Study of the synthesis, antiviral bioactivity and interaction mechanisms of novel chalcone derivatives that contain the 1,1-dichloropropene moiety

Article abstract of DOI:10.1016/j.cclet.2017.03.013

A series of novel chalcone derivatives that contain the 1,1-dichloropropene moiety was designed and synthesized. Bioactivity assays showed that most of the target compounds exhibited moderate to good antiviral activity against tobacco mosaic virus (TMV) at 500?μg/mL. Among the target compounds, compound 7h showed the highest in vivo inactivation activity against TMV with the EC₅₀ and EC₉₀ value of 45.6 and 327.5?μg/mL, respectively, which was similar to that of Ningnanmycin (46.9 and 329.4?μg/mL) and superior to that of Ribavirin (145.1 and 793.1?μg/mL). Meanwhile, the microscale thermophoresis and fluorescence spectroscopy experiments showed that the compound 7h had a strong interaction with the tobacco mosaic virus coat protein.

Full text of DOI:10.1016/j.cclet.2017.03.013

Accepted Manuscript
Title: Study of the synthesis, antiviral bioactivity and
interaction mechanisms of novel chalcone derivatives that
contain the 1,1-dichloropropene moiety
Authors: Liang-Run Dong, De-Yu Hu, Zeng-Xue Wu,
Ji-Xiang Chen, Bao-An Song
PII:
S1001-8417(17)30091-8
DOI:
Reference:
http://dx.doi.org/doi:10.1016/j.cclet.2017.03.013
CCLET 4008
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Chinese Chemical Letters
Received date:
Accepted date:
8-3-2017
8-3-2017
Please cite this article as: Liang-Run Dong, De-Yu Hu, Zeng-Xue Wu, Ji-Xiang Chen,
Bao-An Song, Study of the synthesis, antiviral bioactivity and interaction mechanisms
of novel chalcone derivatives that contain the 1,1-dichloropropene moiety, Chinese
Chemical Lettershttp://dx.doi.org/10.1016/j.cclet.2017.03.013
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Original article
Study of the synthesis, antiviral bioactivity and interaction mechanisms of novel
chalcone derivatives that contain the 1,1-dichloropropene moiety
Liang-Run Dong, De-Yu Hu*, Zeng-Xue Wu, Ji-Xiang Chen, Bao-An Song
State Key Laboratory Breeding Base of Green Pesticide and Agricultural Bioengineering, Key Laboratory of Green Pesticide and Agricultural Bioengineering,
Ministry of Education, Research and Development Center for Fine Chemicals, Guizhou University, Guiyang 550025, China.
 Corresponding author.
E-mail address: basong@gzu.edu.cn (B.-A. Song)
Graphical Abstract
A series of chalcone derivatives that contain the 1,1-dichloropropene moiety was synthesized and assayed for the antiviral activity against
tobacco mosaic virus. Compound 7h showed significant inactivation activity against TMV and strong interaction with the tobacco mosaic
virus coat protein.
ABSTRACT
A series of novel chalcone derivatives that contain the 1,1-dichloropropene moiety was designed and synthesized. Bioactivity assays showed that most of the
target compounds exhibited moderate to good antiviral activity against tobacco mosaic virus (TMV) at 500 μg/mL. Among the target compounds, compound 7h
showed the highest in vivo inactivation activity against TMV with the EC50 and EC90 value of 45.6 and 327.5 μg/mL, respectively, which was similar to that of
Ningnanmycin (46.9 and 329.4 μg/mL) and superior to that of Ribavirin (145.1 and 793.1 μg/mL). Meanwhile, the microscale thermophoresis and fluorescence
spectroscopy experiments showed that the compound 7h had a strong interaction with the tobacco mosaic virus coat protein.
Keywords:
Chalcone derivatives
1,1-Dichloropropene moiety
Synthesis
Antiviral activity
Interaction mechanisms
ARTICLE INFO
Article history:
Received 15 November 2016
Received in revised form 15 December 2016
Accepted 16 January 2017
Available online

1. Introduction
Tobacco mosaic virus (TMV), a positive-sensesingle stranded RNA virus, infects a wide range of plants, especially tobacco,
vegetable and other members of the Solanaceae family. The virus is globally ubiquitous and causes great crop loss [1]. Therefore,
TMV is an uncontrollable form of “plant cancer”. Ribavirin and Ningnanmycin are common treatments for TMV infections under field
conditions. However, Ribavirin has poor inactivation activity, and Ningnanmycin unstable and expensive [2, 3]. Therefore, the
development of a novel, simple, and high-efficiency antiviral agents is a significant challenge in pesticide science [4].
Chalcone is a natural product, that exists in licorice, saffron and other medicinal plants. Chalcones have a wide range of
pharmacological activities [5], such as anti-Alzheimer’s disease [6], antitumor [7], antibacterial [8], insecticidal [9], anti-inflammatory
[10], anti-oxidation [11], antiplatelet [12], and antiviral activities [13–15]. In our previous work, we reported a variety of chalcone
derivatives and evaluated for their antiviral activity [16–18].The results indicated that these compounds have moderate to good
antiviral activity against TMV.
1,3-Dichloropropene is a water-soluble and volatile compound that is mainly used as a pre-planting fumigant and nematicide. It is
widely used in the US and other countries [19]. Its analogue, 1,1-dichloropropene, also displays similar activities [20–23]. Pyridalyl, a
commercially available 3,3-dichloro-2-propenyloxy-substituted compound, has excellent controlling effects on various lepidopterous
and thysanopterous insects. It shows no cross-resistance with the existing insecticides, such as synthetic pyrethroids, organic
phosphates, nicotinic insecticides and ryanodine insecticides [24]. 1,1-Dichloropropene compounds have drawn scientific attention as
an interesting foundation for generating new lead compounds. Recently Liu et al. [25] synthesized several dichloro-allyloxy-phenol
derivatives by introducing various substituted phenyl groups into the pyrazole ring. The resulting insecticidal compounds are active
against P. xylostella at 6.25 μg/mL and are more potent than the reference pyridalyl. In 2015, Yang and co-workers synthesized a series
of the novel 1,1-dichloropropene derivatives bearing diverse heterocycles with insecticidal activity against bollworm and P. litura [26].
Though, the 1,1-dichloropropene group is an important and applicable functional group for designing the insecticide, the corresponding
derivatives that contain the 1, 1-dichloropropene moiety with antiviral activities have not been reported.
In order to evaluate antiviral activity of 1,1-dichloropropene compounds, the novel chalcone derivatives that contain the 1,
1-dichloropropene moiety were designed and synthesized. The physical characteristics were determined by 1H NMR, 13C NMR, IR and
elemental analysis, and their antiviral activities were evaluated. The bioassay results indicated that the target compounds exhibited
moderate to good inactivation activity against TMV. Compounds 7g, 7h, 7m, 7n, 7w, and 7z showed higher antiviral activity against
TMV with EC50 values of 52.9, 45.6, 52.2, 53.8, 50.6, and 53.4 μg/mL, respectively, compared with Ribavirin (145.1 μg/mL).
Furthermore, the interactions between target compounds and tobacco mosaic virus coat protein (TMV CP) were investigated by the
fluorescence spectroscopy and microscale thermophoresis (MST).
2. Results and discussion
2.1. Chemistry
The synthetic route of chalcone derivatives that contain the 1, 1-dichloropropene moiety is summarized in Scheme 1. Intermediate 1
can be easily prepared with 1,1,1,3-tetrachloropropane [21]. Hydroquinone, 1,1,3-trichloroprop-1-ene 1 and K2CO3 were refluxed in
CH3CN for 5 h to obtain intermediate 2 after solvent removal and dichloromethane extraction. Intermediate 2 and diethylamine as a
catalyst were stirred in toluene. The mixture was slowly added to a solution of SO2Cl2. Then, the reaction mixture was stirred at 56 °C
for 5 h to produce intermediate 3 [27]. Intermediate 3, K2CO3, KI, and 1,3-dibromopropane were stirred at room temperature for 12 h
to yield intermediate 4. Intermediates 5 and 6 were prepared in accordance with previously reported methods [25, 26]. Finally,
intermediates 4 and 6 were reacted by etherification to yield the target compounds 7a–7z. The target products were characterized by 1H
NMR, 13C NMR, IR, and elemental analysis. The data of spectroscopic characterization of the target compounds can be found in the
Supporting Information.
2.2. Antiviral activity
The antiviral activities against TMV of the title compounds 7a–7z were determined via the half-leaf method [28,29]. As shown in
Table 1, the bioassay results indicated that all of the target compounds exhibited moderate to good activity against TMV at the
concentration of 500 μg/mL. Among the targets, compounds 7g, 7h, 7m, 7n, 7w, and 7z showed better inactivation activity against
TMV, with the inhibition rates of 93.1%, 94.8%, 93.5%, 92.9%, 94.0%, and 93.1%, respectively. These inhibition rates were similar to
that of Ningnanmycin (94.1%) and superior to that of Ribavirin (72.9%). The other compounds displayed moderate inactivation
activity against TMV.
To establish the structure-activity relationships of the target compounds, the EC50 values of the target compounds’ inactivation
activities against TMV were evaluated. The evaluation results are presented in Table 2. All target compounds displayed excellent

inactivation activity. Compounds 7g, 7h, 7m, 7n, 7w, and 7z showed antiviral activity against TMV with EC50 and EC90 values of 52.9,
45.6, 52.2, 53.8, 50.6, 53.4 and 332.9, 327.5, 331.5, 333.7, 331.0, 333.2 μg/mL, respectively. The inactivation activity of these
compounds was better than that of Ribavirin (145.1 and 793.1 μg/mL). Among these six compounds, compound 7h exhibited the best
inactivation activity against TMV and was similar to that of Ningnanmycin (46.9 and 329.4 μg/mL). Structure-activity relationship
analysis revealed that differences in R groups have an important influence on inactivation activity. When the R groups were
4-OCH3-Ph(7h), 3-OCH3-Ph (7m), 4-OCH2CH3-Ph (7q), and 4-OCF3-Ph (7z), the corresponding compounds displayed good activity
compared with 7e (2-Cl-Ph), 7f (2-Br-Ph), 7g (2-F-Ph) and 7k (2-CF3-Ph). Therefore, the introduction of electron-donating groups
improves the anti-TMV activity of the chalcone compounds.
2.3. Interaction between 7h and TMV-CP
Fluorescence spectroscopy and MST analysis were used to investigate the interactions between target compounds and TMV CP [30].
The test methods are shown in the supporting data. The fluorescence spectrum showed that the binding constant Ka between 7h and
TMV-CP was 2.63 × 105 L/mol, which was similar to that of Ningnanmycin (2.39 × 105 L/mol) and superior to that of Ribavirin (2.57×
103 L/mol). Compound 7r (3.98× 104 L/mol) showed moderate affinity and 7j (6.81× 103 L/mol) showed weak affinity, as shown in Fig.
1 and Table 3. To further confirm the results of fluorescence spectroscopy, the Kd between compound 7h and TMV CP was
investigated with MST. The result of MST analysis was consistent with the data presented by the fluorescence spectrum, as shown in
Fig. 2 and Table 3. The Kd between 7h and TMV-CP was 9.51 μmol/L, which was higher than those of 7r (34.8 μmol/L) and 7j (162
μmol/L). Therefore, the result showed that compound 7h strongly interacted with TMV CP.
3. Conclusion
In summary, we synthesized novel chalcone derivatives that contain a 1,1-dichloropropene moiety. The anti-TMV activity of the
target compounds 7a–7z was evaluated with the in vivo half-leaf method. Bioassay results showed that the target compounds were
better than Ribavirin (145.1 μg/mL) against TMV. In particular, compound 7h possessed excellent inactivation activity against TMV
and had an EC50 value of 45.6 μg/mL, which was similar to that of Ningnanmycin (46.9 μg/mL). In addition, fluorescence spectroscopy
and MST showed that compound 7h strongly interacted with TMV CP. Further studies on the mechanism of inactivation are currently
underway.
4. Experimental
4.1. Synthesis
Unless otherwise stated, allreactants andreagents were purchased from commercial suppliers. The melting points were determined on
an XT-4 binocular microscope (Beijing Tech Instrument, China) and were not corrected. Mass Spectra were obtained with a LC-MS
1
1100/MSD spectrometer (Agilent company, America). H NMR and 13C NMR (solvent CDCl3) spectral analyses were performed on a
1
JEOL ECX 500 NMR spectrometer operated at 500 MHz for H NMR, 125 MHz for 13C NMR at room temperature and TMS as the
internal standard. Elements were performed with an Elemental Vario-III CHN analyzer, and IR spectra were acquired in KBr on a
Bruker VECTOR 22 spectrometer. Analytical thin layer chromatography (TLC) was performed on silica gel GF254
.
General procedure for the preparation of the key intermediate 6a–6z: Intermediates 6a–6z were prepared by performing previously
reported synthetic procedures [24, 25]. A mixture of 4-hydroxychalcone (1.00 g, 4.2 mmol) and nitromethane (5.12 g, 83.9 mmol) in
anhydrous ethanol was stirred. The mixture was refluxed for 2 h after the addition of KOH (0.28 g, 5.04 mmol). The reaction was
monitored to completion via TLC. After the reaction system was cooled, its pH was adjusted with acid. Then, the mixture was filtered
and the supernatant was discarded. The crude solid was purified via TLC on silica gel (petroleum ether/ethyl acetate = 3:1). The
physical and chemical properties and spectroscopic characterization of the target compounds can be found in the Supporting
information.
General synthetic procedures for the title compounds 7a–7z: A mixture of intermediates 4 (1.2 mmol), 6 (1.0 mmol) and potassium
carbonate (2.0 mmol), with KI in acetone as the catalyst, was stirred and refluxed for 8 h. The solvent was evaporated in vacuo and the
residue was isolated via column chromatography on silica gel (petroleum ether/dichloromathane = 1:1) to obtain the title compounds.
The physical and chemical properties and spectroscopic characterization of the target compounds can be found in the Supporting
information.
4.2. Bioassay
The in vivo antiviral activities against TMV of the target compounds were examined. Ningnanmycin was used as the control. EC50 of
the antiviral activity against TMV was determined. The assays were repeated thrice [28, 29].
4.3. Kd of compound 7h to TMV CP
The in vitro binding affinity of the title compounds on TMV-CP was performed by MST and fluorescence spectroscopy
measurement as previously described [30].

Acknowledgment
This research was supported by the National Natural Science Foundation of China (Nos. 21362004, 21562013) and Subsidy Project
for Outstanding Key Laboratory of Guizhou Province in China (20154004).
Appendix A. Supplementary data
Supplementary material related to this article can be found, in the online version, at http://www.sciencedirect.com/
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Fig. 1. Fluorescence emission spectra of TMV-CP in the presence of 7h (A); Ningnanmycin (B); Ribavirin (C); 7r (D); and 7j (E)

Fig. 2. Microscale thermophoresis (MST) of 7h (A); Ningnanmycin (B); Ribavirin (C); 7r (D); and 7j (E)

Scheme 1. Synthesis of compounds 7a to7z

Table 1. In vivo activity against TMV of the target compounds
Compd.
R
Concentration (μg/mL) Inactivation effect (%)
7a
4-CH3-Ph
500
90.9±2.4
4-Cl-Ph
Ph
500
500
500
500
500
500
500
500
500
500
500
500
500
500
500
500
500
500
500
500
500
500
500
500
500
500
500
89.0±3.1
89.9±1.9
86.5±2.0
90.8±3.2
91.5±5.4
93.1±1.6
94.8±2.8
91.4±1.0
78.8±1.1
91.3±3.2
87.4±2.3
93.5±4.1
92.9±1.0
90.8±1.7
90.1±1.6
91.6±1.3
87.7±3.8
88.0±2.9
88.6±6.7
83.2±5.3
91.5±2.1
94.0±1.5
85.3±4.6
81.5±1.5
93.1±2.0
94.1±1.4
72.9±2.4
7b
7c
4-F-Ph
7d
2-Cl-Ph
7e
2-Br-Ph
7f
2-F-Ph
7g
4-OCH3-Ph
2-OCH3-Ph
4-Br-Ph
7h
7i
7j
2-CF3-Ph
3-Br-Ph
7k
7l
3-OCH3-Ph
2,3-diCl-Ph
2-Cl-6-F-Ph
3, 4-diOCH3-Ph
4-OCH2CH3-Ph
4-CH(CH3)2-Ph
furan-2-yl
4-NO2-Ph
2,4-diCl-Ph
3,4-diCl-Ph
pyridin-2-yl
2,6-diCl-Ph
3-NO2-Ph
4-OCF3-Ph
/
7m
7n
7o
7p
7q
7r
7s
7t
7u
7v
7w
7x
7y
7z
Ningnanmycin
Ribavirin
/

Table 2. In vivo EC50 values of compounds 7a-7z against TMV
Compd.
R
EC50/EC90 (μg/mL)
Compd.
R
EC50/EC90 (μg/mL)
4-CH3-Ph
4-Cl-Ph
63.2±2.6
77.7±3.4
7o
7p
2-Cl-6-F-Ph
66.1±2.4
71.7±3.1
7a
7b
3,4-diOCH3-Ph
Ph
74.6±2.6
97.2±1.8
4-OCH2CH3-Ph
4-CH(CH3)2-Ph
furan-2-yl
4-NO2-Ph
2,4-diCl-Ph
3,4-diCl-Ph
pyridin-2-yl
2,6-diCl-Ph
3-NO2-Ph
4-OCF3-Ph
/
56.3±2.8
94.3±2.2
7c
7d
7e
7f
7q
4-F-Ph
7r
2-Cl-Ph
65.5±3.3
90.5±3.6
7s
2-Br-Ph
57.6±1.9
88.7±2.4
7t
2-F-Ph
52.9±3.7/332.9±3.8
45.6±2.8/327.5±2.7
60.8±1.3
106.7±3.8
7g
7h
7i
7u
4-OCH3-Ph
2-OCH3-Ph
4-Br-Ph
59.8±1.3
7v
50.6±2.5/331.0±3.3
102.5±2.1
7w
121.3±2.7
7j
7x
7y
2-CF3-Ph
3-Br-Ph
61.5±3.1
110.6±3.7
7k
7l
95.1±1.6
53.4±1.6/333.2±2.8
46.9±1.5/329.4±3.7
145.1±2.6/793.1±2.3
7z
3-OCH3-Ph
2,3-diCl-Ph
52.2±3.2/331.5±3.1
53.8±1.7/333.7±3.4
Ningnanmycin
Ribavirin
7m
7n
/
Table 3. Binding constant of 7h, 7r, 7j, Ningnanmycin and Ribavirin
MST
(μmol/L)
9.51
Compd.
Fluorescence spectroscopy (L/mol)
2.63×105
7h
7r
3.98× 104
6.81× 103
2.39×105
2.57× 103
34.8
162
9.92
473
7j
Ningnanmycin
Ribavirin