ARTICLE IN PRESS

Journal of Cereal Science 46 (2007) 148–156

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Effect of high-pressure treatment on rheological, thermal and

structural changes in Basmati rice flour slurry
Jasim Ahmed, Hosahalli S. Ramaswamy, Anwer Ayad, Inteaz Alli, Pedro Alvarez
Department of Food Science & Agricultural Chemistry, Macdonald Campus, McGill University, Ste. Anne de Bellevue, PQ, Canada H9X 3V9
Received 10 October 2006; received in revised form 8 January 2007; accepted 10 January 2007

Abstract

Rheological, thermal and structural changes in high pressure (HP) treated Basmati rice flour dispersions were studied as function of
pressure level (350–650 MPa), slurry concentration (with 1:5, 1:3 and 1:2 flour-to-water ratios) and holding time (7.5–15 min). Rice flour
dispersions exhibited a gradual liquid–solid gel transformation as they gelatinized and/or denatured and behaved as viscoelastic fluid
following HP treatment. Mechanical strength (G0 ) of pressurized gel increased with applied pressure and rice concentration. Differential
scanning calorimeter (DSC) thermograms of rice slurry measured after pressure treatment indicated a reduction in peak enthalpy in
proportion with the extent of gelatinization and/or denaturation of starch and proteins. Pressure-treated rice samples had a progressively
lower gelatinization temperature. A 15 min pressure treatment at 550 MPa was found sufficient to complete gelatinization of protein free
isolated rice starch while the slurry required 650 MPa. The presence of proteins might have been responsible for the slower starch
gelatinization in the rice slurry during pressure treatment. The sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE)
and Fourier-transform infrared (FTIR) spectroscopy results indicated some minor changes in protein subunits and secondary structure
of rice protein. This study has provided complementary information on pressure-induced changes in physical (thermal stability, overall
structure) and molecular level (secondary structure) of rice protein.
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Keywords: Rice; Elastic modulus; Complex viscosity; Starch gelatinization; Protein denaturation; Enthalpy; Electrophoresis; FTIR; DSC

1. Introduction processing techniques such as high-pressure pulse-electric

‘Basmati’—a slender, long-grain rice is liked for its
exquisite nut-like fragrance during cooking and a delicate,
almost buttery flavor due to presence of 2-acetyl-1-pyrro-
line (Bergman et al., 2004). It falls in the category of non-
waxy, non-glutinous rice with intermediate amylose con-
tent, and does not stick on cooking. Basmati rice flour
offers potential for traditional and newer product devel-
opment with health benefits due to its higher protein
content and lower glycemic index. Rice flour has been used
to prepare baby foods (especially gluten free), noodles,
soups, thickener, extruded products and blending with
other ingredients. Currently, there is a growing interest in
physical modification of cereal flours using innovative
Corresponding author. Tel.: +1 514 398 8666; fax: +1 514 398 7977.
E-mail addresses: Jasim.ahmed@mcgill.ca, jahmed2k@yahoo.com
(J. Ahmed), hosahalli.ramaswamy@mcgill.ca (H.S. Ramaswamy).
field treatments, microwave, radio frequency, Ohmic
heating for food applications. So far, Basmati rice flour
has not been exploited in that direction.
For more than a decade, high pressure (HP) treatment
has been investigated as an alternative to the traditional
thermal processing. In addition to microbial and enzymatic
inactivation, modifications of food protein and starch
functionality by HP treatment have received considerable
attention. HP treatment has been demonstrated to result in
conformational changes and functionality leading to newer
product development with desirable and novel texture with
retention of sensory attributes (Ledward et al., 1995; Knorr
et al., 2006). HP treatment has been shown to influence the
functional properties of proteins through the disruption
and reformation of hydrogen bonds and hydrophobic
interactions leading to denaturation, aggregation and
gelation (Cheftel, 1992; Lopez-Fandino, 2006). Fourier
transform infrared (FTIR) spectroscopy, differential scanning

0733-5210/$ - see front matter r 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.jcs.2007.01.006
 ARTICLE IN PRESS
J. Ahmed et al. / Journal of Cereal Science 46 (2007) 148–156
calorimetry (DSC), and rheology have been considered
important in evaluating thermal and structural changes.
Rheology and DSC provide information at physical and
macroscopic level indicating changes associated with
gelatinization or denaturation processes. FTIR, a vibra-
tional spectroscopic technique, provides information on
changes associated with the secondary structure proteins.
Thus, FTIR provides complementary information to DSC
on direct structural information about protein at sub-
molecular levels and has been used for conformational
study of globular protein on the amide I region (Ellepola
et al., 2005; Brandes et al., 2006).
French (1984) reported that the crystalline order of
starches could be destroyed by means of HP treatment.
Muhr and Blanshard (1982) found that in presence of
excess water, wheat starch gelatinization could be com-
pleted by HP treatment at 450 MPa and ambient tempera-
ture. The extent of pressure-induced gelatinization was
found to depend on the pressure range, moisture content,
treatment time, temperature, starch concentration, and the
type of starch. It is also reported that B-type starches were
more resistant to pressure gelatinization than A-type and
C-type starches (Muhr and Blanshard, 1982; Ezaki and
Hayashi, 1992; Rubens et al., 1999; Stute et al., 1996;
Katapo et al., 2002). X-ray diffraction studies have
indicated that the pressure treatment of 1 h at 690 MPa
led to conversion of A-type to B-type X-ray diffraction
patterns (Katapo et al., 2002). Cereal starches have been
found to be more sensitive to pressure than tuber starch
(Muhr et al., 1982). DSC studies of wheat starch
suspensions (16% DM) have indicated that starch gelatiniza-
tion was initiated above 300 MPa and was completed at
600 MPa. Recently, Kawai et al. (2007) observed the heat
gelatinization peak (DH) of potato starch–water mixture
disappeared completely when treated at 700 and 1200 MPa
(at 40 1C for 60 min) for 10% and 50% slurries, respectively.
No published information is available on pressure effect on
starch gelatinization and changes in protein secondary
structure of rice flour slurries. Kato et al. (2000) applied
HP treatment (100–400 MPa) to remove allergenic proteins
(mainly 16 kDa albumin and 33 kDa globulin) from rice and
observed that a significant amount of proteins was released in
the pressure range 300–400 MPa. Thus, HP treatment of rice
could have considerable health benefits in addition to
facilitating certain functional properties.
Gelatinization and rheological characteristics of rice
have been studied extensively (Ojeda et al., 2000; Yama-
moto et al., 2006). The major factors influencing the
rheological properties of rice starch gels are the granular
structure and component, plus the amount of leached
amylose (Juliano, 1985). The extent of gelatinization
achieved during the pressure treatment dictates the
rheological properties of the treated product. These are
highly dependent on the starch type and the processing
conditions used, such as pressure, temperature and holding
time (Stolt et al., 2001). Little is known about the pressure
effect on rheological characteristics of rice slurry.
149
The objectives of this work were to study viscoelastic
characteristics of Basmati rice flour slurry as affected by
the HP treatment and to understand the pressure suscept-
ibility of the major rice components (starch and protein)
using rheology, DSC, electrophoresis and FTIR spectro-
photometry.
2. Materials and methods
2.1. Materials
Commercially available milled Basmati rice (Elephant
brand, DSSS, Amritsar, India), product of India, was
purchased from a local source in Montreal and used for
high-pressure treatments. The proximate composition (%)
of rice flour samples (as per analyzed by AOAC method,
1996) are: moisture, 12.0770.31; ash, 0.3170.03; protein
(N  6.25); 7.8770.23; crude fat, 1.2170.10; and, crude
fiber, 2.2270.05. The rice samples were ground with a dry
grinder (Model KSM 2, type 4041, Braun Canada), to
obtain flour and passed through #100 mesh screen aperture
(1.149 mm as per ASTME II). Isolated rice starch was
obtained as the residue from sequential protein extraction
of Basmati rice based on the protein extraction technique
of Hu and Essen (1981) and Kwon et al. (1996) with
modification as reported Ahmed et al. (2007). The starch
sample had 3.770.05% moisture, 0.0570.00% ash,
0.370.02% protein and 0.2570.05% lipid.
2.2. Preparation of sample
Rice flour slurries were prepared at different flour
concentrations yielding 1:5, 1:3 and 1:2 flour/water ratios
(with 16.5, 22.9 and 33% wet basis flour content,
respectively), the final one being the ratio normally used
in cooking of Basmati rice. The required amount of rice
flour and distilled water were placed in a glass beaker, hand
mixed with a glass rod and kept for 3 h at room
temperature (2071 1C) for hydration. All measurements
were conducted with the same lot of rice obtained at one
time. The different flour concentrations were selected to
evaluate changes in rheological properties as influenced by
flour content and HP. Isolated starch slurry was also
prepared at 1:5 (starch-to-water ratio) in similar fashion.
Measurements were made in triplicate.
2.3. HP treatment
Rice slurry and isolated starch dispersions (approx.
20–25 g) were packed in low-density polyethylene bags
(Whirl-Paks, USA) and the bags were heat sealed. The air
in the bag was squeezed out as much as possible before
sealing. Samples were then transferred to the 5 L pressure
treatment chamber (ACIP 6500/5/12VB; ACB Pressure
Systems, Nantes, France) equipped with temperature and
pressure regulators. Water was used as the pressure
transmitting medium. Samples were pressure treated at
 ARTICLE IN PRESS
150 J. Ahmed et al. / Journal of Cereal Science 46 (2007) 148–156
specified pressure level (350, 450, 550 and 650 MPa) for the
stipulated time (7.5 and 15 min). The pressurization rate
was about 4.4 MPa/s and released at 26 MPa/s. The initial
temperature of pressure medium was 18 1C which quickly
increased to 24.8 and 28.5 1C due to the adiabatic effect
during pressurization of 350 and 650 MPa and equilibrated
to 22 and 26 1C during holding period at those pressure
levels. All experiments were carried out in duplicate.
2.4. Rheological measurements
The rheological measurements of pressure-treated rice
flour dispersions were carried out by using a controlled-
stress rheometer (AR 2000, TA Instruments, New Castle,
DE). A parallel plate geometry (60-mm diameter) was used
and the gap between two plates was 1000 mm. The sample
temperature was internally controlled by a peltier system
(20–200 1C with an accuracy of 70.1 1C) attached to a
water circulation unit. A platinum resistance thermometer
positioned at the center of the plate ensured proper
temperature control and measurement. For each test, a
measured volume of thoroughly mixed sample (approxi-
mately 2 mL) was placed between the rheometer plates for
5 min for stress relaxation and temperature equilibration
before the actual measurements.
Small-amplitude oscillation (SAOS) measurements of
pressure-treated sample were carried out within the linear
viscoelastic region after temperature equilibration to 20 1C
over a frequency range between 0.1 and 10 Hz which
enables the material to retain the structure. In another set
of experiment, rice slurry of 1:5 ratio (flour-to-water) was
quickly heated to 90 1C and held for 30 min in the
rheometer plate in situ followed by immediate cooling to
20 1C with the internal peltier heating system. Solvent trap
was used during isothermal heating at 90 1C and as an
additional precaution to prevent moisture loss; the edges of
the plates were covered with a thin layer of light paraffin oil
(Ahmed et al., 2007). Each time, a new sample was used for
measurement. Rheological parameters (elastic modulus, G0 ;
viscous modulus, G00 ; phase angle, d; and complex
viscosity, Z ) were directly obtained from the manufacturer
supplied computer software (Rheology Advantage Data
Analysis Program, TA Instruments, New Castle, DE). The
deviations did not exceed 5% between duplicate runs, as
the experiment was repeated. The average of the duplicate
runs was reported as the measured value.
2.5. Differential scanning calorimetric (DSC) measurement
A DSC (TA Q100, TA Instruments, Newcastle, DE) was
employed to measure the thermal analysis for pressure
treated slurries. The DSC was calibrated with indium and
sapphire for temperature and heat capacity values (TA
Instruments, 2002). Slurry samples (about 12–15 mg) were
accurately weighed into aluminum DSC pans and herme-
tically sealed. Thermal scans were performed from 30 to
130 1C at a heating rate of 5 1C/min. An empty pan was
used as a reference. The DSC measurements were done in
triplicate. Thermal transitions of rice slurries were mea-
sured in terms of onset (T0) and peak (Tm) gelatinization
temperatures. The enthalpy (DH) of the transition (asso-
ciated with starch gelatinization and protein denaturation)
was calculated from the area of the peak endotherm using
the Universal Analysis Software (version 3.6C, TA Instru-
ments, New Castle, DE).
2.6. FTIR spectroscopy
The FTIR transmission spectra were recorded on a
Nicolet 8210E FTIR spectrometer (Nicolet, WI) with a
deuterated triglycine sulphate (DTGS) detector. The
spectrometer was continuously purged with dried air
(Balston, MA, USA) to reduce water vapor. Approxi-
mately 8 mL of rice slurry (1:5 flour to D2O) was placed
between two CaF2 windows separated by a 50 mm thick
TeflonTM spacer. The measurement was carried out at
25 1C in triplicates. The temperature of the cell was
regulated by an Omega temperature controller, with
0.5 1C accuracy (Omega Engineering, Stanford, CT). A
total of 512 scans were coadded at 4 cm1 resolution. The
absorbance spectra were subjected to band narrowing
techniques using Fourier self deconvolution (FSD) employ-
ing a bandwidth of 20 cm1 (w) and enhancement factor of
2.4 (k) followed by a two-point baseline correction starting
at 1710 cm1 and ending at 1590 cm1 using Omnic 6.0
software (Nicolet, WI). Samples were prepared 24 h in
advance and kept at 4 1C to allow for a complete
hydrogen–deuterium exchange.
2.7. Electrophoresis
SDS-PAGE was performed using the method of Fling
and Gregerson (1986) to demonstrate the effects of HP on
the rice protein. Stacking gel and resolving gel of 4% and
12% acrylamide were used for SDS-PAGE. Slab gels (0.75-
mm thickness) were run at a constant current of 15 mA/gel;
the electrophoresis was performed with a Bio-Rad Mini
Trans blotting system following the method described by
Ahmed et al. (2003) earlier.
2.8. Data analysis
Microsoft Excel software package (Microsoft Corpora-
tion, USA) was used to carry out the statistical analysis
and linear regressions of experimental data.
3. Results and discussion
3.1. Mechanical spectra of Basmati rice slurry
Mechanical spectra obtained at 20 1C for control (with-
out pressure treatment) and pressure treated (450 MPa
for 15 min) samples of 1:5 Basmati rice flour–water
slurry are shown in Fig. 1. Rheological properties of the
 ARTICLE IN PRESS
J. Ahmed et al. / Journal of Cereal Science 46 (2007) 148–156 151

pressure-treated sample differed considerably from those of 1000

Control
the control sample. The elastic modulus (G0 ) predominated 350 MPa
over viscous component (G00 ) in the selected frequency 450 MPa
550 MPa
range (0.1–10 Hz) indicating solid-like properties for both 650 MPa

control and pressure-treated rice samples. A small change

in G0 with frequency suggests the elastic nature of the gel

G' (Pa)
network (Ferry, 1970; Ross-Murphy, 1984). Both G0 and 100
G00 values of the slurry significantly (Po0.05) increased
after the pressure-treatment. Phase angle (measure of the
ratio of energy lost to energy stored in a cyclic deforma-
tion) decreased significantly for pressure-treated samples
indicating again the more solid-like characteristic of the gel
as compared to control sample. 10
0.1 1 10
ω (Hz)
3.2. Effect of pressure on mechanical strength
Fig. 2. Effect of high pressure on mechanical strength of 1:5 rice flour
slurry at 20 1C.
Since pressure treatment resulted in a significant increase
in G0 (indicator of mechanical strength of gel) of rice slurry
as compared to G00 (liquid-like property indicator), the Table 1
study of the pressure effect was focussed more on G0 in this Slope of power-type relationship ln G0 ¼ ln K þ n ln o of Basmati rice
paper. Effect of HP treatment on mechanical strength of slurry at selected pressure levels
rice slurry is shown in Fig. 2 as related to different pressure Pressure (MPa) Slope R2 SE
levels. Mechanical strength of rice slurry gel increased with
an increase in pressure levels. Elastic modulus (G0 ) and 0.101 0.140 0.724 0.132
frequency (o) data of pressure-treated gels were fitted to a 350 0.105 0.853 0.065
450 0.089 0.914 0.043
power-type relationship (G0 ¼ Kon ). Slopes of the equation 550 0.086 0.940 0.032
after linearization (ln G 0 ¼ ln K þ n ln o) are presented in 650 0.063 0.907 0.030
Table 1. The slope coefficient of the logarithmic elastic
modulus frequency decreased as the pressure levels
increased, demonstrating greater pressure sensitivity at
both ends. The samples exhibited increasingly solid-like 3.3. Effect of pressure holding time on elasticity and
characteristics with pressure. It has been reported that a viscoelasticity
true gel is characterized by a zero slope of the power law
model (Ross-Murphy, 1984). Pressure-treated gels below Elasticity of rice gel increased with pressure holding time
450 MPa were relatively weak. Similar mechanical spectra at a constant pressure. The slopes coefficients of ln (G0 ) vs.
and weak gelation behavior was earlier reported by ln (o) of rice gels slightly decreased with longer holding
Rosalina and Bhattacharya (2002) for various starch period (for example, 0.089 to 0.086 at 550 MPa for 7.5 and
samples. 15 min). However, the increase in G0 at 650 MPa was found
to be independent of holding time (data not shown). Time
dependency of pressurized gel is represented by a complex
100 control G' control G''
20 viscosity (Z ) which is a measure of the overall resistance to
pressurized G' pressurized G''
control delta pressurized delta flow and visco-elasticity. A plot of complex viscosity versus
angular frequency of rice gel as function of pressure
holding time (7.5 and 15 min) at 550 and 650 MPa is
15
illustrated in Fig. 3. Following increasing trends of both G0
G', G'' (Pa)

and G00 , there was systematic increase in Z with the applied

δ (°)

10
pressure treatment. No significant difference (P40.05) of
10 Z value was observed at 650 MPa between 7.5 and 15 min
pressure holding time. It seem to indicate that at lower
pressure levels (p550 MPa), a certain length of holding
time is essential to achieve a significant gel and viscoelas-
1 5 ticity while at 650 MPa, the gel gets set at a short time and
0.1 1 10 the strength does not change with time. This is close in
ω (Hz) agreement with Katapo et al. (2002) who reported no
Fig. 1. Typical mechanical spectra of control and pressure treated significant differences in gel strength for various starch
(450 MPa for 15 min) Basmati rice slurry (1:5 flour-to-water dispersion) samples pressure treated at 690 MPa with holding time
at 20 1C. between 5 and 60 min.
 ARTICLE IN PRESS
152 J. Ahmed et al. / Journal of Cereal Science 46 (2007) 148–156

1000 10000

100
1:5 flour to water
1:3 flour to water

G' (Pa)
1:2 flour to water
η* (Pa.s)

1000
10

550 MPa 7.5 min

1 550 MPa 15 min

650 MPa 7.5 min

650 MPa 15 min 100

0.1 0.1 1 10
0.1 1 10 ω (Hz)
ω (Hz)
Fig. 4. Effect of flour concentration (flour-to-water ratios) on mechanical
Fig. 3. Effect of pressure holding time on visco-elasticity of pressure- strength of pressure-treated samples (650 MPa for 15 min).
treated 1:5 rice flour slurry gel at 20 1C.

1000 1000
3.4. Effect of concentration

Effect of concentration on gel rigidity of pressure-treated

100
samples is shown in Fig. 4. 1:5, for samples of 1:3 and 1:2

η* (Pa.s)
G' (Pa)

rice flour–water slurry. Gel rigidity increased as concentra-

tion increased at any given pressure level which was mostly
contributed by partial gelatinization of starch and dena-
turation of proteins. Flour-to-water ratio below 1:3 showed
insignificant increase in mechanical strength and produced
brittle gels. Water becomes a critical factor below 1:3 flour-
to-water ratio and, therefore, gelatinization process of rice
slurry containing 1:2 flour-to-water ratio remain incom-
plete (insufficient water for gelatinization). This indicates
that rice slurry with higher water content exhibits a higher
degree of gelatinization during pressurization (until the
minimum required water level is reached) compared to
those slurries with moisture limiting flours. Similar
observation was made by Katapo et al. (2002) during
morphological study of selected HP-treated starch samples.
3.5. Comparison between pressure and heat-induced gels
Rice slurry exhibited some structural modifications
through pressure treatment and subsequent rheological
evaluations under different temperature-time regimes.
Mechanical strength and viscoelastic characteristic were
compared between thermal and pressure-induced gels from
1:5 rice flour–water slurries at 20 1C (Fig. 5). An
isothermally heated (90 1C for 30 min) rice gel exhibited
insignificantly (P40.05) higher mechanical strength and
viscoelasticity as compared to pressure-induced gel
(650 MPa for 15 min). Relatively, in terms of gel strength,
the degree of gelatinization of rice starch at 650 MPa for
15 min was almost equivalent to thermal treatment of 90 1C
for 30 min. An increase in temperature leads to swelling of
the starch granule in flour due to hydration of amorphous
Pressure G' 10
Thermal G'
Pressure |n*|
Thermal |n*|
100 1
0.1 1 10
ω (Hz)
Fig. 5. Comparison of rheograms between pressure (650 MPa for 15 min)
and thermal treated (90 1C for 30 min) rice slurry gel (1:5 flour-to-water) at
20 1C.
parts. In excess of water, the neighboring crystalline order
is destabilized by this absorption of water. When gelatini-
zation temperature is reached, these destabilized crystalline
regions melt leading to an irreversible loss of the granule
structure (Cameron and Donald, 1993). It is believed that a
pressure assisted gelatinization mechanism is similar to that
of temperature-induced mechanism (Rubens and Here-
mans, 2000). HP induces a reversible hydration of the
amorphous phase followed by an irreversible distortion of
the crystalline region leading to destruction of the granular
structure. Moreover, the effect of pressure on the starch–
water systems resembles remarkable similarities to that
found for other water–biopolymer systems especially
water–proteins (Muhr and Blanshard, 1982). An increase
in mechanical strength of thermally treated gel over
pressure-induced gel was earlier reported by several
researchers (Molina et al., 2002; Apichartsrangkoon,
2003). The increase in G0 could be contributed by leaching
 ARTICLE IN PRESS
J. Ahmed et al. / Journal of Cereal Science 46 (2007) 148–156 153

of amylose during thermal gelatinization which was isolated rice globulin appeared to show lower thermal
restricted in pressure gelatinization. transition temperature (Td) of 103.8 1C (Fig. 6b). The lower
Td of globulin could be influenced by presence of starch
and fat of the rice flour. Seed globulins have been found to
3.6. Differential scanning calorimetry
possess Td in the range of 83.8–107.8 1C (Marcone et al.,
1998; Ellepola and Ma, 2006).
The DSC thermogram of rice slurry (1:5 flour-to-water)
In order to investigate the thermal effects of HP on the
and isolated protein fraction (sodium chloride isolation)
flour suspensions, samples were scanned immediately after
dispersion (1:5 globulin to water) before pressure treatment
pressure treatment and after 24 h storage at 4 1C. Thermo-
are illustrated in Fig. 6a and b, respectively. Two distinct
grams obtained from samples of immediate scanning were
endothermic melting peaks (Td), associated with gelatiniza-
not significantly different (P40.05) from those obtained
tion/denaturation were observed in the DSC thermogram
with stored samples. It indicated that there was no
for the untreated rice flour dispersion sample. The first
retrogradation in pressure-treated starch samples during
peak (T d1 ¼ 70:6 1C) resembled the gelatinization peak of
the 24 h low temperature storage. The temperature of the
Basmati rice starch and is in agreement with observation
peak maximum (Td) and degree of gelatinization/denatura-
(70.9 1C for milled white rice) of Ellepola and Ma (2006).
tion (%) of pressure-treated samples are presented in
However, isolated Basmati rice starch exhibited higher
Table 2. Results indicated that the rice slurry samples were
gelatinization temperature (73.8 1C) at similar moisture
significantly affected by the HP treatment though complete
level (discussed later). In addition to starch, the thermal
gelatinization/denaturation did not occur. The Td of first
denaturation of albumin is in the same range (E75 1C);
endotherm decreased systematically from 350 to 550 MPa
however, the high solubility of starch in water is likely
after pressure treatment as compared to the control
responsible for insignificant contribution of albumin to
(without pressure treatment) sample. The degree of starch
total thermal transitions of flour dispersion. The second
gelatinization increased as the treatment pressure level
endotherm peak (T d2 ¼ 105:2 1C) of slurry is likely to be
increased. The endothermic peak disappeared with HP
caused by thermal denaturation of globulin. The extracted
treatment at 650 MPa indicating complete gelatinization of
rice starch. The Td of the second peak showed a small
-0.105 change (P40.05) from 350 to 450 MPa whereas an increase
was noticed in higher pressure level (550–650 MPa)
(Table 2). Similar non-systematic thermal denaturation
temperature of potato and wheat starches after HP
treatment was earlier reported by Muhr and Blanshard
Heat flow (W/g)

-0.12
(1982) and Rubens and Heremans (2000). The decrease of
gelatinization temperature at higher pressures has been
attributed to some kind of alteration in granule structure
and crystallinity (Blaszczak et al., 2005). It is also possible
-0.135
that these changes, which are not gross, are also influenced
by partial ‘annealing’ of starch, protein and lipid
105.2 (Marchant and Blanshard, 1978).
70.6
The melting enthalpies (DH), i.e. the areas of the peaks
-0.15 on the DSC thermograms, of the pressure-treated flour
40 60 80 100 120 suspensions are shown in Fig. 7 demonstrating progressive
Temperature (°C) gelatinization as the pressure levels increased. The DH
value decreased as pressure increased. The rice flour
-2.5
dispersion subjected to 650 MPa for 15 min showed no
peak and hence no DH value. Complete gelatinization/
Heat flow (W/g)

-2.55 denaturation resulted in the disappearance of the enthalpy

of the first endotherm peak. The decrease of enthalpy with
-2.6 pressure level has been reported in the literature for various
starches (Muhr et al., 1982; Ezaki and Hayashi, 1992;
-2.65
Kawai et al., 2007). Few starches like barley starch
103.8
granules remained intact after the pressure treatment up
to 550 MPa and no amylose leached out from the granules
-2.7 (Stolt et al., 2001). Recently, Van der Planchen et al. (2007)
90 100 110 120 found similar decreasing trend in enthalpy with pressure
Temperature (°C)
(525–600 MPa) for egg proteins.
Fig. 6. DSC thermograms of Basmati rice slurry (1:5 flour-to-water) The partial change of protein and starch structure during
before pressure treatment. pressure treatment was explained by Hayakawa et al.
 ARTICLE IN PRESS
154 J. Ahmed et al. / Journal of Cereal Science 46 (2007) 148–156

Table 2
Thermal changes in pressure treated (15 min holding time) rice slurry obtained using differential scanning calorimetry

Pressure (MPa) Peak 1 Peak 2

Tp (1C) Gelatinization (%) Tp (1C) Denaturation (%)

0.101 67.671.4 0 108.672.3 0

350 66.071.1 4.8 107.671.2 37.6
450 64.471.3 13.2 108.172.2 47.9
550 62.770.9 47.8 111.171.6 49.1
650 NDa 100 112.070.7 98.4
a
ND, not detected.

-0.2

-0.4
)

Heat flow (W/g)

Control -0.6
Control starch
350 MPa 73.8
Starch 550 MPa/15 min
450 MPa
-0.8
550 MPa
650 MPa
-1
40 50 60 70 80 90
Temperature (°C)
-1.2
Fig. 7. DSC thermograms for pressure treated 1:5 rice flour slurry. 50 70 90 110
Temperature (°C)

Fig. 8. Thermograms of control and pressurized isolated rice starch

(1996) on the basis of increase in water density. Water
density increases with pressure levels and subsequently the
number of water molecules per lattice increases signifi-
cantly from 4.3 molecules per lattice at atmospheric
pressure and at 4 1C to 10 molecules per lattice at
1000 MPa at 25 1C. These changes could be attributed by
cleavage of hydrogen bonds between surface of the protein
and the surroundings, protecting water molecules.
The residual denaturation enthalpy provides a net value
from a combination of endothermic reactions like the
disruption of hydrogen bonds, and exothermic processes,
including the break-up of hydrophobic interactions and
protein aggregation (Privalov and Khechinashvili, 1974;
Ma and Harwalkar, 1991). As a result, DSC-thermograms
of the pressure-treated rice slurry exhibited subsequent
thermal denaturation of the proteins remaining in a native-
like conformation. A reduction in residual enthalpy could
be an indication for a partial loss of protein structure
during HP and the observation is close in agreement with
earlier researchers (Taniguchi et al., 1994; Hayakawa et al.,
1996; Van der Planchen et al., 2007) where rearrangement
of secondary structure was suggested during HP treatment.
DSC thermogram of pressure-treated isolated rice starch
by (Fig. 8) indicated that a pressure treatment of 15 min at
550 MPa was sufficient to completely gelatinize starch
sample. Similar observation for A-type starch was reported
by Katapo et al. (2002) where the authors claimed a total
loss of the native crystalline structure and molecular order
after pressurization. However, Rubens and Heremans
sample.
(2000) suggested that the imperfect packing of amylose
and amylopectin chains in the starch granule played
significant role in the pressure-induced gelatinization
process. It is, therefore, inferred that the observed decrease
in enthalpy for rice slurry after pressure treatment cannot
be attributed solely to starch gelatinization or unfolding of
proteins; rather it results from combination of these
components.
3.7. Electrophoresis
SDS-PAGE analysis showed a gradual decrease in the
intensity of samples A and B (MWE52.2 and 35.0) with
increasing pressure application (Fig. 9). The major subunits
identified in the rice sample were 16.2 (F), 17.5 (E), 20.4
(D) and 22.2 (C) kDa, respectively. The results suggest that
the subunits C, D and F were affected by the pressure
alone. The lower molecular weights (16.2–22.2 kDa) are
likely globulin subunits (Krishnan et al., 1992) while the
higher MW (E35 kDa) could be glutelin subunits
(Komatsu and Hirano, 1992). The results from this study
are similar to those reported by Steenson and Sathe (1995).
3.8. FTIR spectroscopy
The amide I0 absorption region (1700–1600 cm1) in the
infrared spectrum of a protein has been considered to be
 ARTICLE IN PRESS
J. Ahmed et al. / Journal of Cereal Science 46 (2007) 148–156 155

M 1 2 3 Table 3
200 Band assignments of the deconvoluted amide I0 spectral region of rice
116 protein
97
66 Assignment Band position (cm1)
A
45 B b-Structure 1690
Antiparallel b-sheet (aggregation) 1680
31 C b-Turns 1658
a-Helix 1650
21
D Unordered structure (random coil) 1641
b-Strand 1632
14 Anti-parallel b-sheet 1623
E
6.5 Side chain vibrations 1611
KDa F

Fig. 9. SDS-PAGE of Basmati rice slurry (1:5 flour-to-water) (M: marker, 1.0
1: control sample, 2:350 MPa for 15 min and 3:650 MPa for 15 min).
0.9
one of the most useful for secondary structure elucidation 0.8
since the amide I band is the sum of overlapping 0.7

Absorbance
component bands (a-helix, b-sheet, b-turn and randomly 0.6
coiled conformation), which are mainly related to the CdO 0.5
stretching of the peptide bonds influenced by their various 0.4
environment in the different kinds of secondary structure 0.3
(Schwinte et al., 2001). The amide I0 band assignments of 0.2
rice proteins are presented in Table 3 which are based on 0.1
previous work (Ellepola et al., 2005). The average spectrum 0.0
of all the proteins present in rice is shown in Fig. 10.
Although the secondary structure features shown are the 1700 1680 1660 1640 1620 1600
average of features of all the proteins in rice, it still Wavenumbers (cm-1)
resembles the rice globulin spectrum, since this is the major
Fig. 10. Amide I0 region of the FTIR spectra from basmati rice. (Blue line
storage protein in this cereal. It is clear from the FTIR represents control sample and red line is for pressure-treated sample at
spectra that HP treatment of rice proteins at 650 MPa for 650 MPa for 15 min.)
15 min (1:5 flour-to-water) resulted in irreversible changes of
the secondary structures (Fig. 10). A major change in the treatment at 650 MPa resulted in complete gelatinization of
secondary structure (loss of intensity) of pressurized rice rice flour starch while isolated pure starch from rice could
proteins was observed at around 1690 cm1. This frequency be gelatinized at 550 MPa. HP treatment of rice slurry
corresponds to b-structure which was significantly affected by resulted in a decreased gelatinization temperature as
HP treatment. In addition, slight increases in the intensities of compared to untreated sample, and the presence of
other two bands at 1650 and 1641 cm1suggests a transition proteins in rice flour slurry was considered responsible
of the extended structure (1690 cm1) to a-helix and random for this. These observations were supported by DSC data
coil. Our observations differ somewhat with earlier reports on and the SDS-PAGE electrophoresis indicated alterations in
rice proteins (especially the band at 1690 cm1). The protein structures following HP treatment. Pressure-
difference is expected since our study dealt with crude induced and thermal gels could be compared with respect
proteins within a complex matrix (whole rice flour) whereas to their mechanical strength following certain combination
reported studies (Ellepola et al., 2005) were based on pure treatments. FTIR spectroscopy indicated change in sec-
isolated proteins. Gelatinization of starches after HP treat- ondary structure of post process rice proteins.
ment could be playing an important role by limiting the
solvent availability, when starch granules swell up and trap Acknowledgment
water; therefore, restraining the presence of extended
structures of the proteins. This research was partially funded by a Grant from the
Natural Sciences and Engineering Research Council
4. Conclusions (NSERC) of Canada.

HP treatment of Basmati rice slurries resulted partial to

complete gelatinization/denaturation of starch and protein
components and, increased mechanical strength of pressur-
ized the gel. Rice flour slurry below 1:3 (flour-to-water
ratio) did not permit complete gelatinization. Pressure
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