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INTERNATIONAL JOURNAL OF FRONTIERS IN SCIENCE AND TECHNOLOGY www.ijfstonline.org Research Article ISSN 2321 – 0494 Indexed in CAS, OPEN J-gate and GOOGLE SCHOLAR Received on: 10.3.15, Revised and Accepted on:24.3.15

Cyanuric Chloride based Chalcones for Biological Applications: Synthesis, Characterization and Antimicrobial Studies J.Suresh, E.Vakees, S. Karthik, A. Karthikeyan and A.Arun* PG & Research Department of Chemistry, Government Arts College, Tiruvannamalai-606603. Tamilnadu, India.

Abstract An efficient antimicrobial compound is synthesized based on chalcone and their acrylates are synthesized, characterized and tested for their antimicrobial activity against two different microorganisms.3-(4-((4-(4-acetylphenyl)amino)-6-(napthalen-1-yloxy)-1,3,5-triazin-2 yl)oxy)phenyl)-1-(2,4-dichlorophenyl)prop-2-en-1-one (CAP) is synthesized using cyanuric chloride,

2,4-Dichloro-1-ene(4-hydroxyphenyl)

phenone

(DHP),

1-napthol

and

4-

aminoacetophenone. Novel chalcones (CAC 1, CAC2, CAC3, CAC4 and CAC5 ) are synthesized using CAP and substituted benzaldehyde. The synthesized chalcones are characterized by IR, 1

H-NMR and UV-visible spectroscopic techniques. The antimicrobial activities of the

chalcones and acrylates are determined using Minimum Inhibitory Concentration (MIC) method and found to be very high for tested bacterial strains (1.95±0.2µg/ml). Keywords: Cyanuric chloride; Chalcone; Acrylate; Antimicrobial activity. * Corresponding Author, A.Arun .

,E-mail address: [email protected]

I.Introduction The chemistry of chalcones has generated intensive scientific interest due to their biological and industrial applications. Chalcones are natural biocides and are well known intermediates in the synthesis of heterocyclic compounds exhibiting various biological activities. Chalcones and their derivatives possess some interesting biological properties such as Jan-Mar-2015

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antibacterial, antifungal, insecticidal, anesthetic, antiinflammatory, analgesic etc [1-9]. Cynuric chloride is a class of compounds, which has many applications in different field. Antimicrobial Activity: Nagaini et al, successfully synthesized a new

series of chalcone derivatives as

antimicrobial agents and he found that the position of functional group will have a pronounced effect on the anti-bacterial effect against E. coli. [10]. Swamy et al., reported the antimicrobial activity of 3-hydroxy benzofuran substituted chalcones. It was evident that most of the compounds are very weakly active and few are moderately active against Staphylococcus aureus and Escherichia coli but they possessed very good activity against fungi Aspergillusflavus [11]. Similarly, Mayekar et al., reported a series of chalcones and its cyclohexanone derivatives derived from 6-methoxy-2-naphthaldehyde. It showed comparatively good activity against all the bacterial and fungal strains like Escherichia coli, Staphylococcus aureus, Pseudomonas aeruginosa, when compared to standard drugs like Ampicillin [12]. Tavares et al., evaluated a series of new 6-quinolinyl and quinolinyl N-oxide chalcones to tested on the survival and growth of the human cancer cell lines UACC-62 (melanoma), MCF-7 (breast), TK-10 (renal) and leukemic cells, Jurkat and HL60 [13]. Simillarly, Goto et al., reported the synthesize chalcone from commercially available 2, 4, 6-trihydroxytoluene.Which showed a unique highly potent anti-HIV activity. In this article we have synthesized a triazine based chalcones using DHP (14), CAP and substituted benzaldehydes. Antimicrobial activities of (CAC1, CAC2, CAC3, CAC4 and CAC5 ) were tested on four different bacteria strains. 2.

Experimental Methods

2.1.

Materials 2,4-dichloroacetophenone and 4-aminoacetophenone were purchased from Merck and

used as such. 4-hydroxybenzaldehyde and Cyanuric chloride were purchased from Aldrich chemical and used as such. All the solvents were purchased from S.D. Fine chemicals and used as such. Benzaldehyde, 4-nitrobenzaldehyde, 4-bromobenzaldehyde, 4-chlorobenzaldehyde, para N,N dimethylbenzaldehyde and 4-aminobenzaldehyde were received from SD fine chemicals. 2.2.

Synthesis of 2,4-Dichloro-6-(naphthalene-1-yloxy)-1,3,5-triazine (scheme 2) Cyanuric chloride (5g mol) dissolved in acetone was added slowly to the solution of

alpha naphthol (3.908g mol) dissolved in acetone present in a conical flask at 0 0C. The PH of the

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reaction mixture was maintained at 6 by the addition of aqueous Na 2CO3 (2 M), the reaction was carried out for 3h. After completion, the reaction mixture was poured into ice-cold water. The precipitate was separated out by filtration and washed several times by using cold water and dried in vacuum at 50 0C.

IR (KBr,cm-1): 3305 (N-H str.), 808 (C-N str., s-triazine moiety),

1080 cm-1 (C-Cl str.) ; 1H NMR(CDCl3, δ, ppm): 8.03 (1H, d, Ar-CH=), 6.98 –7.91 (17H, m, Ar-H and -NH). 2.3. Synthesis of 3-(4-(4-chloro-6-(naphthalene-1-yloxy)-1,3,5-triazin-2-yl)oxyphenyl)-1(2,4-dichlorophenyl)prop-2-en-1-one (scheme 3) CNP (3g mol) dissolved in acetone was added slowly to DHP (3.008g mol in 30mL acetone) with constant stirring for 3hr at 35 0C and periodically

10% Na2CO3 solution was

added dropwise to be neutralized. After completion, reaction mixture was poured into ice-cold water, the precipitate was separated out by filtration and washed several times by using cold water and dried in vacuum at 50 0C. IR (KBr,cm-1): 3305 (N-H str.), 3070 (=CH str.), 1649 (C=O str.), 1510 (C=C str.), 1025 (C-O-C str.), 808 (C-N str., s-triazine moiety), 1080 cm-1 (CCl str.) ; 1H NMR(CDCl3, δ, ppm): 6.42 (1H, d, -CO-CH=), 8.03 (1H, d, Ar-CH=), 6.98 –7.91 (17H, m, Ar-H and -NH). 2.4.

Synthesis of 3-(4-((4-((4-acetylphenyl) amino)-6-(naphthalen-1-yloxy)-1,3,5-triazin-

2-yl)oxy)phenyl)-1-(2,4-dichlorophenyl)prop-2-en-1-one (scheme 4) CDP (2g mol) dissolved in acetone was added slowly to the solution of 4aminoacetophenone (0.493g mol) dissolved in acetone contained in 250ml RB flask. To this solution, 10% Na2CO3 solution were added and the reaction was carried out 3hr at 60 0C. After completion, reaction mixture was cooled and poured into ice-cold water. The precipitate was separated out by filtration and washed several times by using cold water and dried in vacuum at 50 0C.

IR (KBr,cm-1): 3405 (N-H str.), 3065 (=CH str.), 809 (C-N str., s-triazinemoiety), 825 (

C-H bending), 1645(C=O str, ), 1266 (C-O-C str.), 1095 (C-Cl str.) ; 1H NMR (CDCl3, δ, ppm): 3.81 (3H, s, p-OCH3), 5.1 (2H, s, -NH2), 6.85 (1H, s, -CH).

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Scheme 1. Synthesis of 3-(4-((4-((4-acetylphenyl) amino)-6-(naphthalen-1-yloxy)-1,3,5-triazin-2yl)oxy)phenyl)-1-(2,4-dichlorophenyl)prop-2-en-1-one 2.5.

Synthesis of triazine containing novel chalcones (scheme 5) Grindstone technique Newly synthesized chalcones were prepared by grinding together equivalent amount of

CPE and substituted benzaldehydes in presence of KOH in a porcelain mortar under solvent free conditions for 4-8mins. On completion of reaction, the mixture was diluted with cold water neutralized by dilute HCl and recrystallized from acetic acid.

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Scheme 2. Synthesis of Chalcones CAC1, CAC2, CAC3, CAC4, CAC5 and CAC6. IR (KBr,cm-1): 3405 (N-H str.), 3065 (=CH str.), 809 (C-N str., s-triazinemoiety), 825 ( C-H bending), 1645(C=O str,), 1266 (C-O-C str.), 1H NMR (CDCl3, δ, ppm): 3.81 (3H, s, pOCH3), 5.1 (2H, s, -NH2), 6.85 (1H, s, -CH). 2.6.

Antimicrobial activity All synthesized compounds were screened for their antibacterial activity by MIC method.

The following bacterial strains and procedure were used. S. a – Staphylococcus aureus MTCC3381 E. c – Escherichia coli MTCC739 2.6.1.

Minimum Inhibitory Concentration (MIC)

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The minimum inhibitory concentration (MIC), which is considered as the least concentration of the sample which inhibits the visible growth of a microbe was determined by the broth dilution method. 2.6.2. Preparation of inocula Organisms were subcultured on nutrient agar, followed by incubation for 24h at 37 °C. Inocula were prepared by transferring several colonies of microorganisms to sterile nutrient broth. The suspensions were mixed for 15sec and incubated for 24h at 37 °C. Required volume of suspension culture was diluted to match the turbidity of 0.5 Mc Farland standard (1.5x108CFU/mL). 2.6.3.

Preparation of sample Samples were prepared in dimethylsulphoxide (DMSO) at the concentration of 2 mg/ml.

2.6.4. Broth dilution assay A series of 15 tubes were filled with 0.5 ml sterilized nutrient broth. Sequentially, test tubes 2–14 received an additional 0.5 ml of the sample serially diluted to create a concentration sequence from 500 – 0.06μg. The first tube served as the control. All the tubes received 0.5ml of inoculum. The tubes were vortexed well and incubated for 24h at 37°C. The resulting turbidity was observed, and after 24h MIC was determined to be where growth was no longer visible by assessment of turbidity by optical density readings at 600nm. No chemicals were used. The experiments were triplicates. 3.Result and Discussion Table 1. FT-IR and 1H NMR values, Molecular Weight, Melting Point and % Yield of the synthesized compounds. IR cm-1

Compound

1

H NMR

M/Wt

Melting Point Yield% DHP 297

1643 (CO), 1597 (–CH=CH–, str.), 92-93

82

1338 (C–N), 811 (C–N, s- triazine)

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7-6.91 (d, 2H, –CO–CH=CH]),

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7.15–7.80 (m, 10 H Ar–H).

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780 (C-Cl). CAC1 735.61

7-6.91 (d, 2H, –CO–CH=CH]),

1645 (CO), 1595 (–CH=CH–, str.), 170-172

78

1338 (C–N), 806 (C–N, s- triazine)

7.15–7.80 (m, 23H Ar–H).

780 (C-Cl). CAC2 770.06

7-6.90 (d, 2H, –CO–CH=CH]),

1647 (CO), 1597 (–CH=CH–, str.), 191-192

70

1338 (C–N), 809 (C–N, s- triazine)

7.15–7.85 (m, 22Ar–H).

786 (C-Cl), 1555 (Ar-NO2). CAC3 814.51

7-6.91 (d, 2H, –CO–CH=CH]),

1646 (CO), 1596 (–CH=CH–, str.), 215-216

67

1338 (C–N), 811 (C–N, s- triazine)

7.15–7.80 (m, 22 Ar–H ).

789 (C-Cl), CAC4 750.63

180-182

65

1338 (C–N), 813 (C–N, s- triazine)

7.15–7.80 (m, 22Ar–H and 2NH).

780 (C-Cl). CAC5 778.08

7-6.90 (d, 2H, –CO–CH=CH]),

1648 (CO), 1598 (–CH=CH–, str.),

.

1648 (CO), 1598 (–CH=CH–, str.), 185-187

7-6.90 (d, 2H, –CO–CH=CH]),

65

1338 (C–N), 813 (C–N, s- triazine)

7.15–7.80 (m, 22Ar–H ).

780 (C-Cl). CAC6 780.61

1648 (CO), 1598 (–CH=CH–, str.), 191-193

7-6.90 (d, 2H, –CO–CH=CH]),

65

1338 (C–N), 813 (C–N, s- triazine)

7.15–7.80 (m, 22Ar–H).

780 (C-Cl). 550 (NO2)

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The novel six chalcones were prepared according to scheme 5. All the synthesized compounds were characterized by IR and 1H NMR techniques to confirm the structure; the obtained data’s were presented in the table 1. Melting points of the synthesized compounds were measured using open capillary method and were incorrect. The antimicrobial activities of the synthesized compounds were evaluated using minimum inhibitory concentration (MIC) method. 3.1.

Antimicrobial activity Totally six novel chalcones were synthesized containing with and without chlorine

content in it. All the compounds were derived from the parent cyanuric chloride and DHP. The MIC values reported in the table 2 and data showed that all the synthesized compounds were active against Escherichia coli than the Staphylococcus aureus with the exception for CAC4 sample. There was a remarkable difference in MIC values between the parental cyanuric chloride and synthesized compounds. The synthesized compound CAC3 showed greater activity towards Staphylococcus aureus with a value of 7.81 (g/ml) and compounds, CAC4, CAC5 and CAC6 showed similar high activity against Escherichia coli. The synthesized compound DHP and CAC2 showed highest activity of 1.95 and 3.91 (g/ml) respectively towards gram negative bacteria Escherichia coli clearly suggesting that the presence of chlorine atoms in the chalcone enhance the antimicrobial activity. The synthesized compound CAC4, CAC5 and CAC6 show poor activity than the parental cyanuric chloride, indicate the absence or least availability of chlorine or hydroxyl group in those compounds. Fig 10 shows the antimicrobial activity of the compounds CY, DHP, CAC1, CAC2, CAC3, CAC4, CAC5 and CAC6 against two tested microorganism.

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Table 2: MIC values of the synthesized compounds Minimum Inhibiting Concentration (g/ml) Samples

Melting

Escherichia coli

CY

31.25

31.25

DHA

15.63

3.95

CAC1

15.63

12.91

CAC2

1.95

31.25

CAC3

12.25

7.81

CAC4

62.5

62.5

CAC5

31.25

31.25

CAC6

62.5

31.25

Temperature oC

3.2.

Staphylococcus aureus

Point 1000 MWt

500

MP 0 DHP

CAC1

,CAC2

CAC3

CAC4

CAC5

CAC5

Fig 12: Effect of Molecular weight on the Melting Point of the DHP, CAC1, CAC2, CAC3, CAC4, CAC5 and CAC6.

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Melting points of all the synthesized compounds were determined in an open capillary method. Fig 11 shows correlation between melting point of the synthesized compounds DHP, CAC1, CAC2, CAC3, CAC4, CAC5 and CAC6 with that of the molecular weight. Melting point of the synthesized compound increases by increasing molecular weight. The molecular weight of the compounds were 297, 735.61, 770.06, 814.51, 780.61, 778.08 and 780.61 respectively and the melting point of these compounds were 90-92, 172, 192, 216, 182, 187 and 193oC. 3.3.

Solubility of the synthesized compounds The solubility data of the synthesized compounds are presented in the table 3. The

chalcones DHP, CAC1, CAC2 are almost soluble in all the solvents used for solubility test except water, benzene and n-hexane but freely soluble in methanol, ethanol, dimethylsulfoxide, dimethylformamide and tetrahydrofuran.

MIC (µg/mL)

80 60 40

S.a

20

E.c

0

CHALCONES

Fig. 11: MIC value bar graph of DHP, CAC1, CAC2, CAC3, CAC4, CAC5 and CAC6.

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Table 3: Solubility Data of the novel chalcones at 300C

Polymer

H2 O

MeOH EtOH

CHCl

DMSO DMF

Aceton

C6H6

THF

e

3

DHP

_

+

+

+

+

+

+

CAC1

_

+

+

+

+

+

+

CAC2

_

+

+

+

+

+

CAC3

_

+

±

±

+

CAC4

-

±

±

±

CAC5

_

+

±

CAC6

_

+

±

nHexane

+

_

_

+

_

+

_

+

_

+

+

_

+

_

+

±

±

-

±

_

±

+

+

+

_

+

_

±

+

+

+

_

+

_

+ = Soluble, - = Insoluble and ± = partially soluble. 4.

Conclusion There are six novel compounds were synthesized by substituting various molecules in to

the cyanuric chloride. All the synthesized compounds were characterized by FT-IR, NMR and UV spectroscopy techniques. All the synthesized chalcones were completely soluble in polar protic solvents like acetone, alcohol, DMSO, THF, ether but insoluble in water. Melting points of all the synthesized compounds were determined in an open capillary and found to be dependent on the molecular weight of the compounds. Antimicrobial activity of CY, DHP, Jan-Mar-2015

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CAC1, CAC2, CAC3, CAC4, CAC5 and CAC6 were tested on Staphylococcus aureus and Escherichia coli. The synthesized compound a1 showed excellent activity towards the tested bacterial strains Escherichia coli with the value of 1.95 g/ml. Similarly, compound b111 showed greater activity towards Staphylococcus aureus with a value of 7.81 (g/ml).

5. References 1. Vibhute Y.B. and Basser M.A., Synthesis and activity of a new series of Chalcones asantibacterial agents, Ind.J. of Chem., 2003,42B, 202-205. 2. Bhat B.A., Dhar K.L., Saxena A.K., Shanmugavel M., Synthesis and biological evaluation of Chalcones and their derived Pyrazoles as potential cytotoxic agents, Bioorg. & Med. Chem., 2005, 15 (3), 177-3180 3.

Michael L. Edwards, David M. Stemerick, and Prasad S. Sunkara, Synthesis of Chalcones: A new class of Antimitotic agents, J. of Med. Chem., 1990, 33, 1948- 54.

4. Kalirajan

R.,

Palanivelu

M.,

Rajamanickam

V.,

Vinothapooshan

G.

and

Anandarajagopal K., Synthesis and biological evaluation of some Chalcone derivatives, Int. J. of Chem. Sci,. 2007, 5(1), 73-80 5. P Prasanna raja et al /Int.J. ChemTech Res.2010,2(4) 2004 6. Udupi R. H, Bhat R. and Krishna Kumar, Synthesis and biological activity of Mannich bases of certain 1, 2-Pyrazolines, Indian J. of Het.Chem., 1998, .8, 143-146 7. Alka Pande and Saxena V. K., Synthesis&Antiviral activity of 4 (Arylhydrazono)-3methyl-1-(3, 5-dinitrobrnzoyl)-2-pyrazolin-5-ones, Ind.J.of Chem.,1987,26B,390-392 8. Urmila Gupta, Vineeta Sareen, Vineeta Khatri, Sanjana Chugh, Synthesis and antifungal activity of new Fluorine containing 4-(substituted Phenyl azo) Pyrazoles and Isoxazoles, Indian J. of Het.Chem., 2005; 14:265-266 9. Pandey V.K., Gupta V.D. and Tiwari D.N., Synthesis of Substituted Benzoxazines as potential Antiviral agents, Indian J. of Het.Chem., 2004, 13, 399-400 10. Rakesh Mani Mishra and Abdul Wahab, Synthesis and Fungicidal activity of some new 2, 3-Dihydro-4H-Benzimidazolo [3, 2-b] - [1,3] - Thiazine-4-ones, Indian J. of Het.Chem., 2003, 13, 29-32

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11. Mayekar AN. Synthesis, Characterization and Antimicrobial Study of Some New Derivatives. Inter. J. Chem.2010; 2:114-123. 12. Ruiz C, Haddad M, Alban J Bourdy G, Reategui R, Castillo D, Sauvain M, Deharo E, Estevez Y, Arevalo J, Rojas R. Activity guided isolation of antileishmanial compounds from Piper hispidum. Phytochemistry Letters. 2011; 4:363-366. 13. Ahmed MR, Sasttry VG, Bano N, Ravichantra S, Raghavendra M. Synthesis

and

cytotoxic, anti oxidant activities of new chalcone derivatives. Rasayan J. Chem.2011; 4: 289-294.

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