MANNICH BASES OF 7-HYDROXY-4 -METHYL AND 4, 6, 7-TRIMETHYL COUMARINS AS POTENTIAL FUNGICIDES FOR ASPERGILLUS FLAVUS AND PYRRICULARIA ORZAE

MANNICH BASES OF 7-HYDROXY-4 -METHYL AND 4, 6, 7-TRIMETHYL COUMARINS AS POTENTIAL FUNGICIDES FOR ASPERGILLUS FLAVUS AND PYRRICULARIA ORZAE

Manoj Kumar Srivastava *, Brij Kishore Singh and Anamika Pandey

Department of Chemistry, Mahatma Gandhi P. G. College, Gorakhpur, Uttar Pradesh, India.

ABSTRACT: Coumarin and its derivatives are widely used as scaffolds in synthesizing new heterocyclic systems. Numerous approaches especially involving nano-particle catalysts, have been developed to get new bioactive coumarin derivatives endowed with pharmacological and biological activities. The present work describes the reactivity and the new strategies for the synthesis of coumarin and its derivatives reported in the literature and their biological properties. Coumarins possess a number of biological activities like Anticoagulant, Antimicrobial, Anti-inflammatory, Analgesic, Antioxidant, Anticancer, Antiviral, Anti-malarial etc. Coumarin belongs to a group as benzopyrones, which consists of a benzene ring joined to a pyrone nucleus ¹². Coumarins (2H-1-benzopyran-2-ones) are important oxygen-containing fused heterocycles used in drugs and dyes. A number of coumarin derivatives have been reported to have diverse biological activities like fungicidal^1-4, molluscicidal ², antihelminthic ^{5, 6} and CNS ⁷ stimulants. Mannich Bases of some heterocyclic compounds are also reported Bioactive ^{8, 9}. In view of this Mannich bases of 7 – hydroxyl – 4 – methyl coumrin and 4, 6, 7– trimethyl coumaryl incorporating various DL–amino acids as a basic component were synthesized and evaluated for their fungitoxicity against A. flavus and P. oryzae.

Keywords: Coumarin, Mannich bases, Amino acids, Aldehydes, Fungicidal activities, Aspergillus flavus and Pyricularia oryzae

INTRODUCTION: Coumarins (2H-1-benzopyran -2-ones) are important oxygen-containing fused heterocycles used in drugs ¹³ and dyes. Coumarins be bound their class name to ‘coumarou’ the vernacular name of the Tonka bean (Dipteryx odorata wild, Fabaceae), from which coumarin was isolated in 1820. They are the family of lactones containing benzopyrone skeletal framework that have enjoyed isolation from the plant and total synthesis in the laboratory. The present work describes the reactivity and the new strategies for synthesizing coumarin and its derivatives reported in the literature and their biological properties ¹⁴.

In view of this Mannich bases of 7– hydroxyl – 4 – methyl coumrin and 4,6,7–trimethyl coumaryl incorporating various DL–amino acids as a basic component was synthesized and evaluated for their fungitoxicity against A. flavus and P. oryzae

MATERIAL AND METHODS: The required 7– hydroxyl – 4 – methyl coumarin (1) was prepared following the method of Pachmann and diusherg ¹⁰ by the condensation of ethyl acetoacetate with resorcinol in the presence of Conc. H₂SO₄.

Synthesis of 7-Hydroxy-4-Methyl Coumarin: About 150 ml. of conc. H₂SO₄ in a 500 ml beaker was stirred with external ice water cooling until the temperature of acid became about 5°C – 10°C,  37 gm of powdered resorcinol was added to 45 ml. of ethyl acetoacetate until a complete solution was obtained. Then this solution was added slowly to H₂SO₄. In such a way, the temperature did not rise above 100C, and the stirring was continued for ½ an hour. The mixture is poured into the ice/cold water & the solid product is separated, filtered out and dried. Then the crude product was recrystallized from absolute ethanol. The resultant Yields: 85%, Melting point: 192°C, Molecular Formula: C₁₀H₈O₃, Molecular Mass: 176.17 and Solubility: Methanol, Ethanol, and Pyridine. Condensation of (1) with DL – amino acids or aldehydes in the presence of formaldehyde and methanol, furnished Mannich bases (2) and (3), respectively as shown in Scheme I.

SCHEME I.

The structural assignments of the synthesized compounds have been established by elemental analysis and spectral data. IR spectra recorded on a Perkin-Elmer-15 spectrophotometer and 90 MHz PMR (DMSO d₆) on Varian EM 390 spectrometer (Chemical shift in δ-ppm). Melting points were taken open glass capillary and are uncorrected.

Synthesis of test Compounds: N-[(7-hydroxy – 4 – methyl coumarin – 8 – yl)] methyl glycine (2A): A mixture of N-(7-hydroxy–4–methyl coumarin (0.01M), glycine (0.01M) and formaldehyde (0.01M) in methanol (20 Ml) was reflux for 5 hours. Poured the mixture into ice cold water. Solid thus obtained was filtered, washed with water, and recrystallized from ethanol.

Yield 90%, m.p. 203°C, IR (in KBr) cm^-1ν_max 3470 (OH), 3200 (NH), 1670 (C=O), 1570, 1555, 1430, 1390 (aromatic ring); PMR (DMSO-d₆): δ (s, 3H, CH₃), 3.9 (s, 2H, CH₂NH), 6.6 – 7.8 (m, 4H, ArH and NH). Other compounds 2b – 2j were also synthesized similarly in Table 1.

TABLE 1: PHYSICAL CHARACTERISATION DATA OF COMPOUND (2)

N-[(7 – hydroxyl – 4 – methyl coumarin – 8 – yl)] methyl/ethyl substituted amines (3A) : N-(7-hydroxy – 4 – methyl coumarin (0.01M), dimethyl amine (0.01M) and formaldehyde (0.01M) in methanol (25 Ml) and water (2 Ml) were refluxed for 5 hours, the resulting mixture was cooled and poured into water and recrystallized from ethanol. Yield 72%, m.p. 163°C, IR (in KBr): 3480 (OH), 1670 (C=O), 1600, 1580, 1440 (aromatic ring); PMR (DMSO- d₆) δ 1.7 (s, 3H, CH₃), 2.6 (s, 6H, CH₃-N-CH₃), 3.1 (s, 2H, CH₂-N), 6.1 – 6.9 (m, 3H, ArH). Other compounds 3b – 3h were synthesized by similar methods Table 2.

TABLE 2: PHYSICAL CHARACTERISATION DATA OF COMPOUND (3)

Fungitoxicity: The fungicidal activity was evaluated against Aspergillus flavus and Pyricularia oryzae by Agar growth technique ¹¹ at 100 and 10 ppm concentration by adding a solution of the test compounds in acetone: water (20: 80; v/v) mixture to pre-sterilized Petri-dishes containing Czepek’s agar and mixing thoroughly. One week old culture of the test fungi was inoculated in centre of each Petri-dish. The bio-assay was done in three replicates. The plates were incubated at 28+2°C for 7 days. The percentage inhibition of the mycelia growth or spore germination was calculated by the following formula.

% Inhibition = C – T / C x 100

Where, C = average diameter (in mm) of the fungal colony in control plates, T = average diameter (in mm) of the fungal colony in treated plates,

ED₅₀ – Values were calculated by the log probability method. The commercial fungicide Dithane M-45 was maintained as a reference for comparison in Table 3.

TABLE 3: FUNGICIDAL ACTIVITY OF COMPOUNDS (2 & 3)

RESULTS AND DISCUSSIONS:

Fungicidal Activity: The fungicidal activity of the test compounds of type 2 was comparatively more active than the compound of type 3. The most active compounds were 2e, 2f, 2i, 2j, 3d and 3g. By comparing the data of compound numbers 2 and 3, it was clear that amino acid derivatives played an important role. The presence of groups like –CH₃, and CH₂-CH-(CH₃)₂ accounted for their fungicidal activity.

The most active compounds 2e, 2f, 2i, 2j, 3d and 3g showed comparatively inferior fungi toxicity than the commercial fungicide Dithane M-45 at 100 and 10 ppm against both the test fungi. The activity decrease on dilution.

CONCLUSIONS: From the above results and discussion, Mannich bases with coumarin and its derivatives enhance the fungicidal activities. The presence of groups like –CH₃ and CH₂-CH-(CH₃)₂ accounted for their fungicidal activity.    

ACKNOWLEDGEMENTS: The authors thank the Director (RSIC) CDRI, Lucknow, India, for recording IR and PMR spectra. The financial support received from UGC New Delhi and we are also thankful to our management for moral support and Laboratory facilities.

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    How to cite this article:

    Srivastava MK, Singh BK and Pandey A: Mannich bases of 7-hydroxy-4 -methyl and 4, 6, 7-trimethyl coumarins as potentential fungicides for Aspergillus flavus and Pyrricularia orzae. Int J Pharm Sci & Res 2023; 14(3): 1261-64. doi: 10.13040/IJPSR.0975-8232.14(3).1261-64.

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