with the addition of the two egg white samples was significantly
lower compared to the control at Day 4, confirming the retrogradation
of bread could be retarded by egg white. It was suggested that non-
gluten proteins, as emulsifiers, strengthen the structure of crumb, and
compete with starch for water absorption, and thus, delay the starch
retrogradation (Nunes, Moore, Ryan, & Arendt, 2009). M200 at applied
10% addition level was more effective in hardening retardation.
The springiness of control and breads supplemented with egg white
is shown in Fig. 2B. The springiness of control bread was insignificantly
lower with increasing storage days. Two egg white samples at 10–15%
levels could retard the reduction of crumb springiness and acted in a
dose-dependent manner. And M200 still showed a slightly better in-
hibition than P110 at 10% supplement levels (P > 0.05).
The cohesiveness and resilience of control bread rapidly dropped
down on the first day of storage (Fig. 2C and E). No significant effect of
time could be observed on the two parameters after the first day. The
cohesiveness was also decreased significantly for samples supplemented
with egg white. But the values were greater than that of control samples
at every time point, which may show a better acceptability of sensory
attributes. The same trends were observed on resilience values. And
both parameters for samples with M200 were slightly higher than those
with P110 (P > 0.05).
On the other hand, the chewiness of control bread showed insig-
nificant increase during the storage (Fig. 2D). This change may be in-
hibited by the addition of egg white in the GF breads but the influence

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Fig. 4. Dynamic surface tension of egg white solids at a bulk concentration of 0.1% w/v (A) and 1% (B), respectively. Exemplary plot of surface tension against t-0.5 at
a bulk concentration of 0.1% w/v (C) and 1% w/v (D), respectively. Dash lines are linear regressions with parameters indicated in each panel. M200, black squares;
P110, red circles. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)
of both samples was not significant (P > 0.05).
Generally, for samples supplemented with M200, there was small
variation in the values of all these parameters during storage, showing
that a relatively stable crumb structure could be achieved and main-
tained by adding M200. The storage period had more significant impact
on these parameters of samples with P110. The samples with M200
have generally narrower standard deviation of each parameter than
those with P110, confirming the improvement by M200 on each texture
parameter was highly repeatable. These results further confirmed that
M200 showed better performance than P110 on the improvement of GF
bread quality.
3.3. Effect of egg white solids on the rheological properties of the batter
To identify and enunciate the impacts of the two egg white samples
on changes in GF bread's characteristics, the rheological properties of
batters supplemented with egg white at three levels have been ana-
lyzed. A temperature sweep of the viscoelastic properties was studied to
understand the thermal treatment of the GF batters. One representative
of the storage modulus (G′) value for each sample during heating from
30 °C to 95 °C and cooling from 95 °C to 30 °C is shown in Fig. 3A and
Fig. 3B, respectively. For all samples, G′ values were greater than loss
moduli G′′ (data not shown), indicating the batters showed a more
elastic than viscous character (Romero, Santra, Rose, & Zhang, 2017).
The changes of storage modulus in the GF batters with heating are
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Food Hydrocolloids 87 (2019) 287–296
mainly associated with starch gelatinization and protein coagulation
and gelation (Matos et al., 2014). When the temperature raised gra-
dually, the control batter without protein showed an early onset of
starch gelatinization from 68 to 78 °C (Fig. 3A). Then, a decrease of G′
occurred at 78 °C with prolonged heating due to the melting of the
crystalline region remaining in the swollen starch granule, which de-
formed and loosened the granules (Singh, Singh, Kaur, Sodhi, & Gill,
2003). By adding egg white, the decrease of G′ was postponed to dif-
ferent degrees but the maximum G′ value during heating showed no
significant correlation with either egg white type or concentration. This
is because that the protein unfolding happens in the same temperature
range (e.g. Td of ovalbumin = 84 °C) which may decrease the G′ but the
followed protein coagulation may result in an increase of network
strength (Liu, Oey, Bremer, Carne, & Silcock, 2017; Matos et al., 2014).
For control batter, a very slight increase of G′ from 30 kPa to 55 kPa
was observed immediately as the temperature decreased from 95 °C to
30 °C (Fig. 3B), indicating the formation of a more rigid network mainly
due to the interactions between amylopectin molecules (Thory &
Sandhu, 2017). With egg white supplemented, the G′ values were sig-
nificantly increased during cooling, and the impact of egg white was
dose-dependent. The much higher G′ values can be associated with the
coagulation phenomenon of the egg white proteins. Throughout all
three levels, batters with M200 exhibited higher G′ than those with
P110, reflecting a greater capacity of M200 to build a strong three-
dimensional internal structure (Matos et al., 2014). A stronger gel-like
A. Han et al. Food Hydrocolloids 87 (2019) 287–296

tan δ value indicates that batters with egg white have a higher capacity
to absorb water. It was suggested that egg white would compete with
starch for water absorption, and the protein-starch complexation hinder
starch-starch complexation and hence reduce starch retrogradation
(Nilufer-Erdil, Serventi, Boyacioglu, & Vodovotz, 2012). In addition,
increase in all tan δ values was observed at higher frequency, showing
that the frequency dependence of G′ was lower than G′′. But a sharper
rise of control sample was observed showing that the structure of
control batter was much susceptible to applied stress than samples with
egg white. This could be attributed to the weakening of the gel-like
structure due to extended deformation and rupture of starch granules
(Villanueva, Ronda, Moschakis, Lazaridou, & Biliaderis, 2018), which
could be postponed by addition of egg white. The observation of tan δ
over frequency could help explain the inhibition of bread staling by egg
white during storage.

3.4. Interfacial properties of egg white solids

To better understand the quality improvement of bread by addition

of egg white especially M200, the adsorption of egg white solids to air-
water interface was studied by determining dynamic surface tensions of
egg white solids at the bulk concentrations of 0.1% and 1% w/v
(Fig. 4). At low concentration, the initial surface tension value was
87 mN/m for M200 and 79 mN/m for P110, respectively. Both of them
reached ∼64 mN/m after 10 min measurement (Fig. 4A). When sample
concentration increased to 1% w/v (Fig. 4B), both systems reached
equilibrium more rapidly than that at the low concentration. But M200
showed a lower value than P110 at the end of measurement (57 mN/m
Fig. 5. (A) Surface hydrophobicity (as relative extrinsic fluorescence) of egg
vs 60 mN/m). The decrease in the surface tension with increasing
white solids M200 (black) and P110 (red) at room temperature. (B) Zeta-po-
protein concentration was also observed by others in dairy and legume-
tential of 0.1 mg/mL egg white solids M200 and P110 at different pH condi-
tions. (For interpretation of the references to colour in this figure legend, the
like proteins, and the speed of surface tension decay was also increased
reader is referred to the Web version of this article.) by higher protein concentration because of the higher number of mo-
lecules that transport and adsorb to the interface (Jarpa-Parra et al.,
2015; Tomczynska-Mleko et al., 2014).
structure may have better water absorption and gas retention capa-
To better estimate the surface tension at equilibrium, the steady
cities, which can help explain the above observation: GF breads sup-
state surface tension values (γ∞) were determined by the linear extra-
plemented with M200 had better quality than those with P110.
polation of the dynamic values of γ to t∞. And the γ values reveal a
To further evaluate the effect of egg white in the viscoelastic
linear dependence for longer time range on σ versus t−1/2 plots (Fig. 4C
properties, and hence to predicate the processability of batter and
and D). Therefore, the adsorption process of the two egg white samples
quality of the resultant bread (Tao, Zhang, Wu, Jin, & Xu, 2016), the
could be assumed as diffusion controlled (Bąk & Podgórska, 2016). As
evolution of storage modulus and the ratio of G″ to G′ (tan δ) with in-
shown in Fig. 4C and D, the γ ∞ values of two samples at 0.1% w/v were
creasing frequency were also studied (Fig. 3C and D). At the frequency
similar (61.77 mN/m vs 61.94 mN/m) as expected, while the γ∞ de-
range tested (1–100 Hz), values of G′ showed insignificant increase with
termined for M200 (56.43 mN/m) was lower than γ∞ for P110 at
frequency, indicating the low susceptibility of GF dough structure on
58.72 mN/m at 1% w/v. In addition, M200 showed a lower rate of
applied stress. Following a similar trend at the studied frequency range,
surface tension decay than P110 at 0.1% but a higher rate when the
storage modules of GF batters increased significantly with increasing
concentration increased to 1%. The results obtained in this work in-
egg white. And batters with M200 showed significantly higher G′ values
dicate that M200 showed a better ability than P110 to decrease inter-
than those with P110 at all three levels.
facial tension if the bulk solution contained abundant protein mole-
As shown in Fig. 3D, the loss tangent (tan δ) of all samples was
cules, and hence had a better capacity for gas retention.
lower than 1, suggesting a gel-like behavior maintained after heating-
The behavior of proteins at air–water interface is affected by a
cooling process. The tan δ was also significantly affected by the sup-
combination of physicochemical properties including electrostatic
plement of egg white. At low frequency, the presence of egg white in
charge, molecular size, surface hydrophobicity, chemical reactivity etc.
batter at all three levels increased the tan δ slightly, except M15 sup-
(de Jongh et al., 2004). Therefore, the surface hydrophobicity and zeta-
plemented with 15% M200. For sample M15, significant increase in the
potential of the two protein samples was also measured (Fig. 5A and B).
value of G′ was accompanied with significant increase of G″, which
The surface hydrophobicity of egg white samples was obtained by using
resulted in a much higher tan δ than control and other samples in the
the ANS probe, which binds to hydrophobic regions on protein surface
whole range of frequencies. It is suggested that the addition of both egg
and hence produce an increase in its emission fluorescence intensity
white samples led to structures with less solid like character than the GF
(Wang & Zhang, 2017). As shown in Fig. 5A, M200 showed a much
batter alone, and the effect was much more evident for M200. This
higher S0 value than P110 (71.30 vs 16.82), indicating that M200 had
observation is consistent with previous work done by Matos et al.
more hydrophobic domains exposed to the aqueous phase. A higher
(2014) that animal protein source including casein and egg white in-
surface hydrophobicity of M20 may induce a greater tendency to adsorb
creased values of tan δ. An increase in tan δ would signify a greater
at air-water interface, and hence dominate the better foam stabilizing
contribution to the viscous properties of gluten-free batter (Romero
effect (A. G. Wouters et al., 2017). The zeta potential of M200 was
et al., 2017). At a fixed water content, the water absorption of starch
−28.8 mV at pH 10 and -25.6 mV at pH 7, while P110 was −30.7 mV
granules is one of the factors determining the rheological properties of
and −29.1 mV for pH 10 and 7, respectively (Fig. 5B). The zeta po-
the starch-based batter system after gelatinization. A slightly higher
tential measurement confirmed the surface hydrophobicity

293
A. Han et al.
Fig. 6. Fluorescence intensity (A and B) and circular dichroism spectra (C and D) of egg white solids M200 (left) and P110 (right) without (black line) and with heat
treatment at 85 °C for 5 min (red line) and 10 min (blue line), respectively. (For interpretation of the references to colour in this figure legend, the reader is referred to
the Web version of this article.)
measurement that M200 had a less charged groups but more hydro-
phobic groups on the surface. At the same pH condition, P110 showed a
slightly more negative surface charge (although insignificantly) and
lower surface hydrophobicity than M200, which may favor the hydra-
tion and diffusion of protein molecules in dilute solution, leading a
faster adsorption of P110 at low concentration (0.1%). However, at
high protein concentration, the electrostatic repulsions among abun-
dant protein molecules added up to the energy barrier for adsorption of
protein molecules to the air-water interface leading to slowed adsorp-
tion (Ercili-Cura et al., 2015). Conversely, a more hydrophobic surface
nature of M200 resulted in better orientation of the hydrophobic parts
of M200 toward the gas phase (Jarpa-Parra et al., 2015). Therefore,
M200 possessed more capacity of lowering surface tension and a better
stabilization of air bubbles in water phase or GF batters.
3.5. Conformational changes of egg white solids by heat treatment
Fluorescence spectroscopy was used to evaluate the impact of heat
treatment on the structure of egg white proteins. As shown in Fig. 6A,
the fluorescence intensity of M200 decreased significantly after heat
treatment, and the wavelength of the peak maximum shifted from
340.5 nm to 348 nm. The red shift indicates that the polarity around the
fluorophore residues (mainly tryptophan) increased, showing the
strong impact of heating on the unfolding of egg white protein (Zhang &
Zhong, 2012). In contrast, the intensity of P110 decreased slightly with
thermal treatment and no obvious shift of peak maximum was observed
(Fig. 6B), indicating the proteins in P110 was less heat susceptible than
294
Food Hydrocolloids 87 (2019) 287–296
M200.
The far-UV CD method was used to measure secondary structure
changes of the two egg white samples with and without heat treatment.
Examples of CD spectra are presented in Fig. 6C for M200 and Fig. 6D
for P110, respectively. Both samples showed a negative sharp minima
at ∼230 nm, which represents the typical β-sheet structure of protein.
The prominent β-sheet content in the two samples was also confirmed
by the observation of a positive band at 193 nm (Sheng et al., 2018).
Although, native fresh ovalbumin, as the major protein in egg white,
was known as a high ordering and substantial protein containing both
α-helical and β-sheet structures (Sheng et al., 2018). The absence of α-
helical band in both samples may be attributed to the complex com-
position and the relatively harsh processing conditions (e.g. spray-
drying) of egg white samples. For M200, the intensity of the negative
band decreased significantly and the shoulder at 193 nm disappeared
after heat treatment, indicating that the secondary structure has been
changed. However, the impact of thermal treatment on the CD spectra
of P110 could be ignored. The overall results of fluorescence and CD
spectra suggested that both secondary and tertiary structures of M200
were significantly affected after heating while P110 was more tolerant.
This observation may further explain why M200 showed better gas
bubble retention than P110 in GF bread. During bread baking, M200
turned to be more disordered and flexible that exposed more hydro-
phobic regions to the interface, contributed to the better stability of the
foam at gas-liquid interface (Sheng et al., 2018).
A. Han et al.
Fig. 7. SDS-PAGE of egg white solids. Lane 1 is marker while lane 2 and 3
correspond to M200 and P110, respectively.
Fig. 8. Schematic representation of egg white proteins in P110 and M200 at air-
water interface.
3.6. General discussion and future prospects
Surface activity at the air-water interface is an important factor to
define the protein's foaming ability and stability. To analyze the protein
composition of M200 and P110 and hence to better understand their
different performance, SDS-PAGE was carried. As shown in Fig. 7, an
obvious weaker intensity of the band at 45 kDa in M200 was found,
which suggested the noticeably lower amount of ovalbumin in M200
compared with P110. In contrast, remarkable increase of the intensities
of bands above 97.4 kDa can be observed in M200. As indicated by the
manufacturer, the two egg white solids have the same chemical com-
position from the same egg source but of rather different processing
procedures and conditions. The increase of proteins with high molecule
weight in M200 might indicate the aggregation of proteins such as
ovalbumin during the processing. And these water-soluble aggregates
and macro-aggregates (at the top of the separation gel) formed in M200
295
Food Hydrocolloids 87 (2019) 287–296
could be responsible for the better surface activity, as observed in the
above experiments. It was also reported earlier that water soluble egg
white aggregates formed by dry heat treatment showed increased sur-
face hydrophobicity, contributing to the improvement of functional
properties such as gelling (Handa, Hayashi, Shidara, & Kuroda, 2001).
The protein orientation and adsorption at air-water interface is
shown in Fig. 8. It has been suggested that the protein adsorption at air-
water interface is a multistep process including (i) protein diffusion, (ii)
adsorb to the interface, (iii) change their molecular structure, and (iv)
spread at the interface (Mitropoulos, Mütze, & Fischer, 2014). When a
protein adsorbs to the interface and interacts with other proteins, the
surface tension decreases. At very low concentration, the diffusion of
single proteins in P110 from bulk solution towards the interface was
faster than the big aggregates in M200, and resulted the lower surface
tension at beginning (Fig. 4A). But at relatively high concentration,
diffusion was not the major decision step for the interface adsorption
any more. Generally, a protein can possess hydrophobic patches on its
surface. An increase in the net charges on the protein surface may re-
duce their tendency to adsorb onto the surface of water (Lu, Su, &
Penfold, 1999). In contrast, a higher surface hydrophobicity leads to
more rapid adsorption and larger reduction of interfacial tension
(Mitropoulos et al., 2014). Compared with P110 proteins with a more
native form, the water-soluble aggregates in M200 were more dena-
tured with more hydrophobic regions exposed to the surface (as sup-
ported by the zeta-potential measurement in Fig. 5), which was re-
sponsible for the lower surface tension and faster adsorption rate as
determined in Fig. 4B and D. As shown in Fig. 6, M200 was more
susceptible to thermal treatment than P110, resulting a more irrever-
sible protein denaturation and a more complex structure at the inter-
face. This is favorable to build up a stable and strong viscoelastic net-
work at interfacial layer, which was confirmed by the observation that
GF batters with M200 showed higher G′ values than those of P110 at all
three levels (Fig. 3). In addition, the more significant increase of tan δ
by adding M200 (Fig. 3D), indicating that M200 could better balance
the viscous and elastic responses in the batters (Crockett et al., 2011).
Apparently, once egg white became the primary protein scaffolding in
the batter and resulted bread (supplemented with 10–15%), a more
stable interconnected honeycomb matrix can be formed (especially by
M200), which improved the loaf volume and crumb texture during
storage (Figs. 1 and 2). Thus, the result suggested that both egg white
solids improved the quality of GF breads, but M200 with more water-
soluble protein aggregates exhibited better performance than regular
egg white P110. M200 may be an excellent ingredient to be applied in
other bakery applications.
4. Conclusion
The results obtained demonstrate that the incorporation of egg
white solids improved the batter rheology and initial quality and sto-
rage stability of GF bread significantly. In particular, egg white solids
with more water-soluble protein aggregates exhibited higher surface
hydrophobicity than general standardization egg white sample, pos-
sessed more capacity of lowering surface tension and a better stabili-
zation of air bubbles in water phase or GF batters, and hence showed
better improvement on bread crumb properties. Our work provides an
alternative GF bread formulation with better quality and longer lasting
softness.
Acknowledgements
This study was supported financially by Kewpie Corporation. The
authors are also grateful to Henningsen Foods, Inc. for supplying the
egg white samples, Liyang Xie for the fluorescence and circular di-
chroism measurements.
A. Han et al.
Appendix A. Supplementary data
Supplementary data related to this article can be found at https://
doi.org/10.1016/j.foodhyd.2018.08.022.
References
Anton, A. A., & Artfield, S. D. (2008). Hydrocolloids in gluten-free breads: A review.
International Journal of Food Sciences & Nutrition, 59(1), 11–23.
Bize, M., Smith, B. M., Aramouni, F. M., & Bean, S. R. (2017). The effects of egg and
diacetyl tartaric acid esters of monoglycerides addition on storage stability, texture,
and sensory properties of gluten‐free sorghum bread. Journal of Food Science, 82(1),
194–201.
Bąk, A., & Podgórska, W. (2016). Interfacial and surface tensions of toluene/water and
air/water systems with nonionic surfactants Tween 20 and Tween 80. Colloids and
Surfaces A: Physicochemical and Engineering Aspects, 504, 414–425.
Capriles, V. D., dos Santos, F. G., & Arêas, J. A. G. (2016). Gluten-free breadmaking:
Improving nutritional and bioactive compounds. Journal of Cereal Science, 67, 83–91.
Crockett, R., Ie, P., & Vodovotz, Y. (2011). Effects of soy protein isolate and egg white
solids on the physicochemical properties of gluten-free bread. Food Chemistry, 129(1),
84–91.
Ercili-Cura, D., Miyamoto, A., Paananen, A., Yoshii, H., Poutanen, K., & Partanen, R.
(2015). Adsorption of oat proteins to air–water interface in relation to their colloidal
state. Food Hydrocolloids, 44, 183–190.
Foschia, M., Horstmann, S., Arendt, E. K., & Zannini, E. (2016). Nutritional therapy–fa-
cing the gap between coeliac disease and gluten-free food. International Journal of
Food Microbiology, 239, 113–124.
Giavasis, I. (2014). Bioactive fungal polysaccharides as potential functional ingredients in
food and nutraceuticals. Current Opinion in Biotechnology, 26, 162–173.
Gómez, M., Ronda, F., Caballero, P. A., Blanco, C. A., & Rosell, C. M. (2007). Functionality
of different hydrocolloids on the quality and shelf-life of yellow layer cakes. Food
Hydrocolloids, 21(2), 167–173.
Hager, A.-S., & Arendt, E. K. (2013). Influence of hydroxypropylmethylcellulose (HPMC),
xanthan gum and their combination on loaf specific volume, crumb hardness and
crumb grain characteristics of gluten-free breads based on rice, maize, teff and
buckwheat. Food Hydrocolloids, 32(1), 195–203.
Handa, A., Hayashi, K., Shidara, H., & Kuroda, N. (2001). Correlation of the protein
structure and gelling properties in dried egg white products. Journal of Agricultural
and Food Chemistry, 49(8), 3957–3964.
Horstmann, S. W., Axel, C., & Arendt, E. K. (2018). Water absorption as a prediction tool
for the application of hydrocolloids in potato starch-based bread. Food Hydrocolloids,
81, 129–138.
Jarpa-Parra, M., Bamdad, F., Tian, Z., Zeng, H., Temelli, F., & Chen, L. (2015). Impact of
pH on molecular structure and surface properties of lentil legumin-like protein and its
application as foam stabilizer. Colloids and Surfaces B: Biointerfaces, 132, 45–53.
de Jongh, H. H., Kosters, H. A., Kudryashova, E., Meinders, M. B., Trofimova, D., &
Wierenga, P. A. (2004). Protein adsorption at air–water interfaces: A combination of
details. Biopolymers, 74(1‐2), 131–135.
Lazaridou, A., Duta, D., Papageorgiou, M., Belc, N., & Biliaderis, C. G. (2007). Effects of
hydrocolloids on dough rheology and bread quality parameters in gluten-free for-
mulations. Journal of Food Engineering, 79(3), 1033–1047.
Liu, Y.-F., Oey, I., Bremer, P., Carne, A., & Silcock, P. (2017). Effects of pH, temperature
and pulsed electric fields on the turbidity and protein aggregation of ovomucin-de-
pleted egg white. Food Research International, 91, 161–170.
Lu, J., Su, T., & Penfold, J. (1999). Adsorption of serum albumins at the air/water in-
terface. Langmuir, 15(20), 6975–6983.
Marco, C., & Rosell, C. M. (2008). Breadmaking performance of protein enriched, gluten-
free breads. European Food Research and Technology, 227(4), 1205–1213.
Matos, M. E., Sanz, T., & Rosell, C. M. (2014). Establishing the function of proteins on the
rheological and quality properties of rice based gluten free muffins. Food
Hydrocolloids, 35, 150–158.
296
Food Hydrocolloids 87 (2019) 287–296
Mitropoulos, V., Mütze, A., & Fischer, P. (2014). Mechanical properties of protein ad-
sorption layers at the air/water and oil/water interface: A comparison in light of the
thermodynamical stability of proteins. Advances in Colloid and Interface Science, 206,
195–206.
Morreale, F., Garzón, R., & Rosell, C. M. (2018). Understanding the role of hydrocolloids
viscosity and hydration in developing gluten-free bread. A study with hydro-
xypropylmethylcellulose. Food Hydrocolloids, 77, 629–635.
Nilufer-Erdil, D., Serventi, L., Boyacioglu, D., & Vodovotz, Y. (2012). Effect of soy milk
powder addition on staling of soy bread. Food Chemistry, 131(4), 1132–1139.
Nunes, M. H. B., Moore, M. M., Ryan, L. A., & Arendt, E. K. (2009). Impact of emulsifiers
on the quality and rheological properties of gluten-free breads and batters. European
Food Research and Technology, 228(4), 633–642.
Phongthai, S., D'Amico, S., Schoenlechner, R., & Rawdkuen, S. (2016). Comparative study
of rice bran protein concentrate and egg albumin on gluten-free bread properties.
Journal of Cereal Science, 72, 38–45.
Romero, H. M., Santra, D., Rose, D., & Zhang, Y. (2017). Dough rheological properties and
texture of gluten-free pasta based on proso millet flour. Journal of Cereal Science, 74,
238–243.
Ronda, F., Perez-Quirce, S., Lazaridou, A., & Biliaderis, C. G. (2015). Effect of barley and
oat β-glucan concentrates on gluten-free rice-based doughs and bread characteristics.
Food Hydrocolloids, 48, 197–207.
Sandri, L. T., Santos, F. G., Fratelli, C., & Capriles, V. D. (2017). Development of glu-
ten‐free bread formulations containing whole chia flour with acceptable sensory
properties. Food Sciences and Nutrition, 5(5), 1021–1028.
Sheng, L., Huang, M., Wang, J., Xu, Q., Hammad, H., & Ma, M. (2018). A study of storage
impact on ovalbumin structure of chicken egg. Journal of Food Engineering, 219, 1–7.
Singh, N., Singh, J., Kaur, L., Sodhi, N. S., & Gill, B. S. (2003). Morphological, thermal and
rheological properties of starches from different botanical sources. Food Chemistry,
81(2), 219–231.
Skendi, A., Mouselemidou, P., Papageorgiou, M., & Papastergiadis, E. (2018). Effect of
acorn meal-water combinations on technological properties and fine structure of
gluten-free bread. Food Chemistry, 253, 119–126.
Tao, H., Zhang, B., Wu, F., Jin, Z., & Xu, X. (2016). Effect of multiple freezing/thawing-
modified wheat starch on dough properties and bread quality using a reconstitution
system. Journal of Cereal Science, 69, 132–137.
Thory, R., & Sandhu, K. S. (2017). A Comparison of mango kernel starch with a novel
starch from litchi (Litchi chinensis) kernel: Physicochemical, morphological, pasting,
and rheological properties. International Journal of Food Properties, 20(4), 911–921.
Tomczynska-Mleko, M., Kamysz, E., Sikorska, E., Puchalski, C., Mleko, S., Ozimek, L.,
et al. (2014). Changes of secondary structure and surface tension of whey protein
isolate dispersions upon pH and temperature. Czech Journal of Food Sciences, 32(1),
82–89.
Villanueva, M., Ronda, F., Moschakis, T., Lazaridou, A., & Biliaderis, C. G. (2018). Impact
of acidification and protein fortification on thermal properties of rice, potato and
tapioca starches and rheological behaviour of their gels. Food Hydrocolloids, 79,
20–29.
Wang, L., & Zhang, Y. (2017). Eugenol nanoemulsion stabilized with zein and sodium
caseinate by self-assembly. Journal of Agricultural and Food Chemistry, 65(14),
2990–2998.
Wouters, A., Fierens, E., Rombouts, I., Brijs, K., Blecker, C., & Delcour, J. (2017a). Air-
water interfacial properties of enzymatically hydrolyzed wheat gluten in the presence
of sucrose. Food Hydrocolloids, 73, 284–294.
Wouters, A. G., Rombouts, I., Schoebrechts, N., Fierens, E., Brijs, K., Blecker, C., et al.
(2017b). Foam fractionation as a tool to study the air-water interface structure-
function relationship of wheat gluten hydrolysates. Colloids and Surfaces B:
Biointerfaces, 151, 295–303.
Zhang, Y., & Zhong, Q. X. (2012). Effects of thermal denaturation on binding between
bixin and whey protein. Journal of Agricultural and Food Chemistry, 60(30),
7526–7531.
Ziobro, R., Juszczak, L., Witczak, M., & Korus, J. (2016). Non-gluten proteins as structure
forming agents in gluten free bread. Journal of Food Science & Technology, 53(1),
571–580.