Aryl Chalcones as Efficient Precursors for Deriving Oxazine: Solvent-Free Synthesis and Antimicrobial Activities of some Oxazine-2-amines

A series of some oxazine derivatives has been synthesised by fly-ash:H2SO4 catalyzed solventfree cyclization of aryl chalcones and urea under microwave irradiation. The yields of the oxazines were more than 85 %. The synthesised oxazines were characterized by their physical constants, analytical and spectroscopic data. The antimicrobial activities of these oxazines have been studied using Bauer-Kirby method.


INTRODUCTION
Oxazines are a six membered heterocyclic compounds possess one oxygen and one nitrogen atom. Depend upon the relative position of these tow atoms and the double bond, these molecule exists many isomeric structures such as 1, 2 or 1, 3 or 1, 4 oxazines [1].
Therefore the authors have taken effort to synthesize some 9H-fluorene-2-yl based oxazine 2-amine derivatives and study the antimicrobial activities using Bauer-Kirby method.

1. General
All chemicals were used in this study were purchased from Sigma-Aldrich and Merck Chemical companies. Mettler FP51 melting point apparatus was used for determining the melting point of all synthesized oxazines in open glass capillaries and are uncorrected. The AVATAR-300 Fourier transform spectrophotometer was used for recording infrared spectra (KBr, 4000-400 cm -1 ) of all oxazines in KBr disc.
The Bruker AV400 series type NMR spectrometer was utilized for recording NMR spectra of all oxazines, operating at 400 MHz for 1 H and100 MHz for 13 C spectra in CDCl 3 solvent using TMS as internal standard. Mass spectra of all synthesised oxazines were recorded on SHIMADZU mass spectrometer using chemical ionization technique.

RESULTS AND DISCUSSION
In our organic chemistry research laboratory, the author attempt to synthesize oxazine derivatives by cyclization of chalcones possess electron with-drawing as well as electron donating group as substituents, urea and in the presence of acidic catalyst fly-ash:H 2 SO 4 using microwave irradiation. Hence the authors have synthesized some substituted 1,3oxazine derivatives by the cyclization of 2 mmole of chalcone, 2 mmole of urea under microwave irradiation with 0.4 g of fly-ash:H 2 SO 4 catalyst at 550 W for 4-6 minutes (Samsung Grill, GW73BD Microwave oven, 230 V A/c, 50Hz, 2450 Hz, 100-750 W (IEC-705), (Scheme 1). During the course of this reaction fly-ash:H 2 SO 4 catalyses cyclization between chalcones and urea followed by rearrangement gave the 1, 3-oxazine amines. The yield of the oxazine in this reaction are more than 80 %. The chalcone containing electron donating substituent (OCH 3 ) gave higher yields than electron-withdrawing (halogens, NO 2 )

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Volume 23 substituents. Further we have investigated this cyclization reaction with equimolar quantities of the styryl 9H-fluorennyl ketone (entry 10 ) and urea under the same condition as above. In this reaction the obtained yield was 90 %. The effect of catalyst on this reaction was studied by varying the catalyst quantity from 0.1 g to 1 g. As the catalyst quantity is increased from 0.1 g to 1 g, the percentage of yield of product is increased from 84 to 90 %. Further increase in the catalyst amount beyond 0.4 g, there is no significant increase in the percentage of the product. The effect of catalyst loading is shown in Fig. 1. The optimum quantity of catalyst loading was found to be 0.4 g. The reusability of this catalyst was The reusability of this catalyst was studied for the cyclization of styryl 9H-fluorenyl ketone, and urea (entry 10) is presented in Table 2. From the Table 2, first two runs gave 90 % product. The third, fourth and fifth runs of reactions gave respectively the yields 89.5 %, 89.5 % and 93 % of oxazines.
There was no appreciable loss in its effect of catalytic activity observed up to fifth run. The effect of solvents on the yield was also studied with methanol, ethanol, dichloromethane and tetrahydrofuran from each component of the catalyst (entry 10). Similarly the effect of microwave irradiation was studied on each component of the catalyst. The effect of solvents on the yield of oxazine derivatives was presented in Table 3. From the table highest yield of oxazine obtained from the cyclization of chalcones and urea with the catalyst fly-ash:H 2 SO 4 in microwave irradiation. The infrared and nmr spectroscopic data of these 1-acetyl pyrazolines are summarized in Table 4. Table 3. The effect of solvents in conventional heating and without solvent in microwave irradiation on yield of oxazine amine (entry 11).   . Infrared and NMR spectroscopic data of 4-aryl-5,6-dihydro-6(substituted phenyl)-4H-1,3-oxazine-2-amines.

1. Antimicrobial activities
Antibacterial sensitivity assay of all oxazine amines were performed using Kirby-Bauer [35]  In each Petri plate about 0.5 mL of the test bacterial sample is spread uniformly over the solidified Mueller Hinton agar using sterile glass spreader. Then the discs with 5mm diameter made up of Whatman No.1 filter paper, impregnated with the solution of the compound are placed on the medium using sterile forceps.
The plates are incubated for 24 hours at 37 °C by keeping the plates upside down to prevent the collection of water droplets over the medium. After 24 hours, the plates are visually examined and the diameter values of the zone of inhibition have been measured. Triplicate results are recorded by repeating the same procedure.
The antibacterial screening effect of synthesized oxazine were shown in (Figure 2; Plates 1-12). The zone of inhibition is compared using    The antifungal activity of substituted chalcones synthesized in the present study is shown in Figure 3; Plates 1-6 and the zone of inhibition values of the effect is given in Table  6. From the table the oxazine derivatives 2-4, 6 and 8 shows satisfactory fungal activities against A. niger. Compounds 8, 9, 10 and 5-7 were shows excellent good and satisfactory antifungal activities against E. coli. The oxazine amines 7, 10 and 2, 3, 5, 8 were shows good and satisfactory antifungal activities against T. viride fungal strains.

CONCLUSIONS
Some oxazine amine derivatives including 9H-flurenyl based oxazine amines have been synthesised by solvent free cyclization of aryl chalcones and urea in presence of flyash:H 2 SO 4 catalyst under microwave irradiation. This synthetic methodology offers solventfree cyclization, non-hazardous, shorter reaction time, easy-workup procedure and better yields. The analytical and spectral data were supported for these oxazine derivatives. Most of the oxazine derivatives shows good and moderate antimicrobial activities against the respective bacterial and fungal strains.