Design And Synthesis Of Novel Sulfonamide Derivatives And Their Antimicrobial Study

Sulfonamides containing p-amino benzenesulfonamide structure as the first type of artificial synthetic antimicrobial drugs were extensively used for the prevention and cure of bacterial infections in human biological systems as early as 80 years ago.As the analogues of aminobenzoic acid,sulfonamides could compete with it to availably prevent the synthesis of nucleic acids and proteins,and then inhibit the growth of various microorganisms.Further research has found that the classical sulfonamides possessed not only antibacterial activities,but also antifungal,anticancer,antiparasitic,anti-inflammatory,antidiabetic,antiepileptic and diuretic activities after structural modification.Nowadays,several sulfonamide derivatives has been successfully developed and employed in clinic,including Sulthiame(antiepileptic drug),Celecoxib(painkiller),Furosemide(diuretic drug)and the supramolecular agents of Ag-sulfadiazine and Ag-sulfadiazine(burn drug)and so on.Therefore,numerous efforts have directed toward the further developments of the novel sulfonamide derivatives in other medicine fields.Meanwhile,a great number of azole compounds including various five-membered azoles such as imidazoles,triazoles and tetrazoles,and benzene fused azoles like benzimidazoles and benzotriazoles with aromaticity are able to interact readily with bioactive sites such as enzymes and receptors via coordination bonds,hydrogen bonds and so on,and they could beneficially modulate the physicochemical and pharmacokinetic properties.The design,synthesis and evaluation of antimicrobial activity of azole derivatives have become one of the highly active highlights and many considerable outstanding achievements have been obtained.In this thesis,based on the current situation in the researches of sulfonamides compounds,a series of novel sulfonamide derivatives were designed and synthesized.All the newly synthesized compounds were evaluated for their antibacterial and antifungal activities.The preliminary structure-activity relationships,preparative methods and conditions were also discussed.Further binding behaviors between the synthesized active compounds and human serum albumin(HSA)were investigated by fluorescence and UV-vis absorption spectroscopy to evaluate their transportation and pharmacokinetic properties.The preliminary antimicrobial mechanism was also studied.The main work was summarized as follows:(1)Preparation of novel benzimidazole sulfonamide analogs: Commercially available acetanilide and chlorosulfonic acid produced N-protected sulfonyl chloride II-2 at high yield in ice bath,which was then further treated by sodium sulfite and sodium bicarbonate(the molar ratio is 1)to give p-acetylamino benzenesulfinic acid sodium salt II-3 at 80 oC.The latter was subsequently reacted with chloromethyl benzimidazole to afford benzimidazole intermediate II-4.The N-alkylation of benzimidazole ring of compound II-4 with halobenzyl halides,alkyl bromides and carbazole bromide in acetonitrile using potassium carbonate as base afforded the target sulfonamide analogs II-5a–h,II-6a–i and II-7.Finally,these compounds were further transformed into the deprotected benzimidazole derivatives II-8a–h,II-9a–i and II-10 in ethanol in the presence of sodium hydroxide in order to explore their effect on the bioactivities.(2)Preparation of novel sulfonamide–derived 1,2,4-triazoles: Commercial acetaniline was reacted with chlorosulfonic acid to produce the intermediate N-protected sulfonyl chloride III-2,and then underwent aminolysis to generate p-acetaminobenzene sulfonamide III-3 at 0 oC.Compound III-3 was N-alkylated with a series of halobenzyl halides in the first synthetic route to afford the corresponding secondary amino derivatives III-4a–e,and then further treated with 1,2-dibromoethane to produce the desired tertiary amino sulfonamides III-5a–e.The latter was subsequently reacted with 1,2,4-triazole to afford triazole-based sulfonamide intermediates III-6a–e.The second synthetic route,through protecting the amino group with the Boc moiety and coupling with p-acetaminobenzene sulfonamide III-3,the Boc-protected sulfonamide III-8 and compound III-9 were prepared,and compound III-8 was reacted with 1,2-dibromoethane to afford the tert-N-sulfonamide III-10.Subsequently,compound III-10 was reacted with 1,2,4-triazole through N-alkylation to conveniently afford the wanted sulfonamide intermediate III-11.Moreover,the undeprotected compound III-12 would separate in this procedure.The secondary amine III-11 could produce intermediate III-6f–g through the reaction with a series of halobenzyl halides.Finally,both of the acylated compounds III-6a–g and III-11 could undergo hydrolyzation under reflux to give the deprotected sulfonamides III-7a–g and III-13 without halobenzyl group.(3)All the newly synthesized compounds were characterized by IR,1H NMR,13 C NMR,MS and/or HRMS spectra.(4)All the target compounds were evaluated for their in vitro antimicrobial activities.The biological assays manifested that some prepared compounds exhibited good or even superior antibacterial and antifungal activities against the tested strains to the reference drugs Chloromycin,Norfloxacin and Fluconazole.Among the novel benzimidazole sulfonamide analogs,compounds II-5c and II-5g showed strong inhibitory activities against all tested strains with low inhibitory concentrations in range of 4–64 ?g/mL.Especially,towards Gram-positive bacteria,compound II-5c bearing 4-fluorobenzyl group displayed equivalent suppressive potency against S.aureus(MIC = 4 ?g/mL)and MRSA(MIC = 2 ?g/mL)to Chloromycin.Towards Gram-negative bacteria,compound II-5g bearing 2,4-dichlorobenzyl group activity was 8-fold more potent to clinical Chloromycin against B.typhi(MIC = 4 ?g/mL).Towards fungi C.utilis,deprotected sulfonamide II-8g gave the highest antifungal activity(MIC = 2 ?g/mL)among these prepared conjugates,which was 8-fold more potent than Fluconazole.Moreover,among the novel sulfonamide–derived 1,2,4-triazoles,compound III-7c with 2-chlorobenzyl group showed the strongest inhibitory activities towards MRSA,B.subtilis and E.coli(DH52 and JM109)strains with MIC values ranging from 8 to 64 ?g/mL,which was 2 to 4 fold more potent than the reference drug Chloromycin.It also exhibited equivalent activity with MIC value of 2 ?g/mL against C.mycoderma in comparison with Fluconazole.(5)The preliminary structure-activity relationships showed that the length of alkyl chain and the type,number and position of substituents on benzene ring had significant influence on biological activities.The introduction of benzimidazole and 1,2,4-triazole fragments were significant to the biological activities.(6)The preliminary interactive investigations of compounds II-5c and II-5g with calf thymus DNA by fluorescence and UV-vis spectroscopic methods revealed that compounds could effectively intercalate DNA to form compound II-5c or II-5g–DNA complex which might block DNA replication,and thus exert their antimicrobial activities.In addition,they could also form complexes by non-covalent bonds with HSA.Experimental results also displayed that compound III-7c could combine with DNA form compound III-7c–DNA complex which might further block DNA replication to exert their powerful antibacterial and antifungal activities.(7)The highly bioactive compounds II-5c,II-5g and III-7c were further examined for cytotoxic properties on HepG2 or MCF-7 lines.By applying fluorescence microscopy,it was demonstrated that the target compound III-7c with potent antimicrobial activity showed lower ROS generation than H2O2.With the increase of concentrations of the active compounds II-5c,II-5g and III-7c,there were obviously decreased trends to the cell viability.(8)Molecular docking indicated that the hydrogen atom connected to the nitrogen atom and oxygen atom in the sulfonyl group of compound II-5g formed two hydrogen bonds with the guanine of DNA,thus preventing the formation of hydrogen bond between cytosine in DNA.And the backbone of Ala 317 was inclined to form a hydrogen bond with one of oxygen atoms at the sulfonyl group in compound III-7c,where the distance was 2.70 ?.These results pointed out that the importance of the sulfonyl fragment might be the crucial reason for compounds II-5c,II-5g and III-7c displayed strong inhibitory efficacy against test strains.(9)In computational chemical studies,the highly bioactive compounds II-5c,II-5g and III-7c possessed more negative charge regions(in red)on the nitrogen atom and/or oxygen atom in the sulfonyl group.It might indicate the capability of hydrogen bond formation with active compounds.This was in agreement with the binding mode obtained from above docking study.