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基于1,3-二羰基功能基导向的全新杀菌化学实体的设计合成与结构优化研究
Alternative TitleDesign, synthesis and structure optimization of novel antimicrobial chemical entities based on 1,3-dicarbonyl functional group
周勇
Subtype硕士
Thesis Advisor刘映前
2023-05-29
Degree Grantor兰州大学
Place of Conferral兰州
Degree Name药学硕士
Degree Discipline药学
Keyword构效关系 structure-activity relationship 1,3-二羰基化合物 structural optimization 结构优化 1,3-dicarbonyl compounds 抗菌活性 antimicrobials 作用机制 mechanism of action
Abstract

喹诺酮类抗生素是一类含4-喹诺酮基本结构的广谱抗菌药物,其作为药物研究的优势骨架,在新药研发中发挥着重要作用。喹诺酮类抗生素构效关系表明,其C-3位侧链上的羰基与C-4位上的羰基具有共平面性,是其发挥抗菌药效的活性必需官能团。因此,本论文以喹诺酮类抗生素的1,3-二羰基活性片段为结构单元,采用生物电子等排、氟化策略、“中间体衍生化”法以及氮杂环化等药物设计策略对其进行结构优化,设计合成一系列结构简单、易于合成的全新1,3-二羰基杀菌化学实体,并测试这些1,3-二羰基化合物对植物病原真菌的抗菌活性,此外,该研究还对高活性化合物进行初步作用机制研究。为测试1,3-二羰基化合物的抗菌谱,本研究进一步的测试其对植物病原细菌、人源致病真菌及细菌的抗菌活性,并充分探究其构效关系,为进一步寻找全新小分子抗菌剂奠定理论基础。本论文主要研究内容分述如下:

1. 氟化1,3-二羰基杀菌化学实体的设计合成与杀菌活性评价

通过对喹诺酮类抗生素药效团与抗菌构效关系分析,以其1,3-二羰基活性片段为结构单元,设计合成了一系列1,3-二羰基化合物并测定其对灰葡萄孢菌、禾谷镰孢菌、立枯丝核菌、辣椒疫霉菌、核盘菌和尖孢镰刀菌6种植物病原真菌的体外抗菌活性。研究发现,氟化1,3-二羰基化合物对6种植物病原真菌具有显著的抗菌活性,其中,化合物ET-60对6种植物病原真菌具有最优的广谱抗菌活性,其EC50值在0.635~4.515 μg/mL之间,化合物ET-29对6种植物病原真菌的广谱抗菌活性次之,EC50值在0.790~7.288 μg/mL之间。本章节进一步测试1,3-二羰基化合物的抗菌谱,结果显示,其对植物病原细菌、人源致病真菌及细菌均表现出较好的抗菌活性,其抗菌活性远高于部分商业化杀菌剂。其中,化合物ET-29对柑橘溃疡病菌、马铃薯黑胫菌和水稻白叶枯病菌3种植物病原细菌的MIC90值分别为0.78、1.56、1.56 μg/mL,对人源致病真菌(白色念珠菌)和细菌(金黄色葡萄球菌和大肠杆菌)的MIC90值分别为25、0.39、50 μg/mL,具有显著抗菌活性;化合物ET-60对金黄色葡萄球菌亦表现出优异的抗菌活性,其MIC90值为0.10 μg/mL,显著优于阳性对照药诺氟沙星。通过对1,3-二羰基化合物的体外抗菌活性评价和构效关系探究,我们最终获得两个广谱高活性先导化合物ET-29和ET-60,并将它们作为进一步修饰和优化的目标。

2. 二芳醚-氟化1,3-二羰基杀菌化学实体及其吡唑杂环的设计合成与杀菌活性评价

基于第二章的高活性先导化合物ET-29和ET-60,我们采用“中间体衍生化”法与活性官能团拼合策略引入苯醚甲环唑、氯氟醚菌唑等商业化杀菌剂的二芳醚官能团,设计合成了22个二芳醚-氟化1,3-二羰基化合物。进一步采用氮杂环化策略将部分上述二芳醚-氟化1,3-二羰基化合物的两个羰基进行合环,设计合成了19个二芳醚-吡唑杂环衍生物。通过测试上述41个化合物对灰葡萄孢菌、立枯丝核菌、禾谷镰孢菌、核盘菌和辣椒疫霉菌5种植物病原真菌的体外抗菌活性,研究发现,在化合物ET-29和ET-60中引入二芳醚功能基后,这些化合物对植物病原真菌仍然保持着较好的抗菌活性,其中,化合物EFM-75、EFM-76和EFM-77表现出广谱抗植物病原真菌活性,EC50值分别在1.373~5.773 μg/mL, 1.588~7.323 μg/mL和2.385~7.588 μg/mL之间。本章节进一步测试二芳醚-氟化1,3-二羰基化合物的抗菌谱,结果显示,其对金黄色葡萄球菌仍然保持着较好的抗菌活性,其中,EFM-79和EFM-80对金黄色葡萄球菌的抗菌活性最好,MIC90值均为0.20 μg/mL,显著优于阳性对照药诺氟沙星。此外,合环后的二芳醚-吡唑杂环化合物对立枯丝核菌保持着较好的抗菌活性,其中,化合物BZ-102表现出优异的抗菌活性,其EC50值为3.000 μg/mL。

3. 化合物ET-60对灰葡萄孢菌作用机制初步探究

在上述两章研究工作的基础上,研究发现,所有氟化1,3-二羰基化合物均对灰葡萄孢菌表现出较好的抗菌活性,因此我们选取广谱抗菌活性最好的化合物ET-60为供试药剂,以灰葡萄孢菌为供试菌种,以嘧霉胺(灰葡萄孢菌特效药)和啶酰菌胺(广谱药)为阳性对照药剂,对化合物ET-60进行初步的作用机制探讨与分析。研究结果显示,化合物ET-60使灰葡萄孢菌菌丝发生严重皱缩、细胞膜破损、线粒体功能异常造成灰葡萄孢菌细胞代谢紊乱而发挥抗菌功效。

Other Abstract

Quinolone antibiotics are a class of broad-spectrum antibacterial drugs containing the basic structure of 4-quinolone, which play an important role in the development of new drugs as a dominant framework for drug research. Researches indicate that the carbonyl groups on the C-3 side chain and the C-4 position of quinolones are essential functional groups for biological activities. According to the structure-activity relationship of quinolones, we finally synthesize a series of novel 1,3-dicarbonyl compounds, which possess the characteristics of simple structure and easy synthesis, by adopting drug design strategies such as bioelectronic isotopes in this paper.

The main contents of this paper are as follows:

1. Design, Synthesis and Antimicrobial Activity Evaluation of Fluorinated 1,3-dicarbonyl Compounds

A series of 1,3-dicarbonyl compounds were designed and synthesized by adopting drug design strategies such as bioelectronic isotopes based on the previous pharmacophore analysis of quinolones. By evaluating the antimicrobial activities of these novel 1,3-dicarbonyl compounds in vitro, we found that fluorinated 1,3-dicarbonyl compounds have significantly antimicrobial activities against six plant pathogenic fungi, named Botrytis cinerea, Fusarium graminearum, Rhizoctonia solani, Sclerotinia sclerotiorum, Fusarium oxysporum, and Phytophthora capsici, and against three plant pathogenic bacteria, named Xanthomonas Campestris pv. citr, Pectobacterium atroseptica ACCC 19901, and Xanthomonas oryzae pv. oryzae ACCC 11602, and against one human pathogenic fungus, named Candida albicans ATCC 24433, and against two human pathogenic bacteria, named Staphylococcus aureus Newman and Escherichia coli ATCC 25922. Fortunately, the entire fluorinated 1,3-dicarbonyl compounds exhibited better antimicrobial activity than oxquinic acid, especially for Botrytis cinerea. The EC50 values of compound ET-29 against six plant pathogenic fungi ranged from 0.790 to 5.968 μg/mL. The MIC90 values for three plant pathogenic bacteria were 0.78, 1.56 and 1.56 μg/mL, respectively. And the MIC90 values for human pathogenic fungi and bacteria were 25, 0.39 and 50 μg/mL, respectively. Compound ET-60 had the best broad-spectrum activity against plant pathogenic fungi among these compounds with EC50 values ranging from 0.635 to 4.515 μg/mL. Also, ET-60 had the best antibacterial activity against Staphylococcus aureus Newman with the MIC90 value of 0.10 μg/mL. By exploring the structure-activity relationship of 1,3-dicarbonyl compounds, we finally determined the highly active lead compounds ET-29 and ET-60 for further modification and optimization.

2. Design, Synthesis, and Antimicrobial Activity Evaluation of Diaryl Ether-fluorinated 1,3-dicarbonyl Compounds and the Pyrazole Heterocycles

Based on the above highly active lead compounds ET-29 and ET-60, we designed and synthesized 22 diaryl ether-fluorinated 1,3-dicarbonyl compounds by introducing the diaryl ether functional groups of commercial fungicides such as difluorocycloazole and chlorofluconazole using the "intermediate derivatization" and active functional group splicing methods. Further, using nitrogen heterocyclization to cyclize two carbonyls of some of the above diaryl ether-fluorinated 1,3-dicarbonyl compounds, and 19 diaryl ether-pyrazole heterocyclic derivatives were designed and synthesized. By testing the in vitro antimicrobial activity of the above 41 compounds against five plant pathogenic fungi, four plant pathogenic bacteria as well as three human pathogenic bacteria, which are the same as in the previous chapter except for weeding out Fusarium oxysporum and Pectobacterium atroseptica ACCC19901, and newly added pseudomonas solanacearum, and erwinia carotovora, we found that these compounds, which were designed by introducing diaryl ether functional groups into compounds ET-29 and ET-60, still maintained excellent antimicrobial activities against plant pathogenic fungi and Staphylococcus aureus Newman. And compounds EFM-75, EFM-76, and EFM-77 exhibited broad spectrum antifungal activity with EC50 values ranging from 1.373 to 5.773 μg/mL, 1.588 to 7.323 μg/mL and 2.385 to 7.588 μg/mL, respectively. The MIC90 values of EFM-79 and EFM-80 for Staphylococcus aureus were 0.20 μg/mL as well. Interestingly, the diaryl ether pyrazole heterocyclic derivatives maintained good antifungal activity against Rhizoctonia solani and compound BZ-102 was the best active one with an EC50 value of 3.000 μg/mL。

3. Preliminary Study on the Action Mechanism of Compound ET-60 on Botrytis cinerea

Based on the work in the previous two chapters, we found that all fluorinated 1,3-dicarbonyl compounds exhibited remakable antifungal activities against Botrytis cinerea. Therefore, we selected the compound ET-60 as the test chemical, which possessed the most antifungal activities, to study the preliminary mechanism of action in the Botrytis cinerea strain, and selected Pyrimethanil and Boscalid as the positive control. The results showed that compound ET-60 had certain protective effects on apples infected by Botrytis cinerea. Further researches on the mechanism found that compound ET-60 caused severe shrinkage of Botrytis cinerea hyphae, cell membrane damage, and mitochondrial dysfunction, resulting in metabolic disorders of Botrytis cinerea cells and eventual death.

Subject Area药物化学生物学
MOST Discipline Catalogue医学 - 药学(可授医学、理学学位) - 药物化学
URL查看原文
Language中文
Other Code262010_220200942561
Document Type学位论文
Identifierhttps://ir.lzu.edu.cn/handle/262010/539279
Collection药学院
Affiliation
兰州大学药学院
Recommended Citation
GB/T 7714
周勇. 基于1,3-二羰基功能基导向的全新杀菌化学实体的设计合成与结构优化研究[D]. 兰州. 兰州大学,2023.
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