Design, Synthesis and Bioactivity of Novel Fluoropyrazole Hydrazides

doi:10.6023/cjoc202112021

Abstract

Laccase is a potential target and has important physiological functions in micro organisms. In order to find compounds with high laccase inhibitory activity, 15 cinnamaldehyde hydrazide derivatives containing difluoromethylpyrazole were designed and synthesized by using the principle of active substructure splicing with 4-chlorocinnamaldehyde thiosemicarbazone (PMDD-5Y) as the lead compound. The structures of the compounds were confirmed by ¹H NMR, ¹³C NMR and HRMS. The results of laccase inhibitory activity test showed that all the prepared compounds have good activity, and N'- ((Z)-3-(3,4-dichlorophenyl)-3-phenylallylidene)-3-(difluoromethyl)-1-methyl-1H-pyrazole-4-carbohydrazide (5l) and N'-((Z)- 3-(2-chlorophenyl)-3-(4-chlorophenyl)allylidene)-3-(difluoromethyl)-1-methyl-1H-pyrazole-4-carbohydrazide (5m) had the half-maximal inhibiting concentration (IC₅₀) values of 0.069 and 0.063 mmol/L, respectively, which were significantly better than that of lead compound PMDD-5Y (IC₅₀=0.553 mmol/L) and positive control cysteine (IC₅₀=0.298 mmol/L). The in vitro bioassay showed that the compounds with good laccase inhibitory activity showed certain fungicidal activity against V. mali, F. graminearum and M. grisea at a concentration of 50 μg/mL. In vivo trials against M. grisea demonstrated that compound 5l was effective with curative activity of 78.55% at a concentration of 200 μg/mL, which showed a certain control efficiency in vivo. Further molecular docking study revealed that compound 5l with high laccase inhibitory activity had a good protein binding mode with laccase protein.

Key words: laccase, laccase inhibitor, fluoropyrazole fragment, molecular docking, fungicidal activity

Cite this article

Changkai Wang, Tengda Sun, Xuebo Zhang, Xinling Yang, Xingxing Lu, Huan Xu, Fasheng Shi, Li Zhang, Yun Ling. Design, Synthesis and Bioactivity of Novel Fluoropyrazole Hydrazides[J]. Chinese Journal of Organic Chemistry, 2022, 42(5): 1527-1536.

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Compd.	R¹	R²	IC₅₀/(mmol•L^–1)	95% confidence interval	Regression equation	r
5a	H	H	0.094	0.083~0.110	y=3.00x–4.65	0.98
5b	H	2-CH₃	0.071	0.063~0.081	y=4.85x–7.02	0.97
5c	H	3-CH₃	0.073	0.065~0.087	y=5.77x–8.43	0.99
5d	H	4-CH₃	0.067	0.060~0.079	y=4.22x–6.00	0.93
5e	4-CH₃	4-CH₃	0.111	0.098~0.144	y=5.59x–9.26	0.98
5f	4-OCH₃	4-OCH₃	0.136	0.107~0.259	y=5.23x–9.36	0.99
5g	H	2-F	0.089	0.079~0.103	y=4.31x–6.66	0.96
5h	H	2-Cl	0.086	0.078~0.097	y=6.86x–10.66	0.99
5i	H	2-Br	0.136	0.098~0.592	y=3.13x–5.64	0.98
5j	H	4-Cl	0.089	0.081~0.103	y=7.11x–11.13	0.98
5k	4-Cl	4-Cl	0.089	0.077~0.115	y=3.99x–6.39	0.99
5l	H	3,4-Cl₂	0.069	0.063~0.076	y=4.89x–7.29	0.99
5m	4-Cl	2-Cl	0.063	0.058~0.069	y=4.57x–6.64	0.98
5n	H	4-NO₂	0.091	0.083~0.102	y=7.22x–11.39	0.99
5o	H	2-Ph	0.066	0.058~0.080	y=4.06x–6.00	0.98
PMDD-5Y			0.553	0.491~0.610	y=5.74x–12.18	0.97
半胱氨酸			0.298	0.218~0.483	y=2.77x–4.23	0.98

Compd.	R¹	R²	IC₅₀/(mmol•L^–1)	95% confidence interval	Regression equation	r
5a	H	H	0.094	0.083~0.110	y=3.00x–4.65	0.98
5b	H	2-CH₃	0.071	0.063~0.081	y=4.85x–7.02	0.97
5c	H	3-CH₃	0.073	0.065~0.087	y=5.77x–8.43	0.99
5d	H	4-CH₃	0.067	0.060~0.079	y=4.22x–6.00	0.93
5e	4-CH₃	4-CH₃	0.111	0.098~0.144	y=5.59x–9.26	0.98
5f	4-OCH₃	4-OCH₃	0.136	0.107~0.259	y=5.23x–9.36	0.99
5g	H	2-F	0.089	0.079~0.103	y=4.31x–6.66	0.96
5h	H	2-Cl	0.086	0.078~0.097	y=6.86x–10.66	0.99
5i	H	2-Br	0.136	0.098~0.592	y=3.13x–5.64	0.98
5j	H	4-Cl	0.089	0.081~0.103	y=7.11x–11.13	0.98
5k	4-Cl	4-Cl	0.089	0.077~0.115	y=3.99x–6.39	0.99
5l	H	3,4-Cl₂	0.069	0.063~0.076	y=4.89x–7.29	0.99
5m	4-Cl	2-Cl	0.063	0.058~0.069	y=4.57x–6.64	0.98
5n	H	4-NO₂	0.091	0.083~0.102	y=7.22x–11.39	0.99
5o	H	2-Ph	0.066	0.058~0.080	y=4.06x–6.00	0.98
PMDD-5Y			0.553	0.491~0.610	y=5.74x–12.18	0.97
半胱氨酸			0.298	0.218~0.483	y=2.77x–4.23	0.98

Treatment	Curative activity			Protective activity
Treatment	Morbidity/%	Disease index/%	Control efficiency/%	Morbidity/%	Disease index/%	Control efficiency/%
5l	100	8.73	78.55	100	16.13	60.36
PMDD-5Y	100	11.21	72.45	100	16.97	58.29
IPT^a	100	11.83	70.93	100	12.82	68.49
CK	100	40.69		100	40.69

Treatment	Curative activity			Protective activity
Treatment	Morbidity/%	Disease index/%	Control efficiency/%	Morbidity/%	Disease index/%	Control efficiency/%
5l	100	8.73	78.55	100	16.13	60.36
PMDD-5Y	100	11.21	72.45	100	16.97	58.29
IPT^a	100	11.83	70.93	100	12.82	68.49
CK	100	40.69		100	40.69

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