Design, synthesis, and structure-activity relationship of novel aniline derivatives of chlorothalonil

Article abstract of DOI:10.1021/jf403739e

Chlorothalonil with both low cost and low toxicity is a popularly used fungicide in the agrochemical field. The presence of nucleophilic groups on this compound allows further chemical modifications to obtain novel chlorothalonil derivatives. Fluazinam, another commercially available agent with a broad fungicidal spectrum, has a scaffold of diaryl amine structure. To mimic this backbone structure, a variety of (un)substituted phenyl amines was used as nucleophilic agents to react with chlorothalonil to obtain compounds with a diphenyl amine structure. Via an elegant design, two leads, 2,4,5-trichloro-6- (2,4-dichlorophenylamino)isophthalonitrile (7) and 2,4,5-trichloro-6-(2,4,6- trichlorophenylamino)isophthalonitrile (11), with potential fungicidal activity were discovered after a preliminary bioassay screen. These two leads were further modified to obtain final products by replacing the chlorine groups in the phenyl ring in phenyl amine with other functional groups. These functional groups with various electronic properties and spatial characteristics were considered to explore the relationship between structure and fungicidal activity. The results indicate that the electron-withdrawing group NO ₂ on the 4 position on the right phenyl ring plays a unique role on enhancing the fungicidal activity. The compounds were identified by proton nuclear magnetic resonance and elemental analysis. Bioassays demonstrated that some of the title compounds exhibited excellent fungicidal activities against cucumber downy mildew at 25 mg/L. Compound 20 has been shown as the optimal structure with 85% control against cucumber downy mildew at 6.25 mg/L concentration. The relationship between structure and fungicidal activity is reported. The present work demonstrates that chlorothalonil derivatives can be used as possible lead compounds for developing novel fungicides.

Full text of DOI:10.1021/jf403739e

Article
pubs.acs.org/JAFC
Design, Synthesis, and Structure−Activity Relationship of Novel
Aniline Derivatives of Chlorothalonil
Ai-Ying Guan, Chang-Ling Liu,* Guang Huang, Hui-Chao Li, Shu-Lin Hao, Ying Xu, and Zhi-Nian Li
State Key Laboratory of the Discovery and Development of Novel Pesticide, Shenyang Research Institute of Chemical Industry
Company, Limited, Shenyang 110021, People’s Republic of China
S
* Supporting Information
ABSTRACT: Chlorothalonil with both low cost and low toxicity is a popularly used fungicide in the agrochemical field. The
presence of nucleophilic groups on this compound allows further chemical modifications to obtain novel chlorothalonil
derivatives. Fluazinam, another commercially available agent with a broad fungicidal spectrum, has a scaffold of diaryl amine
structure. To mimic this backbone structure, a variety of (un)substituted phenyl amines was used as nucleophilic agents to
react with chlorothalonil to obtain compounds with a diphenyl amine structure. Via an elegant design, two leads, 2,4,5-trichloro-
6-(2,4-dichlorophenylamino)isophthalonitrile (7) and 2,4,5-trichloro-6-(2,4,6-trichlorophenylamino)isophthalonitrile (11), with
potential fungicidal activity were discovered after a preliminary bioassay screen. These two leads were further modified to obtain
final products by replacing the chlorine groups in the phenyl ring in phenyl amine with other functional groups. These functional
groups with various electronic properties and spatial characteristics were considered to explore the relationship between structure
and fungicidal activity. The results indicate that the electron-withdrawing group NO2 on the 4 position on the right phenyl ring
plays a unique role on enhancing the fungicidal activity. The compounds were identified by proton nuclear magnetic resonance
and elemental analysis. Bioassays demonstrated that some of the title compounds exhibited excellent fungicidal activities against
cucumber downy mildew at 25 mg/L. Compound 20 has been shown as the optimal structure with 85% control against
cucumber downy mildew at 6.25 mg/L concentration. The relationship between structure and fungicidal activity is reported. The
present work demonstrates that chlorothalonil derivatives can be used as possible lead compounds for developing novel
fungicides.
KEYWORDS: Chlorothalonil derivatives, intermediate derivatization methods, fungicidal activities, structure−activity relationship
and/or pharmaceutical products. Another key feature of these
starting compounds is that these small molecules possess
chemically active functional groups amenable to derivatization
and usually very low cost to cut down the manufacturing cost of
the final product. Further, at the very beginning of a project,
three major factors, including low toxicity of environmentally
friendly intermediates, simple process of preparation, and novel
mechanism of action with innovated structure, need to be
systematically considered.
Chlorothalonil possesses reactive groups, Cl and CN, which
can easily be modified. Thus, chlorothalonil is able to be deri-
vatized by typical organic chemical reactions, such as nucleophile
substitution, hydrolysis, and addition reactions. On the basis of
these considerations mentioned above, we launched a
comprehensive project to contribute our efforts to modify
chlorothalonil. As a part of this project, in this paper, we report
an interesting result based on modification of one of the Cl
atoms on chlorothalonil to mimic the diaryl amine backbone
structure of fluazinam, another commercially available agent
with broad spectrum fungicidal activity. A variety of aryl amines
was used as nucleophilic agents to react with chlorothalonil to
obtain compounds with a diaryl amine structure. The detailed
INTRODUCTION
■
Chlorothalonil (2,4,5,6-tetrachloroisophthalonitrile) (Bravo) is
a well-established broad spectrum fungicide with $310 M sales
in 2011 in the agrochemical field. It is effective against fungal
diseases, such as gray mold, early and late blights, leaf spots,
anthracnose, fruit rots, rusts, and downy mildews, that threaten
numerous vegetable, small fruit, stone fruit, ornamental, turf,
and other agricultural crops with both lower manufacturing cost
and relatively low toxicity of LD50 > 10 000 mg/kg orally of
rats.1 Because of its success in crop protection, a great deal of
synthetic work has been performed, aimed at creating various
analogues of chlorothalonil.2−7 Our interest in chlorothalonil
analogues was to apply our new agrochemical discovery
approach, which we call intermediate derivatization methods,
to try to obtain novel fungicidally active compounds.
Intermediate derivatization methods use a three-pronged
approach to agrochemical discovery: common intermediate
method, terminal group replacement method, and active com-
pound derivatization method.8−10 Among these three approaches,
the terminal group replacement method has been proven as an
effective and practical method.11−16 However, the active
compound derivatization method is another strategy with big
potential to discover novel biological active compounds. This
approach requires further optimization based on existing
compounds. Usually, these existing compounds are selected
from small molecules available in nature with particularly potential
biological properties or those used in known agrochemical
Received: August 22, 2013
Revised: November 19, 2013
Accepted: November 20, 2013
Published: November 20, 2013
© 2013 American Chemical Society
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Figure 1. Overview of structure−activity relationships in aniline derivatives of chlorothalonil.
procedures are given below. The yields were not optimized, and
each target compound was identified and verified by H NMR and
elemental analyses.
syntheses, bioassays, and structure−activity relationships of
these compounds are discussed below.
1
Synthesis of Target Compounds (1−28 and 32−49).17−21
Synthesis of 2,4,5-Trichloro-6-(2-chloro-4-nitrophenylamino)-
isophthalonitrile (20, the Optimal Compound; Figures 2, 5, and
6) and General Procedure for Compounds 1−18, 22−28, 32, 34,
and 36−49. 2-Chloro-4-nitroaniline (1.30 g, 7.5 mmol) was dissolved
in 40 mL of N,N-dimethylformamide (DMF), and sodium hydroxide
(0.60 g, 15.0 mmol) was added to the solution. The solution was stirred
for 10 min, and 2,4,5,6-tetrachloroisophthalonitrile (chlorothalonil,
1.99 g, 7.5 mmol) was then added. The reaction mixture was stirred at
room temperature and monitored by thin-layer chromatography
(TLC). After completion of the reaction (5 h), the mixture was added
to 100 mL of water and extracted with ethyl acetate (3 × 200 mL). The
combined extracts were washed with brine, dried (anhydrous magnesium
sulfate), and filtered. The filtrate was evaporated, and the crude
product was purified via silica gel column chromatography, using a
1:4 (v/v) mixture of ethyl acetate and petroleum ether (boiling point
range of 60−90 °C) as the eluting solvent to obtain compound 20 as a
MATERIALS AND METHODS
■
All starting materials and reagents were commercially available and
used without further purification, except as indicated. Melting
points were determined on a Buchi melting point apparatus and are
̈
uncorrected. Proton nuclear magnetic resonance (1H NMR) spectra
were recorded with a Mercury 300 (Varian, 300 MHz) spectrometer
with deuterochloroform as the solvent and tetramethylsilane (TMS) as
the internal standard. Elemental analyses were determined on a
Yanaco MT-3CHN elemental analyzer. All plant and bacteria materials
were obtained from the Agrochemical Discovery Department at the
Shenyang Research Institute of Chemical Industry.
An overview of the synthesis of aniline derivatives of chlorothalonil
and their structure−activity relationships is presented in Figure 1.
1
yellow solid: 2.60 g (86%), mp 220−222 °C. H NMR (300 MHz,
CDCl3) δ: 8.42 (d, 1H, Ph-3-H, J = 2.7 Hz), 8.20 (dd, 1H, Ph-5-H,
4
3J = 9.0 Hz, J = 2.7 Hz), 7.07 (s, 1H, NH), 7.04 (d, 1H, Ph-6-H, J =
8.7 Hz). Anal. Calcd (%) for C14H4Cl4N4O2: C, 41.83; H, 1.00; N,
13.94. Found: C, 41.71; H, 1.05; N, 14.01.
Synthesis of 4-(2-Chloro-4-(trifluoromethyl)phenylamino)-2,5,6-
trifluoroisophthalonitrile (19; Figure 2) and General Procedure for
Compounds 21, 33, and 35. Compound 19 was prepared from
2-chloro-4-(trifluoromethyl)aniline and 2,4,5,6-tetrafluoroisophthalo-
nitrile22,23 using the same procedure as compound 20 as a yellow solid
1
with a yield of 88%, mp 122−124 °C. H NMR (300 MHz, CDCl3)
δ: 7.78 (s, 1H, Ph-3-1H), 7.59 (d, J = 9.0 Hz, 1H, Ph-5-1H), 7.24
(d, J = 9.0 Hz, 1H, Ph-6-1H, J = 8.4 Hz), 6.82 (br, 1H, NH). Anal.
Calcd (%) for C15H4ClF6N3: C, 47.96; H, 1.07; N, 11.19. Found: C,
47.89; H, 1.12; N, 11.22.
Synthesis of 3,5-Dichloro-N-(4-chlorophenyl)-4-(2,3,5-tri-
chloro-4,6-dicyanophenylamino)benzamide (30; Figure 3)
and General Procedure for Compound 29.17,21 Synthesis of
3,5-Dichloro-4-(2,3,5-trichloro-4,6-dicyanophenylamino)benzoic
Acid (Intermediate 1). Compound 28 (13.31 g, 31.0 mmol) was
dissolved in 120 mL of mixture solution of tetrahydrofuran (THF) and
water (1:4, v/v), and sodium hydroxide (2.45 g, 61.0 mmol) was
added to the solution. The solution was stirred in an oil bath at 50 °C
and monitored by TLC. After completion of the reaction (after 5 h),
Figure 2. Synthesis of compounds 1−28 and 32−37.
The general synthetic methods possessing simple reaction and
operation, high yields, and more importantly, very low synthesis cost
for compounds 1−49 are shown in Figures 2−6. Representative
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Figure 3. Synthesis of compounds 29 and 30.
Figure 4. Synthesis of compound 31.
mixture was evaporated under reduced pressure to obtain inter-
mediate 2, which was used without further purification.
Synthesis of 3,5-Dichloro-N-(4-chlorophenyl)-4-(2,3,5-trichloro-
4,6-dicyanophenylamino)benzamide (30). Intermediate 2 (0.40 g,
0.91 mmol) was added to a solution of 4-chloroaniline (0.12 g, 0.909 mmol)
and triethylamine (0.10 g, 1.00 mmol) in 50 mL of THF, and the
reaction mixture was stirred in an oil bath at 45 °C for 5 h. After
reaction completion (TLC), the reaction mixture was poured into
30 mL of saturated brine and extracted with ethyl acetate, and the
extract was dried over anhydrous magnesium sulfate and concentrated
under reduced pressure. The crude product was purified on a silica gel
column [ethyl acetate/petroleum ether (boiling point range of 60−
90 °C) = 1:3, as an eluent] to give compound 30 as a white solid:
0.46 g (92% based on intermediate 2), mp 275−276 °C.
1H NMR (300 MHz, CDCl3) δ: 10.50 (d, 1H, CONH, J = 12.9 Hz),
8.13 (dd, 2H, Ph-2,6-2H, 3J = 15.7 Hz, 4J = 1.2 Hz), 7.81 (d, 2H, 4-Cl-
Ph-3,5-2H, J = 9.0 Hz), 7.31−7.35 (m, 2H, 4-Cl-Ph-2,6-2H). Anal.
Calcd (%) for C21H8Cl6N4O: C, 46.28; H, 1.48; N, 10.28. Found: C,
46.33; H, 1.41; N, 10.26.
Synthesis of 2,4,5-Trichloro-6-(2,6-difluoro-4-nitrophenyl-
amino)isophthalonitrile (31; Figure 4).17,21 Intermediate 2,4,5-
trichloro-6-(2,6-difluorophenylamino)isophthalonitrile was prepared
from 2,6-difluoroaniline and 2,4,5,6-tetrachloroisophthalonitrile using
the same procedure as compound 20 as a yellow solid with a yield
of 83%.
Figure 5. Synthesis of compounds 38 and 39.
To the mixture of 2,4,5-trichloro-6-(2,6-difluorophenylamino)iso-
phthalonitrile (0.68 g, 2.0 mmol) in concentrated sulfuric acid
(20 mL) was added fuming nitric acid (d = 1.52, 10 mL) dropwise for
20 min with sufficient stirring. After stirring at room temperature for
1 h, the reaction mixture was poured into ice water, and the resulting
precipitate was collected by filtration and washed with water to obtain
compound 31 as a pale white solid, mp 204−206 °C. 1H NMR
Figure 6. Synthesis of compounds 40−49.
the mixture was added to 500 mL of water and extracted with ethyl
acetate (3 × 500 mL). The pH of the aqueous phase was adjusted to 5−6
with dilute hydrochloric acid, and the solid was filtered (intermediate 1)
and used without further purification.
Synthesis of 3,5-Dichloro-4-(2,3,5-trichloro-4,6-dicyanophenylamino)-
benzoyl Chloride (Intermediate 2). Intermediate 1 (5.54 g, 12.72 mmol)
was dissolved in 100 mL of petroleum ether followed by 2 drops of
DMF, and then sulfurous dichloride (2.27 g, 19.08 mmol) was added
to the solution. The solution was refluxed in an oil bath at 85 °C
and monitored by TLC. After completion of the reaction (2 h), the
3
(300 MHz, CDCl3) δ: 7.97−8.01 (dd, 2H, Ph-3,5-2H, J = 10.8 Hz,
4J = 3.0 Hz), 6.70 (s, 1H, NH). Anal. Calcd (%) for C14H3Cl3F2N4O2:
C, 41.67; H, 0.75; N, 13.88. Found: C, 41.71; H, 0.81; N, 13.81.
Fungicidal Assay. Each of the test compounds (4 mg) was first
dissolved in 5 mL of mixture of acetone and methanol (1:1 by
volume), and then 5 mL of water containing 0.1% Tween 80 was
added to generate a 10 mL stock solution of 400 mg/L concentration.
Serial test solutions were prepared by diluting the above solution
(testing range of 6.25−400 mg/L).
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Table 1. Chemical Structures and Fungicidal Activity of Aniline Derivatives of Chlorothalonil (Compounds 1−15)
biological activity against CDM
(% control at the given concentration in mg/L)
compound
R1
R2
R3
R4
R5
400
100
50
25
12.5
6.25
1
2
3
4
5
6
H
H
H
H
H
Cl
H
Cl
H
H
Cl
H
Cl
H
Cl
H
H
Cl
H
H
H
H
H
H
H
Cl
Cl
H
H
H
H
Cl
Cl
0
0
a
a
a
a
a
a
a
a
a
a
a
a
a
a
a
a
a
a
a
a
Cl
H
H
H
H
H
0
a
a
a
H
Cl
H
H
0
a
a
a
Cl
Cl
Cl
H
H
100
80
35
a
0
a
a
H
Cl
H
a
20
a
a
a
7 lead number 1
Cl
H
100
60
100
a
a
a
8
H
a
a
9
Cl
H
H
H
100
85
0
a
a
a
10
Cl
Cl
Cl
CH3
Cl
Cl
H
a
a
a
a
11 lead number 2
Cl
Cl
CH3
Cl
H
Cl
H
98
95
a
75
a
15
a
a
12
80
a
13
CH3
H
80
a
a
a
a
14
100
85
0
a
a
a
15
H
a
a
a
a
chlorothalonil
fluazinam
100
100
100
100
80
100
30
100
a
a
100
90
a
No data.
Table 2. Chemical Structures and Fungicidal Activity of Aniline Derivatives of Chlorothalonil (Compounds 16−23)
biological activity against CDM (% control at the given concentration in mg/L)
compound
R
R1
Cl
R2
Cl
400
100
50
25
12.5
6.25
7 lead number 1
Cl
Cl
Cl
Cl
F
100
100
100
100
100
100
100
100
100
100
100
100
0
20
a
a
a
a
a
a
a
16
CH3
NO2
Cl
Cl
17
Cl
70
30
98
98
98
80
80
a
10
a
a
18
CF3
CF3
NO2
NO2
NO2
NO2
100
100
100
95
80
80
98
40
60
a
20
a
a
19
Cl
a
20 optimal structure
Cl
F
Cl
98
a
85
a
21
Cl
22
Cl
Cl
NO2
CN
95
a
a
23
a
a
a
chlorothalonil
fluazinam
100
100
80
100
30
100
a
100
90
a
No data.
Evaluations of fungicidal activities of the synthesized compounds
The test results of the fungicidal activities of compounds 1−49
against CDM are listed in Tables 1−5.
against cucumber downy mildew (CDM) were performed as follows:
Briefly, a whole plant is used in this test, and the testing solution is
sprayed to the host plant by a special plant sprayer. The plant is
inoculated with fungus after 24 h. According to the infecting char-
acteristics of fungus, the plant is stored in a humidity chamber and
then transferred into a greenhouse after infection is finished. The other
plants are placed in a greenhouse directly. The activity of each
compound was estimated by visual inspection after 7 days, and
screening results were reported as a range from 0% (no control) to
100% (complete control).
RESULTS AND DISCUSSION
■
Synthesis. According to the schemes shown in Figures 2−6,
49 title compounds were synthesized with yields of 50−95%, as
shown in Tables 1−5. The synthesized compounds were char-
1
acterized by H NMR and elemental analyses. All spectral and
analytical data were consistent with the assigned structures.
Structure−Activity Relationship (SAR). Discovery of
Lead Compounds. Initially, chlorothalonil was reacted with
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Table 3. Chemical Structures and Fungicidal Activity of Aniline Derivatives of Chlorothalonil (Compounds 24−37)
biological activity against CDM
(% control at the given concentration in mg/L)
compound
R
R1
Cl
R2
R3
Cl
R4
R5
400
100
50
25
12.5
6.25
11 lead number 2
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
F
Cl
H
H
H
H
H
H
H
H
H
H
H
H
H
Cl
Cl
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
Cl
H
98
100
100
50
95
70
100
a
75
40
100
a
15
20
75
a
a
a
a
a
a
a
24
Cl
Cl
Cl
Br
Cl
Cl
Cl
F
Br
Cl
Cl
Cl
Br
Cl
Cl
Cl
F
25
NO2
CF3
OCF3
a
26
a
27
85
a
a
a
a
28
CO2CH3
CONHPh
CONH(4-Cl-Ph)
NO2
20
a
a
a
a
29
98
0
a
a
a
30
55
a
a
a
a
31
100
100
100
100
100
100
100
100
100
100
95
100
98
100
0
100
98
70
100
90
0
98
95
30
100
85
a
70
50
a
40
40
a
32
Cl
Cl
Br
Br
CH3
Cl
Cl
NO2
F
33
NO2
F
34
Cl
F
NO2
Br
Br
NO2
CN
H
30
a
a
35
NO2
a
36
Cl
Cl
Cl
NO2
a
a
37
CN
a
a
a
a
20 optimal structure
NO2
100
98
98
98
85
a
No data.
modify these two leads to obtain compounds with better
fungicidal activity. More importantly, we obtained very useful
structure−activity information based on the preliminary results;
namely, substitution at the 2,4 and 2,4,6 positions of the right
phenyl ring may be key to improving fungicidal activity of the
whole molecule compared to other substituted positions. We
next considered varying the electronic properties and spatial
characteristics of the substituent groups at these positions.
These changes are described below, and data are presented in
Tables 2 and 3.
Table 4. Chemical Structures, Physical Properties. and
Fungicidal Activity of Cyano Isomers of Compound 25
(Compounds 38 and 39)
biological activity against CDM
(% control at the given concentra-
tion in mg/L)
Optimization of Compound 7. Using compound 7 as a
starting point for additional analogues, we turned our attention
to replacing the Cl atoms on the 2 and 4 positions of the right
side phenyl ring with other electron-donating groups, such as
CH3, and/or electron-withdrawing groups, such as CO2CH3,
CF3, NO2, and CN (Table 2). First, we varied substituents on
the 2 position and kept the 4 position fixed as Cl. We syn-
thesized two compounds 16 with an electron-donating group
(CH3) and 17 with an electron-withdrawing group (NO2),
respectively. The bioassay results showed that compound 16
was less efficacious than lead compound 7 (0 versus 100% at
100 mg/L), indicating that the electron-donating group has a
negative effect on bioactivity. In contrast, the fungicidal activity
of compound 17 increased moderately compared to the lead
compound 7 (30 versus 20% at 50 mg/L), implying that the
electron-withdrawing group is helpful for enhancing activity.
Then, we varied the 4 position and kept the 2 position fixes
as Cl. We synthesized two compounds with electron-with-
drawing groups at the 4 position, compound 18 with a CF3
group and compound 20 with a NO2 group (Table 2). To our
surprise, compound 18 exhibited 80% control at 25 mg/L, and
compound 20 gave 98% control at 25 mg/L and 85% control at
6.25 mg/L. Both of these compounds were more efficacious
than chlorothalonil, which only showed 30% control of CDM
at 25 mg/L. Furthermore, to investigate if the fungicidal activity
position of
compound
double CNs
400
100
50
25
12.5
38
39
25
2,3 position
1,4 position
1,3 position
100
100
100
100
100
100
100
100
95
100
100
98
90
100
75
0
20
a
20 optimal structure
98
98
a
No data.
aniline to give compound 1, which did not show any control of
CDM (Table 1). Then, 9 compounds (compounds 2−10) were
synthesized to evaluate the effect of the substituent position of
R1, R2, R3, R4, and R5 on fungicidal activity using the chlorine
atoms as probes. When a single chlorine was introduced into
any position of the right phenyl ring (compounds 2−4), there
was no improvement in fungicidal activity. Next, we synthesized
dichloro (compounds 5−10) and trichloro (compounds 11−15)
analogues. Fungicidal assays identified two lead compounds,
compound 7 with 2,4-Cl2 substituents (R1 = R2 = Cl, and
R3 = R4 = R5 = H) and compound 11 with 2,4,6-Cl3 sub-
stituents on the right phenyl ring, which displayed 100 and 95%
control, respectively, against CDM at 100 mg/L. Although
when screened at the lower concentration of 50 mg/L,
compounds 7 and 11 showed lower fungicidal activity than
the commercial fungicide chlorothalonil (20 and 75%, respectively,
versus 80% for chlorothalonil), we were encouraged to further
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Table 5. Chemical Structures and Fungicidal Activity of Heterocyclic Amine Derivatives of Chlorothalonil (Compounds 40−49)
biological activity against CDM (% control at the given concentration in mg/L)
compound
Het
pyridin-2-yl
400
100
50
25
12.5
6.25
40
41
42
43
44
45
46
47
48
49
0
30
0
a
a
a
a
a
a
a
a
a
a
a
a
a
a
a
a
a
a
a
a
a
a
a
a
a
a
5-Br-pyridin-2-yl
pyridin-3-yl
a
a
a
6-Br-pyridin-3-yl
2-Cl-pyridin-4-yl
pyrimidin-2-yl
80
90
0
a
a
a
80
a
30
a
10
a
4,6-2CH3-pyrimidin-2-yl
4,6-2OCH3-pyrimidin-2-yl
6-Cl-pyrazin-2-yl
6-Cl-pyridazin-3-yl
0
a
a
a
0
a
a
a
98
100
0
a
a
0
a
a
a
No data.
could be improved further when both 2- and 4-Cl atoms were
substituted by electron-withdrawing groups, compounds (com-
pound 22 with 2-NO2 and 4-NO2 and compound 23 with 2-CN
and 4-NO2) were designed and synthesized. However, the
fungicidal activity results showed that neither compound was as
effective as the lead compound 20. The result of the optimiza-
tion of compound 7 is identification of compound 20 with a
2-Cl-4-NO2 group as the optimized structure with greatly improved
fungicidal activity.
properties of the fluorine atom, such as high thermal stability
and lipophilicity,25 further optimization was conducted by
introducing fluorine into the 2 and/or 6 position with the 4
position fixed as NO2 because the NO2 group was demon-
strated to be beneficial in compound 25. To our excitement, as
we expected, these two compounds (31 with 2,6-F2 and 32
with 2-Cl-6-F) had much higher activity than compound 25
(2,6-Cl2). Compound 31 gave 98% control at 25 mg/L
compared to 75% control shown by compound 25, while
compound 32 gave 95% control. When the 2,6-Cl2 atoms in
compound 11 were replaced by Br atoms (compound 34),
fungicidal activity was similar to compounds 31 and 32. With
continuing interest to induce fluorine into the lead compound,
we turned our attention to replacing all Cl atoms of the left
side ring with F atoms, resulting in compounds 33 and 35
(Table 3). However, these two compounds showed lower
fungicidal activity than their corresponding Cl analogues.
The results suggest that it is not always true that the more
fluorine atoms, the better bioactivity. Further, the position of
the fluorine atom in the intact molecule may also play a crucial
role in its bioactivity.
Finally, two compounds 36 and 37 were prepared to evaluate
the effect of including additional substituents in the right side
ring on fungicidal activity (Table 3). The results show that
neither compound 36 nor compound 37 was as efficacious as
lead compound 25. The result of the optimization of compound
11 is identification of compound 31 with a 2,6-F2-4-NO2 group
as the optimized structure with greatly improved fungicidal
activity.
Activity of Cyano Isomers of Compound 25. To deter-
mine if the cyano isomers of compound 25 would show improved
fungicidal activity, we synthesized and screened compounds 38
and 39 (Table 4). Both compounds 38, which gave 90%
control at 25 mg/L, and 39, which showed 100% control at
25 mg/L, were more efficacious than compound 25, which gave
75% control at 25 mg/L.
Optimization of Compound 11. Using compound 11 as a
starting point for additional analogues, we turned our attention
to replacing the Cl atoms on the 2, 4, and 6 positions of the
right side phenyl ring (Table 3). First, we varied substituents on
the 4 position while maintaining the 2 and 6 positions as Cl or
Br. These substituents are typical groups with both electronic
effect and spatial effect simultaneously. While the 4-Br analogue
(compound 24) showed lower fungicidal activity than lead
compound 11 (40 versus 75%, respectively, at 50 mg/L), the
NO2 analogue (compound 25) gave improved activity
(100 versus 75%, respectively, at 50 mg/L). At a 25 mg/L
dose, a larger difference in activity between compounds 25 and
11 was observed (75 versus 15%, respectively). However,
when the 4-Cl atom of compound 11 was replaced with CF3
(compound 26), OCF3 (compound 27), or CO2CH3
(compound 28), weaker fungicidal activity resulted. On the
basis of these observations, the electron-withdrawing group
NO2 rather than CF3, OCF3, and CO2CH3 plays an important
role in enhancing the fungicidal activity. It was a surprising
result that CF3 and OCF3 at the 4 position did not exhibit
excellent bioactivity because it has been shown that CF3 usually
provides a very strong positive contribution to biological
activities.24 In our case, this may be because the 4 postion is not
an optimum location for these fluorine-containing groups. Further-
more, when large spatial groups, such as CONHPh and
CONH(4-Cl-Ph), were induced into the 4 position of lead
compound 11, compounds 29 and 30 exhibited reduced effect
compared to the lead compound 11 as well. A possible explana-
tion for the lower activity associated with large subsitutuents is
that large substituents would block the interaction of the target
enzyme and these bulky compounds.
Activity of Heterocyclic Amino Analogues of Chlor-
othalonil. To determine if replacing the right side phenyl ring
with a nitrogen heterocycle would enhance fungicidal activity in
this class of compounds, we prepared a series of 10 heterocyclic
amino analogues of chlorothalonil, which were derived from
pyridine, pyrimidine, and pyrazine heterocycles (Table 5).
Considering that many fluorine-containing compounds
exhibit significant agricultural bioactivities, owing to the unique
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The only analogue that showed any appreciable activity was com-
pound 44, in which the heterocyclic group was 2-Cl-pyridin-4-yl.
The poor results from these analogues suggest that introduction
of nitrogen in the aromatic ring was detrimental to the fungicidal
activity in this class of compounds.
On the basis of data presented in Tables 1−5, a clear-cut,
well-defined relationship between the chemical structure and
biological activity has taken shape by examining the effect of
different kinds of electron-withdrawing, electron-donating, and
spatially demanding groups on the fungicidal activity of aniline
derivatives of chlorothalonil. 2,4-Disubstituted aniline deri-
vatives, especially compound 20, which possesses a 2-chloro-
4-nitro substitution in the right side phenyl ring, showed
improved fungicidal activity compared to chlorothalonil. 2,4,6-
Trisubstituted aniline derivatives, especially compound 25,
which possesses a 2,6-Cl2-4-NO2 substitution in the right side
phenyl ring, were more efficacious than chlorothalonil. Cyano
isomers of compound 25, namely, compounds 38 and 39, were
slightly more efficacious than compound 25, while nitrogen
heterocyclic amine derivatives were very weak or inactive.
From what has been discussed above, we can draw the
conclusion that compound 20 derived from chlorothalonil with
a simple process and relative low cost, is the optimal structure
with desired activity. It offers a control of 85% against CDM at
6.25 mg/L concentration, much higher than chlorothalonil and
nearly equal to fluazinam. Compound 20, which has also shown
activity against rice blast and gray mold, besides CDM,21 is a
promising candidate for further development. This study
demonstrates the effectiveness of our intermediate deriva-
tization method approach to the discovery of bioactive com-
pounds. Further synthesis of analogues, structure optimization
studies, and field trials of compound 20 are in progress.
ASSOCIATED CONTENT
■
S
* Supporting Information
1H NMR and melting point data for compounds 2, 3, 5−18,
21−29, and 32−49. This material is available free of charge via
the Internet at http://pubs.acs.org.
AUTHOR INFORMATION
Corresponding Author
*Telephone: 86-24-85869078. Fax: 86-24-85869137. E-mail:
liuchangling@vip.163.com.
■
Funding
The project was supported by the National Key Technology
Support Program during the 12th Five-Year Plan Period of
China (Grant 2011BAE06B05).
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Notes
The authors declare no competing financial interest.
ACKNOWLEDGMENTS
■
We thank Dr. Xiaoping Yang at the University of Colorado and
Dr. Mark Dekeyser (Canada) for assistance with manuscript
preparation.
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