International Journal of Biological Macromolecules 75 (2015) 276282

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International Journal of Biological Macromolecules

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

Structural, functional, and ACE inhibitory properties of water-soluble

polysaccharides from chickpea ours
Abir Mokni Ghribi a , Assad Sila b,c , Ines Maklouf Gafsi a , Christophe Blecker d ,
Sabine Danthine d , Hamadi Attia a , Ali Bougatef b , Souhail Besbes a,
a

Universit de Sfax, Ecole Nationale dIngnieurs de Sfax, Laboratoire Analyses Alimentaires, route de Soukra, 3038 Sfax, Tunisia
Universit de Sfax, Ecole Nationale dIngnieurs de Sfax, Unit Enzymes et Bioconversion, route de Soukra, 3038 Sfax, Tunisia
c
Institut Rgional de Recherche en Agroalimentaire et Biotechnologie: Charles Viollette, EA1026, Equipe ProBioGEM, Universit Lille 1, France
d
Universit de Lige, Gembloux Agro Bio-Tech, Unit de Technologie des Industries Agro-Alimentaires, passage des Dports 2, 5030 Gembloux, Belgium
b

a r t i c l e

i n f o

Article history:
Received 15 December 2014
Received in revised form 16 January 2015
Accepted 25 January 2015
Available online 30 January 2015
Keywords:
Chickpea water-soluble polysaccharides
Physico-chemical characteristics
Functional properties
Anti-hypertensive properties

a b s t r a c t
The present study aimed to characterize and investigate the functional and angiotensin-I converting
enzyme (ACE) inhibition activities of chickpea water-soluble polysaccharides (CPWSP). Physico-chemical
characteristics were determined by nuclear magnetic resonance spectroscopy (NMR), Fourier transforminfrared spectroscopy (FT-IR) analysis, and X-ray diffractometry (XRD). Functional properties (water
holding capacity: WHC, water solubility index: WSI, swelling capacity: SC, oil holding capacity: OHC,
foaming, and emulsion properties) and ACE activities were also investigated using well-established procedures. The FT-IR spectra obtained for the CPWSP revealed two signicant peaks, at about 3500 and
500 cm1 , which corresponded to the carbohydrate region and were characteristic of polysaccharides.
All spectra showed the presence of a broad absorption between 1500 and 670 cm1 , which could be
attributed to C H, C O, and O H bands in the polysaccharides. CPWSP had an XRD pattern that was
typical for a semi-crystalline polymer with a major crystalline reection at 19.6 C. They also displayed
important techno-functional properties (SWC, WSI, WHC, and OHC) that can be modulated according
to temperature. The CPWSP were also noted to display good anti-hypertensive activities. Overall, the
results indicate that CPWSP have attractive chemical, biological, and functional properties that make
them potential promising candidates for application as alternative additives in various food, cosmetic,
and pharmaceutical preparations.
2015 Elsevier B.V. All rights reserved.

1. Introduction
Chickpea is an herbaceous annual plant widely cultivated in
several tropical, sub-tropical, and temperate regions around the
world primarily for its seeds, which are used for both food and
herbal or medicinal purposes. Chickpea seeds have often been
reported to have a wide array of medicinal and therapeutic benets, including the treatment of bronchitis, liver, skin diseases, and
ear inammations [1,2]. It has also been described to offer attractive hypoglycemic and hypocholesteremic effects [3]. Chickpea is
a rich source of dietary protein (1722%), fat (6.48%), and carbohydrate (50%) [4]. These high molecular weight polymers are vital
for several biological functions in the human body and are highly
valued for their multipurpose therapeutic properties, including

Corresponding author. Tel.: +216 54174506; fax: +216 74675761.

E-mail address: besbes.s@voila.fr (S. Besbes).
http://dx.doi.org/10.1016/j.ijbiomac.2015.01.037
0141-8130/ 2015 Elsevier B.V. All rights reserved.

their immuno-modulatory, anti-tumor, anti-inamatory, antioxidant, and anti-pathogenic activities [59].

There is increasing evidence in the literature that several
polysaccharides isolated from various plants, herbal, and fungal
origins have promising therapeutic potential with low cytotoxic
side effects [10,11]. However, and to the authors knowledge,
limited data is currently available on the extraction, characterization, and biological activities of polysaccharides from chickpea.
Accordingly, the present study was undertaken to characterize and
investigate the functional properties and ACE inhibitory effects of
chickpea water-soluble polysaccharides (CPWSP) extracted from
chickpea ours. A hot-water extraction method was used to
produce CPWSP. The latter were submitted to preliminarily characterization by Nuclear magnetic resonance spectroscopy (NMR),
Fourier transform-infrared spectroscopy (FT-IR) analysis, and X-ray
diffractometry (XRD). The functional properties and in vitro ACE
inhibitory activities of CPWSP were investigated seeking for novel
functional components for use in the food, cosmetic, and pharmaceutical industries.

A. Mokni Ghribi et al. / International Journal of Biological Macromolecules 75 (2015) 276282

2. Materials and methods

2.1. Reagents
The chemicals and solvents used in the present study were purchased at the analytical grade or highest level of purity available.
Angiotensin converting enzyme (ACE) from rabbit lung and the
ACE synthetic substrate hippuryl-l-histidyl-l-leucine (HHL) were
purchased from Sigma Chemical Co. (St. Louis, MO, USA).
2.2. Materials
Seeds of chickpea cultivar were dehulled and ground to pass
through a 0.5-mm sieve to obtain chickpea our. The samples were
stored in sealed plastic bags at 20 C. To remove fat, 100 g of
chickpea ours were placed in a dark ask and homogenized using
Ultra-Turrax T25 homogenizer set (Janke & Kunkel IKA Labortechnik, Staufen, Germany) with 300 ml of hexane. After mixing for
4 h in a shaker (Selecta, Spain) at a rate of 180 rpm/min, the mixture was centrifuged for 15 min at 3000 g at ambient temperature
(20 C). The extraction procedure was repeated twice and the pellet
was used to produce CPWSP.
2.3. Extraction of CPWSP
CPWSP were recovered by the method of Liu et al. [12]. Chickpea our was pre-extracted with 95% ethanol at room temperature
to remove small molecules. The dry residue was extracted twice
with 20 volumes of deionized water at 90 C for 4 h with stirring. The extract was combined and ltered, and the ltrates
were then evaporated under vacuum. The concentrated liquid
was precipitated with 95% (v/v) ethanol at 4 C for 24 h and then
centrifuged (4500 g) for 15 min using a refrigerated centrifuge
(Hettich Zentrifugen, ROTINA 380R, Germany). The nal precipitate was re-dissolved in double distilled water. The water phase
was dialyzed at 4 C against running tap water for 2 days and then
against distilled water for an extra day, and the dialysate was concentrated by rotary evaporation under reduced pressure (Rotary
evaporator, Heidolph, Germany) and freeze-dried to obtain CPWSP.
2.4. Determination of chemical composition
The moisture and ash content in the samples were determined according to the AOAC standard methods 930.15 and 942.05,
respectively [13]. The fat and protein (N 6.25) contents were estimated using standard analytical methods [14]. Total carbohydrates
were determined by the phenolsulphuric acid method [15]. The
percentage extraction yield (%) was calculated with the formula
below:
Y (%) = 100% W1 /W0
W1 was the polysaccharide weight of extraction (g), and W0 represented dried sample weight (g).
The colors of the samples were evaluated by a spectrophotocolorimeter Mini Scan XETM (HunterLab Inc., Reston, VA, USA) using
the CIE Lab co-ordinates (L*, a*, b*). In this coordinate system, the L*
value refers to a measure of lightness, ranging from 0 (black) to 100
(white); the a* value refers to a chromatic scale ranging from 100
(greenness) to +100 (redness); and the b* value refers to a chromatic
scale ranging from 100 (blueness) to +100 (yellowness).
2.5.

13 C

CP/MAS NMR spectroscopic analysis

Spectroscopic analysis was performed out by 13 C NMR with

the CP/MAS technique (cross-polarization, magic-angle-spinning)

277

using a BRUKER-ASX300 instrument. NMR spectra were recorded at

a 13 C frequency of 75.5 MHz (eld of 7.04 T). The CP/MAS sequence
was used with the following parameters: the 13 C spin lattice relaxation time was 5 s, powdered samples were placed in an alumina
rotor used for the double air-bearing-type MAS system and spun at
a speed of 8 kHz; the contact time was 8 ms.
2.6. Infra-red spectroscopic analysis
The absorption spectra of CPWSP were obtained by FT-IR spectroscopy (Analect Instruments fx-6 160). The FTIR spectra were
recorded between 400 and 4000 cm1 in a NICOET spectrometer.
The transmission spectra of the samples were recorded using a KBr
pallet containing 0.1% of sample.
2.7. X-ray diffraction
The X-ray diffraction (XRD) patterns of the CPWSP powders
was measured by a Brker D8-Advance Diffractometer (Brker,
Germany) equipped with an Anton Paar temperature control system consisting of a TTK450 low-temperature chamber connected
to a water bath and heating device (TCU 110 Temperature Control
Unit) (Anton Paar, Austria). The data were collected in the 2 ranges
580 with a step size of 0.05 and a counting time of 5 s/step.
2.8. Functional properties
2.8.1. Determination of hydration properties
Water-holding capacity (WHC) was measured using a slightly
modied version of the method described by Sosulski [16]. In brief,
CPWSP (1 g) was placed in a centrifuge tube and dispersed in 25 ml
of distilled water. The dispersions were stirred and left at 25, 40, 60,
or 80 C for 1 h, followed by centrifugation for 25 min at 3000 g.
The supernatant was collected, excess of water was removed by
draining for 25 min at 50 C, and the sample was reweighed. WHC
was expressed as grams of water bound per gram of sample on a
dry basis.
Water solubility index (WSI) was determined by the AACC
method No. 4419 [17]. The powder (S1 , g) was dispersed in a centrifuge tube by adding water with a powder/water ratio of 0.02/1
(w/w) at ambient temperature. The dispersion was then incubated
in a water bath at 80 C for 30 min, followed by centrifugation at
1100 g for 10 min. The supernatant was carefully collected in a
pre-weighed evaporating dish (S2 , g) and submitted to drying at
103 2 C. The evaporating dish with residue was weighed again
(S3 , g). WSI was calculated using the following formula:
WSI (%) = (S3 S2 )/S1 100%.
Swelling capacity (SC) was determined according to the method
of Lecumberri et al. [18]. An initial amount of 1 g powder was
recorded when the sample was poured into a graduate cylinder,
and its occupied bed volume (V1 ) was registered. After that, 10 ml
of distilled water was added, and the mixture was shaken until a
homogeneous dispersion achieved. The dispersion was incubated
in a water bath at 25 C for 24 h to allow the complete swelling of
the powder. The new volume (V2 ) of the wetted powder was then
recorded. SC was calculated using the following formula:
SC (ml/g) = (V2 V1 ) /M.
2.8.2. Determination of oil holding capacity (OHC)
OHC was measured by a partially modied version of the
method described by Lin et al. [19]. In brief, CPWSP (0.5 g) were
mixed with 10 ml of soybean oil, and the dispersions were stirred
and left at 25, 40, 60, or 80 C for 1 h at room temperature. The

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A. Mokni Ghribi et al. / International Journal of Biological Macromolecules 75 (2015) 276282

CPWSP solutions were then centrifuged at 5000 g for 20 min. The

oil supernatant was removed and the centrifuge tube was drained
for 30 min on a lter paper after tilting to a 45 angle. Their capacities were calculated as the weight of the contents of the tube after
draining divided by the weight of the dried CPWSP, and expressed
as the weight % of dried CPWSP.
2.8.3. Determination of foaming properties
Foam capacity (FC) and foam stability (FS) were determined
according to a slightly modied version of the method described
by Lin et al. [19]. In brief, 50 ml of sample were mixed in distilled
water and homogenized for 3 min using an ULTRA-TURAX T 25
basic (IKA WERKE). The mixture was immediately transferred into
a graduated cylinder. The volume was recorded before and after
whipping.
Foam capacity (FC) was expressed as foam expansion at 0 min,
which was calculated according to the following equation:
VT V0
FC (%) =
100.
V0
Foam stability was calculated as the volume of foam remaining
after 30 and 60 min.
FS (%) =

Vt V0
100
V0

where VT is the total volume after whipping (ml); V0 is the original

volume before whipping; Vt is the total volume after leaving at
room temperature for different times (30 and 60 min).
2.8.4. Determination of emulsion properties
Emulsifying activity and stability indices of CPWSP were determined by the method described by Pearce and Kinsella [20].
Emulsions were prepared by homogenizing 50 ml of CPWSP solution with 2 ml of soybean oil for 1 min at speed 3 using an
ULTRA-TURAX T 25 basic (IKA WERKE). A volume of 100 l of emulsion sample was immediately taken from the bottom of the tube
and diluted in 7.5 ml of 10 mM sodium phosphate buffer (pH 7.0)
containing 0.1% sodiumdodecyl sulphate (SDS). The solution was
then vortexed for 10 s. An aliquot was taken from the suspension at
10 min, and the absorbance of the diluted emulsion was measured
at 500 nm. Emulsifying activity index (EAI) and the emulsion stability index (ESI) were calculated according the following equations:

EAI (m2 /g) =

ESI (min) =

2.2.303.A0 .N
C..10000

A0
t
A

where, A0 refers to the absorbance of the diluted emulsion immediately after homogenization, N to the dilution factor (150), C to the
weight of CPWSP per volume (g/ml), to the oil volume fraction
of the emulsion, A to the change in absorbance between 0 and
10 min (A0 A10 ), and t to the time interval of 10 min.
2.9. Determination of ACE inhibition activity
The ACE inhibition activity was measured in triplicate as
described by Nakamura et al. [21]. A sample solution (80 l) containing different concentrations (15 mg/ml) of CPWSP was mixed
with 200 l of 5 mM HHL, and then pre-incubated for 3 min at 37 C.
GPHs and HHL were prepared in 100 mM borate buffer (pH 8.3) containing 300 mM NaCl. The reactions were then initiated by adding
20 l of 0.1 U/ml ACE from rabbit lung prepared in the same buffer.
After incubation for 30 min at 37 C, the enzyme reactions were

Table 1
Proximate composition and hydration properties of water-soluble polysaccharides
from chickpea our. Physico-chemical composition was calculated basis on the dry
mater.
Parameters

CPWSP

Yield (%)
Dry matter (%)
Polysaccharides (%)
Proteins (%)
Lipids (%)
Ash (%)
CIE color
L*
a*
b*
Hydration properties
Water holding capacity (g water/g dry weight)
Water solubility index (%)
Swelling capacity (ml/g)

5.56 0.13
95.96 0.7
73.23 0.9
11.22 0.63
5.4 0.18
1.12 0.08
95.6 0.05
2.63 0.04
12.36 0.00
5.14 0.25
22.62 1.25
6.4 0.3

All the data are expressed as mean SD and are the mean of three replicates.

stopped by the addition of 250 l of 0.05 M HCl. The liberated hippuric acid (HA) was extracted with ethyl acetate (1.7 ml) and then
evaporated at 90 C for 10 min. The residue was dissolved in 1 ml
of distilled water, and the absorbance of the extract at 228 nm was
determined using an UVvisible spectrophotometer (UV mini 1240,
UV/VIS spectrophotometer, SHIMDZU, Japan).
The average value from three determinations at each concentration was used to calculate the ACE inhibition rate as follows:

ACE inhibition (%) =

BA
100
BC

where A is the absorbance of HA generated in the presence of

ACE inhibitor, B is the absorbance of HA generated without ACE
inhibitors (100 mM borate buffer pH 8.3 was used instead of
CPWSP), and C is the absorbance of HA generated without ACE
(corresponding to HHL autolysis in the course of enzymatic assay).
2.10. Statistical analysis
Analytical values were carried out using three independent
determinations. Results were expressed as mean values standard
error of three independent determinations. Statistical analyses
were determined using a statistical software program (SPSS for
Windows version 11.0). The data were subjected to analysis of
variance using the general linear model to determine signicant
differences between samples (p < 0.05).
3. Results and discussion
3.1. Chemical composition of CPWSP
In this study, CPWSP were obtained by hot water extraction,
as well as ethanol precipitation, deproteination, and dialysis. As
shown in Table 1, the yield of crude polysaccharide obtained was
5.56%. These results were similar to those previously obtained
from Pharbitis nil seeds [22]. The CPWSP had relatively low moisture (4%) and was rich in protein (11.22%). The high protein
content showed that part of the protein is not removed with
the applied treatment, possibly because the residual protein
is too closely bound to the cell wall structure indicating that
polysaccharideprotein complexes still resided in the CPWSP.
Higher protein levels were reported for water-soluble polysaccharides from Citrus aurantium L. (17.01%) in the work of Wang
et al. [23]. In another study, Wang et al. [24] showed lower protein
content for Phellinus linteus polysaccharides (1.53% 0.12). In fact,
protein content depends on the extraction and deproteination

A. Mokni Ghribi et al. / International Journal of Biological Macromolecules 75 (2015) 276282

279

Fig. 2. FT-IR spectrometry of water-soluble polysaccharides from chickpea our.

Fig. 1. 13 CP/MAS NMR solid-state spectra of water-soluble polysaccharides from

chickpea our.

processes. The ash and fat contents were 1.12% and 5.4%, respectively. Abdul-Hamid and Luan [25] reported an ash content of
7.41 0.01% in polysaccharides prepared from rice bran. The
variation in the ash content of the CPWSP could presumably be
attributed to the solvents used for their preparation.
The colors of the CPWSP expressed in terms of L*, a*, and b* are
shown in Table 1. In fact, color is one of the esthetic properties that
determine the suitability of CPWSP to the application for which it
is intended. The high L* values indicated that CPWSP had a light
color, and the a* values pointed for its slightly red color.
3.2. NMR spectroscopy
The chemical structures of the CPWSP were also examined by
NMR spectroscopy. The 13 C NMR spectra are shown in Fig. 1.
The complexities of the spectra reected the heterogeneity of the
polysaccharides. The 13 C NMR spectra of the CPWSP revealed that
the 1,4-d-galacturonan with methyl-esteried carboxyl group was
present in the sugar chain, giving rise to the anomeric carbon
resonance at 103.07 ppm. Predominant signals at 81.5 ppm were
assigned to C4 of 1,4--d-galactopyranosyluronan [26]. Signals at
53.36 ppm represented the methyl carbons of the methyl ester
(COOCH3 ) [27].
13 C

3.3. FT-IR spectroscopy

FT-IR spectra were performed in the 4000400 cm1 region
to further characterize the polysaccharides present in the CPWSP
under investigation.
The spectrum presented in Fig. 2 shows the characteristic
absorption of CPWSP. The stretching vibration of hydroxyl groups
falls within the wave number range of 36003200 cm1 [28].
The broad and pure peak observed at 3454 cm1 indicated an
intermolecular hydrogen bonding. The small bands recorded at
2910 cm1 were attributed to the C H anti-symmetrical stretching
vibrations. The weak absorption at 2360 cm1 indicated the presence of aliphatic C H bonds [29]. The bands at around 1647 cm1
represented the carboxylate (COO ) stretching band [30], indicating the presence of free carboxyl groups and uronic acids in the
polysaccharides from chickpea. Carbohydrates in the FT-IR spectra
showed broad absorbance in the region of 1200850 cm1 , which
was dominated by ring vibrations that overlapped with stretching
vibrations of the (C OH) side groups and the (C O C) glycosidic
bond vibration [31]. This region, called as a nger print region,

could be related to conformation and surface structure of molecule

[29]. The main absorbance regions between 1000 and 1200 cm1
represented the galacturonic acid in pectic polysaccharides [32].
Those peaks exhibited by CPWSP were characteristic of pectic
polysaccharides, revealing that the CPWSP were rich in uronic acid.
3.4. XRD
XRD analysis was applied to determine the crystalline degree of
the extracted CPWSP. The XRD pattern presented in Fig. 3 was typical for a semi-crystalline polymer and indicated major crystalline
reections at 19.6 . The results showed that the amorphous component of CPWSP prevailed over the crystalline organized one. This
structural arrangement is known to directly affect various properties, including tensile strength, exibility, solubility, swelling, or
opaqueness of the bulk polymer. In fact, physical properties are
dependent on the degree of order within the material [33].
3.5. Functional properties of CPWSP
3.5.1. Hydration properties
Considering their importance with regard to food functionalities, the hydration properties of CPWSP were investigated using
three methods, namely WHC, WSI, and SC. The WHC refers to the
ability of a moist material to retain water when subjected to an
external centrifugal gravity force or compression. It is the sum of
bound water, hydrodynamic water and, mainly, physically trapped
water [34]. The WHC evolution of CPWSP at different temperatures
(25, 40, 60, and 80 C) is shown in Fig. 4.
An increase in the WHC for CPWSP at high temperatures was
observed. This increase could presumably be attributed to the
increase in the solubility of the polysaccharides due to the rise in

Fig. 3. X-ray diffraction pattern of polysaccharides from chickpea our.

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A. Mokni Ghribi et al. / International Journal of Biological Macromolecules 75 (2015) 276282

Fig. 4. Evolution of the water holding capacity (WHC) of water-soluble polysaccharides from chickpea ours (25, 40, 60, and 80 C) (mean SD in g water/g of
dry weight; n = 3). For each temperature different letters (abc) mean signicant
differences (p < 0.05, Duncan test).

temperature. In this study, the WHC values recorded at 25 and 80 C

were about 5.14 and 7.1 g of water per g of dry matter, respectively. While signicant differences (p < 0.05) were observed in
the WHC values recorded at 25, 40, and 60 C, no signicant differences (p > 0.05) were observed at 60 and 80 C. Sciarini et al.
[35] reported high water retention capacities for galactomannans
(15.20 g H2 O/g sample). Although lower than those reported for
other polysaccharides, the water retention capacity displayed by
CPWSP suggested their adequacy for use as functional ingredients
in food products.
The SC and WSI of CPWSP at 25 C were also investigated. Table 1
shows that CPWSP displayed WSI and SC values of 22.62 g/100 g
and 6.4 g/ml, respectively. Ayadi et al. [36] reported that WSI were
related to the presence of soluble molecules. Swelling values ranging from 7.1 to 40.6 ml/g were reported by Kosmala [37] for Plum
fruits. The results of SC and WHC are similar to those of apple
pulp and Spanish edible seaweeds [38,39]. SC depends on several
factors, including structural characteristics, chemical composition,
and particle sizes.
3.5.2. OHC
OHC is an important feature of polysaccharides and a technological property related to the chemical structure of plant
polysaccharides. The evolution of OHC of CPWSP at different temperatures (25, 40, 60, and 80 C) is presented in Fig. 5.
The OHC of the CPWSP powder was noted to undergo a
slight decrease with the increase in temperature. The OHC values
recorded at 25 and 80 C were about 3.15 and 2.12 g/g, respectively.
Sila et al. [40] reported on higher OHC of 8.48 g oil/g of sample and
7.40 0.06 g oil/g sample for pistachio and almond water-soluble
polysaccharides. These differences in terms of OHC values could
be ascribed to the high amount of lipids in the initial matrix. Furthermore, while signicant differences (p < 0.05) were observed
between the OHC values obtained at 25 and 40 C, no signicant
differences (p > 0.05) were observed for those recorded at 40, 60,
and 80 C. This could presumably be linked to the adsorption of
organic compounds in the surface of the substrates [41]. Reported
OHC values of pea (0.13 g/g) and okara (0.27 g/g) [42] are lower than
those found in CPWSP tested in the present work.
3.5.3. Foaming properties
Foams are two-phase systems consisting of a dispersed
phase (usually air) and a continuous phase [43]. Due to their

Fig. 5. Evolution of the oil holding capacity (OHC) of water-soluble polysaccharides

from chickpea ours (25, 40, 60, and 80 C) (mean SD in g water/g of dry weight;
n = 3). For each temperature different letters (abc) mean signicant differences
(p < 0.05, Duncan test).

predominantly hydrophilic characteristics, polysaccharides generally remain in the aqueous subphase, performing as thickeners
and stabilizers [44]. Accordingly, polysaccharides are normally
included as stabilizing and thickening agents in colloid systems.
The foam capacity (FC) and foam stability (FS) of CPWSP at different
concentrations (0.5%, 1%, 2%, and 4%; w/v) are shown in Table 2.
The increase of CPWSP concentration resulted in an increase
in foam stability, which was more pronounced with high concentrations. Liquid drainage was also reduced with the addition
of polysaccharides. The foaming properties were better when
the polysaccharides concentration increased, which could be
attributed to the fact that polysaccharides have the ability to
increase the viscosity of the aqueous phase and to create a network
that stabilizes the interfacial lm (airwater). Adequate CPWSP
concentrations are needed to cover the air bubble surface, create a
rigid lm around it, and, hence, produce more stable foam.
3.5.4. Emulsion properties
Due to their strong hydrophilic nature, polysaccharides are generally considered as non surface-active agents. Conversely, proteins
are used as emulsiers due to their hydrophilic and hydrophobic
side chains, which make them effective surface-active agents. In
fact, the presence of small fractions of proteins in some hydrocolloids has often been reported to be responsible for observable
emulsifying properties [45]. Surface-active polysaccharides can,
therefore, act both as emulsiers and stabilizers. The EAI and ESI
of CPWSP at different concentrations are shown in Table 3.
The EAI values of CPWSP decreased with the increase of polysaccharide concentrations. This decrease could be due to a reduction
in protein surface hydrophobicity [44]. At low polysaccharides concentration, the repulsion between proteins and polysaccharides
Table 2
Foaming properties of water-soluble polysaccharides from chickpea our at different concentrations.
Concentration
(g/100 ml)

FC (%)

0.5
1
2
4

48.25 0.23a
54.62 0.83b
62.5 0.55c
91.16 1.12d

FS (%)
30 min
30.45
43.25
53.17
79.82

0.12a
0.63b
0.56c
1.02d

60 min
14.15
22.64
35.23
65.12

0.45a
0.25b
0.78c
0.69d

Means with the different superscript letters within the same column are signicantly different (p < 0.05). FC, foam capacity; FS, foam stability.

A. Mokni Ghribi et al. / International Journal of Biological Macromolecules 75 (2015) 276282

Table 3
Emulsion activity index (EAI) and emulsion stability index (ESI) of water-soluble
polysaccharides from chickpea our at different concentrations.
Concentration (%)
0.5
1
2
4

EAI (m2 /g)

46.42
31.6
27.72
25.36

0.47a
0.16b
0.12c
0.4d

ESI (min)
34.67
20.06
17.07
15.36

2.12a
1.25b
1.03c
0.98c

Means with the different superscript letters within the same column are significantly different (p < 0.05). EAI, emulsifying activity index; ESI, emulsion stability
index.

in solution could increase the exposure of protein hydrophobic

regions [46].
At high polysaccharide concentrations, the formation of
proteinpolysaccharide complexes could sterically hinder the surface hydrophobic areas on the protein, thus leading to the decrease
of their diffusion rate [47]. It could, therefore, be inferred that different interactions between the proteins and polysaccharides in
the solution could exert a great effect on the emulsion properties.
Due to their ability to form strong interfacial lms, polysaccharides
have often been used as ingredients to stabilize oil-in-water food
emulsions [48,49]. The ndings showed no signicant differences in
the emulsion stability indexes at high concentrations. These results
indicated that the thickness of the adsorbed layer and the steric
repulsion between the oil droplets were preserved.
3.6. ACE inhibitory activities of CPWSP
The inhibition of ACE by dietary anti-hypertensive agents is
an important hypertension management strategy. In fact, ACE
inhibition has often been considered as a useful therapeutic
approach for the treatment of high blood pressure by virtue of
the reninangiotensin system (RAS) and kallikrein kinnin system
(KKS). While ACE inactivates the vasodilator bradykinin in the case
of KKS, it acts as an exopeptidase that cleaves HisLeu from the
C-terminal of decapeptide angiotensin I and produces the potent
vasoconstrictor octapeptide angiotensin II in the case of RAS [50].
Since the synthetic ACE inhibitors may cause adverse side effects,
plant molecules could be used as natural and economical ACE
inhibitors for hypertension treatment and prevention [40].
The results of the present study revealed that the ACE activity of CPWSP was concentration dependent. The values increased

Fig. 6. ACE-inhibitory activities of water-soluble polysaccharides from chickpea

ours at different concentration. Values are given as mean SD from triplicate determinations. Values are given as mean SD from triplicate determinations. Identical
letters above the bars indicate no signicant differences by SPSS test (p 0.05).

281

with increasing concentrations (Fig. 6). The highest ACE inhibitory

activity (87.83 0.8%) was observed at 1 mg/ml for CPWSP. Sila
et al. [40] reported on anti-hypertensive activities of 79.5 2.8%
and 81.78 1.1% at 5 mg/ml for almond and pistachio watersoluble polysaccharides, respectively. The sulfated polysaccharides
extracted from squid skin exhibit strong ACE-inhibitory activity,
with the highest levels (86.3%) being observed at a concentration
of 1 mg/ml [51].
4. Conclusion
The present study was undertaken to characterize water-soluble
polysaccharides isolated from chickpea and investigate their functional properties and anti-hypertensive activities. The results from
the physico-chemical characterization analyses by FT-IR, NMR,
and XRD showed that CPWSP corresponded to a semi-crystalline,
polymer-forming polysaccharide protein complex with good emulsifying activity and stability and high WHC and OHC properties.
CPWSP was also noted to inhibit ACE. Overall, the results indicated that CPWSP could be considered as a promising source of
ACE-inhibitory agents could open new opportunities for the development of effective techno-functional additives for use in a wide
range of food, cosmetic and pharmaceutical formulations. CPWSP,
with their high OHC, may be better suited for confectionery applications requiring oil emulsication. Moreover, CPWPS could be
useful as a thickening or functional agent for food systems such
as baked goods, beverages, ice cream, and yoghurt. Accordingly,
further studies, some of which are currently underway in our laboratories, are needed to investigate the optimal conditions required
for the over production of these promising polysaccharides.
Acknowledgements
The authors would like to express their sincere gratitude to
Mr. Anouar Smaoui and Mrs. Hanen Ben Salem from the English
Language Unit at the Faculty of Science of Sfax for their valuable
proofreading and language polishing services.
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