Discovery of Pimprinine Alkaloids as Novel Agents against a Plant

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Agricultural and Environmental Chemistry

Discovery of Pimprinine Alkaloids as Novel Agents against a Plant Virus Bin Liu, Rui Li, Yanan Li, Songyi Li, Jin Yu, Binfen Zhao, Ancai Liao, Ying Wang, Ziwen Wang, Aidang Lu, Yuxiu Liu, and Qingmin Wang J. Agric. Food Chem., Just Accepted Manuscript • DOI: 10.1021/acs.jafc.8b06175 • Publication Date (Web): 25 Jan 2019 Downloaded from http://pubs.acs.org on January 27, 2019

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Journal of Agricultural and Food Chemistry

Discovery of Pimprinine Alkaloids as Novel Agents against a Plant Virus

Bin Liu,† Rui Li,† Yanan Li,† Songyi Li,† Jin Yu,† Binfen Zhao,† Ancai Liao,† Ying Wang,† Ziwen Wang,†,* Aidang Lu,‡,* Yuxiu Liu§ and 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 Normal University, Tianjin 300387, China

‡

School of Chemical Engineering and Technology, Hebei University of Technology,

Hebei Collaborative Innovation Center of Modern Marine Chemical Technology, Tianjin 300130, 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

* To whom correspondence should be addressed. For Ziwen Wang, E-mail: [email protected]; Phone:

0086-22-23766531;

Fax:

0086-22-23766531;

For

Aidang

[email protected]; Phone: 0086-22-60302812; Fax: 0086-22-60204274.

1

ACS Paragon Plus Environment

Lu,

E-mail:


1

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ABSTRACT: Plant viral diseases cause tremendous decreases in yield and quality.

3

Natural products have always been valuable source for lead discovery in medicinal and

4

agricultural chemistry. A series of pimprinine alkaloids and their derivatives were

5

prepared and identified by nuclear magnetic resonance (NMR) and high-resolution

6

mass spectrometer (HR-MS). The antiviral activities of these alkaloids against tobacco

7

mosaic virus (TMV) were systematically investigated for the first time. Most of the

8

compounds exhibited higher antiviral activities than ribavirin. Compounds 5l, 9h and

9

10h with similar or higher antiviral activities than ningnanmycin (perhaps the most

10

widely used antiviral agent at present) emerged as new antiviral pilot compounds. The

11

systematically structure-activity relationship research lays a foundation for simplifying

12

the structure of these alkaloids. As the ring open products, acylhydrazones 9a–9u were

13

also found to possess good antiviral activities. Moreover, all the synthesized

14

compounds displayed broad-spectrum fungicidal activities. This study provides

15

important information for the research and development of pimprinine alkaloids as

16

novel antiviral agents.

17

2


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KEYWORDS: natural product, alkaloid, pimprinine, anti-TMV activity, fungicidal

20

activity

21

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INTRODUCTION

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The recent population survey reveals that the population growth rate far exceeds

25

the growth rate of land production capacity, which will lead to the urgent need to

26

increase food production.1,2 Plant viruses often drastically damage crops and cause

27

serious reductions in crop yield and quality. Tobacco Mosaic Virus (TMV) is the

28

earliest discovered plant virus and widely used as a model virus in the study of new

29

antiviral agents. It is known to infect more than 400 crops, leading to yield reduction or

30

abortion.3 The management of these viruses has puzzled scientists for years. Few

31

chemical agents for controlling viruses have been commercialized as effective products.

32

As the most widely used antiviral agent at present, ribavirin and ningnanmycin can only

33

show moderate field control effect. The discovery of novel antiviral agents with

34

different scaffolds or modes of action are needed urgently.

35

Natural products and their derivatives have always been valuable source for lead

36

discovery in medicinal and agricultural chemistry, because their novel scaffolds can

37

provide different action modes from the existing agents.4 Some natural products have 3



38

been commercialized5,6 or found as lead compounds7–13 for plant virus control.

39

Pimprinine is an indole alkaloid first isolated from the filtrates of Streptomyces

40

pimprina cultures in 1963. Members of this family (Figure 1) display a range of

41

biological activities, such as anti-epileptic effect14,15, platelet aggregation inhibitory

42

effect16, anti-tumor activity17, and fungicidal activity18. Because of the low natural

43

content and difficult synthesis of these alkaloids, the research on their biological

44

activities is not deep enough, and there is no report on their anti-plant viral activities at

45

present.

46

Our research group has been devoted to the development of new antiviral agents

47

based on natural products for a long time.10–13 In this work, pimprinine was taken as the

48

parent structure to carry out structural optimization for discovering simpler structure

49

analogues with higher activity. As shown in Figure 2, we designed and synthesized a

50

series of pimprinine analogues in which the indole nitrogen, 2-position of oxazole,

51

oxazole ring have been derivatised. The antiviral and fungicidal activities of these

52

compounds were systematically studied.

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MATERIALS AND METHODS

54

Chemicals. All the reagents used are purchased analytical pure reagents.

55

Instruments. The melting points of the synthesized compounds were determined on an

56

X-4 binocular microscope (Beijing Tech Instruments Co., Beijing, China) without

57

temperature correction. 1H NMR and 13C NMR spectra of the compounds were obtained

58

on Bruker AV 400 spectrometer (Bruker Corp., Switzerland) in CDCl3 or DMSO-d6 4


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59

with tetramethylsilane as the internal standard. High-resolution mass spectra were

60

obtained with an FT-ICR MS spectrometer (Ionspec, 7.0 T).

61

Preparation of Compound 2-(1H-Indol-3-yl)-2-oxoacetyl Chloride (2). The ether

62

solution (100 mL) of oxaloyl chloride (0.22 mol) was added to the ether solution (350

63

mL) of indole (0.17 mol) dropwise while maintaining the temperature at 0 oC. The

64

yellowish mixture was stirred at 0 oC for 90 min and filtered, washed with cold ether to

65

give 2 (yield 90%) as a yellow solid, which was directly used for next reaction without

66

purification.

67

Preparation of Compound 2-(1H-Indol-3-yl)-2-oxoacetaldehyde (3). Under Ar

68

atmosphere, the ethyl acetate solution (150 mL) of tributyltin hydride (0.15 mol) was

69

added to ethyl acetate solution (150 mL) of oxoacetyl chloride 2 (0.15 mol) drop by

70

drop at 0 °C. The reaction mixture was stirred at 0 °C for 30 min and at 25 °C for 15 h.

71

To the mixture was added petroleum ether (700 mL), then filtered to afford 2-

72

oxoacetaldehyde 3 as a yellow powder in 80% yield, which was directly used for next

73

reaction without purification.

74

General Methods for Synthesis of Compounds 5a–5l. The mixture of 2-

75

oxoacetaldehyde 3 (5.8 mmol), corresponding amino acids (11.6 mmol) and I2 (5.8

76

mmol) in dimethyl sulfoxide (DMSO, 30 mL) was stirred at 110 °C for 30 min, then

77

added 5% solution of Na2S2O3 (300 mL) and ethyl acetate (400 mL). Then the reaction

78

mixture was stirred for 30 min and separated. The organic layer was washed with

79

saturated NaCl solution (200 mL × 2), dried with anhydrous Na2SO4, filtered and 5



80

concentrated in vacuum. The compounds 5a–5l were obtained by column

81

chromatography on silica gel.

82

Pimprinine (5a). Brown solid; mp. 202−203 oC (lit.14, 204−205 oC); yield 66%; 1H

83

NMR (400 MHz, DMSO-d6) δ 11.54 (s, 1H, NH), 7.84 (d, J = 7.9 Hz, 1H, Ar-H), 7.74

84

(d, J = 2.6 Hz, 1H, Ar-H), 7.47 (d, J = 8.0 Hz, 1H, Ar-H), 7.30 (s, 1H, Ar-H), 7.12–

85

7.23 (m, 2H, Ar-H), 2.49 (s, 3H, CH3); 13C NMR (100 MHz, DMSO-d6) δ 158.2, 147.3,

86

136.3, 123.5, 122.8, 122.0, 120.0, 119.4, 119.2, 112.0, 103.9, 13.6; HRMS (ESI) calcd

87

for C12H11N2O (M+H)+ 199.0866, found 199.0861.

88

5-(1H-Indol-3-yl)oxazole (5b). Brown solid; mp. 152−154 oC (lit.19, 156−157 oC);

89

yield 68%; 1H NMR (400 MHz, DMSO-d6) δ 11.63 (s, 1H, NH), 8.36 (s, 1H, Ar-H),

90

7.88 (d, J = 5.9 Hz, 1H, Ar-H), 7.83 (s, 1H, Ar-H), 7.49 (s, 2H, Ar-H), 7.17–7.23 (m,

91

2H, Ar-H); 13C NMR (100 MHz, DMSO-d6) δ 149.6, 147.6, 136.4, 123.5, 122.1, 120.1,

92

119.4, 118.7, 112.1, 103.6.

93

2-(tert-Butyl)-5-(1H-indol-3-yl)oxazole (5c). Brown solid; mp. 172−174 oC; yield

94

57%; 1H NMR (400 MHz, DMSO-d6) δ 11.56 (s, 1H, NH), 7.84 (d, J = 7.8 Hz, 1H, Ar-

95

H), 7.77 (d, J = 2.6 Hz, 1H, Ar-H), 7.48 (d, J = 7.9 Hz, 1H, Ar-H), 7.26 (s, 1H, Ar-H),

96

7.14–7.23 (m, 2H, Ar-H), 1.41 (s, 9H, CH3); 13C NMR (100 MHz, DMSO-d6) δ 167.7,

97

147.1, 136.3, 123.6, 123.0, 122.0, 120.0, 119.4, 118.8, 112.0, 104.0, 33.2, 28.5; HRMS

98

(ESI) calcd for C15H17N2O (M+H)+ 241.1335, found 241.1339.

99

5-(1H-Indol-3-yl)-2-isopropyloxazole (5d). Brown solid; mp. 125−126 oC (lit.19,

100

125−126 oC); yield 72%; 1H NMR (400 MHz, DMSO-d6) δ 11.56 (s, 1H, NH), 7.85 (s, 6


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101

1H, Ar-H), 7.76 (s, 1H, Ar-H), 7.48 (s, 1H, Ar-H), 7.29 (s, 1H, Ar-H), 7.21 (s, 1H, Ar-

102

H), 7.17 (s, 1H, Ar-H), 3.15 (s, 1H, Ar-CH), 1.36 (s, 6H, CH3); 13C NMR (100 MHz,

103

DMSO-d6) δ 165.5, 147.1, 136.3, 123.6, 122.9, 122.0, 120.0, 119.4, 118.9, 112.0, 103.9,

104

27.6, 20.4.

105

Labradorin 1 (5e). Brown solid; mp. 145−146 oC (lit.19, 147−148 oC); yield 78%; 1H

106

NMR (400 MHz, DMSO-d6) δ 11.55 (s, 1H, NH), 7.84 (d, J = 7.8 Hz, 1H, Ar-H), 7.73

107

(d, J = 2.6 Hz, 1H, Ar-H), 7.47 (d, J = 8.0 Hz, 1H, Ar-H), 7.30 (s, 1H, Ar-H), 7.12–

108

7.22 (m, 2H, Ar-H), 2.69 (d, J = 7.1 Hz, 2H, Ar-CH2), 2.09–2.19 (m, 1H, ArCH2-CH),

109

0.98 (d, J = 6.7 Hz, 6H, CH3); 13C NMR (100 MHz, DMSO-d6) δ 160.9, 147.2, 136.3,

110

123.5, 122.9, 122.0, 120.0, 119.4, 119.1, 112.0, 103.9, 36.2, 27.1, 22.1; HRMS (ESI)

111

calcd for C15H17N2O (M+H)+ 241.1135, found 241.1133.

112

2-Cyclohexyl-5-(1H-indol-3-yl)oxazole (5f). Brown solid; mp. 169−170 oC; yield

113

48%; 1H NMR (400 MHz, DMSO-d6) δ 11.54 (s, 1H, NH), 8.19–8.24 (m, 1H, Ar-H),

114

7.83 (d, J = 7.7 Hz, 1H, Ar-H), 7.74 (d, J = 2.6 Hz, 1H, Ar-H), 7.46 (d, J = 7.9 Hz, 1H,

115

Ar-H), 7.28 (s, 2H, Ar-H), 2.84–2.90 (m, 1H, Ar-CH), 2.05–2.10 (m, 2H, CH-CH2),

116

1.76–1.80 (m, 2H, CH-CH2), 1.13–1.66 (m, 4H, CHCH2-CH2), 1.26–1.45 (m, 2H,

117

CHCH2CH2-CH2); 13C NMR (100 MHz, DMSO-d6) δ 164.6, 146.9, 137.1, 136.3, 122.9,

118

122.0, 120.0, 119.4, 118.9, 112.0, 104.0, 36.5, 30.3, 25.0; HRMS (ESI) calcd for

119

C17H19N2O (M+H)+ 267.1492, found 267.1493.

120

(5-(1H-Indol-3-yl)oxazol-2-yl)methanol (5g). Brown solid; mp. 187−189 oC (lit.19,

121

189−190 oC); yield 39%; 1H NMR (400 MHz, DMSO-d6) δ 11.59 (s, 1H, NH), 7.89 (d, 7



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122

J = 7.7 Hz, 1H, Ar-H), 7.79 (d, J = 1.4 Hz, 1H, Ar-H), 7.50 (d, J = 7.9 Hz, 1H, Ar-H),

123

7.39 (s, 1H, Ar-H), 7.22 (t, J = 7.1 Hz, 1H, Ar-H), 7.16 (t, J = 7.5 Hz, 1H, Ar-H), 5.71

124

(t, J = 5.9 Hz, 1H, OH), 4.59 (d, J = 5.8 Hz, 2H, CH2); 13C NMR (100 MHz, DMSO-

125

d6) δ 160.8, 147.9, 136.3, 123.5, 123.2, 122.1, 120.1, 119.5, 112.1, 103.7, 55.9.

126

5-(1H-Indol-3-yl)-2-(2-(methylthio)ethyl)oxazole (5h). Brown solid; mp. 146−147

127

oC (lit.19, 149−150 oC); yield 47%; 1H NMR (400 MHz, DMSO-d ) δ 11.57 (s, 1H, NH), 6

128

7.86 (d, J = 7.7 Hz, 1H, Ar-H), 7.76 (s, 1H, Ar-H), 7.48 (d, J = 7.9 Hz, 1H, Ar-H), 7.34

129

(s, 1H, Ar-H), 7.13–7.23 (m, 2H, Ar-H), 3.12 (t, J = 6.9 Hz, 2H, CH2), 2.94 (t, J = 7.1

130

Hz, 2H, CH2), 2.11 (s, 3H, CH3); 13C NMR (100 MHz, DMSO-d6) δ 160.0, 147.5, 136.3,

131

123.5, 123.0, 122.1, 120.0, 119.4, 119.1, 112.0, 103.8, 30.5, 27.9, 14.6.

132

5-(1H-Indol-3-yl)-2-phenyloxazole (5i). Brown solid; mp. 226−228

133

226−228 oC); yield 83%; 1H NMR (400 MHz, DMSO-d6) δ 11.70 (s, 1H, NH), 8.12 (d,

134

J = 1.4 Hz, 1H, Ar-H), 8.10 (s, 1H, Ar-H), 7.99 (s, 1H, Ar-H), 7.98 (d, J = 5.9 Hz, 1H,

135

Ar-H), 7.62 (s, 1H, Ar-H), 7.58 (t, J = 6.9 Hz, 2H, Ar-H), 7.52 (t, J = 6.8 Hz, 2H, Ar-

136

H), 7.19–7.27 (m, 2H, Ar-H); 13C NMR (100 MHz, DMSO-d6) δ 158.0, 148.3, 136.4,

137

130.0, 129.1, 127.2, 125.5, 123.8, 123.5, 122.2, 120.8, 120.3, 119.5, 112.1, 103.6.

138

2-(4-Fluorophenyl)-5-(1H-indol-3-yl)oxazole (5j). Brown solid; mp. 205−206 oC;

139

yield 51%; 1H NMR (400 MHz, DMSO-d6) δ 11.69 (s, 1H, NH), 8.13 (dd, J = 6.3, 9.0

140

Hz, 2H, Ar-H), 7.97 (s, 1H, Ar-H), 7.96 (d, J = 9.0 Hz, 1H, Ar-H), 7.60 (s, 1H, Ar-H),

141

7.50 (d, J = 7.6 Hz, 1H, Ar-H), 7.41 (t, J = 8.2 Hz, 2H, Ar-H), 7.17–7.25 (m, 2H, Ar-

142

H); 13C NMR (100 MHz, DMSO-d6) δ 164.3, 161.8, 157.2, 148.3, 136.4, 127.94, 127.86, 8


oC

(lit.19,

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143

123.9, 123.5, 122.2, 120.7, 120.3, 119.5, 116.4, 116.1, 112.1, 103.5; HRMS (ESI) calcd

144

for C17H12FN2O (M+H)+ 279.0928, found 279.0933.

145

Pimprinaphine (5k). Brown solid; mp. 199−200 oC (lit.19, 202−203 oC); yield 56%;

146

1H

147

7.74 (s, 1H, Ar-H), 7.47 (d, J = 7.6 Hz, 1H, Ar-H), 7.32–7.40 (m, 5H, Ar-H), 7.29 (s,

148

1H, Ar-H), 7.20 (t, J = 6.9 Hz, 1H, Ar-H), 7.14 (t, J = 7.2 Hz, 1H, Ar-H), 4.22 (s, 2H,

149

Ar-CH2); 13C NMR (100 MHz, DMSO-d6) δ 160.0, 147.8, 136.3, 136.2, 128.7, 128.6,

150

127.2, 126.8, 123.5, 123.1, 122.1, 120.0, 119.4, 119.3, 112.0, 103.7, 33.7; HRMS (ESI)

151

calcd for C18H15N2O (M+H)+ 275.1179, found 275.1177.

152

4-((5-(1H-Indol-3-yl)oxazol-2-yl)methyl)phenol (5l). Brown solid; mp. 165−166 oC;

153

yield 44%; 1H NMR (400 MHz, DMSO-d6) δ 11.54 (s, 1H, NH), 9.45 (s, 1H, OH), 8.01

154

(s, 1H, Ar-H), 7.81 (d, J = 7.8 Hz, 1H, Ar-H), 7.72 (d, J = 2.4 Hz, 1H, Ar-H), 7.46 (d,

155

J = 7.5 Hz, 1H, Ar-H), 7.31 (s, 1H, Ar-H), 7.16 (d, J = 8.9 Hz, 2H, Ar-H), 6.75 (d, J =

156

8.3 Hz, 2H, Ar-H), 4.07 (s, 2H, Ar-CH2); 13C NMR (100 MHz, DMSO-d6) δ 166.4,

157

161.0, 156.6, 148.1, 136.9, 136.8, 132.8, 130.2, 126.7, 123.5, 122.6, 122.5, 121.0, 119.9,

158

115.8, 112.5, 104.2, 33.4; HRMS (ESI) calcd for C18H15N2O2 (M+H)+ 291.1128, found

159

291.1129.

160

General Methods for the Synthesis of Compounds 6aa and 6ab. The 60% NaH (7

161

mmol) powder was batched into tetrahydrofuran solution of methyloxazole 5a (2 mmol)

162

at room temperature and stirred for 30 min. To the reaction solution was added

163

iodomethane or benzyl chloride (6 mmol) and stirred for 1 h at room temperature and

NMR (400 MHz, DMSO-d6) δ 11.57 (s, 1H, NH), 7.82 (d, J = 7.4 Hz, 1H, Ar-H),

9



164

refluxed for 1 h. Water (200 mL) was added and extracted with CH2Cl2 (80 mL × 3).

165

The organic layer was washed with saturated NaCl solution (50 mL), dried with

166

anhydrous Na2SO4, filtered and concentrated in vacuum. The compounds 6aa and 6ab

167

were obtained by column chromatography on silica gel.

168

2-Methyl-5-(1-methyl-1H-indol-3-yl)oxazole (6aa). Yellow solid; mp. 71−73 oC;

169

yield 86%; 1H NMR (400 MHz, DMSO-d6) δ 7.86 (d, J = 7.8 Hz, 1H, Ar-H), 7.73 (s,

170

1H, Ar-H), 7.50 (d, J = 8.1 Hz, 1H, Ar-H), 7.30 (s, 1H, Ar-H), 7.27 (d, J = 7.5 Hz, 1H,

171

Ar-H), 7.19 (d, J = 7.5 Hz, 1H, Ar-H), 3.83 (s, 3H, N-CH3), 2.48 (s, 3H, Ar-CH3); 13C

172

NMR (100 MHz, DMSO-d6) δ 158.3, 147.0, 136.8, 126.8, 123.7, 122.1, 120.1, 119.6,

173

119.2, 110.3, 103.0, 32.6, 13.5; HRMS (ESI) calcd for C13H13N2O (M+H)+ 213.1022,

174

found 213.1027.

175

5-(1-Benzyl-1H-indol-3-yl)-2-methyloxazole (6ab). Yellow solid; mp. 94−96 oC;

176

yield 88%; 1H NMR (400 MHz, DMSO-d6) δ 7.93 (s, 1H, Ar-H), 7.87 (d, J = 7.7 Hz,

177

1H, Ar-H), 7.55 (d, J = 8.1 Hz, 1H, Ar-H), 7.15–7.32 (m, 7H, Ar-H), 5.48 (s, 2H, Ar-

178

CH2), 2.48 (s, 3H, Ar-CH3); 13C NMR (100 MHz, DMSO-d6) δ 158.4, 146.8, 137.7,

179

136.1, 128.6, 127.5, 127.2, 126.2, 124.0, 122.3, 120.3, 119.8, 119.5, 110.8, 103.7, 49.2,

180

13.5; HRMS (ESI) calcd for C19H17N2O (M+H)+ 289.1335, found 289.1337.

181

General Methods for the Synthesis of Compounds 9a–9u. The reaction mixture of

182

1H-indole-3-carbohydrazide (7, 6 mmol) and corresponding aldehydes 8a–8u (6 mmol)

183

in anhydrous ethanol (80 mL) was stirred and refluxed at 90 oC for 4 h, then cooled to

184

room temperature and filtered to give compounds 9a–9u. 10


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185

(E)-N'-Benzylidene-1H-indole-3-carbohydrazide (9a). White solid; mp. 233−234 oC

186

(lit.20, 232−234 oC); yield 79%; 1H NMR (400 MHz, DMSO-d6) δ 11.74 (s, 1H, NH),

187

11.38 (s, 1H, NH), 8.20–8.33 (m, 3H, Ar-H), 7.72 (d, J = 7.3 Hz, 2H, Ar-H), 7.47 (s,

188

1H, Ar-CH), 7.38–7.49 (m, 3H, Ar-H), 7.14–7.22 (m, 2H, Ar-H); 13C NMR (100 MHz,

189

DMSO-d6) δ 134.7, 129.5, 128.8, 126.7, 122.2, 120.8, 111.9.

190

(E)-N'-(4-Methoxybenzylidene)-1H-indole-3-carbohydrazide (9b). White solid; mp.

191

242−243 oC (lit.20, 239−241 oC); yield 90%; 1H NMR (400 MHz, DMSO-d6) δ 11.72

192

(s, 1H, NH), 11.27 (s, 1H, NH), 8.20–8.29 (m, 3H, Ar-H, Ar-CH), 7.66 (d, J = 8.6 Hz,

193

2H, Ar-H), 7.45 (d, J = 7.8 Hz, 1H, Ar-H), 7.15–7.20 (m, 2H, Ar-H), 7.03 (d, J = 8.7

194

Hz, 2H, Ar-H), 3.82 (s, 3H, OCH3); 13C NMR (100 MHz, DMSO-d6) δ 160.4, 128.3,

195

127.3, 122.2, 120.7, 114.3, 111.9, 55.2.

196

(E)-N'-(2-Bromobenzylidene)-1H-indole-3-carbohydrazide (9c). White solid; mp.

197

294−296 oC; yield 92%; 1H NMR (400 MHz, DMSO-d6) δ 11.81 (s, 1H, NH), 11.71 (s,

198

1H, NH), 8.68 (s, 1H, Ar-H), 8.30 (s, 1H, Ar-CH), 8.23 (d, J = 7.3 Hz, 1H, Ar-H), 8.02

199

(dd, J = 6.6, 1.2 Hz, 1H, Ar-H), 7.70 (d, J = 8.0 Hz, 1H, Ar-H), 7.46–7.51 (m, 2H, Ar-

200

H), 7.33–7.37 (m, 1H, Ar-H), 7.16–7.24 (m, 2H, Ar-H); 13C NMR (100 MHz, DMSO-

201

d6) δ 136.0, 133.5, 130.1, 131.2, 128.0, 127.0, 123.1, 122.3, 120.9, 112.0; HRMS (ESI)

202

calcd for C16H13BrN3O (M+H)+ 342.0237, found 342.0239.

203

(E)-N'-(4-Nitrobenzylidene)-1H-indole-3-carbohydrazide (9d). Yellow solid; mp.

204

287−288 oC (lit.20, 289−291 oC); yield 92%; 1H NMR (400 MHz, DMSO-d6) δ 11.83

205

(d, J = 1.4 Hz, 1H, NH), 11.73 (s, 1H, NH), 8.38–8.43 (m, 1H, Ar-H), 8.31 (d, J = 8.8 11



206

Hz, 2H, Ar-H), 8.29 (s, 1H, Ar-CH), 8.21 (d, J = 7.5 Hz, 1H, Ar-H), 7.98 (d, J = 8.8

207

Hz, 2H, Ar-H), 7.51 (d, J = 7.5 Hz, 1H, Ar-H), 7.16–7.24 (m, 2H, Ar-H); 13C NMR

208

(100 MHz, DMSO-d6) δ 147.4, 141.2, 136.0, 127.6, 124.1, 122.4, 121.1, 120.9, 112.0.

209

(E)-N'-(4-(Trifluoromethyl)benzylidene)-1H-indole-3-carbohydrazide (9e). White

210

solid; mp. 298−300 oC; yield 53%; 1H NMR (400 MHz, DMSO-d6) δ 11.80 (s, 1H,

211

NH), 11.61 (s, 1H, NH), 8.20–8.39 (m, 3H, Ar-H, Ar-CH), 7.94 (d, J = 8.1 Hz, 2H, Ar-

212

H), 7.82 (d, J = 8.2 Hz, 2H, Ar-H), 7.49 (d, J = 8.1 Hz, 1H, Ar-H), 7.15–7.23 (m, 2H,

213

Ar-H); 13C NMR (100 MHz, DMSO-d6) δ 139.2, 129.9, 129.5, 127.8, 126.3, 126.19,

214

126.15, 126.0, 122.9, 121.4, 112.5; HRMS (ESI) calcd for C17H13F3N3O (M+H)+

215

332.1005, found 332.1001.

216

(E)-N'-(Furan-2-ylmethylene)-1H-indole-3-carbohydrazide (9f). White solid; mp.

217


218

1H, NH), 8.10–8.30 (m, 3H, Ar-H, NHN-CH), 7.84 (s, 1H, Ar-H), 7.48 (d, J = 7.8 Hz,

219

1H, Ar-H), 7.13–7.21 (m, 2H, Ar-H), 6.88 (d, J = 3.3 Hz, 1H, Ar-H), 6.64 (dd, J = 3.0,

220

1.6 Hz, 1H, Ar-H); 13C NMR (100 MHz, DMSO-d6) δ 149.8, 144.6, 122.2, 120.8, 112.3,

221

112.1, 111.9; HRMS (ESI) calcd for C14H12N3O2 (M+H)+ 254.0924, found 254.0925.

222

(E)-N'-(Naphthalen-1-ylmethylene)-1H-indole-3-carbohydrazide (9g). White solid;

223

mp. 279−281 oC; yield 86%; 1H NMR (400 MHz, DMSO-d6) δ 11.80 (s, 1H, NH),

224

11.52 (s, 1H, NH), 8.90–9.10 (m, 2H, Ar-H, NCH), 8.20–8.30 (m, 2H, Ar-H), 8.00–

225

8.04 (m, 2H, Ar-H), 7.94 (d, J = 7.0 Hz, 1H, Ar-H), 7.68 (d, J = 7.1 Hz, 1H, Ar-H),

226

7.60–7.63 (m, 2H, Ar-H), 7.50 (d, J = 7.5 Hz, 1H, Ar-H), 7.16–7.24 (m, 2H, Ar-H); 13C 12


Page 12 of 44

Page 13 of 44


227

NMR (100 MHz, DMSO-d6) δ 133.5, 130.1, 130.0, 128.8, 127.1, 126.2, 125.6, 122.3,

228

120.8, 112.0; HRMS (ESI) calcd for C20H16N3O (M+H)+ 314.1288, found 314.1284.

229

(E)-N'-((5-Bromo-1H-indol-3-yl)methylene)-1H-indole-3-carbohydrazide

230

Brown solid; mp. 230−231 oC (lit.21, 232−233 oC); yield 78%; 1H NMR (400 MHz,

231

DMSO-d6) δ 11.74 (s, 2H, NH), 11.19 (s, 1H, NH), 8.53 (d, J = 4.8 Hz, 2H, Ar-H), 8.24

232

(d, J = 7.4 Hz, 1H, Ar-H), 8.20 (s, 1H, Ar-CH), 7.87 (s, 1H, Ar-H), 7.15–7.51 (m, 5H,

233

Ar-H); 13C NMR (100 MHz, DMSO-d6) δ 136.2, 131.3, 126.4, 125.5, 122.6, 121.1.

234

114.3, 113.4, 112.4.

235

(E)-N'-(Cyclohexylmethylene)-1H-indole-3-carbohydrazide (9i). White solid; mp.

236


237

1H, NH), 8.02–8.14 (m, 2H, Ar-H), 7.53 (s, 1H, NHN-CH), 7.45 (d, J = 7.8 Hz, 1H,

238

Ar-H), 7.11–7.19 (m, 2H, Ar-H), 2.22–2.32 (m, 1H, NCH-CH), 1.63–1.82 (m, 5H, CH2),

239

1.19–1.36 (m, 5H, CH2); 13C NMR (100 MHz, DMSO-d6) δ 122.1, 120.6, 111.8, 29.8,

240


241

(E)-N'-Octylidene-1H-indole-3-carbohydrazide (9j). White solid; mp. 260−266 oC;

242

yield 56%; 1H NMR (400 MHz, DMSO-d6) δ 11.66 (s, 1H, NH), 10.93 (s, 1H, NH),

243

8.12–8.16 (m, 2H, Ar-H, NCH), 7.55–7.64 (m, 1H, Ar-H), 7.45 (d, J = 7.9 Hz, 1H, Ar-

244

H), 7.10–7.19 (m, 2H, Ar-H), 2.24–2.28 (m, 2H, CH2), 1.49–1.52 (m, 2H, CH2), 1.20–

245

1.35 (m, 8H, CH2), 0.87 (t, J = 5.5 Hz, 3H, CH3); 13C NMR (100 MHz, DMSO-d6) δ

246

122.1, 120.5, 111.8, 31.9, 31.2, 28.6, 28.5, 26.2, 22.0, 13.9; HRMS (ESI) calcd for

247

C17H24N3O (M+H)+ 286.1914, found 286.1910. 13


(9h).


Page 14 of 44

248

(E)-N'-(3,4,5-Trimethoxybenzylidene)-1H-indole-3-carbohydrazide (9k). White

249

solid; mp. 243−245 oC (lit.21, 245−246 oC); yield 77%; 1H NMR (400 MHz, DMSO-

250

d6) δ 11.79 (s, 1H, NH), 11.43 (s, 1H, NH), 8.21–8.28 (m, 3H, Ar-H, Ar-CH), 7.49 (d,

251

J = 7.9 Hz, 1H, Ar-H), 7.14–7.22 (m, 2H, Ar-H), 7.02 (s, 2H, Ar-H), 3.85 (s, 6H, CH3),

252

3.71 (s, 3H, CH3); 13C NMR (100 MHz, DMSO-d6) δ 153.2, 138.7, 130.3, 122.3, 120.8,

253

112.0, 103.9, 60.1, 55.9.

254

(E)-N'-(3-Hydroxy-4-methoxybenzylidene)-1H-indole-3-carbohydrazide

255

White solid; mp. 248−250 oC; yield 88%; 1H NMR (400 MHz, DMSO-d6) δ 11.74 (s,

256

1H, NH), 11.25 (s, 1H, NH), 9.49 (s, 1H, OH), 8.15–8.30 (m, 3H, Ar-H, NHNC-H),

257

7.48 (d, J = 8.0 Hz, 1H, Ar-H), 7.31 (s, 1H, Ar-H), 7.13–7.21 (m, 2H, Ar-H), 7.07 (dd,

258

J = 8.1, 1.4 Hz, 1H, Ar-H), 6.85 (d, J = 8.1 Hz, 1H, Ar-H), 3.84 (s, 3H, OCH3); 13C

259

NMR (100 MHz, DMSO-d6) δ 148.4, 148.0, 126.2, 122.2, 120.7, 115.4, 111.9, 108.8,

260

55.5; HRMS (ESI) calcd for C17H16N3O3 (M+H)+ 310.1186, found 310.1191.

261

(E)-N'-(4-Hydroxybenzylidene)-1H-indole-3-carbohydrazide (9m). White solid;

262


263

11.22 (s, 1H, NH), 9.88 (s, 1H, OH), 8.12–8.30 (m, 3H, Ar-H, NHNC-H), 7.55 (d, J =

264

8.5 Hz, 2H, Ar-H), 7.48 (d, J = 7.8 Hz, 1H, Ar-H), 7.13–7.21 (m, 2H, Ar-H), 6.85 (d, J

265

= 8.5 Hz, 2H, Ar-H); 13C NMR (100 MHz, DMSO-d6) δ 158.9, 128.5, 125.7, 122.2,

266

120.7, 115.7, 111.9; HRMS (ESI) calcd for C16H14N3O2 (M+H)+ 280.1081, found

267

280.1086.

268

(E)-N'-(4-Fluorobenzylidene)-1H-indole-3-carbohydrazide (9n). White solid; mp. 14


(9l).

Page 15 of 44


269


270

1H, NH), 8.19–8.40 (m, 3H, Ar-H, NHNC-H), 7.78 (dd, J = 8.6, 5.7 Hz, 2H, Ar-H),

271

7.48 (d, J = 7.6 Hz, 1H, Ar-H), 7.27–7.32 (m, 2H, Ar-H), 7.13–7.22 (m, 2H, Ar-H); 13C

272

NMR (100 MHz, DMSO-d6) δ 164.0, 161.5, 131.3, 128.9, 128.8, 122.2, 121.1, 120.8,

273

115.9, 115.7, 111.9; HRMS (ESI) calcd for C16H13FN3O (M+H)+ 282.1037, found

274

282.1042.

275

(E)-N'-(2-Nitrobenzylidene)-1H-indole-3-carbohydrazide (9o). Yellow solid; mp.

276


277

1H, NH), 8.73 (s, 1H, Ar-H), 8.30 (s, 1H, Ar-CH), 8.23 (d, J = 7.5 Hz, 1H, Ar-H), 8.16

278

(d, J = 7.7 Hz, 1H, Ar-H), 8.08 (dd, J = 8.2, 1.0 Hz, 1H, Ar-H), 7.82 (t, J = 7.7 Hz, 1H,

279

Ar-H), 7.64–7.68 (m, 1H, Ar-H), 7.50 (d, J = 7.9 Hz, 1H, Ar-H), 7.16–7.24 (m, 2H, Ar-

280

H); 13C NMR (100 MHz, DMSO-d6) δ 148.0, 136.0, 133.6, 130.1, 129.1, 127.7. 124.6,

281

122.3, 121.1, 120.9, 112.0; HRMS (ESI) calcd for C16H13N4O3 (M+H)+ 309.0982,

282

found 309.0985.

283

(E)-N'-(Quinolin-4-ylmethylene)-1H-indole-3-carbohydrazide (9p). Yellow solid;

284

mp. > 300 oC; yield 77%; 1H NMR (400 MHz, DMSO-d6) δ 11.86 (s, 1H, NH), 11.79

285

(s, 1H, NH), 8.99 (d, J = 4.5 Hz, 1H, Ar-H), 8.97 (s, 1H, Ar-H), 8.78 (s, 1H, NHNC-

286

H), 8.34 (s, 1H, Ar-H), 8.25 (d, J = 7.4 Hz, 1H, Ar-H), 8.12 (d, J = 8.4 Hz, 1H, Ar-H),

287

7.82–7.86 (m, 2H, Ar-H), 7.75 (t, J = 7.9 Hz, 1H, Ar-H), 7.52 (d, J = 7.9 Hz, 1H, Ar-

288

H), 7.18–7.25 (m, 2H, Ar-H); 13C NMR (100 MHz, DMSO-d6) δ 150.8, 148.9, 138.4,

289

136.5, 130.2, 130.1, 127.9, 125.2, 122.9, 121.5, 112.6; HRMS (ESI) calcd for

15



290

C19H15N4O (M+H)+ 315.1240, found 315.1237.

291

(E)-N'-((1H-Pyrrol-2-yl)methylene)-1H-indole-3-carbohydrazide (9q). Gray solid;

292


293

11.45 (s, 1H, NH), 11.09 (s, 1H, NH), 8.13–8.20 (m, 3H, Ar-H, NHNC-H), 7.47 (d, J

294

= 7.8 Hz, 1H, Ar-H), 7.12–7.20 (m, 2H, Ar-H), 6.92 (s, 1H, Ar-H), 6.45 (s, 1H, Ar-H),

295

6.15 (s, 1H, Ar-H); 13C NMR (100 MHz, DMSO-d6) δ 128.0, 122.6, 122.3, 121.1, 112.9,

296


297

(E)-N'-(Thiophen-2-ylmethylene)-1H-indole-3-carbohydrazide (9r). Yellow solid;

298

mp. 265−267 oC (lit.21, 269−271 oC); yield 93%; 1H NMR (400 MHz, DMSO-d6) δ

299

11.78 (s, 1H, NH), 11.36 (s, 1H, NH), 8.58 (s, 1H, Ar-H), 8.21 (s, 2H, Ar-H, NHNC-

300

H), 7.63 (s, 1H, Ar-H), 7.48 (d, J = 7.1 Hz, 1H, Ar-H), 7.43 (s, 1H, Ar-H), 7.14–7.20

301

(m, 3H, Ar-H); 13C NMR (100 MHz, DMSO-d6) δ 128.0, 122.6, 122.3, 121.1, 112.9,

302

112.4, 109.6.

303

(E)-N'-(Pyridin-4-ylmethylene)-1H-indole-3-carbohydrazide (9s). Gray solid; mp.

304

> 300 oC; yield 54%; 1H NMR (400 MHz, DMSO-d6) δ 11.83 (s, 1H, NH), 11.70 (s,

305

1H, NH), 8.65 (d, J = 5.6 Hz, 2H, Ar-H), 8.28–8.39 (m, 2H, Ar-H, NHNC-H), 8.23 (d,

306


307

7.16–7.24 (m, 2H, Ar-H); 13C NMR (100 MHz, DMSO-d6) δ 150.6, 142.4, 122.9, 121.5,

308


309

(E)-N'-Butylidene-1H-indole-3-carbohydrazide (9t). White solid; mp. 238−240 oC;

310

yield 77%; 1H NMR (400 MHz, DMSO-d6) δ 11.67 (s, 1H, NH), 10.95 (s, 1H, NH), 16


Page 16 of 44

Page 17 of 44


311

8.02–8.34 (m, 2H, Ar-H), 7.60 (s, 1H, NHN-CH), 7.45 (d, J = 7.8 Hz, 1H, Ar-H), 7.11–

312

7.19 (m, 2H, Ar-H), 2.25 (dd, J = 6.9, 13.1 Hz, 2H, NCH-CH2), 1.49–1.58 (m, 2H, CH3-

313

CH2), 0.95 (t, J = 7.3 Hz, 3H, CH3); 13C NMR (100 MHz, DMSO-d6) δ 122.1, 120.6,

314

111.8, 33.9, 19.5, 13.6; HRMS (ESI) calcd for C13H16N3O (M+H)+ 230.1288, found

315

230.1283.

316

(E)-N'-(2-Methylpropylidene)-1H-indole-3-carbohydrazide (9u). White solid; mp.

317


318

1H, NH), 8.10–8.30 (m, 2H, Ar-H), 7.16 (s, 1H, NHNC-H), 7.44–7.47 (m, 1H, Ar-H),

319

7.12–7.19 (m, 2H, Ar-H), 2.50–2.55 (m, 1H, (CH3)2C-H), 1.10 (s, 3H, CH3), 1.08 (s,

320

3H, CH3); 13C NMR (100 MHz, DMSO-d6) δ 122.6, 121.1, 112.3, 31.4, 20.2; HRMS

321


322

General Methods for the synthesis of Compounds 10a–10j. The reaction mixture of

323

corresponding carbohydrazides 9a–9j (3 mmol) and iodobenzene diacetate (6 mmol) in

324

acetonitrile (60 mL) was stirred at 50 oC for 2 h. The acetonitrile was removed in

325

vacuum. Ethyl acetate (150 mL) was added and washed with saturated Na2CO3 solution

326

(80 mL), brine (80 mL) and dried with anhydrous Na2SO4, filtered and concentrated in

327

vacuum. The compounds 10a–10j were obtained by column chromatography on silica

328

gel.

329

2-(1H-Indol-3-yl)-5-phenyl-1,3,4-oxadiazole (10a). White solid; mp. 269−271 oC;

330

yield 72%; 1H NMR (400 MHz, DMSO-d6) δ 12.12 (s, 1H, NH), 8.34 (d, J = 2.9 Hz,

331

1H, Ar-H), 8.20 (t, J = 5.0 Hz, 1H, Ar-H), 8.14–8.16 (m, 2H, Ar-H), 7.64–7.66 (m, 3H, 17



Page 18 of 44

13C

332

Ar-H), 7.57–7.59 (m, 1H, Ar-H), 7.29–7.31 (m, 2H, Ar-H);

333

DMSO-d6) δ 162.0, 161.8, 136.5, 131.5, 129.4, 128.6, 126.3, 124.1, 123.7, 122.9, 121.3,

334

120.2, 112.5, 99.4; HRMS (ESI) calcd for C16H12N3O (M+H)+ 262.0975, found

335

262.0971.

336

2-(1H-Indol-3-yl)-5-(4-methoxyphenyl)-1,3,4-oxadiazole (10b). White solid; mp.

337

276−277 oC; yield 69%; 1H NMR (400 MHz, DMSO-d6) δ 12.07 (s, 1H, NH), 8.07 (d,

338


339

7.40 (t, J = 7.4 Hz, 1H, Ar-H), 7.27–7.31 (m, 1H, Ar-H), 7.19 (t, J = 7.7 Hz, 3H, Ar-

340

H), 3.88 (s, 3H, OCH3); 13C NMR (100 MHz, DMSO-d6) δ 172.5, 162.2, 137.6, 135.0,

341

131.1, 128.6, 128.2, 123.8, 122.1, 118.9, 115.3, 113.4, 56.0; HRMS (ESI) calcd for

342

C17H14N3O2 (M+H)+ 292.1081, found 292.1087.

343

2-(2-Bromophenyl)-5-(1H-indol-3-yl)-1,3,4-oxadiazole (10c). Brown solid; mp.

344


345

1H, Ar-H), 8.18 (d, J = 5.3 Hz, 1H, Ar-H), 8.07 (d, J = 7.2 Hz, 1H, Ar-H), 7.92 (d, J =

346

7.7 Hz, 1H, Ar-H), 7.64 (t, J = 6.8 Hz, 1H, Ar-H), 7.57 (d, J = 5.5 Hz, 2H, Ar-H), 7.29

347

(d, J = 5.9 Hz, 2H, Ar-H); 13C NMR (100 MHz, DMSO-d6) δ 162.8, 161.6, 137.0, 134.8,

348

133.4, 132.1, 129.2, 128.7, 125.6, 124.6, 123.4, 121.9, 121.3, 120.6, 113.0, 99.6; HRMS

349

(ESI) calcd for C16H11BrN3O (M+H)+ 340.0080, found 340.0081.

350

2-(1H-Indol-3-yl)-5-(4-nitrophenyl)-1,3,4-oxadiazole (10d). Gray solid; mp. >300 oC;

351

yield 66%; 1H NMR (400 MHz, DMSO-d6) δ 12.18 (s, 1H, NH), 8.47 (s, 1H, Ar-H),

352

8.45 (s, 1H, Ar-H), 8.35–8.39 (m, 3H, Ar-H), 8.18 (t, J = 5.0 Hz, 1H, Ar-H), 7.56–7.58 18


NMR (100 MHz,

Page 19 of 44


353

(m, 1H, Ar-H), 7.29–7.31 (m, 2H, Ar-H); 13C NMR (100 MHz, DMSO-d6) δ 163.4,

354

161.1, 149.3, 137.0, 129.73, 129.70. 128.0, 125.1, 124.5, 123.5, 121.9, 120.7, 113.1,

355

99.5; HRMS (ESI) calcd for C16H11N4O3 (M+H)+ 307.0826, found 307.0822.

356

2-(1H-Indol-3-yl)-5-(4-(trifluoromethyl)phenyl)-1,3,4-oxadiazole

357

yellow solid; mp. 288−290 oC; yield 82%; 1H NMR (400 MHz, DMSO-d6) δ 12.16 (s,

358

1H, NH), 8.37 (d, J = 2.9 Hz, 1H, Ar-H), 8.34 (d, J = 8.1 Hz, 2H, Ar-H), 8.19 (t, J =

359

5.0 Hz, 1H, Ar-H), 8.01 (d, J = 8.3 Hz, 2H, Ar-H), 7.57–7.59 (m, 1H, Ar-H), 7.28–7.33

360

(m, 2H, Ar-H); 13C NMR (100 MHz, DMSO-d6) δ 163.0, 161.4, 136.9, 127.7, 126.8,

361

124.5. 123.5, 121.9, 120.6, 113.1, 99.6; HRMS (ESI) calcd for C17H11F3N3O (M+H)+

362

330.0849, found 330.0844.

363

2-(Furan-2-yl)-5-(1H-indol-3-yl)-1,3,4-oxadiazole (10f). Gray solid; mp. 214−216 oC;

364


365

1H, Ar-H), 8.16 (dd, J = 3.4, 6.5 Hz, 1H, Ar-H), 8.09 (d, J = 1.1 Hz, 1H, Ar-H), 7.57

366

(dd, J = 2.0, 5.5 Hz, 1H, Ar-H), 7.41 (d, J = 3.5 Hz, 1H, Ar-H), 7.26–7.32 (m, 2H, Ar-

367

H), 6.84 (dd, J = 1.8, 3.5 Hz, 1H, Ar-H); 13C NMR (100 MHz, DMSO-d6) δ 161.7,

368

155.4, 147.0, 139.5, 136.9, 129.1. 124.5, 123.4, 121.8, 120.7, 114.3, 113.1, 113.0, 99.5;

369

HRMS (ESI) calcd for C14H10N3O2 (M+H)+ 252.0768, found 252.0765.

370

2-(1H-Indol-3-yl)-5-(naphthalen-1-yl)-1,3,4-oxadiazole (10g). Gray solid; mp.

371

186−188 oC; yield 70%; 1H NMR (400 MHz, CDCl3) δ 9.37 (d, J = 8.6 Hz, 1H, Ar-H),

372

9.23 (s, 1H, NH), 8.38 (d, J = 7.1 Hz, 1H, Ar-H), 8.30 (d, J = 7.2 Hz, 1H, Ar-H), 8.11

373

(d, J = 2.6 Hz, 1H, Ar-H), 8.06 (d, J = 8.3 Hz, 1H, Ar-H), 7.97 (d, J = 8.2 Hz, 1H, Ar19


(10e).

Light


374

H), 7.70–7.76 (m, 1H, Ar-H), 7.61–7.66 (m, 2H, Ar-H), 7.53–7.58 (m, 2H, Ar-H), 7.34–

375

7.39 (m, 1H, Ar-H); 13C NMR (100 MHz, DMSO-d6) δ 161.8, 161.5, 136.5, 133.5,

376

132.2, 129.3, 128.9, 128.2, 128.1, 126.8, 125.7, 125.4, 124.2, 122.9, 121.3, 120.2, 120.0,

377

112.5. 99.3; HRMS (ESI) calcd for C20H14N3O (M+H)+ 312.1131, found 312.1137.

378

2-(5-Bromo-1H-indol-3-yl)-5-(1H-indol-3-yl)-1,3,4-oxadiazole (10h). Gray solid;

379


380

12.04 (s, 1H, NH), 8.34 (s, 2H, Ar-H), 8.26 (s, 1H, Ar-H), 8.22 (d, J = 6.4 Hz, 1H, Ar-

381

H), 7.58 (d, J = 1.9 Hz, 1H, Ar-H), 7.55 (d, J = 8.6 Hz, 1H, Ar-H), 7.43 (d, J = 8.6 Hz,

382

1H, Ar-H), 7.29 (t, J = 3.6 Hz, 2H, Ar-H); 13C NMR (100 MHz, DMSO-d6) δ 160.4,

383

159.7, 137.0, 135.8, 129.7, 126.3. 124.5, 123.4, 123.0, 120.9, 115.0, 112.9, 100.1, 99.9;

384

HRMS (ESI) calcd for C18H12BrN4O (M+H)+ 379.0189, found 379.0181.

385

2-Cyclohexyl-5-(1H-indol-3-yl)-1,3,4-oxadiazole (10i). White solid; mp. 197−198 oC;

386


387

1H, Ar-H), 8.08 (d, J = 7.0 Hz, 1H, Ar-H), 7.53 (dd, J = 6.5, 1.2 Hz, 1H, Ar-H), 7.23–

388

7.27 (m, 2H, Ar-H), 2.98–3.05 (m, 1H, Ar-CH), 2.07–2.11 (m, 2H, ArCH-CH2), 1.76–

389

1.81 (m, 2H, ArCH-CH2), 1.28–1.69 (m, 6H, ArCHCH2-(CH2)3); 13C NMR (100 MHz,

390

DMSO-d6) δ 167.7, 167.1, 136.9, 128.2, 123.2, 121.6. 120.6, 112.8, 100.1, 34.6, 30.2,

391


392

2-Heptyl-5-(1H-indol-3-yl)-1,3,4-oxadiazole (10j). Gray solid; mp. 176−177 oC; yield

393

46%; 1H NMR (400 MHz, CDCl3) δ 9.23 (d, J = 108.1 Hz, 1H, NH), 8.25–8.38 (m, 1H,

394

Ar-H), 7.84 (dd, J = 2.6, 64.6 Hz, 1H, Ar-H), 7.46–7.57 (m, 1H, Ar-H), 7.32–7.35 (m, 20


Page 20 of 44

Page 21 of 44


395

2H, Ar-H), 4.10–4.33 (m, 2H, CH2), 2.80–2.98 (m, 2H, CH2), 1.85–2.09 (m, 3H, CH2),

396

1.25–1.58 (m, 8H, CH2, CH3); 13C NMR (100 MHz, DMSO-d6) δ 164.3, 161.6, 136.4,

397

127.7, 124.1, 122.7, 121.0, 120.2, 112.3, 31.1, 28.3, 28.2, 26.0, 24.4, 22.0, 13.9; HRMS

398


399

Biological Assay. The biological activity of target compounds were tested on the

400

corresponding test tissues. To ensure the accuracy of the data, each group of tests was

401

repeated at least three times. The activity data were expressed as percentage inhibition

402

rates ranging from 0 to 100 (0 means no activity, 100 means total inhibition).

403

Antiviral Biological Assay. The anti-TMV activities of target compounds were obtained

404

by using reported methods10 with tobacco (Nicotianatobaccum var. Xanthi-nc) as the

405

test plant. The detailed procedures also can be seen in Supporting Information.

406

Phytotoxic Activity. The growing 5–6 leaf stage tobaccos (Nicotiana tabacum var

407

Xanthi nc) were selected. The compound solution (500 µg/mL) was smeared on the

408

leaves and calculated the number of lesions after 3-4 days.

409

Antifungal Biological Assay. The fungicidal activities of target compounds were

410

obtained by using literature methods.22 The detailed procedures also can be seen in

411

Supporting Information.

412

RESULTS AND DISCUSSION

413

Chemistry. Till now, the synthesis of pimprinine has been accomplished by Joshi23,

414

Molina24, Wu19 and Zhou25. Considering a series of derivatives of pimprinine would be 21



415

needed, Wu’s method was selected and optimized (Figure 3).25 2-Oxoacetaldehyde 3

416

and corresponding amino acids underwent decarboxylation coupling reaction to give

417

5a–5l in 39%–83% yields. Indole nitrogen substituted products 6aa and 6ab were

418

prepared (Figure 4) to evaluate the influence of the NH on activity.

419

1,3,4-Oxadiazole is a very good bioisostere of amide and ester functional group.

420

Compounds containing a 1,3,4-oxadiazole ring often exhibit broad-spectrum biological

421

activities, such as anticancer26, antifungal27 and anti-HIV28. On the basis of the principle

422

of combination of active structural moieties, a series of pimprinine derivatives

423

containing a 1,3,4-oxadiazole ring were reasonably designed and prepared (Figure 5).

424

The reaction of hydrazide 7 with a number of aldehydes 8a–8u gave acylhydrazones

425

9a–9u (Figures 5 and 6). It is well known that acylhydrazone compounds may exist as

426

isomers (E/Z) due to the carbon-nitrogen double bonds and the rotamers (cis/trans)

427

caused by amide N–C(O) bond.29,30 The hydrazones prepared from aldehydes and

428

hydrazides exists in E-form in solution.29 For acylhydrazones 9a–9u, only E form were

429

detected by NMR in DMSO-d6. Acylhydrazones 9a–9j were transformed into

430

oxadiazoles 10a–10j by reacting with iodobenzene diacetate.

431

Phytotoxic Activity. The first phytotoxic activity evaluation indicated that pimprinine

432

alkaloids and their derivatives 5a–5l, 6aa, 6ab, 7, 9a–9u and 10a–10j displayed no

433

phytotoxic activity to the tested tobaccos at 500 μg/mL. No local lesion appear on the

434

tobacco leaves.

435

Antiviral Activity. The antiviral activities of pimprinine alkaloids and their derivatives 22


Page 22 of 44

Page 23 of 44


436

5a–5l, 6aa, 6ab, 7, 9a–9u and 10a–10j against TMV was shown in Table 1. Currently

437

widely used antivirals ribavirin and ningnanmycin were selected as the controls.

438

In Vitro Anti-TMV Activity. The in vitro anti-TMV activities of pimprinines 5a–5l, 6aa,

439

6ab, 7, 9a–9u and 10a–10j were first tested. Compounds 5g, 5j–5l, 9d, 9e, 9h, 9m, 9o,

440

9p, 9s, 10c, 10d, 10g and 10h exhibited higher in vitro anti-TMV activities than

441

ribavirin, in which, compounds 5l, 9h and 10h displayed similar or higher antiviral

442

activities than ningnanmycin (perhaps the most widely used antiviral agent at present).

443

Pimprinine (5a) showed about similar TMV inhibitory effect with compounds 6aa

444

and 6ab, which indicated that the modification of indole nitrogen is tolerant. The main

445

difference between 5a–5f lies in the size of alkyl substituents, 5c and 5e showed higher

446

inhibitory effect than the others which indicated that increasing steric hindrance is

447

beneficial to the activity. Compounds 5c and 5e exhibited higher inhibitory effect than

448

pimprinine (5a) which indicated that the introduction of hydroxyl and methylthio

449

groups is favorable for the activity. The replacement of methyl with phenyl maintained

450

activity (inhibitory effect: 5a ≈ 5i). However, the replacement of methyl with 4-F

451

phenyl significantly increased activity (inhibitory effect: 5j > 5a) which indicated that

452

the 2-position of pimprinine is a substituent effect sensitive region. Pimprinaphine (5k)

453

displayed significantly higher activity than pimprinine (5a) and 5i which indicated that

454

the benzyl is more beneficial to the activity. Introduction of hydroxy into benzyl further

455

increased antiviral activity (inhibitory effect: 5l > 5k). The replacement of oxazole ring

456

with oxadiazole ring increased antiviral activity (inhibitory effect: 10a > 5i, 10i > 5f).

23



457

The mainly difference between 10a–10e lies in the phenyl substituents, electron

458

deficient substituents are more beneficial to the activity (inhibitory effect: 10d > 10e >

459

10b). Ortho bromophenyl substituent 10c showed higher inhibitory effect than the

460

others. The replacement of phenyl with furany and alkyls decreased antiviral activity

461

(inhibitory effect: 10a > 10f ≈ 10i ≈ 5j). Naphthyl and indolyl substituents 10g and 10h

462

displayed about similar activities with 10c.

463

N-Acylhydrazones also play an important role in medicinal chemistry, as they

464

display broad-spectrum biological activities, such as antiviral31, anticonvulsant32 and

465

antitrypanosomal33. As the ring open products, 7 and 9a–9u were also tested for their

466

antiviral activities. The results revealed that acylhydrazones 9a–9u also showed good

467

antiviral activities. Compounds 9d, 9e, 9h, 9m, 9o, 9p and 9s showed higher activities

468

than ribavirin, especially for compound 9h displayed about similar activity to

469

ningnanmycin. As the ring open products of 10a–10j, 9a–9j displayed lower antiviral

470

activities than 10a–10j except for 9e (inhibitory effect: 9e > 10e) which indicated that

471

the open of oxadiazole ring is bad for antiviral activity. The alkyl substituted derivatives

472

9i, 9j, 9t and 9u exhibited relatively lower activities than the others. Among the phenyl

473

substituted derivatives 9a–9e and 9k–9o, electron deficient substituted compounds 9d,

474

9e, 9o and hydroxyl substituted compound 9m showed relatively higher activities.

475

Heterocyclic substituted derivatives 9f, 9q and 9r displayed about similar activities

476

with 9a. Compounds 9h and 9p exhibited excellent antiviral activities.

477

In Vivo Anti-TMV Activity Pimprinine alkaloids and their derivatives 5a–5l, 6aa, 6ab,

24


Page 24 of 44

Page 25 of 44


478

7, 9a–9u and 10a–10j also were evaluated for their in vivo anti-TMV activities. Most

479

of these compounds displayed good activities. Compounds 5l, 9h and 10h displayed

480

significantly higher in vivo activities than ribavirin. The SARs of in vivo activity are

481

similar to that of in vitro activity. The EC50 values of compounds 5l, 9h and 10h for

482

protection effect were further investigated to further confirm the activity data. As shown

483

in Table 2, ribavirin displayed 694 μg/mL EC50 value which is significantly higher than

484

that of compounds 5l, 9h and 10h. Ningnanmycin showed 203 μg/mL EC50 value which

485

is slightly lower than that of 5l (211 μg/mL) and higher than that of 9h (190 μg/mL)

486

and 10h (180 μg/mL). Compounds 5l, 9h and 10h with similar or higher antiviral

487

activities than ningnanmycin emerged as new lead compounds for antiviral research.

488

Fungicidal Activity. The in vitro fungicidal activities of pimprinine alkaloids and their

489

derivatives 5a–5l, 6aa, 6ab, 7, 9a–9u and 10a–10j were also tested against 14 kinds of

490

phytopathogenic fungi. Currently widely used fungicides carbendazim and

491

chlorothalonil were selected as the controls.

492

As shown in Table 3, all the tested compounds showed broad-spectrum in vitro

493

fungicidal activities at 50 µg/mL. The fungicidal activities of 5d, 6aa and 6ab are

494

higher than that of carbendazim against Cercospora arachidicola Hori. The fungicidal

495

activities of compounds 5b, 5k and 9i are higher than that of carbendazim against

496

Physalospora piricola. Compounds 5d, 5j, 6aa, 6ab and 9a exhibited higher fungicidal

497

activities than did carbendazim against Rhizoctonia cerealis. The fungicidal activities

498

of 5g against Bipolaris maydis were higher than that of chlorothalonil. The fungicidal

25



499

activities of 6ab and 9a against Alternaria solani and 5c, 5g, 6aa, 6ab, 9h, 9i and 9o

500

against Botrytis cinerea were higher than that of carbendazim.

501

In Summary, pimprinine alkaloids and their derivatives 5a–5l, 6aa, 6ab, 7, 9a–9u

502

and 10a–10j were synthesized and evaluated systematically for their antiviral and

503

fungicidal activities. The anti-TMV activities of these compounds were discovered for

504

the first time. Compounds 5l, 9h and 10h with simple structure showed similar or higher

505

in vivo anti-TMV activities than ningnanmycin, thus emerged as new antiviral pilot

506

compounds. The modification of indole nitrogen is tolerant for antiviral research. The

507

replacement of oxazole ring with oxadiazole ring increased antiviral activity. As the

508

ring open products, acylhydrazones 9a–9u were also found to possess good antiviral

509

activity. Moreover, all the synthesized compounds were found to have broad-spectrum

510

in vitro fungicidal activities. This study laid a solid foundation for the application of

511

pimprinine alkaloids and their derivatives in plant protection.

512

FUNDING

513

We gratefully acknowledge assistance from National Natural Science Foundation of

514

China (21772145, 21732002, 21672117), Tianjin Natural Science Foundation

515

(16JCZDJC32400), the Foundation of Development Program of Future Expert in

516

Tianjin Normal University (WLQR201703), Training Program of Outstanding Youth

517

Innovation Team of Tianjin Normal Uiversity and the Program for Innovative Research

518

Team in University of Tianjin (TD13-5074).

519

NOTES 26


Page 26 of 44

Page 27 of 44


520

The authors declare no competing financial interest.

521

SUPPORTING INFORMATION

522

Biological assay procedures, 1H and 13C NMR spectra of pimprinine alkaloids and their

523

derivatives 5a–5l, 6aa, 6ab, 7, 9a–9u and 10a–10j. This material is available free of

524

charge via the Internet at http://pubs.acs.org.

525

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526

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Kumar, D.; Chakrabarti, G. Development of novel bis(indolyl)-hydrazide−hydrazone

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synthesis, and biological activities of arylmethylamine substituted chlorotriazine and

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methylthiotriazine compounds. J. Agric. Food Chem. 2011, 59, 11711−11717.

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synthesis of pimprinine. Tetrahedron 1963, 19, 1437–1439.

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(24) Molina, P.; Fresneda, P. M.; Almendros, P. Iminophosphorane-mediated synthesis

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of 3-(oxazol-5-yl)indoles: application to the preparation of pimprinine type alkaloids.

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Synthesis 1993, 1993, 54–56.

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catalyzed oxidative cross-coupling of azole-4-carboxylates with indoles: an approach 30


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620

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625

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626

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627

628

629

630

Figure Captions

631

Figure 1. Chemical structure of pimprinine alkaloids.

632

Figure 2. Design of pimprinine analogues.

633

Figure 3. Synthesis of compounds 5a–5l.

634

Figure 4. Synthesis of compounds 6aa and 6ab.

635

Figure 5. Synthesis of compounds 9a–9j and 10a–10j.

636

Figure 6. Synthesis of compounds 9k–9u.

637

32


Page 32 of 44

Page 33 of 44


Table 1. Antiviral Activities of Ribavirin, Ningnanmycin and Compounds 5a–5l, 6aa, 6ab, 7, 9a–9u and 10a–10j Against TMV. In vivo

Compd.

Conc. (µg/mL)

In vitro inactivation

curative

protection

effect (%)

effect (%)

effect (%)

inhibition rate (%)

500

27±2

25±3

25±1

20±1

100

0

15±2

10±3

6±3

500

26±4

22±2

21±1

23±3

100

0

10±1

0

9±1

500

34±3

34±4

31±3

32±1

100

11±2

15±2

13±1

9±3

500

31±2

35±1

35±1

39±4

100

0

11±1

9±1

15±3

500

35±1

37±2

34±1

33±1

100

14±2

15±1

10±1

14±2

500

25±3

25±2

26±2

27±2

100

10±2

13±1

12±2

11±3

500

43±2

44±4

41±1

39±1

100

14±2

15±3

10±1

12±2

5a

5b

5c

5d

5e

5f

5g

33



Page 34 of 44

500

35±2

33±2

32±2

34±2

100

12±2

11±1

9±1

10±2

500

28±3

27±2

24±2

23±5

100

0

12±1

12±2

9±2

500

45±3

46±2

38±3

45±3

100

15±1

17±3

18±1

19±2

500

48±1

46±1

48±1

49±1

100

22±3

21±2

19±1

19±2

500

56±1

55±1

56±2

52±1

100

26±3

23±2

25±1

21±1

500

25±2

23±3

27±4

20±1

100

0

11±2

0

8±2

500

25±2

25±2

20±2

26±2

100

0

0

9±2

11±2

500

15±2

10±3

16±1

17±4

100

0

0

0

0

500

27±1

21±4

24±2

26±2

100

0

0

0

0

500

31±2

36±3

32±3

34±2

100

0

8±2

0

11±3

500

38±2

38±4

39±3

40±2

100

15±2

17±2

15±2

18±2

500

42±3

40±2

41±1

40±1

100

16±2

19±4

15±2

12±2

5h

5i

5j

5k

5l

6aa

6ab

7

9a

9b

9c

9d

34


Page 35 of 44


500

46±2

48±3

48±3

46±2

100

20±1

23±2

24±3

18±1

500

26±2

23±1

20±1

24±2

100

10±2

18±1

15±1

11±1

500

33±3

32±1

39±2

31±4

100

9±2

15±2

20±1

12±2

500

50±3

56±3

51±2

55±3

100

19±2

17±3

19±1

21±2

500

21±2

18±4

15±4

27±2

100

0

11±2

6±1

10±1

500

17±3

13±1

10±1

24±2

100

0

0

0

7±2

500

22±2

21±4

28±2

29±3

100

0

14±2

0

0

500

39±2

40±3

42±3

41±1

100

6±3

14±1

18±1

15±4

500

43±3

47±2

41±1

38±2

100

14±2

23±4

18±2

16±1

500

30±2

28±1

29±1

24±1

100

8±3

11±3

12±4

9±1

500

49±2

45±1

40±2

43±1

100

27±2

30±1

15±1

21±2

500

50±3

47±1

48±3

48±2

100

29±2

27±4

21±2

22±1

9e

9f

9g

9h

9i

9j

9k

9l

9m

9n

9o

9p

35



Page 36 of 44

500

22±2

20±3

23±3

26±3

100

0

13±3

7±2

11±3

500

25±5

25±1

22±3

20±2

100

17±2

10±3

10±3

15±4

500

42±2

42±5

48±2

49±3

100

16±2

14±2

15±2

14±2

500

19±2

18±3

22±3

21±1

100

6±3

9±1

8±1

5±1

500

13±3

17±2

11±1

18±2

100

6±2

13±4

8±2

10±1

500

40±2

38±1

39±1

34±1

100

18±3

15±3

18±4

13±1

500

30±1

34±2

33±1

30±2

100

5±1

12±3

14±2

13±2

500

51±3

50±2

48±3

47±3

100

26±1

24±3

21±2

23±1

500

52±2

48±5

53±2

50±3

100

21±2

24±2

22±3

20±2

500

39±2

40±3

42±3

41±1

100

6±3

14±1

18±1

15±4

500

33±3

37±2

31±1

38±2

100

14±2

13±4

18±2

16±1

500

50±2

53±1

49±1

44±1

100

28±3

21±3

24±4

22±1

9q

9r

9s

9t

9u

10a

10b

10c

10d

10e

10f

10g

36


Page 37 of 44


500

59±2

58±1

60±2

63±1

100

27±2

30±1

25±1

27±2

500

30±3

37±1

38±3

34±2

100

13±2

17±4

20±2

19±1

500

32±2

42±5

28±2

29±3

100

0

14±2

0

0

500

40±2

35±1

36±1

38±2

100

13±2

14±3

11±3

15±1

500

55±2

56±1

56±1

58±2

100

24±2

27±3

25±3

27±1

10h

10i

10j

Ribavirin

Ningnanm ycin a

Average of three replicates. All results are expressed as mean ± SD.

Table 2. The EC50 Values of Compounds 5l, 9h, 10h, Ribavirin and Ningnanmycin Against TMV. Protection effect

Compd.

Regression equation

r

5l

y =1.39+1.56x

0.9762

211

9h

y =1.23+1.66x

0.9747

190

10h

y =0.97+1.79x

0.9870

180

Ribavirin

y =1.67+1.17x

0.9945

694

Ningnanmycin

y =1.00+1.73x

0.9803

203

37


EC50 (µg/mL)


Page 38 of 44

Table 3. Fungicidal Activities of Chlorothalonil, Carbendazim and Compounds 5a–5l, 6aa, 6ab, 7, 9a–9u and 10a–10j Against 14 Kinds of Fungi. Fungicidal activitya (%)/ 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

5a

22±1

32±2

14±1

22±2

25±2

38±1

32±2

35±3

48±2

27±2

16±1

28±2

45±1

48±2

5b

18±2

20±3

57±2

23±1

25±2

27±1

18±2

28±2

56±1

35±2

27±2

20±3

37±2

44±1

5c

24±1

37±2

0

12±2

33±1

34±2

36±1

30±2

64±2

27±1

12±2

27±1

30±2

64±1

5d

31±2

57±1

46±2

60±1

36±2

46±3

27±2

34±1

56±2

35±2

32±1

38±2

27±1

27±2

5e

17±2

23±2

32±2

18±2

23±2

39±2

21±2

23±2

66±2

8±2

15±2

23±1

23±2

49±2

5f

30±2

16±2

11±1

33±3

29±2

27±1

30±1

33±2

55±2

12±2

33±1

22±2

41±2

39±3

5g

37±2

27±1

47±2

47±1

55±2

33±3

22±2

49±1

58±2

18±2

23±1

37±2

50±2

58±1

5h

27±1

32±2

20±1

20±2

28±2

28±1

19±2

35±2

48±1

12±2

13±2

13±2

21±3

31±2

5i

14±2

10±3

4±2

23±2

23±3

28±2

22±2

13±1

32±2

8±1

9±2

9±1

14±2

28±3

38


Page 39 of 44


5j

26±2

41±3

20±1

52±2

31±2

30±3

46±2

19±2

45±3

10±2

15±2

28±3

14±2

28±2

5k

20±2

33±1

54±2

48±2

33±1

33±2

41±1

29±2

44±2

18±1

32±2

35±1

37±2

44±2

5l

15±1

46±2

0

41±3

23±2

25±1

15±2

38±1

42±2

14±2

25±1

46±2

37±1

43±2

6aa

22±2

67±1

26±2

72±2

33±3

44±2

30±2

48±1

76±2

55±1

24±2

53±1

40±2

52±1

6ab

22±1

63±2

22±1

73±2

33±3

47±2

30±1

58±2

72±1

68±2

34±2

67±1

37±2

76±1

7

22±2

30±1

22±2

25±1

20±2

42±2

22±1

11±2

10±2

17±1

4±2

25±2

20±3

21±2

9a

7±1

27±2

28±3

56±2

15±1

45±2

15±1

53±1

12±2

17±1

50±3

19±2

19±3

14±1

9b

23±2

23±3

11±2

30±1

15±2

28±1

11±2

33±2

12±1

5±2

35±1

13±2

15±1

25±4

9c

17±1

37±2

27±2

42±3

25±2

25±1

15±2

6±2

23±1

6±2

14±1

42±2

12±1

6±2

9d

10±2

23±3

33±2

14±2

15±2

14±2

15±1

40±1

26±3

6±1

36±2

22±2

27±1

0

9e

12±1

27±2

4±1

14±2

10±1

25±2

22±2

20±1

10±1

6±2

29±2

19±2

8±1

15±1

9f

15±2

20±1

27±2

23±3

15±2

31±1

41±2

13±1

29±3

18±1

7±1

14±2

19±1

44±1

9g

17±1

20±2

37±1

28±2

13±1

18±2

16±1

20±1

16±2

6±1

7±1

11±1

19±2

4±2

9h

22±2

30±2

0

51±1

23±2

42±2

30±2

40±1

48±3

35±1

57±2

47±2

19±1

60±2

9i

32±1

37±2

65±1

32±2

35±1

39±2

37±1

47±2

16±2

18±1

14±2

42±3

62±1

69±1

9j

21±2

10±1

6±2

29±1

19±2

8±1

15±1

20±2

23±1

12±2

36±2

53±1

39±2

6±1

9k

12±2

36±2

38±1

24±2

22±1

32±1

39±2

47±1

19±2

6±1

21±1

11±2

12±1

15±2

9l

19±1

35±2

17±2

42±3

25±2

26±1

25±2

47±1

29±1

47±1

14±1

22±1

19±2

10±1

9m

28±1

33±2

30±1

20±2

28±2

28±1

19±2

33±2

19±1

18±2

7±2

14±2

31±1

10±2

9n

17±1

37±2

14±1

14±2

30±1

25±2

22±2

27±1

13±2

12±2

14±1

11±1

31±2

6±1

9o

18±2

21±1

27±2

33±3

15±2

31±1

41±2

27±1

23±1

18±1

21±1

11±3

19±1

50±1

9p

21±2

33±3

12±2

20±1

17±2

38±1

11±2

7±1

7±1

12±2

29±2

39±2

4±1

13±1

9q

25±1

27±2

19±1

34±2

28±1

28±2

30±1

27±2

32±1

12±1

7±1

14±2

27±1

4±1

9r

14±2

22±2

35±2

28±1

23±2

39±2

21±2

33±1

39±2

18±1

21±1

14±3

19±1

8±1

39



Page 40 of 44

9s

24±2

13±3

21±2

30±1

25±2

28±1

11±2

33±2

26±1

18±1

14±2

22±1

19±2

10±1

9t

29±1

35±2

35±1

31±2

35±1

37±2

37±1

33±2

16±1

18±2

21±1

11±2

23±1

46±2

9u

11±1

19±2

21±2

29±1

19±2

8±1

15±1

33±2

13±1

12±2

21±1

14±2

15±1

15±1

10a

17±1

36±2

38±1

24±2

22±1

32±1

39±2

7±1

16±2

9±1

7±1

19±2

8±2

25±1

10b

22±2

33±2

19±3

37±3

24±2

29±3

23±2

16±3

33±3

18±2

9±2

36±3

12±1

45±3

10c

21±4

31±2

27±1

24±3

22±4

27±1

23±2

11±1

34±1

15±2

11±2

23±3

17±2

41±2

10d

9±1

27±2

14±1

14±2

20±1

25±2

22±2

7±1

7±1

6±1

21±2

14±1

8±1

4±1

10e

11±2

25±1

27±2

33±3

18±2

21±1

41±2

7±1

16±2

6±1

7±1

19±2

4±1

33±1

10f

19±2

33±3

12±2

22±1

17±2

48±1

11±2

27±1

48±2

12±1

14±2

44±1

15±2

46±2

10g

26±1

22±2

19±1

34±2

28±1

38±2

30±1

27±1

23±2

12±1

7±1

39±2

8±2

35±1

10h

11±2

22±2

35±2

38±2

23±2

39±2

21±2

27±2

13±2

6±1

7±1

33±2

12±2

25±2

10i

14±2

13±3

21±2

33±1

25±2

28±1

11±2

47±1

32±2

18±2

36±2

42±1

12±1

35±1

10j

16±2

30±1

44±2

48±1

33±2

42±1

33±2

27±1

19±2

12±2

21±2

42±1

8±1

29±2

chlorothalonilc

100

73±2

100

73±1

Discovery of Pimprinine Alkaloids as Novel Agents against a Plant

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