Multicomponent Reactions : Microwave-assisted Efficient Synthesis of Dihydropyrimidinones ( thiones ) and Quinazolinones under Green Chemistry Protocol as Probes for Antimicrobial Activities

Microwave promoted diammonium hydrogen phosphate, (NH4)2HPO4, catalyzed threecomponent Biginelli reaction between an aldehyde, a 1,3-dicarbonyl compound and urea or thiourea under solvent-free conditions afforded the corresponding dihydropyrimidinones and quinazolinones in high yields. The synthesized compounds have been screened for their antimicrobial activity against Pseudomonas aeruginosa, Staphylococcus aureus, Vibrio choloriae, Shigella dysenteriae, Salmonella typhi bacteria and Aspargilllus flavus, Saccharomyces cerevisiae and Candida albicans fungi respectively. Some of the synthesized compounds exhibited pronounced antimicrobial activities.

Recently, Microwave heating has emerged as a powerful technique to promote a variety of chemical reactions [52].Microwave reactions under solvent-free conditions are attractive in offering reduced pollution, low cost and offer high yields together with simplicity in processing and handling [53].The Biginelli reaction is important for the preparation of dihydropyrimidine derivatives and excellent results are found for reactions carried out with microwave enhancement [54].
Therefore, a great need still exists for versatile, simple and environmentally friendly processes whereby DHPMs may be formed under milder, more practical and microwave conditions.Diammonium hydrogen phosphate, (NH 4 ) 2 HPO 4 , is a very inexpensive, nontoxic and commercially available compound that can be used in the laboratory without special precautions.To the best of our knowledge, there is only one report regarding the application of diammonium hydrogen phosphate in the preparation of organic compounds under thermal condition [55].Here we report, for the first time, the use of diammonium hydrogen phosphate as catalyst in the synthesis of dihydropyrimidinones and quinazolinones under microwave condition.

Physical measurements
Melting points were recorded with electro thermal melting point apparatus and are uncorrected.Thin layer chromatography was performed on Kieselgel GF 254 and visualization was accomplished by iodine vapour or UV Flame.The infrared (IR) spectra were recorded by FTIR spectrophotometer (Model-8900, Shimadzu, Japan) using KBr matrix in the range 4000-200 cm -1 . 1 H-NMR (400 MHz and 500 MHz) and 13 C-NMR (100 MHz and 125 MHz) spectra were recorded on JEOL GS×400, GEOL JNM-AL 400 (400 MHz) and JEOL GS×400, GEOL JNM-AL 400 (100 MHz) spectrometer (internal standard tetramethyl silane) in CDCl 3 CD 3 OD and DMSO-d 6 as solvent.Chemical shifts were reported in δ unit (ppm) with reference to TMS as an internal standard and J values are given in Hz.The carbon, hydrogen and nitrogen percentages in synthesized products were analyzed according to the approved method ASTM D-5291 by employing Leco-CHNS-932 analyzer.All reactions were carried out in a commercially available LG microwave oven (MB -3947C) having a maximum power output of 800 W operating at 2450 MHz.
The tested compounds were dissolved in N,N-dimethylformamide (DMF) to get a solution of 1 mg mL -1 .The inhibition zones were measured in millimeters at the end of an incubation period of 48 hrs at (35±2)°C.DMF alone showed no inhibition.Nutrient agar (NA) and potato dextrose agar (PDA) were used as basal media to test the bacteria and fungi, respectively.Commercial antibacterial Ampicillin and antifungal Nystatin were also tested under similar conditions for comparison.

Results and Discussions
Salehi and co-workers [55] have reported the synthesis of DHPMs under solvent-free conventional heating conditions at 80 o C. Herein we have successfully carried out the same transformation under microwave irradiation in comparatively shortest duration.A variety of aldehydes were condensed with ethyl acetoacetate/acetylacetone/dimedone and urea (or thiourea) and the products were obtained in good to excellent yields (Scheme 1 and Table 3).
Scheme 1. Synthesis of DHPMs.An important feature of this procedure is the survival of variety of functional groups such as nitro and ether under the reaction conditions.Thiourea also reacts under similar reaction conditions to form the corresponding 3,4-dihydropyrimido-2(1H)-thiones in good to excellent yields.The structures of the products were deduced from their IR, 1 H NMR, 13 C NMR and elemental analyses.For example the 1 H-NMR spectrum of the compound 7 displayed a deshielded doublet signal resonated at δ 8.15 (J= 6.40 Hz) corresponding to two aromatic protons, H-3' and H-5' ortho to the nitro group.Another doublet resonated at δ 7.25 (J= 7.80 Hz) corresponding to two aromatic protons were assigned to H-2' and H-6'.One CH (H-4) proton of the quinazolinone ring resonated at δ 5.55 ppm was observed as singlet.A multiplet appeared at δ 2.48-2.36integrated for six protons corresponding to 2×CH 2 and 2×NH.The spectrum showed two three-proton singlets at δ 1.24 and δ 1.12 due to two CH 3 protons located at C-7 position.The 13   The microanalytical data of the compound 7 is also in good agreement with the assigned structure.Similarly the peaks in 1 H-NMR and 13 C-NMR spectra of the rest compounds were accordance with assigned structures.
Amongst the synthesized compounds screened for the antibacterial activity, compound 2 showed highest activity against S. aureus.Some of the compounds showed low antimicrobial activities and some were unable to show inhibition.For the antifungal activity all compounds, except 7, showed excellent results against all fungi.Compound 2, 3 and 5 revealed highest activity against S. cerevisiae, which was also approximately similar to that of the standard antibiotic, Nystatin.The other tested compounds also exhibited good to excellent results against all the fungi.

Conclusion
In this work, we have developed a new solvent-free strategy for the preparation of 3,4dihydropyrimidin-2-(1H)-ones (and -thiones) as biologically interesting compounds via the condensation of aldehyde with 1,3-dicarbonyl compounds and urea or thiourea.The advantages of this method are high yields, relatively short reaction times, low cost, simple experimental and isolation procedures, and finally, it is in agreement with the green chemistry protocols.The activity data obtained during the study will be certainly useful to go for further research for drug designing and synthesizing new dihydropyrimidinone derivatives.

Table 1 .
Antibacterial activity of the synthesized compounds.

Table 2 .
Antifungal activity of the synthesized compounds.
C NMR spectrum of compound 7 showed the presence of thirteen signals attributed to sixteen carbons of corresponding molecular formula C 16 H 17 N 3 O 3 S.The 13 C NMR spectrum showed the existence of a carbonyl group (C=O) at δ 190.89, a thioxo group (C=S) at δ 189.51, besides five quaternary carbons resonated at δ 146.48 (C-8a), 145.97 (C-1', C-4'), 114.79 (C-4a) and 31.37 (C-7).Two CH 3 carbons observed at δ 29.45 and 27.35.At the same time two CH 2 groups and C-4 carbons appeared at δ 46.86, 46.28 and 33.15.On the other hand, signals resonated at δ 127.57and 123.43 were ascribed for the four aromatic carbons.The DEPT-90 spectrum of the compound 7 showed the presence of three signals at δ 127.57, 123.43, 33.15 attributed to five CH groups in the molecule.The DEPT-135 spectrum of the compound 7 showed the presence of four signals at δ 127.57, 123.43 and 31.15attributed to five CH groups and two signals resonated at δ 29.45 and 27.35 attributed to two CH 3 groups located at C-7 in the molecule.Two carbons of two CH 2 groups appeared at δ 46.86 and 46.28 as negative values.