Synthesis, Herbicidal Activity, and QSAR of Novel ... - ACS Publications

Subscriber access provided by UNIVERSITY OF LEEDS

Article

Synthesis, Herbicidal Activity and QSAR of Novel N-Benzothiazolylpyrimidine-2,4-diones as Protoporphyrinogen Oxidase Inhibitors Yang Zuo, Qiongyou Wu, Sun-Wen Su, Cong-Wei Niu, Zhen Xi, and Guang-Fu Yang J. Agric. Food Chem., Just Accepted Manuscript • DOI: 10.1021/acs.jafc.5b05378 • Publication Date (Web): 05 Jan 2016 Downloaded from http://pubs.acs.org on January 9, 2016

Just Accepted “Just Accepted” manuscripts have been peer-reviewed and accepted for publication. They are posted online prior to technical editing, formatting for publication and author proofing. The American Chemical Society provides “Just Accepted” as a free service to the research community to expedite the dissemination of scientific material as soon as possible after acceptance. “Just Accepted” manuscripts appear in full in PDF format accompanied by an HTML abstract. “Just Accepted” manuscripts have been fully peer reviewed, but should not be considered the official version of record. They are accessible to all readers and citable by the Digital Object Identifier (DOI®). “Just Accepted” is an optional service offered to authors. Therefore, the “Just Accepted” Web site may not include all articles that will be published in the journal. After a manuscript is technically edited and formatted, it will be removed from the “Just Accepted” Web site and published as an ASAP article. Note that technical editing may introduce minor changes to the manuscript text and/or graphics which could affect content, and all legal disclaimers and ethical guidelines that apply to the journal pertain. ACS cannot be held responsible for errors or consequences arising from the use of information contained in these “Just Accepted” manuscripts.

Journal of Agricultural and Food Chemistry is published by the American Chemical Society. 1155 Sixteenth Street N.W., Washington, DC 20036 Published by American Chemical Society. Copyright © American Chemical Society. However, no copyright claim is made to original U.S. Government works, or works produced by employees of any Commonwealth realm Crown government in the course of their duties.

Page 1 of 54

Journal of Agricultural and Food Chemistry

1

Synthesis, Herbicidal Activity and QSAR of Novel N-Benzothiazolyl-

2

pyrimidine-2,4-diones as Protoporphyrinogen Oxidase Inhibitors

3 4

Zuo Yang,† Wu Qiongyou,† Su Sun-wen,† Niu Cong-wei,‡ Xi Zhen‡ and Yang Guang-Fu†,*

5 6

†

7

Chemistry, Central China Normal University, Wuhan 430079, P.R.China;

8

‡

9

R. China

Key Laboratory of Pesticide & Chemical Biology of Ministry of Education, College of

State Key Laboratory of Elemento-Organic Chemistry, Nankai University, Tianjin 300071, P.

10 11 12 13 14 15

*Corresponding

author

16

[email protected])

(Tel:

+86-27-67867800;

Fax:

17 18 19 20 21 22

1 / 54

ACS Paragon Plus Environment

+86-27-67867141;

E-mail:


Page 2 of 54

23

ABSTRACT Protoporphyrinogen oxidase (PPO, E.C. 1.3.3.4) is known as a key action

24

target for several structurally diverse herbicides. As a continuation of our research work on

25

the development of new PPO-inhibiting herbicides, a series of novel 3-(2´-halo-5´-

26

substituted-benzothiazol-1´-yl)-1-methyl-6-(trifluoromethyl)pyrimidine-2,4-diones

27

designed and synthesized. The bioassay results indicated that a number of the newly

28

synthesized compounds exhibited higher inhibition activity against tobacco PPO (mtPPO)

29

than the controls, saflufenacil and sulfentrazone. Compound 9F-5 was identified as the most

30

potent inhibitor with Ki value of 0.0072 µM against mtPPO, showing about 4.2-fold and 1.4-

31

fold higher potency than sulfentrazone (Ki = 0.03 µM) and saflufenacil (Ki = 0.01 µM),

32

respectively. Further green house assay demonstrated that compound 9F-6 (Ki = 0.012 µM)

33

displayed the most promising post-emergence herbicidal activity with broad spectrum even at

34

a concentration as low as 37.5 g active ingredient (ai)/ha. Maize exhibits relative tolerance

35

against compound 9F-6 at the dosage of 150 g ai/ha, but it is susceptible to saflufenacil even

36

at 75 g ai/ha. Thus, compound 9F-6 exhibits the potential as a new herbicide for the weed

37

control in maize fields.

38

Keywords: Molecular design; Protoporphyrinogen oxidase; Pyrimidinedione; Benzothiazole;

39

Herbicide.

40 41 2 / 54


9

were

Page 3 of 54

42


Introduction

43

Protoporphyrinogen oxidase (PPO; EC 1.3.3.4), the last common enzyme in the

44

biosynthetic pathway leading to chlorophyll in plant and heme in animal,1-4 has been

45

identified as one of the most significant targets in herbicide discovery.5 During the last thirty

46

years, a number of structurally diverse PPO-inhibiting type herbicides have been

47

commercialized, such as diphenyl ethers, thiadiazoles, phenylpyrazoles, oxadiazoles,

48

triazolinones, oxazolidinedione, N-phenyl-phthalimides and pyrimidinediones.6 These

49

herbicides can cause peroxidative destruction of cellular membrane by blocking the oxidation

50

of protogen which is a prerequisite step for the synthesis of chlorophylls and bleaching of

51

plant tissues in the presence of light.7 In contrast to herbicides with other modes of action,

52

PPO herbicides have many advantages such as low use-rate, broad herbicidal spectrum, quick

53

onset of action, long lasting effect and environmentally benign characteristics.8

54

Among the chemical families mentioned above, pyrimidinedione derivatives have attracted

55

considerable attention in the field of agrochemistry during recent decades.9 Butafenacil,10

56

benzfendizone11-12 and saflufenacil13 (Figure 1) are three representative commercialized

57

pyrimidinedione type herbicides. Among them, butafenacil is a non-selective pre-emergence

58

herbicide, derived from flupropacil by Syngenta in 1998, and has been successfully used for

59

the weed control in the field of vines, cotton, maize and cereal crop. Benzfendizone is a post-

60

emergence herbicide developed by FMC, showing good control for grass and broadleaf weeds 3 / 54



61

in tree fruits and vines, and has been applied in total vegetation control. Saflufenacil is a

62

relatively new herbicide being developed by BASF in 2010 and is marketed under the Kixor

63

trademark. Saflufenacil was used as a pre-emergence dicotyledon weed control herbicide in

64

several crops, especially for cereals. This product achieves the pre-emergence selectivity,

65

mainly based on the stage of treatment and is known to have high potential for reaching

66

surface water via runoff of the rainwater for several weeks after application.

67

The existing pyrimidinedione-type herbicides always possess a common structural feature:

68

a (2′,4′,6′-trisubstituted)-phenyl moiety (Figure 2). Substituting a chlorine or fluorine atom on

69

C2′-position, and/or C4′-position of the benzene ring was found to be most favorable to

70

achieve high PPO inhibition activity, while a variety of groups were found to be acceptable at

71

C6′-position. For the pyrimidine ring unit, it is clear that substituting a methyl group at the

72

N1-position and a trifluoromethyl in the C6-position at the same time always result in better

73

activity.14 Additionally, linking the substituents at 4′- and 6′-positions together or assembling

74

6′-position and 7′-position of the phenyl ring to furnish a benzoheterocyclic ring was

75

demonstrated to be an effective strategy for discovering novel protoporphyrinogen oxidase

76

inhibitors.15 Two notable examples of this ring-closure strategy are commercial herbicides

77

flumioxazin16 and thiadiazimin,17 in which benzooxazin-3-one was introduced by assembling

78

the substituent at C-4′ and C-6′ into a six-member oxazinone ring. Benzothiazole, an

79

important heterocycle with a wide range of biological activities including anticancer, antiviral, 4 / 54


Page 4 of 54

Page 5 of 54


80

antimicrobial, and antifungal activity,18-22 has been widely applied in agrochemistry.23-24

81

Previously, we successfully integrated benzothiazole moiety with diverse heterocycles to

82

develop novel agrochemicals with high-potential fungicidal or herbicidal activities.5,6, 25-31 It

83

was revealed that the incorporation of benzothiazole with heterocycles such as triazolinone or

84

tetrahydro-2H-isoindole-1,3-dione could result in both an enhancement in herbicidal potency

85

and improvement of the crop selectivity. Inspired by these findings, herein, we report the

86

design and synthesis of a series of novel PPO inhibitors, 1-methyl-3-phenyl-6-

87

(trifluoromethyl)pyrimidine-2,4-dione

88

pyrimidine-2,4-dione component (Figure 2). These newly synthesized compounds were

89

characterized by 1H NMR,

90

Their PPO inhibition activities and herbicidal potency were evaluated, and a number of

91

compounds with high PPO-inhibiting activity and broad spectrum herbicidal activity was

92

discovered. Importantly, compound 9F-6, showing high safety towards maize even at the

93

dosage of 150 g ai/ha, was recognized as a potential herbicide candidate for weed control in

94

maize fields.

13

C NMR,

by

19

incorporating

benzothiazole

moiety

with

F NMR, mass spectroscopy and elemental analysis.

95 96

MATERIALS AND METHODS

97

All chemical reagents were commercially available. Solvents, such as dichloromethane,

98

dimethylformamide, tetrahydrofuran, petroleum ether and acetone were purchased from 5 / 54



Page 6 of 54

99

Sinopharm Chemical Reagent Co., Ltd, were dried and redistilled before use. Chemical

100

reagents, such as 2,4-disubstituted aniline, methyl chloroacetate and ethyl 4,4,4-

101

trifluoroacetoacetate were purchased from Alfa aesar.

102

VARIAN Mercury-Plus 600 or 400 spectrometer (Varian, Palo Alto, California). Mass

103

spectral

104

electrosprayionization (ESI-MS) (Thermo Fisher, Sillicon Valley, California). Elemental

105

analyses were performed on a Vario EL III elemental analysis instrument (Elementar, Hanau,

106

Germany). Melting points were taken on a Buchi B-545 melting point apparatus and were

107

uncorrected.

data

were

obtained

on

a

Thermo

1

H NMR spectra were recorded on a

Fisher

Mass

platform

DSQII

by

108 109

Preparation of 2,4-disubtituted phenylcarbamates, 2a-c.

110

To a stirred solution of 2,4-disubstituted aniline (0.05 mol) in 100 mL of anhydrous

111

methylene chloride was added pyridine ( 7.9 g, 0.1 mol) under ice bath, and followed by

112

dropwise addition of a solution of methyl chloroacetate (5.4 g, 0.05 mol) in 20 mL of

113

anhydrous methylene chloride within 30 min. The reaction mixture was stirred for an

114

additional 1 h and extracted with methylene chloride, and the combined extracts were washed

115

with 2N hydrochloric acid. The organic layer was dried over sodium sulfate, filtered, and

116

concentrated under reduced pressure to give 2,4-disubtituted phenylcarbamates.

117 6 / 54


Page 7 of 54


118

Preparation of ethyl 3-amino-4,4,4-trifluorobut-2-enoate, 3.

119

To a stirred solution of ethyl 4,4,4-trifluoroacetoacetate (18.4 g, 0.1 mol) in ethanol (125 mL)

120

was added ammonium acetate (30.8 g, 0.4 mol) and the mixture was refluxed for 8 h. After

121

cooling to room temperature, the solution was poured into water and extracted with methyl

122

chloroacetate. The organic layer was dried over sodium sulfate, filtered, and concentrated

123

under reduced pressure to give ethyl 3-amino-4,4,4-trifluorobut-2-enoate.

124 125

Preparation of 3-(2′,4′-disubstituted phenyl)-1-methyl-6-(trifluoromethyl)pyrimidine-

126

2,4-diones, 5a-c.

127

To a stirred solution of 3 (7.3 g, 0.04 mol) in 100 mL anhydrous dimethylformamide was

128

added a 60% sodium hydride (1.92 g, 0.048mol) at 0℃ and the mixture was stirred for 30 min.

129

Thereafter, a solution of the carbamate, 2, in 50 mL anhydrous dimethylformamide was added

130

dropwise within 30 min, the mixture was stirred for 30 min and heated to 120℃ for 6 h to

131

synthesize the product, 4. The reaction mixture was cooled to room temperature and

132

potassium carbonate was added (6.6 g, 0.048 mol), then the reaction mixture was treated with

133

methyl iodide (11.4 g, 0.08 mol) and stirred overnight. The reaction mixture was poured into

134

800 mL of ice water and the precipitate was collected by filtration, washed with water and

135

dried to give the dione, 5.

136 7 / 54



137

Preparation

138

pyrimidine-2,4-dione, 6a-c.

139

To a stirred solution of 3-(2,4-disubstitutedphenyl)-1-methyl-6-(trifluoromethyl)pyrimidine-

140

2,4-dione (0.026 mol) in 52 mL of concentrated H2SO4 was slowly added 68% nitric acid

141

(3.12 g, 0.031 mol) at 0℃. The temperature was maintained at 0℃ for 1 h, and then the

142

reaction mixture was poured into ice water. The precipitate was collected by filtration, washed

143

with water and dried to give 3-(2′,4′-disubstituted-5′-nitrophenyl)-1-methyl-6-(trifluoro-

144

methyl)pyrimidine-2,4-dione.

of

3-(2′,4′-disubstituted-5′-nitrophenyl)-1-methyl-6-(trifluoromethyl)-

145 146

Preparation

147

pyrimidine-2,4-dione, 7a-c.

148

Iron powder (2.67 g, 0.05 mol) was added portionwise to a stirred solution of 6 (0.025 mol),

149

NH4Cl (2.66 g, 0.05 mol) in a mixture of ethanol (75 mL) and water (10 mL) at reflux

150

temperature. The reaction mixture was refluxed for 6 h and filtered through diatomaceous

151

earth, and the filtrate was concentrated under reduced pressure. The residue was extracted

152

with ethyl acetate, and the organic residue was washed with brine. Then the residue was dried

153

over sodium sulfate, filtered, and concentrated under reduced pressure to give 3-(5-amino-2,4-

154

disubstitutedphenyl)-1-methyl-6-(trifluoromethyl)pyrimidine-2,4-dione.

of

3-(5′-amino-2′,4′-disubstitutedphenyl)-1-methyl-6-(trifluoromethyl)-

155 8 / 54


Page 8 of 54

Page 9 of 54


156

Preparation of 3-(2’-substitued-5′-mercaptobenzo[d]thiazol-1′-yl)-1-methyl-6-(trifluoro-

157

methyl)-pyrimidine-2,4-dione, 8a-c.

158

To a stirred solution of 7 (0.02 mol) in 50 mL of DMF was added potassium O-ethyl

159

dithiocarbonate (8.0 g, 0.05 mol) at 90 ℃. The temperature was maintained at 90 ℃ for 6 h,

160

the reaction mixture was then poured into 500 mL of water and acidified by concentrated HCl

161

solution to pH 3 to induce precipitation. The precipitate was collected by filtration, washed

162

with water and dried to obtain 3-(6-substitued-2-mercaptobenzo- [d]thiazol-5-yl)-1-methyl-6-

163

(trifluoromethyl)pyrimidine-2,4-dione.

164 165

General procedure for the synthesis of the title compounds, 9.

166

To a stirred solution of 1 mmol of the intermediate 8 in acetone (20 mL) was added potassium

167

carbonate (1.2 mmol). The mixture was stirred for 10 min before the addition of the

168

corresponding halide derivative (1 mmol). The progress of the reaction was monitored by

169

TLC (petroleum ether/acetone 5:1, UV for visualization) and when it was completed, the

170

reaction mixture was filtered and concentrated. The residue was purified by column

171

chromatography on silica gel using 10:1 petroleum ether/acetone as an eluent.

172 173

Data for 9F-1. Yield: 68%. Yellow oil. 1H NMR (600 MHz, CDCl3) δ 7.72 (d, J = 6.6 Hz,

174

1H), 7.63 (d, J = 9.0 Hz, 1H), 6.39 (s, 1H), 4.25 (q, J = 7.2 Hz, 2H), 4.16 (s, 2H), 3.57 (s, 3H), 9 / 54



13

175

1.29 (t, J = 7.2 Hz, 3H).

C NMR (150 MHz, CDCl3) δ 167.86, 166.14, 160.13, 155.45,

176

153.79, 150.83, 149.25, 141.60 (q, J= 33 Hz), 137.39 (d, J= 10.5 Hz), 122.24, 121.01 (d, J=

177

16.5Hz), 119.30 (q, J= 273 Hz), 108.55 (d, J= 25.5Hz), 103.04 (d, J= 6Hz), 62.07, 35.14,

178

32.62, 14.03. 19F NMR (376 MHz, CDCl3) δ: -66.94, -125.65. ESI-MS: 486.1(M+Na)+. Anal.

179

Calcd for C17H13F4N3O4S2: C, 44.06; H, 2.83; N, 9.07; Found: C, 44.28; H, 2.61; N, 9.30;

180 181


182

1H), 7.63 (d, J = 9.0 Hz, 1H), 6.40 (s, 1H), 4.19 (q, J = 7.2 Hz, 2H), 3.60-3.58 (m, 5H), 3.59

183

(t, J = 6.0 Hz, 2H), 1.28 (t, J = 7.2 Hz, 3H). 13C NMR (150 MHz, CDCl3) δ 171.38, 167.23,

184

160.13, 155.29, 153.64, 150.83, 149.62, 141.54 (q, J= 33 Hz), 137.27 (d, J= 9 Hz), 122.00,

185

120.84 (d, J= 16.5 Hz), 119.28 (q, J= 273 Hz), 108.44 (d, J= 25.5 Hz), 103.03, 60.80, 34.05,

186

32.60, 28.17, 14.06. 19F NMR (376 MHz, CDCl3) δ: -66.91, -125.43. ESI-MS: 500.0(M+Na)+.

187

Anal. Calcd for C18H15F4N3O4S2: C, 45.28; H, 3.17; N, 8.80; Found: C, 45.34; H, 2.92; N,

188

8.76;

189 190


191

1H), 7.62 (d, J = 8.4 Hz, 1H), 6.40 (s, 1H), 4.15 (q, J = 7.2 Hz, 2H), 3.58 (s, 3H), 3.41 (t, J =

192

7.2 Hz, 2H), 2.50 (t, J = 7.2 Hz, 2H), 2.17 – 2.15 (m, 2H), 1.26 (t, J = 7.2 Hz, 3H). 13C NMR

193

(100 MHz, CDCl3) δ 172.83, 172.48, 167.79, 163.74, 160.08, 156.20, 153.71, 150.84, 149.81, 10 / 54


Page 10 of 54

Page 11 of 54


194

141.58 (q, J= 33 Hz), 138.12 (d, J= 11 Hz), 121.53 (q, J= 273 Hz), 121.15 (q, J= 17 Hz),

195

117.93, 108.29 (d, J= 25 Hz), 103.00, 61.81, 60.41, 54.31, 32.67, 32.56, 26.21, 24.46, 14.09,

196

13.91.

197

for C19H17F4N3O4S2: C, 46.43; H, 3.49; N, 8.55; Found: C, 46.64; H, 3.34; N, 8.43;

19

F NMR (376 MHz, CDCl3) δ: -66.93, -125.57. ESI-MS: 492.1(M+H)+. Anal. Calcd

198 199


200

1H), 7.64 (d, J = 12.6 Hz, 1H), 6.42 (s, 1H), 4.72 (q, J = 9.0 Hz, 1H), 4.22 (q, J = 9.0 Hz, 2H),

201

3.59 (s, 3H), 1.71 (d, J = 9.0 Hz, 2H), 1.28 (t, J = 9.0 Hz, 3H). 13C NMR (100 MHz, CDCl3) δ

202

171.20, 165.61, 160.09, 155.97, 153.48, 150.85, 149.56, 141.62 (q, J= 33 Hz), 137.53 (d, J=

203

10 Hz), 122.41, 121.07 (d, J= 16 Hz), 119.35 (q, J= 273 Hz), 108.45 (q, J= 25 Hz), 103.09,

204

61.86, 45.31, 32.62, 17.93, 14.01.

205

500.1(M+Na)+. Anal. Calcd for C18H15F4N3O4S2: C, 45.28; H, 3.17; N, 8.80; Found: C, 45.54;

206

H, 3.02; N, 8.66;

19

F NMR (376 MHz, CDCl3) δ: -66.89, -125.75. ESI-MS:

207 208


209

1H), 7.63 (d, J = 9.0 Hz, 1H), 6.41 (s, 1H), 4.20 (q, J = 7.2 Hz, 2H), 3.59 (s, 3H), 1.75 (s, 6H),

210

1.24 (t, J = 7.2 Hz, 3H). 13C NMR (100 MHz, CDCl3) δ 172.88, 163.81, 160.12, 156.26,

211

153.77, 150.89, 149.81, 141.65 (q, J= 33 Hz), 138.16 (d, J= 11 Hz), 123.03, 121.21 (d, J= 15

212

Hz), 119.36 (q, J= 273 Hz), 108.34 (q, J= 25 Hz), 103.06, 61.88, 54.38, 32.64, 26.27, 13.97. 11 / 54



213

19

214

C19H17F4N3O4S2: C, 46.43; H, 3.49; N, 8.55; Found: C, 46.59; H, 3.33; N, 8.47;

Page 12 of 54

F NMR (376 MHz, CDCl3) δ: -66.98, -125.68. ESI-MS: 514.1(M+Na)+. Anal. Calcd for

215 216

Data for 9F-6. Yield: 32%. White solid. m.p. 136-138℃. 1H NMR (600 MHz, CDCl3) δ 7.72

217

(d, J = 6.6 Hz, 1H), 7.62 (d, J = 9.0 Hz, 1H), 6.40 (s, 1H), 4.16 (s, 1H), 4.14(t, J = 6.6 Hz, 2H),

218

3.58 (s, 3H), 1.69 – 1.60 (m, 2H), 0.94 (t, J = 7.2 Hz, 4H). 13C NMR (100 MHz, CDCl3) δ

219

167.91, 166.06, 160.09, 155.91, 153.44, 150.86, 149.39, 141.61 (q, J= 33 Hz), 137.45 (d, J=

220

11 Hz), 122.32, 121.07 (d, J= 15 Hz), 119.36 (q, J= 273 Hz), 108.51 (d, J= 25 Hz), 103.06,

221

67.58, 35.13, 32.62, 21.82, 10.20.

222


223

H, 2.94; N, 9.08;

19


224 225


226

1H), 7.62 (d, J = 9.0 Hz, 1H), 6.40 (s, 1H), 5.11-5.07 (m, 1H), 4.11 (s, 2H), 3.58 (s, 3H), 1.26

227

(d, J = 6.6 Hz, 6H). 13C NMR (100 MHz, CDCl3) δ 167.34, 166.11, 160.11, 155.91, 153.42,

228

150.87, 149.48, 141.62 (q, J= 33 Hz), 137.50 (d, J= 10 Hz), 122.29, 121.04 (d, J= 15 Hz),

229

119.35 (q, J= 273 Hz), 108.52 (d, J= 25 Hz), 103.08, 69.80, 35.52, 32.64, 21.64.

230

(376 MHz, CDCl3) δ: -66.95, -125.57. ESI-MS: 500.2(M+Na)+. Anal. Calcd for

231

C18H15F4N3O4S2: C, 45.28; H, 3.17; N, 8.80; Found: C, 45.22; H, 3.42; N, 8.84. 12 / 54


19

F NMR

Page 13 of 54


232 233


234

(d, J = 6.0 Hz, 1H), 7.76 (d, J = 9.0 Hz, 1H), 6.42 (s, 1H), 4.00 (s, 3H), 3.59 (s, 3H).

235

NMR (150 MHz, CDCl3) δ 166.31, 160.18, 159.42, 156.22, 154.55, 150.86, 148.42, 141.71(q,

236

J= 33 Hz), 138.20 (d, J= 9 Hz), 123.91, 121.65, 119.32 (q, J= 273 Hz), 108.49 (d, J= 24 Hz),

237

103.09, 55.64, 32.70.

238


239

H, 2.00; N, 9.77.

19

13

C


240 241


242

(d, J = 6.0 Hz, 1H), 7.76 (d, J = 9.0 Hz, 1H), 6.41 (s, 1H), 4.47 (q, J = 7.2 Hz 2H), 3.59 (s,

243

3H), 1.41 (t, J = 7.2 Hz 3H). 13C NMR (150 MHz, CDCl3) δ 165.61, 160.11, 159.80, 156.15,

244

154.48, 150.82, 148.36, 141.62 (q, J= 33 Hz), 138.11 (d, J= 10.5 Hz), 123.80, 121.52(d, J=

245

16.5 Hz), 119.30 (q, J= 273 Hz), 108. (d, J= 25.5 Hz), 103.08, 65.64, 32.64, 14.14. 19F NMR

246

(376 MHz, CDCl3) δ: -66.91, -124.99. ESI-MS: 472.1(M+Na)+. Anal. Calcd for

247

C16H11F4N3O4S2: C, 42.76; H, 2.47; N, 9.35; Found: C, 42.57; H, 2.44; N, 9.56.

248 249


250

1H), 7.63 (d, J = 9.0 Hz, 1H), 6.40 (s, 1H), 4.18 (s, 2H), 3.79 (s, 3H), 3.58 (s, 3H). 13C NMR 13 / 54



251

(150 MHz, CDCl3) δ 168.41, 165.85, 160.14, 155.42, 153.76, 150.83, 149.34, 141.59 (q, J=

252

33 Hz), 137.46 (d, J= 10.5 Hz), 122.29, 120.96 (d, J= 16.5 Hz), 119.28 (q, J= 273 Hz), 108.53

253

(d, J= 25.5 Hz), 103.04, 103.01, 52.96, 34.74, 32.64. 19F NMR (376 MHz, CDCl3) δ: -66.84, -

254

124.98. ESI-MS: 472.0(M+Na)+. Anal. Calcd for C16H11F4N3O4S2: C, 42.76; H, 2.47; N, 9.35;

255

Found: C, 42.82; H, 2.57; N, 9.22.

256 257


258

1H), 7.63 (d, J = 9.0 Hz, 1H), 6.41 (s, 1H), 4.70 (q, J = 7.2 Hz, 1H), 3.77 (s, 3H), 3.60 (s, 3H),

259

1.71 (d, J = 7.2 Hz, 3H).

260

153.73, 150.74, 149.42, 141.46 (q, J= 33 Hz), 137.39 (d, J= 10.5 Hz), 122.29, 120.94 (d, J=

261

16.5 Hz), 119.21 (q, J= 273 Hz), 108.41 (d, J= 25 Hz), 102.91, 52.77, 44.81, 32.51, 17.75. 19F

262

NMR (376 MHz, CDCl3) δ: -66.90, -125.63. ESI-MS: 488.2(M+Na)+. Anal. Calcd for

263

C17H13F4N3O4S2: C, 44.06; H, 2.83; N, 9.07; Found: C, 44.23; H, 2.67; N, 8.99.

13

C NMR (150 MHz, CDCl3) δ 171.64, 165.35, 160.02, 155.39,

264 265

Data for 9F-12. Yield: 38%. White solid. m.p. 170-172℃. 1H NMR (600 MHz, CDCl3) δ

266

7.79 (d, J = 6.0 Hz, 1H), 7.65 (d, J = 9.0 Hz, 1H), 6.41 (s, 1H), 4.11 (d, J = 2.4 Hz, 2H), 3.59

267

(s, 3H), 2.30 (s, 1H). 13C NMR (150 MHz, CDCl3) δ 165.71, 160.17, 155.48, 153.82, 150.87,

268

149.53, 141.63 (q, J= 33 Hz), 137.48 (q, J= 10.5 Hz), 122.44, 121.04 (d, J= 16.5 Hz), 119.30

269

(q, J= 273 Hz), 108.54 (q, J= 25 Hz), 103.06, 77.95, 72.45, 32.67, 21.66. 19F NMR (376 MHz, 14 / 54


Page 14 of 54

Page 15 of 54


270

CDCl3) δ: -66.94, -125.64.. ESI-MS: 416.1(M+H)+. Anal. Calcd for C16H9F4N3O4S2: C,

271

46.26; H, 2.18; N, 10.12; Found: C, 46.20; H, 2.22; N, 10.39.

272 273

Data for 9F-13. Yield: 41%. White solid. m.p. 126-128℃. 1H NMR (600 MHz, CDCl3) δ

274

7.77 (d, J = 6.6 Hz, 1H), 7.63 (d, J = 9.0 Hz, 1H), 6.41 (s, 1H), 6.03-5.97 (m, 1H), 5.38 (d, J =

275

6.6 Hz, 1H), 5.22 (d, J = 6.6 Hz, 1H), 4.00 (d, J = 7.2 Hz, 2H), 3.59 (s, 3H). 13C NMR (150

276

MHz, CDCl3) δ 167.26, 160.13, 155.35, 153.69, 150.86, 149.69, 141.56 (q, J= 33 Hz), 137.32

277

(d, J= 10.5 Hz), 131.95, 122.12, 120.89 (d, J=16.5 Hz), 119.31 (q, J= 273 Hz), 119.30, 108.41

278

(q, J= 25 Hz), 103.05, 36.20, 32.63. 19F NMR (376 MHz, CDCl3) δ: -66.95, -125.62. ESI-MS:

279

418.0(M+H)+. Anal. Calcd for C16H11F4N3O4S2: C, 46.04; H, 2.66; N, 10.07; Found: C, 46.15;

280

H, 2.45; N, 10.23.

281 282

Data for 9Cl-1. Yield: 44%. White solid. m.p. 133-134℃. 1H NMR (400 MHz, CDCl3) δ

283

7.94 (s, 1H), 7.74 (s, 1H), 6.41 (s, 1H), 4.24 (q, J = 10.8 Hz, 2H), 4.16 (s, 2H), 3.59 (s, 3H),

284

1.29 (t, J = 10.8 Hz, 3H).

285

150.78, 141.62 (q, J= 33 Hz), 137.63, 130.44, 128.11, 122.39, 122.05, 119.35 (q, J = 273 Hz),

286

103.11,

287

C17H13ClF3N3O4S2: C, 42.55; H, 2.73; N, 8.76; Found: C, 42.56; H, 2.59; N, 8.97.

62.03,

35.09,

13

C NMR (100 MHz, CDCl3) δ 167.73, 167.39, 160.08, 152.10,

32.54,

14.03.

ESI-MS:

502.2(M+Na)+.

288 15 / 54


Anal.

Calcd

for


289

Data for 9Cl-2. Yield: 42%. Yellow oil. 1H NMR (400 MHz, CDCl3) δ 7.93 (s, 1H), 7.75 (s,

290

1H), 6.41 (s, 1H), 4.19 (q, J = 7.2 Hz, 2H), 3.61-3.58 (m, 5H) 2.91 (t, J = 7.2 Hz, 2H)., 1.28 (t,

291

J = 7.2 Hz, 3H).

292

141.56 (q, J= 33 Hz), 137.41, 130.24, 127.76, 122.08, 121.94, 119.26 (q, J = 273 Hz), 103.08,

293

60.80, 33.97, 32.49, 28.13, 14.17, 13.90.

294

516.1(M+Na)+. Anal. Calcd for C18H15ClF3N3O4S2: C, 43.77; H, 3.06; N, 8.51; Found: C,

295

43.96; H, 2.89; N, 8.43.

13

C NMR (150 MHz, CDCl3) δ 171.34, 168.65, 160.13, 152.30, 150.74,

19

F NMR (376 MHz, CDCl3) δ: -66.91. ESI-MS:

296 297


298

1H), 6.41 (s, 1H), 4.14 (q, J = 7.2 Hz, 2H), 3.59 (s, 3H), 3.41 (t, J = 7.2 Hz, 2H), 2.50 (t, J =

299

7.2 Hz, 2H), 2.17 – 2.15 (m, 2H), 1.26 (t, J = 7.2 Hz, 3H).

300

172.48, 169.02, 160.09, 152.38, 150.73, 141.53 (q, J= 33 Hz), 137.34, 130.21, 127.68, 122.04,

301

121.87, 119.26 (q, J= 273 Hz), 103.04, 60.42, 32.62, 32.55, 24.35, 14.06. 19F NMR (376 MHz,

302

CDCl3) δ: -66.94. ESI-MS: 530.0(M+Na)+. Anal. Calcd for C19H17ClF3N3O4S2: C, 44.93; H,

303

3.37; N, 8.27; Found: C, 45.12; H, 3.22; N, 8.19.

13

C NMR (150 MHz, CDCl3) δ

304 305


306

1H), 6.41 (s, 1H), 4.69 (q, J = 6.6 Hz, 1H), 4.22 (q, J = 7.2 Hz, 2H), 3.59 (s, 3H), 1.71 (d, J

307

=7.2 Hz, 3H), 1.28 (t, J = 7.2 Hz, 3H). 13C NMR (150 MHz, CDCl3) δ 171.10, 166.99, 160.07, 16 / 54


Page 16 of 54

Page 17 of 54


308

152.17, 150.70, 141.57 (q, J= 33 Hz), 137.57, 130.31, 128.03, 122.33, 121.98, 119.27 (q, J=

309

273 Hz), 103.05, 61.83, 45.17, 32.52, 17.83, 13.94.

310

ESI-MS: 516.2(M+Na)+. Anal. Calcd for C18H15ClF3N3O4S2: C, 43.77; H, 3.06; N, 8.51;

311

Found: C, 44.01; H, 2.86; N, 8.38.

19

F NMR (376 MHz, CDCl3) δ: -66.91.

312 313


314

1H), 6.41 (s, 1H), 4.20 (q, J = 7.2 Hz, 2H), 3.59 (s, 3H), 1.76 (s, 6H), 1.24 (t, J = 7.2 Hz, 3H).

315

13

316

137.81, 130.20, 128.23, 122.63, 121.81, 119.10 (q, J= 273 Hz), 102.86, 61.64, 54.16, 32.36,

317

25.99, 13.74.

318

for C19H17ClF3N3O4S2: C, 44.93; H, 3.37; N, 8.27; Found: C, 45.07; H, 3.23; N, 8.13.

C NMR (150 MHz, CDCl3) δ 172.57, 165.19, 159.89, 152.26, 150.56, 141.24 (q, J= 33 Hz),

19

F NMR (376 MHz, CDCl3) δ: -66.91. ESI-MS: 530.1(M+Na)+. Anal. Calcd

319 320

Data for 9Cl-6. Yield: 26%. White solid, m.p. 120-121℃. 1H NMR (400 MHz, CDCl3) δ

321

7.97 (s, 1H), 7.76 (s, 1H), 6.44 (s, 1H), 4.20-4.15 (m, 5H), 3.62 (s, 3H), 1.71 (q, J =7.2 Hz,

322

2H), 0.97 (t, J = 7.2 Hz, 3H). 13C NMR (150 MHz, CDCl3) δ 167.71, 167.33, 159.97, 151.94,

323

150.62, 141.45 (q, J = 33 Hz), 137.47, 130.26, 127.90, 122.26, 121.98, 119.19 (q, J= 273 Hz),

324

103.00, 67.45, 34.98, 32.48, 21.66, 10.09.

325

516.1(M+Na)+. Anal. Calcd for C18H15ClF3N3O4S2: C, 43.77; H, 3.06; N, 8.51; Found: C,

326

43.65; H, 2.89; N, 8.78.

19


17 / 54



Page 18 of 54

327 328


329

1H), 6.41 (s, 1H), 5.19 – 4.92 (m, 1H), 4.12 (s, 2H), 3.59 (s, 3H), 1.27 (d, J = 6.0 Hz, 6H). 13C

330

NMR (150 MHz, CDCl3) δ 167.46, 167.18, 160.04, 152.04, 150.69, 141.52 (q, J= 33 Hz),

331

137.54, 130.31, 127.94, 122.22, 122.01, 119.25 (q, J= 273 Hz), 103.05, 69.73, 35.36, 32.49,

332

21.53.

333


19

F NMR (376 MHz, CDCl3) δ: -66.90. ESI-MS: 516.1(M+Na)+. Anal. Calcd for

334 335


336

8.09 (s, 1H), 7.89 (s, 1H), 6.42 (s, 1H), 4.01 (s, 3H), 3.59 (s, 3H).

337

CDCl3) δ 166.06, 161.02, 160.13, 150.85, 150.72, 141.63 (q, J= 33 Hz), 137.92, 130.61,

338

129.28, 123.63, 122.22, 119.23 (q, J= 273 Hz), 103.09, 55.62, 32.58.

339

CDCl3) δ: -66.91. ESI-MS: 452.2(M+H)+. Anal. Calcd for C15H9ClF3N3O4S2: C, 39.87; H,

340

2.01; N, 9.30; Found: C, 40.03; H, 2.11; N, 9.15.

13

C NMR (150 MHz,

19

F NMR (376 MHz,

341 342


343

8.08 (s, 1H), 7.89 (s, 1H), 6.42 (s, 1H), 4.47 (q, J = 7.2 Hz 2H), 3.60 (s, 3H), 1.41 (t, J = 7.2

344

Hz 3H). 13C NMR (150 MHz, CDCl3) δ 165.43, 161.41, 160.09, 150.88, 150.73, 141.61 (q, J=

345

33 Hz), 137.90, 130.61, 129.23, 123.62, 122.16, 119.27 (q, J= 273 Hz), 103.11, 65.68, 32.55, 18 / 54


Page 19 of 54


19


346

14.10.

347


348 349


350

7.95 (s, 1H), 7.75 (s, 1H), 6.41 (s, 1H), 4.18 (s, 2H), 3.79 (s, 3H), 3.59 (s, 3H). 13C NMR (150

351

MHz, CDCl3) δ 168.68, 168.49, 160.16, 151.74, 150.69, 140.91 (q, J= 33 Hz), 136.85, 131.27,

352

127.45, 122.62, 119.29 (q, J= 273 Hz), 117.60, 103.10, 52.67, 34.64, 32.39.

353

MHz, CDCl3) δ: -66.93. ESI-MS: 488.1(M+Na)+. Anal. Calcd for C16H11ClF3N3O4S2: C,

354

41.25; H, 2.38; N, 9.02; Found: C, 41.43; H, 2.22; N, 8.87.

19

F NMR (376

355 356

Data for 9Cl-11. Yield: 39%. Yellow oil, 1H NMR (600 MHz, CDCl3) δ 7.95 (s, 1H), 7.76 (s,

357

1H), 6.41 (s, 1H), 4.72 (q, J = 7.2 Hz, 1H), 3.77 (s, 3H), 3.59 (s, 3H), 1.71 (d, J = 7.2 Hz, 3H).

358

13

359

137.53, 130.31, 128.05, 122.34, 121.99, 119.20 (q, J= 273 Hz), 103.03, 52.83, 44.81, 32.50,

360

17.75.

361



19

F NMR (376 MHz, CDCl3) δ: -66.91. ESI-MS: 502.2(M+H)+. Anal. Calcd for

362 363


364

7.96 (s, 1H), 7.79 (s, 1H), 6.41 (s, 1H), 4.12 (d, J = 2.4 Hz, 2H), 3.59 (s, 3H), 2.30 (s, 1H). 13C 19 / 54



365

NMR (150 MHz, CDCl3) δ 167.07, 160.14, 152.25, 150.80, 141.65 (q, J= 33 Hz), 137.64,

366

130.44, 128.16, 122.53, 122.10, 119.34 (q, J= 273 Hz), 103.14, 77.86, 72.49, 32.61, 21.63.

367

19

368


F NMR (376 MHz, CDCl3) δ: -66.91. ESI-MS: 432.1(M+H)+. Anal. Calcd for

369 370

Data for 9Br-1. Yield: 56%. Yellow oil, 1H NMR (600 MHz, CDCl3) δ 8.11 (s, 1H), 7.74 (s,

371

1H), 6.41 (s, 1H), 4.24 (q, J = 7.2 Hz, 2H), 4.16 (s, 2H), 3.59 (s, 3H), 1.29 (t, J = 7.2 Hz, 3H).

372

13

373

137.95, 131.92, 125.13, 122.34, 119.32 (q, J= 33 Hz), 117.57, 103.16, 62.08, 35.07, 32.57,

374

14.04.

375

C17H13BrF3N3O4S2: C, 38.94; H, 2.50; N, 8.01; Found: C, 38.70; H, 2.71; N, 8.23.


19


376 377


378

1H), 6.41 (s, 1H), 4.18 (q, J = 7.2 Hz, 2H), 3.61-3.58 (m, 5H) 2.91 (t, J = 7.2 Hz, 2H)., 1.27 (t,

379

J = 7.2 Hz, 3H). 13C NMR (150 MHz, CDCl3) δ 171.40, 168.88, 160.15, 153.01, 150.79,

380

141.64 (q, J= 33 Hz), 137.84, 131.88, 125.06, 122.18, 119.36 (q, J= 273 Hz), 117.37, 103.22,

381

60.87, 34.07, 32.61, 28.24, 14.13.

382

560.1(M+Na)+. Anal. Calcd for C18H15BrF3N3O4S2: C, 40.16; H, 2.81; N, 7.81; Found: C,

383

40.32; H, 2.71; N, 7.65.

19


20 / 54


Page 20 of 54

Page 21 of 54


384 385


386

1H), 6.41 (s, 1H), 4.15 (q, J = 7.2 Hz, 2H), 3.59 (s, 3H), 3.42 (t, J = 7.2 Hz, 2H), 2.50 (t, J =

387

7.2 Hz, 2H), 2.18 – 2.15 (m, 2H), 1.26 (t, J = 7.2 Hz, 3H). 13C NMR (150 MHz, CDCl3) δ

388

172.46, 169.17, 160.06, 153.03, 150.68, 141.50 (q, J= 33 Hz), 137.67, 131.75, 124.91, 122.05,

389

119.26 (q, J= 273 Hz), 117.17, 106.72, 103.12, 60.41, 32.61, 32.54, 24.34, 14.06.

390

(376 MHz, CDCl3) δ: -66.96. ESI-MS: 574.2(M+Na)+. Anal. Calcd for C19H17BrF3N3O4S2: C,

391

41.31; H, 3.10; N, 7.61; Found: C, 41.47; H, 2.99; N, 7.53.

19

F NMR

392 393


394

1H), 6.41 (s, 1H), 4.69 (q, J = 7.2 Hz, 1H), 4.22 (q, J = 7.2 Hz, 2H), 3.59 (s, 3H), 1.71 (d, J

395

=7.2 Hz, 3H), 1.28 (t, J = 7.2 Hz, 3H). 13C NMR (150 MHz, CDCl3) δ 171.16, 167.23, 160.12,

396

152.91, 150.75, 141.64 (q, J= 33 Hz), 137.99, 131.92, 125.11, 122.42, 119.35 (q, J= 273 Hz),

397

117.63, 103.21, 61.92, 45.25, 32.61, 17.92, 14.03.

398

ESI-MS: 560.1(M+Na)+. Anal. Calcd for C18H15BrF3N3O4S2: C, 40.16; H, 2.81; N, 7.81;

399

Found: C, 40.34; H, 2.68; N, 7.72.

19

F NMR (376 MHz, CDCl3) δ: -66.92.

400 401


402

1H), 6.41 (s, 1H), 4.20 (q, J = 7.2 Hz, 2H), 3.59 (s, 3H), 1.75 (s, 6H), 1.21 (t, J = 7.2 Hz, 3H). 21 / 54



403

13

404

138.28, 131.83, 125.00, 122.73, 119.24 (q, J= 273 Hz), 117.94, 103.10, 61.80, 54.31, 32.48,

405

26.14, 13.88.

406

for C19H17BrF3N3O4S2: C, 41.31; H, 3.10; N, 7.61; Found: C, 41.52; H, 2.91; N, 7.49.


19

F NMR (376 MHz, CDCl3) δ: -66.86. ESI-MS: 574.1(M+Na)+. Anal. Calcd

407 408

Data for 9Br-6. Yield: 70%. White solid, m.p. 106-107℃. 1H NMR (400 MHz, CDCl3) δ

409

8.11 (s, 1H), 7.79 (s, 1H), 6.44 (s, 1H), 4.20-4.15 (m, 5H), 3.62 (s, 3H), 1.71 (q, J =7.2 Hz,

410

2H), 0.98 (t, J = 7.2 Hz, 3H). 13C NMR (150 MHz, CDCl3) δ 167.82, 167.56, 160.09, 152.73,

411

150.73, 141.59 (q, J= 33 Hz), 137.95, 131.93, 125.13, 122.35, 119.33 (q, J= 273 Hz), 117.56,

412

103.17, 67.59, 35.04, 32.57, 21.78, 10.21.

413

538.1(M+H)+. Anal. Calcd for C18H15BrF3N3O4S2: C, 40.16; H, 2.81; N, 7.81; Found: C,

414

40.36; H, 2.62; N, 7.61.

19


415 416


417

8.12 (s, 1H), 7.74 (s, 1H), 6.42 (s, 1H), 5.22 – 5.00 (m, 1H), 4.13 (s, 2H), 3.60 (s, 3H), 1.28 (d,

418

J = 6.2 Hz, 6H). 13C NMR (150 MHz, CDCl3) δ 167.68, 167.24, 160.11, 152.76, 150.74,

419

141.62 (q, J= 33 Hz), 137.97, 131.92, 125.13, 122.31, 119.34 (q, J= 273 Hz), 117.55, 103.18,

420

69.84, 35.45, 21.76, 21.50.

19

F NMR (376 MHz, CDCl3) δ: -66.91. ESI-MS: 560.0(M+Na)+.

22 / 54


Page 22 of 54

Page 23 of 54


421

Anal. Calcd for C18H15BrF3N3O4S2: C, 40.16; H, 2.81; N, 7.81; Found: C, 40.12; H, 2.64; N,

422

7.97.

423 424


425

8.24 (s, 1H), 7.88 (s, 1H), 6.41 (s, 1H), 4.07 (q, J = 6.6 Hz, 2H), 3.59 (s, 3H), 1.41 (t, J = 6.6

426

Hz, 3H). 13C NMR (150 MHz, CDCl3) δ 165.45, 161.64, 160.11, 151.57, 150.73, 141.65 (q,

427

J= 33 Hz), 138.26, 132.16, 125.29, 123.63, 119.33 (q, J= 273 Hz), 118.94, 103.20, 65.73,

428

32.60, 14.15. 19F NMR (376 MHz, CDCl3) δ: -66.89. ESI-MS: 510.0(M+H)+. Anal. Calcd for

429


430 431


432

8.10 (s, 1H), 7.75 (s, 1H), 6.41 (s, 1H), 4.17 (s, 2H), 3.78 (s, 3H), 3.59 (s, 2H). 13C NMR (150

433

MHz, CDCl3) δ 168.31, 167.42, 160.13, 152.71, 150.75, 141.63 (q, J= 33 Hz), 137.99, 131.93,

434

125.15, 122.41, 119.33 (q, J= 273 Hz), 117.60, 103.17, 53.11, 52.90, 34.72.

435

MHz, CDCl3) δ: -66.90. ESI-MS: 532.1(M+Na)+. Anal. Calcd for C16H11BrF3N3O4S2: C,

436

37.66; H, 2.17; N, 8.23; Found: C, 37.81; H, 2.13; N,8.39.

19

F NMR (376

437 438

Data for 9Br-11. Yield: 53%. White solid, m.p. 68-69℃. 1H NMR (400 MHz, CDCl3) δ 8.11

439

(s, 1H), 7.76 (s, 1H), 6.41 (s, 1H), 4.72 (q, J = 7.2 Hz, 1H), 3.77 (s, 3H), 3.59 (s, 3H), 1.71 (d, 23 / 54



13

440

J = 7.2 Hz, 3H).

C NMR (150 MHz, CDCl3) δ 171.64, 167.08, 160.10, 152.80, 150.74,

441

141.60 (q, J= 33 Hz), 137.92, 131.92, 125.10, 122.41, 119.31 (q, J= 273 Hz), 117.64, 103.17,

442

52.90, 44.87, 32.56, 17.83.

443

Anal. Calcd for C17H13BrF3N3O4S2: C, 38.94; H, 2.50; N, 8.01; Found: C, 38.98; H, 2.56; N,

444

8.06.

19

F NMR (376 MHz, CDCl3) δ: -66.91. ESI-MS: 548.1(M+Na)+.

445 446


447

8.12 (s, 1H), 7.79 (s, 1H), 6.41 (s, 1H), 4.12 (d, J = 2.4 Hz, 2H), 3.59 (s, 3H), 2.30 (s, 1H). 13C

448

NMR (150 MHz, CDCl3) δ 167.26, 160.10, 152.82, 150.74, 141.61 (q, J= 33 Hz), 137.91,

449

131.96, 125.14, 122.48, 119.32 (q, J= 273 Hz), 117.66, 103.17, 77.82, 72.50, 32.59, 21.61.

450

19

451


F NMR (376 MHz, CDCl3) δ: -66.90.

ESI-MS: 497.8(M+Na)+. Anal. Calcd for

452 453


454

8.10 (s, 1H), 7.76 (s, 1H), 6.41 (s, 1H), 6.20 – 5.83 (m, 1H), 5.38 (d, J = 17.2 Hz, 1H), 5.22

455

(d, J = 10.0 Hz, 1H), 4.00 (d, J = 7.2 Hz, 2H), 3.59 (s, 3H). 13C NMR (151 MHz, CDCl3) δ

456

168.81, 160.11, 153.08, 150.75, 141.57 (q, J= 33 Hz), 137.83, 131.83, 125.00, 122.22, 119.34

457

(q, J= 273 Hz), 119.39, 117.31, 103.18, 36.12.

19

F NMR (376 MHz, CDCl3) δ: -66.90. ESI-

24 / 54


Page 24 of 54

Page 25 of 54


458

MS: 500.1(M+Na)+. Anal. Calcd for C16H11BrF3N3O4S2: C, 40.18; H, 2.32; N, 8.79; Found: C,

459

40.31; H, 2.20; N, 8.65.

460 461

Evaluation of the kinetics of PPO Inhibition

462

The expression of the tobacco PPO enzyme (mtPPO) was performed according to the

463

established protocol32-34 and purified as described previously.35 The product of the enzymatic

464

reaction had a maximum excitation wavelength at 410 nm and a maximum emission

465

wavelength at 630 nm.35-38 Thus, we monitored the formation of protoporphyrin IX at room

466

temperature using a ﬂuorescence detector with the excitation and emission wavelengths set at

467

410 and 631 nm, respectively. The concentration of protoporphyrinogen IX was determined

468

according to the absorption of protoporphyrin IX and the concentration of protoporphyrin IX

469

was calculated from the calibration graph. In each assay, the stock dimethyl sulfoxide

470

(DMSO) solution (1% total volume) of inhibitor was added into the reaction system. The final

471

inhibitor concentration ranged from 0.005 µM to 250 µM. The enzymatic reaction rate was

472

measured in a reagent buffer of 100 mM potassium phosphate (pH 7.5), 5 mM DTT, 1 mM

473

EDTA, Tween 80 (0.03%, v/v), 200 mM imidazole, 5 µM FAD, and approximately 0–40 µg

474

of protein. The kinetic parameters were evaluated by Sigma Plot software 10.0 (SPSS,

475

Chicago, IL). The IC50 value was determined by measuring PPO activity over a range of

25 / 54



476

inhibitor concentrations at a fixed substrate concentration. IC50 values were calculated by

477

fitting v versus [I] data to a single binding site model (Equation 1)

478

y = min +

max − min 1 + 10log IC50 − x

(Equation 1)

479

where y is the percentage of maximal rate, with max and min being the y values at which the

480

curve levels off, x is the logarithm of inhibitor concentration, and IC50 is the concentration of

481

inhibitor that caused 50% of the total inhibition. Because the inhibitors belong to the class of

482

competitive inhibitors, the calculated Ki value (Table 1) can be obtained by applying the

483

following relationship among Ki, Km, and IC50 at any saturating substrate concentration (S)

484

(Equation 2).

Ki = 485

IC50 S / Km + 1

(Equation 2)

486 487

CoMFA analysis

488

All molecular modeling and 3D-QSAR studies were performed using SYBYL7.3 with

489

TRIPOS Force Field. The 3D structures of all compounds were made using default settings of

490

SYBYL, and the molecules were subjected to energy minimization at a gradient of 1.0

491

kcal/mol with delta energy change of 0.05 cal/mol. The CoMFA descriptors, steric, and

492

electrostatic field energies were calculated using the SYBYL default parameters: 2.0 Å rid

493

points spacing, an sp3 carbon probe atom with +1 charge and a minimum σ(column filtering)

494

of 2.0 kcal/mol, and the energy cutoff of 30.0 kcal/mol. 26 / 54


Page 26 of 54

Page 27 of 54


495 496

Greenhouse Herbicidal Activities

497

The herbicidal activities of compounds 9 against monocotyledon weeds such as Echinochloa

498

crusgalli (Ec), Digitaria sanguinalis (Ds), and Setaria faberi (Sf), Polypogon fugax (Pf),

499

Beckmannia syzigachne (Bs), Poa annua (Pa) and dicotyledon weeds such as Brassica juncea

500

(Bj), Amaranthus retroflexus (Ar), and Chenopodium album (Ca), Eclipta prostata (Ep),

501

Chenopodium serotinum(Cs), Stellaria media(Sm) were evaluated according to the previously

502

reported procedure.44-46 Sulfentrazone and saflufenacil were selected as the positive control.

503

All the test compounds were formulated as 100 g/L emulsified concentrates using DMF as the

504

solvent and Tween-80 as the emulsification reagent. The concentrated formulas were diluted

505

with water to obtain the required concentration and applied to pot-grown plants in a

506

greenhouse. The soil used was a clay soil, pH 6.5, 1.6% organic matter, 37.3% clay particles,

507

and CEC 12.1 mol/kg. The rate of application (g.ai/ha) was calculated by the total amount of

508

active ingredient in the formulation divided by the surface area of the pot. Plastic pots with a

509

diameter of 9.5 cm were filled with soil to a depth of 8 cm. Approximately 20 seeds of the

510

tested weeds were sown in the soil at the depth of 1~3 cm and grown at 15-30 ºC in a

511

greenhouse. The atmospheric relative humidity was about 50%. The diluted formulation

512

solutions were applied for post-emergence treatment. Dicotyledon weeds were treated at the

513

2-leaf stage and monocotyledon weeds were treated at the 1-leaf stage. The post-emergence 27 / 54



514

application rate was 150 g.ai/ha. Untreated seedlings were used as the control group and the

515

solvent (DMF + Tween-80) treated seedlings were used as the solvent control group.

516

Herbicidal activity was evaluated visually at 15 days post treatment. The results of herbicidal

517

activities are shown in Table 2 and 3, three replicates per treatment.

518

Crop Selectivity

519

The conventional rice, soybean, cotton, wheat, rape and maize were planted in plots (diameter

520

= 12 cm) containing test soil and grown in a greenhouse at 20~25 ºC. After the plants had

521

reached the 4-leaf stage, the spraying treatment was conducted at different dosages by diluting

522

the formulation of the test compound with water. The visual injury and growth state of the

523

individual plant were observed at regular intervals. The final evaluation for crop safety of the

524

test compound was conducted by visual observation in 30 days after treatment on the 0~100

525

scale.

526 527

Results and Discussion

528

Synthetic chemistry of the title compounds

529

As shown in Figure 3, the target compounds 9 were prepared by a multi-step synthetic route

530

using disubstituted aniline as the starting materials. The compounds 2 were synthesized from

531

the reaction of 2,4-disubstituted aniline with methyl chloroacetate, then underwent a

532

cyclization reaction with compound 3 to afford the key intermediate 4. Without further 28 / 54


Page 28 of 54

Page 29 of 54


533

purification, compound 4 reacted directly with methyl iodide to give the methylated product 5

534

with a yield of 63-67% in two steps. Subsequently, the nitration of intermediate 5 was

535

achieved by using the mixed solution of HNO3 in concentrated H2SO4 to give regioselectively

536

5-position nitrated compound 6. The nitro group in molecule 6 was then reduced by Fe

537

powder to afford intermediates 7, which underwent ring-closing reaction with potassium O-

538

ethyl dithiocarbonate in DMF solution at 90℃ to produce the key intermediates, 8, in yields

539

of 84-87%. Finally, intermediates 8a-c reacted with diverse alkylation reagent to give the

540

target compounds 9 with the yields of 24-70%. The structures of all intermediate and title

541

compounds were confirmed by 1H NMR,

542

MS spectral data.

13

C NMR,

19

F NMR, elemental analyses and ESI-

543 544

PPO Inhibition Activity

545

For the sake of clarity, the compounds 9 are defined as fluorine, chlorine and bromine

546

analogues, respectively, throughout the text. The inhibition potency (ki) against tobacco PPO

547

of the newly synthesized compounds is summarized in Table 1. Two commercial compounds,

548

sulfentrazone and saflufenacil, were selected as the positive control. From Table 1, we can

549

conclude that the fluorine analogues displayed much higher activities against tobacco PPO

550

than the chlorine and bromine analogues. If the R group is fixed, the fluorine analogues

551

always exhibited 10- to 100-fold higher activity than the corresponding chlorine and bromine 29 / 54



552

analogues. For example, the fluorine compound 9F-1 (ki = 0.0090 µM) showed about 50-fold

553

and 537-fold higher activity than the corresponding chlorine and bromine analogues 9Cl-1 (ki

554

= 0.45 µM) and 9Br-1 (ki = 4.84 µM), respectively. Additionally, most of the fluorine

555

compounds 9F-1-13 displayed higher PPO-inhibiting potency than the commercial control

556

sulfentrazone. In particular, compound 9F-5, identified as the most potent inhibitor with the ki

557

value against tobacco PPO of 0.0072 µM, showed over 4-fold higher PPO-inhibiting potency

558

than sulfentrazone (ki = 0.03 µM). Furthermore, the length of the linker between the

559

benzothiazole-2-thioyl moiety and the terminal ester has an apparent effect on the PPO-

560

inhibiting activity. For instance, for the fluorine series, when the spacer increased from one

561

carbon (9F-9) to two carbons (9F-1), the PPO-inhibiting activity was remarkably increased,

562

their Ki values were improved from 0.065 to 0.0090 µM. Interestingly, further prolongation of

563

the linker length to a three-carbon spacer (9F-2, Ki = 0.0073 µM) led to a further enhancement

564

of the PPO-inhibiting activity. However, an even longer spacer (4 carbon atoms) resulted in a

565

significant decrease of the PPO-inhibition capacity (9F-3, ki = 0.013 µM). Therefore, a three-

566

carbon spacer is the optimum linker to maintain high inhibition activity. It is worth noting that

567

the unsaturated alkyl substituents are also acceptable when comparing the Ki values of

568

compounds 9F-12 and 9F-13 with that of the ester analogues, although relatively lower

569

activity was observed. For the chlorine and bromine series compounds, the regularity and

570

tendency of the linker length with respect to the effect on the PPO-inhibiting activity is 30 / 54


Page 30 of 54

Page 31 of 54


571

similar to that within fluorine compounds. The three-carbon spacers are more favorable and

572

show higher activity than the other linkers.

573 574

CoMFA analysis

575

To understand the substituent effects on the PPO inhibition of the synthesized compounds, the

576

CoMFA method was applied to investigate the quantitative structure-activity relationship. As

577

listed in Table 1, a predicted CoMFA model was established with the conventional coefficient

578

r2 = 0.9847 and the cross-validated coefficient q2 = 0.8477, the predicted non-cross-validated

579

coefficient r2 (pred) = 0.9020. The observed and calculated activity values of all these

580

compounds were listed in Table 1 and the plot of the predicted versus the actual activity

581

values for all of the compounds were shown in Figure 4. The steric and electrostatic

582

contributions obtained from the CoMFA method were illustrated in Figure 5. The contour

583

map of the steric contributions is marked in green and yellow areas (Figure 5A). The green

584

region means a bulky group which is favorable for a higher PPO inhibition activity. By

585

contrast, the yellow region highlights positions where a bulky group would be unfavorable for

586

higher PPO inhibition activities (Figure 5A), Hence, the substituents at this position should

587

have an optimized steric effects. For example, compound 9F-1 (R= CH2COOC2H5, X = F, ki

588

= 0.0090 µM) is more active than 9Cl-1 (R = CH2COOC2H5, X = Cl, ki = 0.45 µM), and

589

compound 9Br-1 (R = CH2COOC2H5 , X = Br, ki = 4.84 µM) with a relatively bulky bromine 31 / 54



590

substituent shows much lower potency. The electrostatic contour maps are shown in Figure

591

5B. The blue color represented that the electro-positive group will be advantageous to the

592

bioactivity of the PPO inhibitors. On the contrary, the electro-negative groups which are

593

disadvantageous to the bioactivity are marked in red. From Figure 5B, we can infer that the

594

interaction of the electrostatic field is ambiguous, namely, the influence of the electrostatic

595

field is minimal to the activity of these compounds.

596 597

Green house herbicidal activities

598

The post-emergence herbicidal activities of compounds 9 were tested in a greenhouse against

599

monocotyledon weeds, such as E. crusgalli, D. sanguinalis, S. faberi, P. fugax Nees, B.

600

syzigachne and P. annua and dicotyledon weeds such as B. juncea, A. retroflexus, A.

601

theophrasti, E. prostrate, C. serotinum and S. media. The commercial herbicide sulfentrazone

602

and saflufenacil were selected as the control. As shown in Table 2, the fluorine series

603

compounds exhibited excellent broad spectrum herbicidal activities at the concentration of

604

150 g ai/ha. Very promisingly, all the fluorine derivatives showed over 90% control against

605

the dicotyledon weeds and, compounds 9F-1, 9F-4, 9F-6, 9F-7, 9F-10 and 9F-11 displayed

606

over 90% control against the tested monocotyledon weeds as well. Other fluorine compounds

607

such as 9F-2, 9F-3, 9F-5, 9F-8 and 9F-13 displayed varied activities ranging from 60-80%

608

control against monocotyledon weeds depending on the substituent. Therefore, those fluorine 32 / 54


Page 32 of 54

Page 33 of 54


609

compounds except for 9F-9, whose herbicidal potency confined to dicotyledon weeds, were

610

further tested at lower dosages. At the concentration of 75 g ai/ha, nine compounds termed

611

9F-1-8 and 9F-11 still showed over 90% control against dicotyledon weeds such as B. juncea,

612

A. retroflexus, A. theophrasti, while all compounds except for 9F-8, 9F-12 and 9F-13

613

displayed over 60% control efficiency against monocotyledon weeds such as E. crusgalli, D.

614

sanguinalis and S. faberi. Very impressively, even when the concentration was reduced to as

615

low as 37.5 g ai/ha, all the tested fluorine compounds retained over 90% control effect against

616

dicotyledon weeds. Among them, compounds 9F-1, 9F-4, 9F-10 and 9F-11 also exhibited

617

over 80% control potency against monocotyledon weeds including E. crusgalli, D.

618

sanguinalis and S. faberi. whereas, compounds 9F-6 and 9F-7 showed total control against

619

monocotyledon weeds D. sanguinalis and S. faberi at the concentration of 37.5 g ai/ha, but

620

did not show significant herbicidal activity against monocotyledon weed species E. crusgalli.

621

Compared with the fluorine series compounds, the chlorine derivatives and especially the

622

bromine derivatives showed much less activity and narrower herbicidal spectrum. Taking the

623

chlorine series as an example, compounds 9Cl-1, 9Cl-3, 9Cl-4, 9Cl-6 and 9Cl-7 displayed

624

total control against the three dicotyledon weeds species, such as B. juncea, A. retroflexus and

625

A. theophrasti, but they showed limited potency toward monocotyledon weeds. The rest of

626

chlorine type compounds displayed herbicidal activity either confined to one or two specific

627

dicotyledon weeds or totally inactive to both monocotyledon and dicotyledon weeds. The 33 / 54



628

bromine series compounds are the worst in terms of their herbicidal activities. Nearly all of

629

them are inactive toward the tested weed species including monocotyledon and dicotyledon

630

weeds. Only several compounds showed certain control potency against specific dicotyledon

631

weeds. The results indicated that bromine substituent is unfavorable for herbicidal activity,

632

which is consistent with our previous results29.

633

Compound 9F-6, which showed excellent broad spectrum herbicidal activity, was selected for

634

further testing of crop selectivity. The results are listed in Table 3 and the commercial

635

herbicide sulfentrazone and saflufenacil were selected as the positive control. Among the

636

tested crops in Table 3, maize exhibited relative tolerance to compound 9F-6 by post-

637

emergence application at the dosage of 150 g ai/ha, whereas cotton, rape, soybean, rice and

638

wheat were susceptible at the same dosage. However, all the six test crops are susceptible to

639

saflufenacil even at the dosage of 75 g ai/ha. These results indicate that compound 9F-6 has a

640

better crop selectivity than saflufenacil, and can be developed as a potential herbicide used for

641

weed control in maize fields.

642 643

In summary, a series of novel 3-(2´-halo-5´-substituted-benzothiazol-1´-yl)-1-methyl-6-

644

(trifluoromethyl)pyrimidine-2,4-diones 9 were designed and synthesized as potential PPO

645

inhibitors. The result of in vitro and greenhouse tests indicate that some newly synthesized

646

compounds has good PPO inhibition activity and herbicidal activities at the concentration of 34 / 54


Page 34 of 54

Page 35 of 54


647

37.5 g ai/ha. Most interestingly, compound 9F-6 showed a good inhibition against mtPPO

648

with ki value of 0.012 µM and broad spectrum herbicidal activity even at a concentration as

649

low as 37.5 g ai/ha. Furthermore, maize exhibited relative tolerance to compound 9F-6 by

650

post-emergence application at the dosage of 150 g ai/ha. These results indicated that

651

compound 9F-6 may have the potential as herbicide candidate for the weed control in maize

652

fields.

653

Supporting Information

654

Supporting information for the data of the intermediates 2a-c, 3, 5a-c, 6a-c and 7a-c is

655

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

656 657 658

ACKNOWLEDGMENT

659

The research was supported in part by the National Natural science Foundation of China (No.

660

21332004 and 21402059).

661 662 663 664 665 35 / 54



666

References

667

1. Poulson, R.; Polglase, W. J. The enzymic conversion of protoporphyrinogen IX to

668

protoporphyrin IX. Protoporphyrinogen oxidase activity in mitochondrial extracts of

669

Saccharomyces cerevisiae. J. Biol. Chem. 1975, 250, 1269-1274.

670 671 672 673 674 675 676 677

2. Poulson, R. The enzymic conversion of protoporphyrinogen IX to protoporphyrin IX in mammalian mitochondria. J. Biol. Chem. 1976, 251, 3730-3733. 3. Ajioka, R. S.; Phillips, J. D.; Kushner, J. P. Biosynthesis of heme in mammals. Biochimica Et Biophysica Acta-Molecular Cell Research. 2006, 1763, 723-726. 4. Heinemann, I. U.; Jahn, M.; Jahn, D. The biochemistry of heme biosynthesis. Arch Biochem Biophy. 2008, 474, 238-251. 5. Hao, G. F.; Zuo, Y.; Yang, S. G.; Yang, G. F. Protoporphyrinogen oxidase inhibitor: an ideal target for herbicide discovery. Chimia 2011, 65, 961-969.

678

6. Jiang, L. L.; Zuo, Y.; Wang, Z. F.; Tan, Y.; Wu, Q. Y.; Xi, Z.; Yang, G. F. Design and

679

syntheses of novel N-(Benzothiazol-5-yl)-4,5,6,7-tetrahydro-1H-isoindole-1,3(2H)-dione

680

and N-(Benzothiazol-5-yl)-isoindoline-1,3-dione as potent protoporphyrinogen oxidase

681

inhibitors. J. Agric. Food. Chem. 2011, 59, 6172-6179.

682

7. Hess, F. D. Light-dependent herbicides: An overview. Weed Sci. 2000, 48, 160-170.

683

8. Theodoridis, G. Protophyrinogen-IX-oxidase inhibitors. In Modern Crop Protection

684

Compounds; WILEY-VCH Verlag GmbH Co. KGaA: Weinheim, Germany, 2007; pp 36 / 54


Page 36 of 54

Page 37 of 54

685


153-182.

686

9. He, L. L.; Wu, Y. Y.; Zhang, H. Y.; Liu, M. Y.; Shi, D. Q. Synthesis and herbicidal

687

activities of novel uracil derivatives containing pyrimidinyl moiety. Chem. Res. Chin.

688

Univ. 2014, 30, 400-404.

689 690 691 692 693 694 695 696

10. Kunz, W.; Siegrist, U.; Baumeister, Process for the preparation of 3-arylurcils. WO 9532952, 1995. 11. Theodoridis, G. Herbicidal 2-[(4-heterocyclic-phenoxymethyl)phenoxy]-alkanoates. US 5344812, 1994. 12. Theodoridis, G.; Bahr, J. T.; Hotzman, F. W.; Sehgel, S.; Suarez, D. P. New generation of protox-inhibiting herbicides. Crop Prot. 2000, 19, 533-535. 13. Klaus, G.; Ricarda, N.; Nicole, C. The herbicide Saflufenacil (Kixor) is a new inhibitor of Protoporphyrinogen IX oxidase activity. Weed Science. 2010, 58, 1-9.

697

14. Theodoridis, G.; Bahr, J. T.; Crawford, S.; Dugan, B.; Hotzman, F. W.; Maravetz, L. L.;

698

Sehgel, S.; Suarez, D. P. Synthesis and Chemistry of Agrochemicals VI, ACS Symposium

699

Series,Vol.800 Eds. American Chemical Society, Washington, D C, 2002, pp 96–107.

700

15. Theodoridis, G.; Liebl, R.; Zagar, C. Protophyrinogen-IX-Oxidase Inhibitors. In Modern

701

Crop Protection Compounds, WILEY-VCH Verlag GmbH Co. KGaA: Weinheim,

702

Germany, 2012; pp 178-181.

703

16. Nagano, E.; Haga, T.; Sato, R.; Morita, K. Tetrahydrophthalimides and their herbicidal 37 / 54



704

use. US 4640707, 1987.

705

17. Ganzer, M.; Franke, W.; Dorfmeister, G.; Johann, G.; Arndt, F.; Rees, R. Heterocyclic

706

substituted azoles and azines, process for their preparation and their use as an agent with

707

herbicide activity. EP 311135, 1989.

708

18. Kashiyama, E.; Hutchinson, I.; Chua, M. S.; Stinson, S. F.; Phillips, L. R.; Kaur, G.;

709

Sausville, E. A.; Bradshaw, T. D.; Westwell, A. D.; Stevens, M. F. Synthesis, metabolic

710

formation, and biological properties of the C- and N-oxidation products of antitumor 2-

711

(4-aminophenyl)benzothiazoles. J. Med. Chem. 1999, 42, 4172-4184.

712

19. Chua, M. S.; Shi, D. F.; Wrigley, S.; Bradshaw, T. D.; Hutchinson, I.; Shaw, P. N; Barrett,

713

D. A.; Stanley, L. A.; Stevens, M. F. Synthesis of 2-(4-acylaminophenyl)benzothiazoles

714

and investigations into the role of acetylation in the antitumor activities of the parent

715

amines. J. Med. Chem. 1999, 42, 381-392.

716

20. Hutchinson, I.; Chua, M. S.; Browne, H. L.; Trapani, V.; Bradshaw, T. D.; Westwell, A.

717

D.; Stevens, M. F. Synthesis and in vitro biological properties of fluorinated 2-(4-

718

aminophenyl)benzothiazoles. J. Med. Chem. 2001, 44, 1446-1455.

719

21. Hutchinson, I.; Jennings, S. A.; Vishnuvajjala, B. R.; Westwell, A. D.; Stevens, M. F.

720

Synthesis and pharmaceutical properties of antitumor 2-(4-aminophenyl)benzothiazole

721

amino acid prodrugs. J. Med. Chem. 2002, 45, 744-747.

722

22. Mortimer, C. G.; Wells, G.; Crochard, J. P.; Stone, E. L.; Bradshaw, T. D.; Stevens, M. F. 38 / 54


Page 38 of 54

Page 39 of 54


723

G.; Westwell, A. D. 2-(3,4-Dimethoxyphenyl)-5-fluorobenzothiazole (GW 610, NSC

724

721648), a simple fluorinated 2-arylbenzothiazole, shows potent and selective inhibitory

725

activity against lung, colon, and breast cancer cell lines. J. Med. Chem. 2006, 49, 179-185.

726

23. Weng, J. Q.; Tan, C. X.; Liu, X. H. Synthesis and fungicidal activity of hydrazones

727

containing 4-methylbenzo[d] thiazole moiety. J. Pestic. Sci. 2012, 37, 164-168.

728

24. Zhao, L. J.; Yang, X. N.; Li, X. Y.; Mu, W.; Liu, F. Antifungal, insecticidal and

729

herbicidal properties of volatile components from paenibacillus polymyxa strain BMP-11.

730

Arg. Sci. China. 2011, 10, 728-736.

731

25. Zuo, Y.; Yang, S. G.; Luo, Y. P.; Tan, Y.; Hao, G. F.; Wu, Q. Y.; Xi, Z.; Yang, G. F.

732

Design

and

synthesis

of

1-(benzothiazol-5-yl)-1H-1,2,4-triazol-5-ones

733

protoporphyrinogen oxidase inhibitors. Bioorg. Med. Chem. 2013, 21, 3245-3255.

as

734

26. Zuo, Y.; Yang, S. G.; Jiang, L. L.; Hao, G. F.; Wang, Z. F.; Wu, Q. Y.; Xi, Z.; Yang, G. F.

735

Quantitative structure–activity relationships of 1,3,4-thiadiazol-2(3H)-ones and 1,3,4-

736

oxadiazol-2(3H)-ones as human protoporphyrinogen oxidase inhibitors. Bioorg. Med.

737

Chem. 2012, 20, 296-304.

738

27. Hao, G. F.; Tan, Y.; Yu, N. X.; Yang, G. F. Structure-activity relationships of diphenyl-

739

ether as protoporphyrinogen oxidase inhibitors: insights from computational simulations.

740

J. Comput. Aided Mol. Des. 2011, 25, 213-222.

741

28. Zhang, L.; Tan, Y.; Wang, N. X.; Wu, Q. Y.; Xi, Z.; Yang, G. F. Design, syntheses and 39 / 54



742

3D-QSAR studies of novel N-phenyl pyrrolidin-2-ones and N-phenyl-1H-pyrrol-2-ones

743

as protoporphyrinogen oxidase inhibitors. Bioorg. Med. Chem. 2010, 18, 7948-7956.

744

29. Jiang, L. L.; Tan, Y.; Zhu, X. L.; Wang, Z. F.; Zuo, Y.; Chen, Q.; Xi, Z.; Yang, G. F.

745

Design, synthesis and 3D-QSAR analysis of novel 1,3,4-oxadiazol-2(3H)-ones as

746

protoporphyrinogen oxidase inhibitors. J. Agric. Food Chem. 2010, 58, 2643-2651.

747

30. Wu, Q. Y.; Jiang, L. L.; Yang, S. G.; Zuo, Y.; Wang, Z. F.; Xi, Z. ; Yang, G. F.

748

Hexahydrophthalimide–benzothiazole hybrids as a new class of protoporphyrinogen

749

oxidase inhibitors: synthesis, structure–activity relationship, and DFT calculations New J.

750

Chem., 2014, 38, 4510-4518.

751

31. Wu, Q. Y.; Jiang, L. L.; Zuo, Y.; Wang, Z. F.; Xi, Z.; Yang, G. F. Synthesis, in vitro

752

protoporphyrinogen oxidase inhibition, and herbicidal activity of N-(benzothiazol-5-

753

yl)hexahydro-1H-isoindole-1,3-diones and N-(benzothiazol-5-yl)hexahydro-1H-isoindol-

754

1-ones. Chem. Biol. Drug Des. 2014, 84,431-442.

755

32. Manelia, M. H.; Corrigalla, A. V.; Klumpb, H. H.; Davidsa, L. M.; Kirscha, R. E.;

756

Meissnera, P. N. Kinetic and physical characterisation of recombinant wild-type and

757

mutant human protoporphyrinogen oxidases. Biochem. Biophys. Acta 2003, 1650, 10-21.

758 759 760

33. Dailey, T. A.; Dailey, T. A. Human protoporphyrinogen oxidase: Expression, purification, and characterization of the cloned enzyme. Protein Sci. 1996, 5, 98-105. 34. Corrigall, A. V.; Siziba, K. B.; Maneli, M. H.; Shephard, E. G.; Ziman, M.; Dailey, T. A.; 40 / 54


Page 40 of 54

Page 41 of 54


761

Dailey, H. A.; Kirsch, R. E.; Meissner, P. N. Purification of and kinetic studies on a

762

cloned protoporphyrinogen oxidase from the aerobic bacterium Bacillus subtilis. Arch.

763

Biochem. Biophys. 1998, 358, 251-256.

764

35. Tan, Y.; Sun, L.; Xi, Z.; Yang, G. F.; Jiang, D. Q.; Yan, X. P.; Yang, X.; Li, H. Y. A

765

capillary electrophoresis assay for protoporphyrinogen oxidase. Anal. Biochem. 2008,

766

383, 200-204.

767

36. Meissner, P. N.; Day, R. S.; Moore, M. R.; Disler, P. B.; Harley, E. Protoporphyrinogen

768

oxidase and porphobilinogen deaminase in variegate porphyria. Eur. J. Clin. Investig.

769

1986, 16, 257-261.

770

37. Hepherd, M.; Dailey, H. A. A continuous ﬂuorimetric assay for protoporphyrinogen

771

oxidase by monitoring porphyrin accumulation. Anal. Biochem. 2005, 344, 115-121.

772

38. Jeong, E.; Houn, T. A point mutation of valine-311 to methionine in Bacillus subtilis

773

protoporphyrinogen oxidase does not greatly increase resistance to the diphenyl ether

774

herbicide oxyfluorfen. Bioorg. Chem. 2003, 31, 389-397.

775 776

39. Sanner, M. F. Python: a programming language for software integration and development. J. Mol. Graphics Mod. 1999, 17, 57-61.

777

40. Morris, G. M.; Goodsell, D. S.; Halliday, R. S.; Huey, R.; Hart, W. E.; Belew, R. K.;

778

Olson, A. J. Automated docking using a Lamarckian genetic algorithm and an empirical

779

binding free energy function. J. Comput. Chem. 1998, 19, 1639-1662. 41 / 54



780

41. Hao, G. F.; Wang, F.; Li, H.; Zhu, X. L.; Yang, W. C.; Huang, L. S.; Wu, J. W.; Berry, E.

781

A.; Yang, G. F. Computational discovery of picomolar Qo site inhibitors of cytochrome

782

bc1 complex. J. Am. Chem. Soc. 2012, 134, 11168-11176.

783

42. Zhao, P. L.; Wang, L.; Zhu, X. L.; Huang, X. Q.; Zhan, C. G.; Wu, J. W.; Yang, G. F.

784

Subnanomolar inhibitor of cytochrome bc1 complex designed via optimizing interaction

785

with conformationally flexible residues. J. Am. Chem. Soc. 2010, 132, 185-194.

786

43. Case, D. A.; Darden, T. A.; Cheatham; Iii, T. E.; Simmerling, C. L.; Wang, J.; Duke, R.

787

E.; Luo, R.; Merz, K. M.; Pearlman, D. A.; Crowley, M. AMBER 9, University of

788

California, San Francisco, 2006.

789

44. Chen, C. N.; Chen, Q.; Liu, Y. C.; Niu, C. W.; Xi, Z.; Yang, G. F. Syntheses and

790

herbicidal activity of new triazolopyrimidine-2-sulfonamides as acetohydroxyacid

791

synthase inhibitors. Bioorg. Med. Chem. 2010, 18, 4897-4904.

792

45. Chen, C. N.; Lv, L. L.; Ji, F. Q.; Chen, Q.; Xu, H.; Niu, C. W.; Xi, Z.; Yang, G. F. Design

793

and synthesis of N-2,6-difluorophenyl-5-methoxyl-1,2,4-triazolo[1,5-a]pyrimidine-2-

794

sulfonamide as acetohydroxyacid synthase inhibitor. Bioorg. Med. Chem. 2009, 17, 3011-

795

3017.

796

46. Li, Y. X.; Luo, Y. P.; Xi, Z.; Niu, C. W.; He, Y. Z.; Yang, G. F. Design and syntheses of

797

novel phthalazin-1(2H)-one derivatives as acetohydroxyacid synthase inhibitors. J. Agric.

798

Food Chem. 2006, 54, 9135-9139. 42 / 54


Page 42 of 54

Page 43 of 54


799 800 801 802 803 804 805 806 807 808 809 810 811 812 813 814 815 816 817 43 / 54



818

FIGURE CAPTIONS

819

Figure 1 Structures of the representative pyrimidinediones-type PPO-inhibiting

820

herbicides

821

Figure 2. Design of the title compounds

822

Figure 3 Synthetic route of the designed compounds. Reagents and conditions: (a)

823

ClCOOEt, Pyridine, CH2Cl2, ice bath ; (b) ammonium acetate, EtOH, reflux; (c)

824

NaH/DMF, ice bath to 120-130℃; (d) CH3I, K2CO3/DMF, rt ; (e) HNO3/H2SO4, ice bath;

825

(f) iron powder, NH4Cl, EtOH/H2O, reflux; (g) EtOC(S)SK, DMF, 90-95℃, con. HCl; (h)

826

RX, acetone, K2CO3, rt.

827

Figure 4. CoMFA predicted versus experimental pKi values.

828

Figure 5 The contour map of the: A. steric; and B. electronic contributions.

829 830 831 832

44 / 54


Page 44 of 54

Page 45 of 54


Table 1. Inhibitory Activity of the Synthesized Compounds Experimental and Calculated pKi Values No. X R Ki(µmol/L) 9F-1 F CH2COOC2H5 0.0090 9F-2 F CH2CH2COOC2H5 0.0073 9F-3 F CH2CH2CH2COOC2H5 0.0130 9F-4 F CH(CH3)COOC2H5 0.0140 9F-5 F C(CH3)2COOC2H5 0.0072 9F-6a F CH2COOCH2CH2CH3 0.0120 9F-7 F CH2COOCH(CH3)2 0.011 9F-8 F COOCH3 0.036 9F-9 F COOC2H5 0.065 a 9F-10 F CH2COOCH3 0.032 9F-11a F CH(CH3)COOCH3 0.016 9F-12 F CH2CCH 0.019 9F-13 F CH2CHCH2 0.014 9Cl-1 Cl CH2COOC2H5 0.45 9Cl-2 Cl CH2CH2COOC2H5 0.12 9Cl-3 Cl CH2CH2CH2COOC2H5 0.21 9Cl-4 Cl CH(CH3)COOC2H5 1.39 9Cl-5a Cl C(CH3)2COOC2H5 5.32 9Cl-6 Cl CH2COOCH2CH2CH3 1.09 9Cl-7 Cl CH2COOCH(CH3)2 0.46 9Cl-8 Cl COOCH3 4.76 9Cl-9 Cl COOC2H5 7.04 a 9Cl-10 Cl CH2COOCH3 4.85 9Cl-11 Cl CH(CH3)COOCH3 8.64 9Cl-12 Cl CH2CCH 4.88 9Br-1a Br CH2COOC2H5 4.84 a 9Br-2 Br CH2CH2COOC2H5 2.17 9Br-3 Br CH2CH2CH2COOC2H5 2.94 9Br-4 Br CH(CH3)COOC2H5 2.25 9Br-5a Br C(CH3)2COOC2H5 1.69 9Br-6 Br CH2COOCH2CH2CH3 66.32 9Br-7 Br CH2COOCH(CH3)2 11.29 9Br-8 Br COOC2H5 81.12 9Br-9 Br CH2COOCH3 98.54 9Br-10 Br CH(CH3)COOCH3 17.23 9Br-11a Br CH2CCH 130.05 9Br-12 Br CH2CHCH2 140.75 sulfentrazone 0.03 saflufenacil 0.01 a These compounds consist of the test set in CoMFA.

45 / 54


Against mtPPO and the pKi,exp 8.0458 8.1367 7.8861 7.8539 8.1427 7.9208 7.9586 7.4437 7.1871 7.4946 7.7959 7.7212 7.8539 6.3468 6.9208 6.6778 5.8570 5.2741 5.9626 6.3382 5.3224 5.1524 5.3143 5.0635 5.3116 5.3152 5.6635 5.5317 5.6479 5.7721 4.1784 4.9473 4.0909 4.0064 4.7637 3.8859 3.8516

pKi,cal 8.0248 8.0475 7.8170 7.8970 8.2138 8.1385 7.9731 7.4586 7.1789 7.5651 7.7565 7.6651 7.7879 6.0475 7.0761 6.6403 6.2717 5.6171 6.0021 6.2399 5.1588 5.2363 5.1067 5.3800 5.3258 4.7685 6.1941 5.3909 5.4238 5.0850 4.1362 5.1253 4.0755 3.8455 4.4896 4.3949 4.2710


Page 46 of 54

Table 2. Herbicidal Activities of the Synthesized Compounds (post-emergency) No. 9F-1

9F-2

9F-3

9F-4

9F-5

9F-6

9F-7

9F-8

9F-9 9F-10

9F-11

9F-12

9F-13

9Cl-1 9Cl-2 9Cl-3 9Cl-4 9Cl-5 9Cl-6 9Cl-7 9Cl-8 9Cl-9 9Cl-10

Dosage (g ai/ha) 150 75 37.5 150 75 37.5 150 75 37.5 150 75 37.5 150 75 37.5 150 75 37.5 150 75 37.5 150 75 37.5 150 150 75 37.5 150 75 37.5 150 75 37.5 150 75 37.5 150 150 150 150 150 150 150 150 150 150

Ec/Pf ++++ +++ +++ ++++ ++ + ++++ ++++ ++ ++++ ++++ +++ ++++ ++ ++ ++++ ++ ++ ++++ ++++ ++ ++ + ++++ +++ +++ ++++ +++ +++ +++ ++ ++ ++ ++ ++ ++ ++ ++ -

Ds/Bs ++++ +++ +++ ++++ ++ + +++ +++ ++ ++++ ++++ +++ +++ +++ ++ ++++ ++++ ++++ ++++ ++++ +++ +++ + ++++ +++ +++ ++++ ++++ ++++ +++ +++ ++ ++ ++++ ++ ++ -

Sf/Pa ++++ +++ +++ ++ ++ ++ +++ ++ + ++++ ++++ +++ ++++ ++ + ++++ ++++ ++++ ++++ ++++ +++ ++ ++ ++++ +++ +++ ++++ ++++ ++++ ++ ++ ++ ++ ++ ++ -

46 / 54


Bj/Ep ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++ ++ ++++ ++++ ++++ ++++ ++++ +++ ++++ ++++ ++ ++++ ++++ +++ ++++ +++ ++++ ++++ ++++ ++++ +++

Ar/Cs ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ +++ +++ ++++ +++ ++++ ++++ ++++ ++++ ++++

At/Sm ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ +++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ +++ ++++ ++++ + ++++ ++++ -

Page 47 of 54


9Cl-11 9Cl-12 9Br-1 9Br-2 9Br-3 9Br-4 9Br-5 9Br-6 9Br-7 9Br-9 9Br-10 9Br-11 9Br-12 9Br-13 sulfentrazone

saflufenacil

150 150 150 150 150 150 150 150 150 150 150 150 150 150 150 75 37.5 150 75 37.5

+ + / ++ ++++ ++++ +++ ++++ ++++ ++++

++ / ++ + ++++ ++++ ++++

/ ++++ +++ ++ ++++ ++++ ++++

++ + ++ ++ ++ ++ ++ + / + ++++ ++++ ++++ ++++ ++++ ++++

++++ ++ +++ ++ ++ ++++ ++ + +++ / +++ ++++ ++ ++++ ++++ ++++ ++++ ++++ ++++

++++ +++ ++++ ++++ ++++ ++++ ++ ++ ++++ ++++ ++++ ++++ ++++ ++++

“/” no test a: Ec for Echinochloa crusgalli, Ds for Digitaria sanguinalis, Sf for Setaria faberi, Pf for Polypogon fugax, Bs for Beckmannia syzigachne, Pa for Poa annua, Bj for Brassica juncea, Ar for Amaranthus retroflexus, At for Abutilon theophrasti, Ep for Eclipta prostrate, Cs for Chenopodium serotinum, Sm for Stellaria media. b: Rating system for the growth inhibition percentage: ++++, ≧90%; +++, 80~89%; ++, 60~79%; +, 50-59%; -,

Synthesis, Herbicidal Activity, and QSAR of Novel ... - ACS Publications

Recommend Documents