Synthesis and antibacterial activity of some new fused chromenes

A new class of pyrano[3,4-c]chromene, benzo[c]chromene, chromeno[3,4-c]pyridine and chromeno[4,3-c]pyrazole has been prepared from 3-benzoyl-2H-chromen-2-one 1. Most of the chromene derivatives showed moderate to high antibacterial activity as compared to the starting material 1.


Introduction
During the last twenty years, the study of the biological activities of chromene derivatives has been the aim of many scientists -10 Recently, the anticoagulant, antibacterial, anti-helminthic, hypothermal and vasodilatory properties of chromene has been reviewed 1 .. Fused chromenes are interesting due to their significant antibacterial [11][12][13][14][15] and novobiocin 16,17 activities.Recently, Selectfluor 18 was used as an alternative to conventional catalysts for the synthesis of substituted chromenes via Pechmann condensation of phenols with β-ketoesters under solvent-free conditions.As part of our studies aimed to develop simple and efficient syntheses of polyfunction heterocyclics from readily obtained starting materials 19,20 , we have reported the synthesis of chromenopyridine and thiopyranochromene derivatives by cycloaddition of active methylene compounds with chromene-3-(4-aminosulfonyl) carbanilide 19 or coumarin-3thiocarboxamide 20 .[23][24]

Scheme 1 Scheme 1
Treatment of compound 1 with ethyl cyanoacetate under basic conditions, unexpectedly afforded ethyl 10-cyano-9-hydroxy-6-oxo-7-phenyl-6H-benzo[c] chromene-8-carboxylate 3. The reaction pathway is assumed to proceed by nucleophilic addition of the carbanion to the ethylenic bond of compound 1 affording the expected pyranochromene.Which in turn reacts with a second equivalent of ethyl cyanoacetate ion.Subsequent ring opening followed by recyclization and elimination of HCN affords the isolated product 3.
ARKAT USA, Inc.The structure of product 3 was supported by IR and 1 H-NMR data.(See Experimental Section).
Similar treatment of compound 1 with a series of compounds possessing active methylene groups in refluxing ethanol under basic conditions afforded the pyranochromene and chromenopyridine heterocycles 4-6.

Scheme 3
When compound 1 was reacted with hydrazine hydrate in refluxing ethanol in the presence of triethylamine 3-phenylchromeno [4,3-c] Attempts to get a 3-cyanocoumarin by treatment of salicyaldehyde with ethyl cyanoacetate 17 in refluxing ethanol containing a catalytic amount of piperidine, failed.In stead ethyl 2-amino-5oxo-5H-chromeno [3,4-c]pyridine-1-carboxylate 8 was formed.Presumably The reaction proceeds by cyclo condensation of ethyl cyanoactate with salicyaldehyde to give 3cyanocoumarin 18 which then reacts with a second molecule of ethyl cyanoacetate followed by rearrangement and cyclization.
ARKAT USA, Inc.The structure of compound 3 was supported by its IR and 1H-NMR spectra.Similar treatment of salicylaldehyde with cyanoacetamide in refluxing ethanol containing a catalytic amount of piperidine afforded 2,5-dioxo-3,5-dihydro-2H-chromeno [3,4-c]pyridine-1carbonitrile 9, most likely 3-cyanocoumarin is formed as an intermediate which reacts with a second equivalent of cyanoacetamide followed by cyclization and eliminating of ammonia and HCN .
The structure of compound 9 was deduced from its IR and 1H-NMR spectra.

Biological evaluation
The chromenes (2-9) obtained were preliminarly evaluated for their in vitro antibacterial activity against a narrow spectrum of bacterial species procured from the Laboratory of Microbial Biochemistry (Chem.Dept., Faculty of Science, Al-Fatah Univ.).The paper disc assay described by Cooper 26 using nutrient agar medium was applied.Suspensions of each microorganism were prepared from their 24h-cultures to obtain approximately 10 6 colony forming units (cfu) per ml for plating.Paper discs (Whatman No.1) of 8 mm diameter were loaded individually with a constant amount (100 µg/disc) of the compounds to be tested.Discs were aseptically transferred and applied onto the dry surface of the inoculated plates and then incubated at 37 0 C for overnight (~18-20h) .This assay was performed in duplicates and the mean diameters of the clear inhibition zones (mm) were recorded disregard a single colony or a faint haze caused by the inoculums.Results of the in vitro assay of the synthesized compounds are shown in Table 1.
Compounds 2 and 3 showed mild to moderate activity against the tested Gram-positive and Gram-negative organisms, meanwhile compounds 4, 7, 8, and 9 exhibited comparative and distinctly high activity.Highest antimicrobial activity was observed when the starting compound 1 was annulated with pyrazole or pyridine, except for the N-cyclohexyl derivative 6.Although both compound 4 and 5 are relatively bulky pyrano derivatives, only the former is active.This might be due to the hydrophobic nature of both the methyl and benzoyl groups adjacent to each other in the pyran ring which may cause retardation of the uptake of this compound by microbial cells.

Experimental Section
General Procedures.All melting points were determined on a Koffler melting points apparatus and are uncorrected.H 1 -NMR spectra were recorded on a Bruker Avance 300 MHz spectrometer using TMS as internal reference (chemical shift in δ ppm), and IR spectra in KBr pellets were obtained on a Bruker FT-IR ISS25 spectrophotometer (λmax in cm -1 ).
Method II.Alternatively a solution of compound 1 (0.01mol) and equimolar amount of cyanoacetamide in ethanol (10 ml) containing 3 drops of piperidine was heated to reflux for 4 hours.The solid product was filtered off and recrystallized from dioxane as yellow crystals, yield: 55%; m. p. > 340 o C; Selected IR frequencies (ν cm