Green Synthesis and In vitro Antioxidant Activity of 6-aryl-Quinazolin-4 ( 3 H )-ones via Suzuki Cross Coupling

Suzuki cross coupling reaction based synthetic method for the preparation of 6-arylquinazolin-4(3H)-ones (3a-h) in ethylene glycol dimethyl ether is described by coupling of 6-iodo2-phenyl-3-methyl-quinazolin-4(3H)-one (1) with various 6-aryl boronic acids 3 under microwave irradiation in between These compounds were screened for anti-oxidant activity.


INTRODUCTION
The recent literature is enriched with progressive findings about quinazolin-4(3H)-one and its derivatives which exhibit a broad range of biological properties such as antioxidant 1 , anti-tumour 2 , anti-inflammatory 3 , anti-malarial 4 , anti-bacterial 5 and anticonvulsant activity 6 .Further, quinazolin-4(3H)-one have also been investigated scaffold for the synthesis of various drugs and their intermediates 7 .
Cross coupling reaction is an significant method of generating carbon-carbon bonds in organic compounds, which is catalyzed by various transition metals.In the past three decades, carboncarbon bond construction reaction has allowed chemists to produce complex molecular structures of various interests including total synthesis of natural products 8 , medicinal chemistry and industrial process development 9 .
In modern days, Microwave-Assisted Organic Synthesis (MAOS) has fascinated the interest of synthetic chemists.The rate of a reaction is speed up under microwave irradiation compared to conventional heating.
Insight of quinazolin-4(3H)-one moiety and cross coupling reaction, we have in the previous reported the successful use of Kumada crosscoupling reactions for the synthesis of 6-aryl-quinazolin-4(3H)-ones derivatives 10 .However, this coupling is related with extended reaction times and with low yields.In search of our previous investigations toward developing more proficient protocol for the synthesis of 6-aryl-quinazolin-4(3H)-one derivatives, we optimized the Suzuki reaction conditions under microwave irradiation and conventional heating.It was envisaged that the microwave irradiation would enhance the rate of reaction, thereby reducing time.
Here in, we report the coupling of 6-iodo-3-methyl-2-phenyl-quinazolin-4(3H)-one (1) with aryl boronic acids in presence of Pd(PPh 3 ) 4 as catalyst derivatives under microwave irradiation as well as conventional heating conditions and evaluated their in vitro antioxidant property.

Experimental section
IR spectra for all the compounds were recorded in solid KBr on infra cold model 337 Perkin-Elmer instrument.Melting points were measured in open capillary tubes and are uncorrected.The 1 H NMR (400 MHz) and 13 C NMR (75 MHz) spectra were recorded on Varian Gemini 400 or Bruker 75 in CDCl 3 using tetramethylsilane as internal standard.The purity of all obtained compounds was checked by thin-layer chromatography.High resolution Mass spectra were recorded on Q-TOF mass spectrometer.Multisynth series microwave system (Milestone) were used for Suzuki reaction.

General procedure for the preparation of 3a-h via Suzuki cross coupling reaction Microwave heating
The 6-iodoquinazolinone 1 (0.5 mmol) and boronic acid (0.55 mmol) were dissolved ethylene glycol dimethyl ether in a microwave vial.Pd(PPh 3 ) 4 (0.02 mmol) and Na 2 CO 3 (1.25 mmol) were added, and the reaction mixture was irradiated in a microwave apparatus at 80°C, 250 W for 20 minute.After the reaction mixture was cooled to ambient temperature, the product was filtered, the filtrate was concentrated, and the crude mixture was purified by silica gel column chromatography using hexane/ ethyl acetate (88/12) as eluent.

Conventional heating
The 6-iodoquinazolinone 1 (0.5 mmol), boronic acid 2 (0.55 mmol) and Na 2 CO 3 (1.25 mmol) were dissolved in ethylene glycol dimethyl ether in a seal tube.After degassing reaction mixture, Pd(PPh 3 ) 4 (0.02 mmol) and were added.And the reaction mixture was heated at 80°C, 250 W for overnight.After the reaction mixture was cooled to ambient temperature, the product was filtered, the filtrate was concentrated, and the crude mixture was purified by silica gel column chromatography using hexane/ethyl acetate (88/12) as eluent.

Assay of anti-oxidant property Lipid peroxidation assay
In vitro lipid peroxidation of the compounds was determined by the described method 11,12 .Briefly, 1 mL of rat liver microsomal fraction was added to 1.0 mL of 150 mM Tris-HCl buffer (pH 7.4) containing various concentrations (50, 100 and 150 µg/ mL) of test compounds, 0.2 mL FeCl 3 (1 mM) and 0.2 mL ascorbic acid (0.5 mM) to induce lipid peroxidation.The mixtures were incubated at 37°C for 30 minutes.At the end of the incubation, 0.5 ml of glacial acetic acid and 0.5 ml of 0.33% TBA were added to each mixture.The mixtures were kept in a water bath at 97°C for 45 min.at cooling, the pink chromogen was extracted with 2 mL of butanol.The absorbance of the organic layer was measured at 535 nm, and the thiobarbituric acid reactive substances produced were estimated using the MDA standard curve.BHA was used as a control substance (20 µg/ml).

DPPH (1, 1-Diphenyl-2-picrylhydrazyl) radical Scavenging activity
The free radical scavenging activity of isolated compounds was measured by 1,1-diphenyl-2-picryl-hydrazyl (DPPH) method described by Blois 1958.0.1 mM solution of DPPH in methanol was prepared and 1ml of this solution was added to 3 mL of various concentrations of (50, 100 and 150 µ g/ ml) test compounds and the reference compound (20 µ g/ml).After 30 min., absorbance was measured at 517 nm.BHA was used as the reference material.All the tests were performed in triplicate.The percentage of inhibition was calculated by comparing the absorbance values of the control and test samples.% DPPH radical scavenging = [(Absorbance of control -Absorbance of test sample)/ (Absorbance of control)] ×100

ABTS*+ (2, 2'-azino-bis (3-ethylben -zthiazoline-6-sulphonic acid) radical cation decolourisation assay
This method was carried out by well described method 13 .ABTS*+ (54.8 mg) was dissolved in 50mL of distilled water to 2mM concentration and potassium persulphate (17 mM, 0.3 mL) was added.The reaction mixture was left to stand at room temperature overnight in dark before use.To 0.2 mL of various concentrations (50, 100 and 150 µ g/ mL) of test samples and reference standard BHA (20 µ g/mL) 1.0 mL of distilled DMSO and 0.16 mL of ABTS*+ solution was added to make a final volume of 1.36mL.Absorbance was measured spectro-photometrically, after 20 min.at 734 nm 14 .