The interaction between pullulan (PU) and tapioca starch (TS) during gelatinization and retrogradation was studied in this paper. TS and PU were prepared into a TS/PU composite system at the ratios of 10/0, 9.5/0.5, 9.0/1.0, 8.5/1.5 and 8.0/2.0 g/g. The addition of PU tended to decrease the peak, breakdown and final viscosity of the composite system. The increasing tanδ of the dynamic viscoelastic measurement suggested that PU enhanced the liquid-like properties of TS gels. The decreased setback values and the slower increase of the storage modulus at 4 °C indicated that the short-term retrogradation of TS was restrained. Meanwhile, the peaks of X-ray diffraction became lower and wider, revealing that PU could inhibit the long-term retrogradation of TS. The FTIR spectra showed that the absorption peak of the O−H stretching gradually redshifted with increasing PU content, suggesting that a strong intermolecular hydrogen binding occurred between TS and PU. From low-field nuclear magnetic resonance, the spin-spin relaxation time decreased from 1687 ms (TS/PU=10.0/0) to 1427 ms (TS/ PU=8.0/2.0), illustrating that the addition of PU promotes the water retention ability of the TS paste. Therefore, PU had an effect on TS gelatinization and retrogradation.

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Tapioca starch-pullulan interaction during gelation and retrogradation
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DOI: 10.1016/j.lwt.2018.05.064
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2. Contents lists available at ScienceDirect
LWT - Food Science and Technology
journal homepage: www.elsevier.com/locate/lwt
Tapioca starch-pullulan interaction during gelation and retrogradation
Long Shenga,b
, Peishan Lia
, Huiqing Wua
, Yuanyuan Liua
, Ke Hana
, Mostafa Goudaa,c
,
Qunyi Tongb
, Meihu Maa,∗
, Yongguo Jina,∗∗
a
National Research and Development Center for Egg Processing, College of Food Science and Technology, Huazhong Agricultural University, Wuhan, Hubei, 430070,
China
b
The State Key Laboratory of Food Science and Technology, School of Food Science and Technology, Jiangnan University, Wuxi, China
c
Department of Human Nutrition & Food Science, National Research Centre, Dokki, Cairo, Egypt
A R T I C L E I N F O
Keywords:
Gelatinization
Intermolecular interaction
Low-field nuclear magnetic resonance
Rapid visco-analysis
A B S T R A C T
The interaction between pullulan (PU) and tapioca starch (TS) during gelatinization and retrogradation was
studied in this paper. TS and PU were prepared into a TS/PU composite system at the ratios of 10/0, 9.5/0.5,
9.0/1.0, 8.5/1.5 and 8.0/2.0 g/g. The addition of PU tended to decrease the peak, breakdown and final viscosity
of the composite system. The increasing tanδ of the dynamic viscoelastic measurement suggested that PU en-
hanced the liquid-like properties of TS gels. The decreased setback values and the slower increase of the storage
modulus at 4 °C indicated that the short-term retrogradation of TS was restrained. Meanwhile, the peaks of X-ray
diffraction became lower and wider, revealing that PU could inhibit the long-term retrogradation of TS. The FTIR
spectra showed that the absorption peak of the O−H stretching gradually redshifted with increasing PU content,
suggesting that a strong intermolecular hydrogen binding occurred between TS and PU. From low-field nuclear
magnetic resonance, the spin-spin relaxation time decreased from 1687 ms (TS/PU = 10.0/0) to 1427 ms (TS/
PU = 8.0/2.0), illustrating that the addition of PU promotes the water retention ability of the TS paste.
Therefore, PU had an effect on TS gelatinization and retrogradation.
1. Introduction
Starch is a versatile and useful polysaccharide that has found nu-
merous applications in many industries due to its environmental
friendliness and availability (Oladebeye, Oshodi, Amoo, & Karim,
2013). Tapioca starch (TS), derived from cassava roots, is an econom-
ical source of energy because of its high carbohydrate content
(Breuninger, Piyachomkwan, & Sriroth, 2009). Cassava is easy to plant
and has low requirements for growth (Chuang, Panyoyai, Shanks, &
Kasapis, 2017). TS has a bland flavor, low price, lower pasting tem-
perature high viscosity, and clear paste appearance compared with
other starches. In addition, compared with other tuber starches such as
potato starch and sweet potato starch, TS has a lower pasting tem-
perature (Zhang, Zhu, Tong, & Ren, 2012). Due to the above ad-
vantages, TS has been widely used in the food industry including in
cakes, bread, noodles, beverages and sauces as a beneficial thickener
(Singh, Gevekea, & Yadav, 2017).
However, native starches suffer limitations under rigorous industrial
processing because of many undesirable properties such as low water-
holding capacity, retrogradation and syneresis (Russ, Zielbauer,
Ghebremedhin, & Vilgis, 2016). The quality of starch granular swelling,
gelatinization and pasting are the crucial actions of the starch as a
factor in food systems for regulating textural and rheological char-
acteristics (Nishinari, Zhang, & Ikeda, 2000). The thermal and pasting
properties of starch granules are affected by several factors including
variety, solvent concentration, amylose content, type of modification
and hydrocolloids (Huang, 2009). To eliminate or reduce such limita-
tions, non-starch polysaccharides have been widely utilized to improve
the quality of foods and extend the shelf life of foods (Korus, Juszczak,
Witczak, & Achremowicz, 2004; Mahmood et al., 2017). The effects of
the addition of various polysaccharides in starch pastes or gels such as
carboxymethyl cellulose, carrageenan, guar gum or xanthan gum have
been studied extensively (Chaisawang & Suphantharika, 2005;
Lascombes et al., 2017; Tischer, Noseda, de Freitas, Sierakowski, &
Duarte, 2006; Zhou, Wang, Zhang, Du, & Zhou, 2008). Many hydro-
colloids have shown the ability to affect the swelling, gelatinization,
retrogradation and freeze-thaw stability of TS (Chen, Fu, & Luo, 2015;
Temsiripong, Pongsawatmanit, Ikeda, & Nishinari, 2005).
https://doi.org/10.1016/j.lwt.2018.05.064
Received 13 December 2017; Received in revised form 25 May 2018; Accepted 27 May 2018
∗
Corresponding author.
∗∗
Corresponding author.
E-mail addresses: mameihuhn@163.com (M. Ma), jinyongguo@mail.hzau.edu.cn (Y. Jin).
Abbreviations: PU, Pullulan; TS, Tapioca starch; RVA, Rapid visco-analysis; G′, Storage modulus; G″, Loss modulu; tanδ, Loss tangent; FT-IR, Fourier transform infrared; XRD, X-ray
diffractometry; LF-NMR, Low-field nuclear magnetic resonance; SEM, Scanning electron microscopy
LWT - Food Science and Technology 96 (2018) 432–438
Available online 28 May 2018
0023-6438/ © 2018 Elsevier Ltd. All rights reserved.
T

3. Pullulan (PU) is widely used in the medical and food industries on
account of many prominent chemical and physical properties, such as
nontoxicity, slow digestibility, high plasticity and excellent film for-
mation (Sheng et al., 2017). In 2002, PU was approved by the FDA and
recorded in GRAS(GRAS Notice No. GRN99). As a food additive, it is
known by the E number E1204. In addition, PU has also been approved
as a food additive in China, Australia and Canada. PU is a linear
homopolysaccharide comprising repeating maltotriose units connected
by (1 → 6)-α linkages (Pan, Yao, Chen, & Wu, 2013). This unique
linkage pattern endows PU with good solubility in water compared with
other polysaccharides. Aqueous solutions of PU are viscous but do not
form gels. It is also stable over a broad range of pH conditions when in
solution (Singh, Saini, & Kennedy, 2008).
Kim, Choi, Kim, and Lim (2014) prepared TS-PU composite films
and found that the use of TS in mixture with PU could effectively im-
prove the mechanical strength and the storage stability under a humid
conditions. However, the interaction between TS and PU during gela-
tinization and retrogradation is still poorly understood. In the present
study, the influences of PU on the pasting, rheological behavior and
retrogradation of TS were investigated to provide new ideas for mod-
ified starch.
2. Materials and methods
2.1. Materials
Pullulan PI20 (molecular weight 200,000 Da) was donated by
Hayashibara Biochemical Laboratories (Okayama, Japan). The
moisture content of the PU was 35 g/kg. Tapioca starch was provided
by Guangxi State Farms Mingyang Biochemical Group, Inc., Nanning,
China. The amylose and moisture content of the TS were 0.171 g/g and
0.134 g/g (dry weight basis), respectively (AACC, 2000).
2.2. Rapid visco-analysis (RVA)
A rapid visco-analyzer (Model RVA-4C, Newport Scientific Pty. Ltd.,
Warriewood, Australia) was used to analyze the viscosity profiles for
the TS/PU composite system. The concentration of TS was fixed at 60 g/
kg, and the TS/PU mixing ratios were 10/0, 9.5/0.5, 9/1, 8.5/1.5 and
8/2 (based on TS weight). For dispersions of the TS and PU mixtures,
powders of TS and PU were placed in a screw-top glass bottle and
dispersed in distilled water by stirring for 2 h under magnetic stirring.
The general pasting method (STD 2) was selected to program the test
profile. The slurries were first held at 50 °C for 1 min, heated to 95 °C
within 7.5 min, held at 95 °C for 5 min, cooled to 50 °C within 7.5 min
and held at 50 °C for 2 min. To assure the uniformity of the diffuseness,
the stirring speed was set at 960 rpm for the first 10 s, and then at
160 rpm for the rest of the measurement. The RVA characteristics, in-
cluding pasting temperature, peak viscosity, breakdown, and setback
were determined from the RVA curves.
2.3. Average particle diameter
A laser diffraction apparatus (Model S3500, Microtrac Inc.,
Montgomeryville, USA) was used to measure the average particle dia-
meter of the starch granules using the same method described before
(Funami et al., 2005). The sample paste prepared by a rapid visco-
analyzer according to the general pasting method (STD 2) was sus-
pended at 10 g/L in an aqueous solutions of 2 g/L sodium hexameta-
phosphate at 20 °C under magnetic stirring.
2.4. Leached amylose
The cooked TS/PU system (25 mL) was centrifuged at 16,000 × g
for 30 min. The supernatant was mixed with 0.33 mol/L NaOH aqueous
solution (6 mL), and subsequently the mixture was heated at 95 °C for
30 min. A concentration of 5 g/L trichloroacetic acid (5 mL) was added
to the mixture (0.1 mL) to adjust the pH of the solution to approxi-
mately 5.5, and then 0.01 mol/L I2–KI aqueous solution (0.05 mL) was
used to dye the mixture. After incubation at 20 °C for 30 min, the re-
sultant blue color was determined spectrophotometrically by reading
the absorption at 620 nm using distilled de-ionized water as a blank
(Chrastil, 1987). The original amylose content was investigated by TS
as a dry ingredient subjected directly to this assay. Potato amylose of
reagent grade (Sigma Chemical Co., St. Louis, MO, USA) was used as a
standard substance for calibration.
2.5. Dynamic rheological measurements
A stress-controlled rheometer (AR 2000, TA Instruments Inc., New
Castle, USA) equipped with parallel plate geometry (serrated plates to
avoid sample slippage, plate diameter 40 mm) was used for the rheo-
logical measurements. The sample paste prepared by a rapid visco-
analyzer according to the general pasting method (STD 2) was im-
mediately placed between the two stainless steel plates (gap width
1000 μm). To prevent moisture vaporization, a thin layer of silicone oil
was used to cover the perimeter of the samples. The samples were
cooled to room temperature and equilibrated at 25 °C for 5 min. For
dynamic viscoelastic determination, an oscillatory frequency sweep was
made at 25 °C over a frequency range of 1–10 Hz at 10 points per
decade. One percent strain amplitude was selected, and it was in the
linear viscoelastic range when observed by the dynamic strain sweep
test. The storage modulus (G′), loss modulus (G″) and loss tangent
(tanδ = G’’/G’) as a function of angular frequency (ω) were recorded as
the mechanical spectra.
For the dynamic time scanning analysis, new pasting samples pre-
pared by a rapid visco-analyzer were cooled to 4 °C. The edge of the
sample was protected from water evaporation by silicon oil during the
measurements. The G’ evolution was recorded as a function of time for
2 h at 4 °C to define the gelling process of the mixture. The strain and
frequency were set at 1.0% and 0.5 Hz, respectively.
TA rheometer Data Analysis software (Version V. 4.20, TA
Instruments Inc.) was used to record the reported results.
2.6. Fourier transform infrared (FT-IR) spectroscopy
The sample paste prepared by a rapid visco-analyzer according to
the general pasting method (STD 2) was immediately freeze-dried
(FreeZone6L, Labconco, Kansas City, USA). The FT-IR spectrum of the
lyophilized samples was recorded in KBr pellets on an infrared spec-
trometer (Perkin-Elmer 16 PC spectrometer, Boston, USA) over a wa-
velength range of 500–4000 cm−1
.
2.7. X-ray diffractometry (XRD)
The sample paste prepared by a rapid visco-analyzer according to
the general pasting method (STD 2) was stored at 4 °C for 20 days and
then freeze-dried. X-ray patterns of the lyophilized samples were de-
termined through a diffractometer (D8 Advance, Bruker AXS,
Karlsruhe, Germany) in the 2θ range at 4–40 °C at a 2° min−1
scan rate,
with the Cu Kα radiation wavelength of 0.1542 nm. The test conditions
were set as 36 kV, 30 mA, one degree scattering slit, one degree di-
vergence slit, and 0.3 mm receiving slit.
2.8. Low-field nuclear magnetic resonance (LF-NMR)
The sample paste prepared by a rapid visco-analyzer according to
the general pasting method (STD 2) was transferred to an NMR glass
tube and stored in 4 °C for 20 days. The LF-NMR experiment was per-
formed using NMI20-015 V-I (NIUMAG Corporation, Shanghai, China)
with a rapid spin-echo pulse sequence echo time (TE) of ¼ 0.8 ms and a
recycle time (TR) of ¼ 400 ms, P1 of ¼ 12 ms and P2 of ¼ 24 ms. A
L. Sheng et al. LWT - Food Science and Technology 96 (2018) 432–438
433

4. good signal-to-noise ratio was achieved with four accumulations. The
pulse repetition time was set to 8 s. All measurements were performed
at room temperature, 25 °C. The peaks were broadened to several
hundred hertz. A single exponential model was used to fit the experi-
mental curves. DLX software program was used for the data analysis
and calculation of the T2 peak area through its respective relative signal
fractions.
2.9. Scanning electron microscopy (SEM)
The TS gels in the presence and absence of PU were cut into seg-
ments approximately 2 mm thick by a thin blade and rapidly freeze-
dried in a freezing dryer (Free-Zone6L, Labconco, Kansas City, USA) for
3 days. The sample was installed on bronze stubs via a double-faced
adhesive tape and then spray coated with a thick layer of gold (10 nm).
The treated sample was observed using SEM (Model S-4800 SEM,
Hitachi Ltd., Tokyo, Japan) at an accelerating beam voltage of 1.0 kV.
2.10. Statistical analysis
Results are presented as mean values, and the reproducibility of the
obtained results was expressed by pooled standard deviation (Pooled
SD) (Box, Hunter, & Hunter, 1978). Duncan's new multiple range test
was used to establish the difference among means at p < 0.05. All
statistical data were analyzed using SPSS software (version 12.0 for
Windows; SPSS Inc., Chicago, IL, USA).
3. Results and discussion
3.1. RVA pasting behaviors
RVA was used to investigate the changes in the pasting character-
istics of the TS/PU composite system (TS/PU = 10.0/0, 9.5/0.5, 9.0/
1.0, 8.5/1.5 and 8.0/2.0) during heating and cooling processes (Fig. 1).
An addition of PU could affect the typical RVA pasting profiles
(Table 1). A strong influence of PU on the pasting behaviors for each
TS/PU mixing ratio was observed.
Under the onset temperature of gelatinization, the TS granules were
non-soluble in water and the viscosity of the aqueous dispersion was
still low. When the starch granules were heated beyond gelatinization,
they absorbed a large amount of water and swelled to many times their
original size, leading to increased viscosity (Newport ScientiWc, 1995).
With increasing PU content, the pasting temperatures of TS slightly
increased. Compared with water alone, the PU addition decreased the
available water in the TS/PU composite system, changed the structure
of water, became more approachable to the hydration layer of the
starch chains and generated an anti-plasticizing effect.
The addition of PU tended to decrease the peak, breakdown and
final viscosity of the composite system. As the concentrations of PU
increased, this effect was more obvious. PU could adsorb onto the
surface of the starch granules and then coat them to prevent friction
between the starch granules and the leaching of some starch compo-
nents from the starch granule (Christianson, 1982). The PU addition
might have resulted in lubricating the TS/PU composite system without
causing adhesiveness because the unique linkage pattern of PU pro-
vided it with inherent low viscosity with Newtonian flow behavior in an
aqueous system (Singh et al., 2008). The setback values decreased in
the presence of PU, indicating that PU inhibited the aggregation of
amylose as the initial gel network structure development (Karim,
Norziah, & Seow, 2000). Therefore, PU could mitigate the short-term
retrogradation of starch.
3.2. Average particle size and leached amylose of the starch granules
The average particle diameter of the TS granules was slightly altered
by the PU addition, while the amount of leached amylose decreased
Fig. 1. Pasting viscosity of 60 g/kg tapioca starch pastes with or without pullulan. 1, tapioca starch/pullulan = 10/0; 2, tapioca starch/pullulan = 9.5/0.5; 3, tapioca
starch/pullulan = 9.0/1.0; 4, tapioca starch/pullulan = 9.5/1.5; 5, tapioca starch/pullulan = 8.0/2.0; and 6, temperature.
Table 1
Pasting properties of 60 g/kg tapioca starch in the presence of various con-
centrations of pullulan.
TS/PU Pasting
temperature
(°C)
Peak
viscosity
(mPa·s)
Breakdown
(mPa·s)
Final
viscosity
(mPa·s)
Setback
(mPa·s)
10/0 71.3e
776a
300a
728a
252a
9.5/0.5 71.7cd
672b
216b
704b
248a
9/1 72.1bc
609c
176c
659c
226b
8.5/1.5 72.5ab
557d
124d
658c
225b
8/2 72.9a
498e
71e
651c
224b
Pooled SD 0.3 15 9 12 4
Means of three replicates. Values followed by different superscript within the
same column are significantly different (P < 0.05). TS: tapioca starch; PU:
pullulan.
L. Sheng et al. LWT - Food Science and Technology 96 (2018) 432–438
434

5. with increasing PU concentrations (Table 2). The process of starch
swelling upon heating in an aqueous circumstance comprises the
leaching of amylose and the melting of amylopectin crystalline struc-
tures, to achieve moisture absorption (Keetels, van Vliet, & Walstra,
1996). PU might inhibit the leaching of starch via the constitution of
the barrier to encircle the surface of starch granules wherefore amylose
diffusion might be controlled by the existence of PU. Nevertheless, PU
could not influence the disordering of amylopectin crystalline struc-
tures; therefore, the swelling of starch still occurred, and the average
particle diameter of the starch granules was not significantly changed.
This phenomenon was consistent with the previous result of a slight
increase in the pasting temperature.
3.3. Dynamic rheological measurements
Fig. 2 shows the dynamic mechanical spectra of TS/PU mixed
pastes. The elastic and the viscous components of the measured samples
was measured by G′ and G″, respectively, during the dynamic viscoe-
lastic test (Fig. 2 a, b). The magnitudes of G″ were much lower than
those of G′ during the measured frequency ranges. With increasing
frequency, the G′ and G″ of all TS/PU mixing ratios increased, in-
dicating that all of the mixtures exhibited the typical behavior of a weak
gel system (Ptaszek et al., 2009). Both G′ and G″ moduli for the dis-
persions of TS alone were higher than those of all TS/PU mixtures at
any frequency, which revealed that the presence of PU interfered the
gelatinization between amylose chains.
When the tanδ values were plotted against the frequency, the tanδ
of the TS gels presented nearly invariable values at approximately
0.6–0.7 at frequencies exceeding 1 rad/s (Fig. 2 c). By contrast, the tanδ
of the TS/PU mixed pastes increased with increasing frequency. In the
low frequency range, the tanδ of the TS/PU mixed pastes was lower
than that of TS alone, demonstrating that PU increased the solid-like
properties of the mixed pastes. The enhancement of the gel network,
attributed to the combination between amylose and PU, might lead to
the increase in solid-like behavior by the PU addition. However, in the
high frequency range, the tanδ of the TS/PU mixtures was higher,
confirming that PU dominated the liquid-like behavior of the mixed
pastes. This phenomenon suggests that the connection between PU and
amylose was weak. In addition, the tanδ of the TS/PU mixed pastes
increased with increasing PU concentration, suggesting the higher li-
quid-like characteristics of PU in the composite systems.
Fig. 2 d shows the changes in G′ as functions of aging time for 2 h at
4 °C which were determined for the TS/PU mixed pastes with varying
PU contents. The pattern of G′ showed a behavior with an initial swift
rise following a slower increase. The G′ values of the TS gel continued
to increase slightly after 1 h without a plateau. In contrast, the G′ values
Table 2
Leached amylose and average particle diameter of the starch granules in the
presence of various concentrations of pullulan.
TS/PU Leached amylose (g/g) Average diameter (μm)
10/0 36.7a
41.4ab
9.5/0.5 35.4a
41.7ab
9/1 33.5b
42.8a
8.5/1.5 32.1bc
40.5b
8/2 30.8c
41.7ab
Pooled SD 1.0 1.0
Means of three replicates. Values followed by different superscript within the
same column are significantly different (P < 0.05). TS: tapioca starch; PU:
pullulan.
Fig. 2. Dynamic mechanical spectra of 60 g/kg tapioca starch pastes in water (control) and in pullulan solutions at various concentrations. (a): G’; (b): G’’; (c): tanδ;
and (d) G′ during aging at 4 °C. ○, tapioca starch/pullulan = 10/0; ■, tapioca starch/pullulan = 9.5/0.5; ▲, tapioca starch/pullulan = 9.0/1.0; ◆, tapioca starch/
pullulan = 9.5/1.5; and ●, tapioca starch/pullulan = 8.0/2.0.
L. Sheng et al. LWT - Food Science and Technology 96 (2018) 432–438
435

6. of the TS/PU mixtures initially increased steadily and then remained
steady afterwards, suggesting that PU inhibited the starch gelation.
Other than the temperature correlation of the general liquid samples,
the aggregation of amylose results in the rapid increase of G′ in the
early stage of retrogradation (several hours); notably, that of amylose
that retrogrades much faster than amylopectin at the early stage (Miles,
Morris, Orford, & Ring, 1985). Nagano, Tamaki, and Funami (2008)
proposed that the lowered amount of leached amylose from the swel-
ling starch granules during pasting decelerated the increase in G’. The
slower increase of the mixture G’ during the first several hours may be
attributed to the incorporation of PU, which reduced the free volume of
water in the starch-water systems and became an anti-plasticizing
agent, facilitating the restriction in mobility of amylose thereafter.
3.4. FTIR analysis
The FT-IR spectra of the TS/PU mixed pastes are compared in Fig. 3.
The spectra for individual TS and other samples exhibited similar fea-
tures. The absorption in 933 cm−1
indicated the presence of α-1,6-D-
glucosidic bonds. A peak at approximately 853 cm−1
was characteristic
of the α-configuration. A strong absorption at approximately 760 cm−1
proved the presence of α-1,4-D-glucosidic bonds. These were because
both TS and Pu belonged to a homopolysaccharide comprising re-
duplicative glucose units connected by α-1,4-linkages and α-1,6-lin-
kages. A strong peak appearing at 1639 cm−1
suggests O−C−O bond
and glycosidic linkage. A sharp band appearing at 1155 cm−1
was due
to C−O stretching from C−O−H. The signals arriving at 1016 cm−1
were attributed to C−O−C stretching. A broad band at approximately
3400 cm−1
was due to O−H stretching (Sheng et al., 2016). Notably,
Fig. 3. FT-IR spectra of the lyophilized powder of tapioca starch pastes with or without pullulan. 1, tapioca starch/pullulan = 10/0; 2, tapioca starch/pull-
ulan = 9.5/0.5; 3, tapioca starch/pullulan = 9.0/1.0; 4, tapioca starch/pullulan = 9.5/1.5; and 5, tapioca starch/pullulan = 8.0/2.0.
Fig. 4. X-ray diffraction patterns of native tapioca starch
powder and lyophilized powder of tapioca starch pastes with
or without pullulan. 1, native tapioca starch; 2, tapioca
starch/pullulan = 10/0; 3, tapioca starch/pullulan = 9.5/
0.5; 4, tapioca starch/pullulan = 9.0/1.0; 5, tapioca starch/
pullulan = 9.5/1.5; and 6, tapioca starch/pullulan = 8.0/
2.0.
L. Sheng et al. LWT - Food Science and Technology 96 (2018) 432–438
436

7. the absorption peak of O−H stretching gradually redshifted with in-
creasing PU content. The O−H peaks have been found to shift slightly
to a lower wavenumber, and the hydrogen-bond interaction of wheat
starch with microcrystalline cellulose forms when the chain segment on
the surface of swollen microcrystalline cellulose granules interacts with
the amylose of wheat starch to decrease the helical formation prob-
ability of wheat starch (Xiong, Li, Shi, & Ye, 2017). This phenomenon
indicates that intermolecular hydrogen binding occurred between
amylose and PU. Therefore, PU could impede the formation of hy-
drogen bonds among amylose and inhibit the short-retrogradation of
TS.
3.5. X-ray diffraction analysis
The diffractograms of the native TS and TS/PU composite system
after 20 days are shown in Fig. 4. The sharp diffraction peaks at
2θ = 15.1°, 17.0°, 17.9° and 22.9° and the weaker absorption peaks at
2θ = 11.3° and 19.9° indicate that native TS exhibits typical A-type
crystallization (Xia et al., 2015). After gelatinization and retrogradation
for 20 days, a diffuse and wide peak at 2θ = 21° appeared on the dif-
fraction curve, suggesting that the original crystal structure of native TS
was broken and the crystal type of TS changed from A-type crystal-
lization to B-type crystallization. Niu, Wu, and Xiao (2017) also found
that the retrogradation of gelatinized rice starch was evident from the
appearance of a B-type pattern at 2θ angles of 16.9° and 20.8° after
storage for 14 d. Interestingly, the diffraction peaks at 2θ = 21° gra-
dually became lower and wider compared with the pure starch paste
group. The result demonstrates that PU possessed competence to inhibit
the long-term retrogradation of TS. Combined with the FTIR results, the
hydrogen bonds between PU and TS can restrain the interaction among
amylopectin and prevent the orderly accumulation of short chains on
the outside of the amylopectin. Therefore, the action of starch re-
crystallization was weakened.
3.6. LF-NMR analysis
LF-NMR is a sensitive method for evaluating the water-holding ca-
pacity and detecting water changes in varied types of food materials
(Marcone et al., 2013). The changed spin–spin relaxation time (T2)
could represent the different molecular mobility. Table 3 shows the T2
of the TS pastes (60 g/kg) without or with PU. A single exponent model
was used to calculate the T2 value on behalf of the overall average
mobility of water. Clearly, T2 declined steadily with increased PU
concentration. T2 decresed from 1687 ms (TS/PU = 10.0/0) to 1427 ms
(TS/PU = 8.0/2.0), suggesting that the free motion of water molecules
in the TS pastes with PU were more constrained. This phenomenon was
consistent with the FT-IR results. The hydrogen bonds between PU and
TS, especially the leached amylose, weakened the interaction among
the amylopectin and transformed the less amorphous area into a re-
crystallization area. Therefore, the loss of bound water molecules was
controlled, and the retrogradation of TS was restrained. In addition, as a
kind of hydrophilic colloid, PU possessed abundant hydroxyl and ex-
cellent solubility, and it could hold a significant amount of water. In
Table 3
The relaxation time (T2) of tapioca starch pastes in the
presence of various concentrations of pullulan.
TS/PU Relaxation time (T2)
10/0 1687a
9.5/0.5 1623ab
9/1 1476bc
8.5/1.5 1465bc
8/2 1427c
Pooled SD 78
Means of three replicates. Values followed by different
superscript within the same column are significantly dif-
ferent (P < 0.05). TS: tapioca starch; PU: pullulan.
Fig. 5. SEM images of tapioca starch gels without and with pullulan. (a), (c): tapioca starch/pullulan = 10/0; (b), (d): tapioca starch/pullulan = 8/2.
L. Sheng et al. LWT - Food Science and Technology 96 (2018) 432–438
437

8. conclusion, the addition of PU would promote the water retention
ability of TS paste. Similar results for compounds such as xanthan (Lee,
Baek, Cha, Park, & Lim, 2002) and konjac glucomannan (Charoenrein,
Tatirat, Rengsutthi, & Thongngam, 2011) have also proved effective in
enhancing the water retention or water holding capacity of starch paste.
3.7. Morphology analysis
SEM was used to investigate the influence of PU addition on the
microstructures of TS gels. TS/PU gels (TS/PU = 10/0 and 8/2) were
selected as typical samples to investigate the relationship between PU
addition and gel textures (Fig. 5). A clear difference was observed in the
surface for the control and TS gels with PU. As could be seen in low
magnification, the size of the honeycomb pores in the TS/PU gels was
smaller and more homogeneous than that of individual TS gels. When
magnified at 10,000 times, it was clear that the micrographs of the TS
gels exhibited a rough surface and larger hole. Nevertheless, the TS gels
with PU were comparatively homogeneous and showed more compact
pores. As a whole, the TS/PU gel presented a compact matrix and good
structural integrity. Therefore, PU can promote water retention through
the interaction with amylose and amylopectin. This result was in good
agreement with the LF-NMR results.
4. Conclusions
The addition of PU apparently changed the gelatinization and ret-
rogradation characteristics of TS. PU decreased the viscosity of the
paste, and the mixtures showed greater liquid-like characteristics.
Meanwhile, PU could inhibit short- and long-term retrogradation. In
addition, the water retention ability of the TS paste was enhanced by
the addition of PU. PU could adsorb onto the surface of the starch
granules and then coat them to prevent friction between the starch
granules and the leaching of some starch components from the starch
granule during gelatinization. The intermolecular hydrogen bonds oc-
curred between TS and PU could impede the formation of hydrogen
bonds among amylose and amylopectin and inhibit the retrogradation
of TS to a certain extent. The results provide more information to im-
prove the quality of TS for application in the food industry.
Acknowledgements
This research was supported by the National Natural Science
Foundation of China (No. 31701622), Hubei Provincial Natural Science
Foundation of China (No. 2018CFB606), the Fundamental Research
Funds for the Central Universities (Program No. 2662018JC022) and
Modern Agro-Industry Technology Research System (Project No. CARS-
41-K23).
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