Optimization, Structure-Activity Relationship and Mode of Action of

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Agricultural and Environmental Chemistry

Optimization, Structure-Activity Relationship and Mode of Action of Nortopsentin Analogues Containing Thiazole and Oxazole Moieties Jincheng Guo, Yanan Hao, Xiaofei Ji, Ziwen Wang, Yuxiu Liu, Dejun Ma, Yongqiang Li, Huailin Pang, Jueping Ni, and Qingmin Wang J. Agric. Food Chem., Just Accepted Manuscript • DOI: 10.1021/acs.jafc.9b04093 • Publication Date (Web): 26 Aug 2019 Downloaded from pubs.acs.org on August 26, 2019

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Page 1 of 51

Journal of Agricultural and Food Chemistry

Optimization, Structure-Activity Relationship and Mode of Action of Nortopsentin Analogues Containing Thiazole and Oxazole Moieties

Jincheng Guo†,‡, Yanan Hao‡, Xiaofei Ji‡, Ziwen Wang†,*, Yuxiu Liu‡,*, Dejun Ma‡, Yongqiang Li‡, Huailin Pang§, Jueping Ni§, Qingmin Wang‡,*

†Tianjin

Key Laboratory of Structure and Performance for Functional Molecules, MOE Key

Laboratory of Inorganic–Organic Hybrid Functional Material Chemistry, College of Chemistry, Tianjin Normal University, Tianjin 300387, China; ‡State

Key Laboratory of Elemento-Organic Chemistry, Research Institute of Elemento-Organic

Chemistry, College of Chemistry, Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), Nankai University, Tianjin 300071, China; §CAC

Nantong Chemical Co. Ltd, Shanghai 226400, China

* To whom correspondence should be addressed. For Ziwen Wang, E-mail: [email protected]; Phone: 0086-22-23766531; Fax: 0086-22-23766531; For Yuxiu Liu, E-mail: [email protected]; Phone:

0086-22-23503792;

Fax:

0086-22-23503792;

For

Qingmin

[email protected]; Phone: 0086-22-23503952; Fax: 0086-22-23503952.

1

ACS Paragon Plus Environment

Wang,

E-mail:


1 2 3

ABSTRACT: Plant diseases seriously endanger plant health, it is very difficult to control

4

them. A series of nortopsentin analogues were designed, synthesized and evaluated for

5

their antiviral activities and fungicidal activities. Most of these compounds displayed

6

higher antiviral activities than ribavirin. Compounds 1d, 1e and 12a with excellent

7

antiviral activities emerged as novel antiviral lead compounds, among which, 1e was

8

selected for further antiviral mechanism research. The mechanism research results

9

indicated that these compounds may play an antiviral role by aggregating viral particles

10

to prevent their movement in plants. Further fungicidal activity tests revealed that

11

nortopsentin analogues displayed broad-spectrum fungicidal activities. Compounds 2p

12

and 2f displayed higher antifungal activities against Alternaria solani than commercial

13

fungicides carbendazim and chlorothalonil. Current research has laid a foundation for the

14

application of nortopsentin analogues in plant protection.

15 16 17

KEYWORDS: structure optimization, nortopsentin analogues, anti-TMV activity,

18

fungicidal activity, mode of action

19 20 21 2


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22

INTRODUCTION

23

As the population continues to grow, the food problem will once again become a

24

rigid demand.1-5 Among the many factors contributing to the food shortage, plant diseases

25

which reduce crop yield cannot be ignored.6 Tobacco mosaic virus (TMV) is the earliest

26

and deepest studied model virus. It can infect more than 400 crops including tobacco,

27

cucumber, banana and so on.7 As the widely used antiviral agent, ribavirin (Figure 1)

28

displayed less than 50% anti-TMV effect at 500 μg/mL. Control of tobacco mosaic virus

29

disease is very challenging. So developing novel structure, remarkable effect and

30

environmentally friendly pesticides are needed urgently.8

31

Leading discovery and optimization based on natural products are crucial means in

32

development of novel pesticides due to their immense structural diversity and wide

33

variety of biological activities.9−11 The biologically oriented synthesis seeks to elaborate

34

structural modifications onto a bio-active natural-product scaffold to improve its

35

physicochemical property or inherent biological activity.12 Till now, only a small amount

36

of natural products are used as antiviral agents.13,14 Very few antivirals have been studied

37

the mechanism of action. The creation of new antiviral agents based on mechanism of

38

action is difficult to carry out. Development of new antiviral agents limited by backward

39

mechanism research. The antiviral mechanisms in plants involve a series of plant

40

signaling pathways, viral replication, nucleoprotein assembly and RNA-induced gene

41

silencing. Gossypol compounds could neither inhibit the multiplication of TMV nor

42

induce the systemic acquired resistance of tobacco plants.15 Antofine was found to be 3



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43

favorable interaction with origin of TMV RNA to exert its virus inhibition.16 Gramine

44

analogues likely exerted their virus inhibition by crosslinking TMV CP and interfered

45

with virus assembly.17

46

The bisindole alkaloids nortopsentins A−C (Figure 1) were isolated from the

47

deep-water marine sponge Spongosorites ruetzleri by Sun firstly and showed in vitro

48

cytotoxicity against P388 cells and antifungal activity against Candida albicans.18 Due to

49

the interesting biological activities and unique chemical structures of the marine bisindole

50

alkaloids, the marine disindole alkaloids as lead compounds for discovery of new drugs

51

have became an attractive field in medicinal chemistry. So a series of nortopsentin

52

analogues containing five-membered heterocycles, such as bis-indolyl-thiophenes,19

53

-oxazoles,20 –pyrazole,21 -furans,22 -isoxazoles,22 -pyrroles,23 -thiazoles,24,25 and

54

-1,2,4-thiadiazoles26 were reported. But the study of nortopsentin analogues mainly

55

focused on antitumor activity.18-26 In previous work, we have found that nortopsentin

56

alkaloids exhibit good activity in plant-disease prevention.27 In addition, thiazole and

57

oxazole as two kinds of important five-membered heterocycles are widely present in

58

pesticide molecules.28,29 Considering the above findings, we designed and synthesized a

59

series

60

compounds to explore the structure-activity relationship (SAR) in this work (Figure 2).

61

Finally, we evaluated for their antiviral, anti-phytopathogenic-fungus activities and

62

studied antiviral mechanism preliminarily by transmission electron microscope (TEM).

63

MATERIALS AND METHODS

of

bis(indolyl)-thiazole,

bis(indolyl)-oxazole

4


and

mono(indolyl)-thiazole

Page 5 of 51


64

Chemicals. The reagents were purchased from commercial sources and were used as

65

received. All anhydrous solvents were dried and purified by standard techniques prior to

66

use. The intermediates 8a and 9o−9q were purchased from J&K Scientific Ltd.

67

Instruments. The melting points of the compounds were tested on an X-4 binocular

68

microscope (Beijing Tech Instruments Company). NMR spectra were obtained with a

69

Bruker AV 400 spectrometer with either CDCl3 or DMSO-d6 as the solvent.

70

High-resolution mass spectra were obtained with an FT-ICR mass spectrometer (Ionspec,

71

7.0 T). The in vitro TMV rod assembly inhibition and 20S CP disk assembly inhibition

72

were tested via transmission electron microscopy (Tecnai G2 F20).

73

General Procedures for the Preparation of Compounds 4a–4c. To a solution of

74

indoles 3a−3c (15 mmol) in CH3CN was added 60% NaH (21 mmol) at 0 °C. The

75

mixture was stirred for 10 min, and TsCl (16.5 mmol) was added. Then, the mixture was

76

allowed to reach room temperature and was stirred for 4 h. The reaction was quenched

77

with a saturated aqueous solution of NH4Cl. The resulting solution was extracted with

78

ethyl acetate (3 × 100 mL). The combined organic phases were washed with brine (100

79

mL), dried over anhydrous Na2SO4, filtered, and concentrated in vacuo to provide the

80

corresponding compounds 4a−4c.

81

N-Tosylindole (4a). Brown solid; yield 99%; mp 76−78 °C; 1H NMR (400 MHz, CDCl3)

82

δ 7.99 (d, J = 8.4 Hz, 1H), 7.76 (d, J = 8.4 Hz, 2H), 7.56 (d, J = 3.6 Hz, 1H), 7.52 (d, J =

83

7.6 Hz, 1H), 7.36−7.27 (m, 1H), 7.24−7.18 (m, 3H), 6.65 (d, J = 3.6 Hz, 1H), 2.33 (s,

84

3H). 5



85

N-Tosyl-6-bromoindole (4b). Brown solid; yield 99%; mp 131−132 °C; 1H NMR (400

86

MHz, CDCl3) δ 8.17 (s, 1H), 7.76 (d, J = 8.4 Hz, 2H), 7.53 (d, J = 3.6 Hz, 1H), 7.38 (d, J

87

= 8.4 Hz, 1H), 7.33 (dd, J = 8.4, 1.6 Hz, 1H), 7.25 (d, J = 8.4 Hz, 2H), 6.61 (d, J = 3.6 Hz,

88

1H), 2.36 (s, 3H).

89

N-Tosyl-5-bromoindole (4c). Brown solid; yield 99%; mp 133−134 °C; 1H NMR (400

90

MHz, CDCl3) δ 7.86 (d, J = 8.8 Hz, 1H), 7.74 (d, J = 8.4 Hz, 2H), 7.65 (d, J = 1.6 Hz,

91

1H), 7.56 (d, J = 3.6 Hz, 1H), 7.39 (dd, J = 8.8, 2.0 Hz, 1H), 7.23 (d, J = 8.0 Hz, 2H),

92

2.35 (s, 3H).

93

General Procedures for the Preparation of Compounds 5a−5c. To a mixture of AlCl3

94

(300 mmol) in dichloromethane was added acetic anhydride (150 mmol) at 0 °C. Then

95

compounds 4a−4c (50 mmol) were added, and the solution was allowed to reach room

96

temperature and stirred for 2 h. The resulting mixture was poured into ice water and

97

extracted with dichloromethane (3 × 200 mL). The combined organic phase was washed

98

with brine (200 mL), dried over anhydrous Na2SO4, filtered, and concentrated in vacuo to

99

provide the corresponding compounds 5a−5c.

100

N-Tosyl-3-acetylindole (5a). Red-brown solid; yield 99%; mp 143−145 °C; 1H NMR

101

(400 MHz, CDCl3) δ 8.33 (dd, J = 6.8, 1.6 Hz, 1H), 8.21 (s, 1H), 7.93 (dd, J = 7.0, 1.6 Hz,

102

1H), 7.84 (d, J = 8.4 Hz, 2H), 7.41−7.31 (m, 2H), 7.29 (d, J = 8.4 Hz, 2H), 2.58 (s, 3H),

103

2.37 (s, 3H).

104

N-Tosyl-3-acetyl-6-bromoindole (5b). Brown solid; yield 96%; mp 159−160 °C; 1H

105

NMR (400 MHz, CDCl3) δ 8.19 (d, J = 8.8 Hz, 1H), 8.16 (s, 1H), 8.10 (d, J = 1.6 Hz, 6


Page 6 of 51

Page 7 of 51


106

1H), 7.83 (d, J = 8.4 Hz, 2H), 7.45 (dd, J = 8.4, 1.6 Hz, 1H), 7.32 (d, J = 8.4 Hz, 2H),

107

2.56 (s, 3H), 2.40 (s, 3H).

108

N-Tosyl-3-acetyl-6-bromoindole (5c). Light-brown solid; yield 98%; mp 159−160 °C;

109

1H

110

7.82–7.78 (m, 3H), 7.49 (dd, J = 8.8, 2.0 Hz, 1H), 7.30 (d, J = 8.0 Hz, 2H), 2.56 (s, 3H),

111

2.39 (s, 3H).

112

Preparation of 3-Acetyl-1H-5- methoxyindole (7). Phosphoryl chloride (78 mmol) was

113

added to ice cold dimethylacetamide under stirring and cooling in ice. After stirring 0.5 h,

114

a solution of 5-methoxy-1H-indole 3d (60 mmol) in dimethylacetamide was added and

115

the reaction mixture was stirred at 90 ℃ for 2 h, then poured over ice and basified with 4

116

N aqueous sodium hydroxide solution. Finally, the suspension was filtered to give 7.

117

Orange solid; yield 51%; mp 208−209 °C; 1H NMR (400 MHz, DMSO-d6) δ 11.81 (s,

118

1H), 8.24 (s, 1H), 7.68 (d, J = 2.0 Hz, 1H), 7.36 (d, J = 8.8 Hz, 1H), 6.84 (dd, J = 8.8, 2.2

119

Hz, 1H), 3.77 (s, 3H), 2.43 (s, 3H).

120

Preparation of N-Tosyl-3-acetyl-6-methoxyindole (5d). To a solution of 7 (5 mmol) in

121

dichloromethane

122

4-dimethylaminopyridine (0.5 mmol) and triethylamine (7.5 mmol), then the mixture was

123

stirred at room temperature for 3 h. The resulting mixture was washed with water (2 × 50

124

mL) and brine (50 mL), dried over anhydrous Na2SO4, filtered, and concentrated in vacuo

125

to provide compound 5d. Brown solid; yield 82%; mp 162−165 °C; 1H NMR (400 MHz,

126

CDCl3) δ 8.16 (s, 1H), 7.84–7.78 (m, 4H), 7.29 (d, J = 8.0Hz, 2H), 6.97 (dd, J = 9.2, 2.8

NMR (400 MHz, CDCl3) δ 8.52 (d, J = 2.0 Hz, 1H), 8.20 (s, 1H), 7.85 (s, 1H),

was

added

4-toluene

sulfonyl

7


chloride

(5.5

mmol),


127

Hz, 1H), 3.85 (s, 3H), 2.56 (s, 3H), 2.37 (s, 3H).

128

General Procedures for the Preparation of Compounds 6a–6d. To a solution of 5a–5d

129

(12.7 mmol) in mixed solvent (ethyl acetate: chloroform = 1:1) was added CuBr2 (28

130

mmol). The mixture was heated at reflux for 20 min and continued to react for 2 h after

131

adding ethanol. The resulting mixture was filtered, and filtrate was washed with water (3

132

× 200 mL) and brine (200 mL). The organic phase was dried over anhydrous Na2SO4,

133

filtered, and concentrated in vacuo. The residue was purified by flash chromatography on

134

a silica gel using petroleum ether and ethyl acetate (5:1, v/v) as the eluent to give 6a−6d.

135

N-Tosyl-3-(α-bromoacetyl)-indole (6a). Yellow solid; yield 89%; mp 118−119 °C; 1H

136

NMR (400 MHz, CDCl3) δ 8.35 (s, 1H), 8.30 (d, J = 7.6 Hz, 1H), 7.93 (d, J = 7.6 Hz,

137

1H), 7.85 (d, J = 8.0 Hz, 2H), 7.45−7.33 (m, 2H), 7.30 (d, J = 8.0 Hz, 2H), 4.36 (s, 2H),

138

2.38 (s, 3H).

139

N-Tosyl-3-(α-bromoacetyl)-6-bromoindole (6b). Brown solid; yield 81%; mp 162−163

140

°C; 1H NMR (400 MHz, CDCl3) δ 8.30 (s, 1H), 8.16 (d, J = 8.4 Hz, 1H), 8.11 (s, 1H),

141

7.84 (d, J = 8.2 Hz, 2H), 7.48 (d, J = 8.5 Hz, 1H), 7.34 (d, J = 8.0 Hz, 2H), 4.33 (s, 2H),

142

2.40 (s, 3H).

143

N-Tosyl-3-(α-bromoacetyl)-5-bromoindole (6c). Brown solid; yield 88%; mp 163−164

144

°C; 1H NMR (400 MHz, CDCl3) δ 8.47 (d, J = 1.4 Hz, 1H), 8.32 (s, 1H), 7.86–7.77 (m,

145

3H), 7.49 (dd, J = 8.8, 1.6 Hz, 1H), 7.31 (d, J = 8.2 Hz, 2H), 4.32 (s, 2H), 2.39 (s, 3H).

146

N-Tosyl-3-(α-bromoacetyl)-5-methoxyindole (6d). Yellow solid; yield 62%; mp

147

171−172 °C; 1H NMR (400 MHz, CDCl3) δ 8.28 (s, 1H), 7.84–7.80 (m, 3H), 7.77 (d, J = 8


Page 8 of 51

Page 9 of 51


148

2.8 Hz, 1H), 7.29 (d, J = 8.4 Hz, 2H), 6.99 (dd, J = 9.2, 2.4 Hz, 1H), 4.35 (s, 2H), 3.85 (s,

149

3H), 2.38 (s, 3H).

150

General Procedures for the Preparation of Compounds 8b−8d. To a solution of

151

indoles 3b−3d (10 mmol) in DMF was added chlorosulfonamide isocyanate (12 mmol) at

152

-50 °C in an argon atmosphere. The temperature was raised to -10 °C, and then the

153

solution was brought to room temperature after it was stirred for 1.5 h. The resulting

154

solution was poured into ice water, and kept for 30 min, and filtered to give 8b−8d.

155

3-Cyano-6-bromoindole (8b). Yellow solid; yield 99%; mp 189−190 °C; 1H NMR (400

156

MHz, DMSO-d6) δ 12.33 (s, 1H), 8.29 (d, J = 2.8 Hz, 1H), 7.76 (d, J = 1.2 Hz, 1H), 7.61

157

(d, J = 8.4 Hz, 1H), 7.38 (dd, J = 8.4, 1.6 Hz, 1H).

158

3-Cyano-5-bromoindole (8c). Yellow solid; yield 97%; mp 178−179 °C; 1H NMR (400

159

MHz, DMSO-d6) δ 12.41 (s, 1H), 8.32 (d, J = 2.8 Hz, 1H), 7.80 (d, J = 1.2 Hz, 1H), 7.53

160

(d, J = 8.8 Hz, 1H), 7.42 (dd, J = 8.4, 1.6 Hz, 1H).

161

3-Cyano-5-methoxyindole (8d). Brown solid; yield 99%; mp 155−156 °C; 1H NMR

162

(400 MHz, DMSO-d6) δ 12.06 (s, 1H), 8.16 (s, 1H), 7.44 (d, J = 8.8 Hz, 1H), 7.07 (s, 1H),

163

6.90 (d, J = 8.6 Hz, 1H), 3.81 (s, 3H).

164

Preparation of N-Tosyl-3-cyano-6-methoxyindole (10). To a solution of 8d (5 mmol)

165

in

166

4-dimethylaminopyridine (0.5 mmol) and triethylamine (7.5 mmol), then the mixture was

167

stirred at room temperature for 3 h. The resulting mixture was washed with water (2 × 50

168

mL) and brine (50 mL), dried over anhydrous Na2SO4, filtered, and concentrated in vacuo

dichloromethane

was

added

4-toluene

sulfonyl

9


chloride

(5.5

mmol),


Page 10 of 51

169

to provide compound 10. Yellow solid; yield 88%; mp 157−158 °C; 1H NMR (400 MHz,

170

CDCl3) δ 8.04 (s, 1H), 7.87 (d, J = 9.2 Hz, 1H), 7.80 (d, J = 8.4 Hz, 2H), 7.30 (d, J = 8.0

171

Hz, 2H), 7.08 (d, J = 2.4 Hz, 1H), 7.05 (s, 1H, Ar-H), 3.84 (s, 3H), 2.39 (s, 3H).

172

General Procedures for the Preparation of Compounds 9a−9c, 11 and 9i−9m. To a

173

slurry of 70% sodium hydrosulfide hydrate (24 mmol) and magnesium chloride

174

hexahydrate (8 mmol) in 10 mL of DMF was added corresponding materials (8 mmol) in

175

one portion, and the mixture was stirred at 40 °C overnight. The resulting green slurry

176

was poured into 200 mL of water, and the resulting precipitates were collected by

177

filtration. The crude product was resuspended in 1 N HCl and stirred for 20 min, then

178

filtered and washed with water to give 9a−9c, 11 and 9i−9m.

179

1H-Indole-3-carbothioamide (9a). Yellow solid; yield 89%; mp 141−142 °C; 1H NMR

180

(400 MHz, DMSO-d6) δ 11.78 (s, 1H), 8.96 (s, 1H), 8.82 (s, 1H), 8.71–8.54 (m, 1H), 8.09

181

(d, J = 2.8 Hz, 1H), 7.49–7.40 (m, 1H), 7.21–7.09 (m, 2H).

182

DMSO-d6) δ 193.6, 136.8, 128.0, 1259, 122.0, 121.8, 120.7, 116.3, 111.9.

183

1H-6-Bromoindole-3-carbothioamide (9b). Yellow solid; yield 87%; mp 195−196 °C;

184

1H

185

8.8 Hz, 1H), 8.10 (d, J = 3.2 Hz, 1H), 7.63 (d, J = 1.6 Hz, 1H), 7.28 (dd, J = 8.8, 2.0 Hz,

186

1H).

187

1H-5-Bromoindole-3-carbothioamide (9c). Yellow solid; yield 85%; mp 141−142 °C;

188

1H

189

8.15 (s, 1H), 7.42 (d, J = 8.8 Hz, 1H), 7.30 (d, J = 8.8 Hz, 1H).

13C

NMR (100 MHz,

NMR (400 MHz, DMSO-d6) δ 11.87 (s, 1H), 9.05 (s, 1H), 8.92 (s, 1H), 8.60 (d, J =

NMR (400 MHz, DMSO-d6) δ 11.98 (s, 1H), 9.04 (s, 1H), 8.94 (s, 1H), 8.90 (s, 1H),

10


Page 11 of 51


190

N-Tosyl-5-methoxyindole-3-carbothioamide (11). Yellow oil; yield 85%; 1H NMR

191

(400 MHz, DMSO-d6) δ 9.62 (s, 1H), 9.41 (s, 1H), 8.36 (s, 1H), 8.09 (d, J = 2.6 Hz, 1H),

192

7.92 (d, J = 8.4 Hz, 2H), 7.82 (d, J = 9.2 Hz, 1H), 7.42 (d, J = 8.4 Hz, 2H), 6.99 (dd, J =

193

8.8, 2.6 Hz, 1H), 3.75 (s, 3H), 2.33 (s, 3H).

194

156.2, 146.1, 133.5, 130.4, 129.4, 128.8, 127.0, 126.9, 121.4, 114.1, 113.8, 105.3, 55.3,

195

21.0.

196

Benzothioamide (9i). Yellow solid; yield 85%; mp 113–115 °C; 1H NMR (400 MHz,

197

DMSO-d6) δ 9.89 (s, 1H), 9.51 (s, 1H), 7.88 (d, J = 8.0 Hz, 2H), 7.50 (t, J = 7.3 Hz, 1H),

198

7.41 (t, J = 7.6 Hz, 2H).

199

4-Fluorobenzothioamide (9j). Yellow solid; yield 76%; mp 143−144 °C; 1H NMR (400

200

MHz, DMSO-d6) δ 9.90 (s, 1H), 9.52 (s, 1H), 7.99–7.95 (m, 2H), 7.27–7.23 (m, 2H).

201

2-(Trifluoromethyl)benzothioamide (9k). Yellow solid; yield 94%; mp 76−77 °C; 1H

202

NMR (400 MHz, DMSO-d6) δ 10.18 (s, 1H), 9.72 (s, 1H), 7.72–7.63 (m, 2H), 7.54 (t, J =

203

7.6 Hz, 1H), 7.39 (d, J = 7.6 Hz, 1H).

204

3, 4-Dichlorobenzothioamide (9l). Yellow solid; yield 89%; mp 133−134 °C; 1H NMR

205

(400 MHz, DMSO-d6) δ 10.11 (s, 1H), 9.68 (s, 1H), 8.09 (s, 1H), 7.86 (d, J = 8.4 Hz, 1H),

206

7.71 (d, J = 8.4 Hz, 1H).

207

4-Penoxybenzothioamide (9m). Yellow solid; yield 89%; mp 158−159 °C; 1H NMR

208

(400 MHz, DMSO-d6) δ 9.78 (s, 1H), 9.43 (s, 1H), 7.96 (d, J = 8.8 Hz, 2H), 7.44 (t, J =

209

7.8 Hz, 2H), 7.22 (t, J = 7.2 Hz, 1H), 7.09 (d, J = 8.2 Hz, 2H), 6.98 (d, J = 8.8 Hz, 2H).

210

Preparation of Heptanethioamide (9n). To a solution of heptanenitrile (3 mmol) in

13C

NMR (100 MHz, DMSO-d6) δ 192.2,

11



211

anhydrous ethanol was added P2S5 (3.15 mmol) and stirred at room temperature overnight.

212

The resulting mixture was quenched with water and extracted with chloroform. The

213

organic fractions were evaporated under reduce pressure and then, the evaporated fraction

214

was recrystallized in hexane to give 9n. White solid; yield 35%; mp 57−58 °C; 1H NMR

215

(400 MHz, CDCl3) δ 7.54 (s, 1H), 6.84 (s, 1H), 2.67 (t, J = 7.8 Hz, 2H), 1.81–1.73(m,

216

2H), 1.41–1.25 (m, 6H), 0.89 (t, J = 6.8 Hz, 3H).

217

General Procedures for the Preparation of Compounds 2a−2q. To a solution of

218

6a−6d (2 mmol) in anhydrous ethanol was added 9a−9c, 11 and 9i−9q (2 mmol). The

219

mixture was stirred at reflux for 0.5 h, then filtered to give 2a−2q.

220

2-(1H-Indol-3-yl)-4-(1-tosyl-1H-indol-3-yl)thiazole (2a). Yellow solid; yield 97%; mp

221

246−248 °C; 1H NMR (400 MHz, DMSO-d6) δ 11.87 (s, 1H), 8.38 (s, 1H), 8.36 (d, J =

222

7.2 Hz, 1H), 8.31–8.26 (m, 1H), 8.23 (d, J = 2.8 Hz, 1H), 8.05 (s, 1H), 8.03 (d, J = 7.2 Hz,

223

1H), 7.95 (d, J = 8.4 Hz, 2H), 7.54 (dd, J = 5.2, 3.6 Hz, 1H), 7.45 (m, 2H), 7.40 (d, J =

224

8.0 Hz, 2H), 7.30–7.24 (m, 2H), 2.30 (s, 3H). 13C NMR (100 MHz, DMSO-d6) δ 163.3,

225

147.5, 146.2, 137.1, 135.2, 134.4, 130.8, 128.5, 127.5, 127.3, 125.7, 124.9, 124.7, 124.5,

226

122.9, 122., 121.4, 120.5, 118.3, 113.9, 112.8, 111.9, 110.7, 21.5. C26H20N3O2S2 [M+H]+

227

470.0991, found (ESI+) 470.1000.

228

2-(6-Bromo-1H-indol-3-yl)-4-(1-tosyl-1H-indol-3-yl)thiazole (2b). Orange solid; yield

229

96%; mp 192−194 °C; 1H NMR (400 MHz, DMSO-d6) δ 11.97 (s, 1H), 8.37 (s, 1H), 8.33

230

(d, J = 7.6 Hz, 1H), 8.26 (s, 1H), 8.24 (d, J = 8.0 Hz, 2H), 8.06 (s, 1H), 8.03 (d, J = 8.0

231

Hz, 1H), 7.95 (d, J = 8.0 Hz, 2H), 7.73 (s, 1H), 7.48–7.37 (m, 5H), 7.39 (d, J = 8.0 Hz, 12


Page 12 of 51

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232

2H), 2.31 (s, 3H). 13C NMR (100 MHz, DMSO-d6) δ 162.7, 147.7, 146.2, 138.0, 135.2,

233

134.3, 130.8, 128.4, 127.3, 125.7, 125.0, 124.5, 124.3, 123.8, 122.4, 122.2, 118.3, 115.6,

234

115.4, 113.9, 112.3, 111.0, 21.5. C26H19BrN3O2S2 [M+H]+ 548.0097, found (ESI+)

235

548.0086.

236

4-(6-Bromo-1-tosyl-1H-indol-3-yl)-2-(1H-indol-3-yl)thiazole (2c). Orange solid; yield

237

84%; mp 236−237 °C; 1H NMR (400 MHz, DMSO-d6) δ 11.87 (s, 1H), 8.44 (s, 1H), 8.36

238

(d, J = 8.4 Hz, 1H), 8.28–8.23 (m, 1H), 8.23 (d, J = 2.8 Hz, 1H), 8.15 (d, J = 1.6 Hz, 1H),

239

8.06 (s, 1H), 7.99 (d, J = 8.4 Hz, 2H), 7.61 (dd, J = 8.4, 1.6 Hz, 1H), 7.56–7.52 (m, 1H),

240

7.44 (d, J = 8.4 Hz, 2H), 7.31–7.22 (m, 2H), 2.33 (s, 3H).

241

DMSO-d6) δ 163.4, 147.1, 146.5, 137.1, 135.8, 134.1, 131.0, 127.7, 127.6, 127.4, 125.5,

242

124.7, 124.3, 122.9, 121.4, 120.5, 118.5, 118.1, 116.2, 112.8, 112.3, 110.7, 21.5.

243

C26H19BrN3O2S2 [M+H]+ 548.0097, found (ESI+) 548.0093.

244

4-(6-Bromo-1-tosyl-1H-indol-3-yl)-2-(6-bromo-1H-indol-3-yl)thiazole (2d). Yellow

245

solid; yield 85%; mp 223−224 °C; 1H NMR (400 MHz, DMSO-d6) δ 11.96 (s, 1H), 8.43

246

(s, 1H), 8.32 (d, J = 8.4 Hz, 1H), 8.24 (d, J = 2.8 Hz, 1H), 8.21 (d, J = 8.4 Hz, 1H), 8.15

247

(d, J = 1.6 Hz, 1H), 8.07 (s, 1H), 7.99 (d, J = 8.4 Hz, 2H), 7.72 (d, J = 1.6 Hz, 1H), 7.61

248

(dd, J = 8.4, 1.6 Hz, 1H), 7.44 (d, J = 8.4 Hz, 2H), 7.41 (dd, J = 8.4, 1.6 Hz, 1H), 2.33 (s,

249

3H). 13C NMR (100 MHz, DMSO-d6) δ 162.9, 147.3, 146.6, 137.9, 135.8, 134.1, 131.0,

250

128.4, 127.6, 127.4, 125.5, 124.4, 124.2, 123.7, 122.4, 118.5, 118.1, 116.2, 115.6, 115.3,

251

112.6, 110.9, 21.5. C26H18Br2N3O2S2 [M+H]+ 625.9202, found (ESI+) 625.9178.

252

4-(5-Bromo-1-tosyl-1H-indol-3-yl)-2-(5-bromo-1H-indol-3-yl)thiazole (2e). Orange 13


13C

NMR (100 MHz,


253


254

(d, J = 1.8 Hz, 1H), 8.50 (s, 1H), 8.49 (d, J = 1.2 Hz, 1H), 8.28 (d, J = 2.8 Hz, 1H), 8.13

255

(s, 1H), 8.00 (d, J = 8.8 Hz, 1H), 7.95 (d, J = 8.4 Hz, 2H), 7.61 (dd, J = 8.8, 2.0 Hz, 1H),

256

7.52 (d, J = 8.6 Hz, 1H), 7.42 (d, J = 8.2 Hz, 2H), 7.38 (dd, J = 8.6, 2.0 Hz, 1H), 2.32 (s,

257

3H). 13C NMR (100 MHz, DMSO-d6) δ 162.6, 146.9, 146.0, 135.3, 133.5, 133.4, 130.4,

258

129.9, 128.5, 127.9, 126.8, 125.9, 125.6, 125.1, 124.4, 122.5, 117.0, 116.8, 115.3, 114.3,

259

113.6, 112.0, 109.8, 21.0. C26H18Br2N3O2S2 [M+H]+ 625.9202, found (ESI+) 625.9194.

260

4-(5-Bromo-1-tosyl-1H-indol-3-yl)-2-(6-bromo-1H-indol-3-yl)thiazole (2f). Orange

261


262

(d, J = 2.0 Hz, 1H), 8.47 (s, 1H), 8.26 (d, J = 8.8Hz, 1H), 8.23 (d, J = 2.8Hz, 1H), 8.11 (s,

263

1H), 7.99 (d, J = 8.8 Hz, 1H), 7.95 (d, J = 8.4 Hz, 2H), 7.72 (d, J = 1.6 Hz, 1H), 7.60 (dd,

264

J = 8.8, 1.6 Hz, 1H), 7.42 (d, J = 8.4 Hz, 2H), 7.37 (dd, J = 8.5, 1.4 Hz, 1H), 2.32 (s, 3H).

265

13C

266

128.3, 127.3, 126.1, 124.1, 123.8, 117.5, 117.3, 115.8, 115.6, 115.4, 112.6, 110.9, 21.5.

267

C26H18Br2N3O2S2 [M+H]+ 625.9202, found (ESI+) 625.9200.

268

2,4-Bis(5-methoxy-1-tosyl-1H-indol-3-yl)thiazole (2g). Red solid; yield 79%; mp

269

236−238 °C; 1H NMR (400 MHz, DMSO-d6) δ 8.57 (s, 1H), 8.39 (s, 1H), 8.26 (s, 1H),

270

8.01−7.88 (m, 7H), 7.80 (d, J = 2.4 Hz, 1H), 7.41 (d, J = 8.4 Hz, 2H), 7.38 (d, J = 8.4 Hz,

271

2H), 7.11 (dd, J = 9.2, 2.8 Hz, 1H), 7.07 (dd, J = 9.2, 2.8 Hz, 1H), 3.84 (s, 3H), 3.82 (s,

272

3H), 2.32 (s, 3H), 2.30 (s, 3H). 13C NMR (100 MHz, DMSO-d6) δ 160.2, 157.3, 157.0,

273

148.4, 146.4, 146.1, 134.3, 134.2, 130.84, 130.76, 129.8, 129.5, 129.4, 128.6, 127.6,

NMR (100 MHz, DMSO-d6) δ 162.9, 147.2, 146.4, 138.0, 134.1, 133.9, 130.9, 128.5,

14


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274

127.5, 127.2, 126.0, 117.9, 116.9, 114.8, 114.4, 114.1, 110.0, 105.2, 105.0, 56.0, 55.8,

275

21.5, 21.4. C35H30N3O6S3 [M+H]+ 684.1291, found (ESI+) 684.1293.

276

2-(6-Bromo-1H-indol-3-yl)-4-(5-methoxy-1-tosyl-1H-indol-3-yl)thiazole (2h). Orange

277


278

(s, 1H), 8.32 (d, J = 8.4 Hz, 1H), 8.25 (d, J = 2.8 Hz, 1H), 8.08 (s, 1H), 7.93−7.89 (m,

279

3H), 7.87 (d, J = 2.4 Hz, 1H), 7.73 (d, J = 1.2 Hz, 1H), 7.39 (d, J = 8.2 Hz, 2H), 7.36 (dd,

280

J = 8.4, 1.6 Hz, 1H), 7.05 (dd, J = 9.0, 2.8 Hz, 1H), 3.87 (s, 3H), 2.30 (s, 3H). 13C NMR

281

(100 MHz, DMSO-d6) δ 162.3, 156.4, 147.4, 145.5, 137.4, 133.8, 130.2, 129.2, 129.1,

282

127.9, 126.7, 125.0, 123.6, 123.3, 122.0, 117.8, 115.1, 114.9, 114.4, 114.3, 111.6, 110.5,

283

103.8, 55.4, 21.0. C27H21BrN3O3S2 [M+H]+ 578.0202, found (ESI+) 578.0202.

284

4-(6-Bromo-1-tosyl-1H-indol-3-yl)-2-phenylthiazole (2i). White solid; yield 75%; mp

285

173−175 °C; 1H NMR (400 MHz, DMSO-d6) δ 8.50 (s, 1H), 8.33 (d, J = 8.8 Hz, 1H),

286

8.29 (s, 1H), 8.14 (d, J = 1.6 Hz, 1H), 8.07 (dd, J = 7.8, 1.8 Hz, 2H), 7.98 (d, J = 8.4 Hz,

287

2H), 7.61–7.53 (m, 4H), 7.44 (d, J = 8.2 Hz, 2H), 2.33 (s, 3H). 13C NMR (100 MHz,

288

DMSO-d6) δ 167.5, 148.6, 146.5, 135.7, 134.2, 133.2, 130.98, 130.95, 129.8, 127.6,

289

127.4, 127.3, 126.7, 125.8, 124.3, 118.5, 117.7, 116.2, 116.1, 21.5. C24H18BrN2O2S2

290

[M+H]+ 508.9988, found (ESI+) 508.9984.

291

4-(6-Bromo-1-tosyl-1H-indol-3-yl)-2-(4-fluorophenyl)thiazole (2j). White solid; yield

292

82%; mp 229−230 °C; 1H NMR (400 MHz, DMSO-d6) δ 8.49 (s, 1H), 8.32 (d, J = 8.4 Hz,

293

1H), 8.29 (s, 1H), 8.14–8.11 (m, 3H), 7.98 (d, J = 8.4 Hz, 2H), 7.58 (dd, J = 8.4, 1.6 Hz,

294

1H), 7.47–7.37 (m, 4H), 2.33 (s, 3H). 13C NMR (100 MHz, DMSO-d6) δ 166.3, 163.8 (d, 15



295

J = 248.0 Hz), 148.6, 146.6, 135.7, 134.1, 131.0, 129.9, 129.1 (d, J = 7.8 Hz), 127.7,

296

127.6, 127.4, 125.9, 124.3, 118.6, 117.6, 116.8 (d, J = 22.0 Hz), 116.2, 110.0, 21.5.

297

C24H17BrFN2O2S2 [M+H]+ 526.9893, found (ESI+) 526.9888.

298

4-(6-Bromo-1-tosyl-1H-indol-3-yl)-2-(2-(trifluoromethyl)phenyl)thiazole (2k). White

299

solid; yield 64%; mp 152−153 °C; 1H NMR (400 MHz, DMSO-d6) δ 8.48 (d, J = 7.4 Hz,

300

2H), 8.25 (d, J = 8.6 Hz, 1H), 8.12 (s, 1H), 8.00–7.94 (m, 3H), 7.86 (d, J = 4.0 Hz, 2H),

301

7.81–7.78(m, 1H), 7.53 (dd, J = 8.4, 1.6 Hz, 1H), 7.44 (d, J = 8.2 Hz, 2H), 2.33 (s, 3H).

302

13C

303

(q, J = 2.0 Hz), 131.0, 128.3, 127.5 (q, J = 30.0 Hz),127.4, 127.3, 125.8, 124.2, 124.1(q, J

304

= 272.0 Hz), 118.5, 118.2, 117.5, 116.2, 21.5. C25H17BrF3N2O2S2 [M+H]+ 576.9861,

305

found (ESI+) 576.9860.

306

4-(6-Bromo-1-tosyl-1H-indol-3-yl)-2-(3,4-dichlorophenyl)thiazole 2l. White solid;

307

yield 97%; mp 211−212 °C; 1H NMR (400 MHz, DMSO-d6) δ 8.51 (s, 1H), 8.35 (s, 1H),

308

8.28–8.26 (m, 2H), 8.13 (d, J = 1.6 Hz, 1H), 8.04–8.01 (m, 1H), 7.98 (d, J = 8.4 Hz, 2H),

309

7.80 (d, J = 8.4 Hz, 1H), 7.58 (d, J = 8.4 Hz, 1H), 7.44 (d, J = 8.2 Hz, 2H), 2.33 (s, 3H).

310

13C

311

131.4, 130.5, 127.5, 127.1, 126.9, 126.8, 126.3, 125.6, 123.6, 118.0, 116.9, 116.8, 115.7,

312

99.5, 21.0. C24H16BrCl2N2O2S2 [M+H]+ 576.9208, found (ESI+) 576.9203.

313

4-(6-Bromo-1-tosyl-1H-indol-3-yl)-2-(4-phenoxyphenyl)thiazole (2m). White solid;

314

yield 68%; mp 189−190 °C; 1H NMR (400 MHz, DMSO-d6) δ 8.47 (s, 1H), 8.31 (d, J =

315

8.6 Hz, 1H), 8.24 (s, 1H), 8.13 (d, J = 1.4 Hz, 1H), 8.07 (d, J = 8.8 Hz, 2H), 7.97 (d, J =

NMR (100 MHz, DMSO-d6) δ 164.2, 148.3, 146.6, 135.7, 134.1, 133.3, 132.9, 132.2

NMR (100 MHz, DMSO-d6) δ 164.1, 148.4, 146.1, 135.2), 133.7, 133.1, 132.8, 132.1,

16


Page 16 of 51

Page 17 of 51


316

8.4 Hz, 2H), 7.57 (dd, J = 8.4, 1.6 Hz, 1H), 7.49–7.42 (m, 4H), 7.23 (t, J = 7.4 Hz, 1H),

317

7.15−7.12 (m, 4H), 2.33 (s, 3H). 13C NMR (100 MHz, DMSO-d6) δ 166.4, 158. 8, 155.7,

318

148.0, 146.1, 135.2, 133.7, 130.5, 130.2, 128.2, 127.9, 127.1, 127.0, 126.8, 125.3, 124.2,

319

123.8, 119.4, 118.6, 118.0, 117.2, 115.7, 115.2, 21.0. C30H22BrN2O3S2 [M+H]+ 601.0250,

320

found (ESI+) 601.0247.

321

4-(6-Bromo-1-tosyl-1H-indol-3-yl)-2-(4-hexylphenyl)thiazole (2n). White solid; yield

322

84%; mp 163−165 °C; 1H NMR (400 MHz, DMSO-d6) δ 8.35 (s, 1H), 8.21 (d, J = 8.6 Hz,

323

1H), 8.14 (d, J = 1.6 Hz, 1H), 8.08 (s, 1H), 7.95 (d, J = 8.4 Hz, 2H), 7.54 (dd, J = 8.5, 1.7

324

Hz, 1H), 7.41 (d, J = 8.3 Hz, 2H), 3.04 (t, J = 7.6 Hz, 2H), 2.31 (s, 3H), 1.73–1.75 (m,

325

2H), 1.35–1.27 (m, 6H), 0.85 (t, J = 7.0 Hz, 3H).

326

171.3, 146.7, 146.5, 135.7, 134.1, 130.9, 127.4, 127.3, 125.3, 124.1, 118.4, 117.8, 116.2,

327

115.0, 33.0, 31.3, 29.8, 28.5, 22.4, 21.5, 14.3. C24H26BrN2O2S2 [M+H]+ 517.0614, found

328

(ESI+) 517.0620.

329

4-(6-Bromo-1-tosyl-1H-indol-3-yl)-2-(p-tolyl)thiazole (2o). White solid; yield 94%; mp

330


331

8.11 (d, J = 1.6 Hz, 1H), 8.04 (s, 1H), 7.95 (d, J = 8.4 Hz, 2H), 7.54 (dd, J = 8.6, 1.8 Hz,

332

1H), 7.43 (d, J = 8.0 Hz, 2H), 2.74 (s, 3H), 2.33 (s, 3H). 13C NMR (100 MHz, DMSO-d6)

333

δ 166.0, 146.0, 135.2, 133.6, 130.5, 126.9, 126.8, 125.0, 123.6, 118.0, 117.1, 115.7, 115.0,

334

21.0, 18.72. C19H16BrN2O2S2 [M+H]+ 446.9831, found (ESI+) 446.9824.

335

4-(6-Bromo-1-tosyl-1H-indol-3-yl)-2-(4-(tert-butyl)phenyl)thiazole (2p). White solid;

336


13C

NMR (100 MHz, DMSO-d6) δ

17



337

8.8 Hz, 1H), 8.13 (d, J = 1.6 Hz, 1H), 8.06 (s, 1H), 7.95 (d, J = 8.4 Hz, 2H), 7.55 (dd, J =

338

8.6, 2.0 Hz, 1H), 7.41 (d, J = 8.0 Hz, 2H), 2.31 (s, 3H), 1.45 (s, 9H). 13C NMR (100 MHz,

339

DMSO-d6) δ 180.8, 146.8, 146.4, 135.7, 134.1, 131.0, 127.7, 127.5, 127.3, 125.2, 124.3,

340

118.4, 118.1, 116.2, 114.6, 37.8, 31.0, 26.8, 21.5. C22H22BrN2O2S2 [M+H]+ 489.0301,

341

found (ESI+) 489.0299.

342

Ethyl 2-(4-(4-(6-bromo-1-tosyl-1H-indol-3-yl)thiazol-2-yl)phenyl)acetate (2q). White

343

solid; yield 68%; mp 138−139 °C; 1H NMR (400 MHz, DMSO-d6) δ 8.34 (s, 1H), 8.18 (d,

344

J = 8.8 Hz, 1H), 8.17(s, 1H), 8.11 (d, J = 1.6 Hz, 1H), 7.95 (d, J = 8.4 Hz, 2H), 7.54 (dd,

345

J = 8.6, 1.8 Hz, 1H), 7.43 (d, J = 8.4 Hz, 2H), 4.27 (s, 2H), 4.17 (q, J = 7.2 Hz, 2H), 2.33

346

(s, 3H), 1.23 (t, J = 7.2 Hz, 3H). 13C NMR (100 MHz, DMSO-d6) δ 168.9, 162.1, 146.4,

347

146.0, 135.2, 133.6, 130.5, 126.93, 126.90, 126.8, 124.9, 123.6, 118.0, 117.2, 116.3,

348

115.7, 61.0, 38.1, 21.0, 14.0. C22H20BrN2O4S2 [M+H]+ 519.0042, found (ESI+) 519.0046.

349

General Procedures for the Preparation of Compounds 1a−1p. To a solution of

350

compounds 2a−2p (1 mmol) in MeOH (40 mL) was added 2 N NaOH (10 mL). The

351

mixture was heated at reflux for 1.5 h, cooled to room temperature, and extracted with

352

dichloromethane (3 × 50 mL). The combined organic phases were washed with brine

353

(100 mL), dried over anhydrous Na2SO4, filtered, and concentrated in vacuo to give

354

1a−1p.

355

2,4-Di(1H-indol-3-yl)thiazole (1a). Yellow solid; yield 88%; mp 282−283 °C; 1H NMR

356

(400 MHz, DMSO-d6) δ 11.81 (s, 1H), 11.46 (s, 1H), 8.4–8.30 (m, 1H), 8.22 (d, J = 7.2

357

Hz, 1H), 8.14 (d, J = 2.0 Hz, 1H), 8.00 (d, J = 1.6 Hz, 1H), 7.60 (s, 1H), 7.54–7.50 (m, 18


Page 18 of 51

Page 19 of 51


13C

358

1H), 7.49 (d, J = 8.0 Hz, 1H), 7.25 (dd, J = 5.6, 3.2 Hz, 2H), 7.22–7.11 (m, 2H).

359

NMR (100 MHz, DMSO-d6) δ 162.2, 151.0, 137.1, 137.1, 126.8, 125.3, 125.2, 124.9,

360

122.8, 122.0, 121.2, 120.9, 120.6, 120.1, 112.7, 112.4, 111.8, 111.3, 106.4. C19H14N3S

361

[M+H]+ 316.0903, found (ESI+) 316.0909.

362

2-(6-Bromo-1H-indol-3-yl)-4-(1H-indol-3-yl)thiazole (1b). Light-green solid; yield

363

76%; mp 258−260 °C; 1H NMR (400 MHz, DMSO-d6) δ 11.87 (s, 1H), 11.42 (s, 1H),

364

8.35 (d, J = 8.4 Hz, 1H),8.21–8.18 (m, 2H), 8.01 (d, J = 2.0 Hz, 1H), 7.71 (s, 1H), 7.62 (s,

365

1H), 7.49 (d, J = 7.2 Hz, 1H), 7.39 (d, J = 8.4 Hz, 1H), 7.27–7.12 (m, 2H).

366

(100 MHz, DMSO-d6) δ 161.7, 151.1, 137.9, 137.1, 127.8, 125.4, 125.1, 124.1, 123.9,

367

122.8, 122.0, 120.6, 120.2, 115.5, 115.2, 112.4, 111.7, 111.5, 106.7. C19H13BrN3S

368

[M+H]+ 394.0008, found (ESI+) 394.0008.

369

4-(6-Bromo-1H-indol-3-yl)-2-(1H-indol-3-yl)thiazole (1c). Light-pink solid; yield 97%;

370

mp 266−267 °C; 1H NMR (400 MHz, DMSO-d6) δ 11.78 (s, 1H), 11.56 (s, 1H),

371

8.41–8.31 (m, 1H), 8.21 (d, J = 8.4 Hz, 1H), 8.16 (d, J = 2.8 Hz, 1H), 8.04 (d, J = 2.4 Hz,

372

1H), 7.69 (d, J = 1.6 Hz, 1H), 7.63 (s, 1H), 7.60–7.46 (m, 1H), 7.30 (dd, J = 8.4, 1.6 Hz,

373

1H), 7.28–7.22 (m, 2H). 13C NMR (100 MHz, DMSO-d6) δ 162.5, 150.4, 138.0, 137.1,

374

126.9, 126.2, 124.8, 124.3, 123.0, 122.8, 122.5, 121.2, 120.9, 114.9 114.8, 112.7, 112.1,

375

111.2, 107.1. C19H13BrN3S [M+H]+ 394.0008, found (ESI+) 394.0001.

376

2,4-Bis(6-bromo-1H-indol-3-yl)thiazole (1d). Light-pink solid; yield 88%; mp 265−266

377

°C; 1H NMR (400 MHz, DMSO-d6) δ 11.87 (s, 1H), 11.55 (s, 1H), 8.31 (d, J = 8.4 Hz,

378

1H), 8.18 (d, J = 2.8 Hz, 1H), 8.17 (d, J = 2.8 Hz, 1H). 8.03 (d, J = 2.8 Hz, 1H), 7.70 (d, J 19


13C

NMR


379

= 1.6 Hz, 1H), 7.67 (d, J = 1.6 Hz, 1H), 7.66 (s, 1H), 7.39 (dd, J = 8.4, 1.6 Hz, 1H), 7.29

380

(dd, J = 8.4, 1.6 Hz, 1H). 13C NMR (100 MHz, DMSO-d6) δ 161.9, 150.5, 138.0, 137.9,

381

127.9, 126.2, 124.2, 124.1, 123.9, 123.0, 122.7, 122.4, 115.5, 115.2, 114.9, 114.8, 111.9,

382

111.4, 107.4. C19H12Br2N3S [M+H]+ 471.9113, found (ESI+) 471.9106.

383

2,4-Bis(5-bromo-1H-indol-3-yl)thiazole (1e). Light-brown solid; yield 96%; mp

384


385

1.7 Hz, 1H), 8.47 (d, J = 1.7 Hz, 1H), 8.21 (d, J = 2.8 Hz, 1H), 8.03 (d, J = 2.6 Hz, 1H),

386

7.68 (s, 1H), 7.49 (d, J = 8.6 Hz, 1H), 7.46 (d, J = 8.6 Hz, 1H), 7.38 (dd, J = 8.6, 1.9 Hz,

387

1H), 7.31 (dd, J = 8.6, 1.9 Hz, 1H). 13C NMR (100 MHz, DMSO-d6) δ 162.1, 150.6,

388

135.8, 135.7, 128.5, 127.1, 126.5, 126.4, 125.5, 124.6, 123.3,123.1, 114.7, 114.3, 113.9,

389

112.9, 111.3, 110.8, 107.3. C19H12Br2N3S [M+H]+ 471.9113, found (ESI+) 471.9101.

390

4-(5-Bromo-1H-indol-3-yl)-2-(6-bromo-1H-indol-3-yl)thiazole (1f). Light-brown solid;

391

yield 83%; mp 219−221 °C; 1H NMR (400 MHz, DMSO-d6) δ 11.88 (s, 1H), 11.64 (s,

392

1H), 8.46 (s, 1H), 8.36 (d, J = 8.5 Hz, 1H), 8.18 (d, J = 2.6 Hz, 1H), 8.06 (d, J = 2.3 Hz,

393

1H), 7.72 (d, J = 1.1 Hz, 1H), 7.69 (s, 1H), 7.46 (d, J = 8.6 Hz, 1H), 7.37 (dd, J = 8.5, 1.5

394

Hz, 1H), 7.31 (dd, J = 8.6, 1.7 Hz, 1H). 13C NMR (100 MHz, DMSO-d6) δ 161.5, 149.9,

395

137.5, 135.2, 127.4, 126.5, 126.1, 124.0, 123.5, 123.4, 122.6, 122.2, 115.0, 114.8, 113.9,

396

112.4, 110.9,110.8, 106.9. C19H12Br2N3S [M+H]+ 471.9113, found (ESI+) 471.9121.

397

2,4-Bis(5-methoxy-1H-indol-3-yl)thiazole (1g). Light-brown solid; yield 91%; mp

398


399

2.7 Hz, 1H), 7.97 (d, J = 2.3 Hz, 1H), 7.93 (d, J = 2.5 Hz, 1H), 7.79 (d, J = 2.2 Hz, 1H), 20


Page 20 of 51

Page 21 of 51


400

7.56 (s, 1H), 7.42 (d, J = 8.8 Hz, 1H), 7.38 (d, J = 8.8 Hz, 1H), 6.92 (dd, J = 8.8, 2.4 Hz,

401

1H), 6.86 (dd, J = 8.8, 2.3 Hz, 1H), 3.88 (s, 3H), 3.87 (s, 3H). 13C NMR (100 MHz,

402

DMSO-d6) δ 162.0, 154.6, 153.9, 150.7, 131.8, 131.7, 126.8, 125.3, 125.1, 125.0, 112.8,

403

112.4, 112.0, 111.3, 111.2, 110.7, 105.2, 102.8, 102.7, 55.5, 55.4. C21H18N3O2S [M+H]+

404

376.1114, found (ESI+) 376.1112.

405

2-(6-Bromo-1H-indol-3-yl)-4-(5-methoxy-1H-indol-3-yl)thiazole (1h). Light-brown

406


407

(s, 1H), 8.40 (d, J = 8.5 Hz, 1H), 8.15 (s, 1H), 7.93 (s, 1H), 7.75 (s, 1H), 7.70 (s, 1H),

408

7.58 (s, 1H), 7.35 (m, 2H), 6.83 (d, J = 8.7 Hz, 1H), 3.88 (s, 3H). 13C NMR (100 MHz,

409

DMSO-d6) δ 161.2, 154.0, 150.9, 137.5, 131.7, 127.3, 125.2, 125.1, 123.5, 123.4, 122.3,

410

115.0, 114.8, 112.5,111.8, 111.1, 111.0, 105.8, 101.9, 99.5, 55.3. C20H15BrN3OS [M+H]+

411

424.0114, found (ESI+) 424.0118.

412

4-(6-Bromo-1H-indol-3-yl)-2-phenylthiazole (1i). White solid; yield 94%; mp 233−234

413

°C; 1H NMR (400 MHz, DMSO-d6) δ 11.61 (s, 1H), 8.21 (d, J = 8.6 Hz, 1H), 8.07 (dd, J

414

= 8.1, 1.4 Hz, 2H), 8.04 (d, J = 2.6 Hz, 1H), 7.88 (s, 1H), 7.68 (d, J = 1.7 Hz, 1H),

415

7.58–7.52 (m, 3H), 7.30 (dd, J = 8.5, 1.8 Hz, 1H). 13C NMR (100 MHz, DMSO-d6) δ

416

166.0, 151.4, 137.4, 133.2, 130.1, 129.2, 126.1, 125.8, 123.7, 122.6, 122.1, 114.42,

417

114.36, 111.0, 110.6. C17H12BrN2S [M+H]+ 354.9899, found (ESI+) 354.9894.

418

4-(6-Bromo-1H-indol-3-yl)-2-(4-fluorophenyl)thiazole (1j). White solid; yield 97%;

419

mp 220−221 °C; 1H NMR (400 MHz, DMSO-d6) δ 11.60 (s, 1H), 8.19 (d, J = 8.5 Hz, 1H),

420

8.12 (d, J = 6.0 Hz, 2H), 8.02 (s, 1H), 7.87 (s, 1H), 7.66 (s, 1H), 7.40 (t, J = 8.4 Hz, 2H), 21



421

7.28 (d, J = 8.5 Hz, 1H). 13C NMR (100 MHz, DMSO-d6) δ 165.4, 163.6 (d, J = 246.0

422

Hz), 151.8, 137.9, 130.3 (d, J = 3.1 Hz), 128.9 (d, J = 8.6 Hz), 126.3, 124.2, 123.1, 122.6,

423

116.8 (d, J = 22.0 Hz), 114.9, 114.8, 111.4, 111.2. C17H11BrFN2S [M+H]+ 372.9805,

424

found (ESI+) 372.9800.

425

4-(6-Bromo-1H-indol-3-yl)-2-(2-(trifluoromethyl)phenyl)thiazole (1k). White solid;

426


427

8.5 Hz, 1H), 8.04–7.97 (m, 3H), 7.90–7.76 (m, 3H), 7.69 (s, 1H), 7.27 (d, J = 8.4 Hz, 1H).

428

13C

429

5.5 Hz), 126.3, 124.3(q, J = 272.1 Hz), 124.2, 123.0, 122.5, 114.9, 113.0, 111.4.

430

C18H11BrF3N2S [M+H]+ 422.9733, found (ESI+) 422.9767.

431

4-(6-Bromo-1H-indol-3-yl)-2-(3,4-dichlorophenyl)thiazole (1l). White solid; yield 99%;

432


433

8.16 (d, J = 8.6 Hz, 1H), 8.06 (s, 1H), 8.03 (dd, J = 8.4, 2.1 Hz, 1H), 7.97 (s, 1H), 7.82 (d,

434

J = 8.4 Hz, 1H), 7.66 (d, J = 1.7 Hz, 1H), 7.29 (dd, J = 8.5, 1.8 Hz, 1H). 13C NMR (100

435

MHz, DMSO-d6) δ 163.2, 151.7, 137.4, 133.5, 132.4, 132.0, 131.4, 127.3, 126.1, 126.1,

436

123.6, 122.6, 121.9, 114.5, 114.4, 111.7, 110.7. C17H10BrCl2N2S [M+H]+ 422.9120,

437

found (ESI+) 442.9122.

438

4-(6-Bromo-1H-indol-3-yl)-2-(4-phenoxyphenyl)thiazole (1m). White solid; yield 90%;

439


440

8.06 (d, J = 8.2 Hz, 2H), 7.99 (s, 1H), 7.82 (s, 1H), 7.65 (s, 1H), 7.46 (t, J = 7.7 Hz, 2H),

441

7.29–7.19 (m, 2H), 7.14 (t, J = 6.9 Hz, 4H). 13C NMR (100 MHz, DMSO-d6) δ 165.5,

NMR (100 MHz, DMSO-d6) δ 163.2, 151.5, 137.9, 133.2, 132.7, 130.7, 127.5 (q, J =

22


Page 22 of 51

Page 23 of 51


442

158.5, 155.8, 151.2, 137.40, 130.2, 128.4, 128.0, 125.8, 124.2, 123.7, 122.6, 122.1, 119.3,

443

118.6, 114.4, 114.4, 111.1, 110.2. C23H16BrN2OS [M+H]+ 447.0161, found (ESI+)

444

447.0157.

445

4-(6-Bromo-1H-indol-3-yl)-2-(4-hexylphenyl)thiazole (1n). White solid; yield 83%; mp

446

79−80 °C; 1H NMR (400 MHz, CDCl3) δ 8.38 (s, 1H), 7.90 (d, J = 8.8 Hz, 1H), 7.68 (d, J

447

= 2.8 Hz, 1H), 7.55 (d, J = 1.6 Hz, 1H), 7.32 (dd, J = 8.4, 1.6 Hz, 1H), 7.22 (s, 1H),

448

3.11–3.02 (m, 2H), 1.89–1.81 (m, 2H), 1.49–1.42 (m, 2H), 1.39–1.28 (m, 4H), 0.90 (t, J

449

= 6.9 Hz, 3H).

450

118.6, 116.3, 110.7, 109.1, 107.9, 104.5, 28.3, 26.3, 24.9, 23.6, 17.3, 8.8. C17H20BrN2S

451

[M+H]+363.0525, found (ESI+) 363.0533.

452

4-(6-Bromo-1H-indol-3-yl)-2-(p-tolyl)thiazole (1o). White solid; yield 87%; mp

453


454

7.88 (s, 1H), 7.62 (d, J = 1.7 Hz, 1H), 7.61 (s, 1H), 7.23 (dd, J = 8.5, 1.8 Hz, 1H), 2.72 (s,

455

3H). 13C NMR (100 MHz, DMSO-d6) δ 164.6, 149.6, 137.4, 125.4, 123.6, 122.4, 121.9,

456

114.3, 114.2, 111.2, 109.6, 18.8. C12H10BrN2S [M+H]+ 292.9743, found (ESI+) 292.9736.

457

4-(6-Bromo-1H-indol-3-yl)-2-(4-(tert-butyl)phenyl)thiazole (1p). Brown solid; yield

458

78%; mp 129−131 °C; 1H NMR (400 MHz, DMSO-d6) δ 11.49 (s, 1H), 8.09 (d, J = 8.5

459

Hz, 1H), 7.88 (d, J = 2.4 Hz, 1H), 7.63 (s, 1H), 7.62 (d, J = 1.3 Hz, 1H), 7.23 (dd, J = 8.5,

460

1.5 Hz, 1H), 1.46 (s, 9H). 13C NMR (100 MHz, DMSO-d6) δ 179.2, 149.5, 137.3, 125.3,

461

123.8, 122.4, 122.1, 114.3, 114.2, 111.42, 108.9, 37.2, 30.6. C15H16BrN2S [M+H]+

462

335.0212, found (ESI+) 335.0213.

13C

NMR (100 MHz, DMSO-d6) δ 165.9, 144.3, 132.0, 118.9, 118.7,

23



463

General Procedures for the Preparation of Compounds 14a−14b. To a solution of

464

indoles 3a−3b (20 mmol) in anhydrous ether was added (COCl)2 (26 mmol) dropwise at

465

0 °C. The mixture was stirred for 1.5 h, and filtered to give 13a−13b, which were used

466

for the next step directly. To a solution of compounds 13a−13b in anhydrous ether was

467

added ammonia dropwise at room temperature. The mixture was stirred for 5 min, and

468

then filtered. Filter cake was washed with water and cold anhydrous ether to provide

469

14a−14b.

470

1H-Indole-3-oxacin (14a). Yellow solid; yield 84%; mp 225−256 °C; 1H NMR (400

471

MHz, DMSO-d6) δ 12.21 (s, 1H), 8.69 (d, J = 3.2 Hz, 1H), 8.27–8.18 (m, 1H), 8.09 (s,

472

1H), 7.73 (s, 1H), 7.54–7.52 (m, 1H), 7.28–7.23 (m, 2H).

473

1H-6-Bromoindole-3-oxacin (14b). Yellow solid; yield 75%; mp 265−266 °C; 1H NMR

474

(400 MHz, DMSO-d6) δ 12.28 (s, 1H), 8.71 (d, J = 2.4 Hz, 1H), 8.14 (d, J = 8.8 Hz, 1H),

475

8.12 (s, 1H), 7.76 (s, 1H), 7.73 (d, J = 1.2 Hz, 1H), 7.40 (dd, J = 8.4, 1.6 Hz, 1H).

476

General Procedures for the Preparation of Compounds 15a–15b. To a solution of

477

compounds 14a−14b (0.5 mmol) in DMF was added SOCl2 (0.75 mmol) dropwise under

478

inert atmosphere. The mixture was stirred for 30 min at 0 ℃, then poured into 50 mL

479

water and extracted with EtOAc (3 × 30 mL). The combined organic phase was washed

480

with brine (3 mL), dried over anhydrous Na2SO4, filtered, and concentrated in vacuo to

481

give 15a−15b.

482

1H-Indole-3-carbonyl cyanide (15a). Yellow solid; yield 82%; mp 162−163 °C; 1H

483

NMR (400 MHz, DMSO-d6) δ 12.94 (s, 1H), 8.65 (d, J = 2.8 Hz, 1H), 8.05 (d, J = 7.6 Hz, 24


Page 24 of 51

Page 25 of 51


484

1H), 7.60 (d, J = 7.6 Hz, 1H), 7.41–7.31 (m, 2H).

485

1H-6-Bromoindole-3-carbonyl cyanide (15b). Yellow solid; yield 71%; mp 131−133

486

°C; 1H NMR (400 MHz, DMSO-d6) δ 12.98 (s, 1H), 8.67 (s, 1H), 7.96 (d, J = 8.4 Hz, 1H),

487

7.77 (d, J = 1.6 Hz, 1H), 7.48 (dd, J = 8.4, 1.6 Hz, 1H).

488

Preparation of 2-amino-1-(1H-indol-3-yl) ethan-1-one acetate (16). To a solution of

489

compound 15a (1.47 mmol) in AcOH was added Pb/C. The mixture was stirred for 1 h at

490

room temperature in a hydrogen atmosphere under atmospheric pressure, and

491

concentrated in vacuo, recrystallized with ether to give compound 16. Yellow solid; yield

492

78%; mp 146−148 °C; 1H NMR (400 MHz, DMSO-d6) δ 8.35 (s, 1H), 8.22–8.15 (m, 1H),

493

7.53–7.45 (m, 1H), 7.26–7.16 (m, 2H), 3.98 (s, 2H), 1.86 (s, 3H). 13C NMR (100 MHz,

494

DMSO-d6) δ 192.4, 172.9, 137.0, 134.3, 125.7, 123.3, 122.3, 121.5, 114.2, 112.7, 46.9,

495

22.2.

496

General Procedures for the Preparation of Compounds 17a−17b. To a solution of

497

compound 16 (1 mmol) in DCM was added TEA. After stirred for 10 minutes,

498

compounds 15a−15b were added in two portions. The resulting mixture was allowed to

499

stir for 5 h at room temperature. Then the mixture was diluted with water and extracted

500

with dichloromethane (3 × 50 mL). The combined organic phase was washed with brine

501

(3 mL), dried over anhydrous Na2SO4, filtered, and concentrated in vacuo to give

502

compounds 17a−17b.

503

N-(2-(1H-Indol-3-yl)-2-oxoethyl)-1H-indole-3-carboxamide (17a). Yellow solid; yield

504

98%; mp 285−287 °C; 1H NMR (400 MHz, DMSO-d6) δ 12.03 (s, 1H), 11.61 (s, 1H), 25



Page 26 of 51

505

8.52 (d, J = 2.8 Hz, 1H), 8.23 (t, J = 5.6 Hz, 1H), 8.19 (d, J = 7.6 Hz, 1H), 8.15 (d, J =

506

8.0 Hz, 1H), 8.13 (d, J = 2.8 Hz, 1H), 7.50 (d, J = 7.6 Hz, 1H), 7.45 (d, J = 8.0 Hz, 1H),

507

7.27–7.19 (m, 2H), 7.18–7.06 (m, 2H), 4.66 (d, J = 5.6 Hz, 2H).

508

DMSO-d6) δ 191.2, 164.7, 136.4, 136.1, 133.5, 128.1, 126.0, 125.5, 122.8, 121.8, 121.8,

509

121.2, 120.9, 120.4, 114.2, 112.1, 111.8, 110.4, 45.6. C19H16N3O2 [M+H]+ 318.1237,

510

found (ESI+) 318.1240.

511

N-(2-(1H-Indol-3-yl)-2-oxoethyl)-6-bromo-1H-indole-3-carboxamide (17b). Yellow

512


513

(s, 1H), 8.52 (s, 1H), 8.34 (t, J = 5.6 Hz, 1H), 8.23–8.14 (m, 2H), 8.09 (d, J = 8.6 Hz, 1H),

514

7.66 (s, 1H), 7.50 (d, J = 7.6 Hz, 1H), 7.23 (m, 3H), 4.65 (d, J = 5.6 Hz, 2H). 13C NMR

515

(100 MHz, DMSO-d6) δ 191.0, 164.3, 137.0, 136., 133.5, 128.8, 125.4, 125.2, 123.3,

516

122.8, 122.7, 121.8, 121.2, 114.6, 114.5, 114.1, 112.2, 110.6, 45.6. C19H15BrN3O2

517

[M+H]+ 396.0342, found (ESI+) 396.0342.

518

General Procedures for the Preparation of Compounds 12a−12b. The compounds

519

17a−17b (1 mmol) was dissolved in POCl3. The resulting mixture was heated to reflux

520

and stirred for about 1 h. After the reaction was completed, the mixture was poured into

521

water, extracted with ethyl acetate and purified via column chromatography (PE: EA = 2:

522

1).

523

2,5-Di(1H-indol-3-yl)oxazole (12a). Yellow solid; yield 84%; mp 249–251 °C. 1H NMR

524

(400 MHz, DMSO-d6) δ 11.79 (s, 1H), 11.60 (s, 1H), 8.28 (d, J = 6.8 Hz, 1H), 8.14 (d, J

525

= 2.0 Hz, 1H), 7.96 (d, J = 7.6 Hz, 1H), 7.90 (d, J = 1.6 Hz, 1H), 7.59–7.44 (m, 3H), 26


13C

NMR (100 MHz,

Page 27 of 51


526

7.30–7.14 (m, 4H). 13C NMR (100 MHz, DMSO-d6) δ 156.8, 145.4, 136.4, 136.4, 126.2,

527

124.2, 123.5, 122.9, 122.3, 122.1, 120.6, 120.4, 120.0, 119.6, 112.1, 112.1, 104.1, 104.1.

528

C19H14N3O [M+H]+ 300.1131, found (ESI+) 300.1129.

529

2-(6-Bromo-1H-indol-3-yl)-5-(1H-indol-3-yl)oxazole (12b). Yellow solid; yield 76%;

530

mp 259–260 °C; 1H NMR (400 MHz, DMSO-d6) δ 11.90 (s, 1H), 11.61 (s, 1H), 8.21 (d, J

531

= 7.6 Hz, 1H), 8.17 (s, 1H), 7.96 (d, J = 6.4 Hz, 1H), 7.91 (s, 1H), 7.72 (s, 1H), 7.51 (m,

532

2H), 7.37 (d, J = 7.2 Hz, 1H), 7.28–7.13 (m, 2H).

533

156.7, 146.2, 137.7, 136.91, 127.6, 124.0, 123.8, 123.6, 122.7, 122.6, 120.6, 120.1, 115.6,

534

115.2, 112.6, 104.8, 104.5. C19H13BrN3O [M+H]+ 378.0237, found (ESI+) 378.0236.

535

Biological Assay. Each test was repeated three times at 25±1 °C. Active effect expressed

536

in percentage scale of 0−100 (0: no activity; 100: total inhibited).

537

Specific steps for the anti-TMV30 and fungicidal31 activities were carried out in

538

accordance with the literature method, also can be seen in Supporting Information.

539

Mode of Action Studies. The mode of action studies were carried out in accordance with

540

the literature method,17 also can be seen in Supporting Information.

541

RESULTS AND DISCUSSION

13C

NMR (100 MHz, DMSO-d6) δ

542

Chemistry.

543

Till now, only one synthetic method of the corresponding bis-(indolyl)thiazole

544

analogues has been reported by Jiang24 and Parrino25. However, it has some shortcomings,

545

namely low yields and many steps. The route needs improvement. As shown in Figure 3,

546

commercially available indoles (3a−3c) were reacted with tosyl chloride to give the 27



Page 28 of 51

547

protected indoles (4a−4c), which were treated with acetic anhydride and brominated to

548

provide the key intermediate α-bromoketones (6a−6c). Due to the influence of electronic

549

effect, 5-methoxyindole (3d) can not be converted into 6d by the above steps.

550

3-Acetyl-1H-5-methoxyindole

551

methoxyindole (3d) and dimethylacetamide, was reacted with tosyl chloride to give 5d.

552

Bromination of 5d gave N-tosyl-3-(α-bromoacetyl)-5-methoxyindole (6d). As depicted in

553

Figures 4 and 5, treatment of the indoles (3b−3d) with chlorosulfonyl isocyanate (CSI)

554

gave 3-cyanoindoles (8b−8d). Cyanoindoles (8a−8c) and nitriles 8i−8m were reacted

555

with sodium hydrosulfide hydrate and magnesium chloride hexahydrate to give the key

556

intermediate thioamides (9a−9c and 9i−9m). However, fatty nitriles are not suitable for

557

this method. Intermediate thioamide 9n was prepared from 8n using P2S5 as vulcanization

558

reagent. Cyanoindole (8d) needs to protect NH with tosyl chloride before it can be

559

converted to thioamide (11). As shown in Figures 6 and 7, the protected

560

bis-(indolyl)thiazole compounds (2a−2h) and indole-thiazole compounds (2i−2q) were

561

provided by condensation of the α-bromoketones (6) with the thioamides (9) using

562

Hantzsch reaction.24,25 The removal of the Ts group was carried out under refluxing

563

conditions in a solution of 2 N NaOH to afford bis-(indolyl)thiazole compounds (1a−1h)

564

and indole-thiazole compounds (1i−1p). The corresponding bis-(indolyl)oxazole

565

analogues (12a−12b) were synthesized by Horne’s method (Figures 8−10), which is

566

described in the literature.20

567

(7),

provided

by

Vilsmeier–Haack

reaction

of

Phytotoxic Activity. The phytotoxic-activity tests showed that the nortopsentins 28


Page 29 of 51


568

analogues, namely bis-(indolyl)thiazole compounds 1a−1p, indole-thiazole compounds

569

2a−2q and bis-(indolyl)oxazole compounds 12a, 12b were safe for testing on plants at

570

500 μg/mL. The detailed test procedures can be seen in Supporting Information.

571

Antiviral Activity in vivo. The results of the anti-TMV activities of the nortopsentin

572

analogues containing thiazole 1a−1p and 2a−2q, oxazole 12a−12b are listed in Table 1

573

with nortopsentins A−D and the commercial plant virucide ribavirin as the controls. In

574

order to improve the efficiency of the antiviral activity test, we first test the inactive

575

activities of all compounds listed in Table 1 at 500 μg/mL, and further test the curative

576

activities and protective activities of compounds with good inactive activities (inactive

577

effect > 40%).

578

As depicted in Table 1, the nortopsentin analogues containing thiazole 1a−1p and

579

2a−2q, oxazole 12a−12b showed good antiviral activities in vivo. Most of the analogues

580

exhibited higher in vivo TMV inhibitory effects than ribavirin, especially compounds 1d,

581

1e and 12a. The substitution of sulfur or oxygen atoms for nitrogen atom obviously

582

changed the structure-activity relationship of these compounds. The activities of these

583

alkaloids were significantly increased when the imidazole ring was changed to thiazole

584

ring, except compound 1a (inhibitory effect: 1a < nortopsentin D). Compound 12a

585

containing oxazole ring showed the same level of biological activity as nortopsentin D

586

but higher activity than 1a. Oxazole ring containing compound 12b showed the same

587

level of biological activity as 1b but higher activity than nortopsentin B. The mainly

588

difference between 1a−1h lies in substitutes on indole rings. Compound 1a without 29



589

substituents and 1b, 1c with single substituents in the indole ring showed relatively poor

590

antiviral activities, which are about similar with that of ribavirin. Bibrominated

591

compounds 1d and 1e exhibit excellent antiviral activities. The replacement of bromine

592

atom with methoxy group or two bromine atoms at different positions will lead to the

593

decrease of biological activity (inhibitory effect: 1d and 1e > 1f−1h), which indicated that

594

the substitution effect on indole ring is very obvious. Substitution of aryl or alkyl groups

595

for 6-bromoindole substituents at the 2-position of thiazole resulted in a significant

596

decrease in biological activity. Compounds 1k−1m displayed relatively high biological

597

activity. For bisindole thiazole compounds 1a−1h, substitution of indole ring N-H with

598

Ts maintained or decreased antiviral activity. For thiazole 2-aryl substituted compounds

599

1i−1m, the effect of TS substitution of indole ring N-H on antiviral activity is complex

600

(inhibitory effect: 1i and 1j < 2i−2j respectively, 1k and 1m > 2k−2m respectively). For

601

thiazole 2-alkyl substituted compounds 1n−1p, TS substitution of indole ring N-H is

602

beneficial to antiviral activity. Compounds 1d, 1e and 12a with excellent antiviral

603

activities emerged as novel antiviral lead compounds, among which, 1e was selected for

604

further antiviral mechanism research.

605

Preliminary Mode of Action. The preliminary antiviral mechanism research on

606

compound 1e was carried out via TEM with RNA inhibitor antofine16 and CP disks

607

assembly inhibitor NK020932 as controls. TMV is a single stranded RNA virus. It

608

contains a single stranded RNA encapsulated in 2130 CP monomers, showing about 300

609

nm length.33 The test results revealed that 20S CP Disk can be well formed (Figure 11, A) 30


Page 30 of 51

Page 31 of 51


610

and assembled into full-length virus particles (Figure 11, B) with RNA. The use of a

611

small amount of DMSO does not affect assembly (Figure 11, C). As controls, antofine

612

and NK0209 can significantly inhibit the assembly of TMV rod (Figure 11, D and E).

613

The above experiments verify the feasibility of our method. Although compound 1e did

614

not inhibit the assembly of viruses, it enabled the viruses to gather together (Figure 11, F).

615

Further 20S CP Disk assembly inhibition tests were carried out to evaluate the interaction

616

of 1e with TMV CP. The 20S CP Disk can be well formed (Figure 12, A). A small

617

amount of DMSO (Figure 12, B) and the TMV RNA inhibitor antofine (Figure 12, C)

618

displayed no impact on 20S CP Disk assembly. CP disks assembly inhibitor NK0209

619

showed significant impact on 20S CP Disk (Figure 12, D). Compound 1e can effectively

620

induce CP Disks aggregation and fusion (Figure 12, E). The above results indicated that

621

compound 1e may play an antiviral role by aggregating viral particles to prevent their

622

movement in plants, thus slowing down the spread of viruses.

623

Fungicidal Activity. Thiazole 1a−1p and 2a−2q, oxazole 12a−12b were also

624

evaluated for their fungicidal activities with nortopsentins A−D and commercial

625

fungicides carbendazim and chlorothalonil as controls.

626

In Vitro Fungicidal Activity. The in vitro fungicidal activities of 1a−1p, 2a−2q and

627

12a−12b were evaluated in mycelial growth tests31 conducted in artificial media against

628

14 plant pathogens at a rate of 50 μg/mL. The results showed that these compounds also

629

exhibited broad-spectrum fungicidal activities (Table 2). Compounds 1a−1d displayed

630

about similar level antifungal activities with nortopsentins A−D. Most of these 31



631

compounds showed good antifungal activities against Sclerotinia sclerotiorum.

632

Compounds 2p and 2f displayed higher antifungal activities against Alternaria solani

633

than commercial fungicides carbendazim and chlorothalonil. Compound 2f showed same

634

level of antifungal activity against Fusarium graminearum as commercial fungicide

635

carbendazim. The fungicidal activity of 1o against Phytophthora capsici was same as

636

commercial fungicide carbendazim and higher than that of commercial fungicide

637

chlorothalonil.

638

In Vivo Fungicidal Activity. Compounds 1a−1p, 2a−2q and 12a−12b were further

639

evaluated for their in vivo fungicidal activities with commercial fungicide azoxystrobin as

640

control.31 The pathogens tested in this screen were Sclerotinia sclerotiorum on rape,

641

Rhizoctonia cerealis on cerealis, Phytophthora capsici on capsici. The results (Table 3)

642

revealed that lots of compounds also displayed in vivo fungicidal activities.

643

In summary, nortopsentin analogues 1a−1p, 2a−2q and 12a−12b were designed

644

synthesized and evaluated for their antiviral activities and fungicidal activities. Most of

645

these compounds displayed higher antiviral activities than ribavirin. Compounds 1d, 1e

646

and 12a with significantly higher antiviral activities than ribavirin emerged as new

647

antiviral lead compounds for further research. The substitution of sulfur or oxygen atoms

648

for nitrogen atom obviously changed the structure-activity relationship of these

649

compounds. The preliminary mode of action studies revealed that these compounds may

650

play an antiviral role by aggregating viral particles to prevent their movement in plants.

651

Further fungicidal test revealed that these compounds displayed broad-spectrum 32


Page 32 of 51

Page 33 of 51


652

fungicidal activities against 14 kinds of plant fungi at 50 μg/mL. Compounds 2p and 2f

653

displayed higher antifungal activities against Alternaria solani than commercial

654

fungicides carbendazim and chlorothalonil. Current research has laid a foundation for the

655

application of nortopsentin analogues in plant protection.

656

ASSOCIATED CONTENT

657

Supporting Information

658

The detailed bio-assay procedures. The spectra data of nortopsentin analogues 1a−1p,

659

2a−2q and 12a−12b. This material is available free of charge via the Internet at

660

http://pubs.acs.org.

661

AUTHOR INFORMATION

662

Corresponding Authors

663

*(Z.W.)

664

0086-22-23766531.

665

*(Y.L.)

666

0086-22-23503792.

667

*(Q.W.)

668

0086-22-23503952.

669

Funding

670

This study was supported by Natural Science Fund of China (21772145, 21732002,

671

21772104, 31760527), Training Program of Outstanding Youth Innovation Team of

672

Tianjin Normal Uiversity and the Program for Innovative Research Team in University of

E-mail:

E-mail:

E-mail:

[email protected];

Phone:

0086-22-23766531;

Fax:

[email protected];

Phone:

0086-22-23503792;

Fax:

[email protected].

Phone:

0086-22-23503952.

Fax:

33



673

Tianjin (TD13-5074).

674

Notes

675

The authors declare no competing financial interest.

676

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677

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Figure Captions Figure 1. Structures of Ribavirin and Nortopsentins A–D. 38


Page 38 of 51

Page 39 of 51


Figure 2. Design of Nortopsentin Analogues. Figure 3. Synthesis of Intermediates 6a–6d. Figure 4. Synthesis of Intermediate 9a–9c and 11. Figure 5. Synthesis of Intermediate 9i–9n. Figure 6. Synthesis of 1a–1h. Figure 7. Synthesis of 1i–1p. Figure 8. Synthesis of 15a–15b. Figure 9. Synthesis of 16. Figure 10. Synthesis of 12a–12b. Figure 11. TMV Rod Assembly Inhibition of Compounds 1e, NK0209, and Antofine: (A) 20S CP disk (100 nm scale bar), (B) 20S CP disk + RNA (100 nm scale bar), (C) 20S CP disk + RNA + 1/100 DMSO (200 nm scale bar), (D) 20S CP disk + RNA + 10 μM antofine (200 nm scale bar), (E) 20S CP disk + RNA + 10 μM NK0209 (100 nm scale bar), (F) 20S CP disk + RNA + 10 μM 15 (200 nm scale bar). Figure 12. 20S CP Disk Assembly Inhibition of Compounds 1e, NK0209, and Antofine (100 nm scale bar): (A) CP, (B) CP + 1/100 DMSO, (C) CP + 10 μM antofine, (D) CP + 10 μM NK0209, (E) CP + 10 μM 1e.

Table 1. In Vivo Antiviral Activities of Compounds 1a−1p, 2a−2q and 12a, 12b, Nortopsentins A–D and Ribavirin against TMV. 39



Page 40 of 51

Compd

Concn (μg/mL)

Inactive effect (%)a

Curative effect (%)a

Protective effect (%)a

Compd

Concn (μg/mL)

Inactive effect (%)a

Curative effect (%)a

Protective effect (%)a

1a

500

32±3

—

—

2e

500

33±3

—

—

1b

500 100

44±1 0

37±2 7±1

49±3 14±1

2f

500 100

46±1 10±1

32±3 6±2

38±3 17±1

500

47±1

40±4

44±3

100

11±1

18±1

15±1

2g

500

34±3

—

—

500

59±4

64±2

55±5

100

22±4

30±2

26±1

2h

500

31±3

—

—

500 100 500 100 500 100 500 100

54±3 19±1 47±2 22±3 50±2 9±2 51±1 15±1

50±3 17±1 45±3 14±1 53±3 20±2 38±5 5±3

53±2 12±3 50±2 18±1 38±4 5±1 48±4 11±1

500 100 500 100

43±4 13±1 49±1 6±2

39±3 6±2 33±1 0

44±2 11±1 35±4 10±1

2k

500

36±2

—

—

2l

500

29±3

—

—

1i

500

26±4

—

—

2m

1j

500 500 100 500 100 500 100

37±4 48±3 9±1 48±1 6±3 50±4 20±1

— 42±4 19±1 36±3 0 41±1 8±2

— 40±2 0 38±1 0 46±3 15±2

2n

1n

500

32±1

—

—

12a

500 100 500 500 100 500 100 500 100 500 100

40±1 4±2 39±3 47±3 19±1 43±4 10±1 41±1 8±2 54±3 16±3

35±5 0 — 41±3 11±2 36±1 13±3 45±2 15±1 59±4 21±1

43±2 0 — 50±1 17±2 38±2 0 33±4 5±2 52±4 10±1

1o

500

16±5

—

—

12b

500

33±2

—

—

1p

500

28±2

—

—

Nortopsentin A

500 100

48±3 16±1

51±2 12±1

44±4 10±3

2a

500

19±2

—

—

Nortopsentin B

500

32±2

—

—

500

48±2

43±2

35±5

100

0

7±1

8±2

Nortopsentin C

500

35±1

—

—

2c

500 100

42±2 21±2

58±4 15±4

56±2 21±1

Nortopsentin D

2d

500

20±2

—

—

Ribavirin

500 100 500 100

54±3 26±3 38±1 12±1

47±4 18±1 36±2 11±1

49±1 13±2 39±1 13±1

1c 1d 1e 1f 1g 1h

1k 1l 1m

2b

a

2i 2j

2o 2p 2q

Average of three replicates; All results are expressed as mean ± SD; Activity Data with prominent were presented in

40


Page 41 of 51


pink bold.

Table 2. In Vitro Fungicidal Activities of Compounds 1a−1p, 2a−2q and 12a, 12b, Carbendazim and Chlorothalonil against 14 Kinds of Fungi. Fungicidal activities (%)a/ 50 μg/mL

Compd F.Cb

C.Hb

P.Pb

R.Cb

B.Mb

W.Ab

F.Mb

A.Sb

F.Gb

P.Ib

P.Cb

S.Sb

R.Sb

B.Cb

1a

11±1

10±2

27±2

9±1

2±1

19±2

24±3

13±1

16±2

5±1

3±1

23±2

16±1

14±1

1b

13±1

7±1

36±3

7±1

11±2

30±3

30±1

40±3

11±2

5±1

7±1

12±1

16±2

11±2

1c

9±1

3±1

50±3

9±1

2±1

12±2

30±2

27±2

21±3

5±1

10±1

14±1

16±2

28±2

1d

13±1

0

25±1

6±1

0

23±1

24±2

7±1

21±2

16±1

3±1

23±2

16±1

8±1

1e

26±2

16±1

8±1

37±2

14±2

27±1

11±1

11±1

25±1

17±2

13±2

68±3

19±1

26±2

1f

23±1

11±1

13±1

33±1

17±1

30±2

16±1

22±1

56±2

8±1

5±1

85±2

19±1

48±2

1g

34±2

53±2

25±2

75±1

28±3

37±1

16±1

44±3

31±1

17±1

25±1

66±1

19±1

63±1

1h

29±2

21±1

25±1

45±2

25±1

33±1

21±2

22±2

25±2

8±1

5±1

73±3

30±2

50±2

1i

49±2

47±2

17±1

63±2

33±1

33±2

26±1

22±2

13±1

8±1

13±1

68±2

8±1

9±1

1j

23±1

11±1

4±1

26±1

25±2

27±1

5±1

33±1

25±2

8±1

25±2

73±1

15±1

11±1

1k

26±2

37±2

29±2

53±1

22±1

20±2

16±2

22±1

38±1

8±1

13±1

37±2

12±1

39±2

1l

40±1

42±1

29±1

39±2

14±1

10±1

6±1

33±2

19±1

8±1

5±1

24±1

12±1

9±1

1m

26±2

16±2

13±2

28±1

14±2

30±1

26±2

33±1

13±1

8±1

5±1

61±1

15±1

11±1

1n

4±1

14±1

28±1

4±1

9±1

9±1

6±1

6±1

24±1

33±2

7±1

7±2

31±2

17±1

1o

29±2

16±1

13±2

45±2

47±1

33±3

47±1

44±2

50±2

50±1

90±3

81±2

50±1

85±2

1p

31±1

16±1

17±1

45±1

28±2

33±1

42±2

44±1

25±1

17±1

38±1

78±1

19±1

39±2

2a

9±1

0

38±1

0

0

26±1

18±1

13±1

10±1

5±1

13±1

12±1

5±1

6±1

2b

13±2

10±1

25±2

0

11±1

14±1

21±1

33±1

11±1

5±1

7±1

12±1

24±2

14±1

2c

21±1

3±1

28±1

0

13±1

9±1

24±2

13±2

16±2

5±1

3±1

12±2

16±2

28±2

2d

11±1

7±1

20±1

0

9±1

21±3

42±3

7±1

16±1

5±1

7±1

17±1

24±3

28±1

2e

31±2

11±2

13±2

33±1

28±1

30±1

11±1

44±2

50±1

17±2

25±2

12±1

12±1

48±1

2f

40±1

5±1

46±1

24±1

22±1

40±2

16±1

89±1

88±2

42±1

50±1

37±2

19±1

46±2

2g

40±2

21±1

50±2

33±2

33±2

40±1

21±2

11±1

13±1

8±1

5±1

12±1

19±2

11±1

2h

34±1

21±2

33±1

41±1

25±1

37±1

26±2

11±1

13±1

8±1

5±1

78±1

27±1

15±2

2i

31±2

47±1

21±1

63±2

33±1

43±2

21±1

11±2

25±2

8±1

13±2

12±2

15±2

13±1

2j

23±1

21±1

8±1

41±1

14±2

30±1

32±1

11±1

13±1

8±1

13±1

12±1

8±1

9±1

2k

31±2

42±1

13±1

73±2

19±1

27±2

47±2

11±2

38±1

8±1

5±1

12±2

4±1

7±1

2l

31±1

37±3

38±1

29±1

22±1

13±1

84±1

33±2

25±1

17±1

25±1

46±1

35±2

15±2

41



Page 42 of 51

2m

31±2

21±1

33±1

39±3

33±2

40±2

26±1

11±1

19±2

8±1

5±1

12±1

31±1

17±2

2n

13±1

7±1

42±3

25±1

9±1

5±1

6±1

6±1

12±1

33±3

7±1

7±1

39±2

17±1

2o

26±1

0

4±1

33±2

22±1

23±1

37±2

22±1

38±2

17±1

25±1

63±2

46±1

63±3

2p

31±2

21±1

33±2

28±1

17±1

40±1

16±1

67±1

75±1

33±1

50±1

73±1

27±1

52±1

2q

11±1

16±2

54±1

43±3

22±2

30±2

11±1

11±2

25±1

8±1

5±1

32±2

15±1

17±1

12a

19±1

29±1

47±2

39±1

19±1

28±2

18±2

7±1

24±2

11±1

10±1

25±2

5±1

19±2

12b

13±2

3±1

44±2

6±1

9±1

30±1

18±1

13±1

14±1

16±2

13±2

85±1

36±2

33±1

Nortopsentin A

21±1

13±2

36±1

7±1

13±2

26±1

30±2

13±2

5±1

16±2

3±1

12±1

24±2

22±2

Nortopsentin B

15±2

52±1

47±1

35±3

11±1

28±2

12±1

33±3

10±1

16±1

7±2

45±1

27±2

9±1

Nortopsentin C

28±2

32±3

34±1

11±2

21±1

58±1

21±3

27±1

0

26±2

7±1

32±1

17±3

9±3

Nortopsentin D

26±1

29±2

58±1

26±2

21±1

42±2

9±1

33±1

10±1

16±2

3±1

39±2

14±1

7±1

Carbendazimc

77±2

58±2

54±1

78±2

72±2

90±1

84±1

56±2

88±2

83±1

90±2

100

100

96±1

Chlorothalonilc

97±1

11±2

96±2

98±1

97±2

97±1

79±2

56±1

100

100

55±2

100

27±1

100

aAverage

of three replicates; All results are expressed as mean ± SD. bF.C, Fusarium oxysporum f. sp. cucumeris; C.H,

Cercospora arachidicola Hori; P.P, Physalospora piricola; R.C, Rhizoctonia cerealis; B.M, Bipolaris maydis; W.A, Watermelon anthracnose; F.M, Fusarium moniliforme; A.S, Alternaria solani; F.G, Fusarium graminearum; P.I, Phytophthora infestans; P.C, Phytophthora capsici; S.S, Sclerotinia sclerotiorum; R.S, Rhizoctonia solani; B.C, Botrytis cinerea.. cThe commercial agricultural fungicides were used for comparison of antifungal activity; Activity Data with prominent were presented in bold.

Table 3. In Vivo Fungicidal Activities of Compounds 1a−1p, 2a−2q and 12a, 12b and Azoxystrobin against 3 Kinds of Fungi. compd

inhibition rate (%)a 200 µg/mL 42


Page 43 of 51


S.Sb

R.Sb

P.Cb

1a

12±2

5±1

0

1b

4±1

3±1

8±2

1c

11±1

4±1

9±1

1d

16±2

9±2

0

1e

21±2

13±1

0

1f

32±1

9±2

0

1g

21±1

8±1

0

1h

21±2

8±1

0

1i

18±1

8±1

0

1j

12±2

13±2

5±1

1k

10±1

8±1

5±1

1l

17±2

20±2

0

1m

15±1

4±1

0

1n

8±1

6±1

0

1o

34±1

16±1

30±2

1p

14±2

8±1

10±1

2a

12±2

5±1

12±1

2b

8±1

12±2

8±1

2c

10±1

6±1

10±2

2d

13±2

16±2

13±1

2e

11±1

9±1

10±2

2f

18±2

21±2

30±1

2g

8±1

5±1

0

2h

28±2

16±1

0

2i

8±1

15±2

0

2j

9±2

8±1

0

2k

9±1

4±1

0

2l

14±1

16±1

5±1

2m

8±2

22±2

0

2n

6±1

6±2

0

2o

18±2

18±1

5±1

43



aAverages

Page 44 of 51

2p

23±2

8±1

5±1

2q

9±1

10±2

0

12a

15±2

5±1

11±2

12b

26±2

17±2

13±1

azoxystrobinc

100

100

86±2

of three replicates. bS.S, Sclerotinia sclerotiorum; R.S, Rhizoctonia solani; P.C,

Phytophthora capsici. c The dilution of azoxystrobin is 1000 times.

Figure 1.

44


Page 45 of 51


O

HOH2C

N

HO

R1

N

R2

N

CONH2

N

N H

N H

N H

OH

1

R = R2 = Br: Nortopsentin A R1 = H; R2 = Br: Nortopsentin B R1 = Br; R2 = H: Nortopsentin C R1 = R2 = H: Nortopsentin D

Ribavirin

Figure 2. R2

N R1

N H

Lead discovery

N H

N H R1 = R2 = Br: Nortopsentin A R1 = H; R2 = Br: Nortopsentin B R1 = Br; R2 = H: Nortopsentin C R1 = R2 = H: Nortopsentin D Structural

S N

Br

R

optimization

R

S R

N

N H

O

N H

N H

N H

New mechanism discovery

TMV RNA

N

1e

TMV CP

Figure 3.

45


R N H


Page 46 of 51

O

O TsCl, NaH

R1 N H

CuBr2

AA, AlCl3

R1 N Ts

MeCN, r.t.

R1

DCM, r.t.

4a 99% 4b 99% 4c 99%

3a 3b 3c

O MeO

POCl3 N H

DMA 0 oC-90oC

N Ts 6a 85% 6b 81% 6c 88% 6d 62%

O

DCM, r.t.

N H

3d

R1

5a 99% 5b 96% 5c 98%

TsCl, 4-DMAP Et3N MeO

MeO

EA/CCl3H reflux

N Ts

Br

N Ts

7 51%

5d 82%

a: R1 = H; b: R1 = 6-Br; c: R1 = 5-Br; d: R1 = 5-OMe

Figure 4. S NaHS, MgCl2 6H2O

R2 N H

DMF, r.t. CN CSI

R2 N H

DMF, Ar -50- -10 oC

8a 8b 8c

9a 89% 9b 87% 9c 85%

R2 N H 8d

3b 3c 3d

NH2

8b 99% 8c 97% 8d 99%

TsCl, 4-DMAP, Et3N

MeO

CN

N Ts 10 88%

DCM, r.t.

a: R2 = H; b: R2 = 6-Br; c: R2 = 5-Br; d: R2 = 5-OMe

Figure 5.

46


S NaHS, MgCl2 6H2O DMF, r.t.

MeO N Ts 11 85%

NH2

Page 47 of 51


S

NaHS, MgCl2 6H2O 8i-m

N

R2

DMF, r.t.

9i-m 76%--94%

2

R

8n

8i-n

NH2

S

P2S5, EtOH

R2

r.t.

NH2

9n 35% j:

i:

R2:

k:

F

CF3

l:

m:

Cl

O

n: 5

Cl

Figure 6. R2

O

S

Br +

1

R

2

R

N R3

N Ts 1

6a R = H 6a R1 = H 6b R1 = 6-Br 6b R1 = 6-Br 6c R1 = 5-Br 6c R1 = 5-Br 6d R1 = 5-OMe 6d R1 = 5-OMe

2

3

NH2

S

EtOH reflux

9a R = R = H 9b R2 = 6-Br; R3 = H 9a R2 = H; R3 = H 9b R2 = 6-Br; R3 = H 9c R2 = 5-Br; R3 = H 9b R2 = 6-Br; R3 = H 11 R2 = 5-OMe; R3 = Ts 9b R2 = 6-Br; R3 = H

R2

N R3 N

R1

MeOH reflux

N Ts 1

S

NaOH

NH

N

R1 N H

2

3

2a 97% R = R = R = H 2b 96% R1 = R3 = H; R2 = 6-Br 2c 84% R1 = 6-Br; R2 = R3 = H 2d 85% R1 = R2 = 6-Br; R3 = H 2e 78% R1 = R2 = 5-Br; R3 = H 2f 91% R1 = 5-Br; R2 = 6-Br; R3 = H 2g 79% R1 = R2 = 5-OMe; R3 = Ts 2h 80% R1 = 5-OMe; R2 = 6-Br; R3 = H

Figure 7.

47


1a 88% R1 = R2 = R3 = H 1b 76% R1 = R3 = H; R2 = 6-Br 1c 97% R1 = 6-Br; R2 = R3 = H 1d 88% R1 = R2 = 6-Br; R3 = H 1e 89% R1 = R2 = 5-Br; R3 = H 1f 93% R1 = 5-Br; R2 = 6-Br; R3 = H 1g 81% R1 = R2 = 5-OMe; R3 = Ts 1h 86% R1 = 5-OMe; R2 = 6-Br; R3 = H


O

Br

S EtOH

S + N Ts

Br

j:

i:

reflux

NH2

S

R2 NaOH

N 2i-2p

Br N Ts

6b

R2:

R2

9i-9q

k:

m:

MeOH reflux

Br N H 1i-1p

n:

Cl

CF3

R2 N

2i-2q

l:

F

Page 48 of 51

o:

p:

O

5

O

O

q:

Cl

Figure 8. O

0 oC

N H

O O

(COCl)2 , Et2O R

Cl

O

sat.aq. NH4Cl, KOH R

3a 3b

N H

r.t.

13a 13b

R

N H 14a 84% 14b 75%

a: R = H; b: R = Br

Figure 9. O

O

CN

NH2

Pd/C, H2 N H 15a

MeOH-HOAc, r.t.

NH2

HOAc N H 16 78%

Figure 10.

48


O SOCl2

Argon, DMF, R 0 oC

N H 15a 82% 15b 71%

CN

Page 49 of 51


O

R

NH2

O

16

O

Et3N

HAc + N H

CN

N H

R 15a 15b

DCM, r.t.

HN

R H N

N

POCl3 NH

O 17a 98% 17b 96%

a: R = H; b: R = Br

Figure 11.

49


reflux

O HN 12a 84% 12b 76%

NH


Figure 12.

50


Page 50 of 51

Page 51 of 51


TOC graphic

Agrochemical Bioregulators R2

N R1 Lead discovery

N H

N H

N H R1 = R2 = Br: Nortopsentin A R1 = H; R2 = Br: Nortopsentin B R1 = Br; R2 = H: Nortopsentin C R1 = R2 = H: Nortopsentin D Structural

S N

Br

R

optimization

R

S R

N

N H

O

N H

N H

N H

New mechanism discovery

TMV RNA

N

1e

TMV CP

51


R N H

Optimization, Structure-Activity Relationship and Mode of Action of

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