Design, synthesis and biological activity of novel pyrazole-quinazoline-2,4-diones as 4-hydroxyphenylpyruvate dioxygenase inhibitors
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摘要: 对羟基苯丙酮酸双加氧酶 (HPPD) 抑制剂近年来因其高活性和低抗性风险成为了除草剂领域的研究热点。喹唑啉二酮被证明是一类具有潜力的骨架结构，为了继续发挥该骨架的优势，在前期工作的基础上，结合已有的构效关系研究结果，设计并合成了30个新型含喹唑啉二酮结构的吡唑类HPPD抑制剂，其结构均经过了高分辨质谱 (HRMS)、核磁共振氢谱 (1H NMR) 和碳谱 (13C NMR) 的表征。酶水平和活体活性测试结果表明，大部分目标化合物展现出了与对照药剂喹草酮相当甚至更优异的酶抑制活性。温室除草活性测试结果表明，目标化合物对6种供试杂草均有一定的生长抑制作用，特别是化合物 9-28 ，在有效成分150 g/hm2剂量下，对6种杂草的防治效果均在80%以上，其中对稗草和马唐的防治效果达100%。最后，培养了代表性化合物 9-28 与拟南芥HPPD的复合物晶体结构，从分子层面说明了该类抑制剂与靶标的结合模式，也为后续该类抑制剂的开发提供了一定的分子基础和设计思路。
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关键词:
- 对羟基苯丙酮酸双加氧酶 /
- 喹唑啉二酮 /
- 吡唑类 /
- 除草活性 /
- 晶体结构
Abstract: In recent years, inhibitors targeting 4-hydroxyphenylpyruvate dioxygenase (HPPD) have become the hot research in the field of herbicide due to their high activity and low risk of resistance. Quinazoline-2,4-dione has been proved to be a potent scaffold. To further exploit its advantage, 30 novel pyrazole-quinazoline-2,4-dione derivatives were designed and synthesized based on the previous work and existing structure-activity relationship, all the structure of which were determined by HRMS, 1H NMR and 13C NMR. Furthermore, the in vitro and in vivo activity of the synthesized compounds were evaluated. The result indicated that most of the synthesized compounds exhibited comparable or even better enzyme inhibitory activity than that of the positive control benquitrione. The greenhouse herbicidal activity experiments revealed that the tested compounds showed growth inhibition against the six tested weeds, especially for compound 9-28 , which achieved more than 80% inhibition against the six species of target weeds at the dosage of 150 g/hm2 and 100% inhibition against Echinochloa crus-galli and Digitaria sanguinalis. Furthermore, the AtHPPD- 9-28 complex was obtained, which explained the binding mode between the inhibitor and target enzyme, and brought a novel thought to facilitate a detailed understanding for the discovery of new HPPD inhibitors with improved performance. -
图 1 目标化合物9-28与AtHPPD的复合物晶体结构
A. 化合物9-28与AtHPPD的结合模式；B. 化合物9-28的电子云密度轮廓图；C. 化合物9-28与活性腔腔口处氨基酸残基的相互作用图；D. AtHPPD-9-28与AtHPPD-喹草酮的叠合图。
Figure 1. Co-crystal structure of AtHPPD-9-28
A. Binding mode of compound 9-28 with AtHPPD; B. 2Fo-Fc map (contoured at 1.0 σ) of compound 9-28; C. Interaction between compound 9-28 and the residues around the entrance of the active pocket; D. Superposition of the binding modes of compound 9-28 and benquitrione in AtHPPD.
表 1 目标化合物9-1~9-30的温室除草活性 (150 g/hm2) 和对AtHPPD的IC50值
Table 1. Greenhouse herbicidal activities of compounds 9-1-9-30 at 150 g/hm2 and IC50 values against AtHPPD
化合物
Compd.R 抑制率b
Inhibition rateb/%AtHPPD 抑制活性
AtHPPD inhibitory activity稗草
E. c.a狗尾草
S. v.马唐
D. s.苋菜
A. r.藜
C. s.苘麻
A. t.IC50/(μmol/L) 9-1 -CH3 B C C D B B 0.295 ± 0.002 9-2 -CH2CH3 C A C A B C 0.227 ± 0.022 9-3 -CH2CH2CH3 B B B C C B 0.208 ± 0.010 9-4 -CH2CH2CH2CH3 D E D D E E 0.311 ± 0.009 9-5 -CH(CH3)2 C C C B B A 0.183 ± 0.009 9-6 -CH(CH3)CH2CH3 D B E B B B 0.172 ± 0.014 9-7 -CH2CH2F C A B D D E 0.254 ± 0.018 9-8 -CH2CHF2 B B B D D D 0.274 ± 0.018 9-9 -CH2CF3 C B B D D E 0.336 ± 0.033 9-10 -CH2CH2CH2F B A B C D B 0.228 ± 0.007 9-11 -CH2CH2CF3 B B B D E E 0.294 ± 0.014 9-12 -CH2CH2OCH3 C D D D D A 0.253 ± 0.017 9-13 -CH2CH=CH2 B B E B A B 0.176 ± 0.001 9-14 -CH2C≡CH A B C B B A 0.190 ± 0.002 9-15 -CH2CH=C(CH3)2 D E F E A F 0.182 ± 0.016 9-16 
E B D C E E 0.463 ± 0.015 9-17 
B B E C D D 0.238 ± 0.005 9-18 
B B B A D B 0.157 ± 0.008 9-19 
A B B B A C 0.169 ± 0.002 9-20 
D C E C E E 0.172 ± 0.011 9-21 
C D E D D D 0.208 ± 0.010 9-22 
E E C D D D 0.332 ± 0.010 9-23 
D D E D A E 0.211 ± 0.002 9-24 
E F E D E E 0.136 ± 0.003 9-25 
F F F F F F 0.155 ± 0.008 9-26 
E D E D D D 0.160 ± 0.012 9-27 
D E D F D D 0.183 ± 0.017 9-28 
A B A B B B 0.370 ± 0.026 9-29 
B B D A A C 0.199 ± 0.010 9-30 
B B B A A C 0.247 ± 0.013 喹草酮 B B B B A B 0.375 ± 0.012 注(Note)：a Echinochloa crus-galli (E. c.), Setaria viridis (S. v.), Digitaria sanguinalis (D. s.), Amaranthus retroflexus (A. r.), Chenopodium serotinum (C. s.), and Abutilon theophrasti (A. t.). b A = 100%; B ≥ 80%; C ≥ 60%; D ≥ 40%; E ≥ 20%; F < 20%. 
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