Diversity-oriented synthesis and antifungal activities of novel pimprinine derivative bearing a 1,3,4-oxadiazole-5-thioether moiety

Molecular Diversity

https://doi.org/10.1007/s11030-020-10048-8

ORIGINAL ARTICLE

Diversity‑oriented synthesis and antifungal activities of novel

pimprinine derivative bearing a 1,3,4‑oxadiazole‑5‑thioether moiety

1

2

3

4

5

Zi‑Long Song · Yun Zhu · Jing‑Rui Liu · Shu‑Ke Guo · Yu‑Cheng Gu · Xinya Han · Hong‑Qiang Dong · Qi Sun ·

1

Wei‑Hua Zhang · Ming‑Zhi Zhang

Received: 24 October 2019 / Accepted: 5 February 2020

Springer Nature Switzerland AG 2020

©

Abstract

Based on the strategy of diversity-oriented synthesis and the structures of natural product pimprinine and streptochlorin, two

series of novel pimprinine derivatives containing 1,3,4-oxadiazole-5-thioether moieties were eꢀciently synthesized under

the optimized reaction conditions. Biological assays conducted at Syngenta showed the designed derivatives displayed an

altered pattern of biological activity, of which 5h was identiꢁed as the most promising compound with strong activity against

Pythium dissimile and also a broad antifungal spectrum in primary screening. Further structural optimization of pimprinine

and streptochlorin derivatives is well under way, aiming to discover synthetic analogues with improved antifungal activity.

Graphic abstract

Two series of novel pimprinine derivatives containing 1,3,4-oxadiazole-5-thioether moieties wereeꢀciently synthesized

through diversity-oriented synthesis strategy under the optimizedconditions. Biological assays showed the designed deriva-

tives exhibited potential activity.

Keywords Pimprinine · 1,3,4-oxadiazole-5-thioether · Diversity-oriented synthesis · Antifungal activity · SAR

Electronic supplementary material The online version of this

article (https://doi.org/10.1007/s11030-020-10048-8) contains

supplementary material, which is available to authorized users.

3

*

Xinya Han

School of Chemistry and Chemical Engineering, Anhui

University of Technology, Ma’anshan 243002, China

xinyahan@ahut.edu.cn

4

5

Wei-Hua Zhang

College of Plant Science, Tarim University, Alaer 843300,

Xinjiang, China

njzhangwh@126.com

Ming-Zhi Zhang

Key Laboratory for Green Chemical Process of Ministry

of Education, School of Chemistry and Environmental

Engineering, Wuhan Institute of Technology, Wuhan 430205,

China

mzzhang@njau.edu.cn

1

2

Jiangsu Key Laboratory of Pesticide Science,

College of Sciences, Nanjing Agricultural University,

Nanjing 210095, China

Syngenta Jealott’s Hill International Research Centre,

Bracknell, Berkshire RG42 6EY, UK

Vol.:(0

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123456789)

3

Molecular Diversity

Introduction

by 1,3,4-oxadiazole [19], as this scaꢂold is an impor-

tant pharmacophore in agricultural and medicinal ꢁelds

[20–22], and replacing the indole ring by other active het-

erocycles [23]. The SAR study demonstrated that indole

ring is an essential moiety for the antifungal activity of

pimprinine derivatives, and spectrum of antifungal activ-

ity could be expanded by introducing a methylene bridge

between the two heterocyclic rings.

Pimprinine, an indole alkaloid originally isolated from the

ꢁ

ltrates of cultures of Streptomyces pimprina in 1963 [1,

2

], belongs to the class of naturally occurring 5-(3-indolyl)

oxazoles [3, 4]. A range of related natural products has

now been isolated from plants and marine microorgan-

isms, such as pimprinethine, pimprinaphine, WS-30581

A, WS-30581 B, labradorin 1, labradorin 2, streptochlo-

rin, martefragin A, 5-hydroxy-2′-isobutyl-streptochlorin

Therefore, in a continuation of our studies aimed at the

discovery of novel pimprinine analogues though step-by-

step optimization, we now describe this study which has

focused on replacement of the oxazole ring of pimprinine

with 1,3,4-oxadiazole-5-thioether (ODATE). 1,3,4-oxadia-

zole-5-thioether is an important building block for the new

drugs discovery due to its numerous biological activities

[24, 25], primarily as antiviral, antimalarial, antimicrobial,

antidiabetic, anticancer, antioxidant, anticonvulsant and anti-

inꢃammatory. We brieꢃy summarize the latest publication

on 1,3,4-oxadiazole-5-thioether derivatives, as shown in

Fig. 2, and emphasis is placed on the diverse array of bio-

logical activities.

(

HIS) and diazonamide A [5–8] (Fig. 1). Among which,

streptochlorin, as a 4-chloro-analogue of pimprinine, also

an indole natural product of this family [9]. Pimprinine

and streptochlorin are considered as promising drug leads

to various biological activities [5, 10–12], such as inhibit

TNF-α-induced NF-κB activation and mast cell degranula-

tion [13, 14], and show selective cytotoxic eꢂects against

cancer cell lines and potent anti-allergic eꢂects in vivo

[

15–17]. It is worth noting that pimprinine and strep-

tochlorin showed activity against the following fungi on

Syngenta’s screens against Pythium dissimile, Botrytis

cinerea, Septoria tritici, Pyricularia oryzae, Fusarium

culmorum and Rhizoctonia solani [2]. However, that

the tests in which activity was seen are highly sensitive,

so pimprinine has a good spectrum of activity, but low

potency.

ODATE 1 was reported to display a broad-spectrum anti-

viral activity, and it is shown to inhibit the replication of

inꢃuenza viruses A and B, HCV and HIV at noncytotoxic

concentrations [26]. ODATE 2 presented antimalarial activ-

ity against cultured P. falciparum [27]. Based on active natu-

ral products, an uncharged catechol-derivative compound

ODATE 3 showed an IC50 of 3.09 μM against protein tyros-

ine phosphatases 1B (PTP1B) [28]. The scorpion-shaped

compound ODATE 4 showed high activity against GSK-

3α/β, which is believed to be central to the pathogenesis

of Alzheimer’s disease [29]. As a potential focal adhe-

sion kinase (FAK) inhibitor, ODATE 5 exhibited potent

in vitro anticancer activities [30]. It is reported ODATE 6

In order to discover new lead candidates from natural

products, for further structural optimization and investi-

gation of targeting enzyme, various structural modiꢁca-

tions to pimprinine were conducted in our previous study,

including diꢂerent substituents at the indole ring and the

nitrogen of the indole ring [2, 5, 18], replacing the Cl on

the oxazole ring by Br or H, replacing the oxazole ring

Fig. 1 Chemical structures of 5-(3-indolyl)oxazole natural products

1

3

Molecular Diversity

Fig. 2 Structures of active 1,3,4-oxadiazole-5-thioether derivatives

demonstrated impressive anticancer activity in vitro and in

vivo as a sphingosine kinase (SK) inhibitor [31]. The pre-

liminary bioassay results indicated that the inhibition rate

of compound ODATE 7 against B. cinerea and R. solani at

pimprinine derivatives bearing 1,3,4-oxadiazole-5-thioether

moieties are designed and synthesized through DOS strat-

egy, aiming at the discovery of natural product derivatives

with improved antifungal activity, as shown in Fig. 3.

5

0 mg/L reached above 85% [32]. The insecticidal activities

of ODATE 8 displayed 71.43% activity against P. xylostella

at 0.4 μM and 33.33% against S. exigua at 1.0 μM [33]. Hit

molecule ODATE 9 was identiꢁed as the inhibitor of glu-

taminyl cyclase (hQC), which has emerged as a new poten-

tial target for the treatment of Alzheimer’s disease [34].

ODATE 10 exhibited unsurpassed antimicrobial activities

against all the experimental bacteria and fungi, and it pro-

vided a facile strategy to develop a new type of agricultural

fungicides [35]. ODATE 11 exhibited remarkable anti-TMV

activity with the EC values of 33.66 mg/mL, which was

Materials and methods

Chemistry

All chemicals were purchased from commercial sources

(e.g., Alfa Aesar Co.) and were all analytically pure. The

solvents and liquid reagents were dried by standard meth-

ods in advance and distilled before use. Melting points were

taken on a Büchi M-560 melting point apparatus (uncor-

rected), and FTIR spectra were recorded on a Thermo Nico-

5

0

comparable to that of Ningnanmycin (36.85 mg/mL) and

1

superior to that of Ribavirin (88.52 mg/mL) [36].

let 380 FTIR spectrometer. H NMR spectra were recorded

Based on the structure of natural product pimprinine and

the previous SAR conclusions [2, 18, 19, 23], indole ring is

an essential moiety for the antifungal activity of pimprin-

ine derivatives, and spectrum of antifungal activity could

be expanded by introducing a methylene bridge between

the two heterocyclic rings. Therefore, two series of novel

on a VARIAN Mercury-Plus 600 spectrometer in CDCl

3

1

3

or DMSO-d with TMS as the internal reference. C NMR

6

spectra were recorded in CDCl or DMSO-d on a Varian

3

6

Mercury 600 MHz or a Bruker Avance 400 MHz spectrom-

eter, and chemical shifts (δ) are given in ppm relative to the

center line of a triplet at 77.0 ppm of CDCl or 39.5 ppm of

3

Fig. 3 Design strategy of target

compounds

1

3

Molecular Diversity

DMSO-d . Elemental analyses were performed on a Vario

temperature, and the results of preliminary screening are

summarized in Table 1. Under the optimized conditions, the

ratio of thiol, substituted benzyl bromide and triethylamine

is 1:1.1:2, the DMF solution was heated under 40 °C and was

monitored by TLC, cooled down after the reaction ꢁnished

and then poured into icy water, and the solid obtained was

ꢁltered, washed with water and small quantity of petroleum

ether, dried under high vacuum to give target compounds 5.

6

EL III elemental analysis instrument. Mass spectra were

determined using a ThermoFisher Mass platform DSQII by

electrospray ionization (ESI-MS), and the signals were given

in m/z. Reaction yields were not optimized.

General synthetic procedures for compounds 2–4

Compounds 2, 3 and 4 were synthesized from indole-

3

-carboxylic acid in three steps including esteriꢁcation,

2‑(ethylthio)‑5‑(1H‑indol‑3‑yl)‑1,3,4‑oxadiazole (5a) Yield,

1

hydrazidation and cyclization, and the synthetic methods

were described in previous publications [29, 33, 37–39], as

shown in Scheme 1.

70%. mp, 142–144 °C. H NMR (600 MHz, DMSO-d ):

6

δ 1.45 (t, J=7.2 Hz, 3H),3.32 (m, 2H), 7.29 (m, 2H), 7.58

(d, J=7.8 Hz, 1H), 8.12 (d, J=7.8 Hz, 1H), 8.21 (s, 1H),

1

3

1

2.08 (s, 1H). CNMR(150 MHz, DMSO-d ): δ14.9, 26.7,

6

methyl 1H‑indole‑3‑carboxylate (2) Yield, 89%.mp, 149–

99.2, 112.9, 120.7, 121.7, 123.3, 127.6, 128.7, 136.4, 160.7,

1

+

1

50 °C. H NMR (400 MHz, DMSO-d ) δ3.82 (s, 3H), 7.26

163.0. EI-MS: m/z (%) 245 (M , 65), 184 (6), 217 (4), 160

6

(

m, 2H), 7.60 (m, 1H), 8.02 (dt, J=6.8, 2.9 Hz, 1H), 8.10

(18), 144 (100), 116 (10), 89 (11), 44 (7). Anal.Calcd for

C H N OS: C, 58.76; H, 4.52; N, 17.13; S, 13.07; Found:

s, 1H), 11.97 (s, 1H).

1

2

11

3

C, 58.66; H, 4.523; N, 17.46; S, 13.13.

1

H‑indole‑3‑carbohydrazide (3) Yield, 95%.mp, 234–236 °C.

1

H NMR (600 MHz, DMSO-d ): δ4.34 (s, 2H), 7.11 (m, 2H),

2‑(butylthio)‑5‑(1H‑indol‑3‑yl)‑1,3,4‑oxadiazole (5b) Yield,

6

1

7

.43 (t,J=7.2 Hz,1H), 7.98 (m, 1H), 8.17 (s, 1H), 9.18 (s,

70%. mp, 142–144 °C. H NMR (600 MHz, DMSO-d ):

6

+

1

H), 11.55(s, 1H). EI-MS: m/z (%) 175 (M , 22), 144 (100),

δ 0.94 (t, J = 7.2 Hz, 3H),1.51 (m, 2H), 1.69 (m, 2H),

16 (17), 89 (11).

3.12 (m, 2H), 7.28 (m, 2H), 7.56 (d, J=7.2 Hz, 1H), 8.14

(

d, J = 7.8 Hz, 1H), 8.25 (d, J = 2.4 Hz, 1H), 11.89 (s,

1

3

5

2

7

1

‑(1H‑indol‑3‑yl)‑1,3,4‑oxadiazole‑2‑thiol (4) Yield, 97%.mp,

1H). CNMR(150 MHz, DMSO-d ):12.4, 22.1, 32.0, 35.9,

6

1

46–248 °C. H NMR (600 MHz, DMSO-d ): δ7.22 (m, 1H),

98.6,111.1,119.8, 121.4, 121.7, 127.4,128.5, 136.6, 160.4,

6

.26 (m, 1H), 7.52 (d, J=7.8 Hz, 1H), 7.91 (d, J=7.8 Hz,

164.5. Anal.Calcd for C H N OS: C, 61.51; H, 5.53; N,

1

4

15

3

1

3

H), 8.19 (s, 1H), 12.09 (s, 1H), 14.49 (s, 1H). C NMR

15.37; S, 11.73; Found: C, 61.58; H, 5.26; N, 15.30; S,

(

150 MHz, DMSO-d ): δ 98.4, 119.9, 120.0, 123.1, 123.5,

11.57.

6

+

1

28.9, 129.0, 136.5, 159.1, 176.0. EI-MS: m/z (%) 217 (M ,

00), 157 (28), 129 (32), 78 (18).

2‑(allylthio)‑5‑(1H‑indol‑3‑yl)‑1,3,4‑oxadiazole (5c) Yield,

1

7

9%. mp, 126–128 °C. H NMR (600 MHz, CDCl ): δ

3

Preparation

3.92 (d, J=7.2 Hz, 2H), 5.21 (d, J=9.6 Hz, 1H), 5.38 (d,

of 2‑(1H‑indol‑3‑yl)‑5‑(substituted‑thio)‑1,3,4‑oxadiazole 5

J = 16.8 Hz, 1H), 6.02 (m, 1H),7.30 (d, J = 3.9 Hz, 2H),

7

.50 (d, J=3.6 Hz, 1H), 7.88 (s, 1H), 8.24 (d, J=7.8 Hz,

1

3

In our initial study, the reaction of 5-(1H-indol-3-yl)-1,3,4-

oxadiazole-2-thiol (4) with 3-Fluorobenzyl bromide in

the presence of base was chosen as a model to optimize

the reaction conditions, including base, ratio, solvent and

1H), 9.55 (s, 1H). CNMR(150 MHz, DMSO-d ): δ 35.4,

6

101.1, 111.9, 120.9, 121.8, 122.0, 123.5, 124.2, 126.6,

+

126.7, 136.2, 161.3, 163.6. EI-MS: m/z (%) 257 (M , 42),

144 (100), 116 (15), 89 (13). Anal.Calcd for C H11N OS:

1

3

Scheme 1 Synthetic route

of 2-(1H-indol-3-yl)-

5

-thioether-1,3,4-oxadiazoles

1

3

Molecular Diversity

Table 1 Reaction condition optimization for compound 5p

Ratio^a

Solvent

Temperature

Time (h)

b

Yield^c

Entry

Base

1

2

3

4

5

6

K CO

1:1.1:5

1:1.1:2

1:1.1:4

1:1.1:2

Acetone

DMF

Reꢃux

130 °C

40 °C

1

6

1

2

4

1

21%

29%

78%

–^d

82%

99%

2

3

Cs CO

2

3

Et N

3

K CO

2

3

K CO

2

Et N

40 °C

3

a

The ratio of thiol: benzyl bromide: base

Time to ꢁnish the reaction monitored by TLC

Isolated yields

b

c

d

Complex reaction system

C, 60.68; H, 4.31; N, 16.33; S, 12.46; Found: C, 60.61; H,

1H), 3.46 (m, 1H), 3.56 (m, 1H), 7.33 (m, 2H), 7.48 (d,

J=7.8 Hz, 1H), 7.88 (s, 1H), 8.26 (d, J=6.6 Hz, 1H), 8.76

4

.26; N, 16.10; S, 12.42.

1

3

(

s, 1H). CNMR(150 MHz, DMSO-d ): δ 34.4, 46.8, 50.1,

6

2

‑((3,3‑dichloroallyl)thio)‑5‑(1H‑indol‑3‑yl)‑1,3,4‑oxadiazole

94.8, 112.4, 120.1, 121.2, 122.9, 123.9, 128.4, 136.4, 160.1,

1

+

(

5d) Yield, 65%. mp, 146–148 °C. H NMR (600 MHz,

162.2. EI-MS: m/z (%) 273 (M , 100), 241 (34), 227 (28),

CDCl ): δ4.01 (d, J = 11.4 Hz, 2H),6.33 (m, 1H), 7.26

201 (18), 115 (13), 89 (13), 73 (23), 57 (14), 44 (47). Anal.

Calcd for C H N O S: C, 57.13; H, 4.06; N, 15.37; S,

3

(

s, 1H), 7.33 (t, J = 6.0 Hz, 1H), 7.48 (t, J = 6.6 Hz, 1H),

1

3

11

3

2

7

1

.89 (d,J = 3.6 Hz, 1H), 8.26 (d, J = 7.2 Hz, 1H), 8.94 (s,

11.73; Found: C, 57.33; H, 4.08; N, 15.17; S, 11.97.

1

3

H). CNMR(150 MHz, DMSO-d ): δ 31.0, 99.1, 113.0,

6

19.6, 120.7, 121.8, 122.8, 123.9, 125.6, 126.7, 129.0,

methyl 2‑((5‑(1H‑indol‑3‑yl)‑1,3,4‑oxadiazol‑2‑yl)thio)acetate

+

1

36.4, 159.4, 163.6. EI-MS: m/z (%) 325 (M , 31), 290

(5g) Yield, 75%. mp, 166–168 °C. H NMR (600 MHz,

(

21), 216 (8), 144 (100), 116 (13), 89 (10). Anal.Calcd for

DMSO-d ): δ 3.73 (s, 3H),4.32 (s, 2H), 7.27 (m, 2H),7.56

6

C H Cl N OS: C, 47.87; H, 2.78; N, 12.88; S, 9.83; Found:

(d, J=7.8 Hz, 1H), 8.06 (d, J=7.2 Hz, 1H), 8.18 (s, 1H),

13

9

2

3

1

3

C, 47.94; H, 2.82; N, 12.76; S, 10.09.

12.08 (s, 1H). CNMR(150 MHz, DMSO-d ): δ 33.8 (m,

6

1

C), 52.6 (m, 1C), 99.0, 120.6, 121.8, 123.4, 123.9, 127.8,

+

2

‑((cyclopropylmethyl)thio)‑5‑(1H‑indol‑3‑yl)‑1,3,4‑oxa-

128.9, 136.5, 159.9, 163.2, 168.4. EI-MS: m/z (%) 289 (M ,

46), 184 (8), 160 (12), 144 (100), 116 (9), 89 (7), 43 (22).

Anal.Calcd for C H N O S: C, 53.97; H, 3.83; N, 14.52;

1

diazole (5e) Yield, 89%. mp, 143–145 °C. H NMR

(

600 MHz, DMSO-d ): δ 0.37 (d, J=1.8 Hz, 2H), 0.67 (d,

6

13 11

3

J=6.6 Hz, 2H), 1.28 (m, 1H),3.24 (d, J=7.8 Hz, 2H), 7.29

S, 11.08; Found: C, 53.95; H, 4.08; N, 14.57; S, 11.13.

(

t, J = 3.0 Hz, 2H), 7.50 (t, J = 3.0 Hz, 1H), 7.88 (s, 1H),

1

3

8

.24 (t, J = 3.0 Hz, 1H), 9.75 (s, 1H). CNMR(150 MHz,

= 61.1 Hz, 1C),

isopropyl 2‑((5‑(1H‑indol‑3‑yl)‑1,3,4‑oxadiazol‑2‑yl)thio)

1

DMSO-d ): δ 5.6 (m, 1C), 11.0 (t, J

acetate (5h) Yield, 93%. mp, 147–149 °C. H NMR

6

C–C

3

1

8.0, 101.0, 111.9, 120.4, 121.9, 123.4, 124.2, 126.7, 136.2,

(600 MHz, DMSO-d ): δ1.20 (d, J = 6.0 Hz, 6H), 2.24

6

+

62.2, 163.4. EI-MS: m/z (%) 271 (M , 10), 224 (32), 217

(s, 2H),4.97 (m,1H), 7.27 (m, 1H), 7.55 (d, J = 7.8 Hz,

(

19), 144 (100), 116 (15), 89 (11), 55(20). Anal.Calcd for

1H), 8.05 (d, J = 7.8 Hz, 1H), 8.17 (s, 1H), 12.05 (s,

1

3

C H N OS: C, 61.97; H, 4.83; N, 15.49; S, 11.82; Found:

1H). CNMR(150 MHz,DMSO-d ): δ 21.4, 34.1, 69.2,

1

4

13

3

6

C, 61.81; H, 4.95; N, 15.32; S, 11.66.

99.0, 112.4, 120.1, 121.2, 122.9, 123.8, 128.2, 136.4, 159.8,

+

1

63.1, 167.2. EI-MS: m/z (%) 317 (M , 49), 184 (11), 156

2

‑(1H‑indol‑3‑yl)‑5‑((oxiran‑2‑ylmethyl)thio)‑1,3,4‑oxa‑

(14), 144 (100), 116 (8), 89 (6), 44 (15). Anal.Calcd for

C H N O S: C, 56.77; H, 4.76; N, 13.24; S, 10.10; Found:

1

diazole (5f) Yield, 40%. mp, 125–127 °C. H NMR

15

3

(

600 MHz, CDCl ): δ 2.80 (m, 1H), 2.93 (m, 1H), 3.40(m,

C, 56.76; H, 4.89; N, 13.03; S, 10.28.

3

1

3

Molecular Diversity

+

tert‑butyl 2‑((5‑(1H‑indol‑3‑yl)‑1,3,4‑oxadiazol‑2‑yl)thio)ace-

EI-MS: m/z (%) 307 (M , 48), 144 (100), 116 (9), 91 (74),

1

tate (5i) Yield, 92%. mp, 149–151 °C. H NMR (600 MHz,

89 (11), 65 (9). Anal.Calcd for C H N OS: C, 66.43; H,

1

7

13

3

DMSO-d ): δ1.40 (s, 9H), 4.17 (s, 2H), 7.26 (m, 2H), 7.55

4.26; N, 13.67; S, 10.43; Found: C, 66.68; H, 4.11; N, 13.49;

6

(

d, J = 7.8 Hz, 1H), 8.06 (d, J = 7.8 Hz, 1H),8.17 (s, 1H),

S, 10.42.

1

3

1

8

1

2.05 (s, 1H). CNMR(150 MHz, DMSO-d ): δ 27.5, 34.8,

6

1.9, 99.0, 112.4, 120.1, 121.2, 122.9, 123.8, 128.2, 136.4,

2‑(1H‑indol‑3‑yl)‑5‑((2‑methylbenzyl)thio)‑1,3,4‑oxadiazole

+

1

59.9, 163.1, 166.7. m/z (%) 331 (M , 42), 275 (35), 160 (7),

(5n) Yield, 78%. mp, 146–148 °C. H NMR (600 MHz,

44 (100), 116 (9), 89 (6), 57 (16), 41 (23). Anal.Calcd for

DMSO-d ): δ2.42 (s, 3H), 4.60 (s, 2H), 7.15 (s, 1H),7.22

6

C H N O S: C, 57.99; H, 5.17; N, 12.68; S, 9.68; Found:

(s, 2H), 7.27 (m, 2H), 7.42 (d, J = 5.4 Hz, 1H), 7.57 (d,

1

6

17

3

C, 57.83; H, 5.24; N, 12.58; S, 9.71.

J=6.6 Hz, 1H), 8.08 (d, J=6.0 Hz, 1H), 8.20 (s, 1H), 12.09

1

3

(

s, 1H). CNMR(150 MHz, DMSO-d ): δ 18.8, 34.7, 99.1,

6

ethyl 4‑((5‑(1H‑indol‑3‑yl)‑1,3,4‑oxadiazol‑2‑yl)thio)butanoate

112.5, 120.2, 121.3, 122.9, 123.9, 126.1, 128.1, 128.4,

1

(

5j) Yield, 83%. mp, 109–111 °C. H NMR (600 MHz,

130.0, 130.5, 134.0, 136.4, 136.8, 160.0, 163.2. EI-MS: m/z

+

DMSO-d ): δ1.31 (t, J=7.2 Hz, 3H), 2.10 (m, 2H), 2.32 (m,

(%) 321 (M , 42), 288 (27), 144 (91), 116 (8), 105 (100),

6

2

7

1

2

1

H), 3.11 (m, 2H), 4.14 (q, J = 7.2 Hz, 2H), 7.30 (m,2H),

89 (9), 79 (11). Anal.Calcd for C H N OS: C, 67.27; H,

1

8

15

3

.55 (d, J=7.2 Hz, 1H), 8.11 (d, J=7.2 Hz, 1H), 8.19 (s,

4.70; N, 13.07; S, 9.98; Found: C, 67.39; H, 4.82; N, 12.96;

1

3

H), 12.08 (s, 1H). CNMR(150 MHz,DMSO-d ): δ 13.9,

S, 10.10.

6

4.7, 32.7, 35.8, 61.5, 111.1, 119.5, 120.0, 121.7, 122.9,

23.9, 128.4, 136.6, 157.5, 164.5, 172.1. Anal.Calcd for

2‑((2‑fluorobenzyl)thio)‑5‑(1H‑indol‑3‑yl)‑1,3,4‑oxadiazole

1

C H N O S: C, 57.99; H, 5.17; N, 12.68; S, 9.68; Found:

(5o) Yield, 81%. mp, 155–157 °C. H NMR (600 MHz,

1

6

17

3

C, 57.82; H, 5.19; N, 12.73; S, 9.77.

DMSO-d ): δ 4.60 (s, 2H), 7.20 (d, J = 7.2 Hz, 1H),7.26

6

(

m, 3H), 7.37 (d, J = 4.8 Hz, 1H), 7.57 (d, J = 7.8 Hz,

ethyl 2‑((5‑(1H‑indol‑3‑yl)‑1,3,4‑oxadiazol‑2‑yl)thio)‑2‑meth-

2H), 8.07 (d, J = 6.6 Hz, 1H), 8.19 (s, 1H), 12.09 (s,

1

13

ylpropanoate (5k) Yield, 42%. mp, 107–109 °C. H NMR

1H). CNMR(150 MHz, DMSO-d ): δ30.0, 99.1, 112.5,

6

(

600 MHz, DMSO-d ): δ1.10 (t, J = 7.2 Hz, 3H), 1.68

115.5 (d, J = 21.9 Hz, 1C), 120.2, 121.3, 122.9, 123.7,

6

C–F

s, 6H), 4.10 (m, 2H),7.28 (m,2H), 7.57 (d, J = 7.8 Hz,

123.9 (d, J = 9.3 Hz, 1C), 124.6, 128.4, 130.1, 131.4,

C-F

1

6

1

H), 8.08 (d, J = 7.2 Hz, 1H), 8.21 (s, 1H), 12.12 (s,

136.4, 159.6(d, J =15.0 Hz, 1C), 161.2, 163.4. EI-MS:

C–F

1

3

+

H). CNMR(150 MHz,DMSO-d ): δ 13.6, 25.7, 53.1,

m/z (%) 325 (M , 51), 144 (100), 116 (10), 109 (49), 89 (10),

6

1.5, 99.0, 112.5, 120.0, 121.4, 122.9, 123.9, 128.7, 136.4,

44 (32). Anal.Calcd for C H FN O : C, 62.76; H, 3.72; N,

1

7

12

3

S

+

57.5, 164.5, 171.6. EI-MS: m/z (%) 331 (M , 42), 217

12.91; S, 9.86; Found: C, 62.71; H, 3.93; N, 12.80; S, 9.91.

(

65), 157 (19), 144 (100), 116 (10), 89 (7). Anal.Calcd for

C H N O S: C, 57.99; H, 5.17; N, 12.68; S, 9.68; Found:

2‑((3‑fluorobenzyl)thio)‑5‑(1H‑indol‑3‑yl)‑1,3,4‑oxadiazole

1

6

17

3

1

C, 57.93; H, 5.22; N, 12.59; S, 9.48.

(5p) Yield, 82%. mp, 164–166 °C. H NMR (600 MHz,

DMSO-d ): δ4.60 (s, 2H), 7.19 (m, 1H),7.27 (m, 3H), 7.37

6

(

E)‑ethyl 4‑((5‑(1H‑indol‑3‑yl)‑1,3,4‑oxadiazol‑2‑yl)thio)

(m, 1H), 7.57 (d, J=7.8 Hz, 2H), 8.06 (d, J=7.8 Hz, 1H),

1

13

but‑2‑enoate (5l) Yield, 86%. mp, 101–103 °C. H NMR

8.19(s, 1H), 12.09(s, 1H). CNMR(150 MHz, DMSO-d ):

6

(

600 MHz, DMSO-d ): δ1.19 (t, J=7.2 Hz, 3H), 4.12 (m,

δ30.0, 99.1, 112.5, 114.5, 115.4-115.9 (d, J = 14.6 Hz,

6

C–F

4

7

1

3

1

3

H), 6.16 (d, J=15.6 Hz, 1H), 7.00 (m, 1H),7.27 (m, 2H),

1C), 120.2, 121.3, 122.9, 123.8-123.9(d, J = 15.4 Hz,

C–F

.56 (d, J = 7.8 Hz, 1H), 8.07 (d, J = 7.2 Hz, 1H),8.18 (s,

1C), 124.6, 128.4, 130.2, 131.4, 136.4, 159.6-159.7 (d,

1

3

+

H), 12.07 (s, 1H). CNMR(150 MHz,DMSO-d ): δ 14.0,

J

=10.0 Hz, 1C), 160.0, 163.0. EI-MS: m/z (%) 325 (M ,

6

C–F

2.6, 60.1, 99.0, 112.4, 120.1, 121.2, 122.9, 123.9, 128.4,

57), 144 (100), 116 (15), 109 (47), 89 (10), 44 (20). Anal.

+

36.4, 142.3, 159.6, 163.3, 165.1. EI-MS: m/z (%) 329 (M ,

Calcd for C H FN OS: C, 62.76; H, 3.72; N, 12.91; S,

1

7

12

3

7), 256 (18), 184 (12), 144 (100), 116 (15), 89 (7). Anal.

9.86; Found: C, 62.60; H, 3.59; N, 13.00; S, 9.90.

Calcd for C H N O S: C, 58.34; H, 4.59; N, 12.76; S, 9.74;

16

15

3

Found: C, 58.30; H, 4.46; N, 12.50; S, 9.54.

2‑(1H‑indol‑3‑yl)‑5‑((3‑methylbenzyl)thio)‑1,3,4‑oxadiazole

1

(

5q) Yield, 19%. mp, 136–138 °C. H NMR (600 MHz,

2

‑(benzylthio)‑5‑(1H‑indol‑3‑yl)‑1,3,4‑oxadiazole (5m) Yield,

DMSO-d ): δ2.28 (s, 2H), 4.54 (s, 2H), 7.1 (d, J=6.0 Hz,

6

1

5

5%. mp, 167–169 °C. H NMR (600 MHz, DMSO-d ):

1H),7.23 (m, 2H), 7.29 (m, 3H), 7.56 (d, J = 7.8 Hz, 1H),

6

δ4.58 (s, 2H), 7.27 (m, 3H),7.35 (m, 2H), 7.50 (m, 2H),

8.06 (d, J=7.8 Hz, 1H), 8.19 (d, J=1.8 Hz, 1H), 12.09 (s,

1

3

7

1

9

1

.56 (m, 1H), 8.06 (d, J=7.8 Hz, 1H), 8.19 (d, J=1.8 Hz,

1H). CNMR(150 MHz, DMSO-d ): δ 20.9, 36.1, 99.1,

6

1

3

H), 12.08 (s, 1H). CNMR(150 MHz, DMSO-d ): δ 36.1,

112.5, 120.2, 121.2, 122.9, 123.9, 126.1, 128.3, 128.4,

6

9.1, 113.0, 120.7, 121.8, 123.4, 123.9, 127.7, 128.0, 128.2,

128.5, 129.6, 136.4, 136.5, 137.7, 160.2, 163.1. EI-MS: m/z

+

28.5, 128.9, 129.1, 129.5, 136.4, 136.7, 160.2, 163.2.

(%) 321 (M , 57), 142 (100), 116 (10), 105 (93), 89 (5),

1

3

Molecular Diversity

4

1

4 (11). Anal.Calcd for C H N OS: C, 67.27; H, 4.70; N,

(s, 1H), 7.63 (s, 1H), 8.04 (s, 1H), 8.18(s, 1H), 12.09 (s,

1

8

15

3

1

3

3.07; S, 9.98; Found: C, 67.21; H, 4.92; N, 12.93; S, 9.91.

1H). CNMR(150 MHz, DMSO-d ): δ 29.5, 94.9, 99.0,

6

1

04.2 (d, J = 20.7 Hz, 1C), 111.7, 112.5, 120.4, 121.4,

C-F

2

‑(1H‑indol‑3‑yl)‑5‑((3‑nitrobenzyl)thio)‑1,3,4‑oxadiazole

123.0, 123.9, 128.2, 128.6, 132.6, 136.4, 159.6, 163.5.

1

+

(

5r) Yield, 9%. mp, 163–165 °C. H NMR (600 MHz,

EI-MS: m/z (%) 343 (M , 36), 144 (100), 127 (83), 116 (10),

CDCl ): δ4.59 (s, 2H), 7.25 (m, 2H),7.49 (d, J=7.8 Hz, 2H),

89 (7), 44 (7). Anal.Calcd for C H F N OS: C, 59.47; H,

3

17 11

2

3

7

.54 (s, 1H), 7.84 (s, 1H), 7.89 (s, 1H), 8.38 (s, 1H), 11.35

3.23; N, 12.24; S, 9.34; Found: C, 59.59; H, 3.33; N, 12.04;

1

3

(

s, 1H). CNMR(150 MHz, CDCl ): δ 35.0, 99.3, 111.1,

3

S, 9.35.

1

3

12.1, 120.5, 121.3, 122.6, 123.3, 123.6, 127.2, 128.6,

29.6, 136.0, 138.1, 147.6, 159.4, 163.3. EI-MS: m/z (%)

2‑(1H‑indol‑3‑yl)‑5‑((4‑(trifluoromethox y)benz yl)

thio)‑1,3,4‑oxadiazole (5w) )Yield, 80%. mp, 151–153 °C.

+

52 (M , 40), 144 (100), 136 (16), 116 (9), 89 (9), 44 (19).

1

Anal.Calcd for C H N O S: C, 57.95; H, 3.43; N, 15.90;

H NMR (600 MHz, DMSO-d ): δ4.62 (s, 2H), 7.26 (m,

1

7

12

4

3

6

S, 9.10; Found: C, 57.88; H, 3.30; N, 15.64; S, 9.32.

2H),7.28 (d, J=7.2 Hz,2H), 7.36 (d, J=6.6 Hz, 1H), 7.65

(

d, J=7.8 Hz, 2H), 8.05 (d, J=7.2 Hz, 1H), 8.18 (s, 1H),

1

3

2

‑(1H‑indol‑3‑yl)‑5‑((4‑methylbenzyl)thio)‑1,3,4‑oxadiazole

12.09 (s, 1H). CNMR(150 MHz, DMSO-d ): δ 35.1, 94.9,

6

1

(

5s) Yield, 80%. mp, 178–180 °C. H NMR (600 MHz,

99.1, 112.6, 119.2, 120.0, 120.3, 120.9, 121.1 (m, 1C),

DMSO-d ): δ2.53 (s, 3H), 4.54 (s, 2H), 7.15 (d, J=4.8 Hz,

122.8, 123.9, 128.3, 128.7, 131.0 (d, J = 9.2 Hz, 1C),

6

C–F

2

H),7.28 (m, 2H), 7.38 (d, J = 5.4 Hz, 2H), 7.58 (d,

132.1, 136.6 (d, J = 12.8 Hz, 1C), 147.7, 160.0, 163.3.

C–F

+

J=6.6 Hz, 1H), 8.09 (d, J=6.0 Hz, 1H), 8.21 (s, 1H), 12.10

EI-MS: m/z (%) 391 (M , 43), 175 (100), 144 (99), 116

1

3

(

s, 1H). CNMR(150 MHz, DMSO-d ): δ 20.7, 35.9, 99.2,

(10), 89 (7), 44 (14). Anal.Calcd for C H F N O S: C,

6

18 12

3

2

1

4

12.5, 120.2, 121.2, 122.9, 123.9, 128.3, 128.9, 129.1,

55.24; H, 3.09; N, 10.74; S, 8.19; Found: C, 55.05; H, 3.07;

+

33.6, 136.5, 137.0, 160.2, 163.1. EI-MS: m/z (%) 321 (M ,

N, 10.65; S, 8.10.

2), 180 (11), 144 (64), 116 (9), 105 (100), 44 (26). Anal.

Calcd for C H N OS: C, 67.27; H, 4.70; N, 13.07; S, 9.98;

methyl 4‑(((5‑(1H‑indol‑3‑yl)‑1,3,4‑oxadiazol‑2‑yl)thio)

18

15

3

1

Found: C, 67.37; H, 4.69; N, 12.99; S, 9.65.

methyl)benzoate (5x) Yield, 79%. mp, 180–182 °C. H

NMR (600 MHz, DMSO-d ): δ 3.44 (s, 3H), 4.65 (s, 2H),

6

2

‑((4‑fluorobenzyl)thio)‑5‑(1H‑indol‑3‑yl)‑1,3,4‑oxadiazole

7.25 (d, J = 7.2 Hz, 1H),7.29 (d, J = 7.2 Hz, 1H), 7.57 (d,

J=7.8 Hz, 1H), 7.66 (d, J=7.8 Hz, 2H), 7.96 (d, J=7.2 Hz,

2H), 8.03 (d, J = 7.2 Hz, 1H), 8.19 (s, 1H), 12.09 (s,

1

(

5t) Yield,78%. mp, 194–196 °C. H NMR (600 MHz,

DMSO-d ): δ4.55 (s, 2H), 7.26 (m, 2H),7.46 (d,

6

1

3

J=7.8 Hz,2H), 7.55 (d, J=7.8 Hz, 3H), 8.02(d, J=7.8 Hz,

1H). CNMR(150 MHz, DMSO-d ): δ35.6, 99.1, 112.5,

6

13

1

H), 8.18 (s, 1H), 12.09 (s, 1H). CNMR(150 MHz, DMSO-

120.1, 121.2, 122.9, 123.9, 128.4, 128.9, 129.3, 129.4,

+

d ): δ 35.3, 99.0, 112.4, 120.1, 120.9, 121.2, 122.9, 123.9,

136.4, 142.6, 159.9, 163.3, 165.9. EI-MS: m/z (%) 365 (M ,

6

1

28.3, 131.2, 131.4, 136.4, 136.5, 160.0, 163.2. Anal.Calcd

36), 149 (27), 142 (100), 121 (13), 89 (11), 44 (24). Anal.

Calcd for C H N O S: C, 62.45; H, 4.14; N, 11.50; S, 8.78;

for C H FN OS: C, 62.76; H, 3.72; N, 12.91; S, 9.86;

1

7

12

3

19 15

3

Found: C, 62.82; H, 3.62; N, 12.89; S, 9.73.

Found: C, 62.44; H, 4.00; N, 11.40; S, 8.76.

2

‑(1H‑indol‑3‑yl)‑5‑((4‑(triꢀuoromethyl)benzyl)thio)‑1,3,4‑oxa-

4‑(((5‑(1H‑indol‑3‑yl)‑1,3,4‑oxadiazol‑2‑yl)thio)methyl)

1

diazole (5u) Yield, 86%. mp, 129–131 °C. H NMR

benzoic acid (5y) Yield, 46%. mp, 249–251 °C. H NMR

(

600 MHz, DMSO-d ): δ4.53 (s, 2H), 7.21–7.23 (m,

(600 MHz, DMSO-d ): δ7.22 (m, 1H), 7.26 (m, 1H), 7.53 (d,

6

1

7

1

9

1

3

H),7.25–7.27 (m, 1H), 7.52–7.53 (d, J = 7.8 Hz, 1H),

J=8.4 Hz, 1H), 7.57 (d, J=7.8 Hz, 2H),7.91 (d, J=7.8 Hz,

.91–7.92 (d, J=7.8 Hz, 1H), 8.19 (s, 1H), 12.09 (s, 1H),

2H), 7.98 (d, J=7.8 Hz, 1H), 8.15 (d, J=2.4 Hz, 1H), 12.09

1

3

13

4.49 (s, 1H). CNMR(150 MHz, DMSO-d ): δ35.3, 94.9,

(s, 1H). CNMR(150 MHz, DMSO-d ): δ 35.7, 99.0, 112.5,

6

7.4, 98.8, 120.1, 121.2, 123.3, 124.0, 125.4, 129.4, 129.8,

120.1, 121.2, 122.9, 123.9, 128.3, 129.0, 129.5, 136.4,

+

36.8, 142.1, 147.8, 159.8, 163.4, 166.8. EI-MS: m/z (%)

141.4, 160.0, 163.3. EI-MS: m/z (%) 351 (M , 24), 144

+

75 (M , 51), 159 (37), 144 (100), 116 (11), 89 (10), 44

(100), 134 (22). Anal.Calcd for C H N O S: C, 61.53; H,

1

8

13

3

+

(

20). EI-MS: m/z (%) 375 (M , 51), 159 (37), 144 (100),

3.73; N, 11.96; S, 9.13; Found: C, 61.19; H, 3.51; N, 11.70;

1

16 (11), 89 (10), 44 (20). Anal.Calcd for C H F N OS:

18 12 3 3

S, 9.20.

C, 57.59; H, 3.22; N, 11.19; S, 8.54; Found: C, 57.80; H,

3

.05; N, 10.99; S, 8.72.

General synthetic procedures for compounds 7–10

2

‑((2,4‑diꢀuorobenzyl)thio)‑5‑(1H‑indol‑3‑yl)‑1,3,4‑oxadiazole

Compounds 7–10 were synthesized as shown in Scheme 2

using previously described methods [40, 41], similar to the

synthetic route for compounds 2–5.

1

(

5v) Yield, 70%. mp, 156–158 °C. H NMR (600 MHz,

DMSO-d ): δ4.57 (s, 2H), 7.10 (s, 1H),7.27 (m, 3H), 7.56

6

1

3

Molecular Diversity

Scheme 2 Synthetic route of

2

5

-((1H-indol-3-yl)methyl)-

-thioether-1,3,4-oxadiazoles

ethyl 2‑(1H‑indol‑3‑yl)acetate(7) Yield 98%, mp 46–47 °C.

(10). Anal.Calcd for C H Cl N OS: C, 49.42; H, 3.26; N,

1

4

11

2

3

1

H NMR (600 MHz, DMSO-d ) δ1.43 (t, J = 7.8 Hz,3H),

12.35; S, 9.42; Found: C, 49.43; H, 3.24; N, 12.08; S, 9.56.

6

3

1

.72 (s, 2H), 4.11 (d, J = 7.2 Hz, 2H), 6.99 (t, J = 7.2 Hz,

H), 7.08 (t, J = 7.2 Hz, 1H), 7.24 (s, 1H), 7.35 (d,

2 ‑ ( ( 1 H ‑ i n d o l ‑ 3 ‑ y l ) m e t hy l ) ‑ 5 ‑ ( (c yc l o p ro py l m e t hy l )

1

J=8.0 Hz, 1H), 7.49 (d, J=7.8 Hz, 1H), 10.94 (s, 1H).

thio)‑1,3,4‑oxadiazole (10c) Yield, 87%. mp, 70–72 °C. H

NMR (600 MHz, DMSO-d ): δ0.25 (t, J=2.4 Hz, 2H), 0.50

6

2

‑(1H‑indol‑3‑yl)acetohydrazide(8) Yield 95%, mp 125–

(m, 2H),1.13 (m, 1H), 3.12 (m, 2H), 4.34 (s, 2H), 7.00 (m,

1

27 °C. H NMR (600 MHz, DMSO-d ): δ 3.46 (s, 2H), 4.20

1H),7.10 (m, 1H), 7.34 (s, 1H), 7.38 (d, J = 8.4 Hz, 1H),

6

1

3

(

s, 2H), 6.98 (t, J=7.8 Hz, 1H), 7.07 (t, J=7.8 Hz, 1H), 7.18

s, 1H), 7.34 (d, J=7.8 Hz, 1H), 7.58 (d, J=7.8 Hz, 1H),

7.51 (d, J=8.4 Hz, 1H), 11.04 (s, 1H). CNMR(150 MHz,

(

DMSO-d ): δ 5.6, 10.8, 21.4, 37.7, 106.6, 111.6, 118.2,

6

1

3

9

.16 (s, 1H), 10.87 (s, 1H). C NMR (150 MHz,DMSO-

118.7, 121.3, 124.1, 126.6, 136.2, 163.3, 167.0. EI-MS:

+

d ): δ30.8, 108.7, 111.3, 118.3, 118.7, 120.8, 123.6, 127.2,

m/z (%) 285 (M , 39), 130 (100), 55 (12). Anal.Calcd for

6

+

1

36.1, 170.3. EI-MS: m/z (%) 189 (M , 26), 144 (14), 130

C H N OS: C, 63.13; H, 5.30; N, 14.73; S, 11.24; Found:

1

5

15

3

(

100), 77 (7).

C, 63.30; H, 5.14; N, 14.65; S, 11.27.

5

‑((1H‑indol‑3‑yl)methyl)‑1,3,4‑oxadiazole‑2‑thiol(9) Yield

2 ‑ ( ( 1 H ‑ i n d o l ‑ 3 ‑ y l ) m e t hy l ) ‑ 5 ‑ ( (ox i ra n ‑ 2 ‑ y l m e t hy l )

1

7

4%. mp, 85–87 °C. H NMR (600 MHz, DMSO-d ): δ4.23

thio)‑1,3,4‑oxadiazole (10d) Yield, 21%. mp, 83–85 °C.

6

1

(

s, 2H), 7.01 (m, 1H), 7.11 (m, 1H), 7.36 (s, 1H), 7.39 (m,

H NMR (600 MHz, DMSO-d ): δ2.54 (s, 1H), 2.64 (t,

6

1

H), 7.51 (d, J=8.4 Hz, 1H), 11.09 (s, 1H), 14.35 (s, 1H).

J=2.4 Hz, 1H), 3.28 (t, J=1.2 Hz, 1H),3.37 (m, 1H), 3.44

(m, 1H), 4.38 (s, 2H), 7.04 (m, 1H),7.14 (m, 1H), 7.39 (s,

1H), 7.42 (t, J=7.8 Hz, 1H), 7.55 (d, J=7.8 Hz, 1H), 11.09

1

3

C NMR (150 MHz, DMSO-d ): δ 21.8, 105.7, 111.7,

6

1

18.1, 118.8, 121.4, 124.5, 126.6, 136.2, 163.4, 177.8.

1

3

(

s, 1H). CNMR(150 MHz, DMSO-d ): δ 26.6, 39.3, 111.7,

6

2

1

‑((1H‑indol‑3‑yl)methyl)‑5‑(butylthio)‑1,3,4‑oxadiazole(

116.8, 123.3, 123.9, 126.5, 129.3, 131.7, 141.3, 167.8,

1

+

0a) Yield, 78%. mp, 127–129 °C. H NMR (600 MHz,

172.5. EI-MS: m/z (%) 287 (M , 57), 130 (100). Anal.Calcd

DMSO-d ): δ 0.92 (t, J = 7.8 Hz, 3H),1.48 (m, 2H), 1.67

for C H N O S: C, 58.52; H, 4.56; N, 14.62; S, 11.16;

6

14 13

3

2

(

m, 2H), 3.11 (m, 2H), 4.20 (s, 2H), 7.28 (m, 2H), 7.55

Found: C, 58.66; H, 4.56; N, 14.53; S, 11.35.

d, J = 7.2 Hz, 1H), 8.16 (d, J = 7.8 Hz, 1H), 8.23 (d,

1

3

J=2.4 Hz, 1H), 12.1 (s, 1H). CNMR(150 MHz, DMSO-

ethyl 2‑((5‑((1H‑indol‑3‑yl)methyl)‑1,3,4‑oxadiazol‑2‑yl)

1

d ):12.4, 21.6,22.1, 32.0, 35.9, 98.6,111.1,119.8, 121.4,

thio)acetate (10e) Yield, 37%. mp, 96–98 °C. H NMR

6

1

21.7, 127.4,128.5, 136.6, 160.4, 164.5.Anal.Calcd for

(600 MHz, DMSO-d ): δ 1.13 (t, J=5.4 Hz, 3H), 4.07 (q,

6

C H N OS: C, 62.69; H, 5.96; N, 14.62; S, 11.16; Found:

J = 5.4 Hz, 2H), 4.15 (s, 2H), 4.33 (s, 2H), 7.00 (s, 1H),

1

5

17

3

C, 62.53; H, 5.72; N, 14.62; S, 11.12.

7.10 (s, 1H),7.32 (s, 1H), 7.38 (d, J=6.6 Hz, 1H), 7.50 (d,

1

3

J=6.6 Hz, 1H),11.05 (s, 1H). CNMR(150 MHz, DMSO-

2

‑((1H‑indol‑3‑yl)methyl)‑5‑((3,3‑dichloroallyl)thio)‑1,3,4

d ): δ 13.8, 21.4, 33.7, 61.5, 106.5, 111.6, 118.1, 118.7,

6

1

‑

oxadiazole(10b) Yield, 81%. mp, 135–137 °C. H NMR

121.3, 126.5, 136.2, 162.4, 167.2, 167.5. EI-MS: m/z (%)

+

(

600 MHz, CDCl ): δ4.01 (d, J = 11.4 Hz, 2H),4.20 (s,

317 (M , 46), 130 (100). Anal.Calcd for C H N O S: C,

3

15 15

3

2

7

H),6.33 (m, 1H), 7.29 (s, 1H), 7.34 (t, J = 6.0 Hz, 1H),

56.77; H, 4.76; N, 13.24; S, 10.10; Found: C, 56.66; H, 4.69;

.51 (t, J = 7.2 Hz, 1H), 7.95 (d,J = 3.6 Hz, 1H), 8.29 (d,

N, 13.20; S, 10.15.

1

3

J=7.2 Hz, 1H), 11.88 (s, 1H). CNMR(150 MHz, DMSO-

d ): δ22.1, 31.4, 99.7, 112.6, 119.8, 121.7, 122.8, 123.7,

isopropyl 2‑((5‑((1H‑indol‑3‑yl)methyl)‑1,3,4‑oxadiazol‑2‑yl)

6

1

3

25.9, 126.7, 129.1, 136.5, 160.1, 163.9. EI-MS: m/z (%)

thio)acetate (10f) Yield, 71%. mp, 74–76 °C. H NMR

+

25 (M , 31), 290 (21), 216 (8), 144 (100), 116 (13), 89

(600 MHz, DMSO-d ): δ1.12 (d, J = 6.0 Hz, 6H),4.11 (s,

6

1

3

Molecular Diversity

+

2

H), 4.33 (s, 2H), 4.88 (m, 1H), 7.00 (t, J=7.8 Hz, 1H),7.10

136.2, 136.5, 162.7, 167.3. EI-MS: m/z (%) 321 (M , 61),

(

t, J = 7.8 Hz, 1H), 7.32 (s, 1H), 7.38 (d, J = 8.4 Hz, 1H),

156 (35), 130 (100), 91 (53), 77 (8), 44 (8). Anal.Calcd for

C H N OS: C, 67.27; H, 4.70; N, 13.07; S, 9.98; Found:

1

3

7

.50 (d, J=8.4 Hz, 1H), 11.05 (s, 1H). CNMR(150 MHz,

1

8

15

3

DMSO-d ): δ 21.3, 21.4, 33.9, 69.2, 106.5, 111.6, 118.1,

C, 67.37; H, 4.69; N, 12.93; S, 9.93.

6

1

18.7, 121.3, 124.1, 126.5, 136.2, 162.4, 167.0, 167.2.

+

EI-MS: m/z (%) 331 (M , 48), 130 (100). Anal.Calcd for

2‑((1H‑indol‑3‑yl)methyl)‑5‑((2‑methylbenzyl)thio)‑1,3,4‑oxa-

1

C H N O S: C, 57.99; H, 5.17; N, 12.68; S, 9.68; Found:

diazole (10k) Yield, 80%. mp, 129–131 °C. H NMR

1

6

17

3

C, 57.88; H, 5.05; N, 12.56; S, 9.66.

(600 MHz, DMSO-d ): δ 2.31 (s, 3H), 4.34 (s, 2H), 4.42

6

(

s, 2H), 7.01 (m, 2H),7.12 (d, J=7.2 Hz, 1H), 7.15 (m, 3H),

tert‑butyl 2‑((5‑((1H‑indol‑3‑yl)methyl)‑1,3,4‑oxadiazol‑2‑yl)

thio)acetate (10g) Yield, 89%. mp, 100–102 °C. H NMR

7.34 (s, 1H), 7.40 (d, J=7.8 Hz, 1H), 7.51 (d, J=7.8 Hz,

1

13

1H),11.08 (s, 1H). CNMR(150 MHz, DMSO-d ): δ 18.6,

6

(

600 MHz, DMSO-d ): δ1.33 (s, 9H),4.03 (s, 2H), 4.33 (s,

21.5, 34.4, 106.5, 111.6, 118.2, 118.8, 121.3, 124.2, 126.0,

6

2

H), 7.00 (t, J=7.2 Hz, 1H),7.10 (t, J=7.8 Hz, 1H), 7.33

126.6, 128.1, 129.8, 130.4, 133.7, 136.2, 136.6, 162.5,

+

(

s, 1H), 7.38 (d, J=7.8 Hz, 1H), 7.51 (d, J=7.8 Hz, 1H),

167.3. EI-MS: m/z (%) 335 (M , 53), 320 (4), 231 (9), 205

13

1

3

1

2

1.05 (s, 1H). C NMR (150 MHz, DMSO-d ): δ 21.4, 27.4,

(6), 156 (39), 130 (21), 105 (100), 77 (16), 44 (19). Anal.

Calcd for C H N OS: C, 68.03; H, 5.11; N, 12.53; S, 9.56;

6

4.6, 81.9, 106.5, 111.6, 118.1, 118.7, 121.3, 124.1, 126.5,

19

17

3

+

36.2, 162.5, 166.5, 167.1. EI-MS: m/z (%) 345 (M , 45),

Found: C, 67.92; H, 5.30; N, 12.37; S, 9.31.

89 (25), 130 (100). Anal.Calcd for C H N O S: C, 59.11;

17

19

3

H, 5.54; N, 12.17; S, 9.28; Found: C, 59.15; H, 5.53; N,

2‑((1H‑indol‑3‑yl)methyl)‑5‑((2‑ꢀuorobenzyl)thio)‑1,3,4‑oxa-

1

2.07; S, 9.47.

diazole (10l) Yield, 88%. mp, 101–103 °C. H NMR

(

600 MHz, DMSO-d ): δ 4.33 (s, 2H), 4.43 (s, 2H), 7.01

6

ethyl 2‑((5‑((1H‑indol‑3‑yl)methyl)‑1,3,4‑oxadiazol‑2‑yl)

(m, 2H),7.14 (m, 2H), 7.31 (m, 3H), 7.39 (d, J=7.8 Hz, 1H),

1

3

thio)‑2‑methylpropanoate (10h) Yield, 32%. mp, 126-

7.49 (d, J=7.2 Hz, 1H), 11.07 (s, 1H). CNMR(150 MHz,

1

28 °C. H NMR (600 MHz, DMSO-d ): δ0.98 (t, J=7.2 Hz,

DMSO-d ): δ21.4, 29.7, 106.5, 111.6, 115.3, 115.5,

6

3

H), 1.53 (m, 6H),3.91 (q, J=7.2 Hz, 2H), 4.37 (s, 2H), 7.00

118.1, 118.7, 121.3, 123.4 (d, J = 13.5 Hz, 1C), 124.3

C–F

(

t, J = 7.8 Hz, 1H),7.10 (t, J = 7.8 Hz, 1H), 7.33 (s, 1H),

(d, J = 27.6 Hz, 1C), 126.6, 130.1 (d, J = 8.1 Hz,

C–F

7

1

5

1

5

.38 (d, J=7.8 Hz, 1H), 7.50 (d, J=7.8 Hz, 1H), 11.06 (s,

1C), 131.2 (d, J = 2.3 Hz, 1C), 136.2, 159.4, 161.1,

C–F

1

3

+

H). C NMR (150 MHz, DMSO-d ): δ 13.5, 21.6, 25.7,

162.2, 167.5. EI-MS: m/z (%) 339 (M , 48), 238 (11), 156

6

3.2, 61.4, 106.4, 111.5, 118.1, 118.7, 121.3, 124.1, 126.5,

(25), 130 (21), 109 (34), 86 (18), 77 (5). Anal.Calcd for

C H FN OS: C, 63.70; H, 4.16; N, 12.38; S, 9.45; Found:

36.2, 160.2, 168.6, 171.4. Anal.Calcd for C H N O S: C,

17

19

3

18 14

3

9.11; H, 5.54; N, 12.17; S, 9.28; Found: C, 59.34; H, 5.46;

C, 63.83; H, 4.02; N, 12.16; S, 9.66.

N, 12.41; S, 9.44.

2

‑((1H‑indol‑3‑yl)methyl)‑5‑((3‑ꢀuorobenzyl)thio)‑1,3,4‑oxadi-

1

(

E)‑ethyl 4‑((5‑((1H‑indol‑3‑yl)methyl)‑1,3,4‑oxadiazol‑2‑yl)

azole (10m) Yield, 65%. mp, 85–87 °C. H NMR (600 MHz,

1

thio)but‑2‑enoate (10i) Yield, 30%. mp, 86–88 °C. H NMR

DMSO-d ): δ4.33 (s, 2H), 4.43 (s, 2H), 7.00 (m, 1H),7.08

6

(

600 MHz, DMSO-d ): δ1.20 (s, 3H),4.01 (d, J = 6.0 Hz,

(m, 3H), 7.24 (m, 2H), 7.26 (s, 1H),7.38 (m, 1H), 7.49 (d,

6

1

3

2

H), 4.11 (d, J = 6.6 Hz, 2H), 4.34 (s, 2H), 6.02 (d,

J=7.8 Hz, 1H), 11.08 (s, 1H). CNMR(150 MHz, DMSO-

d ): δ 21.5, 35.1, 106.5, 111.6, 114.6 (d, J =23.7 Hz, 1C),

J=15.0 Hz, 1H), 6.88 (t, J=7.8 Hz, 1H),7.01 (d, J=6.6 Hz,

6

C–F

1

7

H), 7.10 (s, 1H), 7.34 (s, 1H), 7.38 (d, J = 7.8 Hz, 1H),

115.7 (d, J = 22.2 Hz, 1C), 118.2, 118.8, 121.4, 124.2,

C–F

1

3

.50 (d, J=6.6 Hz, 1H), 11.05 (s, 1H). CNMR(150 MHz,

125.0 (d, J =2.25 Hz, 1C), 126.6, 130.4(d, J =8.1 Hz,

C–F

DMSO-d ): δ 14.0, 21.4, 32.3, 60.1, 106.5, 111.6, 118.1,

1C), 136.2, 139.5 (d, J = 8.1 Hz, 1C), 161.1, 162.6 (d,

6

C–F

+

1

18.7, 121.3, 124.0, 124.2, 126.5, 136.2, 141.9, 162.2,

J

=27.6 Hz, 1C), 167.4. EI-MS: m/z (%) 339 (M , 56),

C–F

+

65.0, 167.4. EI-MS: m/z (%) 343 (M , 20), 230 (79), 130

238 (8), 156 (17), 130 (100), 109 (28), 77 (5). Anal.Calcd for

(

100). Anal.Calcd for C H N O S: C, 59.46; H, 4.99; N,

C H FN OS: C, 63.70; H, 4.16; N, 12.38; S, 9.45; Found:

1

7

17

3

18 14

3

1

2.24; S, 9.34; Found: C, 59.39; H, 5.08; N, 12.12; S, 9.57.

C, 63.74; H, 4.33; N, 12.33; S, 9.59.

2

‑((1H‑indol‑3‑yl)methyl)‑5‑(benzylthio)‑1,3,4‑oxadiazole

2‑((1H‑indol‑3‑yl)methyl)‑5‑((3‑methylbenzyl)thio)‑1,3,4‑oxa-

1

(

10j) Yield, 92%. mp, 117–119 °C. H NMR (600 MHz,

diazole (10n) Yield, 50%. mp, 97–99 °C. H NMR

DMSO-d ): δ4.34 (s, 2H), 4.40 (s, 2H), 7.01 (m, 1H),7.11

(600 MHz, DMSO-d ): δ2.50 (s, 3H), 4.33 (s, 2H), 4.37 (s,

6

(

m, 1H), 7.22 (s, 3H), 7.27 (s, 2H),7.34 (s, 1H), 7.40

2H), 7.01 (t, J=7.8 Hz, 1H),7.04 (d, J=7.8 Hz, 1H), 7.06 (s,

(

d, J = 8.4 Hz, 1H), 7.50 (d, J = 7.2 Hz, 1H), 11.09 (s,

1H), 7.10 (m, 3H), 7.33 (s, 1H), 7.39 (d, J=7.8 Hz, 1H),7.51

1

3

13

1

H). CNMR(150 MHz, DMSO-d ): δ21.5, 35.8, 106.6,

(d, J = 7.8 Hz, 1H), 11.06 (s, 1H). CNMR(150 MHz,

6

11.6, 118.8, 121.4, 124.3, 126.6, 128.7, 128.5, 128.9,

DMSO-d ): δ20.8, 21.4, 35.8, 106.6, 111.6, 118.1, 118.8,

6

1

3

Molecular Diversity

1

21.3, 124.2, 126.0, 126.6, 128.3, 129.4, 136.2, 136.3,

159.6, 161.1 (d, J = 12.6 Hz, 1C), 162.1, 162.7, 167.5.

C-F

+

37.7, 162.7, 167.2. EI-MS: m/z (%) 335 (M , 68), 156

EI-MS: m/z (%) 357 (M , 42), 156 (24), 130 (100), 127 (62),

(

53), 130 (100), 105 (97), 77 (14), 44 (12). Anal.Calcd for

77 (7). Anal.Calcd for C H F N OS: C, 60.49; H, 3.67; N,

1

8

13

2

3

C H N OS: C, 68.03; H, 5.11; N, 12.53; S, 9.56; Found:

11.76; S, 8.97; Found: C, 60.67; H, 3.60; N, 11.57; S, 9.10.

1

9

17

3

C, 68.14; H, 5.16; N, 12.46; S, 9.63.

2

‑((1H‑indol‑3‑yl)methyl)‑5‑((4‑(trifluoromethoxy)benzyl)

2

‑((1H‑indol‑3‑yl)methyl)‑5‑((3‑nitrobenzyl)thio)‑1,3,4‑oxa-

thio)‑1,3,4‑oxadiazole (10s) Yield, 89%. mp, 113–115 °C.

1

diazole (10o) Yield, 83%. mp, 135–137 °C. H NMR

H NMR (600 MHz, DMSO-d ): δ 4.34 (s, 2H), 4.43 (s, 2H),

6

(

600 MHz, DMSO-d ): δ4.33 (s, 2H), 4.57 (s, 2H), 6.99

7.00 (m, 1H),7.11 (t, J = 7.2 Hz, 1H), 7.17 (d, J = 7.8 Hz,

6

t, J = 7.2 Hz, 1H),7.11 (d, J = 7.8 Hz, 1H), 7.33 (s, 1H),

2H), 7.35 (s, 1H), 7.39 (m, 3H), 7.49 (s, J = 7.8 Hz, 1H),

1

3

7

1

.39 (d, J=8.4 Hz, 1H), 7.48 (m, 1H), 7.52 (d, J=7.8 Hz,

11.08 (s, 1H). C NMR (150 MHz, DMSO-d ): δ 21.4,

6

13

H),8.09 (d, J=8.4 Hz, 1H), 8.33 (s, 1H), 11.07 (s, 1H). C

34.8, 106.5, 111.6, 118.1, 118.7, 119.1, 120.9, 121.3, 124.2,

NMR (150 MHz, DMSO-d ): δ 21.4, 34.7, 106.5, 111.6,

126.6, 130.8, 136.2, 136.3, 147.6, 162.5, 167.3. EI-MS: m/z

6

+

1

3

18.1, 118.7, 121.3, 122.5, 123.7, 124.2, 126.5, 129.8,

(%) 405 (M , 53), 175 (90), 156 (37), 130 (100), 77 (6).

35.5, 136.2, 139.3, 147.6,,162.4, 167.4. EI-MS: m/z (%)

Anal.Calcd for C H F N O S: C, 56.29; H, 3.48; N, 10.37;

1

9

14

3

2

+

66 (M , 31), 349 (10), 230 (7), 136 (11), 130 (100), 103

S, 7.91; Found: C, 56.12; H, 3.62; N, 10.33; S, 7.84.

(

7), 44 (8). Anal.Calcd for C H N O S: C, 59.01; H, 3.85;

18 14 4 3

N, 15.29; S, 8.75; Found: C, 59.07; H, 3.62; N, 15.10; S,

methyl 4‑(((5‑((1H‑indol‑3‑yl)methyl)‑1,3,4‑oxadiazol‑2‑yl)

8

.51.

thio)methyl)benzoate (10t) Yield, 93%. mp, 127–129 °C.

1

H NMR (600 MHz, DMSO-d ): δ3.84 (s, 3H), 4.33 (s, 2H),

6

2

‑((1H‑indol‑3‑yl)methyl)‑5‑((4‑methylbenzyl)thio)‑1,3,4‑oxa-

4.46 (s, 2H), 7.00 (t, J=7.2 Hz, 1H),7.11 (d, J=7.8 Hz, 1H),

1

diazole (10p) Yield, 74%. mp, 127–129 °C. H NMR

7.34 (s, 1H), 7.40 (d, J=8.4 Hz, 3H), 7.48 (d, J=8.4 Hz,

1

3

(

600 MHz, DMSO-d ): δ 2.50 (s, 3H), 4.33 (s, 2H), 4.35

1H), 7.78 (d, J = 7.8 Hz, 2H),11.07 (s, 1H). C NMR

6

s, 2H), 7.00 (m, 3H),7.12 (m, 3H), 7.34 (d, J=1.2 Hz, 1H),

(150 MHz, DMSO-d ): δ 21.4, 35.3, 52.1, 106.5, 111.6,

6

7

1

3

.40 (d, J=8.4 Hz, 1H),7.50 (d, J=7.8 Hz, 1H), 11.07 (s,

118.1, 118.8, 121.3, 124.2, 126.5, 128.8, 129.1, 129.3,

1

3

+

H). CNMR(150 MHz, DMSO-d ): δ20.6, 21.4, 35.6,

136.2, 142.2, 162.3, 165.8, 167.4. EI-MS: m/z (%) 379 (M ,

6

06.6, 111.6, 118.2, 118.7, 121.3, 124.2, 126.6, 128.8,

53), 230 (35), 156 (15), 149 (26), 130 (100), 121 (9), 90 (8),

44 (8). Anal.Calcd for C H N O S: C, 63.31; H, 4.52; N,

29.0, 133.3, 136.2, 136.9, 162.7, 167.2. EI-MS: m/z (%)

2

0

17

3

+

35 (M , 38), 156 (40), 130 (56), 105 (100). Anal.Calcd for

11.07; S, 8.45; Found: C, 63.19; H, 4.55; N, 10.93; S, 8.61.

C H N OS: C, 68.03; H, 5.11; N, 12.53; S, 9.56; Found:

1

9

17

3

C, 68.26; H, 5.22; N, 12.25; S, 9.59.

X‑ray difraction analysis

2

‑((1H‑indol‑3‑yl)methyl)‑5‑((4‑(trifluoromethyl)benzyl)

The crystallographic data of representative compound 5d

have been deposited with the Cambridge Crystallographic

Data Centre (CCDC) as supplementary publication number:

thio)‑1,3,4‑oxadiazole (10q) Yield, 91%. mp, 127–129 °C.

1

H NMR (600 MHz, DMSO-d ): δ 4.35 (s, 2H), 4.48 (s,

6

2

H), 7.01 (t, J=7.8 Hz, 1H),7.12 (t, J=7.2 Hz, 1H), 7.36

9

21313 (Fig. 4 ). Copy of the data can be obtained free of

(

s, 1H), 7.41 (d, J = 7.8 Hz, 1H), 7.49 (m, 3H), 7.53 (d,

charge via http://www.ccdc.cam.ac.uk/, or on application to

CCDC, 12 Union Road, Cambridge CB12 1EZ, UK (Fax

13

J=7.8 Hz,2H), 11.09 (s, 1H). C NMR (150 MHz, DMSO-

d ): δ 21.5,35.0,106.5, 111.6, 118.1, 118.8, 121.3, 124.3,

6

+

44 1223 336033 or E-mail: deposit@ccdc.cam.ac.uk).

1

25.2, 126.6, 128.1 (d, J = 31.1 Hz, 1C), 129.6, 136.2,

C-F

+

41.7, 162.4, 167.4. EI-MS: m/z (%) 389 (M , 65), 159 (14),

30 (100). Anal.Calcd for C H F N OS: C, 58.60; H, 3.62;

Biological assays

19

14

3

N, 10.79; S, 8.23; Found: C, 58.85; H, 3.53; N, 10.56; S,

8

.42.

Leaf‑piece assays

2

‑((1H‑indol‑3‑yl)methyl)‑5‑((2, 4‑difluorobenz yl)

Water agar of 200–300 μL per well was dispensed into the

96-well assay plates, and 6-mm-diameter leaf pieces (6 mm

length for wheat) were cut and placed on the surface of the

agar. Compounds to be tested were formulated to the appro-

priate rate, and 10 μL samples were dispensed onto the leaf

pieces. The plates were inoculated with spore suspensions

of the appropriate pathogen at a set rate (Table 2), using a

handheld spray gun. The plates were covered with lids and

1

thio)‑1,3,4‑oxadiazole (10r) Yield, 59%. mp, 75–77 °C. H

NMR (600 MHz, DMSO-d ): δ 4.34 (s, 2H), 4.41 (s, 2H),

6

.85 (m, 1H),7.01 (m, 1H), 7.11 (m, 1H), 7.18 (m, 1H), 7.31

(

m, 2H), 7.40 (d, J = 7.8 Hz,1H), 7.49 (d, J = 7.8 Hz,1H),

1

3

1

1.07 (s, 1H). C NMR (150 MHz, DMSO-d ): δ 21.4, 29.2,

6

04.0 (t, J =25.3 Hz, 1C), 106.5, 111.5 (t, J =19.0 Hz,

C-F

C), 118.1, 118.7, 120.0, 121.3, 124.2, 126.6, 132.2, 136.2,

1

3

Molecular Diversity

Fig. 4 X-ray diꢂraction struc-

ture of 5d

Table 2 Details of the primary-

_C_h_e_m_i_c_a_l_r_a_t_e_sa

_R_e_p_l_i_c_a_t_e_sb

_S_p_o_r_e_r_a_t_e_sc

Pathogen

Host

level leaf-piece assays

Phytophthora infestans

Septoria tritici

Uromyces viciae-fabae

Tomato

Wheat

Field bean

200, 60

100

2

3

2

150,000

1,000,000

c

100

0.3

a

Rates are given as parts per million a.i. in the formulated sample applied to the leaf piece

Number of replicates per rate

Spore rates are sporangia per ml for P. infestans, conidia per ml for S. tritici, and mg per mL for U. viciae-

b

c

fabae

placed in controlled environment cabinets for between 5 and

4 days, depending on the species being tested.

90 μL spore suspension, to give the chemical rates detailed

in Table 3). The plates were then stored in controlled envi-

ronments set to appropriate conditions, until assessment

between 5 and 14 days later, depending on the species.

1

Artiꢀcial media assays

Spore suspensions were prepared from stock plates of the

target pathogens (Table 3) for all species apart from P. dis-

simile, for which a suspension of fragmented mycelia was

used. These suspensions were made into a 3% agar and dis-

pensed into 96-well plates containing formulated samples of

the compounds to be tested (10 μL formulated sample and

Assessment of biological assays

Assessment for all assays was carried out by eye, with each

individual well scored on a three-band pattern with addi-

tional result types for assay failure and phytotoxic activity

on the leaf piece (Table 4). The resulting data were collated

Table 3 Details of the Primary-

level assays conducted in

artiꢁcial media

_C_h_e_m_i_c_a_l_r_a_t_e_sa

_R_e_p_l_i_c_a_t_e_sb

_S_p_o_r_e_r_a_t_ec

₀_.₅c

20,000

15,000

10,000

Pathogen

Pythium dissimile

Alternaria solani

Botryotinia fuckeliana (Botrytis cinerea)

Gibberella zeae (Fusarium graminearum)

20, 2

2

a

Rates are given as parts per million a.i. in the ꢁnal artiꢁcial media mixture

Number of replicates per rate

Spore rates are given in spores per ml, with the exception of P. dissimile which is given as an optical den-

b

c

sity reading at 425 nm

1

3

Molecular Diversity

Table 4 Assessment criteria for the biological assays

3-Fluorobenzyl bromide was more than 1.1 equivalents.

Therefore, a focused library of 25 thioethers varying from

alkyl, cycloalkyl, isopentene, dichloropropene and ester

to substituted benzyl substituent was set up efficiently

with this optimized method, as shown in Table 5. Based

on the initial design and optimized synthetic method, the

second series of 20 target molecules with a methylene

bridge between indole ring and 1,3,4-oxadiazole ring

was synthesized subsequently, as shown in Scheme 2 and

Table 6.

Scoring band

Well condition

0

5

9

0–49% control of disease or pathogen

50–80% control of disease or pathogen

81–100% control of disease or pathogen

Not captured (too phytotoxic)

5

9

NCH

NC

Not captured (test failure)

All of the target molecules listed in Tables 5 and 6

were characterized on the basis of physical and spectral

data. In addition, compound 5d was further confirmed

by X-ray diffraction crystallography, as shown in Fig. 4.

for each compound, and averages across replicates were

used to make a judgment of the overall activity level of the

compound.

Antiꢀungal activity and the structure–activity

relationships (SAR)

Results and discussion

The results of the biological testing against seven phytopath-

ogenic fungi are shown in Table 7. For the purposes of an

analysis of structure–activity relationships, the antifungal

activities of compounds 5 and 10 were compared to the lead

compounds streptochlorin and pimprinine.

Synthetic chemistry

Based on the strategy of diversity-oriented synthesis

or DOS, which aims to develop relatively small library

with an emphasis on substituent or skeleton diversity

for the discovery of biologically active molecules [42,

Although the antifungal activity of most of 1,3,4-oxadia-

zole-5-thioether-containing pimprinine derivatives has been

proven to be rather poor, making it hard to extract a clear

structure–activity relationship summary, some preliminary

conclusions can still be drawn.

4

1

3], two series of pimprinine derivatives containing

,3,4-oxadiazole-5-thioether moieties were smoothly

synthesized in high yields and then were screened for

their antifungal activity. The first series of target mol-

ecules 5 were prepared in a four-step process involving

sequential conversion of indole-3-carboxylic acid 1 into

methyl indole-3-carboxylate 2, indole-3-carbohydrazide

Firstly, it is noticeable that the antifungal activity of target

compounds lacks potency. Though some compounds (5h,

10a) showed broader antifungal spectrum and more anti-

fungal potential than pimprinine, most of the target com-

pounds were inactive against the tested fungi, compared

with lead compound streptochlorin. Secondly, 1,3,4-oxadi-

azole-5-thioether-containing pimprinine derivatives were

relatively more active against P. dissimile, Uromyces viciae-

fabae and S. tritici rather than the other tested fungi, four

compounds (5b, 5h, 5j and 5l) possessed eꢂective or even

excellent control against P. dissimile, and 5h was identiꢁed

as the most promising candidate, as it possessed signiꢁcant

activity against P. dissimile and also a broad antifungal spec-

trum. Thirdly, the overall eꢂect on the antifungal activity of

methylene bridge in 1,3,4-oxadiazole-5-thioether-containing

pimprinine derivatives seems unhelpful due to the general

3

and 5-(3′-indolyl)-1,3,4-oxadiazole-2-thiol 4 in high

yields, as shown in Scheme 1. The reaction conditions

of intermediate 4 and 3-Fluorobenzyl bromide, including

temperature, solvent, base and ration, were optimized for

the subsequent substitution to –SH group, as shown in

Table 1, and compound 5p could be prepared in almost

quantitative yield under 40 °C, DMF as the solvent, Et N

3

as the base, and the ratio 1:1.1:2 (thiol: benzyl bromide:

base). It is worth noting that the N-substituted compounds

did not form during the synthesis of compounds 5 from

4

, showing excellent selectivity between –SH and NH,

while N-substituted product could be isolated when

1

3

Molecular Diversity

Table 5 Yields for the synthesis of compounds of 5

1

3

Molecular Diversity

Table 6 Yields for the synthesis of compounds of 10

absence of biological activity, though compounds 10a and

Biological assays conducted at Syngenta showed that

the synthesized derivatives displayed an altered pattern of

biological activity compared to the natural product pimprin-

ine, and 5h was identiꢁed as the most promising candidate,

as it possessed signiꢁcant activity to P. dissimile and also

a broad antifungal spectrum in primary screening, though

the antifungal activity of target compounds lacks potency,

methylene bridge in 1,3,4-oxadiazole-5-thioether-containing

pimprinine derivatives seems unhelpful to improve the anti-

fungal activity.

1

0o showed improved antifungal activity, compared with

their unbridged counterparts.

Conclusions

Based on the strategy of diversity-oriented synthesis (DOS)

and the structures of natural product pimprinine and strep-

tochlorin, we have eꢀciently synthesized two series of pim-

prinine derivatives containing 1,3,4-oxadiazole-5-thioether

moieties under the optimized reaction condition.

Further structural optimization of pimprinine and strep-

tochlorin derivatives is well under way, alongside more

1

3

Molecular Diversity

Table 7 Antifungal activity of

_P_H_Ya

_S_E_Pa

_U_R_Oa

_P_Y_Ta

20

_A_L_Ta

_B_O_Ta

_G_I_Ba

Species

Compd

pimprinine derivatives

Rate

200

60

100

2

20

2

20

2

20

2

5

1

a

b

c

d

e

f

g

h

i

j

k

l

m

n

o

p

q

r

s

t

u

v

w

x

y

0^b

0

49

0

49

0

33

18

0

33

51

0

36

51

0

NC

0

55

0

27

0

49

0

77

0

99

0

27

0

99

0

77

0

77

0

55

0

55

0

99

0

55

0

99

0

99

0

d

0

27

0

27

0

49

0

c

0a

0b

0c

0d

0e

1

0f

0g

0h

0i

0j

0k

0l

0m

0n

0o

0p

0q

0r

0s

0

1

0t

Streptochlorin

Pimprinine

55

27

99

0

99

0

a

PHY Phytophthora infestans (on tomato leaf pieces), SEP Septoria tritici (on wheat leaf pieces), URO

Uromyces viciae-fabae (on bean leaf pieces), PYT Pythium dissimile, ALT Alternaria solani, BOT Botryo-

tinia fuckeliana, GIB Gibberella zeae (all in artiꢁcial media)

The data are the mean of three replicates

data not captured for this replicate (test failure)

b

c

d

Bold means above 50% control

1

3

Molecular Diversity

detailed biological testing of the active compounds, with

the aim of improving their levels of antifungal activity.

Devel Ther 9:2201–2214. https://doi.org/10.2147/DDDT.S8020

5

1

3. Choi IK, Shin HJ, Lee HS, Kwon HJ (2007) Streptochlorin, a

marine natural product, inhibits NF-kappaB activation and sup-

presses angiogenesis in vitro. J Microbiol Biotechnol 17(8):1338–

Acknowledgements We are grateful to the financial sup-

port for this work from the National Natural Science Founda-

tion of China (21602110), National Key R&D Program of China

1

343. https://doi.org/10.1371/journal.pone.0074194

4. Lee SH, Shin HJ, Kim DY, Shim DW, Kim TJ, Ye SK, Won

HS, Koppula S, Kang TB, Lee KH (2013) Streptochlorin sup-

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of Lyn/Fyn and Syk signaling pathways in cellular and mouse

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al.pone.0074194

(

2018YFC1602804), Jiangsu Provincial Science Foundation for Youths

BK20160734) and the Fundamental Research Funds and for the Cen-

(

tral Universities (KYTZ201604, KYLH201908). We thank the Biology

Team at Syngenta for their kind help in screening the compounds for

biological activity.

1

5. Park C, Shin HJ, Kim GY, Kwon TK, Nam TJ, Kim SK, Cheong J,

Choi IW, Choi YH (2008) Induction of apoptosis by streptochlorin

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Article Doi

Diversity-oriented synthesis and antifungal activities of novel pimprinine derivative bearing a 1,3,4-oxadiazole-5-thioether moiety

DOI: 10.1007/s11030-020-10048-8

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