Synthesis, Characterization and Antimicrobial Activity of New 2-Phenylquinoline-4(3H)-one Derivatives

Introduction

Quinazolinone is one of the flourishing compounds in pharmaceutical fields. Quinazolines and its derivatives represent one of the most active classes of compounds. Medically it has been exhibited various activites as anti-bacterial ^1,2 ,cytotoxic ³, analgesic , anti-inflammatory ⁴, anticonvusant ⁵,antitubercular ^6,7 , anticancer⁸, and antioxidant⁹ , anti-HIV¹⁰,antitumour¹¹,and anti-microbial ¹². Quinazolinones derivatives were exhibit substantial in treating of leukemia than the common drugs¹³. Recent studies demonstrated the significant effect of quinazolinones derivatives via breast cancer in vitro study^14-16.

Aim of the present study is to synthesis  a new phenylquinazoline derivatives as   antibacterial agents.

Material and Method

Chemistry

Melting points were determined using electrothermal melting point apparatus. A SHMADZU-FT-IR-8400 spectrometer was used to record the Infrared spectra atyrhemiss KBr disc. All ¹H, ¹³C-NMR spectra measurement were made at Al-Albayt University, Jordan and Cairo University, Egypt on a Bruker 300 MHz and Avance III HD spectrometers using DMSO-d₆ a solvent and TMS as internal reference. The progress of the reaction was monitored by TLC using aluminum silica gel plates. Grade solvent and reagents were purchased from commercial suppliers.

2-phenyl-4H-benzo[d][1,3] oxazin-4-one

0.01 mol. of C₇H₅OCl was gradually added to a 30 ml solution of 3.33×10^-4 M of anthranilic acid in pyridine with continuous stirring at about 15˚C for 30 min. The formed product was separated while the reaction mixture stirred for 3 hours room temperature. Finally, a pale-yellow solid was yielded upon the addition of 15 ml. of 10% NaHCO₃ was added to the solution. The formed precipitate was filtered off and washed with distilled water, dried and ethanol was used for recrystallization. Yield (68%), m.p. (118-120)˚C¹⁷.

Synthesis of 3-(1H-indol-3-yl)-2-(4-oxo-2-phenylquinazolin-3(4H)-yl) propanoic acid

(0.001mol.) of 2-phenyl-4H-benzo[d][1,3] oxazin-4-one was mixed with tryptophan in 15 ml  of glacial acetic acid. The mixture was refluxed for (3-4) hr., followed by the addition of (25 ml) of Icy distilled water. The product was separated off by filtration and washed with distilled water, the dried.  Recrystallization of the obtained product was carried out from ethanol. Yield (70%), m.p. (165-167)˚C¹⁸.

(1H-indol-3-yl)-2-(4-oxo-2-phenylquinazolin-3(4H)-yl)propanoyl chloride

A solution of compound 2 (0.01 mol.) and SOCl₂ (2 ml.) in (20 ml.) benzene(dry) was refluxed for 4 hr. Crystals of the product were obtained by removing the excess of thionyl chloride and benzene under vacuum. yield (75%), m.p. (158-160)˚C ¹⁹.

3-(1H-indol-3-yl)-2-(4-oxo-2-phenylquinazolin-3(4H)-yl)propanehydrazide

0.01 mol.  of compound  3 was mixed with 80% N₂H₄.H₂O solution (0.5 ml., 5 mmol.) in dry benzene (10 ml.). After refluxing the mixture for five hours, the mixture was let to cool down to room temperature before removing the excess solvent and hydrazine under reduce pressure. The precipitate that obtained was washed with ether, and purified by recrystallized from ethanol. Yield (85%), m.p. (178-180)˚C ¹⁹.

(3-(1H-indol-3-yl)-1-(3-methyl-5-oxo-4,5-dihydro-1H-pyrazol-1-yl)-1-oxopropan-2-yl)-phenylquinazolin-4(3H)-one

This compound was synthesized by refluxing (0.01 mol.) of compound 4 and ethyl acetoacetate (0.01 mol.) in 20 ml. glacial acetic acid for 6 hrs. After then, the mixture was cooled and 25 ml of icy distilled water was added, the reaction product (compound 5) was filtered out and dried. Yield (80%), m.p. (221-223)˚C ²⁰.

3-(2-(1H-indol-3-yl)-1-(5-mercapto-1,3,4-oxadiazole-2-yl)ethyl)-2- phenylquinazolin-4(3H)-one

Twenty  milliliters of ethanolic solution of (0.015 mol.) NaOH was mixed with (0.01 mol.) of compound 4 with continuous stirring for 15 min. Two milliliters of CS₂ was then added before heating the mixture under reflux for 8 hrs. After concentrating the mixture, the reaction product was collected by acidification with dilute HCl solution. The product was then washed with water and recrystallized from ethanol. Yield (76%), m.p. (229-231)˚C ²¹.

2-hydrazinyl-5- (2-(1H-indol-3-yl) ethyl)-2-phenylquinazolin-4(3H)-one

To 20 ml  of the ethanolic solution of compound 6 (0.01 mol), two milliliters of  N₂H₄.H₂O solution were added. After 7 hrs of reflux, the reaction mixture was left to cool down to room temperature.  The pure yellow crystals of the product were obtained after removing the solvent under vacuum and recrystallization from ethanol. Yield (80%), m.p. (154-156)˚C ²¹.

 2-(3-(1H-indol-3-yl)-2-(4-oxo-2-phenyl quinazolin-3(4H)-yl)propanoyl)-2,3-dihydrophthalazine-1,4-dione

This compound was obtained by refluxing a solution of an equimolar mixture (0.001 mol.) of compound 4 and phthalic anhydride in glacial acetic acid.  After the addition of 20 ml. icy distilled water, the product was filtered and dried. Yield (80%), m.p. (121-123)˚C ²².

General Method for Synthesis of Schiff Bases 9a-d

An equimolar mixture (0.01 mol.) of and compound 4 with an appropriate aldehyde in (25 ml.) of absolute ethanol was refluxed in water bath for 8 hr after the addition of three drops of anhydrous CH₃COOH.  The formed product was collected by filtration and dried, after cooling the reaction mixture to room temperature²³. The molecular formulas, melting points, and the yields of the synthesized compounds are presented in Table (1).

General Method for Synthesis of Oxazepines Compounds 10a,b

An equimolar mixture (0.01 mol.) of compounds  9a,b and phthalic anhydride was dissolved in (10 ml) of dry benzene and refluxed in a water bath for 10 hr. The solution was then cooled and treated with sodium bicarbonate to formed compound 10a,b. The reaction product i.e. 10a,b was separated by filtration, dried and recrystallized from ethanol²⁴. Table (1) shows the data for compound (10a,b).

Biological Activity

The antimicrobial activity of the synthesized phenylquinazoline on two kinds of bacteria namely, staphylococcus aureous and Escherichia coli was investigated. The investigation was carried out via cup method with nutrient agar as a media for bacterial growth using 0.1ml of DMF as a solvent for all samples.The agar was scoop up from the sterilized cork borer cups and transferred into a bacterially inoculated petri dish. After then, a 0.1 ml aliquot of the solution of the investigated compound was introduced to each dish and the dishes were incubated for 24 hrs at 37°C.

Scheme 1 Click here to View scheme

 

Results and Discussion

Chemistry

The starting materials 2-amino-3-(1H-indol-3-yl) propanoic acid 2 , used in this work was prepared by refluxing of tryptophan with 2-phenyl-4H-benzo[d][1,3]oxazin-4-one (1) in glacial acetic acid. Analytical and spectral data were used to elucidate the structure of compound 2. The FTIR absorption spectrum of the compound displayed bands at 1685 cm^-1 for carbonyl of acid, 2500-3200 cm^-1 due to OH group of acid.

Compound 3 was obtained upon treatment of compound 2 with thionyl chloride in dry benzene. The presence of acyl chloride group was confirmed using its FTIR spectrum  which display a strong absorption band at 1762 cm^-1. The reaction of acyl chloride derivative 3 with hydrazide hydrate in refluxing ethanol gave the corresponding hydrazide derivative 4. The compound shows several bands at 1660 cm^-1 which belong to carbonyl of amide, and at 3307, 3215 cm^-1 for NH₂ group. Moreover, its The ¹H-NMR spectrum, fig. (1), shows characteristics chemical shifts, i.e.  singlet signals at 5.67, 10.30, and 12.70 ppm which could be related to the protons of NH₂ group, proton of NH group and the proton of OH group present in two tautomeric forms.

Figure 1  Click here to View figure

 

The triplet signal of proton for CH group  at 4.30 ppm.  Also showed the multiple signal at 6.92-8.68 ppm for aromatic protons of three benzene rings. The two  peaks at at 164 and 167 in the ¹³C NMR spectrum of compound 4. fig. (2), indicate the presence of two resonating carbonyl groups of amide, while peaks at (127.40, 128.90) ppm are due to carbon of C=N group.

Figure 2 Click here to View figure

 

Compound 4 was treated with ethyl acetoacetate in boiling glacial acetic acid to give compound 5. Bands at 1726, 1678 cm-1, 1637 cm-1, and 3309 cm-1 in the FTIR spectrum of compound  5 indicate the presence of CO amide, C=N, NH groups. The treatment of compound  4 with CS₂ in presence of NaOH yields compound 6. The appearance of two peaks at (1616, 1600) cm-1 in the FTIR spectrum of the formed compound could be attributable to the presence of C=N of cyclic imine.   Moreover, the ¹H NMR spectrum of compound 6,  fig. (3), exhibited two singlets at δ 11.69 and 11.71 which are specific for NH and SH protons, a triplet signal at δ 3.45 for proton of CH proton, a multiplet at δ 7.45-8.69 related to the aromatic-protons, a singlet at δ 8.69 for NH of cyclic tryptophan.  Singlet   signal at δ 4.38 for CH of cyclic tryptophan.

Figure 3 Click here to View figure

 

Reaction of compound 6 with hydrazine hydrate in refluxing ethanol gave the corresponding compound 7 The FTIR spectrum of compound 7 confirmed the presence of intense absorption bands at 3329, 3105, 1662, and 1620 cm^-1 due to amino, carbonyl amide and imine groups respectively. The ¹H NMR spectrum of compound 7 ,fig.(4), showed two signals, triplet signal at δ 3.44 for CH proton and doublet signal at δ 2.68 for proton of CH₂ group. A singlet signal at δ 10.45 for proton of NH, a multiple signal at δ 7.18-8.20 region related to aromatic protons, a signal at δ 8.40 for proton of NH of tryptophan.

Figure 4 Click here to View figure

 

Compound 8 was synthesized by the condensation of compound 4 with phthalic anhydride in dry benzene. The chemical structure of this compound was determined by FTIR and ¹H NMR spectroscopic techniques. The recorded infrared spectrum of the compound depicts the disappearance of νs and νas of NH2 group at (3317, 3165) cm-1, the appearance of two bands (1703, 1786) cm-1 for carbonyl group of cyclic amide, and a band rise at (1678) cm-1 due to carbonyl of amide. On the other hand, the proton-NMR spectrum of compound 8, fig. (5), shows many signals, a triplet at δ 3.59 and a doublet at δ 1.19 for CH and CH₂ protons respectively, a singlet at δ 4.39 for the tryptophan five membered ring CH, two singlets at δ 11.76 and δ 12.01 for NH and OH protons in the two tautomeric forms, a multiple at δ 7.26-8.79 for the aromatic protons, and a singlet  at δ 9.65 which corresponds to the tryptophan NH group.

Figure 5 Click here to View figure

 

Compound 4   condensed easily with different aldehydes in ethanol containing piperidine at reflux temperature affording the corresponding Schiffʼs bases compounds 9a-d. The FTIR spectrum of compounds 9a-d   showed disappearance of amino group and exhibited peak at region (1604-1649) cm^-1 belonging to (C=N) group. The ¹H NMR spectrum of compound 9a ,fig.(6),showed singlet signals at δ 10.57, 11.78, 11.95, and 12.13 corresponding to CH=N, NH of tryptophan, NH and OH protons in two tautomeric forms respectively. A multiple signals at δ 7.27-8.60 region owing to aromatic protons.The¹³CNMR spectrum of compound 9a, fig. (7), (DMSO): δ123.10-132.58(benzene rings), (127.24, 128.89) ppm due to two carbon of C=N groups. While (164.49-164.94) ppm due to two carbonyls of amide groups.

Figure 6   Click here to View table

Figure 7 Click here to View figure

Figure 8 Click here to View figure

Figure 9 Click here to View figure

 

Synthesis of compounds 10a,b was carried out via condensation of compounds 9a,b with phthalic anhydride in dry benzene. The purity structural formula of compounds 10a,b were confirmed by FTIR and ¹H, ¹³C NMR spectroscopic techniques. The FTIR of compounds 10a,b showed a peak at (1664-1691) cm^-1 belong to carbonyl of cyclic ester. The carbonyl absorption band of amide was observed at (1656-1691) cm^-1.  The ¹H-NMR spectrum of compound  10 b, fig. (8), display two singles at δ 11.83 and 12.09 for NH and OH protons in the two tautomeric forms, singlet   at δ 8.61 owing to protons of NHof tryptophan group, a multiple signal at δ (6.75-8.58) region attributed to aromatic protons. Two signals, triplet signal at 3.32 δ for CH proton and doublet signal at 2.86 δ for protons of CH₂ group, singlet signals at 3.84 δ for CH of tryptophan. Singlet signal at 8.58 δ for  (-CH-O-N) group. The ¹³C NMR spectrum of compound 10b, fig.(9), (DMSO): δ 28.23 (CH₂), 123.01-132.33 (benzene rings), 164.40 (CO amide), 195.63 (CO of cyclic ester).

Table 1: Physical properties for Schiffʼs bases and 1,3-oxazepine compounds.

Comp. No.	Ar	Arˋ	Molecular formula	M. p. (˚C)	Yield (%)
9a	4-BrC6H4–		C32H25N5O2Br	240-242	70
9b	4,4-N,N(CH3)2C6H4–		C34H31N6O2	220-222	75
9c	4-ClC6H4–		C32H25N5O2Cl	217-219	80
9d	C6H5–		C32H26N2O2	198-200	78
10a		4-BrC6H4–	C40H28N5O5Br	211-213	60
10b		4,4-N,N(CH3)2C6H4–	C42H34N6O5	206-208	70

Table 2: The FTIR absorption bands for Schiffʼs bases and 1,3-oxazepine compounds.

Comp. No	N-H	C-H ar.	C-H ali.	C=O amide	C=O  cyclic amide	C=N	C=C
9a	3192	3053	2988	1649		1623	1600
9b	3186	3062	2970	1676		1635	1589
9c	3227	3057	2960	1680		1604	1591
9d	3221	3062	2983	1660		1649	1600
10a	3223	3050	2966	1656	1693		1604
10b	3180	3047	2970	1664	1691		1600

Biological Activity

In this study the antimicrobial activity of the synthesized phenylquinazoline on two kinds of bacteria namely, staphylococcus aureous and Escherichia coli was investigated. Table (3) shows the antibacterial activity of some synthesized compounds and the obtained zones .Results shows that compound 7 exhibit highly antibacterial activity against Escherichia coli and  Staphylococcus aureus than other compounds.

Table 3 : Antibacterial activity of some synthezised compounds

	Zone of inhibition	(mm)
Comp. No	Escherichia coli	Staphylococcus aureus
5	16	15
6	25	25
7	40	28
8	14	16
9	12	14

 

Conclusions

A series of newly phenylquinazoline compounds were successfully synthesized and characterized. Some of the newly compounds were tested for antibacterial activity which compound 7 exhibit highly antibacterial activity against Escherichia coli and  Staphylococcus aureus than other compounds.

Acknowledgements

The author would like to thank the University of Baghdad for supporting this study and provide the grant for this study. We are also grateful to Dr.Zeinab M. AlRubaei for her contributions.

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