Synthesis of 1,2,3-triazole hydrazide derivatives exhibiting anti-phytopathogenic activity

Article abstract of DOI:10.1016/j.ejmech.2016.10.006

A series of new 1,2,3-triazole derivatives have been prepared and screened for their antifungal activity against phytopathogenic fungi using the mycelium growth inhibition method in?vitro. The results indicated that the 1,2,3-triazole hydrazide scaffold d

Full text of DOI:10.1016/j.ejmech.2016.10.006

Accepted Manuscript
Synthesis of 1,2,3-triazole hydrazide derivatives exhibiting anti-phytopathogenic
activity
Xing Wang, Zhi-Cheng Dai, Yong-Fei Chen, Ling-Ling Cao, Wei Yan, Sheng-Kun Li,
Jian-Xin Wang, Zheng-Guang Zhang, Yong-Hao Ye
PII:
S0223-5234(16)30868-6
10.1016/j.ejmech.2016.10.006
EJMECH 8971
DOI:
Reference:
To appear in: European Journal of Medicinal Chemistry
Received Date: 12 May 2016
Revised Date: 27 September 2016
Accepted Date: 3 October 2016
Please cite this article as: X. Wang, Z.-C. Dai, Y.-F. Chen, L.-L. Cao, W. Yan, S.-K. Li, J.-X. Wang, Z.-
G. Zhang, Y.-H. Ye, Synthesis of 1,2,3-triazole hydrazide derivatives exhibiting anti-phytopathogenic
activity, European Journal of Medicinal Chemistry (2016), doi: 10.1016/j.ejmech.2016.10.006.
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Graphic Abstract

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Synthesis of 1,2,3-triazole hydrazide derivatives exhibiting anti-phytopathogenic
activity
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Xing Wang , Zhi-Cheng Dai , Yong-Fei Chen , Ling-Ling Cao , Wei Yan ,
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Sheng-Kun Li , Jian-Xin Wang , Zheng-Guang Zhang , Yong-Hao Ye
1. College of Plant Protection, State & Local Joint Engineering Research Center of
Green Pesticide Invention and Application, Nanjing Agricultural University, Nanjing
210095, P. R. China.
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2. Key Laboratory of Integrated Management of Crop Diseases and Pests, Ministry of
Education, Nanjing 210095, P. R. China
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X. Wang and Z. C. Dai contributed equally to this paper.
* Corresponding author. Tel.: +86-25-84399753. Fax: +86-25-84399753. E-mail
address: yeyh@njau.edu.cn

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Abstract:
A series of new 1,2,3-triazole derivatives have been prepared and screened for their
antifungal activity against phytopathogenic fungi using the mycelium growth
inhibition method in vitro. The results indicated that the 1,2,3-triazole hydrazide
scaffold displayed significant antifungal activity. Compound 6ad exhibited the most
potent anti-phytopathogenic activity, with EC values of 0.18, 0.35, 0.37 and 2.25
5
0
-1
µg·mL against Rhizoctonia solani, Sclerotinia sclerotiorum, Fusarium graminearum,
and Magnaporthe oryzae, respectively. In vivo testing demonstrated that 6ad was
effective for the control of rice sheath blight, rape sclerotinia rot, fusarium head blight
and rice blast caused by the aforementioned phytopathogens. This work suggests that
the combination of 1,2,3-triazole and hydrazide moiety could be a promising
fungicide scaffold in the future.
KEY WORDS: 1,2,3-triazole; hydrazide; antifungal activity; fungicide

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1. Introduction
Plant diseases, such as rice sheath blight (RSB), rape sclerotinia rot (RSR),
fusarium head blight (FHB) and rice blast (RB), result in severe yield losses and
decreases in the quality of agricultural products around the world[1]. With the
increasing use of pesticides, the emergence of pesticide resistance and pollution
problems associated with conventional agrochemicals, there is an urgent need to
develop novel agrochemicals with high activity, good selectivity and eco-friendly
properties.
1,2,3-Triazoles have been widely studied due to their broad spectrum of
biological activities, which include anti-bacterial[2], anti-tubercular[3],
anti-neoplastic[4], anti-HIV[5], and antifungal[6] activities. It is synthesized via click
chemistry with the Huisgen 1,3-dipolar cycloaddition reaction as the key step[7], in
which Cu(I) accelerates the reaction rate dramatically and makes the reaction highly
regioselective[8]. Our previous work showed that 1,2,3-triazole phenylhydrazone
derivatives displayed significant antifungal activity[9]. The 3D-QSAR model
indicated that halogen substitution of R at an ortho position showed advantage over
1
substitution at meta or para positions (Fig. 1).
In addition, molecules with acylhydrazone, oxime ether and hydrazide scaffolds
exhibit a broad range of biological and pharmacological activities[10-14] and have
been widely used as frameworks in drug design. For example, the acylhydrazone
derivative benquinox[15] showed moderate efficacy against Pythium aphanidermatum;
the oxime ether derivative pyrifenox[16] had special effects against pathogenic

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bacteria on fruits and vegetables; the hydrazide derivative daminozide[17] was used
for plant growth inhibition; and the hydrazide derivative RH-5992[18] exhibited
moderate insecticidal activity against cotton bollworm (Fig. 2).
Keeping the biological importance of acylhydrazone, oxime ether, hydrazide and
1,2,3-triazole derivatives in mind, we designed and synthesized target compounds 6a,
7a, and 9a as part of our continuing research on new antifungal development (Fig. 1).
Antifungal activity tests demonstrated that 6a performed better than 7a and 9a.
Consequently, a series of 1,2,3-triazole hydrazide derivatives 6ab-6fg were
synthesized and evaluated in vitro and in vivo for their antifungal activities against
four important phytopathogenic fungi (R. solani, S. sclerotiorum, F. graminearum,
and M. oryzae). Systemic translocation of the most active compound in wheat leaves
was also evaluated.
2. Results and discussion
2.1. Synthetic chemistry
The synthesis of the 1, 2, 3-triazole derivatives is outlined in Scheme 1. Different
aromatic amines (1a-f) were diazotized by sodium nitrite and were subsequently
turned into azides by sodium azide with yields ranging from 60% to 90%. Then, the
mixture was extracted with ethyl acetate and the organic phase was washed with
diluted hydrochloric acid for three times to remove residual phenylamine. Aromatic
azides (2a-f) were employed in 1,3-dipolar cycloaddition with propargyl alcohol,
which was catalyzed by Cu(I). Copper sulfate and sodium ascorbate guided
regioselectivity to obtain 1,4-disubstituted 1,2,3-triazoles at yields of 60-80%.

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1,2,3-Triazole alcohols (3a-f) were further oxidized to aldehyde (4a) and acids
(5a-f). The compounds (5a-f) were first transformed into triazole chlorides in the
presence of SOCl and then reacted with and appropriate phenylhydrazine to generate
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the target compounds 6a-6fg at yields of 62-83%.
Compound 7a was synthesized with an excellent yield through the reaction of
aldehyde (4a) with the corresponding benzoyl hydrazine in refluxing ethanol with 95%
yields.
1,2,3-Triazole aldehyde (4a) was transformed to the corresponding oxime (8a)
with hydroxylamine hydrochloride by refluxing the compound under ethanol (90%
yield), then stirred with benzyl bromide in DMF at r.t. to create the target compound
9a with a 93% yield.
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The results of H-NMR spectroscopy, ESI-MS spectra and elemental analysis
data of the target compounds reflected the assigned structures of the compounds.
2.2. Antifungal activities in vitro
The compound concentration for initial antifungal activity screening in vitro was
-1
set as 25 µg·mL . Various concentrations of the compounds were used to calculate
their median effective concentration (EC ) values and determine if the inhibitory rate
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was greater than 50%. Validamycin A and carbendazim were co-assayed as positive
controls. The results are shown in Table 1. Hydrazone derivative 7a and oxime ether
derivative 9a showed weak activity against the four tested strains. Hydrazide
derivatives 6a-6fg exhibited high antifungal effects, especially against R. solani, with
-1
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00
26 compounds displaying potent activity with EC values lower than 1 µg·mL . The
50

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EC value for carbendazim was 1.32 µg·mL . Compound 6ad had the lowest EC
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for R. solani at 0.18 µg·mL , whereas 6fd against S. sclerotiorum had an EC of 0.19
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µg·mL (carbendazim was 0.12 µg·mL ), 6fg against F. graminearum had an EC of
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0.36 µg·mL (carbendazim is 0.60 µg·mL ), 6fe against M. oryzae had an EC of
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0.56 µg·mL (carbendazim is 0.29µg·mL ). Compared comprehensively, we chose
6ad as the most potent compound, which inhibited the mycelia growth of S.
sclerotiorum, F. graminearum and M. oryzae with EC values of 0.35, 0.37 and 2.25
50
-1
µg·mL , respectively.
A series of hydrazide derivatives was determined to be synthesized by comparing
the antifungal activities of 6a, 7a and 9a. At first, the substituents of R were fixed at
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o-Cl, m-Cl, p-Cl to synthesize compounds 6ab-6ed, and the EC50 values of these
compounds indicated that substituents of R at the ortho position, especially o-Cl
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(6ab-6ad), always showed an advantage over those at the meta or para positions.
Subsequently, the substituents of R were fixed at halogen substituents Cl and F to
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synthesize compounds 6ae-6fg. The EC50 values indicated that an electron
withdrawing group for R showed an advantage over an electron-donating group.
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When R was a para substituent, the compounds were more beneficial than the ortho
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or meta positions (the results for M. oryzae were not as obvious compared with the
other three pathogens). The conclusions were reinforced by the predicted result of
3D-QSAR of 1,2,3-triazole phenylhydrazone derivatives in our previous work[9]
.
In general, the structure activity relationships of these compounds can be
summarized: halogen substituents of R at the ortho position and halogen substituents
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of R at the para substituent (6ad, 6ag, 6fd and 6fg) formed the optimal
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conformations. Due to the higher price of fluoro-substituted aniline compared to
chlorine-substituted aniline, 6ad was chosen to test antifungal activity in vivo despite
the similar antifungal activity produced by other compounds.
2.3. Antifungal activities in vivo
2.3.1. Protective activity of 6ad against rice sheath blight
6ad was selected to evaluate the protective activity against R. solani under
greenhouse conditions. The results are shown in Fig. 3 and Table 2. The negative
control, a brown disease spot, was observed on the rice sheath seven days after
inoculating the cultivars with R. solani and the lesion length reached 16.4 mm. At a
-1
concentration of 200 µg·mL , the protective effect of 6ad in vivo could reach 60.4%,
which was comparable to validamycin A (64.6%). When the concentration to 100
-1
µg·mL , the protective effect of 6ad (55.9%) was significantly higher than
validamycin A (43.3%).
2.3.2. Protective activity of 6ad against rape sclerotinia rot
The efficacy of the treatment is shown in Fig. 4. The untreated negative control
(pathogen only) resulted in 100% disease incidence (0% healthy plant standard) 24 h
-1
after transplantation. Treatment with 500 µg·mL 6ad resulted in 52.9% healthy plant
standard after 36 h of treatment (Table 3), whereas carbendazim treatment resulted in
68.1%. Therefore, compound 6ad not only significantly inhibited the mycelia growth
of S. sclerotiorum in vitro but also successfully suppressed disease development in S.
sclerotiorum infected cole in vivo.

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2.3.3. Therapeutic activity of 6ad against fusarium head blight
The efficacy of the therapeutic activity experiment two weeks after spraying
compound 6ad is shown in Table 4. The untreated negative control resulted in 100%
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FHB disease incidence, whereas 250 µg·mL 6ad and carbendazim resulted in 47.4%
and 61.7% respectively (Fig. 5). The result indicated that 6ad had a certain
therapeutic activity against FHB.
2.3.4. Protective activity against rice blast and inhibition of conidium germination by
6ad
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In vivo, 6ad at 100 µg mL could control the development of RB in a similar
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manner to 50 µg mL carbendazim. The level of disease on rice leaves is shown Fig.
6A, and the efficacy of the treatment is shown in Table 4. Conidia of M. oryza could
not form the germ tube and appressorium germinate in the 6ad solution at a
-1
concentration of 1.0 µg mL (Fig. 6B). Therefore, 6ad showed protective activity
against RB.
2.4. Systemic translocation of 6ad in wheat
The effect of 6ad on the treatment areas of wheat leaves was 100% (Table 6). At
-1
the terminal parts of the treated leaves, the control effect of 200 µg mL 6ad reached
75.4%, which was comparable to the control effect of triadimefon at 97.1%. At the
stem base parts of the treated leaves, 6ad and triadimefon showed a poor effect
against powdery mildew. At the back of the treated leaves, the control effect of 200 µg
-1
mL 6ad reached 66.0%, which is comparable to triadimefon at 86.1%. The result
indicated that 6ad had the effect of upward translocation and osmosis as well as no

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downward translocation effect (Fig. 7).
3. Conclusions
In this study, new hydrazide derivatives of 1,2,3-triazole were synthesized and
investigated for the antifungal activities against R. solani, S. sclerotiorum, F.
graminearum, and M. oryzae. The results showed that all these target compounds
displayed significant antifungal activity. Among them, compound 6ad exhibited the
most potent anti-phytopathogenic activity with EC values of 0.18, 0.35, 0.37 and
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-1
2.25 µg mL against four fungi respectively. In vivo testing also demonstrated that
6ad was effective for the control of rice sheath blight, rape sclerotinia rot, fusarium
head blight and rice blast caused by the aforementioned phytopathogens. Due to the
excellent activities, 6ad can be considered as a promising fungicide in the future. The
mechanism of 1,2,3-triazole hydrazide derivatives is unreported at present, but some
attempts had been made, such as the effect of triazole-hydrazine derivative on
respiration system of mycelium was tested. The result showed that compounds had
inhibitory active on respiration system (data not shown). Our follow-up studies may
clarify the mechanism.
4. Materials and methods
All chemicals and reagents were analytical grade (provided by Shanghai crystal
pure biological technology Co. Ltd.). The reactions were monitored by TLC using
pre-coated silica gel GF254 plates from Qingdao Haiyang Chemical, China. UV
detections were conducted at a wavelength of 254 nm. Melting points (uncorrected)
were measured with a SPSIC WRS-1B digital melting-point apparatus (Shanghai,

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China). HR-ESI-MS spectra were recorded on a Brucker UHR-TOF maxis mass
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spectrometer. H-NMR spectra were collected on a Bruker Avance III 400 NMR
spectrometer at room temperature using DMSO-d as a solvent. The chemical shifts (δ)
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were reported in ppm with reference to internal TMS, and coupling constants (J) were
given in Hz. Elemental analyses were performed on a CHN-O-Rapid instrument and
were within ± 0.4 % of the theoretical values.
4.1. General procedure for the synthesis of compounds 2a-f
Substituted aniline (20 mmol) was added to a stirred solution of HCl (3N, 20 mL)
in an ice bath. NaNO (30 mmol) dissolved in 30 mL water was added dropwise. The
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mixture was stirred for 30 min. Sodium azide (40 mmol) dissolved in 50 mL water
was added dropwise. Then, the system was stirred for another 3 h at r.t.[19]. The
mixture was extracted with ethyl acetate and the organic phase was washed with
diluted hydrochloric acid for three times to remove residual phenylamine and
concentrated in vacuo.
4.2. General procedure for the synthesis of compounds 3a-f
In a round-bottom flask equipped with a magnetic stirring bar, 2a-f (10 mmol)
was added to a solvent of tert-butanol/H O (1:1) along with propargyl alcohol (15
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mmol), CuSO pentahydrate (0.4 mmol), and sodium ascorbate (1 mmol) [20-22]. The
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reaction mixture was stirred for 48 h at r.t. and subsequently extracted with ethyl
acetate for three times. The organic phase was washed with saturated salt water, dried
over anhydrous Na SO , filtered, and concentrated in vacuo. Further purification was
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performed with silica gel column chromatography using petroleum ether/EtOAc (1:2)

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as the eluent to obtain the pure derivative 3a-f (in yields of 60-80%).
4.3. General procedure for the synthesis of compounds 5a-f
A stirring mixture of 3a-f (10.0 mmol) and Na CO (6 mmol) was dissolved in
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H O, and then the mixture was heated to 80-85 ºC for 10 min. Afterwards, KMnO4
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(12 mmol) was added slowly[23]. The reaction was monitored with thin layer
chromatography (TLC). After the reaction finished, the solution was filtered, and the
aqueous solution was acidified with 2N HCl (pH=2) to precipitate the desired
compounds, which were collected by filtration, washed with cold water, and allowed
to air-dry overnight to generate 5a-f (in yields of 90- 95%) as a white solid.
4.4. General procedure for the synthesis of compounds 6a-f
A mixture of compound 5a-f (1 mmol) and SOCl (10 mL) was stirred and
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refluxed for 2 h. Remaining SOCl was removed by distillation under reduced
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pressure. After SOCl was completely removed, the residue was directly dissolved in
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anhydrous CH Cl (20 mL). The synthesis of substituted phenylhydrazines was
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performed according to protocols from the literature[24]. Appropriate substituted
phenylhydrazine (1 mmol) and TEA (2.5 mmol) was added dropwise, and the mixture
was stirred at r.t. for 1 h[25].
The reaction mixture was washed with saturated salt water several times. The
CH Cl solution was collected and dried with anhydrous Na SO . Further purification
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was carried out with silica gel column chromatography using petroleum ether/EtOAc
(2:1) as the eluent. The final compounds 6a and 6ab-6fg were obtained.
4.4.1 1-(2-chlorophenyl)-N'-phenyl-1H-1,2,3-triazole-4-carbohydrazide 6a

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Light yellow powder. Yield, 76.3%; mp, 193–194 ºC. H NMR (400 MHz, DMSO-d )
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δ 10.61 (s, 1H, -CONH-), 9.13 (s, 1H, -C N H-), 8.01 (s, 1H, -NH-), 7.81 (dd, J =
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11.3, 8.0 Hz, 2H, -C H Cl), 7.70 (t, J = 7.7 Hz, 1H, -C H Cl), 7.64 (t, J = 7.6 Hz, 1H,
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-C H Cl), 7.17 (t, J = 7.7 Hz, 2H, -C H ), 6.79 (d, J = 8.1 Hz, 2H, -C H ), 6.73 (t, J =
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7.2 Hz, 1H, -C H ). HRMS Calcd. for 336.0628 (C H ClN NaO, [M+Na] ), found
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15 12
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336.0623. Anal. Calcd. For C H ClN O: C, 57.42; H, 3.86; N, 22.32. Found: C,
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57.37; H, 3.65; N, 22.12.
4.4.2 N',1-bis(2-chlorophenyl)-1H-1,2,3-triazole-4-carbohydrazide 6ab
1
White powder, Yield, 83.0%; mp, 179–180 ºC. H NMR (400 MHz, DMSO-d ) δ
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10.74 (d, J = 2.0 Hz, 1H, -CONH-), 9.17 (d, J = 8.4 Hz, 1H, -C N H-), 7.83 (dd, J =
2
3
8.0, 1.3 Hz, 1H, -C H Cl), 7.80 (dd, J = 7.7, 1.7 Hz, 1H, -C H Cl), 7.73 (d, J = 1.8 Hz,
6
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6
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1H, -NH-), 7.71–7.61 (m, 2H, -C H Cl), 7.33 (dd, J = 7.9, 1.2 Hz, 1H, -C H Cl), 7.18
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(t, J = 7.7 Hz, 1H, -C H Cl), 6.86–6.75 (m, 2H, -C H Cl). HRMS Calcd. for
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+
370.0238 (C H Cl N NaO, [M+Na] ), found 370.0233. Anal. Calcd. For
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C H Cl N O: C, 51.74; H, 3.18; N, 20.11. Found: C, 51.57; H, 3.05; N, 20.24.
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4.4.3.
1-(2-chlorophenyl)-N'-(3-chlorophenyl)-1H-1,2,3-triazole-4-carbohydrazide
6ac
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Yellow powder, Yield, 70.0%; mp, 168–169 ºC. H NMR (400 MHz, DMSO-d ) δ
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10.69 (d, J = 2.3 Hz, 1H, -CONH-), 9.15 (s, 1H, -C N H-), 8.31 (d, J = 2.3 Hz, 1H,
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-NH-), 7.84–7.78 (m, 2H, -C H Cl), 7.70–7.66 (m, 2H, -C H Cl), 7.19 (t, J = 8.2 Hz,
6
4
6
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1H, -C H Cl), 6.75 (dd, J = 6.9, 2.8 Hz, 3H, -C H Cl). HRMS Calcd for 370.0238
6
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(C H Cl N NaO, [M+Na] ), found 370.0233. Anal. Calcd. For C H Cl N O: C,
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51.74; H, 3.18; N, 20.11. Found: C, 51.45; H, 3.25; N, 20.01.
4.4.4.
1-(2-chlorophenyl)-N'-(4-chlorophenyl)-1H-1,2,3-triazole-4-carbohydrazide
6ad
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Yellow powder, Yield, 71.6%; mp, 161–162 ºC. H NMR (400 MHz, DMSO-d ) δ
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10.67 (d, J = 2.3 Hz, 1H, -CONH-), 9.14 (s, 1H, -C N H-), 8.20 (d, J = 2.3 Hz, 1H,
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-NH-),7.82 (dd, J = 8.0, 1.3 Hz, 1H, -C H Cl), 7.79 (dd, J = 7.7, 1.6 Hz, 1H, -C H Cl),
6
4
6
4
7.69 (td, J = 7.8, 1.7 Hz, 1H, -C H Cl), 7.63 (td, J = 7.6, 1.4 Hz, 1H, -C H Cl), 7.20 (t,
6
4
6
4
J =6.0 Hz, 2H, -C H Cl), 6.82–6.74 (m, 2H, -C H Cl). HRMS Calcd for 370.0238
6
4
6
4
+
(C H Cl N NaO, [M+Na] ), found 370.0233. Anal. Calcd. For C H Cl N O: C,
1
5
11
2
5
15 11
2
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51.74; H, 3.18; N, 20.11. Found: C, 51.60; H, 3.11; N, 20.25.
4.4.5. N'-(2-chlorophenyl)-1-(3- chlorophenyl)-1H-1,2,3-triazole-4-carbohydrazide
6bb
1
Light yellow powder, Yield, 80.3%; mp, 205–206 ºC. H NMR (400 MHz, DMSO-d )
6
δ 10.73 (d, J = 1.9 Hz, 1H, -CONH-), 9.47 (s, 1H, -C N H-), 8.15 (t, J = 1.8 Hz, 1H,
2
3
-NH-), 8.01 (d, J = 7.9 Hz, 1H, -C H Cl), 7.73 (d, J = 1.8 Hz, 1H, -C H Cl), 7.70–
6
4
6
4
7.60 (m, 2H, -C H Cl), 7.32 (d, J = 7.8 Hz, 1H, -C H Cl), 7.16 (t, J = 7.7 Hz, 1H,
6
4
6
4
-C H Cl), 6.85–6.74 (m, 2H, -C H Cl). HRMS Calcd for 370.0238 (C H Cl N NaO,
6
4
6
4
15 11
2
5
+
[M+Na] ), found 370.0233. Anal. Calcd. For C H Cl N O: C, 51.74; H, 3.18; N,
1
5
11
2
5
20.11. Found: C, 51.63; H, 3.09; N, 20.18.
4.4.6. N',1-bis(3- chlorophenyl)-1H-1,2,3-triazole-4-carbohydrazide 6bc

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Light yellow powder, Yield, 65.5%; mp, 171–172 ºC. H NMR (400 MHz, DMSO-d )
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δ 10.69 (d, J = 2.1 Hz, 1H, -CONH-), 9.46 (s, 1H, -C N H-), 8.32 (d, J = 1.9 Hz, 1H,
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-NH-), 8.14 (t, J = 1.8 Hz, 1H, -C H Cl), 8.03–7.97 (m, 1H, -C H Cl), 7.67 (t, J = 8.0
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-C H Cl), 6.74 (dd, J = 6.2, 4.7 Hz, 3H, -C H Cl). HRMS Calcd for 370.0238
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+
(C H Cl N NaO, [M+Na] ), found 370.0233. Anal. Calcd. For C H Cl N O: C,
1
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11
2
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15 11
2
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51.74; H, 3.18; N, 20.11. Found: C, 51.58; H, 3.20; N, 20.15.
4.4.7. 1-(3- chlorophenyl)-N'-(4- chlorophenyl)-1H-1,2,3-triazole-4-carbohydrazide
6bd
1
White powder, Yield, 75.0%; mp, 194–195 ºC. H NMR (400 MHz, DMSO-d ) δ
6
10.66 (s, 1H, -CONH-), 9.45 (s, 1H, -C N H-), 8.20 (s, 1H, -NH-), 8.14 (t, J = 1.9 Hz,
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1H, -C H Cl), 8.03–7.98 (m, 1H, -C H Cl), 7.67 (t, J = 8.0 Hz, 1H, -C H Cl), 7.64–
6
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6
4
6
4
7.59 (m, 1H, -C H Cl), 7.22–7.18 (m, 2H, -C H Cl), 6.80–6.76 (m, 2H, -C H Cl).
6
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4
6
4
+
HRMS Calcd for 370.0238 (C H Cl N NaO, [M+Na] ), found 370.0233. Anal.
1
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11
2
5
Calcd. For C H Cl N O: C, 51.74; H, 3.18; N, 20.11. Found: C, 51.50; H, 3.26; N,
1
5
11
2
5
20.19.
4.4.8. N'-(2-chlorophenyl)-1-(4- chlorophenyl)-1H-1,2,3-triazole-4-carbohydrazide
6cb
1
Light yellow powder, Yield, 79.4%; mp, 208–209 ºC. H NMR (400 MHz, DMSO-d )
6
δ 10.72 (d, J = 2.0 Hz, 1H, -CONH-), 9.42 (s, 1H, -C N H-), 8.06–8.02 (m, 2H,
2
3
-C H Cl), 7.72 (d, J = 2.1 Hz, 2H, -C H Cl), 7.71 (d, J = 2.0 Hz, 1H, -NH-), 7.32 (dd,
6
4
6
4
J = 7.9, 1.2 Hz, 1H, -C H Cl), 7.16 (t, J = 7.2 Hz, 1H, -C H Cl), 6.84–6.74 (m, 2H,
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-C H Cl). HRMS Calcd for 370.0238 (C H Cl N NaO, [M+Na] ), found 370.0233.
6 4 15 11 2 5
Anal. Calcd. For C H Cl N O: C, 51.74; H, 3.18; N, 20.11. Found: C, 51.66; H, 3.28;
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N, 20.27.
4.4.9. N'-(3-chlorophenyl)-1-(4- chlorophenyl)-1H-1,2,3-triazole-4-carbohydrazide
6cc
1
Light yellow powder, Yield, 64.7%; mp, 195–196 ºC. H NMR (400 MHz, DMSO-d )
6
δ 10.67 (s, 1H, -CONH-), 9.41 (s, 1H, -C N H-), 8.30 (s, 1H, -NH-), 7.98 (d, J = 8.9
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Hz, 2H, -C H Cl), 7.72 (d, J = 1.9 Hz, 2H, -C H Cl), 7.21–7.14 (m, 1H, -C H Cl),
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6.74 (dd, J = 6.1, 4.5 Hz, 3H, -C H Cl). HRMS Calcd for 370.0238 (C H Cl N NaO,
6
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15 11
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+
[M+Na] ), found 370.0233. Anal. Calcd. For C H Cl N O: C, 51.74; H, 3.18; N,
1
5
11
2
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20.11. Found: C, 51.71; H, 3.28; N, 20.01.
4.4.10. N',1-bis(4- chlorophenyl)-1H-1,2,3-triazole-4-carbohydrazide 6cd
1
Light yellow powder, Yield, 72.5%; mp, 210–211 ºC. H NMR (400 MHz, DMSO-d )
6
δ 10.66 (d, J = 2.5 Hz, 1H, -CONH-), 9.42 (s, 1H, -C N H-), 8.20 (d, J = 2.4 Hz, 1H,
2
3
-NH-), 8.03 (d, J = 8.9 Hz, 2H, -C H Cl), 7.72 (d, J = 8.9 Hz, 2H, -C H Cl), 7.20 (d, J
6
4
6
4
= 8.8 Hz, 2H, -C H Cl), 6.77 (d, J = 8.8 Hz, 2H, -C H Cl). HRMS Calcd for 370.0238
6
4
6
4
+
(C H Cl N NaO, [M+Na] ), found 370.0233. Anal. Calcd. For C H Cl N O: C,
1
5
11
2
5
15 11
2
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51.74; H, 3.18; N, 20.11. Found: C, 51.61; H, 3.11; N, 20.20.
4.4.11. N'-(2-chlorophenyl)-1-(2-methoxyphenyl)-1H-1,2,3-triazole-4-carbohydrazide
6db
1
Light yellow powder, Yield, 75.0%; mp, 192–193 ºC. H NMR (400 MHz, DMSO-d )
6
δ 10.66 (s, 1H, -CONH-), 8.98 (s, 1H, -C N H-), 7.70 (s, 1H, -NH-), 7.68 (d, 1H,
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-C H OMe), 7.59 (t, J = 7.9 Hz, 1H, -C H Cl), 7.34 (dd, J = 15.7, 8.1 Hz, 2H,
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-C H OMe), 7.17 (dd, J = 14.8, 7.3 Hz, 2H, -C H Cl)), 6.83 (d, J = 7.6 Hz, 1H,
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-C H OMe), 6.77 (t, J = 7.6 Hz, 1H, -C H Cl), 3.88 (s, 3H, -OCH ). HRMS Calcd for
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+
366.0734 (C H ClN NaO , [M+Na] ), found 366.0728. Anal. Calcd. For
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14
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C H ClN O : C, 55.90; H, 4.11; N, 20.37. Found: C, 55.71; H, 4.21; N, 20.29.
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4.4.12. N'-(3-chlorophenyl)-1-(2-methoxyphenyl)-1H-1,2,3-triazole-4-carbohydrazide
6dc
1
Yellow powder, Yield, 66.8%; mp, 161–162 ºC. H NMR (400 MHz, DMSO-d ) δ
6
10.61 (d, J = 1.5 Hz, 1H, -CONH-), 8.98 (s, 1H, -C N H-), 8.28 (d, J = 1.6 Hz, 1H,
2
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-NH-), 7.69 (dd, J = 7.8, 1.4 Hz, 1H, -C H OMe), 7.59 (dd, J = 11.4, 4.5 Hz, 1H,
6
4
-C H Cl)), 7.36 (d, J = 8.4 Hz, 1H, -C H OMe), 7.18 (t, J = 7.2 Hz, 2H, -C H OMe),
6
4
6
4
6
4
6.78–6.70 (m, 3H, -C H Cl), 3.88 (s, 3H, -OCH ). HRMS Calcd for 366.0734
6
4
3
+
(C H ClN NaO , [M+Na] ), found 366.0728. Anal. Calcd. For C H ClN O : C,
1
6
14
5
2
16 14
5
2
55.90; H, 4.11; N, 20.37. Found: C, 55.84; H, 4.15; N, 20.30.
4.4.13. N'-(4-chlorophenyl)-1-(2-methoxyphenyl)-1H-1,2,3-triazole-4-carbohydrazide
6dd
1
Light yellow powder, Yield, 70.3%; mp, 182–183 ºC. H NMR (400 MHz, DMSO-d )
6
δ 10.57 (s, 1H, -CONH-), 8.96 (s, 1H, -C N H-), 8.16 (s, 1H, -NH-), 7.68 (dd, J = 7.8,
2
3
1.5 Hz, 1H, -C H OMe), 7.61–7.55 (m, 1H, -C H OMe), 7.35 (d, J = 8.2 Hz, 1H,
6
4
6
4
-C H OMe), 7.20 (dd, J = 12.4, 6.1 Hz, 2H, -C H Cl), 7.16–7.19 (m, 1H, -C H OMe),
6
4
6
4
6
4
6.78 (d, J = 8.8 Hz, 2H, -C H Cl), 3.88 (s, 3H, -OCH ). HRMS Calcd for 366.0734
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(C H ClN NaO , [M+Na] ), found 366.0728. Anal. Calcd. For C H ClN O : C,
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16 14
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55.90; H, 4.11; N, 20.37. Found: C, 55.75; H, 4.21; N, 20.40.
4.4.14. N'-(2-chlorophenyl)-1-(p-tolyl)-1H-1,2,3-triazole-4-carbohydrazide 6eb
1
White powder, Yield, 81.0%; mp, 211–212 ºC. H NMR (400 MHz, DMSO-d ) δ
6
10.68 (s, 1H, -CONH-), 9.34 (s, 1H, -C N H-), 7.87 (d, J = 8.3 Hz, 2H, -C H Me),
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7.71 (s, 1H, -NH-), 7.43 (d, J = 8.3 Hz, 2H, -C H Me), 7.32 (d, J = 7.9 Hz, 1H,
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-C H Cl), 7.16 (t, J = 7.7 Hz, 1H, -C H Cl), 6.83 (d, J = 8.1 Hz, 1H, -C H Cl), 6.77 (t,
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6
4
6
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J = 7.6 Hz, 1H, -C H Cl), 2.40 (s, 3H, -CH ). HRMS Calcd for 350.0784
6
4
3
+
(C H ClN NaO, [M+Na] ), found 350.0779. Anal. Calcd. For C H ClN O: C,
1
6
14
5
16 14
5
58.63; H, 4.31; N, 21.37. Found: C, 58.55; H, 4.21; N, 21.40.
4.4.15. N'-(3-chlorophenyl)-1-(p-tolyl)-1H-1,2,3-triazole-4-carbohydrazide 6ec
1
Light yellow powder, Yield, 72.3%; mp, 204–205 ºC. H NMR (400 MHz, DMSO-d )
6
δ 10.63 (s, 1H, -CONH-), 9.32 (s, 1H, -C N H-), 8.29 (s, 1H, -NH-), 7.86 (d, J = 8.4
2
3
Hz, 2H, -C H Me), 7.43 (d, J = 8.3 Hz, 2H, -C H Me), 7.17 (dd, J = 11.2, 5.2 Hz, 1H,
6
4
6
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-C H Cl), 6.76–6.71 (m, 3H, -C H Cl), 2.40 (s, 3H, -CH ). HRMS Calcd for 350.0784
6
4
6
4
3
+
(C H ClN NaO, [M+Na] ), found 350.0779. Anal. Calcd. For C H ClN O: C,
1
6
14
5
16 14
5
58.63; H, 4.31; N, 21.37. Found: C, 58.49; H, 4.39; N, 21.45.
4.4.16. N'-(4-chlorophenyl)-1-(p-tolyl)-1H-1,2,3-triazole-4-carbohydrazide 6ed
1
Light yellow powder, Yield, 75.4%; mp, 224–225 ºC. H NMR (400 MHz, DMSO-d )
6
δ 10.61 (d, J = 2.4 Hz, 1H, -CONH-), 9.32 (s, 1H, -C N H-), 8.18 (d, J = 2.3 Hz, 1H,
2
3
-NH-), 7.86 (d, J = 8.4 Hz, 2H, -C H Me), 7.43 (d, J = 8.2 Hz, 2H, -C H Me), 7.20 (d,
6
4
6
4
J = 8.8 Hz, 2H, -C H Cl), 6.78 (d, J = 8.8 Hz, 2H, -C H Cl), 2.52–2.49 (m, 3H, -CH ).
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HRMS Calcd for 350.0784 (C H ClN NaO, [M+Na] ), found 350.0779. Anal. Calcd.
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6
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For C H ClN O: C, 58.63; H, 4.31; N, 21.37. Found: C, 58.45; H, 4.29; N, 21.55.
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4.4.17. 1-(2-chlorophenyl)-N'-(2-fluorophenyl)-1H-1,2,3-triazole-4-carbohydrazide
6ae
1
Light yellow powder, Yield, 79.0%; mp, 161–162 ºC. H NMR (400 MHz, DMSO-d )
6
δ 10.66 (s, 1H, -CONH-), 9.14 (s, 1H, -C N H-), 7.93 (s, 1H, -NH-), 7.83 (d, J = 4.4
2
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Hz, 2H, -C H Cl), 7.78 (d, J = 1.6 Hz, 1H, -C H Cl), 7.69 (d, J = 1.5 Hz, 1H,
6
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6
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-C H Cl), 7.14–7.07 (m, 1H, -C H F), 7.03–6.98 (m, 1H, -C H F), 6.86–6.80 (m, 1H,
6
4
6
4
6
4
-C H F), 6.75 (dd, J = 15.4, 6.4 Hz, 1H, -C H F). HRMS Calcd for 354.0534
6
4
6
4
+
(C H ClFN NaO, [M+Na] ), found 354.0529. Anal. Calcd. For C H ClFN O: C,
1
5
11
5
15 11
5
54.31; H, 3.34; N, 21.11. Found: C, 54.45; H, 3.23; N, 21.05.
4.4.18. 1-(2-chlorophenyl)-N'-(3-fluorophenyl)-1H-1,2,3-triazole-4-carbohydrazide
6af
1
Yellow powder, Yield, 74.8%; mp, 154–155 ºC. H NMR (400 MHz, DMSO-d ) δ
6
10.67 (s, 1H, -CONH-), 9.14 (s, 1H, -C N H-), 8.31 (s, 1H, -NH-), 7.82 (dd, J = 8.0,
2
3
1.2 Hz, 1H, -C H Cl), 7.79 (dd, J = 7.7, 1.6 Hz, 1H, -C H Cl), 7.69 (td, J = 7.8, 1.7
6
4
6
4
Hz, 1H, -C H Cl), 7.63 (td, J = 7.6, 1.4 Hz, 1H, -C H Cl), 7.18 (dd, J = 15.3, 8.3 Hz,
6
4
6
4
1H, -C H F), 6.62 (d, J = 7.8 Hz, 1H, -C H F), 6.54–6.47 (m, 2H, -C H F). HRMS
6
4
6
4
6
4
+
Calcd for 354.0534 (C H ClFN NaO, [M+Na] ), found 354.0529. Anal. Calcd. For
1
5
11
5
C H ClFN O: C, 54.31; H, 3.34; N, 21.11. Found: C, 54.25; H, 3.33; N, 21.25.
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4.4.19. 1-(2-chlorophenyl)-N'-(4-fluorophenyl)-1H-1,2,3-triazole-4-carbohydrazide
6ag

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Light yellow powder, Yield, 74.0%; mp, 169–170 ºC. H NMR (400 MHz, DMSO-d )
6
δ 10.62 (d, J = 2.9 Hz, 1H, -CONH-), 9.11 (s, 1H, -C N H-), 7.97 (d, J = 2.9 Hz, 1H,
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-NH-), 7.82 (dd, J = 8.0, 1.3 Hz, 1H, -C H Cl), 7.78 (dd, J = 7.7, 1.7 Hz, 1H,
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-C H Cl), 7.01 (t, J = 8.9 Hz, 2H, -C H F), 6.82–6.76 (m, 2H, -C H F). HRMS Calcd
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+
for 354.0534 (C H ClFN NaO, [M+Na] ), found 354.0529. Anal. Calcd. For
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11
5
C H ClFN O: C, 54.31; H, 3.34; N, 21.11. Found: C, 54.39; H, 3.23; N, 21.17.
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5
4.4.20. 1-(2-chlorophenyl)-N'-(2-methoxyphenyl)-1H-1,2,3-triazole-4-carbohydrazide
6ah
1
White powder, Yield, 73.1%; mp, 188–189 ºC. H NMR (400 MHz, DMSO-d ) δ
6
10.63 (s, 1H, -CONH-), 9.10 (s, 1H, -C N H-), 7.81 (dd, J = 8.0, 1.3 Hz, 1H,
2
3
-C H Cl), 7.78 (dd, J = 7.7, 1.7 Hz, 1H, -C H Cl), 7.68 (td, J = 7.7, 1.8 Hz, 1H,
6
4
6
4
-C H Cl), 7.62 (td, J = 7.6, 1.4 Hz, 1H, -C H Cl), 7.19 (s, 1H, -NH-), 6.92 (d, J = 7.9
6
4
6
4
Hz, 1H, -C H OMe), 6.84–6.71 (m, 3H, -C H OMe), 3.85 (s, 3H, -OCH ). HRMS
6
4
6
4
3
+
Calcd for 366.0734 (C H ClN NaO , [M+Na] ), found 366.0728. Anal. Calcd. For
16
14
5
2
C H ClN O : C, 55.90; H, 4.11; N, 20.37. Found: C, 55.65; H, 4.23; N, 20.25.
16
14
5
2
4.4.21. 1-(2-chlorophenyl)-N'-(3-methoxyphenyl)-1H-1,2,3-triazole-4-carbohydrazide
6ai
1
Brown powder, Yield, 62.5%; mp, 154–155 ºC. H NMR (400 MHz, DMSO-d ) δ
6
10.67 (s, 1H, -CONH-), 9.60 (s, 1H, -C N H-), 7.79 (dd, J = 7.8, 1.4 Hz, 1H,
2
3
-C H Cl), 7.63 (dd, J = 11.4, 4.5 Hz, 1H, -C H Cl), 7.42 (s, 1H, -NH-), 7.36 (d, J =
6
4
6
4
8.4 Hz, 1H, -C H OMe), 7.28 (t, J = 7.2 Hz, 2H, -C H Cl), 6.78–6.70 (m, 3H,
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-C H OMe), 3.81 (s, 3H, -OCH ). HRMS Calcd for 366.0734 (C H ClN NaO ,
6
4
3
16 14
5
2
+
[M+Na] ), found 366.0728. Anal. Calcd. For C H ClN O : C, 55.90; H, 4.11; N,
1
6
14
5
2
20.37. Found: C, 55.73; H, 4.08; N, 20.30.
4.4.22. 1-(2-chlorophenyl)-N'-(4-methoxyphenyl)-1H-1,2,3-triazole-4-carbohydrazide
6aj
1
White powder, Yield, 68.0%; mp, 158–159 ºC. H NMR (400 MHz, DMSO-d ) δ
6
10.55 (s, 1H, -CONH-), 9.09 (s, 1H, -C N H-), 7.82 (s, 1H, -NH-), 7.80 (d, J = 5.5 Hz,
2
3
1H, -C H Cl), 7.77 (s, 1H, -C H Cl), 7.67 (d, J = 7.9 Hz, 1H, -C H Cl), 7.63 (d, J =
6
4
6
4
6
4
7.6 Hz, 1H, -C H Cl), 6.80–6.75 (m, 4H, -C H OMe), 3.67 (s, 3H, -OCH ). HRMS
6
4
6
4
3
+
Calcd for 366.0734 (C H ClN NaO , [M+Na] ), found 366.0728. Anal. Calcd. For
16
14
5
2
C H ClN O : C, 55.90; H, 4.11; N, 20.37. Found: C, 55.79; H, 4.15; N, 20.19.
16
14
5
2
4.4.23.
1-(2-chlorophenyl)-N'-(2-nitrophenyl)-1H-1,2,3-triazole-4-carbohydrazide
6ak
1
Orange powder, Yield, 65.0%; mp, 201–202 ºC. H NMR (400 MHz, DMSO-d ) δ
6
10.80 (s, 1H, -CONH-), 9.22 (d, J = 0.9 Hz, 1H, -C N H-), 7.91 (d, J = 8.5 Hz, 2H,
2
3
-C H Cl), 7.83 (s, 1H, -NH-), 7.72–7.67 (m, 2H, -C H Cl), 7.64 (dt, J = 5.2, 2.7 Hz,
6
4
6
4
2H, -C H NO ), 7.43 (d, J = 8.6 Hz, 2H, -C H NO ). HRMS Calcd for 381.0479
6
4
2
6
4
2
+
(C H ClN NaO , [M+Na] ), found 381.0473. Anal. Calcd. For C H ClN O : C,
1
5
11
6
3
15 11
6
3
50.22; H, 3.09; N, 23.43. Found: C, 50.39; H, 3.15; N, 23.27.
4.4.24. 1-(2-chlorophenyl)-N'-(3-nitrophenyl)-1H-1,2,3-triazole-4-carbohydrazide 6al
1
Orange powder, Yield, 67.1%; mp, 189–190 ºC. H NMR (400 MHz, DMSO-d ) δ
6
10.64 (s, 1H, -CONH-), 9.22 (s, 1H, -C N H-), 8.40 (s, 1H, -NH-), 7.83–7.79 (m, 2H,
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-C H Cl), 7.71–7.64 (m, 2H, -C H Cl), 7.59 (dd, J = 8.1, 1.9 Hz, 2H, -C H NO ),
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6
4
6
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7.46 (t, J = 8.1 Hz, 1H, -C H NO ), 7.35 (t, J = 8.1 Hz, 1H, -C H NO ). HRMS Calcd
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4
2
6
4
2
+
for 381.0479 (C H ClN NaO , [M+Na] ), found 381.0473. Anal. Calcd. For
1
5
11
6
3
C H ClN O : C, 50.22; H, 3.09; N, 23.43. Found: C, 50.18; H, 3.22; N, 23.25.
15
11
6
3
4.4.25.
1-(2-chlorophenyl)-N'-(4-nitrophenyl)-1H-1,2,3-triazole-4-carbohydrazide
6am
1
Orange powder, Yield, 70.4%; mp, 216–217 ºC. H NMR (400 MHz, DMSO-d ) δ
6
10.98 (s, 1H, -CONH-), 9.20 (s, 1H, -C N H-), 8.10 (d, J = 9.1 Hz, 2H, -C H Cl),
2
3
6
4
7.83 (s, 1H, -NH-), 7.82–7.78 (m, 2H, -C H Cl), 7.70–7.60 (m, 2H, -C H NO ), 6.84
6
4
6
4
2
(d, J = 9.2 Hz, 2H, -C H NO ). HRMS Calcd for 381.0479 (C H ClN NaO ,
6
4
2
15 11
6
3
+
[M+Na] ), found 381.0473. Anal. Calcd. For C H ClN O : C, 50.22; H, 3.09; N,
1
5
11
6
3
23.43. Found: C, 50.31; H, 3.29; N, 23.30.
4.4.26. N'-(2-chlorophenyl)-1-(2-fluorophenyl)-1H-1,2,3-triazole-4-carbohydrazide
6fb
1
White powder, Yield, 80.5%; mp, 166–167 ºC. H NMR (400 MHz, DMSO-d ) δ
6
10.73 (d, J = 2.0 Hz, 1H, -CONH-), 9.17 (d, J = 1.6 Hz, 1H, -C N H-), 7.91 (td, J =
2
3
7.8, 1.5 Hz, 1H, -C H F), 7.73 (d, J = 1.9 Hz, 1H, -NH-), 7.70–7.64 (m, 1H, -C H Cl),
6
4
6
4
7.65–7.59 (m, 1H, -C H F), 7.49 (t, J = 7.5 Hz, 1H, -C H F), 7.32 (dd, J = 7.9, 1.2 Hz,
6
4
6
4
1H, -C H F), 7.16 (t, J = 7.3 Hz, 1H, -C H Cl), 6.84 (dd, J = 8.1, 1.1 Hz, 1H,
6
4
6
4
-C H Cl), 6.80–6.74 (m, 1H, -C H Cl). HRMS Calcd for 354.0534 (C H ClFN NaO,
6
4
6
4
15 11
5
+
[M+Na] ), found 354.0529. Anal. Calcd. For C H ClFN O: C, 54.31; H, 3.34; N,
1
5
11
5
21.11. Found: C, 54.65; H, 3.20; N, 21.05.

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4.4.27. N'-(3-chlorophenyl)-1-(2-fluorophenyl)-1H-1,2,3-triazole-4-carbohydrazide
6fc
1
Light yellow powder, Yield, 71.6%; mp, 169–170 ºC. H NMR (400 MHz, DMSO-d )
6
δ 10.68 (d, J = 2.3 Hz, 1H, -CONH-), 9.16 (d, J = 1.6 Hz, 1H, -C N H-), 8.31 (d, J =
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2.3 Hz, 1H, -NH-), 7.90 (td, J = 7.8, 1.5 Hz, 1H, -C H F), 7.71–7.57 (m, 2H, -C H F),
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4
6
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7.48 (dd, J = 11.6, 4.6 Hz, 1H, -C H F), 7.18 (t, J = 8.1 Hz, 1H, -C H Cl), 6.80–6.69
6
4
6
4
+
(m, 3H, -C H Cl). HRMS Calcd for 354.0534 (C H ClFN NaO, [M+Na] ), found
6
4
15 11
5
354.0529. Anal. Calcd. For C H ClFN O: C, 54.31; H, 3.34; N, 21.11. Found: C,
1
5
11
5
54.25; H, 3.10; N, 21.32.
4.4.28. N'-(4-chlorophenyl)-1-(2-fluorophenyl)-1H-1,2,3-triazole-4-carbohydrazide
6fd
1
Yellow powder, Yield, 69.8%; mp, 179–180 ºC. H NMR (400 MHz, DMSO-d ) δ
6
10.67 (s, 1H, -CONH-), 9.16 (d, J = 1.5 Hz, 1H, -C N H-), 8.19 (s, 1H, -NH-), 7.90
2
3
(td, J = 7.9, 1.4 Hz, 1H, -C H F), 7.72–7.58 (m, 2H, -C H F), 7.49 (t, J = 7.6 Hz, 1H,
6
4
6
4
-C H F), 7.20 (d, J = 8.8 Hz, 2H, -C H Cl), 6.78 (d, J = 8.8 Hz, 2H, -C H Cl). HRMS
6
4
6
4
6
4
+
Calcd for 354.0534 (C H ClFN NaO, [M+Na] ), found 354.0529. Anal. Calcd. For
1
5
11
5
C H ClFN O: C, 54.31; H, 3.34; N, 21.11. Found: C, 54.35; H, 3.16; N, 21.27.
15
11
5
4.4.29. N',1-bis(2- fluorophenyl)-1H-1,2,3-triazole-4-carbohydrazide 6fe
1
Yellow powder, Yield, 81.3%; mp, 154–155 ºC. H NMR (400 MHz, DMSO-d ) δ
6
10.64 (d, J = 1.8 Hz, 1H, -CONH-), 9.15 (d, J = 1.5 Hz, 1H, -C N H-), 7.93 (s, 1H,
2
3
-NH-), 7.89 (dd, J = 7.8, 1.3 Hz, 1H, -C H F), 7.71–7.57 (m, 2H, -C H F), 7.49 (t, J =
6
4
6
4
7.6 Hz, 1H, -C H F), 7.11 (dd, J = 12.1, 8.1 Hz, 1H, -C H F), 7.00 (t, J = 7.7 Hz, 1H,
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-C H F), 6.84 (t, J = 7.9 Hz, 1H, -C H F), 6.78–6.70 (m, 1H, -C H F). HRMS Calcd
6
4
6
4
6
4
+
for 338.0829 (C H F N NaO, [M+Na] ), found 338.0824. Anal. Calcd. For
1
5
11
2
5
C H F N O: C, 57.14; H, 3.52; N, 22.21. Found: C, 57.35; H, 3.36; N, 21.27.
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11
2
5
4.4.30. 1-(2- fluorophenyl)-N'-(3-fluorophenyl)-1H-1,2,3-triazole-4-carbohydrazide
6ff
1
Light yellow powder, Yield, 65.6%; mp, 151–152 ºC. H NMR (400 MHz, DMSO-d )
6
δ 10.66 (d, J = 2.0 Hz, 1H, -CONH-), 9.15 (d, J = 1.4 Hz, 1H, -C N H-), 8.30 (d, J =
2
3
1.5 Hz, 1H, -NH-), 7.90 (t, J = 7.1 Hz, 1H, -C H F), 7.71–7.64 (m, 1H, -C H F),
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4
6
4
7.64–7.58 (m, 1H, -C H F), 7.48 (t, J = 7.5 Hz, 1H, -C H F), 7.18 (dd, J = 15.4, 8.3
6
4
6
4
Hz, 1H, -C H F), 6.61 (d, J = 8.6 Hz, 1H, -C H F), 6.53–6.47 (m, 2H, -C H F).
6
4
6
4
6
4
+
HRMS Calcd for 338.0829 (C H F N NaO, [M+Na] ), found 338.0824. Anal. Calcd.
1
5
11
2
5
For C H F N O: C, 57.14; H, 3.52; N, 22.21. Found: C, 57.25; H, 3.46; N, 21.17.
1
5
11
2
5
4.4.31. 1-(2- fluorophenyl)-N'-(4-fluorophenyl)-1H-1,2,3-triazole-4-carbohydrazide
6fg
1
Light yellow powder, Yield, 77.6%; mp, 187–188 ºC. H NMR (400 MHz, DMSO-d )
6
δ 10.62 (d, J = 3.0 Hz, 1H, -CONH-), 9.13 (d, J = 1.7 Hz, 1H, -C N H-), 7.97 (d, J =
2
3
3.0 Hz, 1H, -NH-), 7.90 (td, J = 7.8, 1.5 Hz, 1H, -C H F), 7.71–7.57 (m, 2H, -C H F),
6
4
6
4
7.48 (dd, J = 11.7, 4.6 Hz, 1H, -C H F), 7.05–6.96 (m, 2H, -C H F), 6.83–6.75 (m,
6
4
6
4
+
2H, -C H F). HRMS Calcd for 338.0829 (C H F N NaO, [M+Na] ), found
6
4
15 11
2
5
338.0824. Anal. Calcd. For C H F N O: C, 57.14; H, 3.52; N, 22.21. Found: C,
1
5
11
2
5
57.09; H, 3.40; N, 21.31.
4.5. General procedure for the synthesis of compounds 7a

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In a round-bottom flask equipped with a magnetic stirring bar, MnO (150 mmol)
2
and 10 mmol of 1,2,3-triazole 3a were added to ethyl acetate (30 mL), the mixture
was heated at reflux for 1 h. Afterwards, the solution was filtered and concentrated in
vacuo to obtain pure aldehyde 4a with a yield of 80.0%.
Both equimolar aldehyde and benzoyl hydrazine were mixed in ethanol and
heated at reflux for approximately 2 h[26]. The reaction mixture was cooled to
ambient temperature and massive crystal particles were generated and separated out of
the solution to create 7a.
4.5.1. N'-((1-(2-chlorophenyl)-1H-1,2,3-triazol-4-yl)methylene)benzohydrazide 7a
1
White powder, Yield, 95.0%; mp, 201–202 ºC. H NMR (400 MHz, DMSO-d ) δ
6
11.94 (s, 1H, -NH-), 9.32 (d, J = 2.4 Hz, 1H, -C N H-), 8.66 (s, 1H, -CH=N-), 8.04 (d,
2
3
J = 7.8 Hz, 2H, -C H ), 8.00 (d, J = 7.7 Hz, 1H, -C H ), 7.94 (d, J = 7.4 Hz, 2H,
6
5
6
5
-C H ), 7.61 (t, J = 7.7 Hz, 1H, -C H Cl), 7.46 (dd, J = 11.3, 8.0 Hz, 2H, -C H Cl),
6
5
6
4
6
4
7.38 (t, J = 7.6 Hz, 1H, -C H Cl). HRMS Calcd for 348.0628 (C H ClN NaO,
6
4
16 12
5
+
[M+Na] ), found 348.0623. Anal. Calcd. For C H ClN O: C, 55.10; H, 3.47; N,
1
6
12
5
20.08. Found: C, 55.19; H, 3.25; N, 20.31.
4.6. General procedure for the synthesis of compounds 9a
Solid hydroxylamine hydrochloride (3 mmol) was added to a stirred solution of
4a (1 mmol) in ethanol. The mixture was heated at reflux for 30 min, then sodium
bicarbonate (3.6 mmol) dissolved in water was added dropwise. The reaction was
monitored by thin layer chromatography (TLC). Ethanol was removed by distillation
under reduced pressure. The 1, 2, 3-triazole oxime 8a was separated out and collected

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by filtration (yield of 90.0%).
Benzyl bromide (1.2 mmol) and KOH (3 mmol) was added to a solution of 8a (1
mmol) in DMF (1 mL) and stirred at r.t. for an hour[27]. Afterwards, the mixture was
washed with saturated salt water, then extracted with ethyl acetate three times. The
organic phase was dried over anhydrous Na SO , filtered, and concentrated in vacuo.
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4
Further purification was carried out with silica gel column chromatography using
petroleum ether/EtOAc (2:1) as the eluent to obtain the pure derivative 9a.
4.6.1. 1-(2-chlorophenyl)-1H-1,2,3-triazole-4-carbaldehyde O-benzyl oxime 9a
1
Light yellow powder, Yield, 93.0%; mp, 146–147 ºC. H NMR (400 MHz, DMSO-d )
6
δ 8.98 (d, J = 1.8 Hz, 1H, -C N H-), 8.66 (s, 1H, -CH=N-), 7.82–7.77 (m, 1H,
2
3
-C H Cl), 7.76–7.70 (m, 1H, -C H Cl), 7.65 (d, J = 8.1 Hz, 1H, -C H Cl), 7.62–7.57
6
4
6
4
6
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(m, 1H, -C H Cl), 7.44 (d, J = 7.8 Hz, 2H, -C H ), 7.41 (d, J = 8.1 Hz, 2H, -C H ),
6
4
6
5
6
5
7.32 (t, J = 7.2 Hz, 1H, -C H ), 5.20 (s, 2H, -CH -). HRMS Calcd for 335.0676
6
5
2
+
(C H ClN NaO, [M+Na] ), found 335.0671. Anal. Calcd. For C H ClN O: C,
1
6
13
4
16 13
4
57.24; H, 3.90; N, 16.69. Found: C, 57.19; H, 3.45; N, 16.41.
4.7. Biological activities assay
The test fungi Rhizoctonia solani, Sclerotinia sclerotiorum, Fusarium
graminearum, Magnaporthe oryzae were provided by the Laboratory of Plant Disease
Control at Nanjing Agricultural University. After removing strains from the storage
tube, the strains were incubated in potato dextrose agar (PDA) at 25 °C for a week to
develop new mycelia for the antifungal assay.
4.7.1. Antifungal bioassay

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Fungicidal activity was tested using the mycelia growth inhibition method on
PDA in vitro against four plant-pathogenic fungi (R. solani, S. sclerotiorum, F.
graminearum, M. oryzae). The synthesized compounds were dissolved in DMSO to
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-1
prepare the 1.0 × 10 µg mL stock solution before mixing with molten PDA below
60 ºC. The initial concentration used for antifungal activity screening in vitro was set
−
1
as 25 µg mL to find active compounds. After the appropriate number of days (2 d
for R. solani, 2.5 d for S. sclerotiorum, 3 d for F. graminearum, and 8 d for M. oryzae)
at 25 ºC in the dark, the colony diameter of each strain was measured. Percentage
inhibition was calculated as (1-a/b) ×100%, where a is the colony diameter in petri
dishes with compounds and b is the mean colony diameter in petri dishes without
tested compounds[28]. DMSO served as a negative control and commercially
available agricultural fungicides carbendazim and validamycin A were used as
positive controls. The EC values were calculated through linear-regression analysis
50
(Table 1).
4.7.2. In vivo protective activity of 6ad against RSB
The tested rice cultivar was grown in plastic pots (diameter of 18 cm × height of
16 cm) in the greenhouse, which simulated natural conditions, and artificially
inoculated the cultivar with the strain R. solani using a previously reported
method[29]. Validamycin A (20% S.P., Zhejiang Tonglu HSBC Biochemical Co., Ltd.,
China) was used as the positive control. The length of symptoms was measured after
cultivation at 25 ºC for 7 days (different concentrations were sprayed on plants 24 h
before inoculation). The protective control efficacies were calculated as follows:

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Protection efficacy = [(average lesion length of control − average lesion length of
treated group) / average lesion length of control] × 100%.
4.7.3. In vivo protective activity of 6ad against RSR
The strain S. sclerotiorum and susceptible cole leaves were used to measure the
efficacy of compounds in vivo. Healthy cole leaves were sprayed with compounds
(0.5 mg/mL) and subsequently cultivated at 20 °C for 24 h before artificial inoculation
with the strain S. sclerotiorum. The diameters of symptoms were observed after
cultivation at 20 ºC for 36 h. Carbendazim (50% W.P., Jiangsu Rotam Lanfeng
Biochemical Co., Ltd., China) was co-assayed as the positive control. The efficacy of
disease control was calculated as (1 − c/d) × 100, where c is the diameter of the
treatment and d is the diameter of the negative control[30].
4.7.4. In vivo therapeuti activity of 6ad against FHB
The F. graminearum susceptible wheat cultivar Yangmai 2 was used to evaluate
the therapeutic activity of 6ad in a greenhouse at Nanjing Agricultural University.
When anthesis approached, plants were artificially inoculated with the strain F.
graminearum for 24 h before they were sprayed with the test compounds. Inoculation
was achieved by injecting plant spikes with a dosage of 10µL per spike of the
prepared conidial suspension using a pipette. Carbendazim (50% WP, Jiangsu Rotam
Lanfeng Biochemical Co., Ltd., China) was co-assayed as the positive control. The
2
solution was applied at 75 mL/m by using a precision hand sprayer. Visual disease
assessment was conducted 15 days after spraying. The therapeutic control efficacy
was calculated as follows[31]: Therapeutic efficacy = [1-(Disease index of the

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treatment) / (Disease index of control) ] × 100 %.
4.7.5. In vivo protective activity and inhibition conidium germination of 6ad against
RB
6ad was further measured in vivo against RB in rice (Shanyou 63). 6ad (100 µg
-1
5
mL ) was sprayed on two-week-old rice-leaf seedlings. A concentration of 1 × 10
-1
RB spores mL in sterile water with 0.2% (w/v) gelatin was sprayed on leaves after
24 hours. Inoculated plants were kept in a 25 °C with 90% humidity and in a 12/12
hours light/dark cycle. Lesion formation was observed daily and photographed 7 days
-1
after inoculation[32]. The positive control was 50 µg mL Carbendazim (50% WP,
Jiangsu Rotam Lanfeng Biochemical Co., Ltd., China). The protective control efficacy
was calculated as follows: Protective efficacy = [(number of disease spots of control −
number of disease spots of treated group) / number of disease spots of control] ×
100%.
The wild type M. oryzae Guy11 was used to evaluate the inhibition of conidium
germination of 6ad. The mycelia were incubated on a straw decoction (10%) and corn
(4%) medium at 28 °C in darkness for 7 days, followed by 3-4 days of constant
illumination to induce the production of conidia. Conidia were harvested after 10 days,
filtered through three layers of lens paper, and resuspended to a concentration of 5 ×
4
10 spores per milliliter in sterile water. For conidium germination, conidial
suspensions (20 µL) were placed on plastic cover slips and dropped into 6ad solution
-1
at the concentration of 1.0 µg mL , then incubated under humid conditions at room
temperature[33]. Appressorium formation percentage was examined after incubation