Design,Synthesis And Antifungal Activity Evaluation Of Phloroglucinol Derivatives

In recent decades,although great progress has been made in antifungal therapy,antifungal drug resistance has become more and more serious.Infectious diseases caused by fungal infections are still one of the major health problems in the world.Therefore,discovery of broad-spectrum and low toxicity antifungal drugs is an important target for new drug research.Pseudoaspidinol is a phloroglucinols derivative with antifungal activity and is the main active ingredient of Dryopteris fragrans and Dryopteris crassirhizoma.A class of phloroglucinol derivatives represented by Pseudoaspidinol is highly active against Trichophyton rubrum,Trichophyton mentas,Candida albicans,Staphylococcus aureus,Escherichia coli,etc,and have selective inhibition on epidermal-like cancer cells and have antioxidant effects.Although there are various extracted methods of phloroglucin derivatives,it is necessary to synthesize Pseudoaspidinol owing to the limited content of Pseudoaspidinol in Dryopteris fragrans.However,there are very few reported synthetic methods for phloroglucinols derivatives.Total synthesis of natural products is one of the hotspots concerned by chemists nowadays.In previous study,the research group synthesized Pseudoaspidinol and phloroglucin derivatives,which showed good antifungal effects.This study further optimizes the synthetic route of Pseudoaspidinol,and designe phloroglucin derivatives using Pseudoaspidinol and its key intermediate as lead compounds,synthesize these compounds,investigate their antifungal activity,and explore their interaction with related proteins by molecular docking.The main contents and results are as follows:The 2,4,6-trihydroxy benzaldehyde was synthesized using phloroglucin,phosphorus oxychloride as raw materials,N,N-dimethylformamide as catalyst,then was reducted to2-methylbenzene-1,3,5-triol by sodium cyanoborohydride.The2-methylbenzene-1,3,5-triol was transferred to 1-?2,4,6-trihydroxy-3-methylphenyl?butan-1-one through acylation reaction under anhydrous aluminum trichloride and nitrobenzene,which replaces 3-position and5-position hydrogen groups to synthesize 3,5-bis?benzyloxy?-2-?but-1-en-2-yloxy?-6-methylphenol by the action of benzyl chloride,then to obtain 2-methoxy-4,6-dibenzyloxy-3-methylphenyl-butanone under anhydrous potassium carbonate and dimethyl sulfate,which was reducted to Pseudoaspidinol by hydrogen reduction.The improving synthetic route of Pseudoaspidinol is as follows:1.2,4,6-trihydroxybenzaldehyde use zinc powder as electronating agent,diethyl ether and ethyl acetate as the mixed solvent.2.The inversion of1-?2,4,6-trihydroxy-3-methylphenyl?butan-1-one to 4-butyryl-5-methoxy-6-methyl-1,3-phenylene bis?2-methylpropanoate?use EDCI and DMAP as catalysts,isobutyric acid as protective group.3.The 4-butyryl-5-methoxy-6-methyl-1,3-phenylenebis?2-methylpropanoate?,3-phenylenebis?2-methylpropionate?was synthesized under the principle of acid-base neutralization.The intermediates 2-methylresorcinol of Pseudoaspidinol and Naftifine were used as the lead compounds,and the C1-C4 acyl group was introduced at the 6-position to obtain 2,4,6-trihydroxy-3-methylbenzaldehyde,1-?2,4,6-Trihydroxy-3-methylphenyl?ethan-1-one,1-?2,4,6-Trihydroxy-3-methylphenyl?propan-1-one.Subsequently,The?E?-2-?4-Aminobut-2-en-1-yl?-4-methylbenzene-1,3,5-triol is obtained by introduction of cis-4-chloro-2-butene amine group to the 4-position of2-methylbenzene-1,3,5-triolby Friedel-Calcylation reaction,then introduce a C1-C4 acyl group at the 6-position to give?E?-3-?4-Aminobut-2-en-1-yl?-2,4,6-trihydroxy-5-methyl benzaldehyde,?E?-1-?3-?4-Aminobut-2-en-1-yl?-2,4,6-trihydroxy-5-methylphenyl?ethan-1-one,?E?-1-?3-?4-Ami nobut-2-en-1-yl?-2,4,6-trihydroxy-5-methylphenyl?propan-1-one.Then the introduction of methyl and methylnaphthalene on the nitrogen of?E?-1-?3-?4-aminobut-2-en-1-yl?-2,4,6-trihydroxy-5-methylphenyl?butan-1-one is to obtain?E?-1-?2,4,6-trihydroxy-3-methyl-5-?4-?methylamino?but-2-en-1-yl?phenyl?butan-1-one,?E?-1-?3-[4-?dimethylamino?but-2-en-1-yl]-2,4,6-trihydroxy-5-methylphenyl?butan-1-one,?E?-1-?2,4,6-trihydroxy-3-methyl-5-?4-??naphthalen-2-ylmethyl?amino?but-2-en-1-yl?phenyl?butan-1-one,?E?-1-?3-?4-?bis?naphthalen-2-ylmethyl?amino?but-2-en-1-yl?-2,4,6-trihydroxy-5-methylphenyl?butan-1-one and the target compound?E?-1-(2,4,6-trihydroxy-3-methyl-5-?4-?methyl?naphthalen-2-ylmethyl?ami nbut-2-en-1-yl?phenyl?butan-1-one.The structures of compounds were confirmed by MS,¹H NMR,¹³C NMR,and the synthetic route has been optimized to some extent.The antifungal activity of phloroglucinol derivatives against Trichophyton rubrum and Trichophyton mentagrophytes was determined by MTT method.The results showed that most of the compounds are more active than Pseudoaspidinol,of which?E?-1-?2,4,6-trihydroxy-3-methyl-5-?4-??naphthalen-2-ylmethyl?amino?but-2-en-1-yl?phenyl?butan-1-one has strong activity against two species of fungi with the minimum inhibitory concentration against T.rubrum of 5.12?g/mL and T.mentagrophytes of 8.13?g/mL.The target compound?E?-1-(2,4,6-Trihydroxy-3-methyl-5-?4-?methyl?naphthalen-2-ylmethyl?aminbut-2-en-1-yl?phenyl?butan-1-one has the strongest antifungal activity against Trichophyton rubrum and Trichophyton mentagrophytes with IC₅₀ of3.05?g/mL and 5.16?g/mL,respectively.The final result proves that the phloroglucin skeleton and butyryl group of Pseudoaspidinol,as well as the allylamine group of Naftifine and methylnaphthalene contribute to the antifungal activity.According to the effects of phloroglucin derivatives on the three target enzymes such as lanosterol 14?-demethylase,squalene epoxidase and? -1,3-glucan synthase,based on the evaluation of the antifungal activity of a series of phloroglucin derivatives including Pseudoaspidinol,molecular docking experiment were carried out to explore interaction with related proteins.The results showed that most of the phloroglucin derivatives had interaction with lanosterol 14?-demethylase,squalene epoxidase and?-1,3 glucan synthase.Among them,?E?-1-(2,4,6-Trihydroxy-3-methyl-5-?4-?methyl?naphthalen-2-ylmethyl? aminbut-2-en-1-yl?phenyl?butan-1-one has the strongest docking ability.The compounds interact with hydrogen bonds,hydrophobic interactions,and Pi-Pi.In order to understand the binding mode in the dynamic environment,the molecular dynamics simulation of the protein crystal structure of the target compound was carried out.It was confirmed by the ligand root mean square deviation?RMSD?that the target compound exhibited a stable binding conformation throughout the MD operation.It is consistent with the results of antifungal activity experiments.In conclusion,this topic has fully synthesized Pseudoaspidinol,and effectively optimized the total synthetic route.Using the intermediates2-methylbenzene-1,3,5-triol and Naftifine as lead compounds,22phloroglucin derivatives are synthesized,in which 10 novel phloroglucinol derivatives are unreported.The synthesis methods were optimized with the best synthesis method and good yield.The antifungal activity of the synthetic phloroglucinol derivatives against Trichophyton rubrum and Trichophyton mentagrophytes were evaluated,and most of the compounds had strong antifungal activity.