Synthesis and biological activity of novel 1,3,4-oxadiazole derivatives containing a pyrazole moiety

Article abstract of DOI:10.1007/s11164-019-04015-8

Several new 1,3,4-oxadiazole derivatives containing a pyrazole ring were designed and synthesized from ethyl acetoacetate and triethyl orthoformate as starting materials via multi-step reactions. The compound structures were confirmed by melting point, ¹H NMR and HRMS. They were evaluated for fungicidal and herbicidal activities. Four of the compounds exhibited moderate fungicidal activity against Colletotrichum species. Most of the compounds had moderate-to-good activity as a herbicide.

Full text of DOI:10.1007/s11164-019-04015-8

Research on Chemical Intermediates
https://doi.org/10.1007/s11164-019-04015-8
Synthesis and biological activity of novel 1,3,4‑oxadiazole
derivatives containing a pyrazole moiety
Wei Yu1 · Zhi‑Wen Zhai1 · David E. Wedge2 · Stephen O. Duke2 · Hong‑Ke Wu1 ·
Jian‑Quan Weng1 · Cheng‑Xia Tan1 · Yong‑Gang Zhang3 · Xing‑Hai Liu1
Received: 2 June 2019 / Accepted: 1 August 2019
© Springer Nature B.V. 2019
Abstract
Several new 1,3,4-oxadiazole derivatives containing a pyrazole ring were designed
and synthesized from ethyl acetoacetate and triethyl orthoformate as starting mate-
rials via multi-step reactions. The compound structures were confrmed by melt-
1
ing point, H NMR and HRMS. They were evaluated for fungicidal and herbicidal
activities. Four of the compounds exhibited moderate fungicidal activity against
Colletotrichum species. Most of the compounds had moderate-to-good activity as a
herbicide.
Keywords Oxadiazole · Pyrazole · Synthesis · Fungicide · Herbicide
Introduction
Heterocycles play an important role in natural product chemistry [1–3], medicinal
chemistry [4–6], organic synthetic chemistry [7, 8] and pesticidal chemistry [9]. The
pyrazole ring, a fve-membered heterocyclic compound, has been used considerably
in synthetic medicines and pesticides [10–13]. Pyrazole compounds possess diverse
biological activities, such as herbicidal [14, 15], antioxidant [16], antimicrobial
[17], anticancer [18], nematocidal [10, 19–27], fungicidal [28–37] and insecticidal
*
Xing-Hai Liu
xhliu@zjut.edu.cn
Stephen O. Duke
sduke@olemiss.edu
Yong-Gang Zhang
zhangygcq@163.com
1
College of Chemical Engineering, Zhejiang University of Technology, Hangzhou 310014,
Zhejiang, China
2
3
USDA ARS, Natural Products Utilization Research Unit, University, MS 38677, USA
Biology Institute, Qilu University of Technology (Shandong Academy of Sciences), Jinan,
Shandong, China
Vol.:(0123456789)
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W. Yu et al.
activities [38]. Most of the succinate dehydrogenase inhibitor (SDHI) fungicides, an
important fungicide class, have pyrazole rings. In addition, many references report
1,3,4-oxadiazole compounds with diverse biological activities, such as ketol-acid
reductoisomerase inhibitors [39] and antiviral [40], anti-tumor [41], fungicidal [42]
and antiproliferative activities [43].
Penfufen has good activity against Rhizoctonia solani and was developed com-
mercially as a seed treatment, potato tuber treatment and soil application to protect
seeds and seedlings against a wide range of fungal pathogens in feld crops. Our
interest in this study was to evaluate this group of pyrazole derivatives for possible
activity against any of the three Colletotrichum species common to another high-
value feld crop, strawberry (Fragaria×ananassa Duchesne). In line with our con-
tinued eforts on the synthesis of novel lead compounds for pesticides [44–55], the
commercial carboxamide fungicide penfufen was selected as a lead compound. The
scafold of this lead compound is maintained at the 1-methyl-3-methyl-1H-pyrazole
moiety of the molecule. The fuorine atom was replaced by hydrogen atom, and the
carboxamide group was cyclized by 1,3,4-oxidiazole ring. Our original strategy is
depicted in Fig. 1. It is possible that pyrazole amide derivatives possess fungicidal
activity. Most of them exhibited good herbicidal activity against a monocot, creep-
ing bentgrass (Agrostis stolonifera).
Experimental
General information
The chemical intermediates were prepared in our laboratory, and reagents were of
analytical grade. Melting points were recorded on an X-4 apparatus and were uncor-
1
rected. H-NMR spectra were determined on an Avance 500 MHz spectrometer
using CDCl3 as solvent. HRMS were recorded on an Agilent TOF–MS instrument.
H
3C
F
O
N
N
N
R
N
CH3
S
NH
cyclized amide group
O
N
N
5a~5l
Lead compound
Fungicides
Fig. 1 Design strategy of the pyrazole compounds with 1,3,4-oxadiazole moiety
1 3

Synthesis and biological activity of novel 1,3,4-oxadiazole…
Synthesis
Ethyl 1,3‑dimethyl‑1H‑pyrazole‑4‑carboxylate (2)
In a 1000-mL four-necked fask, ethyl 3-oxobutanoate (130 g, 1 mol), triethyl ortho-
formate (296 g, 2 mol) and acetic anhydride (255 g, 2.5 mol) were added and heated
at 120 °C for 10 h. Then, the solvent was removed and the product was then distilled
under vacuum over a column to give a brown liquid. Then, the CH3NHNH2 was
added into the solution of brown liquid in EtOH (550 mL) at 0 °C, and the mixture
was stirred at room temperature for 3 h. The solvent was evaporated, and the result-
ing white solid was produced with a yield of 51.2%, 86.1 g, m.p. 48–50 °C.
1,3‑Dimethyl‑1H‑pyrazole‑4‑carbohydrazide (3)
To a solution of ethyl 1,3-dimethyl-1H-pyrazole-4-carboxylate (1.68 g, 10 mmol)
in EtOH (10 mL), 80% hydrazine hydrate (5.63 g, 90 mmol) was added dropwise.
Then, the mixture solution was stirred and refuxed for 8 h. The solvent was evapo-
rated and cooled, and a white solid was given with a yield of 92.1%, 1.42 g, m.p.
140–141 °C.
5‑(1,3‑Dimethyl‑1H‑pyrazol‑4‑yl)‑1,3,4‑oxadiazole‑2‑thiol (4)
The solution of 1,3-dimethyl-1H-pyrazole-4-carbohydrazide (15.4 g) and KOH
(7 g) in EtOH (300 mL) was stirred at 0 °C for 15 min; then, the CS2 (22.8 g) was
added dropwise. The mixture was refuxed for 8 h. The EtOH was evaporated, and
the residue was dissolved in water. The solution was acidifed by conc. HCl, and a
fnal intermediate 4 was produced and fltered, giving a yield of 84%, 16.56 g, m.p.
237–238 °C, 1H NMR (500 MHz, CDCl3) δ: ppm 7.84 (s, 1H, pyrazole-H), 3.92 (s,
3H, N-CH3), 2.48 (s, 3H, pyrazole-CH3).
General procedure for the synthesis of compounds 5a–5m
N,N-Dimethylformamide (5 mL), 5-(1,3-dimethyl-1H-pyrazol-4-yl)-1,3,4-oxadia-
zole-2-thiol (0.25 g, 1 mmol), NaOH (0.05 g, 1.2 mmol) and RCH2Cl (1.1 mmol)
were put into a pressure-rated vial (10 mL). Then, the mixture was reacted at 90 °C
for 15 min under a microwave synthesizer. After the mixture temperature was below
50 °C, it was poured into crushed ice, and the expected 1,3,4-oxadiazole was col-
lected and recrystallized.
2‑(Benzylthio)‑5‑(1,3‑dimethyl‑1H‑pyrazol‑4‑yl)‑1,3,4‑oxadiazole 5a
1
White solid, m.p. 82–85 °C, yield 28.0%, H NMR (500 MHz, CDCl3) δ: ppm
7.82 (s, 1H, pyrazole-H), 7.45–7.44(m, 2H, Ar–H), 7.36–7.33(m, 2H, Ar–H),
7.31–7.29(m, 1H, Ar–H), 4.59(s, 2H, –CH2–), 3.91(s, 3H, N-CH3), 2.49(s, 3H,
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W. Yu et al.
pyrazole-CH3); ESI-HRMS calcd for C14H14N4OS 287.0961, found 287.0961
[M+H]+.
2‑((4‑Bromobenzyl)thio)‑5‑(1,3‑dimethyl‑1H‑pyrazol‑4‑yl)‑1,3,4‑oxadiazole 5b
Light yellow solid, m.p. 94–98 °C, yield 70.1%, 1H NMR (500 MHz, CDCl3) δ: ppm
7.79 (s, 1H, pyrazole-H), 7.45 (d, J=8.3 Hz, 2H, Ar–H), 7.33 (d, J=8.3 Hz, 2H,
Ar–H), 4.41 (s, 2H, –CH2–), 3.88 (s, 3H, N-CH3), 2.51 (s, 3H, pyrazole-CH3); ESI-
HRMS calcd for C14H13BrN4OS 365.0066, found 365.0066 [M+H]+.
2‑((3,4‑Dichlorobenzyl)thio)‑5‑(1,3‑dimethyl‑1H‑pyrazol‑4‑yl)‑1,3,4‑oxadiazole 5c
White solid, m.p. 124–126 °C, yield 89.0%, 1H NMR (500 MHz, CDCl3) δ: ppm 7.82
(s, 1H, pyrazole-H), 7.58 (d, J=2.01 Hz, 1H, Ar–H), 7.42 (d, J=8.3 Hz, 1H, Ar–H),
7.34 (dd, J=8.26, 2.01 Hz, 1H, Ar–H), 4.43 (s, 2H, –CH2–), 3.91 (s, 3H, N-CH3),
2.53 (s, 3H, pyrazole-CH3); ESI-HRMS calcd for C14H12Cl2N4OS 355.0182, found
355.0178 [M+H]+.
2‑((2‑Chlorobenzyl)thio)‑5‑(1,3‑dimethyl‑1H‑pyrazol‑4‑yl)‑1,3,4‑oxadiazole 5d
1
White solid, m.p. 105–106 °C, yield 85.9%, H NMR (500 MHz, CDCl3) δ: ppm
7.82 (s, 1H, pyrazole-H), 7.64 (d, J=7.0 Hz, 1H, Ar–H), 7.42 (d, J=7.5 Hz, 1H,
Ar–H), 7.31–7.21 (m, 2H, Ar–H), 4.61 (s, 2H, –CH2–), 3.91 (s, 3H, N-CH3), 2.53 (s,
3H, pyrazole-CH3); ESI-HRMS calcd for C14H13ClN4OS 321.0571, found 321.0571
[M+H]+.
2‑((3‑Chlorobenzyl)thio)‑5‑(1,3‑dimethyl‑1H‑pyrazol‑4‑yl)‑1,3,4‑oxadiazole 5e
White solid, m.p. 70–71 °C, yield 75.0%, 1H NMR (500 MHz, CDCl3) δ: ppm 7.77
(s, 1H, pyrazole-H), 7.45–7.18 (m, 4H, Ar–H), 4.40 (s, 2H, –CH2–), 3.86 (s, 3H,
N-CH3), 2.48 (s, 3H, pyrazole-CH3); ESI-HRMS calcd for C14H13ClN4OS 321.0571,
found 321.0570 [M+H]+.
2‑((4‑Chlorobenzyl)thio)‑5‑(1,3‑dimethyl‑1H‑pyrazol‑4‑yl)‑1,3,4‑oxadiazole 5f
White solid, m.p. 88–89 °C, yield 67.2%, 1H NMR (500 MHz, CDCl3) δ: ppm 7.81
(s, 1H, pyrazole-H), 7.42 (d, J=8.1 Hz, 2H, Ar–H), 7.34–7.26 (m, 2H, Ar–H), 4.45
(s, 2H, –CH2–), 3.91 (s, 3H, N-CH3), 2.53 (s, 3H, pyrazole-CH3); ESI-HRMS calcd
for C14H13ClN4OS 321.0571, found 321.0569 [M+H]+.
2‑((2,4‑Dichlorobenzyl)thio)‑5‑(1,3‑dimethyl‑1H‑pyrazol‑4‑yl)‑1,3,4‑oxadiazole 5g
1
Light yellow solid, m.p. 109–110 °C, yield 94.6%, H NMR (500 MHz, CDCl3)
δ: ppm 7.80 (s, 1H, pyrazole-H), 7.60 (d, J=8.3 Hz, 1H, Ar–H)), 7.41 (s, 1H,
Ar–H)), 7.20 (d, J=7.3 Hz, 1H, Ar–H), 4.54 (s, 2H, –CH2–), 3.89 (s, 3H, N-CH3),
1 3

Synthesis and biological activity of novel 1,3,4-oxadiazole…
2.51 (s, 3H, pyrazole-CH3); ESI-HRMS calcd for C14H12Cl2N4OS 355.0182, found
355.0182 [M+H]+.
2‑(((6‑Chloropyridin‑3‑yl)methyl)
thio)‑5‑(1,3‑dimethyl‑1H‑pyrazol‑4‑yl)‑1,3,4‑oxadiazole 5h
Light yellow solid, m.p. 130–132 °C, yield 31.2%, 1H NMR (500 MHz, CDCl3) δ: ppm
8.50 (s, 1H, pyrazole-H), 7.90–7.79 (m, 2H, pyridine-H), 7.32 (d, J=8.2 Hz, 1H, pyr-
idine-H), 4.45 (s, 2H, –CH2–), 3.91 (s, 3H, N-CH3), 2.53 (s, 3H, pyrazole-CH3); ESI-
HRMS calcd for C14H14N4OS 322.0524, found 322.0518 [M+H]+.
2‑(1,3‑Dimethyl‑1H‑pyrazol‑4‑yl)‑5‑((2‑fuorobenzyl)thio)‑1,3,4‑oxadiazole 5i
White solid, m.p. 120–123 °C, yield 82.2%, 1H NMR (500 MHz, CDCl3) δ: ppm 7.82
(s, 1H, pyrazole-H), 7.58–7.53 (m, 1H, Ar–H), 7.34–7.27 (m, 1H, Ar–H), 7.14–7.05
(m, 2H, Ar–H), 4.52 (s, 2H, –CH2–), 3.91 (s, 3H, N-CH3), 2.53 (s, 3H, pyrazole-CH3);
ESI-HRMS calcd for C14H13FN4OS 305.0867, found 305.0868 [M+H]+.
4‑(((5‑(1,3‑Dimethyl‑1H‑pyrazol‑4‑yl)‑1,3,4‑oxadiazol‑2‑yl)thio)methyl)benzonitrile 5j
Light yellow solid, m.p. 118–119 °C, yield 72.3%, 1H NMR (500 MHz, CDCl3) δ: ppm
7.81 (s, 1H, pyrazole-H), 7.68–7.58 (m, 4H, Ar–H), 4.50 (s, 2H, –CH2–), 3.90 (s, 3H,
N-CH3), 2.51 (s, 3H, pyrazole-CH3); ESI-HRMS calcd for C14H14N4OS 312.0914,
found 312.0914 [M+H]+.
2‑((5‑(1,3‑Dimethyl‑1H‑pyrazol‑4‑yl)‑1,3,4‑oxadiazol‑2‑yl)thio)acetonitrile 5k
Light yellow solid, m.p. 158–160 °C, yield 27.7%, 1H NMR (500 MHz, CDCl3) δ: ppm
7.88 (s, 1H, pyrazole-H), 4.09 (s, 2H, –CH2–), 3.93 (s, 3H, N-CH3), 2.56 (s, 3H, pyra-
zole-CH3); ESI-HRMS calcd for C15H13N5OS 236.0601, found 236.0600 [M+H]+.
2‑(((2‑Chlorothiazol‑5‑yl)methyl)
thio)‑5‑(1,3‑dimethyl‑1H‑pyrazol‑4‑yl)‑1,3,4‑oxadiazole 5l
1
Brown solid, m.p. 115–117 °C, yield 47.4%, H NMR (500 MHz, CDCl3) δ: ppm
7.84 (s, 1H, pyrazole-H), 7.57 (s, 1H, thiazole-H), 4.63 (s, 2H, –CH2–), 3.92 (s, 3H,
N-CH3), 2.55 (s, 3H, pyrazole-CH3); ESI-HRMS calcd for C11H10ClN5OS2 328.0088,
found 328.0088 [M+H]+.
Fungicidal activity
Fungal pathogen production
Isolates of Colletotrichum acutatum Simmonds, C. fragariae Brooks, C. gloeospori-
oides (Penz.) Penz. & Sacc. in Penz. were obtained from B. J. Smith, USDA–ARS,
1 3

W. Yu et al.
Small Fruit Research Station, Poplarville, MS. The three Colletotrichum species were
isolated from strawberry (Fragaria×ananassa Duchesne).
Bioautography assay
Direct bioautography is a technique to screen large numbers of crude extracts, or
pure compounds. Bioautography procedures were described in our previous studies
[56, 57]. Pure compounds were evaluated for antifungal activity against strawberry
anthracnose-causing plant pathogens, Colletotrichum acutatum, C. fragariae and C.
gloeosporioides using the direct overlay bioautography. Technical grade commer-
cial fungicide standards benomyl, cyprodinil, azoxystrobin and captan (with difer-
ent modes of action) were used at 0.9–1.61 µg/µL concentrations in 95% ethanol.
After sample application, each TLC plate was subsequently sprayed with a spore
suspension (3.0×105 spores/mL) of the fungus of interest and incubated in a mois-
ture chamber for 4 days at 26 °C with a 12-h photoperiod. Clear zones of fungal
growth inhibition on the TLC plate indicate the presence of antifungal constituents
in each extract or of the pure compounds.
Micro‑dilution antifungal assay
A standardized 96-well micro-dilution broth assay developed by Wedge and Kahajek
[58] was used to evaluate antifungal activity of pure compounds from extracts that
were identifed as active by bioautography [59]. This assay was used to evaluate anti-
fungal activity of pure compounds toward Colletotrichum acutatum Simmonds, C.
fragariae Brooks, C. gloeosporioides (Penz.) Penz. & Sacc. in Penz., in comparison
with known fungicide standards [60–62]. Commercial, technical grade azoxystrobin
and captan (without formulation) were used as validation controls in all micro-dilu-
tion broth assays. Each fungus was treated with 50, 75 and 150 µM of each com-
pound. Captan and azoxystrobin standards were run at 0.3, 3.0 and 30.0 µM. Micro-
titer plates (Nunc MicroWell, untreated; Roskilde, Denmark) were covered with a
plastic lid and incubated in a growth chamber as described previously for fungal
growth. Fungal growth was then evaluated by measuring absorbance of each well at
620 nm using a microplate photometer (Packard Spectra Count, Packard Instrument
Co., Downers Grove, IL). Mean absorbance values with standard errors were used to
evaluate fungal growth at 48 and 72 h. Means for percent inhibition/stimulation of
each fungus at each dose of test compound relative to the untreated positive growth
controls were used to evaluate fungal growth. The SAS Proc ANOVA was used to
identify signifcant factors, and Fisher’s protected LSD was used to separate means.
All experiments were repeated at least once in time.
Herbicidal activity
The method of Dayan et al. [63] was used for bioassay of a monocot and dicot plant
species. Bioassays were done in duplicate in sterile non-pyrogenic polystyrene
24-well cell culture plates (CoStar 3524, Corning Incorporated). One flter paper
1 3

Synthesis and biological activity of novel 1,3,4-oxadiazole…
disk (Whatman Grade 1, 1.5 cm) was placed in each well. The control wells con-
tained 200 µL of distilled water. The control+solvent well contained 180 µL of
water and 20 µL of the solvent. All sample wells contained 180 µL of water and
20 mL of the appropriate dilution of the sample. Water was always pipetted into the
well before the sample or solvent. All plate preparation was done in a sterile envi-
ronment to reduce chances of microbial contamination.
For lettuce (Lactuca sativa L.cv. Iceberg A Crisphead; Burpee seeds, W. Atlee
Burpee & Co., Warminster, PA, USA), fve seeds were placed in each well, and
20–30 seeds per well of creeping bentgrass (Agrostis stolonifera L. cv. Penncross;
Turf Seed Inc., Hubbard, OR, USA) were used. Lids were sealed with Paraflm. The
plates were incubated in a Percival Scientifc CU-36L5 incubator under continuous
light conditions at 26 °C and 120 µmol/s m2 photon fux of photosynthetically active
radiation average light intensity. Plates were incubated for at least 7 days. Ranking
of plant growth was subjective.
On a scale of 0–5, a ranking of 0 indicated no apparent inhibition (sample well
plants looked identical to the control+solvent well plants). A ranking of 5 indi-
cated no growth or complete inhibition. A ranking of 5 was given only if no seeds
germinated.
Results and discussion
Synthesis and spectra
The studied 1,3,4-oxadiazole derivatives are outlined in Scheme 1. The key inter-
mediate pyrazole ring was synthesized using ethyl 2-(ethoxymethylene)-3-oxob-
utanoate and CH3NHNH2 according to the reference. Also, we also successfully
used NH2NH2·H2O instead of CH3NHNH2 to synthesize the pyrazole ring. Unfor-
tunately, when we used CH3I as the methylation reagent to synthesize intermediate
2, it did not work well. The pyrazole carbohydrazide was prepared from a pyrazole
ester and NH2NH2·H2O. The 1,3,4-oxadiazole ring was cyclized using pyrazole car-
bohydrazide in CS2 and KOH under refux conditions with high yield. Lastly, the
Scheme 1 Synthetic route of compounds 5a–5l
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W. Yu et al.
1,3,4-oxadiazole thione reacted with diferent RCH2Cl compounds in water under
microwave irradiation, and the fnal products were easily produced. Compounds
5a–5l had 28–89% yields.
1
All the pyrazole compounds were confrmed by H-NMR and HRMS. From the
1H-NMR results, the SCH2 single was observed at 4.09–4.63 ppm, and the CH pro-
ton signal of pyrazole ring was found around 7.8 ppm as a single peak. The two
methyl groups on the pyrazole ring appeared at 3.9 and 2.5 ppm, respectively. Mean-
while, all the pyrazole derivatives showed the M+H+ peak in the HRMS data.
Antifungal activity
The bioautography assay results are listed in Table 1. The micro-dilution broth assay
of test compounds demonstrated that compound 5b was the most active against C.
acutatum (Fig. 2). However, inhibitory activity of this compound decreased between
48 and 72 h 45% to 26%, respectively (72 h data not shown). Loss of fungal inhibi-
tory activity over time is usually due to compound instability in the test solution or
to metabolic degradation within the fungus. At the highest concentration of 150 µM,
none of the test compounds possessed antifungal activity near that of azoxystrobin
or captan standards that were tested at lower concentrations. As seen for compound
5l, growth stimulation by fungicides at subtoxic concentrations (hormesis) in fungi
is common [64]. Compounds 5b, 5c and 5f were the most active analogs against
C. gloeosporioides. Compound 5f was the most active analog against C. fragariae;
however, it only produced 28% inhibition. The structure–activity relationship of
tested compounds indicated that when the halogen was substituted on the para- or
meta-position of the benzene ring, the compounds had good antifungal activity, such
as compounds 5b, 5c, 5e and 5f. Also, if there was no substitution on the benzene
ring, the activity was still good.
Phytotoxicity
The herbicidal activity of the 1,3,4-oxadiazole compounds is listed in Table 1. As
shown in Table 2, most of the 1,3,4-oxadiazole compounds had varying degrees
of phytotoxicity against lettuce and Agrostis at 1 mM, except compound 5c. Com-
pound 5c was not phytotoxic against lettuce and bentgrass at 1 mM. All the active
compounds were more phytotoxic to Agrostis than to lettuce, with 5k showing high
phytotoxicity. The most herbicidal compound 5k exhibited the same activity against
bentgrass as control aminotriazole (ranking 5), but it had lower activity (ranking 3)
against lettuce than that of aminotriazole (ranking 4). Compounds 5b, 5h, 5i, 5j and
5l also had good herbicidal activity against bentgrass (ranking 4). Only compounds
1 3

Synthesis and biological activity of novel 1,3,4-oxadiazole…
A. Colletotrichum acutatum
100.0%
50 µM
75 µM
150 µM
80.0%
60.0%
40.0%
15.2%
20.0%
0.0%
(20.0%)
(13.7%)
(40.0%)
(31.4%)
(36.1%)
(44.6%)
(60.0%)
(80.0%)
(87.3%)
(88.9%)
(100.0%)
5a
5b
5c
5e
5f
Azoxystrobin Captan
B. Colletotrichum fragariae
100.0%
50 µM
75 µM
150 µM
80.0%
60.0%
40.0%
20.0%
5.1%
0.0%
(20.0%)
(15.2%)
(24.3%)
(27.7%)
(28.4%)
(40.0%)
(60.0%)
(80.0%)
(100.0%)
(70.3%)
(88.2%)
5a
5b
5c
5e
5f
Azoxystrobin Captan
C. Colletotrichum gloeosporoides
100.0%
50 µM
75 µM
150 µM
80.0%
60.0%
40.0%
20.0%
0.0%
(20.0%)
(18.6%)
(40.0%)
(37.8%)
(60.0%)
(52.8%)
(58.5%)
(62.2%)
(80.0%)
(100.0%)
(93.4%)
(96.4%)
5a
5b
5c
5e
5f
Azoxystrobin Captan
Fig. 2 Micro-dilution broth assay of fve compounds against three Colletotrichum species
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W. Yu et al.
Table 1 Antifungal activity of
fve pyrazole derivatives against
Colletotrichum test species
using direct bioautography at
12 mM with 8 µL
Compounds
Mean zone diameter of fungal growth inhibi-
tion in mm
C. acutatum
C. fragariae C. gloe-
ospori-
oides
5a
13
6
9
15
Difuse
Difuse
12
Difuse
Difuse
6
5
6
12
Difuse
Difuse
15
Difuse
21
Difuse
Difuse
Difuse
10
Difuse
Difuse
16
5b
5c
5e
5f
Benomyl*
Captan*
Cyprodinil*
Azoxystrobin*
Difuse
17
* is the note the control concentration is 2 mM and applied 2 µL
Table 2 Phytotoxicity of title
No.
R
Lettuce
Agrostis
compounds at 1 mM
5a
Ph
2
1
0
3
2
2
0
4
4
1
3
3
4
3
4
0
3
3
3
2
4
4
4
5
4
5
5b
4-BrPh
3,4-Cl2Ph
2-ClPh
3-ClPh
4-ClPh
2,4-Cl2Ph
2-ClPy
2-FPh
5c
5d
5e
5f
5g
5h
5i
5j
4-CNPh
CN
2-Cl-thiazole
5k
5l
Aminotriazole
0=no efect, 5=no growth
5d, 5e and 5f displayed moderate herbicidal activity against bentgrass (ranking 3).
For lettuce, only compounds 5h and 5i exhibited good herbicidal activity (ranking
4), which is the same as the positive control aminotriazole (ranking 4). Comparing
herbicidal activity among 5a–5l in Table 2, the compounds with electron withdraw-
ing group at para-position of benzene ring exhibited good herbicidal activity against
bentgrass. The compound with a cyanide group had the best activity against the two
species. When the R is a heterocycle, the herbicidal activity is higher than that of
phenyl compounds, such as 5h and 5l.
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Synthesis and biological activity of novel 1,3,4-oxadiazole…
Conclusions
Twelve new 1,3,4-oxadiazole derivatives containing a pyrazole moiety were eval-
uated for fungicidal activity. Low levels of both activities were found, but none of
the activities approached those of commercial fungicides. All of the tested com-
pounds had little-to-moderate growth inhibition on dicots and major activity on
monocots. The complete lack of phytotoxicity of 5c coupled with moderate fun-
gicidal activity indicates that it could be applied directly to plants for control of
some diseases. The antifungal activities reported here are sufcient for further
structure manipulation to improve the antifungal activity.
Acknowledgements This work was funded by Zhejiang Provincial Natural Science Foundation of China
(No. LY19C140002), Research Fund of Department of Education of Zhejiang Province (Y201941832)
and the Opening Foundation of the Key Laboratory of Green Pesticide and Agricultural Bioengineering,
Ministry of Education, Guizhou University, Grant No. 2018GDGP0104.
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