Synthesis and Spectroscopic Characterization of Novel Hybrid Antibacterial Molecules of Fluorene and Triazole

INTRODUCTION

Heterocyclic compounds are important in the recent years for development in science and technology taking on much more advances of both theoretical and practical relevance which shows many pharmaceutical, medicinal and biological activities. Heterocyclic compounds offer many opportunities for synthetic organic chemist. From wide variety of heterocycles that have been explored developing pharmaceutically important molecule such as fluorene and triazoles have played important role in medicinal chemistry. They are reported to possess broad spectrum of biological activities as antiulcer, anticonvulsant, antihistaminic, antiviral, anti-parasitic, cardiovascular agent as well as antihelmintic.^1-15

Experimental

Material and Methods

Melting points were determined by melting point apparatus and are uncorrected. These all compounds were routinely checked by for their homogeneity by Thin layer chromatography on silica gel G plates. ¹H NMR spectra were recorded on BRUKER spectrometer on a 400 MHz using TMS as internal standard, IR spectra were recorded by Perkin Elmer FT IR spectrophotometer and Mass spectra recorded on (FAB mass).The bacterial strains studied for activities are identified strains.

General Procedure

Preparation of 9 H-fluorene-4-carbonyl chlorides

A mixture of 9H-fluorene-4-carboxylic acid (20g, 0.12mole) in dichloromethane (100ml) and thionyl chloride (14.65g, 0.12mole) was fitted with a condenser and nitrogen purging, The reaction mixture was refluxed with slow nitrogen flush and allowed to stir for 12 h at 35°C. After completion of reaction solvent an excess thionyl chloride distilled out at 50°C under vacuum to obtained solid recrystallize from isopropanol to get pure compound with 90% yield. Analysis observed: C-73.48%, H-4.05%.Calculated for C₁₄H₉ClO is C-73.53.

1-(9H-fluorene-4-carbonyl)-3-phenyl-thiourea

To a suspension of NH₄SCN (4.05g, 0.11 mol) with acetone (100ml) added slowly 9H-fluorene-4-carbonyl chloride (17g, 0.074 mol) under dry condition in 20 min. After addition over reaction mass was stirred at refluxed for 30min. A solution of aniline (6.9g, 0.1 mol) in acetone (50ml) was added to above stirred reaction mass at reflux. After completion of addition reaction mixture was refluxed for further for 30 min. Reaction mixture cooled to RT and poured in the water and resulting solid separated by filtration. Solid was recrystallized in isopropyl alcohol giving pure compound with (20g) yield 78%. Analysis observed C73.20%, H-4.63%, N-8.10% and calculated for C₂₁H₁₆N₂OS is   C73.23%, H-64%, N 8.13%, S-7.41%.

Preparation of 1-(9H-Fluorene-4-carbonyl)-2-methyl-3-phenyl-isothiourea

A mixture of 1-(9H-fluorene-4-carbonyl)-3-phenyl-thiourea (18g, 0.052 mol), methyl iodide (8.1g, 0.057 mol) and anhydrous potassium carbonate (10.8g, 0.078 mol) in DMF (150 ml) with stirring. The resulting off white solid was filtered, washed with water and dried to give compound 1-(9H-Fluorene-4-carbonyl)-2-methyl-3-phenyl-isothiourea Further crystallised in ethanol to give off white solid. Yield 14.3g (81%). Analysis Observation: C-73.68%, H-5.01%, N-7.78%; Calc for C₂₂H₁₈N₂OS: C-73.71,H-5.06%, N-7.82%.

Preparation of [5-1-(9H-Fluoren-4-yl)-2H-[1,2,4] triazol-3-yl]-phenyl -amine

To a suspension of 1-(9H-Fluorene-4-carbonyl)-2-methyl-3-phenyl-isothiourea (12g, 0.033mol)hydrazine hydrate (2.51g, 0.050 mol) in ethanol (80ml) was refluxed for 5hr. Reaction mixture was cool to RT and dumped in water, further obtained light yellow solid separated by filtration. Solid was recrystallized in ethanol to give compound [5-1-(9H-Fluoren-4-yl)-2H-[1,2,4] triazol-3-yl]-phenyl-amine in yield 8.8g, (80%). Analysis observation: C-77.70%, H-4.95%, N-17.22%; Calc for C₂₁H₁₆N₄:C77.76%, H-4.97%, N-17.27%.

Preparation of 3-(9-(4-methlbenylidine)-9H-Fluoren-4-yl]-N-phenyl-1H-1,2,4 triazol-5-amine

To a suspension of [5-1-(9H-Fluoren-4-yl)-2H-[1,2,4] triazol-3-yl]-phenyl-amine (8g, 0.024 mol), potassium hydroxide (0.7g, 0.012 mol) and paratolualdehyde (4.32g, 0.036 mol) were dissolved in N,N-Dimethylformamide (75 ml) and heated to 60-70°C for 24 hr. Water (800ml) to added and reaction mass extracted with ethylacetate (3 X 200ml) and the combine extracted organic layers treated with sodium sulphate. Distilled out organic layer under vacuum to obtained residue further crystallize from ethanol to obtained pure off white compound 3-(9-(4-methlbenylidine)-9H-Fluoren-4-yl]-N-phenyl-1H-1,2,4 triazol-5-amine in 76% yield. Analysis Observed is C-81.48%, H-4.85%, N-13.53%; Calculated for C₂₈H₂₀N₄ is C-81.53%, H-4.89%, N-13.58%.

The progress of reaction and purity of compound checked by TLC aluminum sheet silica gel 60 F254 (E.Merck) using Hexane:acetate (60:40 V/V) and visualized in U. V (254nm).

Similarly other substituted hybrid molecules of fluorene and triazole have been prepared. The physical data are recorded in Table-1.

Biological Evaluation

Antibacterial activity of 3-(9-(4-methylbenzylidene)-9H-fluoren-yl)-N-Aryl-1H-1,2,4-triazole-5-amine. Novel synthesized hybrid compounds of fluorene and triazole have been tested for their antibacterial activity against gram positive and gram negative bacteria B.coccus, S aureus, E.aerogenes, P.aeruginosa by using borer medium and filled with 0.04 ml (40µg) solution of sample in DMF and Amoxicillin, Benzoylpenicillin, Ciprofloxacin and erythromycin were used as a reference compound A1, A2, A3, A4, A5, A6, A7, A8, A9 and A10 have shown moderate activity. The Zone of inhibition of the bacterial growth were measured in millimeter and are recorded in table no 2.

Table 1: Physical constant of 3-(9-(4-methylbenzylidene)-9H-fluoren-yl)-N-Aryl-1H-1,2,4-triazole-5-amine

Sr. No.	Ar	Molecular formula	M P	Yield	C, H, N Elemental analysis
					% C		% H		% N
			°C	%	Cal.	Obs.	Cal.	Obs.	Cal.	Obs.
A-01	-C6H5	C28H20N4	113-117	66	81.53	81.50	4.89	4.88	13.58	13.57
A-02	-C6H4-CH3	C29H22N4	106-111	69	81.66	81.67	5.20	5.22	13.14	13.10
A-03	-C6H4-Br	C28H19BrN4	96-101	59	68.44	68.45	3.90	3.89	11.40	11.39
A-04	-C6H4CN	C29H19N5	106-112	63	79.61	79.59	4.38	4.39	16.01	17.99
A-05	-C6H4OH	C28H20N4O	95-100	59	78.49	78.48	4.70	4.69	13.08	13.06
A-06	-CH2-C6H5	C29H22N4	88-95	62	81.66	81.69	5.20	5.24	13.14	13.15
A-07	-C6H4-C4H9	C31H26N4	113-118	57	81.91	81.88	5.77	5.78	12.33	12.29
A-08	-C6H4-C3H7	C30H24N4	89-93	65	81.79	81.80	5.49	5.50	12.72	12.69
A-09	-C5H4N	C27H19N5	98-103	55	78.43	78.45	4.63	4.59	16.94	16.95
A-10	-C6H3-F2	C28H18F2N4	103-108	69	74.99	75.01	4.05	4.03	12.49	12.45

Reaction Scheme

 

Table 2: Antibacterial activity of 3-(9-(4-methylbenzylidene)-9H-fluoren-yl)-N-Aryl-1H-1,2,4-triazole-5-amine

Compounds	Antibacterial and antimicrobial activity MIC in µg/ml
	B. coccus	P. aeruginosa	S. aureus	E,aerogenes	A.niger	C.albicans
A-01	13	15	17	16	16	13
A-02	21	17	18	16	14	12
A-03	19	19	16	17	19	18
A-04	16	20	17	19	18	17
A-05	15	18	15	15	19	18
A-06	14	13	12	14	16	11
A-07	19	17	16	19	18	17
A-08	14	16	17	20	16	10
A-09	20	15	19	17	18	18
A-10	14	14	14	20	17	09
Amoxicillin	25	22	25	20	00	00
Benzoyl penicillin	19	21	19	21	00	00
Erythromycin	22	23	21	19	00	00
Ciprofloxacin	20	16	15	22	00	00

Antibacterial Activity Chart

RESULTS AND DISCUSSION

Novel synthesized hybrid molecules of fluorene and triazole with various aryl groups having yield between 57 to 69%. The structure of novel compounds are characterized and confirmed by 1H NMR, IR and Mass spectral data which are further supported by elemental analysis data. Novel compounds A1 to A10 were shown significant activities.

ACKNOWLEDGEMENT

The authors are thankful to Biotechnology department of North Gujrat University, Patan for biological activity

REFERENCES

1. M. Burns, john Iball (1954), “Molecular structure of Fluorene Nature.” volume 173, p.635.
2. Gerkin, R. E., Lundstedt, A. P., & Reppart, W. J. (1984). Structure of fluorene, C13H10, at 159 K. Acta Crystallographica Section C: Crystal Structure Communications, 40(11), 1892-1894.
3. Kurdyukova, I. V., & Ishchenko, A. A. (2012). Organic dyes based on fluorene and its derivatives. Russian Chemical Reviews, 81(3), 258.
4. Belfield, K. D., Hagan, D. J., Van Stryland, E. W., Schafer, K. J., & Negres, R. A. (1999). New two-photon absorbing fluorene derivatives: Synthesis and nonlinear optical characterization. Organic Letters, 1(10), 1575-1578.
5. Meng, S., Sun, N., Su, K., Zhao, X., Wang, D., Zhou, H., & Chen, C. (2018). Novel organosoluble polyarylates based on diphenylamine-fluorene units: Synthesis, electrochromic, and electrofluorescent properties. High Performance Polymers, 30(7), 864-871.
6. Yadavi, M., Badiei, A., Ziarani, G., & Abbasi, A. (2013). Synthesis of novel fluorene-functionalised nanoporous silica and its luminescence behaviour in acidic media. Chemical papers, 67(7), 751-758.
7. Jonas Alves Fernandes, Yasuhiro Morisaki & Yoshiki Chujo. (2011). Aromatic-ring-layered polymers composed of fluorene and xanthene. Polymer Journal, 43, 733-737.
8. Al-Soud, Y. A., Al-Dweri, M. N., & Al-Masoudi, N. A. (2004). Synthesis, antitumor and antiviral properties of some 1, 2, 4-triazole derivatives. Il Farmaco, 59(10), 775-783.
9. Seelam, N., Shrivastava, S. P., Prasanthi, S., & Gupta, S. (2016). Synthesis and in vitro study of some fused 1, 2, 4-triazole derivatives as antimycobacterial agents. Journal of Saudi Chemical Society, 20(4), 411-418.
10. Mishra, R., Kumar, R., Kumar, S., Majeed, J., Rashid, M., & Sharma, S. (2010). Synthesis and in vitro antimicrobial activity of some triazole derivatives. Journal of the Chilean Chemical Society, 55(3), 359-362.
11. Karaali, N., Mentese, E., Imaz, F. Y., Usta, A., & Kahveci, B. (2013). Microwave-assisted synthesis of some 1H-1, 2, 4-triazol-3-one derivatives. South African Journal of Chemistry, 66(1), 72-76.
12. Karaali, N., Mentese, E., Imaz, F. Y., Usta, A., & Kahveci, B. (2013). Microwave-assisted synthesis of some 1H-1, 2, 4-triazol-3-one derivatives. South African Journal of Chemistry, 66(1), 72-76.
13. Wu, J., Ni, T., Chai, X., Wang, T., Wang, H., Chen, J., & Jiang, Y. (2018). Molecular docking, design, synthesis and antifungal activity study of novel triazole derivatives. European journal of medicinal chemistry, 143, 1840-1846.
14. Wu, J., Ni, T., Chai, X., Wang, T., Wang, H., Chen, J., & Jiang, Y. (2018). Molecular docking, design, synthesis and antifungal activity study of novel triazole derivatives. European journal of medicinal chemistry, 143, 1840-1846.
15. New class of triazole Derivatives and their antimicrobial activity, Letters in drug Design & Discovery, 9(7), 2012 PP. 687-693.