Design , synthesis and antimicrobial evaluation of s-triazinyl urea and thiourea derivatives

A series of urea and thiourea derivatives of s-triazine have been developed based on high yielding nucleophilic substitution of 2,4,6-trichloro-1,3,5-triazine by 4-hydroxy coumarin, cyclopropylamine and ammonia at suitable conditions. These were further treated with various substituted aryl isocyanate and aryl isothiocyanate. All the synthesized compounds were evaluated for their antibacterial activities against various Gram-positive and Gram-negative strains of bacteria. A few compounds showed good to superior in vitro antibacterial activity against S.aureus, B.subtilis, E.coli and P.aeruginosa respectively. The new synthesized compounds were characterized using IR, H-NMR and elemental analysis.


Introduction
The exploration of heterocycles as privileged structures in drug discovery is, beyond doubt, one of the major areas in medicinal chemistry.These privileged structures represent a class of molecules that act as ligands for various biological receptors with a high degree of binding affinity.Problems of multi-drug resistant microorganisms have reached on alarming level in many countries around the world.A numbers of recent clinical reports describe the increasing occurrence of meticillin-resistant S. aureus and other antibiotic-resistant human pathogenic microorganisms in United State and European countries.Infections caused by those microorganisms pose a serious challenge to the medical community and the need for an effective therapy has led to a search for novel antimicrobial agents.Exploitation of these molecules should allow us to rapidly discover new biologically active compounds across a broad range of therapeutic areas in a shorter time scale.
In this work, we report the synthesis and biological activity of some 1,3,5-triazinyl urea and thiourea analogues a class of privileged structures that have a wide range of biological properties.6] The above literature survey led us to consider the s-triazine nucleus as a possible scaffold.
Coumarins constitute an important class of compounds with several types of pharmaceutical agents possessing anticancer, anti-HIV, anticoagulant and antiproliferative activity. 17] In the design of new compounds, development of hybrid molecules through the combination of different pharmacophores in one structure may lead to compounds with increased antimicrobial activity.Therefore, these observations prompted us to synthesize new s-triazine derivatives which were attached with coumarin ring through an oxygen bridge.Then, the synthesized compounds were tested against two Gram-positive bacteria (Staphylococcus aureus, Bacillus subtilis), two Gram-negative bacteria (Pseudomonas aeruginosa, Escherichia coli) using the broth microdilution method.

Materials and Methods
All reagents were of analytical grade and use directly.All the melting points were determined in open glass capillary and are uncorrected.Progress of reaction was monitored by thin layer chromatography (TLC) using silica gel-G coated aluminium plates (0.5 mm thickness, Merck) and spots were visualized under UV radiation, purified by recrystallisation and column chromatography.The IR spectra were recorded on BRUKER TENSOR Series using KBr pellets. 1 H NMR spectra were recorded on 300MHz BRUKER ULTRASHIELD using DMSO-d 6 as a solvent and TMS as an internal reference and chemical shift values were expressed in δ ppm.

General procedure for the preparation of 2-(Coumarinyl-4-oxy)-4-(cyclopropylamino)-6-(phenyl thioureido)-s-triazine (6a-j)
A mixture of (4) (1.5 g, 0.005 mol) and aryl isothiocyanate (0.005 mol) in ethanol (30 mL) was refluxed for 5-6 hours.The progress of reaction was monitored by TLC using toluene: acetone (6:4) as eluent.After the completion of reaction, the solvent was removed by distillation and the resulting solid was recrystallized from methanol.For antibacterial activity, in present protocol 25 µg mL -1 is considered as moderate activity, 12.5 µg mL -1 is considered as good activity and 6.25 µg mL -1 is considered as excellent active compound to the standard drug.

Results and Discussion
The present paper is focused on the synthesis of novel heterocyclic compounds as possible antibacterial agents.The minimum inhibitory concentration (MICs, µg mL -1 ) of tested compounds against bacteria are shown in Table I.All the synthesized compounds exhibited moderate to excellent inhibitory effect with MIC values against four strains of bacteria (Grampositive, Gram-negative).The ureido linkage two chloro substituted (5c and 5d) and methoxy substituted (5i) compounds showed excellent activity against Gram-positive B. subtillus and S. aureus bacteria.Among the thioureido linkage compounds, (6d and 6f) displayed excellent activity against Gram-positive B. subtillis and S. aureus organism.In general, it is concluded that the presence of electron withdrawing group increases the antibacterial activities compare to the electron donating group to the aromatic ring.Based upon this observation it becomes necessary to optimize lead compound with chloro substitution for augmenting antibacterial prob.Based upon the results, it will also be necessary to optimize by the substituting a series of electronwithdrawing groups on aromatic ring and selectively modifying the s-triazine nucleus.The substitution in the C-3 and C-4 positions of the phenyl ring seems to be very important for antibacterial effect, as well as the presence and the position of oxo-linkage in the connecting linker between the aromatic rings seems to be very important for antibacterial effect.
20with gradual dilution starting from (100, 50, 25, 12.5, 6.25, 3.125) µg/mL.Penicillin and Streptomycin were used as reference antibacterial agents.Solution of the test compounds and reference drugs were dissolved in DMSO.