Research Article

Received: 1 February 2020 Revised: 29 April 2020 Accepted article published: 7 May 2020 Published online in Wiley Online Library: 16 May 2020

(wileyonlinelibrary.com) DOI 10.1002/ps.5885

Encapsulation of thiabendazole
in hydroxypropyl-⊎-cyclodextrin
nanofibers via polymer-free
electrospinning and its characterization
Shuang Gao, Yanyan Liu, Jingyu Jiang, Xiaoming Li, Lixia Zhao, Ying Fu*
and Fei Ye*

Abstract
BACKGROUND: Thiabendazole (TBZ) is a poorly water-soluble benzimidazole fungicide. However, the water solubility of TBZ
can be significantly enhanced by inclusion complexation with cyclodextrins. In this study, a thiabendazole/hydroxypropyl-
⊎-cyclodextrin (TBZ/HP⊎CD) complex was synthesized and electrospinning was performed to produce a TBZ/HP⊎CD nanofi-
brous (TBZ/HP⊎CD-NF) complex that improved water solubility and antifungal activity.
RESULTS: The formation of TBZ/HP⊎CD-NF was characterized by Fourier transform infrared spectroscopy, X-ray diffraction and
nuclear magnetic resonance. The morphology of TBZ/HP⊎CD-NF was studied by scanning electron microscopy. A phase solubil-
ity experiment showed that HP⊎CD exerted a great solubilization effect on TBZ, and TBZ/HP⊎CD-NF had better antifungal activ-
ity compared to that of TBZ alone.
CONCLUSIONS: In summary, the solid fungicidal nanodispersion prepared in the present study is a new type of formulation that
can enhance the water solubility of TBZ. This formulation, which demonstrated potential as a new fast dissolving formulation
type with increased efficacy, is expected to be conducive to the sustainable development of agriculture.
© 2020 Society of Chemical Industry

Keywords: thiabendazole; hydroxypropyl-⊎-cyclodextrin; inclusion complex; electrospinning; nanofibrous

1 INTRODUCTION a host molecule such as cyclodextrin. In the cyclodextrin family,

A correlation has been observed between the use of pesticides
and their effect on humans and the environment.1 In recent years,
the annual output of fungicides has increased. Benzimidazole fun-
gicides are one of the most widely used agricultural fungicides.2
Thiabendazole [2-(4-thiazolyl) benzimidazole] (TBZ) is a low-
toxicity broad-spectrum water-insoluble benzimidazole fungi-
cide. Its mechanism of action is to inhibit the formation of tubulin
during the mitosis of plant pathogenic fungi.3 TBZ can be used to
prevent and treat diseases of fruits and vegetables during storage,
such as citrus green mold, blue mold and banana crown rot. It
possesses anti-corrosion properties and maintains freshness, as
well as preventing and treating mushroom brown rot, apple tree
ring rot and grape black pox. It is the most important fungicide
for the production, storage and freshness of various fruits. The
TBZ is commonly applied either as a suspension concentrate or
wettable powder, however, the latter is a risk to the environment
due to its high dust potential. Therefore, the development of an
innovative formulation concept that increases the biological effi-
cacy of TBZ is of great importance.
Nanosolids are regarded as new pesticide formulations that can
effectively improve the water solubility of pesticides.4,5 The nano-
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there are three main members, containing different amounts of
glucopyranose units (six, seven and eight units),6 which are
named ⊍-cyclodextrin, ⊎-cyclodextrin and γ-cyclodextrin,7 respec-
tively. Cyclodextrin has a conical, hollow, cylindrical three-
dimensional structure. The ‘hydrophobic inside and hydrophilic
outside’ structure can absorb hydrophobic small molecular sub-
stances or groups of a certain size and shape to produce a stable
inclusion complex.8 By forming inclusion complexes with hydro-
phobic molecules, cyclodextrin can change the physical and
chemical properties of the incorporated molecules.
Hydroxypropyl-⊎-cyclodextrin (HP⊎CD) (Fig. 1) is an etherified
derivative of ⊎-cyclodextrin, and its water solubility has been
greatly improved.9,10 It can form inclusion complexes with various
hydrophobic substances, thus improving the water solubility, sta-
bility, oxidation resistance and photolysis resistance of the guest
molecules.11,12 Furthermore, such complexes can show slow-
* Correspondence to: F Ye or Y Fu, Department of Applied Chemistry, Northeast
Agricultural University, Harbin 150030, China. E-mail: yefei@neau.edu.cn (Ye);
E-mail: fuying@neau.edu.cn (Fu)

Department of Applied Chemistry, Northeast Agricultural University, Harbin,

solid dispersions can be made by complexation of the active with China

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Encapsulation of thiabendazole in hydroxypropyl-⊎-cyclodextrin nanofibers
Figure 1 The molecular structure of HP⊎CD.
release properties and induce three-dimensional separation
effects.13,14
For a long time, electrospinning nanofiber technology has
played a critical role in the fields of agriculture and food due to
its ability to achieve encapsulation of active substances such as
food additives and drugs.15,16 Various attempts have been made
to improve drug properties through electrospinning technol-
ogy.17,18 Most of these attempts were based on the characteristics
of cyclodextrin and its derivatives. Asli Celebioglu et al.19 reported
a nanofiber web inclusion complex, which was prepared by elec-
trospinning with a modified cyclodextrin and essential oil com-
plex (thymol). The volatility of thymol was inhibited via the
formation of the inclusion complex, while most thymol
(88–100%, w/w) was stored in the nanofibers of the thymol/cyclo-
dextrin inclusion complex, which exhibited improved water solu-
bility and high thermal stability as well as antioxidant activity.
Zeynep Aytac and coworkers20 successfully realized the prepara-
tion of a polymer-free nanofiber electrospinning inclusion com-
plex with modified cyclodextrin and camphor. Experiments
proved that after spinning, camphor is slowly released, and its
water solubility is enhanced. Eleni Kavetsou et al.21 proposed a
green method of wrapping mint essential oil in baker's yeast
and evaluated its potential as an insecticide to control the insect
pest Myzus persicae, representing the successful development of
an environment-friendly pesticide. Previous experiments have
proven that ⊎-cyclodextrin and HP⊎CD can expand the value of
chlorothalonil in agriculture and commerce as good carriers to
enhance the water solubility and fungicidal activity of
chlorothalonil.22
Different varieties of bioactive compounds (e.g. essential oils,
vitamins, flavors, food supplements, etc.) have been encapsulated
within nanofiber by electrospinning for food related applications.
However, so far, there have been few studies on the application of
electrospinning to pesticide formulations, in fact there has been
no study reported related to encapsulation of TBZ in electrospun
nanofiber. In order to broaden the application range of electro-
spinning technology and improve the water solubility of TBZ,
we have carried out innovative research on the formulation of
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water-insoluble TBZ and found a water-soluble pesticide formula-
tion with nanometer level. In this study, the raw materials for elec-
trostatic spinning were generated by mixing HP⊎CD and TBZ in an
equimolar ratio, and then nanofibrous webs of the thiabendazole/
hydroxypropyl-⊎-cyclodextrin nanofibrous (TBZ/HP⊎CD-NF) were
obtained via electrospinning. A phase solubility test was carried
out to determine the change in solubility of TBZ after adding dif-
ferent concentrations of HP⊎CD. The morphology of TBZ/HP⊎CD-
NF was observed by scanning electron microscopy (SEM), and the
average diameter of the nanofibers was calculated. The chemical,
structural and fluorescence characteristics of TBZ and
TBZ/HP⊎CD-NF were characterized by proton nuclear magnetic
resonance (1H-NMR), Fourier transform infrared (FTIR) spectros-
copy, X-ray diffraction (XRD) and fluorescence spectroscopy. Anti-
fungal experiments showed that the fungicidal activity was also
enhanced due to the improved water solubility of TBZ. In a word,
the formation of TBZ/HP⊎CD-NF improved the water solubility of
TBZ, which in turn increased the fungicidal efficacy. This research
represents an effective attempt at the development of environ-
mentally friendly, efficient and water-based modern pesticide
formulations.
2 MATERIALS AND METHODS
2.1 Materials
TBZ was purchased from Dalian Meilun (≥98.0%, Dalian, China).
HP⊎CD was provided by Aladdin Reagent Co., Ltd (C51H88O38,
degree of substitution: ~0.6, ≥ 98.0%, Shanghai, China). Deuter-
ated dimethylsulfoxide (DMSO-d6, deuteration degree minimum
99.8% for NMR spectroscopy, Aladdin Reagent Co., Ltd). Potas-
sium bromide (KBr, 99%, FTIR grade, Aladdin Reagent Co., Ltd).
Gibberella sp (Strain number: CICC 2498, Beijing, China). All other
reagents were purchased from Aladdin Reagent Co., Ltd.
2.2 Preparation of a physical mixture
Equimolar HP⊎CD and TBZ were mixed in a mortar and ground for
20 min. After grinding evenly, vacuum drying was carried out at
room temperature to obtain a physical mixture.
2.3 Preparation of solutions for electrospinning
TBZ/HP⊎CD solution (1 mL) was prepared by mixing HP⊎CD and
TBZ in an aqueous solution at a molar ratio of 1:1. TBZ powder
(0.133 g) was dispersed in water, and then HP⊎CD (1.6 g) was
added into the dispersion. The solution was obtained after being
stirred at room temperature for 12 h. Electrospinning was per-
formed to prepare TBZ/HP⊎CD nanofibers in the form of self-
supporting nanofiber webs.
2.4 Electrospinning of nanofibers
The distance between the syringe pump and the receiving plate
was adjusted to 12–16 cm, and a 1 mL plastic syringe (the inner
diameter of the metal needle of the syringe was 0.1 mm) was
filled with TBZ/HP⊎CD solution and installed on the syringe pump.
An electric field (15–20 kV) was applied and the solution was
sprayed on the aluminum-covered grounded metal current col-
lector foil at a speed of 0.5 mL/h. After electrospinning, the
TBZ/HP⊎CD nanofiber web was placed in a refrigerator at 4 °C.
2.5 Measurements and characterization
2.5.1 Phase solubility studies
The experiments were performed as described by Higuchi and
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Connors theory:23 25 mL of HP⊎CD solution at different
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concentrations (ranging from 0 to 10 mmol/L) was mixed with
excess TBZ (2.00 g) in a flask. The mixture of HP⊎CD and TBZ
was shaken for 3 days at 33 °C to dissolve the HP⊎CD and TBZ uni-
formly. Subsequently, the solution was filtered using a 0.45 μm fil-
ter membrane. The content of TBZ dissolved in each filtrate was
determined spectrophotometrically at 361 nm using a UV-2550
spectrophotometer (Shimadzu, Suzhou, China). The solubility of
TBZ in different concentrations of HP⊎CD was calculated.
The correlative constant value (Kf) was obtained by the follow-
ing formula:24
Slope
Kf = ð1Þ
SO ð1−SlopeÞ
where ‘Slope’ is the slope of phase solubility curve and S0 is the
solubility of TBZ in water.
Complexation efficiency (CE) was calculated via the following
formula:
CðTBZ=HP⊎CD slope
CE = S0 × Kf = = ð2Þ
C ðHP⊎CDÞ 1−slope
where C(TBZ/HP⊎CD) represents the concentration of TBZ/HP⊎CD
and C(HP⊎CD) expresses the uncombined HP⊎CD concentration.
Studies of phase solubility offer guiding significance for analyzing
the influence of inclusion formation on TBZ water solubility.25
2.5.2 Fourier transform infrared spectroscopy (FTIR)
Infrared spectra were measured with a Shimadzu 8400S FTIR spec-
trometer (Shimadzu, Kyoto, Japan). FTIR spectra of TBZ, HP⊎CD,
physical mixture and TBZ/HP⊎CD-NF were measured after they
were mixed with KBr respectively. FTIR spectra were recorded in
the range 4000–400 cm −1, with a resolution of 4 cm −1 and
16 scans.26,27
2.5.3 X-ray diffraction (XRD)
An XRD spectral diagram was measured with a scanning angle
range of 5° to 90° and scanning rate of 2°/min by a Phillips X-
ray diffractometer (Eindhoven, The Netherlands) using a 30 mA
current, Cu-K⊍ (⊗ = 1.5406 Å) radiation and 40 kV voltage.28,29
2.5.4 Scanning electron microscopy (SEM)
After the TBZ/HP⊎CD-NF was fixed on the sample rack, a gold
layer was sprayed at a voltage of 12.5 kV under high vacuum con-
ditions. The morphological appearance of the sample was
recorded by a Su-8010 SEM environmental system (Hitachi, Tokyo,
Japan).
2.5.5 Nuclear magnetic resonance (NMR) spectroscopy
The scanning range of 1H-NMR was set as −1 ppm to approxima-
tely 14 ppm in this experiment. The 1H-NMR spectra of HP⊎CD
and TBZ/HP⊎CD-NF were generated using an AVANVE 300 MHz
(Bruker, Beijing, China) with tetramethylsilane (TMS) acting as an
internal standard and DMSO-d6 as a solvent. In addition, NMR
spectra were analyzed and generated by MestReNova
software.30,31
2.6 Fluorescent spectrometry
Fluorescence emission spectra were recorded on a PerkinElmer
LS55 fluorescence spectrometer (PerkinElmer, Beaconsfield, UK).
In three identical 10 mL colorimetric tubes, equal concentrations
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of TBZ and TBZ/HP⊎CD-NF solution (1.0 × 10−6 mol/L) were
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S Gao et al.
accurately prepared. At room temperature, the fluorescence spec-
trum was measured with a 1 cm quartz cell at ⊗ex/
⊗em = 361 nm/469 nm, and the entrance and emission slits were
both 5 nm.32
2.7 Molecular modeling method
The high-performance three-dimensional molecular structures of
TBZ and HP⊎CD were generated by the sketch module (Tripos, St
Louis, MO, USA) of the SYBYL-X 2.0 software package. Subse-
quently, after reasonable optimization of molecules, the
Gasteiger–Huckel charges were calculated accurately. A semiflex-
ible docking method was performed on receptor and ligand mol-
ecules with Accelrys Discovery Studio 2.5 (Accelrys Inc., San Diego,
CA, USA).33 According to the principles of CDOCKER-ENERGY, the
most stable structure of receptor and ligand was determined.
Before docking, the water in receptor HP⊎CD was removed, and
a CHARMM force field was applied to the receptor structure.34
After HP⊎CD was prepared, the active site for the docking study
was determined, and the subset region was 13.0 A from the
known ligand center. The maximum hit was set to 100 while other
parameters were set as default. The obtained HP⊎CD was used as
the ‘Input Receptor’.35 TBZ was chosen as the ‘Input Ligands’. The
successful binding of the small molecule ligand-cyclodextrin
receptor complex was regarded as an index for molecular docking
evaluation, which represented the maximum negative expression
of the conformation with the highest stability.36
2.8 Bioassay
The antifungal activities of original TBZ and TBZ/HP⊎CD-NF
against Gibberella sp. was studied, referring to the method of Yuan
et al.37 Sample solutions containing different concentrations
(1 mL) of TBZ and TBZ/HP⊎CD-NF were prepared with sterile
water. The final TBZ contents in the original drug and
TBZ/HP⊎CD-NF solutions were 0.025, 0.05, 0.10, 0.20 and
0.50 μg/mL. Next, 10 mL of potato glucose agar medium were
rapidly stirred with the prepared solution at 55 °C and then
placed in a culture dish. After the culture medium was solidified
and completely cooled, a 6 mm hypha plate was incubated with
the solidified plate culture medium. It was incubated and inverted
for 48 h at 28 °C. The experiment was repeated three times in
each group. After 48 h, the mycelial elongation diameter was
recorded. Sample-free mixed medium was used as a blank con-
trol. The inhibition rate was obtained using Eqn (3):
C−T
Ið%Þ = ×100 ð3Þ
C
where I represents the inhibition rate (%), T and C show the aver-
age diameter (in millimeters) of the experimental group and con-
trol group, respectively. The EC50 (concentration for 50%
inhibition) values of the original TBZ and TBZ/HP⊎CD-NF were
determined according to the connection between concentration
and inhibition rate.
2.9 Statistical analyses
The results of the experiments performed at least in three repli-
cates, were expressed as mean values and standard deviations.
Statistical analyses were carried out by one-way or two-way anal-
ysis of variance (ANOVA), whichever was applicable. The ANOVA
analyses were carried out using Origin Lab at a 0.05 level of
probability.
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3 RESULTS AND DISCUSSION

3.1 Phase solubility studies
The solubility of TBZ was calculated by plotting the change with
HP⊎CD concentration as shown in Fig. 2. The R2 value of the
extrapolated curve was 0.98763, indicating that a significant cor-
relation between TBZ solubility and HP⊎CD concentration could
be confirmed. Moreover, TBZ solubility was positively correlated
with the increase in HP⊎CD concentration. According to Eqns
(1) and (2) and the slope of the phase solubility curve, Kf and CE
of TBZ/HP⊎CD were calculated. The Kf value of TBZ/HP⊎CD
(997 L/mol) indicated that HP⊎CD and TBZ formed an inclusion
complex with favorable stability. The CE obtained in the present
study was 0.57, which indicated that HP⊎CD could promote the
dissolution of TBZ in water. According to Higuchi and Connors'
theory,23 the A-type phase solubility diagram has subtypes of
AL-type, AN-type, and AP-type which stands for linear increases
in guest solubility as a function of cyclodextrin concentration,
Figure 3 Results of FT-IR: A, TBZ; B, physical mixture of HP⊎CD and TBZ; C,
positive deviation of isotherms, and negative deviation of iso-
TBZ/HP⊎CD-NF; D, HP⊎CD.
therms, respectively.38 In our case, the phase solubility diagram
exhibits the AL-type pattern, suggesting the stoichiometric ratio
of the formed complex is 1:1 (HP⊎CD: TBZ). The earlier-mentioned process of TBZ and HP⊎CD. It is confirmed that the binding of
results show that complexation of TBZ with HP⊎CD can signifi- TBZ with HP⊎CD was through intermolecular forces, such as
cantly improve the water solubility of TBZ. hydrogen bonds, van der waals forces, etc.39

3.3 Analysis of XRD

3.2 FTIR analysis
As shown in Fig. 4, TBZ is a crystalline drug, which has character-
The infrared absorption spectrum of TBZ in Fig. 3 was analyzed.
istic diffractions at 13.1°, 17.7°, 21.2°, and 26.5° (2⊔). The diffrac-
The extension of the N─H contraction vibration absorption peak
tion pattern of the physical mixture is primarily a simple
was at 3457 cm−1. The contraction vibration absorption peak of
coincidence of the diffraction patterns of HP⊎CD and TBZ, indicat-
C─H of the benzene ring was at 3065 cm−1. The contraction vibra-
ing that a simple physical mixture of HP⊎CD and TBZ cannot form
tion absorption peak of C═C was at 1667 cm−1, and the skeleton
an inclusion complex. HP⊎CD is an amorphous cyclodextrin type,
vibration absorption peaks of the benzene ring were at 1643,
having a broad XRD pattern without any sharp peaks.40 The XRD
1538, 1466 and 1486 cm−1. Figure 3 shows that the spectrogram
pattern of TBZ/HP⊎CD-NF is very similar to that of pristine HP⊎CD,
of the physical mixture of TBZ and HP⊎CD simply superimposes
but the diffraction peaks at 21.2° and 26.5° were observed in a
the characteristic peaks of the two components. The infrared
very low intensity, suggesting that a low amount of TBZ crystals
spectrum of TBZ/HP⊎CD-NF was almost similar to that of HP⊎CD,
were present in this sample (Fig. 4(C)). The XRD pattern of TBZ
However, the characteristic absorption peak of TBZ was not
has sharp crystalline peaks, while the XRD pattern of HP⊎CD is
observed. The interaction of TBZ molecules with HP⊎CD cavities
amorphous. In the case of TBZ/HP⊎CD-NF sample, most of the
could lead to shifts, disappearance or attenuation in the charac-
TBZ/HP⊎CD-NF was in the amorphous state, indicating that TBZ
teristic peaks of the guest molecules. Therefore, TBZ entered the
molecules were separated from each other after they were encap-
hydrophobic cavity of HP⊎CD, with the result that the infrared
sulated in the cavity of HP⊎CD. Although there are very few
characteristic peaks of TBZ were masked or weakened by HP⊎CD.
uncomplexed TBZ crystals in TBZ/HP⊎CD-NF sample, the forma-
No new covalent bonds were formed during the complexing
tion of an inclusion complex can be well proved. In addition, it is
worth mentioning that amorphization of drug molecules is
desired for their fast-dissolution. Here, the amorphous nature of
the inclusion complex of TBZ encapsulated in HP⊎CD fiber matri-
ces along with highly porous and very large surface area of
TBZ/HP⊎CD-NF webs greatly help the fast-dissolving behavior of
these samples as discussed later.

3.4 Analysis of 1H-NMR spectroscopy

The 1H-NMR was applied to characterize the inclusion complex by
comparison of the NMR spectra of TBZ, TBZ/HP⊎CD-NF and
HP⊎CD (Fig. 5). Tables 1 and 2 summarize the chemical shifts of
TBZ and HP⊎CD in the 1H-NMR spectra after the formation of
TBZ/HP⊎CD-NF. The chemical shifts of H-3 and H-5 for HP⊎CD
were significantly changed after the formation of TBZ/HP⊎CD-
NF. Chemical shifts all migrated to high fields. This phenomenon
could be due to the presence of H-3 and H-5 in the inside cavity
of HP⊎CD molecules. When TBZ/HP⊎CD-NF was generated, the
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Figure 2 Phase solubility curve of TBZ in the presence of HP⊎CD. chemical shift values of H-3 and H-5 varied significantly, while

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Figure 4 Results of XRD: A, TBZ; B, physical mixture of HP⊎CD and TBZ; C,

TBZ/HP⊎CD-NF; D, HP⊎CD.

the values of H-2, H-4 and H-6 varied slightly. Additionally, the
chemical displacement of TBZ moved to a high field to different
degrees. When the small guest molecule was combined with
the large host molecule, the small molecule entered the cavity
of the large molecule, and thus the original water molecule in
the HP⊎CD cavity was forced out. At this time, the magnetic field
was shielded by an electron cloud with high density in the cavity,
so the actual magnetic field felt by the nucleus was lower than the
original magnetic field, which indicated that the NMR detection
moved to the high magnetic field. Protons in the HP⊎CD cavity
were transferred to the high field to different degrees, and the
peak shape was changed, indicating that an inclusion complex
of TBZ and HP⊎CD was formed.
The complexation stoichiometric ratio could be obtained by
comparing the integral area under the H1 proton for the HP⊎CD
with that of the H1 protons from the TBZ in the inclusion complex
spectrum (Fig. 5(B)). Peaks corresponding to protons of TBZ and Figure 5 1H NMR spectra: (A) TBZ; (B) TBZ/HP⊎CD-NF; (C) HP⊎CD.
HP⊎CD (Fig. 5(A, C)) were initially assigned. The intensity of the
HP⊎CD peak was arbitrarily assigned to 1, and the program auto-
matically calculated the strength of the TBZ proton. Therefore, the enhancing fluorescence intensity. Therefore, the enhancement
inclusion complex stoichiometry was obtained as follows: of TBZ fluorescence intensity can effectively prove the formation
of an inclusion complex.
HP⊎CD intensity =1:00 ð11 protonsÞ
3.6 Results of molecular modeling
TBZ intensity=0:09 ðone protonÞ Molecular models refer to the use of theoretical methods and
Ratio intensities = ð0:09=1Þ=ð1=11Þ=0:99 computational techniques to simulate the appearance or proper-
ties of chemical molecules and predict the binding mode of
Therefore, the stoichiometric ratio obtained from the 1 H-NMR receptors and ligands. Figure 7 shows the minimum energy
data was approximately 1, which could be mutually corroborated model generated through molecular modeling. The results
with the TBZ of the inclusion ratio. proved that the ligand TBZ and the receptor HP⊎CD were effec-
tively combined. In Fig. 7, TBZ moved into the active site of HP⊎CD
3.5 Study on fluorescence spectra as shown in the cross-section. Additionally, ligands and receptors
TBZ is a fungicide with fluorescent properties. According to the formed multiple hydrogen bonds to maintain a stable supramo-
fluorescence spectrum of TBZ and TBZ/HP⊎CD-NF (Fig. 6), strong lecular conformation.41
fluorescence excitation and emission peaks appear at ⊗ex/
⊗em = 361 nm/409 nm. The shape of TBZ/HP⊎CD-NF fluorescence 3.7 Fungicidal activity
spectrum was similar to TBZ, but the fluorescence intensity was After forming TBZ/HP⊎CD-NF, the mycelial elongation diameter of
higher than TBZ. The reason is that when TBZ and HP⊎CD form fungal settlements for TBZ was obviously inhibited (Fig. 8), reflect-
TBZ/HP⊎CD-NF by complex action, the hydrophobic cavity of ing that the formation of TBZ/HP⊎CD-NF significantly improved
HP⊎CD protected TBZ molecule, reducing fluorescence quench- antifungal activity.
ing of TBZ, freedom of movement and deactivation collision Table 3 demonstrates that the EC50 values of TBZ and
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between molecules, further improving quantum yield and TBZ/HP⊎CD-NF against Gibberella sp. were 0.406 and 0.222 μg/

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Table 1 Chemical shift of HP⊎CD after forming TBZ/HP⊎CD-NF

Chemical shift (⊐)

Substances H-1 H-2 H-3 H-4 H-6 H-5

HP⊎CD 4.838 3.301 3.763 3.220 3.607 3.578

TBZ/HP⊎CD-NF 4.829 3.303 3.737 3.219 3.608 3.548
4⊐ −0.009 0.002 −0.026 −0.001 0.001 −0.030

Table 2 Chemical shift of TBZ after forming TBZ/HP⊎CD-NF

Chemical shift (⊐)

Substances H-a H-b H-c H-d H-e H-f H-g

TBZ 12.967 9.343 8.448 7.671 7.652 7.231 7.199

TBZ/HP⊎CD-NF 12.946 9.328 8.444 7.642 7.509 7.230 7.184
4⊐ −0.021 −0.015 −0.004 −0.029 −0.116 −0.001 −0.015

Figure 7 The docking modeling of TBZ and HP⊎CD.

Figure 6 The fluorescence spectra of TBZ and TBZ/HP⊎CD-NF.

mentioning that the polymer-free electrospinning was successful,

mL, respectively. Compared to the original TBZ, the fungicidal
activity of TBZ/HP⊎CD-NF was 1.83 times higher. To sum up, the
formation of TBZ/HP⊎CD-NF is mainly through the interaction of
van der Waals force and hydrogen bonds, and does not change
the chemical characteristics of the fungicide itself. Therefore, it
does not affect the bactericidal mechanism of TBZ or the bacteri-
cidal activity in aqueous solution. It could be concluded that
TBZ/HP⊎CD-NF had better fungicidal activity than TBZ due to
the improvement of water solubility.37
3.8 Morphology analysis of nanofibers
Figure 9 shows a representative SEM image of electrospun nano-
fibers. As shown in the SEM images, TBZ and HP⊎CD were success-
fully electrospun into beadless and uniform nanofibers in an
aqueous system. According to the SEM images, the fiber diameter
distribution diagram of TBZ/HP⊎CD-NF was calculated by soft-
ware nano measurer (Fig. 9). The fiber diameter distribution of
TBZ/HP⊎CD-NF was between 90 and 750 nm having an average
and no carrier polymer matrix was used in this electrospinning
system. The homogeneous morphology of the nanofibers indi-
cated that the spinning conditions were properly controlled. In
general, the spinning conditions were effective and could pro-
duce uniform TBZ/HP⊎CD-NF. The fast-dissolution of TBZ/
HP⊎CD-NF was visually tested by adding 1 mL of water to a glass
vessel (Fig. 10). Due to the poor water solubility of TBZ, its powder
will not dissolve when water is added to the vessel. However,
HP⊎CD belongs to the type of cyclodextrin with high water solu-
bility, thus the pristine HP⊎CD web dissolves immediately upon
contact with water. When TBZ/HP⊎CD-NF was exposed to water,
TBZ/HP⊎CD-NF also completely dissolved immediately, there
was no indication of undissolved TBZ when TBZ/HP⊎CD-NF was
dissolved confirming that the water solubility of TBZ was
improved compared to its powder form. From XRD data, there
was a few amounts of crystalline uncomplexed TBZ in TBZ/
HP⊎CD-NF, and dissolution tests indicated that the uncomplexed
TBZ in TBZ/HP⊎CD-NF could not dissolve in the dissolution
3269

fiber diameter of 370 ± 198 nm. In addition, it is worth medium. Even so, we did not detect any undissolved TBZ during

Pest Manag Sci 2020; 76: 3264–3272 © 2020 Society of Chemical Industry wileyonlinelibrary.com/journal/ps
 www.soci.org S Gao et al.

Figure 8 The fungicidal activity of TBZ (A) and TBZ/HP⊎CD-NF (B) against Gibberella sp. (CTBZ = 0.10 μg/mL).

Table 3 The toxicity equations and EC50 of TBZ and TBZ/HP⊎CD-NF against Gibberella sp.

Toxic regression equation Correlation coefficient (R) EC50 (μg/mL) 95% Confidence limit Improved fold

TBZ y = 0.7913x + 5.3097 0.9996 0.406 0.379 ~ 0.420 —

TBZ/HP⊎CD-NF y = 1.0232x + 5.6688 0.9979 0.222 0.220 ~ 0.226 1.83

Figure 9 (A) SEM image of TBZ/HP⊎CD-NF and (B) the fiber diameter distribution graphs of TBZ/HP⊎CD-NF.
3270

Figure 10 Presentation of the solubility behavior.

wileyonlinelibrary.com/journal/ps © 2020 Society of Chemical Industry Pest Manag Sci 2020; 76: 3264–3272
Encapsulation of thiabendazole in hydroxypropyl-⊎-cyclodextrin nanofibers
the visual evaluation of fast-dissolution behavior. This might be
due to the homogenous distribution of TBZ through the nanofi-
brous webs even though it is in an uncomplexed crystal form.42
Enhanced water solubility is conducive to full drug efficacy and
reduction of residual drug.
4 CONCLUSION
In this study it was shown that HPßCD and the water-insoluble
fungicide TBZ could produce a self-supporting nanofiber web
without using a polymer matrix. From the phase solubility dia-
gram, it can be concluded that the solubility of TBZ increases with
increased HP⊎CD concentration. SEM images showed that TBZ
had a beadless morphology. The formation of TBZ/HP⊎CD-NF
was confirmed by 1H-NMR, FTIR, XRD and fluorescence analysis.
In addition, the fluorescence of TBZ was enhanced after an inclu-
sion complex was formed. Antifungal activity test results proved
that TBZ/HP⊎CD-NF showed high antifungal activity against Gib-
berella sp. Finally, TBZ/HP⊎CD-NF was observed to dissolve rapidly
in water. In summary, these results indicated that TBZ/HP⊎CD-NF
has great potential for improving the subsequent development of
TBZ and pesticide residue detection due to the enhanced solubil-
ity, and high antifungal activity of the large amounts of stored
TBZ. Additionally, the fungicide solid nanodispersion that we pre-
pared is a new pesticide formulation that can prevent the wide-
spread use of organic solvents, thus making great contributions
to the subsequent development of agriculture and pesticide resi-
due detection, and it conforms to the economic development
trends related to high efficiency, green technology and environ-
mental protection advocated by the state.
ACKNOWLEDGEMENTS
This work was supported by the National Natural Science Founda-
tion of China (31801784, 31572042), the Natural Science Founda-
tion of Heilongjiang Province (ZD2017002), the Heilongjiang
Province Postdoctoral Science Foundation (LBH-Z16030), and
the Research Science Foundation in Technology Innovation of
Harbin (2017RAQXJ017).
CONFLICT OF INTEREST
The authors declare no conflict of interest.
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