Synthesis, Identification and Antibacterial Potency of Azo Dyes having Quinolin-8-ol and Active Methylene Moiety

Kanchan R. Damade1*, Jyotsana D. Shinde2, Dinesh J. Bhojsing2, Vijay S. Patil2, Bhaiya S. Chauhan2, Kiran R. Kapadane2 & Chandan I. Rajput2

1, 2 & * Department of Chemistry, Arts, Commerce and Science College Bodwad, Distt Jalgaon, Maharashtra , INDIA

* Correspondence: E-mail: Kanchan_Damade@rediffmail.com

(Received 09 June, 2018; Accepted 25 June, 2018; Published 30 June, 2018 )

ABSTRACT: In the present study we have reacted the five aryl diazonium salts prepared from nitro substituted anilines and amino substituted benzoic acids and coupled with Quinolin-8-ol and Active methylene group of pent-1,4-dione respectively. Synthesized compounds are then identified by FTIR Spectroscopical method. The final product, I to V formed has potential to use as azo dyes and as an intermediate in other synthetic procedures or transformations along with this they exhibits good biological activities .

Keywords: Active methylene group; diazonium salt; antibacterial activity; nitroanilines and azo dye.

INTRODUCTION: Azo compounds were under study from the long time for their remarkable partaking in pharmaceuticals, cosmetics and textiles industries. They have immense important biological activities and high therapeutic values for all mammals reported by Sisley and Porsche, 19111 . reductive cleavage of Azo group gives great therapeutic properties for treatment of serious disorders in human beings1,2,3 .Biological outcomes from an enzymatic metabolism (occurs invivo)4, 5 for example in skin bacteria such as Staphylococcus aureus6 which involves reduction of azo group (-N=N- ) to produce toxic or nontoxic resultant amines7 which may produce carcinogenic effects 8, 9. Despite of having negative role for environment and human health azo groups have attracted medical attentions. Later studies showed health hazards from these azo families in the form of carcinogenic and mutagenic properties, even though many of the synthetic strategies have been developed to produce these classes of compounds for their specific physico-chemical and biological activities. The azo coupling reaction with AMG results into formation of bioactive compounds like cinnolines which is one of the important class of natural products having remarkable pharmaceutical and biological importance10. Due to simple process in aqueous media we can produce wide varieties of azo dyes and can determined biological actions. In our study we synthesized novel azo compound in two different schemes forming diazonium salt of o and m substituted nitro anilines and substituted amino benzoic acids followed by coupling reaction with quinolin-8-ol and active methylene group of acetyl acetone respectively, so here in we reported some novel azo compounds and testing out their antibacterial potency on selected strains of bacterias like Staphylococcus aureus andBacillus subtilis with discussion of their infections .

Many authors have reported about the infections registered through S. aureus and B. subtilis bacterias in human body. Logan et al. 1988 stated the infections with bacilli species in impotent patients that is who are immunologically compromised. Similarly Kiss et al.1988 also reported the B. subtilis infections in enervate patients. Donzis et al. 1988 reported B. subtilis eye infections due to contaminated lenses. Penington et al.1976 described infections in patients having blood cancer.

Staphylococcus aureus is also one of the dangerous pathogen found in human. Rates of S.aureus infections are high among patients with type-1 diabetes,12 intravenous drug users,13 patients facing hemodialysis,14 patients having done with major surgery,15,16 and who are HIV positive17. Patients with leukaemia are also at increased risk for staphylococcal disease18. So therefore many studies have been done for preparing antibiotics for these organisms.

MATERIALS AND METHODS: All the chemicals were of standard grade and used without further purifications. Silica gel was used to monitor the progress of reactions by TLC and visualized under Iodine chamber. The colours of resultant dyes are recorded visually and melting point range were recorded by one end open capillary tube method. The purity of compounds was noted by melting point determination and silica gel-G TLC, elemental analysis (C,H,N), FTIR spectral data. The bacterial strains, Staphylococcus aureus and Bacillus subtilis are purchased from National Centre for Cell Science (NCCS), Pune, India and maintained at Smt. G. G. Khadse College, to determine the antibacterial activity of synthesized all five azo compounds (I-V).

Scheme 1: General procedure for synthesis of azo dyes20 (I-III)

Stage 1: Preparation of Diazonium Salt - In 100 ml Capacity beaker add 2 ml aniline (or its derivative) and to this mixture of 5ml conc. HCl and 10 ml water were added and stirred with the glass rod to get clear solution. Cool, the solution upto 0 C by keeping in freezing mixture. Dissolving (1gm) sodium nitrite in 10 ml water. Allowed to cool the solution in ice bath to 0 C, after attaining 0 C we added NaNO2 solution into aniline hydro chloride (or derivatives) solution drop wise with constant stirring (kept maintained temperature below 50C during addition). After decompose of excess of nitrous acid by adding pinch of urea filtered the solution and collect the filtrate which was diazonium salt of aniline and its derivatives.

img2

X= -H (I), 2-NO2 (II), 3-NO2 (III)

Stage 2: Diazonium coupling reaction with quinolin-8-ol - Prepared a mixture of solution of 1.5gm quinolin-8-ol in 10 ml 10% NaOH and allowed to cool up to 0C, after attaining 0C ,we added solution of diazonium salt dropwise in to quinolone-8-ol in NaOH solution with constant stirring, after complete addition allowed reaction mixture to stand for 10 min in ice bath, filtered the azo colourant and washed it with cold water, dry, weight and noted the yield of clear crude azo product then recrystallized by using solvent ethanol. Recorded the dried weight and the color , melting point range of compounds.

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(E)-5-[(nitro substituted phenyl) diazenyl] quinolin-8-ol

Scheme 2: General procedure for synthesis of azo dyes (IV & V)

Stage 1: preparation of diazonium salts of amino substituted benzoic acids - Same as discussed in scheme-1.

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Stage 2: Diazonium coupling reaction with AMG of Acetyl acetone (pent-2,4-dione) - Add (-o/-p) carboxy aryl diazonium salt solution drop wise, to the well cooled mixture of, Pentane-2,4-dione (1.8ml) which is dissolved in 5 ml ethanol and sodium acetate , 8-10 gm in 10-15ml of water and maintained the 00C temperature , a coloured precipitate is separated, then adding 20 ml of con. HCl the product is filtered, then by checking the absence of ester, the product obtained is recrystallized by using ethanol, dried it. Record the dried weight (in gms) and then melting point range of the compounds VI & V.

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2, 4-dioxopentan-3-yl) diazenyl) Substituted benzoic acid (IV & V)

Substitution on benzoic acid: 2-NH2 (IV) & 4-NH2 (V)


The synthesized azo compounds (I-V) were tested for their anti-bacterial activity against two strains of bacterias named Staphylococcus aureus and Bacillus subtilis by disc diffusion assay using solutions of azo compounds at two different concentrations of 500 & 1000 g/ml.

RESULTS AND DISCUSSION: In the present study diazonium salts of aniline (and nitro derivatives) in scheme-1 is coupled with Quinolin-8-ol which results into formation of compounds I19 (B.E. Ezema et,al. 2014) II & III 5-[(nitro substituted phenyl) diazenyl] quinolin-8-ol. Similarly in scheme-2, carboxy diazonium salt is coupled with AMG of Pent-2, 4-dione to form products IV & V (2,4-dioxopentan-3-yl)diazenyl) substituted benzoic acids. The synthesised compounds then screened for antibacterial activity at two antibiotic concentrations of 500 and 1000 g/ml. All the compounds were of high purity and ascertained by melting point determinations as well as by silica gel TLC. The probable structural of the compounds (I-V) was assigned by FTIR spectroscopical method. The I.R frequencies of the presents functional groups are shown in Table 1 The name of the compound, practical yield, melting point range and color are shown in Table 2 .The photographical view of antibacterial action (zone of inhibition shown in Table 3) of azo compounds on two strains Staphylococcus aureus and Bacillus subtilis has been figured as a, b, c, d and e. The antibacterial activity test shown that the synthesised compounds I and II is active against both of the bacterial strains , and compound IV is active towards only S. aureus at both of the concentrations while the azo compounds III and V has no action towards any of the tested strains. Results outcomes has also confirms that the compound II is most active against both of the bacterial strains forming growth inhibition zone upto 18mm at 1000 g/ml while at lower concentration of 500 g/ml there is occurrence of smaller inhibition zone of 11mm and 15mm in S.aureus and B.subtilis respectively. This concludes that azo colorant II can exhibit better antibiotic properties against S.aureus as well as B. subtilis with increasing concentration.


Table 1: FTIR spectral data for the synthesized azo compounds (I-V)

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Table 2: Analytical and physical data of azo colourants I-V.


Azo compounds

Practical

Yield

Melting point range

Colour

(E)-5-(phenyldiazenyl)quinolin-8-ol (I)

81.73%

222- 226 C

Glittering red

(E)-5-((2-nitrophenyl)diazenyl)quinolin-8-ol (II)

87.16%

259- 261 C

Reddish brown

(E)-5-((3-nitrophenyl)diazenyl)quinolin-8-ol (III)

79.49%

243 -246 C

Yellowish

(E)-2-((2,4-dioxopentan-3-yl)diazenyl) benzoic acid (IV)

68.84%

259-260 C

Bright yellow

(E)-4-((2,4-dioxopentan-3-yl)diazenyl) benzoic acid (V)

81.52%

240-242 C

Greenish yellow

Table 3: Showing antibacterial actions of the compounds in mm of zone of inhibition.

Compound


Bacterial

strain

I

II

III

IV

V

Concentration

Concentration

Concentration

Concentration

concentration

500

g/ml

1000

g/ml

500

g/ml

1000

g/ml

500

g/ml

1000

g/ml

500

g/ml

1000

g/ml

500

g/ml

1000

g/ml

zone diameter (mm)

zone diameter (mm)

zone diameter (mm)

zone diameter (mm)

zone diameter (mm)

B. subtilis

06

08

11

18

-

-

-

-

-

-

S. aureus.

06

08

15

18

-

-

08

08

-

-

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Figure 1: FTIR spectrum for azo dyes I-V shown in a-e.

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Figure 2: Photographical view for antibacterial action of synthesised dyes I-V on bacterial strain S. aureus (a-e) and B. subtilis (a-e).


CONCLUSION: No report has been found for the synthesis of Dye II-V whereas the Dye I has been reported by Ezema, 2014. In present work we reported biological potency of all dyes I-V. The results outcome has shown that Dye II is most active toward inhibition of S.aureus as well as B.subtilis bacterial strain with increasing antibiotic concentration from 500 g/ml to 1000 g/ml, this concludes dye-II produces growth inhibiting activity with increasing concentrations while both of the bacterial strains shown resistance against dye-III &V, this might be due to permeability problem of the compounds to reach up to the cell organelles or degradation potency of the bacterial cells.

ACKNOWLEDGEMENT: Authors are thankful to the Department of Chemistry, Arts, Commerce and Science College Bodwad of The Bodwad Sarv. Co. Op Education Society Ltd. Bodwad for providing us lab facility and motivations.

REFERENCES:

1. Sisley P., Poscher C. (1911) Du sort des matieres colorant dans iorganisme animal. C.r. hebd.seanc.acad.sci .Paris. 152. 1062

2. Trefouel J., Trefouel J., Nitti F. Bovet D. (1935) Activity of p-minosulfamide sur les infections stretococciques Esperimental de la souris et du lapin. C.r.Seanc. Sco.Biol.120.756

3. Fuller A. T. (1937) Is p-amino benzene-sulphanoamide the active agent in prontosil therapy? Lancet, 194.

4. Rinde E. and Troll W. (1975) Metabolic reduction of benzidine azo dyes to benzidine in the Rhesus monkey, J. Natl. Cancer Inst., 1975, 55, 181.

5. Robens J. F., Dill G. S., Ward J. M., Joiner J. R., Griesemer R. A., Douglas J. F (1980) Thirteen-week subchronic toxicity studies of Direct Blue 6, Direct Black 38 and Direct Brown 95 dyes, Toxicol. Appl. Pharmacol, 54, 431.

6. Platzek T., Lang C., Grohmann G., Gi U. S., Baltes W. (1999) Formation of a carcinogenic aromatic amine from an azo dye by human skin bacteria in vitro, Hum. Exp. Toxicol., 18, 552

7. Martin C. N., Kennelly J. C. (1985) Metabolsism, mutagenicity and DNA binding of biphenyl-based azo dyes, Drug. Metab. Rev., 16, 89.

8. Collier S. W., Storm J. E., Bronaugh R. L (1993) Reduction of azo dyes during in vitro percutaneous absorption, Toxicol. Appl. Pharmacol, 118, 73.

9. Levine W. G. (1991) Metabolism of azo dyes: Implications for detoxification and activation, Drug Metab, 23, 253.

10. Parasuraman P., Shanmugarajan R. S., Aravazhi T., Nehru K., Mathiazhagan T. and Rajakumari R. (2012) Int. J. Pharm. Life Sci., 3(2), 1430.

11. Goksu S., Uguz M. T.,Ozdemir H. and Secen H. A. (2005) Truk J Chem ., 29, 199-205.

12. Tuazon C. U., Perez A., Kishaba T., Sheagren J. N. (1975) Staphylococcus aureus among insulin-injecting diabetic patients: an increased carrier rate, JAMA, 231, 1272.

13. Tuazon C.U., Sheagren J.N. (1974) Increased rate of carriage of Staphylococcus aureus among narcotic addicts, J Infect Dis, 129, 725-7.

14. Yu V. L., Goetz A., Wagener M. (1986) Staphylococcus aureus nasal carriage and infection in patients on hemodialysis: efficacy of antibiotic prophylaxis, N Engl J Med, 315, 91-6.

15. Weinstein H. J. (1959) The relation between the nasal-staphylococcal-carrier state and the incidence of postoperative complications, N Engl J Med, 260, 1303-8.

16. Kluytmans J. A. J. W., Mouton J. W, Ijzerman E. P. F. (1995) Nasal carriage of Staphylococcus aureus as a major risk factor for wound infections after cardiac surgery, J Infect Dis, 171, 216-9.

17. Weinke T., Schiller R., Fehrenbach F. J., Pohle H. D. (1992) Association between Staphylococcus aureus nasopharyngeal colonization and septicemia in patients infected with the human immunodeficiency virus, Eur J Clin Microbiol Infect Dis, 11, 985-9.

18. Waldvogel F. A. (1995) Staphylococcus aureus (including toxic shock syndrome). In: Mandell GL, Bennett JE, Dolin R, eds. Mandell, Douglas and Bennetts principles and practice of infectious diseases. 4th ed. Vol. 2. New York: Churchill Livingstone, 1754-77.

19. Ezema B. E., Ezema, David C. G., Ugwu Jude. Ayogu (2014) Synthesis of Heterocyclic Azo Dyes from Quinolin-8-ol, Chemistry and Materials Research, 6(9), 1-6.

20. Patil C. J., Patil M. C.,Rane V., Mahajan K. Nehete C. A. (2015) Coupling Reactions Involving Reactions of Aryldiazonium Salt: Part-III. Chemoselective Condensation with b-Naphthol to Synthesize Sudan-I, its Nitro Derivatives and Antibacterial Potential, Journal of Chemical, Biological and Physical Sciences, 5(4), 3860-3867.

21. Patil C. J., Talele D. S., Talele S. P., Pohekar P. R, Patil A. S. (2017) Coupling Reactions Involving Reactions of Aryldiazonium Salt: Part-IV. Chemoselective Synthesis and Antibacterial Activity of 3-(Substituted-phenylazo)-pentane-2,4-diones, Int. J. Pharm. Sci. Rev. Res., 45(1), 64-73.