Synthesis And Activity Evaluation Of Difluorobenzonitrile Derivatives

Among numerous compounds,nitrogen-containing and nitrogen-containing heterocyclic compounds have become a highly attractive class of compounds due to their diverse biological activities.Many drugs,such as antibacterial and anticancer drugs,contain pyrazole ring and isoxazole ring in their molecular structures.Therefore,the modification,synthesis,and activity research of nitrogen heterocycles or nitrogen containing side chains in compounds has become an important research field.3,4-difluorobenzonitrile is the prodrug of cyhalofop,an acetyl coenzyme A carboxylase inhibitor.Its molecule contains fluorine atoms and cyano groups,which can improve the hydrophilicity and biological activity of the drug structure.Therefore,it is a strategy worth considering to design and synthesize a series of drug molecules with different biological activities using 3,4-difluorobenzonitrile as the parent core.This paper synthesized a series of 3,4-difluorobenzonitrile derivatives containing pyrazole and isoxazole rings,as well as a derivative containing a biguanidine group.The inhibitory effect and structure-activity relationship of the compound on dihydrofolate reductase and COX-2 enzyme were analyzed through molecular docking,and the inference was verified through antibacterial experiments,anti-tumor cell activity experiments,and insecticidal experiments.The main content is as follows:(1)A series of compounds containing pyrazole rings were synthesized using 3,4-difluorobenzonitrile as the starting compound.On the basis of compounds containing pyrazole rings,the side chains of isoxazole rings were expanded,and a compound containing a biguanidine group was also attempted for synthesis.Preliminary identification and content determination of compounds were carried out using thin layer chromatography or high-performance liquid chromatography,and the structure of the compounds was characterized using nuclear magnetic resonance hydrogen spectroscopy and infrared spectroscopy.(2)Molecular docking simulations were conducted using MOE(2015.10 version)software to simulate the compounds with dihydrofolate reductase and COX-2 enzyme,providing a theoretical basis for subsequent evaluation of antibacterial and anti-tumor activities.The results showed that compounds 5-6,12-14,and 15-17 had high binding rates with dihydrofolate reductase,which may have good antibacterial biological activity.Compounds 6 and 8,12-14 and 15-17 have high binding rates with COX-2 enzymes,which may have good anti-tumor cell biological activity.(3)The inhibitory activity of the synthetic compound against six strains of E.coli,S.aureus,P.aeruginosa,B.subtilis,S.typhimurium and C.albicans was evaluated using the Oxford cup method.The compounds 12-14 modified with pyrazole rings containing side chains of-3-en-2-one and compounds 15-17 containing isoxazole rings have lower MIC values and relatively high antibacterial activity against six tested strains.(4)The MTT method was used to evaluate the cell survival rate of synthetic compounds in Hep G2 liver cancer cells and A549 lung cancer cells.The results showed that pyrazole compounds had a relatively stronger inhibitory effect or cytotoxicity at low concentrations,while isoxazole compounds had a higher inhibitory effect or cytotoxicity than pyrazole compounds at high concentrations.(5)The insecticidal activity of compounds 1-17 against diamondback moth was evaluated using fipronil as a positive control drug.The results showed that compounds15,16,and 17 containing isoxazole rings had a higher killing rate against diamondback moth,while pyrazole ring compounds 12,13,and 14 with side chains of-3-en-2-one structure also exhibited good inhibitory activity.