Synthesis, bioactivity, action mode and 3D-QSAR of novel anthranilic diamide derivatives

Article abstract of DOI:10.1016/j.cclet.2018.05.023

To study the pesticide effect, action mode, structure-activity relationships (SARs) of anthranilic diamide insecticide and screen highly active pesticides, novel anthranilic diamide derivatives were synthesized. Bioassays indicated that all of the title compounds displayed 100% mortality against diamondback moth and oriental armyworm at 100 mg/L, among which 12v and 12w showed 100% insecticidal acitvity at 5 mg/L. Surprisingly compound 12w exhibited better insecticidal acitvity than commercialized chlorantraniliprole against Pyrausta nubilalis (0.1 mg/L) and Cnaphalocrocis Medinalis (2 mg/L). 3D-QSAR and SARs statistical analysis revealed that title compounds with R² fixed as methoxy had the highest probability possessing high activity. The calcium fluorescence measurements on neurons revealed that E series compounds containing pyrazinyl may have a molecular target different from caffeine on ryanodine receptors rather than the voltage-gated calcium channel present on cytomembran.

Full text of DOI:10.1016/j.cclet.2018.05.023

G Model
CCLET 4555 No. of Pages 4
Chinese Chemical Letters xxx (2018) xxx–xxx
Contents lists available at ScienceDirect
Chinese Chemical Letters
journal homepage: www.elsevier.com/locate/cclet
Communication
Synthesis, bioactivity, action mode and 3D-QSAR of novel anthranilic
diamide derivatives
*
*
Weijie Liu, Jiao Li, Kai He, Fangfang Huang, Yi Ma, Yuxin Li, Qingshan Li , Fengbo Xu
State Key Laboratory of Elemento-Organic Chemistry, College of Chemistry, Nankai University, Tianjin 300071, China
A R T I C L E I N F O
A B S T R A C T
Article history:
To study the pesticide effect, action mode, structure-activity relationships (SARs) of anthranilic diamide
insecticide and screen highly active pesticides, novel anthranilic diamide derivatives were synthesized.
Bioassays indicated that all of the title compounds displayed 100% mortality against diamondback moth
and oriental armyworm at 100 mg/L, among which 12v and 12 w showed 100% insecticidal acitvity at 5
mg/L. Surprisingly compound 12 w exhibited better insecticidal acitvity than commercialized
chlorantraniliprole against Pyrausta nubilalis (0.1 mg/L) and Cnaphalocrocis Medinalis (2 mg/L). 3D-QSAR
and SARs statistical analysis revealed that title compounds with R2 fixed as methoxy had the highest
probability possessing high activity. The calcium fluorescence measurements on neurons revealed that E
series compounds containing pyrazinyl may have a molecular target different from caffeine on ryanodine
receptors rather than the voltage-gated calcium channel present on cytomembran.
Received 16 April 2018
Received in revised form 9 May 2018
Accepted 11 May 2018
Available online xxx
Keywords:
Anthranilic diamide
Ryanodine receptor
Insecticide
Action mode
3D-QSAR
© 2018 Chinese Chemical Society and Institute of Materia Medica, Chinese Academy of Medical Sciences.
Published by Elsevier B.V. All rights reserved.
To overcome resistance and ecological problems involved with
conventional insecticides, there was an urgent need to discover
novel potent insecticides with a new mode of action [1]. Ryanodine
receptors (RyR) are members of a family of intra-cellular Ca2+
release channel proteins present on endoplasmic reticulum (ER) or
sarcoplasmic reticulum (SR) membranes, deriving the name from
its high affinity with ryania [2,3]. Ryanodine receptors insecticides
had already become a focus of pesticide discovery and research
because of its good insecticidal activity and high safety for non-
target creatures such as mammal [4–6]. Natural ryanodine
receptors insecticides include caffeine, spiganthine, 9,21-dehy-
droryanodine, ryanodol, etc. [7]. Commercial ryanodine receptors
inscticides include phthaldiamide such as flubendiamide (A)
discovered by Nihon Nohyaku in 1993, anthranilic diamide such
as cyantraniliprole (B) and chlorantraniliprole (C) discovered by
DuPont [8–11] (Fig. 1).
In this article, we divided the chemical structure of chloran-
traniliprole (Fig.1D) into three parts: N-pyridylpyrazole moiety (a),
anthraniloyl moiety (b), and aliphatic moiety (c). In our previous
work, o-carboxamidobenzamide compounds containing 2-(substi-
tuted phenyl)oxazole group (Fig. 1C) in part a were synthesized,
but unfortunately most of the compounds did not showed higher
bioactivity than commercialized chlorantraniliprole [12]. Based on
this and encouraged by the extensive and excellent biological
activity of pyrazine derivants in the fields of medicine [13],
pesticide [14,15], flavouring [16], etc., N-pyrazinylpyrazole was
introduced into part a according to the principle of bioelectronics
to give the series of E compounds (Fig. 1E). SARs analysis revealed
that E series containing N-pyrazinylpyrazole in part a exhibited
inferior bioactivity compared with chlorantraniliprole containing
N-pyridylpyrazole, in addition, title compounds with methoxyl-
carbamoyl in patrt c had the highest probability possessing high
activity, so the series of F compounds (12v and 12 w) containing
methoxylcarbamoyl on part c and N-pyridylpyrazole in part a were
further designed (Fig. 1F and Scheme 1). Three dimensional
quantitative structure-activity relationship (3D-QSAR) and action
mode of title compounds were also investigated in this paper.
The synthesis of compounds 12a–w is shown in Scheme 1. The
pure product of 2-chloro-6-hydrazinylpyrazine (2a) can easily be
obtained after filtration instead of extraction and distillation by the
reaction of 2,6-dichloropyraine with 80% hydrazine hydrate at
present of ethanol. Compound 4a was synthesized according the
reported method for chlorantraniliprole synthesis with minor
improvements [5]. During which process, it was found that 2-
hydrazinylpyrazine and 2-chloro-6-hydrazinylprazine can suc-
cessfully be reacted with diethyl maleate by controlling the
reaction temperature, but 3-chloro-6-hydrazinylpyrazine cannot
be reacted. Compound 4a was bromized with phosphorus
oxybromide to give compound 5a. Then 5a was hydrolysed with
* Corresponding authors.
E-mail addresses: qli@nankai.edu.cn (Q. Li), xufb@nankai.edu.cn (F. Xu).
https://doi.org/10.1016/j.cclet.2018.05.023
1001-8417/© 2018 Chinese Chemical Society and Institute of Materia Medica, Chinese Academy of Medical Sciences. Published by Elsevier B.V. All rights reserved.
Please cite this article in press as: W. Liu, et al., Synthesis, bioactivity, action mode and 3D-QSAR of novel anthranilic diamide derivatives, Chin.
Chem. Lett. (2018), https://doi.org/10.1016/j.cclet.2018.05.023

G Model
CCLET 4555 No. of Pages 4
2
W. Liu et al. / Chinese Chemical Letters xxx (2018) xxx–xxx
Fig. 2. (A) Superimposition of the training and test sets. (B) Electrostatic map
form CoMFA model.
Fig. 1. Chemical structures of compounds A–F.
N-pyrazinylpyrazole against diamondback moth at 50 mg/L, 12a-
u were categoried and sorted into three types: H (high activity with
percentage 70–100), M (moderate activity with percentage 40–70),
L (low activity with percentage 0–40). As shown in Table S1 and
Fig. S1 in Supporting information, seven of E series fell into H
category when R2 fixed as OCH3, so 87.5% were H, similarly, 75% are
H as CH3, 40% as OEt and 0% as cyclopropyl, consequengtly the
order from high to low was OCH3, CH3, OEt, cyclopropyl. As a result,
title compounds with methoxylcarbamoyl in patrt c had the
highest probability possessing high activity, in addition, keeping R,
R1, R3 same with each other, compounds with the highest activity
in each series were 12c,12d,12e,12f,12k,12l and 12n, respectively.
It was found that 5/7 of these had a methoxylcarbamoyl in part a.
Bulky substituent in part a could lead to a decrease activity, for
example, 12 m with R2 fixed as CH3 exhibited 80% mortality while
12t only showed 10% with cyclopropyl. The results analysed above
did explain that the later designed 12v and 12 w bearing
methoxylcarbamoyl in part c and N-pyridylpyrazole in part a
could show relatively higher activity than 12a-u and why keeping
the other chemical structure consistent with each other 12 w could
exhibit higher actvity than chlorantraniliprole. For R3, the
corresponding sequence was OCH3 > Br > Cl, for R1, it was Cl
> CH3. It was also found that title compound with R and R1 fixed as
Cl and Cl showed higher activity than those fixed as Cl and CH3
respectively, which was demonstrated by the sequence 12d > 12 l,
12e > 12s, 12f > 12p, 12 g > 12n, 12i > 12 u.
NaOH to give the corresponding carboxylic acid 6a. Compound 6a
was converted into the acyl chloride 8a by treatment with thionyl
chloride and pyrazolone ring was constructed at the same time.
Methoxy substituted intermediates 7a can directly be obtained by
refluxing compound 6a in methanol, which was not reported
before.
Compound 9a was cyclized by triphosgene to give the
corresponding six-membered lactam 10a at a high yield of 98%.
Amine hydrochloride derivatives were firstly deprotonated with
triethylamine to release their nucleophilicity, and then were used
for ring opening reaction of lactam 10a to give 2-amino-5-
chlorobenzamide derivatives 11. Title compounds 12a-w were
synthesized with acyl chloride
8 and amine 11 throungh
nucleophilic substitution reaction in the presence of organic base
triethylamine.
The insecticidal activities of 12a-w against diamondback moth
and oriental armyworm were shown in Table 1. All of title
compounds showed 100% insecticidal activity against diamond-
back moth and oriental armyworm at 100 mg/L. Title compounds
12c, 12v and 12 w exhibited 100% mortality at 50 mg/L, but when
the concentration was reduced to 5 mg/L, only 12 w keep 100%
mortality against both diamondback moth and oriental army-
worm. For compound 12 w with excellent activity, its insecticidal
activity against Mythimnaseparata walker, Pyrausta nubilalis,
Cnaphalocrocis Medinalis at concentrations below 5 mg/l were
further investaged. The results were listed in Table 2, it was worth
mentioning that 12 w showed better activity than contrast
compound chlorantraniliprole against Pyrausta nubilalis (0.1 mg/
L) and Cnaphalocrocis Medinalis (2 mg/L).
Comparative molecular field analysis (CoMFA) is an effective
computer implemented methodology of 3D-QSAR employing both
the interactive graphics and statistical techniques for correlating
structure of molecules with observed biological properties, and
thus is widely used in drug design [17,18]. The results of CoMFA
computation was summarized in Table S2 (Supporting informa-
tion). The cross-validated coefficient q2 obtained by leave-one-out
method is 0.658, where a value over 0.5 indicates the model is a
In order to evaluate the SARs of title compounds systematically,
according to the insecticidal activity of
E series bearing
Scheme 1. Synthetic route of 12a-w. Reagent and conditions: (a) NH2NH2, EtOH, 80 ꢀC; (b) Na, EtOH, HAc, 50 ꢀC; (c) P(O)X3, CH3CN; (d) NaOH, H2O, EtOH 65 ꢀC; (e) SOCl2,
65 ꢀC; (f) triphosgene, 1,4-dioxane, 40 ꢀC; (g) R2NH2, CH3COOEt, 40 ꢀC; (h) Et3N, CH3CN, r.t.; (i) CH3CN.
Please cite this article in press as: W. Liu, et al., Synthesis, bioactivity, action mode and 3D-QSAR of novel anthranilic diamide derivatives, Chin.
Chem. Lett. (2018), https://doi.org/10.1016/j.cclet.2018.05.023

G Model
CCLET 4555 No. of Pages 4
W. Liu et al. / Chinese Chemical Letters xxx (2018) xxx–xxx
3
Table 1
Table 2
Insecticidal activity (%) against diamondback moth and oriental armyworm.
Insecticidal activity (%) of 12 w against different insects.
Insects
Compd.
Concentration (mg/L)
0.1
0.5
1
2
4
Mythimnaseparata walker
Pyrausta nubilalis
12 w
100
100
60
50
–
100
100
100
100
–
100
100
100
100
–
100
100
100
100
26
100
100
100
100
59
Controla
12 w
Controla
12 w
Cnaphalocrocis medinalis
Controla
–
–
–
19
28
Compd.
R
R1
R2
R3
Insecticidal activity (%)
aChlorantraniliprole.
-: Not tested.
Diamondback
moth
Oriental
armyworm
100b 50b 5b
100b
5b
12a
12b
12c
12d
12e
12f
12 g
12h
12i
12j
12k
12 l
12 m
12n
12o
12p
12q
12r
H
H
H
Cl Cl
Cl Cl
Cl Cl
Cl Cl
Cl Cl
Cl Cl
Cl Cl
Cl CH3 CH3
Cl CH3 OCH3
Cl CH3 CH3
Cl CH3 OCH3
Cl CH3 CH3
Cl CH3 OEt
Cl
OCH3
Br
Br
Br
100
100
100
90
30
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
–
higher activity, as shown in the sequence OCH3 > Br > Cl discussed
above. Alkoxycarbamoyl in part c displayed higher activity because
they occupy the red region, and a relatively positive-charged alkyl
part in alkoxycarbamoyl was favorable for the increase of activity.
Title compounds 12c, 12d, 12e and chlorantraniliprole at a
concentration of 100 mg/L were applied to neurons of beet
armyworm, after which the effect of that on free cytosolic calcium
concentration ([Ca2+]c) in beet armyworm neurons was shown in
Fig. 3A. It was observed that the application of 12c,12d and 12e can
induce a rapid increase of [Ca2+]c in single neuron. Previous studies
suggested that calcium signal was generated by calcium ion store
present on ER in neurons of insects. Ryanodine receptors, which
locates at ER membranes of insect nerve cells, is the ligand-gated
calcium channel regraluting release of calcium ion [2]. It was
reported that the transfer of calcium from ER to cytosol in insect
neurons can indirectly evoke the muscle contraction [19]. In view
of the description above, the action mode could be correctly
conjectured (Fig. 3B). The combination of tested compounds with
RyR resulted in the sustained release of calcium ion from ER to
cytoplasm, which induced the stress contraction of insect muscle
fiber. Ultimately, pests died from symptoms of body contraction,
paralysis, feeding cessation, etc. [20]. It was concluded that the
target of E series of compounds is the ligand-gated calcium
channel present on ER, and thus they may be RyR activators.
To explore the effect of E series of compounds on voltage-gated
calcium channel present on cytomembrane of neurons. Title
compound 12e (100 mg/L) was applied to beet armyworm neurons
at the presence and absence of Ca2+ in extracellular fluid and its
CH3 CH3
CH3 OCH3
100 50
100 60
100 60
100 80
95
95
90
80
50
–
40
–
OCH3
OCH3
OEt
OCH3
CH2CN
OEt
OCH3 100
Cl
Br
Br
Br
100
100
100
100
–
50
40
30
20
0
–
–
–
OCH3 100
Cl
Cl
OCH3 100
OCH3 100
Br
Br
Br
–
OEt
100
100
–
100 60
–
90
80
80
60
60
60
55
50
10
10
30
0
0
0
0
0
0
0
–
–
–
100
100
100
100
100
100
–
–
–
Cl CH3 Morpholin-1-yl Br
Cl CH3 Cyclopropyl
Cl CH3 OCH3
Cl CH3 Cyclopropyl
Cl CH3 OEt
–
Br
Cl
–
12s
–
12 t
12 u
12v
12 w
Controla
OCH3 100
OCH3 100
100
–
–
–
100 100 100
100 100 100
100 100 100
60
100
100
100
100
-: Not tested.
a
Chlorantraniliprole.
Concentration of tested compound (mg/L).
b
great predictor of the insecticidal activity. The results of structural
alignment and isocontour diagrams of the electrostatic field
contributions obtained from the CoMFA analysis were illustrated in
Fig. 2, in which a postive charged group in the blue region will
increase the activity but a negative charged substituent in the red
region will be favorable.
Title compounds bearing an electron-withdrawing group at 3-
or 4- position and an electron-donating group at 6- position of the
pyrazine or pyridine ring, such as 12v, 12 w, and 12c, displayed
higher activity. At the 3-position of the imidazole ring (R3),
compounds bearing an electron-donating groups would display
effect on [Ca2+
] was recorded (Fig. 3C). After beet armyworm
c
neurons were loaded by incubation with 100 mL Fluo-3-AM for 3 h,
F/F0 (%) reached a maximum of about 109 % at 5 min after
application under both calcium-rich and calcium-free conditon in
extracellular fluid. As a result, title compound 12e cannot activate
the voltage-gated calcium channel present on cytomembrane of
neurons and have no molecular target on it.
After beet armyworm neurons were loaded by incubation with
100
mL Fluo-3-AM for 3 h, under calcium-free condition in
extracellular fluid, the effect of the application of 12e (100 mg/L)
Fig. 3. (A) Effect of 12c, 12d, 12e and chlorantraniliprole (100 mg/L) on [Ca2+]c in beet armyworm neurons. (B) Schematic illuatration of the molecular target and action mode
of E series of compounds. (C) Effect of Ca2+ in extracellular fluid on [Ca2+
c in beet armyworm neurons after the application of 12e (100 mg/L). (D) Effect of the application of
12e (100 mg/L) and caffeine (30 in beet armyworm neurons.
mol/L) during different time intervals on [Ca2+
]
m
]
c
Please cite this article in press as: W. Liu, et al., Synthesis, bioactivity, action mode and 3D-QSAR of novel anthranilic diamide derivatives, Chin.
Chem. Lett. (2018), https://doi.org/10.1016/j.cclet.2018.05.023

G Model
CCLET 4555 No. of Pages 4
4
W. Liu et al. / Chinese Chemical Letters xxx (2018) xxx–xxx
and caffeine (30
m
mol/L) during different intervals on [Ca2+
]
in
Appendix A. Supplementary data
c
beet armyworm neurons was recorded (Fig. 3D). It was observed
that the prior application of 12e had no effect on [Ca2+]c change of
Supplementary material related to this article can be found, in
the online version, at doi: https://doi.org/10.1016/j.cclet.2018.05.
023.
caffeine application and the subsequent [Ca2+
] change of 12e
c
application was not affected by prior application of caffeine
neither. If the molecular target of caffeine was occupied by the
priorly applied 12e, a rise on [Ca2+
]
would not be observed, in
References
c
other words, title compound 12e has a molecular target on RyR
different from natural ryanodine receptors insecticide caffeine.
In summary, we synthesized 21 novel anthranilic diamide derivatives
containing pyrazinyl, 2 derivatives containing methoxylcarbamoyl in the
aliphatic part of chlorantraniliprole. Bioassays indicated that all of the title
compounds displayed 100% mortality against diamondback moth and oriental
armyworm at 100 mg/L, based on analysis, we obtained title compound 12 w
with better insecticidal activity than chlorantraniliprole against
Pyrausta nubilalis (0.1 mg/L) and Cnaphalocrocis Medinalis (2 mg/L).
The 3D-QSAR results provided significant information for the
structure optimization of anthranilic diamide insecticides, espe-
cially, 12 w can serve as lead compound to screen highly active
pesticide and have the value of further research. The calcium
fluorescence measurements results revealed that E series com-
pounds containing pyrazinyl may possess a molecular target
different from caffeine on RyR rather than the voltage-gated
calcium channel present on cytomembrane.
[1] Q. Feng, Z.L. Liu, L.X. Xiong, et al., J. Agric. Food Chem. 58 (2010) 12327–12336.
[2] S. Treves, H. Jungbluth, F. Muntoni, F. Zorzato, Curr. Opin. Pharmacol. 8 (2008)
319–326.
[3] C.W. Taylor, D.L. Prole, T. Rahman, Biochemistry 48 (2009) 12062–12080.
[4] D.B. Sattelle, D. Cordova, T.R. Cheek, Invert. Neurosci. 8 (2008) 107–119.
[5] G.P. Lahm, D. Cordova, J.D. Barry, Bioorg. Med. Chem. 17 (2009) 4127–4133.
[6] H. Wang, M.I. Na, Z.J. Fan, N.P. Belskaia, V.A. Bakulev, J. Sichuan Normal Univ. 34
(2011) 427–434.
[7] M. Porta, A.V. Zima, A. Nani, et al., Biophysics 100 (2011) 931–938.
[8] M. Tohnishi, H. Nakao, T. Furuya, et al., J. Pestic. Sci. 30 (2005) 354–360.
[9] L.G. Philip, T.M. Stevenson, B.J. Myers, T.P. Selby, US 20000191242, 2003.
[10] T.M. Stevenson, G.P. Lahm, R.J. Pasteris, WO 03106427(A2), 2003.
[11] G.P. Lahm, T.P. Selby, WO 0248137(A2), 2003.
[12] M.M. Wang, Q.Q. Zhang, K. Yue, Q.S. Li, F.B. Xu, Chin. J. Org. Chem. 37 (2017)
1774–1780.
[13] M. Ibrahim, K. Andries, N. Lounis, et al., Agents Chem. 51 (2007)
1011–1015.
[14] H. Malipeddi, V. Malipeddi, S. Mathur, et al., Int. J. Res. Ayurveda Pharm. 1
(2010) 180–185.
[15] R.C. Reynolds, A. Tiwari, J.E. Harwell, et al., J. Med. Chem. 43 (2000) 1484–1488.
[16] Q.S. Ma, X.H. Liu, J.Q. Weng, et al., Chin. J. Org. Chem. 33 (2013) 1749–1754.
[17] R.D.C. Iii, S.B. Wold,US 5307287 (A)., 1991.
[18] Q. Chen, X.L. Zhu, L.L. Jiang, Z.M. Liu, G.F. Yang, Eur. J. Med. Chem. 39 (2010)
595–603.
Acknowledgment
[19] U. Ebbinghaus-Kintscher, K. Raming, T. Masaki, N. Yasokawa, Pflanzenschutz-
Nachrichten Bayer 60 (2007) 117–140.
[20] T. Masaki, N. Yasokawa, S. Fujioka, K. Motoba, M. Tohnishi, et al., J. Pestic. Sci. 34
(2009) 37–42.
This work was supported by the National Natural Science
Foundation of China (Nos. 20772060 and 21172114).
Please cite this article in press as: W. Liu, et al., Synthesis, bioactivity, action mode and 3D-QSAR of novel anthranilic diamide derivatives, Chin.
Chem. Lett. (2018), https://doi.org/10.1016/j.cclet.2018.05.023