Accepted Manuscript

Synthesis and biological evaluation of novel 2, 4, 6-triazine derivatives as an‐

timicrobial agents

Sandip N. Gavade, Vijay L. Markad, Kisan M. Kodam, Murlidhar S. Shingare,

Dhananjay V. Mane

PII: S0960-894X(12)00723-8

DOI: http://dx.doi.org/10.1016/j.bmcl.2012.05.111

Reference: BMCL 19156

To appear in: Bioorganic & Medicinal Chemistry Letters

Received Date: 2 April 2012

Revised Date: 16 May 2012

Accepted Date: 30 May 2012

Please cite this article as: Gavade, S.N., Markad, V.L., Kodam, K.M., Shingare, M.S., Mane, D.V., Synthesis and

biological evaluation of novel 2, 4, 6-triazine derivatives as antimicrobial agents, Bioorganic & Medicinal Chemistry

Letters (2012), doi: http://dx.doi.org/10.1016/j.bmcl.2012.05.111

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1

Synthesis and biological evaluation of novel 2, 4, 6-triazine derivatives as

antimicrobial agents

Sandip N. Gavade a, Vijay L. Markad

b, Kisan M. Kodam

b, Murlidhar S. Shingare

c,

Dhananjay V. Mane a,*

a P.G. Department of Chemistry and Research Center, S.C.S. College, Omerga-413 606, India.

b Biochemistry Division, Department of Chemistry, University of Pune, Pune-411007, India.

c Department of Chemistry, Dr. Babasaheb Ambedkar Marathwada University, Aurangabad-431 004, India

*Corresponding author

Tel. +91 240 2403134 Fax: +91 240 2403134;

E-mail: dr_dvmane@rediffmail.com

2

Abstract

A series of 2, 4, 6-trisubstituted [1, 3, 5] triazines were synthesized and evaluated for

their antimicrobial activity against two representative Gram-positive, Gram-negative

bacteria and two fungi. Biological data revealed that among all the compounds screened,

compounds 3f, 3g, 3h, 3i, 3m, 3o and 3p found to have promising antimicrobial activity

against all the selected pathogenic bacteria and fungi. Out of the synthesized compounds

seven analogues have shown MIC in the range of 6.25-12.5 µg/mL. These compounds

were generally nontoxic and may prove useful as antimicrobial agents.

Keywords

Triazine derivatives, Anti-bacterial activity, Anti-fungal activity, MIC.

3

The treatment of bacterial infections remains a challenging therapeutic problem

because of emerging infectious diseases and the increasing number of multidrug-resistant

microbial pathogens. Despite the many antibiotics and chemotherapeutics available, the

emergence of old and new antibiotic-resistant bacterial strains in the last decades leads to

a substantial need for new classes of anti-bacterial agents.1 The increasing incidence of

infection caused by the rapid development of bacterial resistance to most of the known

antibiotics is a serious health problem.2 While many factors may be responsible for

mutations in microbial genomes, it has been widely demonstrated that the incorrect use of

antibiotics can greatly increase the development of resistant genotypes.3 As multidrug-

resistant bacterial strains proliferate, the necessity for effective therapy has stimulated

research into the design and synthesis of novel antimicrobial molecules. The chemistry of

[1,3,5]triazine compounds has been studied intensively and is the subject of many

reviews.4 The triazine scaffold has provided the basis for the design of biologically

relevant molecules with broad biomedical value as therapeutics. For example, triazine

compounds possess potent antiprotozoal,5 anticancer,

6 antimalarial,

7 and antiviral

activity.8 Also, it was reported that some of these compounds possess potent

antimicrobial activity.9 These compounds have been used in the treatment of

depression,10

and hence received a considerable therapeutic importance. These are

valuable bases for estrogen receptor modulators11a

and also used as bridging agents to

synthesize herbicides.11b

Further substituted s-triazines have been used as NLO materials,

which have a wide range of applications in optoelectronics and telecommunications.12

The starting material for these compounds is cyanuric chloride. This is an inexpensive

commercially available reagent which makes its use more attractive. Increased interest in

this scaffold lies in the different reactivities of the substituent chlorine atoms, which are

controlled by temperature.4 This allows sequential introduction of various substituents

into the [1,3,5]triazine ring. In order to further expand the scope of triazine derivatives as

privileged medicinal scaffold, we have been engaged in design, synthesis and

antimicrobial activity evaluation of novel 2, 4, 6-trisubstituted [1,3,5] triazine derivatives.

We encouraged by recently reported antimicrobial activity of 2-fluorophenyl-4,6-

disubstituted [1,3,5]triazines13

and new S-triazine based chalcones and their derivatives.14

4

Herein we disclose our results antibacterial and antifungal activity of novel 2, 4, 6-

triamino [1,3,5] triazine derivatives.

Our synthetic strategy for the novel 2,4,6-trisubstituted-[1,3,5]-triazine derivatives

is illustrated in Scheme 1. The key intermediates 5-[(4,6-dichloro-1,3,5-triazin-2-

yl)amino]pyridine-2-carbonitrile 1 was synthesized by 2,4,6-trichloro-1,3,5-triazine

(cyanuric chloride) by reacting them with 5-amino 2-cyano pyridine in the presence of

acetone at 0 ⁰C – rt for 6h in 78% yield. The trisubstituted triazines (2a-2r) were

synthesized by reaction of the 5-[(4,6-dichloro-1,3,5-triazin-2-yl)amino]pyridine-2-

carbonitrile (1) with different nucleophiles (R, Fig. 1) in the presence of K2CO3 in DMF

at rt for 5h in 63-86% yield. Trisubstituted triazine hydrochloride (3a-3r) was made by

using 50% HCl in Dioxane at rt for 2h. The purity of the compounds was checked by

TLC and HPLC. Spectral data 1H NMR,

13C NMR and MS of the newly synthesized

compounds (3a-3r) was consistent with the proposed structures.

In order to search for the potent compound, the newly synthesized compounds

(3a-3r) were evaluated for their antibacterial and antifungal activity against various

Gram-positive, Gram-negative bacteria and fungal strains using agar well diffusion

method. The antimicrobial evaluation data is represented in Table 1. As can be seen from

our results, many compounds from the newly synthesized series found to be potent

antibacterial and antifungal agents. Thus the compounds 3d, 3f, 3g, 3h, 3i, 3l, 3m, 3n, 3o

and 3p (Table 1) exhibited comparable antibacterial and antifungal activity than the

standard Streptomycin and Nystatin respectively, against all the tested bacteria or fungi.

Interestingly, the compounds 3f, 3g, 3h, 3i, 3m, 3o and 3p exhibited potent to same

antibacterial and antifungal activity respectively as that of the standard Streptomycin and

Nystatin. The compounds 3f, 3g, 3h, 3m and 3o found to be potent among the series and

even comparable with standard Streptomycin as antibacterial agent against some bacteria

viz. Bacillus subtilis (MTCC 441), Staphylococcus aureus (MTCC 96), Escherichia coli

(MTCC 1650) and Pseudomonas aeruginosa (MTCC 1688). Compounds 3f, 3g, 3i and

3o found to be good antifungal agents and was comparable with standard Nystatin as

antifungal agent against some fungi viz. Aspergillus niger (MTCC 1781) and Candida

albicans (MTCC 227). The remaining compounds of this series were found to have

moderate or low activity.

5

Among all the compounds tested, seven compounds showed MIC in the range of

6.25–12.5 µg/mL whereas six compounds showed a MIC of 12.5-50 µg/mL. The results

showed a good structure–activity relationship. Results also shows the importance of

lipophilicity and basicity in the antibacterial and antifungal activity of the synthesized

compounds (3a–r). Antibacterial activity of 1,3,5-Triazine having R as N-methyl

piperazine 3d showed a MIC of 12.5 µg/mL whereas when the methyl group was

replaced with acetyl 3k, benzoyl 3l and phenyl group 3m it showed a MIC of 50 µg/mL,

12.5 µg/mL and 6.25 µg/mL respectively. Compound 3m has good activity as compare to

3d and 3k, it means that activity increases with increase in lipophilicity and decreasing

basicity of the compounds. Morpholine substituted compound 3a showed a MIC of 50

µg/mL whereas when morpholine was replaced with 4-(2-aminoethyl) morpholine 3j

MIC increased to 25 µg/mL. When the morpholine group in 3i and 3o was replaced with

1H-pyrazol-3-amine 3i and 1-(furan-2-yl)methanamine 3o the activity increased showing

a MIC of 6.25 µg/mL. These results emphasize the role of aromaticity and better efficacy

of 1H-pyrazol-3-amine and 1-(furan-2-yl)methanamine moiety over morpholine moiety

in the antibacterial activity. In the compounds 3a and 3j the same relation between

lipophilicity and basicity with activity was observed. With R as cyclohexylamine 3f

showed a good MIC whereas slightly decreasing the ring size to cyclopentylamine 3h

activity reduced against some bacteria. Branching in the alkyl chain also favour the

activity. When R as a 2-methoxy ethyl amine 3g and N,N-dimethyl ethyl amine 3n

showed a increase in activity having a MIC of 6.25 and 12.5 µg/mL. When R as a

aromatic amine electron donating group 3p showed more antibacterial activity than the

electron withdrawing group 3q.

In case of antifungal activity of 1,3,5-Triazine having R as cyclohexylamine 3f

showed a MIC of 12.5 µg/mL and whereas R as cyclopentylamine 3h showed a MIC of

25 µg/mL. In the compounds 3g, 3i and 3o the same relation between lipophilicity,

basicity and aromaticity with activity was observed. 3g, 3i and 3o showed athe MIC of

12.5 µg/mL.

The twenty 2,4,6-trisubstituted-1,3,5-triazines (3a–r) were synthesized evaluated

for their antimicrobial activity. Out of the synthesized compounds seven analogues have

shown MIC in the range of 6.25-12.5 µg/mL. The compounds 3f, 3g, 3h, 3i, 3m, 3o and

6

3p were found to potent antibacterial and antifungal agents. Thus the presence of

aromaticity and lipophilicity found to have strong relevance to the antimicrobial activity.

These identified triazines can be new leads in antimicrobial chemotherapy. These

molecules are very useful for further optimization work in microbial chemotherapy.

Acknowledgments

The authors acknowledge the financial support from UGC, [F. No. 39-774/2010

(SR)], New Delhi and are grateful to Principal of S.C.S. College, Omerga for providing

the necessary laboratory facilities.

References and notes

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Table 1. Antimicrobial activity of novel 2, 4, 6-triazine derivatives (MICa values are in

µg/mL)

Comp.

No.

Antibacterial activity

Antifungal activity

Grampositive bacteria

Gramnegative bacteria

Bacillus

subtilis

Staphylococcus

aureus

Escherichia

coli

Pseudomonas

aeruginosa

Aspergillus

niger

Candida

albicans

3a

50 (12)

50 (12)

50 (12)

50 (10)

100 (14)

50 (14)

3b 50 (10) 50 (12) 25 (14) 25 (12) 25 (14) 50 (10)

3c 50 (12) 50 (12) 50 (12) 50 (10) 100 (14) 25 (14)

3d 12.5 (12) 12.5 (12) 12.5 (12) 12.5 (10) 50 (12) 50 (10)

3e 100 (12) 100 (12) 50 (12) 100 (12) 50 (14) 50 (12)

3f 6.25 (12) 6.25 (12) 6.25 (12) 6.25 (10) 12.5 (14) 12.5 (14)

3g 6.25 (12) 12.5 (12) 6.25 (12) 6.25 (12) 12.5 (14) 12.5 (12)

3h 6.25 (12) 6.25 (12) 6.25 (10) 12.5 (12) 25 (12) 25 (12)

3i 6.25 (10) 12.5 (12) 6.25 (12) 12.5 (14) 12.5 (14) 12.5 (14)

3j 25 (10) 50 (12) 25 (14) 25 (12) 50 (14) 25 (12)

3k 50 (12) 50 (12) 50 (12) 50 (10) 100 (14) 25 (14)

3l 12.5 (10) 12.5 (12) 25 (12) 25 (14) 50 (12) 50 (14)

3m 6.25 (10) 6.25 (12) 6.25 (12) 6.25 (10) 25 (14) 25 (14)

3n 25 (12) 12.5 (12) 12.5 (14) 12.5 (12) 25 (12) 25 (14)

3o 6.25 (10) 6.25 (12) 6.25 (14) 6.25 (12) 12.5 (12) 12.5 (12)

3p 12.5 (10) 6.25 (12) 6.25 (14) 12.5 (12) 25 (12) 25 (12)

3q 25 (10) 25 (12) 25 (14) 50 (12) 25 (12) 25 (12)

3r 25 (12) 12.5 (12) 12.5 (12) 25 (10) 100 (14) 25 (14)

Streptomycin 6.25 (22) 6.25 (20) 6.25 (22) 6.25 (24) NTb NT

Nystatin NT NT NT NT 6.25 (20) 6.25 (20)

a MIC values were evaluated at concentration range between 6.25 and 200µg/ml.

b NT: Not Tested.

MIC (µg/mL) and zone of inhibition (mm) in parenthesis.

Figure: 1

Scheme 1. Synthesis of novel 2, 4, 6-triazine derivatives.

Synthesis and biological evaluation of novel 2, 4, 6-triazine derivatives as

antimicrobial agents

Sandip N. Gavade a, Vijay L. Markad

b, Kisan M. Kodam

b, Murlidhar S. Shingare

c, Dhananjay

V. Mane a,*

a P.G. Department of Chemistry and Research Center, S.C.S. College, Omerga-413 606, India.

b Biochemistry Division, Department of Chemistry, University of Pune, Pune-411007, India.

c Department of Chemistry, Dr. Babasaheb Ambedkar Marathwada University, Aurangabad-431 004, India

A novel series of 2, 4, 6-triazine derivatives have been designed, synthesized and evaluated for

antimicrobial (antifungal and antibacterial) activities against selected pathogenic strains.