Discovery of bis-aromatic ring neonicotinoid analogues fixed as cis-configuration: Synthesis, insecticidal activities, and molecular docking studies

Article abstract of DOI:10.1016/j.bmcl.2010.04.050

A novel series of bis-aromatic ring neonicotinoid analogues (1a-1l, 2a-2c), were designed and prepared by introducing a new substituted aromatic ring into nitenpyram and forming a tetrahydropyrimidine ring, with the cis-configuration confirmed by X-ray diffraction. Preliminary bioassays showed most analogues exhibited good insecticidal activities at 100 mg/L, and compound 1d and 2a were highly potent even at 10 mg/L. Modeling the ligand-receptor complexes by molecular docking study explained the structure-activity relationships observed in vitro, which may provide some useful information for future design of new insecticides.

Full text of DOI:10.1016/j.bmcl.2010.04.050

Bioorganic & Medicinal Chemistry Letters 20 (2010) 3301–3305
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Discovery of bis-aromatic ring neonicotinoid analogues fixed
as cis-configuration: Synthesis, insecticidal activities, and molecular
docking studies
a
a
a
a
b
Chuanwen Sun a, , Jia Jin , Jun Zhu , Haifeng Wang , Dingrong Yang , Jiahua Xing
*
a College of Life and Environment Sciences, Shanghai Normal University, Shanghai 200234, China
b Bioassay department, Branch of National Pesticide R&D South Center, Hangzhou 310023, China
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 31 January 2010
Revised 5 April 2010
Accepted 12 April 2010
Available online 14 April 2010
A novel series of bis-aromatic ring neonicotinoid analogues (1a–1l, 2a–2c), were designed and prepared
by introducing a new substituted aromatic ring into nitenpyram and forming a tetrahydropyrimidine
ring, with the cis-configuration confirmed by X-ray diffraction. Preliminary bioassays showed most
analogues exhibited good insecticidal activities at 100 mg/L, and compound 1d and 2a were highly potent
even at 10 mg/L. Modeling the ligand–receptor complexes by molecular docking study explained the
structure–activity relationships observed in vitro, which may provide some useful information for future
design of new insecticides.
Keywords:
Bis-aromatic ring
Nitenpyram
Ó 2010 Elsevier Ltd. All rights reserved.
Docking model
Insecticidal
cis-Configuration
The discovery of neonicotinoid insecticides (NNs) can be
considered as a milestone in agrochemical research over the past
three decades. NNs have recently received global interests from
both agricultural chemistry and medicinal fields, due to their broad
spectrum of biological activities and high selectivity for public
health applications1 and crop protection. In 2006, the neonicoti-
noid family accounted for worldwide annual sales of around $US
1.56 billion, representing nearly 17% of the global insecticide
market.2 Neonicotinoids represented by imidacloprid3 act on the
insect nicotinic acetylcholine receptors (nAChRs), with a much
lower affinity for the mammalian nAChR,4,5 which makes this class
of insecticides particularly attractive to the primarily controlled
sap-feeding pests6–8 and relatively safe toward mammals.9,10
However, during the past decades, frequent field applications
have inevitably led to insects resistance to the major class of
NNs.11–13 Significant increases in cross-resistance have also been
observed in a range of species,14–18 with more recently collected
strains exhibiting more than 100-fold resistance to imidacloprid
and comparable levels of resistance to thiamethoxam and acetam-
iprid.19,20 Therefore, the development of new potent neonicoti-
noids with novel chemical structures and low resistance is of an
urgent desire.
Until recently, most of the researches about structure optimiza-
tion of NNs are based on cyclic neonicotinoid insecticides, such as
imidacloprid. However, few studies have been focused on the
structural modification of acyclic NNS, such as nitenpyram and
acctamiprid. Encouraged by this, we developed a new structure
developing strategy, using nitenpyram as the lead compound
(Fig. 1), and a series of novel substituted-1,2,3,6-tetrahydropyrim-
idine derivatives were designed by introducing a new substituted
aromatic ring into nitenpyram and forming a tetrahydropyrimidine
ring fixed as cis-configuration. As a result, most of these analogues
exhibited significant insecticidal activities against Nilaparvata
legen, and some of them had >90% mortality at 10 mg/L. This paper
describes the syntheses, bioactivities and the structure–activity
relationship. The interactions between these new compounds with
S
NO2
( )
O2N
N
Ar
n
N
N
N
H
N
Cl
N
Cl
N
Nitenpyram
s
=H,CH3,COOC2H5 n=1~2
Ar=aryl,cycloalkyi
* Corresponding author. Tel./fax: +86 021 64321926.
E-mail addresses: willin@shnu.edu.cn, sunwillin6@shnu.edu.cn (C. Sun).
Figure 1. Chemical structures of the target compounds.
0960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.04.050

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C. Sun et al. / Bioorg. Med. Chem. Lett. 20 (2010) 3301–3305
Scheme 1. Preparation of substituted 1,2,3,6-tetrahydropyrimidines (1a–1l, 2a–2c). Reagents and conditions: (a) ethanamine (42%); (b) 1,1,1-trichloro-2-nitroethane/CHCl3
2–7 °C (65%); (c) methanamine 3–7 °C (58%); (d) various substituted amine, 37% HCHO, EtOH, 25 °C (65–80%); (e)
Et3N/EtOH, 70 °C, refluxing (71–78%).
L-a-amino acid ethyl ester hydrochloride, 37% HCHO,
Figure 2. Molecular structure of compound 1d (CCDC number 746864) with atom-labeling and the packing diagram.
Cl
Cl
N
N
N
N
N
N
O
N
O
N
N
O
N
O
ii
i
Scheme 2. Resonance of the structures i/ii.

C. Sun et al. / Bioorg. Med. Chem. Lett. 20 (2010) 3301–3305
3303
Table 1
their target nAChRs were also investigated by molecular docking,
which may provide some useful information for future design of
new insecticides.
Insecticidal activities of the target compounds (1a–1l, 2a–2c) against Nilaparvata
legen
Synthesis procedures for these new substituted-1,2,3,6-tetrahy-
dropyrimidine neonicotinoid analogues are summarized in Scheme
1. Nitenpyram, which was prepared based on the procedures pre-
viously reported,21 was reacted with various substituted amine
and formaldehyde in acetate acid, affording desired compounds
1a–1l with cis-configuration fixed by 1,2,3,6-tetrahydro-pyrimi-
dine. Meanwhile, compounds 2a–2c with cis-configuration were
also obtained by Mannich reaction. The mixture of Nitenpyram,
Compd
R
Concn.a (mg/L)
100
500
10
L
-a-amino acid ethyl ester hydrochlorides and formaldehyde was
1a
1b
++++++
+++++
+++
stirred for 30 min in a microwave reactor, which may speed up
reactions with good yields. The structures of the target compounds
were well characterized by 1H NMR, MS, EA, IR, and elemental
analysis.
++++++
++++++
++++++
++++
++
F
OCH3
To further obtain precise three-dimensional structural informa-
tion, compound 1d was recrystallized by slow evaporation from
mixed solution of compound 1d in ethyl acetate/petroleum ether
(v:v = 1:4), and its single-crystal structure was confirmed by
X-ray crystallographic data analysis (Fig. 2).22 The crystal data have
been listed in the Supplementary data associated with this Letter.
Due to the lone-pair electrons transferring on the amines to
C(9)@C(10), the C9–N2 and C9–N3 bond lengths, both 1.36 Å, are
remarkably shorter than typical C–N single bond (1.47 Å) but close
1c
++++++
1d
1e
++++++
++++++
++++++
+++
+++++
+
1f
+++++
+
n.t.b
to C@N (imine) (1.33 Å).23 A coplanar olefin–amine
p-electron
OCH3
network (Scheme 2) is formed, owing to the delocalization of the
electrons which extend to the strong electron-withdrawing group,
–NO2. In addition, the bond lengths of C9@C10 and C10–N5 are
1.40 and 1.39 Å, which are longer than that of pure C@C (1.34 Å)
and shorter than that of typical C–N in C–NO2 (1.49 Å). In the sym-
metric unit, an intramolecular hydrogen bond (C3–H3Á Á ÁO1) form-
ing a nine-membered ring (O1, N5, C10, C9, N2, C6, C4, C3, H3) was
observed, and two intermolecular hydrogen-bonds are found be-
tween molecules (Fig. 2b), forming a three-dimensional interaction
network, which may help stabilize the crystal structure. Besides,
The most remarkable feature of this molecule is the cis-configura-
tion of its nitro group, and the preference for this geometry is
mostly due to the newly introduced tetrahydropyridine ring that
fixes the nitro moiety in the cis position. Since most of the crystal
structures of neonicotinoid insecticides were determined with
trans-configuration, which all the studies of mechanism were
based on, this new unique cis-configuration may endue these
new compounds with a novel mode of action.
The insecticidal activities of these novel analogues were evalu-
ated against Nilaparvata legen.24 As depicted in Table 1, most com-
pounds exhibited good in vitro insecticidal activities against
Nilaparvata legen and had >80% mortality at 100 mg/L. Among
these compounds, 1d and 2a showed significant potency that is
close to nitenpyram at 10 mg/L, and the bioactivities of (1b, 1g,
1j, and 1k) were slightly weaker than that of nitenpyram. Com-
pared with 1a, the introduction of cycle alkyl (1l) in stead of aro-
matic groups showed decreasing tendency in insecticidal activity.
Interestingly, the introduction of F and Cl elements into the aro-
matic groups resulted in increased insecticidal activity (1a vs 1b,
1i vs 1j), and compounds containing phenyl displayed higher
inhibitory activity than the corresponding derivatives with hetero-
cycle such as pyridinyl (1d vs 1i).
1gc
++++++
++++++
++++
1hd
+++++
++++
+++
1i
1j
++++++
++++++
++++
+++
N
++++++
++++
N
Cl
1k
1l
++++++
+++++
++++++
++++
++++
++
OC2H5
2a
++++++
++++++
+++++
O
i-C4H9
OC2H5
2bd
+++
+
À
O
2cd
++++
+++
++
O
C2H5O
Nitenpyramb
++++++
++++++
++++++
a
Rating system for the mortality percentage: ++++++, 100%; +++++, ꢀ90%; ++++,
ꢀ80%; +++, ꢀ70%; ++, ꢀ60%; +, ꢀ50%; À, <50%.
b
n.t. = not tested.
c
(+)-.
( )-.
d
For the effects of the flexible linkage at 1-position of tetrahydro-
pyrimidine ring, the insecticidal activities of the corresponding
analogues decreased in the order benzyl (1d) > phenylethyl
(1k) > phenyl (1a), and the introduction of a large ester group into
the linkage resulted in remarkably decreased insecticidal activity
(2c vs 1k). These observations clearly suggested that the length
and flexibility of the linkage was strongly related to their insecti-
cidal potency.
To further explore the structural features for better activities,
models of these new compounds–receptor complexes were inves-
tigated by docking studies with AutoDock version 4.0.25 Since the
amino acids forming the active sites are both structurally and func-
tionally consistent in the diverse nAChRs and AchBPs, the pub-
lished crystal structure of a Lymnaea stagnalis-AChBP (Ls-AChBP)
co-crystallized with imidacloprid (PDB ID: 2zju)26 was used as

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C. Sun et al. / Bioorg. Med. Chem. Lett. 20 (2010) 3301–3305
Figure 3. Modeling of the docking results of compounds 1d, 2a in the extracellular domain of nAChR. (a) 2a nestled in the interfacial agonist-binding pocket between the
(+)-face (primary, yellow) and (À)-face (complementary, blue) subunits of the Ls-AChBP (PDB ID: 2zju), as a structural surrogate of the insect nAChR; (b) nAChR-2a binding
site interactions (zoomed-in) featuring hydrogen-bonding between 2a and the active site residues; (c) nAChR-1d interactions. The second structure of the protein is
represented as line ribbon.
the template of receptor. The docking was carried out through the
graphical user interface AUTODOCKTOOLS (ADT 1.4.6). The only modifi-
cation was the number of docking runs that was set to 200 (previ-
ously 100) for more accuracy.
determined by X-ray diffraction, with the cis-configuration of its
C9@C10 confirmed. In addition, molecular docking studies were
also carried out to model the ligand-nAChR complexes and analyze
their interactions for improved activity. The docking results re-
vealed a unique binding mode other than nitenpyram, and were
in good agreement with their high insecticidal potential, which
also explained the structure–activity relationships observed
in vitro. Further researches are underway to verify the nAChR tar-
get and evaluate their inhibitory activities against resistant insect
species. The study herein has shed a light on the mechanism of ac-
tion of these bis-aromatic ring neonicotinoid analogues, thereby
prompting some useful information for future design of new NNs
pesticides.
As a result, the scoring function of the docking program ranked
the compounds in the same general order observed experimentally
(data not shown), and all active analogues exhibited significant
hydrogen-bonding interactions with the nAChR target. As ex-
pected, the most potent compound 2a is nicely accommodated
within the subunit interfacial binding pocket between the two
faces of adjacent subunits (Fig. 3a). Its binding conformation exhib-
ited important hydrogen bond between its nitro O21 and H–O of
Tyr192 (O–HÁ Á ÁN: 2.85 Å, 138.6°) (Fig. 3b), and the O24 of its ester
group hydrogen-bonds the side chain O of Gln55, while the chloro-
pyridine interacts primarily with Arg104. Other interactions in the
active site region may be mediated via water(s), since these resi-
dues are near the surface of the receptor. In addition, analogue
1d binds to the nAChR with comparable affinity to 2a (Fig. 3c),
which is consistent with its high insecticidal activity. Besides,
important additional hydrophobic interactions have been found
between the side chain of Leu112 and its phenyl, which is newly
introduced to these bis-aromatic ring neonicotinoid analogues.
These observations have also explained the structure–activity rela-
tionships observed in vitro.
Acknowledgments
This research was supported by the National Natural Science
Foundation of China (No. 20672073), Innovation Program of
Shanghai Municipal Education Commission (09YZ157 and
ssd08013), and Leading Academic Discipline Project of Shanghai
Normal University (DZL808).
Supplementary data
Furthermore, most of the other active derivatives (1b, 1g, 1j, 1k)
shared a quite similar binding mode with 2a, and many of them
exhibited more than two hydrogen-bonds with different amino
acids of the active pocket between the nAChR subunits (unpub-
lished result). These amino acids (Gln73, Trp 143, Cys187,
Cys188, and Glu190) were different from the ones that interacted
with imidacloprid, which suggested a novel mechanism of the
insecticidal effect by these new compounds. Thereby, the newly
introduced substituents of the designed analogues presumably
played important roles in ligand recognition and binding interac-
tions, which may further enhance their activities and contribute
to the selectivity as well. Based on these, further target inhibitory
tests and advanced insecticide design are underway.
In summary, a series of novel bis-aromatic ring neonicotinoid
analogues, which were designed based on the acyclic NNS
(nitenpyram), were synthesized and tested for their insecticidal
activity against Nilaparvata legen. All the target compounds
presented good insecticidal activity at 100 mg/L. Among these ana-
logues, 1d and 2a afforded the best in vitro activity, and had >90%
mortality at 10 mg/L. The single-crystal structure of 1d was further
Supplementary data (experimental details include full synthesis
procedures, analytical characterization of all the compounds and
the crystallographic data of compound 1d) associated with this
article can be found, in the online version, at doi:10.1016/
j.bmcl.2010.04.050.
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