Synthesis And Antibacterial Activity Of Cajaninstilbene Acid And Its Structural Analogue Benzyl Phenyl Sulfide Derivatives

It's an urgent need to develop new classes of antibacterial agents with distinct mechanism of action due to the increasing incidence of bacterial resistance to conventional antibiotics.Cajaninstilbene acid?CSA?is an important nature product which exhibits a wide range of important physiological properties.Recently,it was reported that CSA exhibits potent antibacterial activity against gram-positive bacteria such as a kind of“superbug”MRSA.CSA was thought as a new antibacterial chemical entity whose molecular structure is different from other antibacterial agents.The short supply of CSA,however,rendered a detailed biological assessment difficult.Moreover,the CSA itself has a limited antibacterial activity with a quite cytotoxicity,thus it's impossible to taking CSA as a medicine.Therefore,it's of great significance to synthesis CSA and its analogues that are effective against drug-resistant bacteria with a low cytotoxicity for the treatment of drug-resistant bacterial infections.Objective:Recent years,many studies about total synthesis of CSA and its analogues had been reported.Previously,a series of CSA derivatives have been synthesized in our group and many of them exhibited enhanced activity than CSA.However,the former synthetic routes are deficient and produced an unsatisfactory yield that remains to be improved.And our former medical chemistry study was far from sufficiency.Moreover,the potential cytotoxicity of many potent synthetic analogues is the major obstacle for developing CSA analogues as therapeutics.CSA shares a gross structural similarity with recently reported membrane-targeting antimicrobials i.e.they are all amphiphilic molecules.Similarly,the structure-activity relationships of our former synthetic CSA analogues indicated that the antibacterial activity deeply rely on the molecular amphiphilicity.Thus,we try to describe the mechanism of action of CSA analogues as a membrane-targeting antimicrobial.Under this condition,this study was focus on two parts:Firstly,taking a new strategy for effective synthesis of CSA and its analogues with high yields and short routes;Secondly,based on the previous study,we continued to synthesis several CSA analogues for finding compounds with better activity and lower toxicity and for preliminarily proving the hypothesis about the antibacterial mechanism of synthetic CSA analogues.Methods:Firstly,a new route for synthesis of CSA and its analogues has been designed.The difficulty for the synthesis of CSA is what has come to be known as the unsatisfactory C-prenylation reaction.Focusing on this,we adopted a strategy of pre-prenylation,i.e.the prenyl was introduced before the construction of the benzyl ring A.Started with a cheap commercial material methyl acetoacetate,a prenylation reaction on C4 site was carried out under excessive base.Then a TiCl₄-mediated cyclocondensation was performed as a key reaction to construct the ring A,and the products was hydrolyzed to affording a building block for synthesis of CSA.The building block was coupling with various aromatic aldehydes under a modified McMurry reaction condition to introduce the ring B.Then CSA and its derivatives were obtained by hydrolysis of the products of McMurry reaction.Secondly,after extensive literature review,it was found that the stilbene skeleton of CSA may be the main cause for its cytotoxicity,for this structure was found to exhibit remarkable DNA damage activity.For reducing the cytotoxicity and improving the antibacterial activity,the stilbene skeleton of CSA was changed by introducing of S atom to get benzyl?phenyl?sulfide skeleton.By using the organic synthesis,a series of benzyl?phenyl?sulfide derivatives were obtained.And their antibacterial activity was evaluated by minimum inhibitory concentration?MIC?assay.The in vitro cytotoxicity of representative benzyl?phenyl?sulfide derivatives were examined in human cardiomyocytes AC16 in terms of a maximum tolerated test?MTT?.The permeabilization of the bacterial cell membrane was studied using propidium iodide?PI?assay for proving the hypothesis about the antibacterial mechanism of benzyl?phenyl?sulfide derivatives.Results:?1?A three-step synthesis of a substituted salicylate that can be used as building block for late-stage introduction of styryl groups to synthesis CSA and its derivatives is completed.The synthesis of CSA was accomplished in six steps with an overall yield of 20%by a synthesis whose key steps are TiCl₄-mediated cyclization and McMurry coupling.?2?Totally,33 novel benzyl?phenyl?sulfides was designed and prepared,the antibacterial activities for Gram-positives and Gram-negative bacteria of those analogues have been evaluated.Results reveal that all the derivatives shown no activity to Gram-negative species,however,many of them displayed significant antibacterial activities against a panel of Gram-positives species,including MRSA.The MIC values of the most potent compounds 13f and 13h were in the2-8?g/mL rang,better than positive controls penicillin and norfloxacin.Importantly,13f,13h shown a lower cytotoxicity compared to their stilbene counterparts.?3?The mechanism of action investigation on 13f,13h revealed bacteria cell membrane permeabilization in MRSA 43300.Conclusions We had completed a six steps facile synthesis of CSA and its derivatives by using a pre-prenylation strategy.Our synthetic route is the most concise method with the higher yield for synthesis of CSA,and no protecting group was used.Currently,we were able to obtain the CSA and its derivatives easily,which will support further medicinal chemistry studies of CSA.The antibacterial activity and cytotoxicity of CSA structuralanalogue benzyl?phenyl?sulfide derivatives shown that the stilbene skeleton isn't the necessary structure for antibacterial activity however caused the cytotoxicity thus better to be replaced.The structure-activity analysis of synthetic benzyl?phenyl?sulfide derivatives and the bacterial membrane permeabilization activity of representative compounds indicated that the probable bacteriological mechanism of action is membrane-targeting mode of action.