Synthesis and evaluation of antibacterial activity of some Benzopyran-4-one derivatives

 

Sukhen Som¹* and K.N. Jayaveera²

¹Department of Pharma. Chemistry, M.M.U College of Pharmacy, K. K. Doddi, Ramanagara- 571511Karnataka

²Department of Chemistry, Jawaharlal Nehru Technological University Anantapur, Anantapur, Andhra Pradesh,

*Corresponding Author E-mail: sukhen18@rediffmail.com

 

In view of the general observations that Benzopyrone derivatives are associated with antibacterial, anti-inflammatory, anticancer, antihepatotoxic activity etc. it was our interest to prepare different substituted Benzopyrone derivatives and evaluate them for their antibacterial activity.A series of substituted benzopyran-4-one derivatives were synthesized through various steps starting from 2-benzoyloxy-4-substituted acetophenone. The structures of the synthesized compounds were confirmed by their spectral and analytical data. Out of all synthesized compounds 6a and 6b and compounds 6f, 6i and 6k showed maximum activity against E.coli and S.aureus.

 

 

 

INTRODUCTION:

The inevitable consequence of the widespread use of antibacterial agents has been the emergence of antibiotic resistant pathogens, fuelling an over-increasing need for new drugs. However the pace of antimicrobial drug development has slowed dramatically with only a handful of new agents being introduced in the clinical practice each year. The major stream of investigation that validate the biologically active compounds reveals that of late the heterocyclic compounds have been increasingly important not only in the medicinal chemistry but in the field of agriculture and industry too. In view of the general observation that pharmacological activity is invariably associated with the diverse category of heterocyclic compounds, the present investigation of some heterocycle belonging to certain class of compound was undertaken.

 

Coumarins, chromones and flavones are pharmacologically important classes of plant products. Among these the flavonoids are considered to be potential for human health as well as constitute a significant part of daily human diet. Chemically flavonoids are very closely related to anthocyanins and occur as hydroxylated derivatives of 2-phenyl-4-chromone.

 

These are polyhydroxylated compounds and they are capable of selectively reacting with free radicals on systems related to the induction of inflammatory process. Quercetin and related flavonoids are known to inhibit the growth of tumour cells and potentiates the cytotoxicity of DNA damaging anticancer drugs such as cisplatin¹. Compounds having chromone moiety are associated with interesting physiological activities such as antioxidant^2-5, anticancer^6-8, antileukemic⁹, CNS depressant¹⁰ etc. Some reports are available on the influence of lipophilic substituents on the antioxidant or anti-inflammatory activities of this class of natural products¹¹. Certain bromoflavones are found to significantly induce quinine reductase activity which is an important mechanism of chemoprevention¹².

 

In view of the concerned observations and reliancing the above literatures in this study we report the synthesis and evaluation of antibacterial activity of some benzopyrone derivatives condensed with various amines.

 

MATERIALS AND METHODS:

The melting points were determined in open capillary tubes and are uncorrected. The IR spectra of the compounds were recorded in the region 4000-400 cm^­1 using KBr discs on a Shimadzu 8400S FTIR spectrophotometer. ¹HNMR spectra were recorded at 60 MHz on a Brukar Spectrospin-200 spectrometer using TMS as internal standard. Mass spectral studies were done in JEOL GC mate. Purity of the compounds was checked by TLC using precoated silica gel G plates, benzene: ethyl acetate (1:1) as mobile phase and iodine vapour and UV chamber as detection method.

 

Scheme of synthesis:

 

Table-1. Physical data of the synthesized compounds

 

 

 

Synthesis of 2, 4-dihydroxy-(3-chloro)dibenzoylmethane (2a)

2-(3-chlorobenzoyloxy)-4-benzoyloxyacetophenone (0.02 mol) was dissolved in 25ml of dry pyridine and heated to around 60⁰C. Then 0.03 mol of freshly fused and powdered potassium hydroxide was added. The resulting solution was stirred for about half an hour and then set aside for 2 hours to cool. Then it was acidified with 10% acetic acid. Precipitated product was filtered and washed 5 times with 20ml each portions of water and little cold alcohol, dried and then crystallized with methanol. Melting point125⁰C, yield 69%, R_f value 0.71. In a similar manner compounds 2b-f were synthesized. 2a IR (cm^-1): 3575 (O-H str), 3080 (C-H str aromatic), 2950 (C-H str aliphatic), 1685 (C=O str ketone), 1487 (C=C str aromatic), 750 (C-Cl str). 2a ¹H NMR (δ): 6.7-7.0 4H m Ar H, 6.7-7.0 1H methine proton (enol form), 7.4-7.6 3H m Ar H, 11.9-12.0 1H s phenolic OH, 12.2-12.3 1H s phenolic OH, 15.1-15.2 1H s enolic OH.

 

Synthesis of 7-hydroxy-2-(3-chlorophenyl)benzopyran-4-ones (3a)

0.025 mol of 2,4-dihydroxy-(3-chloro)dibenzoylmethane (2a) was dissolved in 35ml of glacial acetic acid and then added with shaking 2ml of concentrated sulphuric acid. With intermittent shaking the solution was then refluxed for 2 hours. The reaction mixture was then poured with stirrinng on crushed ice. The product was filtered and washed repeatedly with a large volume of water and dried. It was recrystallized from petroleum ether (60-80⁰C). Melting point 122⁰C, yield 63%, R_f value 0.81. Similarly compounds 3b-f were synthesized. 3a IR (cm^-1): 3475 (OH str), 3065 (C-H str aromatic), 1606 (C=O str ketone), 1573 (C=C vibration C2 and C3), 1488 (C=C str aromatic), 748 (C-Cl str). 3a ¹H NMR (δ): 6.6 1H s C-3 H, 7.3-7.5 4H m Ar H, 7.8-8.0 3H m Ar H, 11.8-11.9 1H s phenolic OH.

 

Synthesis of 6-acetyl-7-hydroxy-2-(3-chlorophenyl)benzopyran-4-ones (4a)

0.01 mol of 3a was added to 30 ml of nitrobenzene. To this added 0.01 mol of acetic anhydride and freshly powdered 0.022 mol of anhydrous aluminium chloride and then it was heated in an oil bath for around 4 hours. The flask was then removed, cooled. Added to this flask 75 gm of crushed ice followed by 4 ml of concentrated hydrochloric acid. The product obtained was filtered and recrystallized from 95% ethanol to get pure 4a. Melting point 205⁰C, yield 67%, R_f value 0.78. In a similar manner compounds 4b-f were synthesized. 4a IR (cm^-1): 3480 (O-H str), 3070 (C-H str aromatic), 2950 (C-H str aliphatic), 1610 (C=O str ketone), 1575 (C=C vibration C2 and C3), 1490 (C=C str aromatic), 752 (C-Cl str). 4a ¹H NMR (δ): 2.8-2.9 3H s CH₃, 6.3 1H s C-3 H, 7.0-7.2 4H m Ar H, 7.5-7.7 2H m Ar H, 11.8-11.91H s phenolic OH.

 

Synthesis of bromo derivative of 6-acetyl-7-hydroxy-2-(3-chlorophenyl)benzopyran-4-ones (5a)

0.01 mol of 4a was added to 50 ml of glacial acetic acid taken in beaker kept on a magnetic stirrer. To this solution added bromine drop wise over a period of 1 hour. After complete addition the resulting solution was stirred for around 6 hours till the evolution of hydrogen bromide ceases. The resulting solution was then poured on around 250 gm of crushed ice. Crystals were collected by filtration. The product was then recrystallized from 50% alcohol to give the pure compound. Melting point 165⁰C, yield 59%, R_f value 0.72. Similarly compounds 5b-f were synthesized. 5a IR (cm^-1): 3485 (OH str), 3068 (C-H str aromatic), 2948 (C-H str aliphatic), 1630 (C=O str ketone), 1574 (C=C str C2 and C3), 1485 (C=C str aromatic), 760 (C-Cl str), 575 (C-Br str). 5a ¹H NMR (δ): 3.5-3.6 2H s alkyl H, 6.7 1H s C-3 H, 7.4-7.6 4H m Ar H, 7.8-8.0 2H m Ar H, 12.1 1H s phenolic OH.

 

Synthesis of 6-acetylamino-7-hydroxy-2-(3-chlorophenyl)benzopyran-4-ones (6a)

To a solution of bromo derivative (5a, 0.01 mol) in absolute alcohol (25 ml) was added the amine (0.01 mol) and the mixture was heated under reflux for about 2 hours on water bath. A crystalline solid was separated out on standing the above solution overnight in cold condition. Filtered, washed with cold water and dried. The compounds 6b-l were synthesized in a similar manner. Physical data are reported in table 1. 6a IR (cm^-1): 3490 (OH str), 3068 (C-H str aromatic), 2950 (C-H str aliphatic), 1632 (C=O str ketone), 1575 (C=C str C2 and C3), 1482 (C=C str aromatic), 1120 (C-N str amine), 770 (C-Cl str). 6a ¹H NMR (δ): 3.6-3.7 2H s alkyl H, 4.1-4.3 8H m aliphatic H, 6.7 1H s C-3 H, 7.45-7.67 4H m Ar H, 7.79-8.01 2H m Ar H. 12.1 1H s phenolic OH.

 

Antibacterial Activity

All the synthesized compounds were screened in their DMF solution for their antibacterial activity by cup plate method against Staphylococcus aureus (gram +ve) and Escherichia coli (gram -ve). Nutrient agar was used as a culture medium. Test solutions were prepared by dissolving 1mg of the test compound in 1ml of DMF and 0.1ml of this solution was used for testing the antibacterial activity. The zones of inhibition formed after an incubation for 24 hrs at 37⁰ C, were measured in mm and are represented by (+), (++) and (+++) depending upon the diameter of the inhibited microbial growth zone. The control (CHCl₃) with solvent (DMF) under identical condition showed no activity.  Norfloxacin showed a zone of inhibition of 24 mm against both the bacteria Staphylococcus aureus and Escherichia coli.

 

RESULTS AND DISCUSSION:

The convenient starting material for the synthesis of the titled compounds was 2-bezoyloxy-4-substituted acetophenone (1a-f). The structure of 2,4-dihydroxy-(3-chloro)dibenzoylmethane (2a) synthesized from 1a was confirmed by the NMR spectrum, where it exhibited keto-enol tautomerism and as a result instead of appearing two methylene protons, it appeared as one methine proton at δ 6.7-7.0 and one enolic proton at δ 15.1-15.2. The NMR spectra of 3a revealed the disappearance of enolic proton and showed a proton at δ 6.6 which is identified as the C-3 proton of the benzopyrone ring, thus confirming the cyclisation of 2a to give 3a. Appearance of three alkyl protons as a sharp intense singlet at δ 2.8-2.9 in NMR spectrum of 4a confirms the acetylation of the benzopyrone (3a-f). In further step the brominated derivative (5a) of the acetylated compound showed a downfield shift of the signals for two protons at δ 3.5. Continued thereafter the title compounds 6a-l were synthesized by reacting morpholine and piperidine with different derivatives of bromoacetyl benzopyrone. All the synthesized compounds showed their characteristic absorption band i.e. 3475-3575 cm^-1 (OH str), 3065-3080 cm^-1 (C-H str aromatic), 1606-1685 cm^-1 (C=O) & 1482-1490 cm^-1 (C=C str aromatic) and proton signals in IR and ¹HNMR spectra respectively.

 

The preliminary antibacterial screening results revealed that many of the test compounds were effective against both the two strains of bacteria, however with a degree of variation. Compounds 6a & 6b exhibited highest antibacterial activity against E.coli. Compounds 6f, 6i & 6k were highly active against S. aureus, while compounds 6c, 6d, 6e & 6j displayed moderate activity and the rest of the compounds were weakly active.

 

Table-2. Antibacterial activity of the synthesized compounds

Compound code	Zone of inhibition
	S. aureus	E. coli
6a	++	+++
6b	++	+++
6c	++	++
6d	++	++
6e	++	+
6f	+++	+
6g	+	+
6h	+	+
6i	+++	++
6j	++	++
6k	+++	++
6l	+	+

+ = weakly active (9-13 mm), ++ = moderately active (14-18 mm), +++ = highly active (19-23 mm)

 

CONCLUSION:

It can be concluded from antibacterial screening that for the substituted benzopyrone derivatives the antibacterial activity is appreciable although further studies are required to determine their toxicity.

 

ACKNOWLEDGEMENT:

One of the author is thankful to the management specially Prof. Mohamed Khaleel, Principal, M.M.U College of Pharmacy, Ramanagara, Karnataka for providing necessary facilities to carry out the research work.

 

REFERENCES:

1.       Fritz B et al. Selected Novel Flavones Inhibit the DNA Binding or the DNA Religation Step of Eukaryotic Topoisomerase I. American Soc. of Biochem. and Mol. Bio. 271(4);1996:2262-70

2.       Cholbi MR et al. Inhibitory effects of phenolic compounds on CCl4-induced microsomal lipid peroxidation. Experientia.  47; 1991: 195-199.

3.       Mora A et al. Toxication of MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine) and analogs by monoamine oxidase: A structure-reactivity relationship study. Biochem Pharmacol. 40; 1990: 793-797

4.       Rios JL et al. Antioxidant activity of flavonoids from Sideritis javalambrensis. Phytochemistry. 31; 1992: 1947-1950

5.       Sanz MJ et al. Influence of a series of natural flavonoids on free radical generating systems and oxidative stress. Xenobiotica. 24; 1994: 689-699

6.       Cassady J M et al. Natural Products as a Source of Potential Cancer Chemotherapeutic and Chemopreventive Agents. J of Nat Prod. 53 (1);1990; 23-41

7.       Ono K et al. Differential inhibitory effects of various flavonoids on the activities of reverse transcriptase and cellular DNA and RNA polymerases Eur. J. of biochem. 190: 1990: 469–476

8.       Cushman M et al. Synthesis and Evaluation of Hydroxylated Flavones and Related Compounds as Potential Inhibitors of the Protein-Tyrosine Kinase. J of Nat Prod. 54 (5); 1991: 1345-1352

9.       Jiang Su. New medical college dictionary of traditional Chinese drugs. 1985; 5469

10.     Rastogi MK et al. Synthesis of 2-methyl-3-N substituted Amino Chromones as Potential CNS Agents. Ind. J of Chem. 16B; 1978: 895-897

11.     I L Finar. Organic Chemistry, Vol 2, 5^th ed. Pearson Education Asia Ltd

12.     Lynda L et al. Cancer Chemopreventive Activity Mediated by 4′-Bromoflavone, a Potent Inducer of Phase II Detoxification Enzymes. Cancer Research. 59; 1999: 578-585

 

 

 

Received on 27.02.2012         Modified on 07.03.2012

Accepted on 14.03.2012         © AJRC All right reserved