Food Hydrocolloids 67 (2017) 54e62

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

Effects of axseed gum concentrations and pH values on the stability

of oil-in-water emulsions
Meng Wang a, Mei-qin Feng b, Kun Jia a, Jian Sun a, *, Xing-lian Xu a, Guang-hong Zhou a
a
National Center of Meat Quality and Safety Control, College of Food Science and Technology, Nanjing Agricultural University, and with Jiangsu
Collaborative Innovation Center of Meat Production and Processing, Quality and Safety Control, Nanjing, 210095, PR China
b
College of Animal Science and Technology, Jinling Institute of Technology, Nanjing, 210038, PR China

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

Article history: This study investigated the effects of axseed gum (FG) concentrations (0.1e0.5%, w/w) and pH values
Received 1 September 2016 (5.0, 6.0 and 7.0) on the stability of 10% (w/w) liquid lard emulsions. The emulsions were analyzed for
Received in revised form particle size, rheological properties, creaming stability, optical microscopy and nuclear magnetic reso-
29 December 2016
nance (NMR) measurement. With the increase of FG concentration, there was an improved packing of
Accepted 2 January 2017
Available online 3 January 2017
polysaccharide onto the droplet surface, which caused a signicant reduction in particle size, with the
smallest oil droplets being formed in 0.5% (w/w) FG emulsions. Rheological and creaming stability
measurements showed that FG solutions had thickening and gelling properties. When FG concentration
Keywords:
Flaxseed gum
increased to greater than 0.3% (w/w), the samples no longer exhibited creaming behavior, and pH values
Emulsions no longer had signicant effect on the emulsion stability. Shorter spin-spin relaxation time (T2, by low-
Particle size eld 1H NMR) were observed for emulsions containing 0.1% (w/w) and 0.2% (w/w) FG, suggesting more
Rheology restriction of mobility of oil droplets. The increased line-width in high-eld 1H, 13C NMR spectra with
Creaming increasing FG concentration indicated an increased interaction between FG and oil molecules. These
Nuclear magnetic resonance results showed that FG had the potential to substitute for protein emulsiers which is sensitive to pH to
stabilize the emulsion-type products.
2017 Elsevier Ltd. All rights reserved.

1. Introduction more research is needed to develop different and novel

Hydrocolloids, a heterogeneous group of high molecular weight
hydrophilic polymers, have a wide array of functional properties in
foods. One of the key functional roles is to control the micro-
structure and shelf-life of emulsions (Dickinson, 2003; Garti, 1999).
Previous studies have conrmed that some hydrocolloids can act as
emulsifying agents, which are absorbed at the surface of the freshly
formed ne droplets and thus prevent them from coalescing with
their neighbors to form larger droplets again (Dickinson, 2009).
Hydrocolloids are also used as stabilizers due to their water-
thickening property which is conrmed vital in stabilizing emul-
sions (Saha & Bhattacharya, 2010). Compared with protein, a nat-
ural and commonly used food emulsifying agent, hydrocolloids are
less sensitive to the change of environmental conditions (i.e., pH,
ionic strength, etc.). With the growing demand for hydrocolloids
indicated by the MicroMarket Monitor's report (Li & Nie, 2016),
* Corresponding author. 1 Wei Gang, Nanjing, Jiangsu, 210095, PR China.
E-mail address: sunjian01@njau.edu.cn (J. Sun).
hydrocolloids.
Flax (Linum usitatissimum L.) is one of the oldest crops known to
man (Oomah & Mazza, 1993). Flaxseed, the seed of ax, contains
many functional components such as lignans, protein, poly-
unsaturated fatty acid and gum (Cardoso Carraro, Dantas, Espeschit,
Martino, & Ribeiro, 2012; Wang et al., 2007; Zhang et al., 2007). FG,
which accounts for 8% of the seed, is of special interest owing to its
functional properties when dispersed in aqueous solutions (Liu,
Shim, Poth, & Reaney, 2016). Wang et al. (2008) found that both
the apparent viscosity and consistency index of native maize starch
are increased with the increasing addition of FG. Similar results
were also reported by other authors (Derkach, Zhabyko, Voron'ko,
Maklakova, & Dyakina, 2015; Hussain et al., 2015; Wang, Wang,
Li, Xue, & Mao, 2009). Like most hydrocolloids, FG plays an
important role in stabilizing oil-in-water emulsions, and has good
water-holding and thickening capacities. More interestingly, FG
shows weak gel property thus we can use it to replace the non-
gelling gums (Cui & Mazza, 1996). However, there is little infor-
mation on application of FG in the food industry, especially in the
emulsied meat products.

http://dx.doi.org/10.1016/j.foodhyd.2017.01.004
0268-005X/ 2017 Elsevier Ltd. All rights reserved.
 M. Wang et al. / Food Hydrocolloids 67 (2017) 54e62
Emulsion-type meat products are made using crush and emul-
sication processing methods. Fat in meat products plays an
important role in textural property, water holding capacity, cooking
loss and meat emulsion stability (Choe, Kim, Lee, Kim, & Kim, 2013).
Nevertheless, high fat intake is associated with cardiovascular
disease, hypertension and obesity. Therefore, more and more hy-
drocolloids have been used in the meat products to replace fat and
improve the product texture and stability. To characterize the
properties and stability of emulsions, many traditional techniques,
such as rheology, particle size distribution and microscopy have
been studied (Garti, Slavin, & Aserin, 1999; Hosseini-Parvar, Osano,
& Matia Merino, 2016; Nakauma et al., 2008). Recently, NMR, an
important tool to research the emulsion-type food, is a fast and
accurate technique that has been proved immensely valuable. And
another important advantage of the NMR application is that it can
be used without dilution and pre-treatment of the emulsions. It has
been used to determine the total fat and moisture content in meat
(Srland, Larsen, Lundby, Rudi, & Guiheneuf, 2004), the stability of
emulsions (Opedal, Srland, & Sjoblom, 2010), the droplet size
distributions (Hollingsworth, Sederman, Buckley, Gladden, & Johns,
2004; Lingwood, Chandrasekera, Kolz, Fridjonsson, & Johns, 2012)
and the inuence of freezing on emulsions (Hindmarsh,
Hollingsworth, Wilson, & Johns, 2004).
The aim of this research was to investigate the effects of FG
concentrations and pH values on the stability of oil-in-water
emulsions. To simulate the meat system, liquid lard was supplied
to prepare the emulsions. Except for particle size, creaming index,
rheology and microscopy, the classic indicators to measure the
stability of emulsions, we also employed NMR spectroscopy to
provide further information to reveal the mechanism of FG in sta-
bilizing emulsions.
2. Materials and methods
2.1. Materials
FG (powder, purity 99.8%, containing 16.09% protein, 5.10% wa-
ter, 4.35% ash) was purchased from Xinjiang Lvqi Biotechnology Co.,
Ltd. (Xinjiang uygur autonomous region, China). Fresh pork back fat
was obtained from a local supermarket. It was vacuum packed,
frozen and stored at
20  C until use. Deuterium oxide was pur-
chased from Aladdin Industrial Corporation (Shanghai, China). All
other chemicals were of analytical grade.
2.2. Sample preparation
Frozen pork back fat was thawed for 24 h at 4  C prior to use. The
samples were cut into small pieces and heated at 100e120  C with
constant agitation. Four layers of cheesecloth were used to remove
impurities of the liquid lard.
FG was dissolved in deionised water to give a nal concentration
of 0.1%, 0.2%, 0.3%, 0.4% and 0.5% (w/w) and gently stirred with
magnetic stirrer overnight to assure complete hydration. The
Table 1
The average particle size D4,3 of FG-stabilized oil-in-water emulsions.
pH FG concentration (%)
0.1 0.2
5.0 61.00 1.40aA 43.33 0.15bA
6.0 50.27 0.15aB 37.53 0.21bB
7.0 45.30 0.26aC 36.77 0.45bC
Note: values were mean of triplicate values standard deviations.
a ee
different letters in the same raw indicate statistically signicant differences at P < 0.05.
A
C
different letters in the same column of treatment indicate statistically signicant differences at P < 0.05.
55
obtained gum solutions were then adjusted to pH 5.0, 6.0 and 7.0
with HCl or NaOH.
Pre-emulsions were made by adding 10% (w/w) liquid lard to the
prepared FG solutions and were homogenized using a high-speed
homogenizer (IKA T25-Digital Ultra Turrax, Staufen, Germany) for
2 min at 12,500 rpm. The pre-emulsions were then passed through
a high-pressure homogenizer at 500 bar for the rst pass and
300 bar for further pass. All of these procedures were carried out at
about 40  C to prevent the solidication of liquid lard in the
emulsion preparation process. The nal composition of the emul-
sions was 10% (w/w) lard and 0.1e0.5% (w/w) FG. Before further
analysis, the prepared emulsions were kept in a water bath at 50  C
for 30 min and analyzed within 24 h.
2.3. Measurement of particle size
The measurement of particle size distribution of the emulsions
was performed using a Malvern Mastersizer 3000 instrument
(Malvern Instruments Ltd, Worcestershire, UK) and the data were
analyzed using the Mastersizer 3000 software. The Mastersizer
determines the droplet size distribution of emulsions by using the
Mie theory (Hattrem, Dille, Seternes, & Draget, 2014). The average
droplet size was represented by mean volume diameter D4,3 which
was more sensitive to fat droplet occulation/coalescence than
D3,2 (Ahmed, Taher, Mulla, Al-Hazza, & Luciano, 2016). Samples
were dispersed into recirculating water until the obscuration was
among 6%e10%. The refractive and absorption index of the sample
particles were set as 1.520 and 0.001, respectively. The dispersant
was water and it's refractive index was taken as 1.330. The mea-
surement was conducted immediately after the emulsions were
prepared. Average of three readings was obtained for each test.
2.4. Rheological properties
Rheological measurements were carried out using a Physica
MCR 301 rheometer (Anton Paar, Graz, Austria), which was tted
with a steel parallel plate geometry (25 mm diameter, 1 mm gap).
The frequency sweep tests were performed at 25  C over the
angular frequency range of 1e100 rad/s. The strain amplitude for
frequency sweep was selected as 2%. Storage modulus (G0 ) and loss
modulus (G00 ) were recorded. Continuous shear tests were per-
formed at 25  C over the shear rate range of 0.01e100 s
1 to
measure the apparent viscosity.
2.5. Creaming stability measurement
Ten milliliters of emulsions were transferred into a glass test
tube and maintained at ambient temperature for 24 h. During the
storage, the emulsions were separated into a cream layer at the top
and a serum layer at the bottom. The total height of the emulsions
(HE) and the height of the serum layer (HS) were measured. The
extent of creaming was calculated by creaming index
(%) 100  (HS/HE).
0.3 0.4 0.5
30.37 0.38cA 25.03 0.35dA 23.23 0.31eA
29.47 0.40cB 23.37 0.35dB 20.03 0.35eB
27.23 0.21cC 23.33 0.50dB 20.63 0.25eB
56 M. Wang et al. / Food Hydrocolloids 67 (2017) 54e62

2.6. Optical microscopy
Light microscope was used to observe the state of the samples.
The diluted emulsions (10 times) were visualized under
400  optical microscope. A drop of the diluted emulsion sample
was placed on a microscope slide, and covered with a cover slip.
The microscope was equipped with a CCD camera to capture
images.
2.7. Low-eld NMR (LF-NMR) and high-eld NMR (HF-NMR)
measurements
addition, the rate of hydrocolloids migrate to the oil-water interface
would determine the nal droplet size distribution. Slow migrate
rate and low hydrocolloids concentration are not contributing to
the formation of stable emulsions (Dickinson, 2009). FG is a
mixture of protein and natural polysaccharide mainly composed of
D-xylose, L-galactose, D-rhamnose and D-galacturonic acid (Mazza &
Biliaderis, 1989; Oomah, Kenaschuk, Cui, & Mazza, 1995; Zhao et al.,
2015). . Previous studies reported that the crude and dialyzed FG
contained 9.1% and 6.3% protein respectively, and the content was

Emulsions taken for NMR relaxation measurements were made

with D2O in order to minimize the resonance of water. The LF-NMR
experiments were made using a MicroMR-22MHz spectrometer
(Niumag, MicroMR, shanghai, China). The spin-spin relaxation time
(T2) was measured using the Carr-Purcell-Meiboom-Gill (CPMG)
pulse sequence. Approximately 2.00 g sample was placed in a
15 mm (diameter) NMR tube. Each sample was acquired with 32
scans and a repetition time of 13000 ms, the 90e180 pulse spacing
(t) and EchoCnt were set to 800ms and 6000 ms, respectively. The
CPMG exponential decay curve was carried out with the inversion
software of the instrument. High-eld 1H and 13C NMR spectra
were carried out on a 300 MHz Bruker NMR spectrometer Spec-
trometer (Bruker Group, Fallanden, Switzerland).

2.8. Statistical analysis

SAS 9.2 for windows was used to analyze the obtained results.
Analysis of variance (ANOVA) and a Duncan's multiple range test
were performed on the data and it was presented as the mean
value standard deviation. P<0.05 was as the signicance level. All
of the results were conducted in triplicate.

3. Results and discussion

3.1. Measurement of particle size

The effects of FG concentrations and pH values on the mean

volume diameter D4,3 are shown in Table 1. Increasing the con-
centration of FG from 0.1% (w/w) to 0.5% (w/w) at any given pH
values signicantly (P<0.05) reduced the average droplet size. The
reduction was particularly pronounced when FG concentration
below 0.3% (w/w). For example, the reduction of droplet size was
43.83% 4.56% when the addition of FG elevated from 0.1% (w/w)
to 0.3% (w/w) while the reduction was 11.02% 2.93% when FG
concentration increased from 0.4% (w/w) to 0.5% (w/w). The in-
crease in pH value, from 5.0 to 7.0, also resulted in the reduction of
droplet size. Nevertheles, D4,3 was not signicant (P > 0.05)
different at pH 6.0 and pH 7.0 when the FG concentration exceeded
0.3% (w/w), which indicated that the mean diameter of the droplets
was no longer dependent on the pH values. Farshchi, Ettelaie, and
Holmes (2013) have reported that samples with higher locust
bean gum concentration had a smaller D4,3 and when compared
with pH 5.0, 5.5 and 6.5, emulsions at pH 5.0 exhibited the largest
particle size, which was consistent with our results. However,
Nakauma et al. (2008) found that the droplets size (D3,2) of oil-in-
water emulsions stabilized by sugar beet pectin, soybean soluble
polysaccharide and arabic gum at 1.5%, 4.0%, 10.0%, respectively,
were signicantly independent on pH from 3.0 to 6.0. This could be
caused by the relatively higher concentration of stabilizer used.
The decrease in the droplet size with increasing concentration
may be due to the adsorption of FG at the surface of the emulsion
droplets, which prevents the occulation and coalescence. As a Fig. 1. The apparent viscosity of emulsions with different concentration of FG at pH 5.0
stabilizer, FG can migrate to the interface of oil and water. In (A), 6.0 (B) and 7.0 (C).
 M. Wang et al. / Food Hydrocolloids 67 (2017) 54e62
Fig. 2. The storage modulus (G0 , A) and loss modulus (G00 , B) of emulsions with different concentration of FG at various pH values.
governed by various parameters, such as the origin, the species and
the extraction process of FG (Cui, Mazza, & Biliaderis, 1994). It is
well known that proteins in FG play an important role in its
emulsifying properties. Therefore, at pH 5.0, which close to the
protein's isoelectric point, the emulsions tend to aggregate to form
larger particles as observed in our studies.
3.2. Rheological properties
FG emulsions showed shear thinning behavior as the apparent
viscosity decreased with the increase in shear rate (Fig. 1), sug-
gesting that the samples exhibit non-Newtonian behavior. With the
increase of FG concentration, the viscosity of samples increased
rapidly. The emulsions with 0.1% (w/w) FG concentration showed
the lowest viscosities which were nearly 10% of the viscosities of
0.5% (w/w) FG samples under high shear rates. The ow curves
obtained at different FG concentrations are in good agreement with
previous studies (Surh, Decker, & McClements, 2006; Zhao et al.,
2015). In contrast, samples with different pH exhibited insigni-
cant differences. The viscosity of samples almost overlapped
especially at high FG concentration, which suggested that the so-
lution itself was resistant to pH changes. Hosseini-Parvar et al.
(2016) also found that the basil seed gum stabilized emulsions
was resistant to pH and ionic strength. The complex viscosity only
became signicantly higher when pH shifted below 2.0, which re-
ects the aggregation of droplets. Emulsion shear thinning
behavior was not only related to the non-Newtonian behavior of
the continuous phase, but also associated with the droplet deoc-
culation of disperse phase (Lorenzo, Zaritzky, & Califano, 2008). At
low shear rates, the stretching hydrocolloid molecules intertwined
to form aggregates, with higher apparent viscosity attributing to
the ow resistance of the continuous phase. Improved shear rate
destroys molecular aggregation and arrangement along the ow
57
direction. Furthermore, uids ow resistance also declines with
increasing shear rate, leading to the decrease of apparent viscosity
(Oh, So, & Yang, 1999).
Dynamic frequency sweep tests of G0 and G00 were performed to
illuminate the structure of emulsion system. It shows that both G0
and G00 of all emulsions increased with increasing angular fre-
quency (0.1e100 rad/s) (Fig. 2). In other words, G0 and G00 values
demonstrated that the viscoelastic properties increased with the
addition of FG, indicating an increased rigidity and a stronger
structure of the samples. Similar trend of G0 and G00 was extensively
observed in emulsions stabilized by various hydrocolloids (Emine
Alben & Esra, 2009; Lorenzo et al., 2008). According to the
research by Emine Alben and Esra (2009), both G0 and G00 increased
with the addition of guar gum and crossing each other at high
frequencies in whey protein isolate stabilized emulsions. The
decreased macromolecular mobility within the network due to the
presence of neutral polysaccharide was considered as one of the
reasons for this phenomenon. The results further illuminated that
the structure of emulsions can be enhanced by the addition of
hydrocolloids. However, the variation of G0 and G00 with frequency
for samples at different pH values were not signicant.
The stronger structure can be attributed to the formation of a
highly occulated droplet network structure or the formation of a
gel-like structure in the continuous phase. Particularly, solutions
with 0.5% (w/w) FG showed a gel-like behavior as G0 was higher
than G00 throughout the frequency range (Fig. 3). Other emulsions,
by contrast, exhibited a liquid-like behavior. Fig. 3 also showed that
G'/G00 decreased slightly as the frequency increased, indicating the
emulsions were weaker than those gel samples which is less
dependent on the frequency. All of these observations suggest that
the rheology is controlled mostly by the FG present in the emul-
sions. Emulsions under different pHs showed no signicant dif-
ference in both G0 and G00 as a function of angular frequency.
58 M. Wang et al. / Food Hydrocolloids 67 (2017) 54e62

3.3. Creaming stability measurement

Emulsion itself is an unstable thermodynamic system. The two

phases trends to separate to form a more stable state to minimize
the free energy. Flocculation and aggregation of oil droplets, Ost-
wald ripening and creaming phenomenon are the factors contrib-
uting to the instability of emulsions. Creaming stability, referred to
the ability of emulsions to resist creaming, coalescence and oc-
culation, was used in our research to measure the emulsion sta-
bility (Sun, Zhou, Sun, & Zhao, 2013).
Fig. 4 showed the effect of FG concentrations and pH values on
the creaming stability of samples. It was noted that the addition of
FG had an obvious inuence on the creaming index of emulsions.
Interestingly, it was found when FG concentration was greater than
0.3% (w/w), emulsions did not exhibit any creaming behavior. In
contrast, a signicant creaming phenomenon occurred in the
diluted emulsions and the creaming index values progressively
decreased with addition of FG. The highest creaming index (70%)
was obtained at 0.1% (w/w) FG, with the creaming phenomenon
immediately occurred after the preparation of samples. This result
is consistent with the consensus that at higher FG concentrations,
there are sufcient hydrocolloids absorbed to the oil/water inter-
face of oil droplets during the emulsication, thus the formed
emulsions are more stable against occulation or coalescence. A
similar result has been found by Zhao et al. (2015) for sodium
caseinate-axseed gum system. Bigger oil droplets were formed at
low FG concentrations due to the insufcient of caseinate-axseed
gum complexes to coat the surface of oil droplets.
Unlike the inuence of FG concentrations, in the present study,
the effect of pH on the creaming index was not so signicantly.
Emulsions with 0.3%e0.5% (w/w) FG had no difference in creaming
index from pH 5.0 to 7.0. This can be explained that hydrocolloids
are commonly perceived to improve emulsion stability by thick-
ening the continuous phase. Concentrated emulsions at different
pH values are stable due to the viscoelastic property of the Fig. 3. Effects of FG concentrations and pH values on frequency sweep of oil-in water
continuous phase that behaves like a gel-like network. Moreover, as emulsions.
shown in Fig. 4, creaming index of emulsions with low FG con-
centration (0.1%, w/w and 0.2%, w/w) decreased with the increased
pH values, suggesting more stable emulsions were formed at higher samples containing FG above 0.3% (w/w), the droplets were
pH values. These results suggest that emulsions prepared at pH 5.0 completely wrapped and distributed evenly to form a stable
are extremely susceptible to the gravitational separation (Farshchi emulsion. These results are consistent with another study from Hu
et al., 2013). et al. (2016) who found that the emulsions stabilized by sodium
caseinate containing low concentration (0.5%) of regenerated cel-
3.4. Optical microscopy lulose (RC) had lager droplets and a greater occulation. However,
particle size and occulation were both decreased and inhibited in
As mentioned above, different FG concentration is the most the samples containing RC from 0.5% to 1.5%. In addition, our results
signicant factor for the creaming stability. Thus, the microscopic
images of the FG emulsions prepared at pH 6.0 are chosen and
shown in Fig. 5. The emulsions showed signicant structural dif-
ferences as a function of FG concentration. It is observed from Fig. 5
that, after being treated with high pressure homogenization, oil
droplets were wrapped by FG particles, forming a stable or unstable
emulsions. Optical microscopy showed emulsions with 0.1% (w/w)
FG had the largest particle size and the 0.5% (w/w) samples had the
smallest particle size, corroborating with the results obtained from
particle size distribution (Table 1). Flocculation was observed in the
samples at low FG concentration. With the increase of FG concen-
tration, the degree of occulation descended down gradually. In
emulsions with low concentrations of FG, the particles gathered
together and dispersed insufciently, which results in combining of
oil droplets to form large masses. This phenomenon could be
explained the added hydrocolloid has a destabilizing effect on the
emulsion at very low concentrations. Since the depletion occula-
tion induced by the non-adsorbing hydrocolloid causes enhanced
serum separation of the emulsion (Dickinson, 2009). In contrast, in Fig. 4. Creaming index of emulsions stabilized by FG at different pH values.
 M. Wang et al. / Food Hydrocolloids 67 (2017) 54e62 59

Fig. 5. Light micrographs of emulsions containing (A) 0.1%, (B) 0.2%, (C) 0.3%, (D) 0.4% and (E) 0.5% FG at pH 6.0.

also conrmed the existence of interaction between oil droplets Burgar, & Augustin, 2007). The decrease in T2 indicates more re-
and FG and established the foundation for NMR measurements. stricts of movement of the oil droplets, which probably occurred
occulation, irrespective of whether it is depletion or bridging

3.5. LF-NMR and HF-NMR measurements

3.5.1. LF-NMR measurement

LF-NMR is a useful instrument that can be used to analyze the
emulsifying mechanism and characteristics among FG and oil
droplets in FG-stabilized emulsions. The emulsions used for LF-
NMR measurement had certain liquid lard (10%, w/w) and the
FG concentration increased from 0.1% (w/w) to 0.5% (w/w). The
samples prepared with D2O can eliminate strong proton signals
from water, so oil droplets were considered to dominate the LF-
NMR spectrum due to the shorter T2 values of FG in CPMG
sequence when compared with the pulse spacing that can be
ignored.
In addition to the translational diffusion of oil droplets, the
motion of oil molecules within the droplets could also contribute to
the mobility of oil droplets. Thus, the T2 values obtained from LF-
NMR not only provide information about the mobility of oil drop- Fig. 6. Distribution of relaxation time (T2) obtained by CPMG sequence for emulsions
lets, but also the structure of the bulk emulsions (Day, Xu, Hoobin, containing 10% liquid lard with different concentrations of FG.
60 M. Wang et al. / Food Hydrocolloids 67 (2017) 54e62
Fig. 7. The high-eld 1H NMR spectra of the emulsions containing 10% liquid lard with
FG concentration of: (A) 0.2%, (B) 0.3%, (C) 0.4% and (D) 0.5%.
occulation, or aggregation (Day et al., 2007). As shown in Fig. 6,
the T2 distribution curves shifted rightward with the increase of FG
concentration. The shorter T2 of oil molecules in the 0.1% (w/w) and
0.2% (w/w) FG emulsions suggested that more droplets aggregated
to restrict the motion of oil. Emulsions with higher FG (0.3%, and
above) had the similar bi-modal T2-amplitude distribution, lager
peak area was obtained and attributed to the high mobility of oil
droplets. Therefore, compared with dilute solutions, solutions with
0.3% (w/w) or higher FG concentration had better emulsion prop-
erties and stabilities. The oil surface was fully covered by the
existing FG, thus prevented the aggregation of droplets. This is
consistent with our ndings of the microscopic images of
Table 2
Chemical shifts obtained in high-eld 1H NMR for the groups on the oil molecules

ndez et al., 2002; Sacchi et al., 1996; Segre & Mannina, 1997).
(Ferna
Signal Chemical shift (ppm) Functional group
1 0.78 eCH3 (acyl group)
2 1.18 e(CH2)ne (acyl group)
3 1.47 eCH2eCH2eCOOR (acyl groups)
4 1.91 eCH2eCH]CHe (acyl groups)
5 2.13 eCH2eCH]CHe (acyl groups)
6 2.65 ]CHeCH2eCH] (acyl groups)
7 3.97 eCH2OCOR (glyceryl group)
8 4.17 eCH2OCOR (glyceryl group)
9 4.68 Water
10 5.20 eCHOCOR (glyceryl group)
emulsions (Fig. 5). However, the presence of excessive of FG may
result in depletion occulation of oil droplets, which is demon-
strated by Fig. 6 that samples of 0.4% (w/w), 0.5% (w/w) FG were
shifted towards shorter T2 values compared with 0.3% (w/w) FG,
indicating more restricts between oil molecules due to the deple-
tion occulation. Various kinds of hydrocolloids, such as xanthan
gum, gum arabic, modied starch, maltodextrin, pectin, carra-
geenan, have previously been shown to be capable of inducing
depletion occulation when being added with sufciently high
concentrations (McClements, 2004).
3.5.2. HF-NMR measurement
The high-eld 1H NMR can measure, directly and simulta-
neously, different acyl groups in oil and fats and has the unique
ability to give structural information on the emulsions detected
(Guille
n & Ruiz, 2003). It has been shown to be a powerful
research tool to study the dynamics of oil droplets molecule of
the emulsions. The high-eld 1H NMR spectra distribution of
emulsions with FG from 0.2% (w/w) to 0.5% (w/w) are shown in
Fig. 7. Samples had the same proton spectra but with varying
line-width, suggesting the interaction between oil and FG at the
oil-water interface was relatively weak when compared with
interaction between oil molecules. Increasing line-width with
increasing FG concentration was evidence of the increased
interaction between FG and oil molecules. The chemical shifts
obtained in high-eld 1H NMR for the groups on the oil mole-
cules are shown in Table 2. The chemical shifts of the 1H signals
of the acyl groups are well known (Fern
andez, Hilty, Wider, &
Wthrich, 2002; Sacchi et al., 1996; Segre & Mannina, 1997).
Chemical shifts from 1H NMR spectra showed that lard was
mainly composed of saturated fatty acids. When compared with
other chemical shifts of the triacylglycerol molecule, e(CH2)
negroups (d 1.18 ppm) had more freedom of mobility (Fig. 7
and Table 2). It also suggested that e(CH2)negroups was
further away from the interface of FG and oil molecules as it had
more space to stretch. This result is different from that of Day
et al. (2007) who studied the sh oil emulsions stabilized by
sodium caseinate and reported eCH3 had the greater mobility
compared to other triacylglycerol molecules. This could be
related to the fact that lard has more saturated fatty acids while
sh oil is mainly composed of unsaturated fatty acids.
It has been conrmed that high-eld 13C NMR is an effective
technique that is able to provide useful information about the
positional distribution of acyl groups of different oils (Zamora,
Navarro, & Hidalgo, 1994). 13C NMR has its unique advantages
and disadvantages when compared to high-eld 1H spectroscopy.
The major advantage of 13C spectrum is that it can be used
to conrm the acyl positional distribution of oils. Nevertheless,
high-eld 1H NMR spectrum cannot provide us such information.
It is also noting that due to the low sensitivity of 13C NMR
 M. Wang et al. / Food Hydrocolloids 67 (2017) 54e62
Fig. 8. The high-eld 13C NMR spectra of the emulsions containing 10% liquid lard with
FG concentration of: (A) 0.2%, (B) 0.3%, (C) 0.4% and (D) 0.5%.
signals, the strength loss cannot be ignored and therefore it is
time-consuming. Moreover, the spectra only shows signals from
the main component of oils, the glycerol esters (Mannin,
Luchinat, Emanuele, & Segre, 1999). The 13C NMR spectra of
emulsions with FG at different concentrations are shown in
Fig. 8. These signals showed that the liquid lard played a domi-
nant role as it was the most mobile components in the samples.
All spectra of studied samples appear very similar, with differ-
ences being apparent only in the line-width and height of the
peak. The line-width increased with the increase of FG concen-
tration. The results were in accordance with the ndings ob-
tained in the high-eld 1H NMR signals, indicating the increased
interaction between FG and oil molecules at higher concentra-
tions of FG.
61
4. Conclusions
This research found that FG acted as an effective stabilizer for
oil-in-water emulsions. The results showed that FG concentrations
had signicant inuence on the particle size of emulsions, with
smaller droplets being observed with the increase of FG concen-
trations. A reduction of droplet size also observed with the increase
of pH values. Nevertheles, when the FG concentration exceeded
0.3% (w/w), samples at pH 6.0 and 7.0 had no signicant difference.
The apparent viscosity of all samples was independent of pH values
but increased with FG concentration, which was in good agreement
with the results of emulsion stability. G0 and G00 had the same
trends with apparent viscosity and samples with 0.5% (w/w) FG
behaved like a viscoelastic solid. The emulsions with higher than
0.3% (w/w) FG had no creaming behavior at all given pH values,
indicating that 0.3% (w/w) was a critical concentration for FG to
stabilize emulsions. The microscopic images also conrmed that
bridging occulation contributed to the aggregation of droplets at
low FG concentration. NMR spectra provided information about oil
mobility and the interaction between oil and FG at the interface. T2
values, which reect the mobility of oil droplets, became shorter
with the decrease of FG concentration. This is because FG is
insufcient to coat the surface of oil droplets, suggesting increased
restricted mobility of oils. The high-eld 1H and 13C NMR spectra
had increased line-width with the increased of FG concentration,
conrming that the interaction between FG and oil molecules
increased. The chemical shift obtained from high-eld 1H NMR for
the individual groups of oil molecules, indicating that e(CH2)
newas the most mobile and furthest away from the oil/FG inter-
face. These ndings indicate that FG above 0.3% (w/w) can stabilize
oil-in-water emulsions (10% liquid lard) well at pH 5.0 to 7.0. These
results suggest that it is possible to use FG to replace or combine
with protein emulsiers which are sensitive to pH values for the
food industry.
Acknowledgments
This work was co-nanced by the National Natural Science
Foundation of China (No. 31401516) and Fundamental Research
Funds for the Central Universities (Y0201400114). The authors are
deeply grateful to the technical support provided by the National
Center of Meat Quality and Safety Control of Nanjing Agricultural
University.
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