Design And Synthesis Of Novel Triazoles And Their Antimicrobial Study

Triazole compounds are an important class of nitrogen-containing aromatic heterocycles with electron-rich system,which are widely present in many biologically active compounds.This type of unique structure endows triazole-based derivatives to readily interact with various enzymes and receptors in biological system via weak non-covalent interactions,thus displaying a wide range of biological activities.Ever since the development of fluconazole,a large number of predominantly triazole-based compounds have been frequently used in clinic as drugs or condidates for the treatment of various types of diseases,which have shown their huge development value and great potential as medicinal agents.Their important clinical uses have been motivating extensive efforts to construct new bioactive molecules based on the triazole fragment.The triazole ring can be identified as an important isostere of imidazole,oxazole,pyrazole,thiazole,benzimidazoles and amide moiety etc.and widely used in designing and developing versatile bioacitive molecules.Triazole ring has been extensively used as an attractive linker to combine different pharmacophore fragments to produce innovative bifunctional drug molecules and triazole ring with mutiple binding sites is extensively employed in designing supramolecular drugs.Based on the current situation in the researches of triaozle compounds,a series of novel triazole derivatives were designed and synthesized.All the new compounds were evaluated for antimicrobial activities.The preliminary structure-activity relationships,preparative methods were discussed.To evaluate their transportation and pharmacokinetic properties,further binding behaviors between the synthesized active molecule and human serum albumin(HSA)were investigated by UV-vis absorption and fluorescence spectroscopy.The preliminary antimicrobial mechanism was also discussed.The main work was summarized as follows:(1)Preparation of novel Schiff base-bridged triazolyl berberine derivatives:Berberine has been commonly used in clinics to treat intestinal infections and the incidence of resistance to berberine is reported rarely despite its long term clinical use and possesses especially widespread application prospects in medicinal chemistry.Starting from commercially available berberine hydrochloride,berberrubine II–2 was easily obtained by the 9-demethylation of berberine hydrochloride II–1 at 190 °C under vacuum,and was further reduced by NaBH4 in methanol to give tetrahydroprotoberberine(THPB)II–3 at 0 °C.The formylation of compound II–3 by hexamethylenetetramine(HMTA)in trifluoroacetic acid(TFA)under reflux and by hydrolysis with 10% H2SO4 at 90 °C produced the important intermediate II–4 without further purification.The latter was condensed with 4H-1,2,4-triazol-4-amine in ethanol under reflux using a catalytic amount of glacial acetic acid to afford the target Schiff base II–5.The further structural modification of compound II–5 by alkyl or aralkyl halides generated the alkyl THPBs II–6a-f and aralkyl THPBs II–7a-h in DMF at 80 °C using potassium carbonate as base.(2)Preparation of novel Schiff base-bridged triazole carbazoles:Carbazole heterocycle is present in a variety of naturally occurring medicinally active substances and also exhibit the large potential applications of carbazole-based derivatives.Starting from commercial carbazole material,the N-alkylation of carbazole III–1 with various alkyl bromides in DMF at room temperature using sodium hydride as base respectively afforded the N-alkyl carbazole compounds III–2a–h.The formylation of N-alkylcarbazoles III–2a–h was carried out by direct Vilsmeier–Haack reaction employing DMF and phosphorus oxychloride to afford N-ethyl-carbazole-carbaldehyde III–5 and N-alky-carbazole-dicarbaldehydes III–3a–h.Further Shiff base-linked target bistriazole derivatives III–4a–j and monotriazole derivatives III–6a–b and III–7a–d were smoothly obtained by condensation reaction of formyl carbazole with 4H-1,2,4-triazol-4-amine and 4-amino-4H-1,2,4-triazole-3-thiol respectively.Besides,aldehyde III–3a reacted with hydroxylamine to give the corresponding oxime derivative III–8 and further treatment of the comppound III–8 with 4H-1,2,4-triazol-4-amine obtained the target derivative III–9.(3)All the newly synthesized compounds were characterized by 1H NMR?13C NMR?IR and HRMS spectra.(4)The in vitro antibacterial and antifungal assays showed that most target THPB triazoles in the series II gave better inhibitory potency than the precursor berberine and some of them were much more active than the reference drugs.Among the alkyl derivatives,the butyl compound II–6b and the hexyl analogue II–6c gave broader antibacterial spectrum and better activities with MIC values of 1–32 ?g/mL.Compound II–7a without substitution on the phenyl ring gave the best antibacterial efficiencies with MIC values of 1–32?g/mL towards the corresponding bacteria,especially compound II–7a exhibited 8-and 4-fold more activity towards MRSA with lower concentration(MIC = 2 ?g/mL)than chloromycin and norfloxacin,respectively,which had the potency to be a lead molecule in the development of more effective antimicrobial agents with a broad spectrum.The N-butyl-carbazole-triazole compound III–4i could significantly inhibit growth of A.flavus(MIC = 1 ?g/mL),which was more potent than clinical fluconazole.It also showed effective anti-MRSA activity(MIC = 4 ?g/mL),better than chloromycin(MIC = 16 ?g/mL)and norfloxacin(MIC = 8 ?g/mL).The N-penty-lcarbazole triazole derivative III–4d gave the best inhibitory potency against M.luteus with MIC value of 0.5 ?g/mL.(5)The structure-activity relationship(SAR)study revealed that the combination of the triazole fragment and THPB was beneficial to exert biological activity,and the substituents on the benzyl moiety as well as the length of the alkyl chain at the 9-position of THPB influenced the antibacterial potency.Bis-triazolyl carbazoles possessed better antibacterial efficacy than mono-triazolyl ones.Proper length of alkyl chain exerted an important influence in enhancing bioactivity and either the decrease or increase of the chain length was unfavorable for the bioactivity.(6)Time-kill kinetic assays manifested that compound II–7a could rapidly kill MRSA and bacterial resistance assays against MRSA indicated that MRSA was more difficult to develop resistance against compound II–7a than the clinical drug norfloxacin.Molecular docking indicated that target compound II–7a could bind with MRSA DNA through hydrogen bonds.The binding investigation of compound II–7a with HSA revealed that this molecule could be effectively transported by HSA.The preliminary exploration for the antimicrobial mechanism disclosed that compound II–7a could not only form a stable II–7a–DNA complex with calf thymus DNA by an intercalating mode,but also directly cleave pUC19 DNA by formation of a complex with a Zn2+ ion,thus exerting antibacterial activity.(7)The highly bioactive compound III–4i was examined for cytotoxic properties using MTT assay,which showed low toxicity to human hepatocyte LO2 lines.The interaction of high active molecule III–4i with calf thymus DNA displayed compound III–4i could intercalate into the double helix of DNA by substituting with neutral red(NR)in the DNA-NR complex which might block DNA replication to lead to the death of the strain.Molecular docking indicated that compound III–4i could bind with DNA topoisomerase IB through hydrogen bonds,which correlated with the inhibitory effect.Fifty three compounds were successfully synthesized in this thesis.Thirty one compounds were new,including fifteen THPB triazoles,sixteen Schiff base linked N-alkylcarbazole triazoles.