Solvent-free microwave-assisted synthesis and biological evaluation of aurones and flavanones based on 2,2-dimethylchroman-4-one

A series of novel 8-aryl-2,2-dimethyl-2,3,7,8-tetrahydropyrano[3,2-g]chromene-4,6-dione and 2-arylidene-7,7-dimethyl-6,7-dihydro-2H-furo[3,2-g]chromene-3,5-dione derivatives have been synthesized by oxidation of the corresponding chalcones under microwave irradiation. All newly synthesized compounds were characterized by IR, ¹H, ¹³C NMR, and mass spectra and valuated for their in vitro antibacterial and antifungal activities.

Synthesis of heterocyclic compounds has emerged as a powerful technique for generating new molecules useful for drug design.1 In particular, five- and six-membered heterocycles occupy a unique position due to dominant pharmacological applications.2 Flavonoids and their subclass flavanones occur in nature as polyphenolic compounds that have been reported to possess anticancer or anticarcinogenic (antimutagenic) activities.3 ^, 4 In this family, flavanones have been widely employed as important intermediates in the synthesis of many natural products and medicinal agents. They are widely distributed in nature displaying diverse range of biological activities. Aurones, containing a 1-benzofuran ring system, constitute a less studied subclass of flavonoids, which occur rarely in nature. Aurones are biologically important compounds due to their anticancer,5 antibacterial,6 and anti-inflammatory7 activities.

Chroman-4-ones and chromen-4-ones (chromones) that contain a 1-benzopyran ring system and constitute the principal structural motif of flavonoids have been also used as scaffolds for the development of peptidomimetics. Depending on the substitution pattern these chromene derivatives show different biological effects.8 ^– 10 In addition, the 2,2-dimethyl-substituted chromane ring system is found in many widely varied natural compounds. For example, vitamin E (D-α-tocopherol)11 is a natural radical scavenger that suppresses cellular membrane phospholipid degradation and also exhibits antioxidant, anticancer, and cardioprotective activities.12 This has determined our interest in the synthesis and evaluation of the antimicrobial activity of diverse classes of compounds containing condensed pyran framework.

In recent years, microwave-assisted organic synthesis has attracted the attention of synthetic chemists as a new tool in organic synthesis.13 This technique offers simple, clean, fast, and efficient synthesis of a large number of organic molecules.

In view of the biological importance of chromanone, flavanones, and aurones, we describe in this report the synthesis of a series of substituted flavanones (8-aryl-2,2-dimethyl-2,3,7,8-tetrahydropyrano[3,2-g]chromene-4,6-diones) and aurones ((Z)-2-arylidene-7,7-dimethyl-6,7-dihydro-2H-furo-[3,2-g]chromene-3,5-diones) under solvent-free microwave (MW) irradiation, as well as preliminary tests of their antimicrobial activity. The target flavanones and aurones were obtained in three steps. The chromene synthons 6-acetyl-7-hydroxy-2,2-dimethylchroman-4-one (2a)14 and 2,2,8,8-tetramethyl-2,3,7,8-tetrahydropyrano[3,2-g]chromene-4,6-dione (2b)14 were prepared from 4,6-diacetylresorcinol-(1,1'-(4,6-dihydroxybenzene-1,3-diyl)diethanone) (1) and acetone in the presence of pyrrolidine and ethanol as shown in Scheme 1. Compounds 2a and 2b were isolated by column chromatography in the ratio of 3:1. Compound 2a was then subjected to the Claisen–Schmidt condensation with different aromatic aldehydes 3a–h in 40% aq KOH and ethanol to yield the corresponding chalcones 4a–h (Scheme 2).

Scheme 1

Scheme 2

Further, chalcones 4a–h were cyclized in the presence of trifluoroacetic acid (TFA) under MW irradiation to afford the corresponding flavanones 5a–g in good yields. Aurones 6a–h were synthesized by the oxidation of chalcones 4a–h with mercuric acetate under MW irradiation in moderate to good yields (Table 1).

Table 1 Synthesis of flavanones 5a–g and 6a–h

The structures of compounds 5a–g and 6a–h were characterized by IR, ¹H and ¹³C NMR, and mass spectroscopy, as well as elemental analysis. IR spectra of compounds 5a–g featured the signal of carbonyl groups at 1679–1704 cm^–1. The ¹H NMR spectra of compounds 5a–g showed three signals characteristic of an ABX proton system consisting of diastereotopic protons of the 7-CH₂ group and 8-CH proton at the chiral carbon atom of the flavanone pyran ring. The methylene proton which is cisoriented relative to the 8-CH proton is observed as a doublet of doublets (J _AB = 16.9, J _AX = 2.6–3.2 Hz) at a higher field (2.92–2.87 ppm) than the trans-oriented methylene proton (J _BA = 16.9, J _BX = 12.5–12.9 Hz) (3.08–3.00 ppm). The 8-CH proton (5.46–5.53 ppm) signal had both matching vicinal spin-spin interaction constants (J _AX and J _BX). The proposed structure of compounds 5a–g was further supported by the ¹³C NMR spectrum, which contained the signals at 42.7–44.3 and 48.6–48.8 ppm that could be attributed to the 7-CH₂ and 3-CH₂ groups, respectively (only the latter signal is observed in the spectra of compounds 4a–h and 6a–h).

The IR spectra of compounds 6a–h showed a characteristic absorption band carbonyl groups (1671–1709 cm^–1). The ¹H NMR spectra of the representative compounds 6a–h showed a characteristic singlet at 6.72–6.85 ppm due to the benzylidene methine proton. In the ¹³C NMR spectrum, the signal of carbonyl carbon in the aurone five-membered ring appeared at 181.0–186.5 ppm. The NMR spectroscopic data of the synthesized aurones prove that this oxidative cyclization procedure yields one isomer. The GC-MS spectra of all compounds 5a–g and 6a–h exhibited [M+H]+ peaks with the expected m/z values.

All synthesized compounds 5a–g and 6a–h were screened for in vitro antibacterial activity against two Grampositive bacterial strains Staphylococcus aureus (ATCC-6538), Bacillus subtilis (ATCC-6633) and two Gram-negative bacterial strains Escherichia coli (ATCC-25922) and Klebsiella pneumoniae (ATCC-13883) by the disc diffusion method15 at different concentrations (20 and 40 μg/ml). Nutrient agar medium was used for the antibacterial screening. The zone of inhibition (in mm) was compared with that of standard drug ciprofloxacin. The results are presented in Table 2.

Table 2 Antimicrobial and antifungal activity of aurones 5a–g and 6a–h (zone of inhibition, mm)

All the synthesized compounds showed a better activity against Gram-negative bacterial strains than Gram-positive bacterial strains at the concentrations 20 and 40 μg/ml. Flavanones 5a,e–g and aurones 6a,e–h containing an aryl group with electron donor properties showed a promising activity against all bacterial strains. Compounds 5b–d and 6b–d with electron-withdrawing halogen substituents at the phenyl ring showed moderate zone of inhibition against all the bacterial strains. Compounds 5f, 6f, and 6h showed zone of inhibition (31.5, 33.0, and 32.0 mm, respectively) comparable to the standard drug ciprofloxacin (33.4 mm) against B. subtilis at the concentration 40 μg/ml. Compounds 5f (23.2 mm) and 6h (25.0 mm) showed a good activity against the Gram-negative bacterial strain K. pneumoniae at the concentration 20 μg/ml.

All the synthesised compounds were screened for their antifungal activity against two pathogenic fungi, Aspergillus niger and Fusarium oxysporum by the poison plate technique.16 The results of the antifungal screening were compared with standard antifungal drugs amphotericin B and hymexazol. All the compounds showed moderate activity against the tested fungal strains. The activity of compounds 6f,g against A. niger was slightly higher than that of standard drug amphotericin B.

In summary, we have successfully developed synthesis of novel aurone and flavanone derivatives under solventfree microwave irradiation. The present method should be economically feasible because the products can be obtained through a facile, two-step reaction with commercially available and relatively inexpensive chemicals. Some of the synthesized compounds show promising antimicrobial activities compared with commercial drugs while all the rest show moderate activity against the tested organisms. Electron-donating substituents on the flavanone or aurone phenyl group appear to diminish the antimicrobial activity compared to other substituents.

Experimental

FT-IR spectra were recorded in KBr pellets on a Perkin Elmer spectrophotometer. ¹H and ¹³C NMR spectra were recorded on a Brucker DRX-400 instrument (400 and 100 MHz, respectively) in CDCl₃. Chemical shift values are reported relative to TMS as internal reference. Mass spectra were obtained on a Jeol SX-102 mass spectrometer (ESI). Elemental analyses were recorded on a Karlo Erba 1106 elemental analyzer. Melting points were determined in open capillary tubes and are uncorrected. Microwave reactions were carried out in a Milestone multi SYNTH series (ATC-FO 300) multimode microwave reactor with a twin magnetron (2 × 800 W, 2.45 GHz) and the maximum delivered power of 1,000 W in 10 W increments (pulsed irradiation). All the reactions were monitored by TLC on Merck Kieselgel 60 F524, visualized by UV light and/or spraying a 5% H₂SO₄ in EtOH followed by heating. Column chromatography was performed on Silica Gel 60 (60–120 mesh). All the available reagent grade chemicals were purchased from Sigma-Aldrich or Spectrochem Pvt. Ltd. and used without further purification. Solvents were dried according to standard methods.

Synthesis of 6-acetyl-7-hydroxy-2,2-dimethylchroman-4-one (2a) and 2,2,8,8-tetramethyl-2,3,7,8-tetrahydropyrano-[3,2-g]chromene-4,6-dione (2b) followed a published procedure.14

6-Acetyl-7-hydroxy-2,2-dimethylchroman-4-one (2a). ¹H NMR spectrum, δ, ppm: 1.40 (6H, s, 2CH₃); 2.40 (3H, s, CH₃); 2.74 (2H, s, CH₂); 6.50 (1H, s, H Ar); 7. 55 (1H, s, H Ar).

2,2,8,8-Tetramethyl-2,3,7,8-tetrahydropyrano[3,2- g ]-chromene-4,6-dione (2b). ¹H NMR spectrum, δ, ppm: 1.45 (12H, s, 4CH₃); 2.60 (4H, s, 2CH₂); 6.30 (1H, s, H Ar); 8.33 (1H, s, H Ar).

Synthesis of chalcones 4a–h by Claisen–Schmidt condensation (General method). 6-Acetyl-7-hydroxy-2,2-dimethylchroman-4-one (2a) (0.5 g, 0.002 mol) and an aromatic aldehyde 3a–h (0.002 mol) were dissolved in minimum amount of ethanol. Aqueous potassium hydroxide solution (40%, 1.2 ml, 0.015 mol) was added slowly, and the reaction mixture was stirred at room temperature for 24 h. The progress of the reaction was monitored by TLC (eluent n-hexane–AcOEt, 6:4). After completion of reaction, the reaction mixture was poured onto crushed ice, carefully neutralized with 3 N HCl, and extracted with EtOAc (15 ml). The organic layer was concentrated in vacuum and purified by column chromatography on silica gel (eluent n-hexane–AcOEt, 3:1).

7-Hydroxy-6-[(2 E )-3-phenylprop-2-enoyl]-2,2-dimethylchroman-4-one (4a). Yield 65%, yellow solid, mp 148–150°C. IR spectrum, ν, cm^–1: 1203, 1367, 1564 (C=C), 1637 (C=O), 1683, 2970 (O–H). 1H NMR spectrum, δ, ppm (J, Hz): 1.49 (6H, s, 2CH₃); 2.75 (2H, s, CH₂); 6.46 (1H, s, H Ar); 7.45–7.46 (3H, m, H Ar); 7.69–7.71 (3H, m, =CH, H Ar); 7.94 (1H, d, J = 15.6, =CH); 8.58 (1H, s, H Ar); 13.54 (1H, s, OH). ¹³C NMR spectrum, δ, ppm: 26.8; 48.6; 80.2; 105.9; 113.4; 114.1; 118.0; 122.3; 127.5; 128.5; 130.8; 135.2; 145.1; 167.8; 170.3; 190.8; 192.8. Mass spectrum, m/z: 323 [M+H]⁺. Found, %: C 74.50; H 5.60. C20H18O4. Calculated, %: C 74.52; H 5.63.

6-[(2 E )-3-(4-Fluorophenyl)prop-2-enoyl]-7-hydroxy-2,2-dimethylchroman-4-one (4b). Yield 77%, yellow solid, mp 195–197°C. IR spectrum, ν, cm^–1: 1201, 1359, 1558 (C=C), 1636 (C=O), 1679, 2974 (O–H). ¹H NMR spectrum, δ, ppm (J, Hz): 1.49 (6H, s, 2CH3); 2.74 (2H, s, CH2); 6.46 (1H, s, H Ar); 7.19 (2H, s, H Ar); 7.71 (2H, s, H Ar); 7.56 (1H, d, J = 15.3, =CH); 8.04 (1H, d, J = 15.6, =CH); 8.55 (1H, s, H Ar); 13.62 (1H, s, OH). ¹³C NMR spectrum, δ, ppm: 26.8; 48.6; 80.2; 105.9; 113.4; 114.1; 115.4; 118.7; 130.8 (2C); 131.0; 135.2; 145.1; 162.1; 167.8; 170.3; 190.9, 192.8. Mass spectrum, m/z: 341 [M+H]⁺.

6-[(2 E )-3-(4-Chlorophenyl)prop-2-enoyl]-7-hydroxy-2,2-dimethylchroman-4-one (4c). Yield 67%, yellow solid, mp 220–222°C. IR spectrum, ν, cm^–1: 1204, 1368, 1569 (C=C), 1632 (C=O), 1691, 2971 (O–H). ¹H NMR spectrum, δ, ppm (J, Hz): 1.49 (6H, s, 2CH₃); 2.74 (2H, s, CH₂); 6.46 (1H, s, H Ar); 7.44 (2H, s, H Ar); 7.59–7.68 (3H, m, =CH, H Ar); 8.05 (1H, d, J = 15.5, =CH); 8.57 (1H, s, H Ar); 13.55 (1H, s, OH). ¹³C NMR spectrum, δ, ppm: 26.8; 48.6; 80.2; 105.7; 113.4; 114.1; 118.7; 128.7; 129.0; 130.7; 133.3; 133.5; 145.1; 162.1; 168.8; 170.3; 190.9; 192.8. Mass spectrum, m/z: 357 [M(³⁵Cl)+H]⁺. Found, %: C 67.30; H 4.75. C₂₀H₁₇ClO₄. Calculated, %: C 67.32; H 4.80.

6-[(2 E )-3-(4-Bromophenyl)prop-2-enoyl]-7-hydroxy-2,2-dimethylchroman-4-one (4d). Yield 90%, yellow solid, mp 215–217°C. IR spectrum, ν, cm^–1: 1211, 1375, 1572 (C=C), 1645 (C=O), 1689, 2978 (O–H). ¹H NMR spectrum, δ, ppm (J, Hz): 1.49 (6H, s, 2CH₃); 2.74 (2H, s, CH₂); 6.44 (1H, s, H Ar); 7.50 (2H, s, H Ar); 7.59–7.61 (3H, m, =CH, H Ar); 7.98 (1H, d, J = 15.5, =CH); 8.56 (1H, s, H Ar); 13.64 (1H, s, OH). ¹³C NMR spectrum, δ, ppm: 26.8; 48.6; 80.2; 105.7; 113.1; 114.1; 118.7; 122.3; 128.7; 130.7; 131.3; 134.2; 145.1; 168.8; 170.3; 190.9; 192.8. Mass spectrum, m/z: 401 [M(⁷⁹Br)+H]⁺. Found, %: C 59.80; H 4.82. C₂₀H₁₇BrO₄. Calculated, %: C 59.87; H 4.27.

7-Hydroxy-2,2-dimethyl-6-[(2 E )-3-(4-methylphenyl)-prop-2-enoyl]chroman-4-one (4e). Yield 70%, yellow solid, mp 180–183°C. IR spectrum, ν, cm^–1: 1204, 1370, 1568 (C=C), 1640 (C=O), 1689, 2971 (O–H). ¹H NMR spectrum, δ, ppm (J, Hz): 1.49 (6H, s, 2CH₃); 2.41 (3H, s, CH₃); 2.74 (2H, s, CH₂); 6.46 (1H, s, H Ar); 7.25–7.27 (2H, m, H Ar); 7.59–7.65 (3H, m, =CH, H Ar); 7.92 (1H, d, J = 15.3, =CH); 8.57 (1H, s, H Ar); 13.52 (1H, s, OH). ¹³C NMR spectrum, δ, ppm: 21.3; 26.8; 48.6; 80.2; 105.9; 113.4; 114.1; 118.5; 128.5; 128.9; 130.7; 132.2; 137.6; 145.1; 167.8; 170.3; 190.8; 192.8. Mass spectrum, m/z: 337 [M+H]⁺. Found, %: C 74.92; H 5.94. C₂₁H₂₀O₄. Calculated, %: C 74.98; H 5.99.

7-Hydroxy-6-[(2 E )-3-(4-methoxyphenyl)prop-2-enoyl]-2,2-dimethylchroman-4-one (4f). Yield 75%, yellow solid, mp 176–178°C. IR spectrum, ν, cm^–1: 1207, 1369, 1552 (C=C), 1633 (C=O), 1687, 2972 (O–H). ¹H NMR spectrum, δ, ppm (J, Hz): 1.49 (6H, s, 2CH₃); 2.74 (2H, s, CH₂); 3.88 (3H, s, OCH₃); 6.45 (1H, s, H Ar); 6.91 (2H, d, J = 8.8, H Ar); 7.55 (1H, d, J = 15.3, =CH); 7.66 (2H, d, J = 8.8, H Ar); 7.91 (1H, d, J = 15.3, =CH); 8.57 (1H, s, H Ar); 13.71 (1H, s, OH). ¹³C NMR spectrum, δ, ppm: 26.8; 48.6; 55.8; 80.2; 105.7; 113.4; 114.1; 114.2; 118.7; 127.7; 130.2; 130.7; 145.2; 168.8; 170.3; 190.9; 192.8. Mass spectrum, m/z: 353 [M+H]⁺. Found, %: C 71.54; H 5.70. C₂₁H₂₀O₅. Calculated, %: C 71.58; H 5.72.

7-Hydroxy-6-[(2 E )-3-(4-isopropylphenyl)prop-2-enoyl]-2,2-dimethylchroman-4-one (4g). Yield 70%, yellow solid, mp 153–155°C. IR spectrum, ν, cm^–1: 1213, 1364,

1555 (C=C), 1643 (C=O), 1678, 2984 (O–H). ¹H NMR spectrum, δ, ppm (J, Hz): 1.28 (6H, d, J = 6.8, (CH ₃)₂CH); 1.52 (6H, s, 2CH₃); 2.78 (2H, s, CH₂); 2.96 (1H, sept, J = 6.8, (CH₃)₂CH); 6.79 (1H, s, H Ar); 7.32 (2H, d, J = 7.9, H Ar); 7.82 (2H, d, J = 7.9, H Ar); 7.90 (1H, d, J = 15.3, =CH); 7.52 (1H, d, J = 15.3, =CH); 8.58 (1H, s, H Ar); 13.72 (1H, s, OH). ¹³C NMR spectrum, δ, ppm: 23.3; 26.7; 33.2; 48.3; 80.2; 105.7; 113.4; 114.1; 118.7; 126.0; 128.3; 130.7; 132.4; 145.1; 147.6; 168.8; 170.3; 190.9; 192.8. Mass spectrum, m/z: 365 [M+H]⁺. Found, %: C 75.76; H 6.60. C₂₃H₂₄O₄. Calculated, %: C 75.80; H 6.64.

7-Hydroxy-2,2-dimethyl-6-[(2 E )-3-(naphthalen-1-yl)-prop-2-enoyl]chroman-4-one (4h). Yield 66%, yellow solid, mp 169–171°C. IR spectrum, ν, cm^–1: 1201, 1367, 1556 (C=C), 1631 (C=O), 1683, 2978 (O–H). ¹H NMR spectrum, δ, ppm (J, Hz): 1.49 (6H, s, 2CH3); 2.75 (2H, s, CH2); 6.47 (1H, s, H Ar); 6.92 (2H, d, J = 8.7, H Ar); 7.48–7.51 (3H, m, H Ar); 7.55 (1H, d, J = 15.3, =CH); 7.91 (1H, d, J = 15.3, =CH); 8.21 (1H, d, J = 8.4, H Ar); 8.28 (1H, d, J = 7.3, H Ar); 8.57 (1H, s, H Ar); 13.71 (1H, s, OH). ¹³C NMR spectrum, δ, ppm: 26.8; 48.6; 80.7; 105.7; 113.4; 114.1; 121.3; 122.6; 124.0; 126.0; 126.3; 126.9; 126.9; 128.3; 128.8; 130.7; 132.0; 133.5; 133.6; 135.6; 168.8; 170.3; 190.9; 192.8. Mass spectrum, m/z: 373 [M+H]⁺. Found, %: C 77.35; H 5.35. C₂₄H₂₀O₄. Calculated, %: C 77.40; H 5.41.

Synthesis of flavanones 5a–g (General method). A mixture of chalcone 4a–g (10 mmol) and TFA (2 ml) was placed in a quartz tube which was inserted into a screw-capped Teflon vial and then was subjected to microwave irradiation at 320 W for 3–5 min. After completion of the reaction (as indicated by TLC), the reaction mixture was poured into ice-cold water, extracted with dichloromethane (2×30 ml), and the organic layer was dried over Na₂SO₄, purified by column chromatography (eluent n-hexane–AcOEt, 9:1).

2,2-Dimethyl-8-phenyl-2,3,7,8-tetrahydro-4 H ,6 H pyrano[3,2- g ]chromene-4,6-dione (5a). IR spectrum, ν, cm^–1: 2981, 1688 (C=O), 1600, 1464, 1229, 1149. ¹H NMR spectrum, δ, ppm (J, Hz): 1.46 (3H, s, CH3); 1.48 (3H, s, CH3); 2.72 (2H, s, CH2); 2.89 (1H, dd, J = 16.9, J = 3.1) and 3.06 (1H, dd, J = 16.9, J = 12.8, 7-CH2); 5.50 (1H, dd, J = 12.8, J = 3.0, 8-CH); 6.53 (1H, s, H Ar); 7.40–7.48 (5H, m, H Ph); 8.56 (1H, s, H Ar). ¹³C NMR spectrum, δ, ppm: 26.6; 27.0; 44.3; 48.7; 79.9; 80.5; 105.4; 115.9; 126.2; 128.4; 128.9; 129.0; 129.7; 138.2; 165.3; 166.5; 189.8; 190.3. Mass spectrum, m/z: 323 [M+H]⁺. Found, %: C 74.48; H 5.60. C₂₀H₁₈O₄. Calculated, %: C 74.52; H 5.63.

8-(4-Fluorophenyl)-2,2-dimethyl-2,3,7,8-tetrahydro-4 H ,6 H -pyrano[3,2- g ]chromene-4,6-dione (5b). IR spectrum, ν, cm^–1: 2990, 1690 (C=O), 1605, 1469, 1231, 1165. ¹H NMR spectrum, δ, ppm (J, Hz): 1.46 (3H, s, CH₃); 1.48 (3H, s, CH₃); 2.72 (2H, s, CH₂); 2.87 (1H, dd, J = 16.9, J = 3.1) and 3.03 (1H, dd, J = 16.9, J = 12.7, 7-CH₂); 5.48 (1H, dd, J = 12.7, J = 3.0); 6.52 (1H, s, H Ar), 7.09–7.17 (2H, m, H Ar); 7.42–7.48 (2H, m, H Ar); 8.56 (1H, s, H Ar). ¹³C NMR spectrum, δ, ppm: 26.6; 27.0; 44.3; 48.6; 79.2; 80.6; 105.4; 115.8; 116.0; 128.1; 128.4; 134.0; 134.1; 161.7; 164.2; 165.4; 166.3; 189.6; 190.4. Mass spectrum, m/z: 341 [M+H]⁺. Found, %: C 70.51; H 5.00. C₂₀H₁₇FO₄. Calculated, %: C 70.58; H 5.03.

8-(4-Chlorophenyl)-2,2-dimethyl-2,3,7,8-tetrahydro-4 H ,6 H -pyrano[3,2- g ]chromene-4,6-dione (5c). IR spectrum, ν, cm^–1: 2983, 1704 (C=O), 1602, 1461, 1227, 1157. ¹H NMR spectrum, δ, ppm (J, Hz): 1.46 (3H, s, CH₃); 1.48 (3H, s, CH₃); 2.72 (2H, s, CH₂); 2.88 (1H, dd, J = 16.9, J = 3.2) and 3.01 (1H, dd, J = 16.9, J = 12.5, 7-CH2); 5.48 (1H, dd, J = 12.5, J = 3.2, 8-CH); 6.52 (1H, s, H Ar); 7.36–7.45 (4H, m, H Ar), 8.56 (s, 1H, Ar-H). ¹³C NMR spectrum, δ, ppm: 26.7; 27.0; 42.7; 48.6; 79.8; 80.8; 104.6; 115.9; 116.8; 126.9; 128.0; 128.6; 129.0; 133.2; 136.4; 166.4; 167.6; 189.9; 190.6. Mass spectrum, m/z: 357 [M+H]⁺. Found, %: C 67.30; H 4.75. C₂₀H₁₇ClO₄. Calculated, %: C 67.32; H 4.80.

8-(4-Bromophenyl)-2,2-dimethyl-2,3,7,8-tetrahydro-4 H ,6 H -pyrano[3,2- g ]chromene-4,6-dione (5d). IR spectrum, ν, cm^–1: 2963, 1679 (C=O), 1604, 1463, 1231, 1160. ¹H NMR spectrum, δ, ppm (J, Hz): 1.46 (3H, s, CH₃); 1.48 (3H, s, CH₃); 2.73 (2H, s, CH₂); 2.88 (1H, dd, J = 16.9, J = 3.2) and 3.00 (1H, dd, J = 16.9, J = 12.5, 7-CH₂); 5.47 (1H, dd, J = 12.5, J = 3.2, 8-CH); 6.52 (1H, s, H Ar); 7.32–7.36 (2H, m, H Ar); 7.56–7.59 (2H, m, H Ar); 8.55 (1H, s, H Ar). ¹³C NMR spectrum, δ, ppm: 26.7; 27.0; 42.7; 48.6; 80.8; 81.8; 105.1; 116.0; 116.3; 122.0; 127.2; 128.8; 131.8; 137.3; 166.6; 167.8; 189.9; 190.6. Mass spectrum, m/z: 401 [M(⁷⁹Br)+H]⁺. Found, %: C 59.82; H 4.22. C₂₀H₁₇BrO₄. Calculated, %: C 59.87; H 4.27.

2,2-Dimethyl-8-(4-methylphenyl)-2,3,7,8-tetrahydro-4 H ,6 H -pyrano[3,2- g ]chromene-4,6-dione (5e). IR spectrum, ν, cm^–1: 2981, 1691 (C=O), 1604, 1467, 1229, 1158. ¹H NMR spectrum, δ, ppm (J, Hz): 1.46 (3H, s, CH₃); 1.48 (3H, s, CH₃); 2.39 (3H, s, CH₃); 2.72 (2H, s, CH₂); 2.87 (1H, dd, J = 16.9, J = 2.9) and 3.06 (1H, dd, J = 16.9, J = 12.8, 7-CH₂); 5.46 (1H, dd, J = 12.7, J = 2.6, 8-CH); 6.51 (1H, s, H Ar); 7.24 (2H, d, J = 8.1, H Ar); 7.34 (2H, d, J = 8.0, H Ar); 8.55 (1H, s, H Ar). ¹³C NMR spectrum, δ, ppm: 21.1; 26.6; 27.0; 44.2; 48.7; 79.9; 80.5; 105.4; 115.8; 115.8, 126.2; 128.3; 129.5; 135.2; 138.9; 165.3; 166.6; 190.0; 190.4. Mass spectrum, m/z: 337 [M+H]⁺. Found, %: C 74.93; H 5.96. C₂₁H₂₀O₄. Calculated, %: C 74.98; H 5.99.

8-(4-Methoxyphenyl)-2,2-dimethyl-2,3,7,8-tetrahydro-4 H ,6 H -pyrano[3,2- g ]chromene-4,6-dione (5f). IR spectrum, ν, cm^–1: 2979, 1690 (C=O), 1602, 1459, 1230, 1145. ¹H NMR spectrum, δ, ppm (J, Hz): 1.46 (3H, s, CH₃); 1.48 (3H, s, CH₃); 2.71 (2H, s, CH₂); 2.88 (1H, dd, J = 16.9, J = 2.9) and 3.08 (1H, dd, J = 16.9, J = 12.8); 3.25 (3H, s, OCH3); 5.53 (1H, dd, J = 12.7, J = 2.6, 8-CH); 6.51 (1H, s, H Ar); 7.24 (2H, d, J = 8.1, H Ar); 7.35 (2H, d, J = 8.0, H Ar); 8.57 (1H, s, H Ar). ¹³C NMR spectrum, δ, ppm: 26.6; 27.0; 44.2; 48.8; 53.8; 79.8; 80.4; 105.4; 115.4; 115.7; 126.5; 128.1; 129.4; 135.2; 139.0; 165.1; 166.8; 189.9; 190.7. Mass spectrum, m/z: 353 [M+H]⁺. Found, %: C 71.52; H 5.69. C₂₁H₂₀O₅. Calculated, %: C 71.58; H 5.72.

8-(4-Isopropylphenyl)-2,2-dimethyl-2,3,7,8-tetrahydro-4 H ,6 H -pyrano[3,2- g ]chromene-4,6-dione (5g). IR spectrum, ν, cm^–1: 2964, 1690 (C=O), 1601, 1465, 1224, 1152. ¹H NMR spectrum, δ, ppm (J, Hz): 1.27 (6H, d, J = 6.3, (CH ₃)₂CH); 1.46 (3H, s, CH₃); 1.48 (3H, s, CH₃); 2.72 (2H, s, CH₂); 2.88 (1H, dd, J = 16.9, J = 2.9) and 3.08 (1H, dd, J = 16.9, J = 12.9, CH₂); 2.95 (1H, sept, J = 6.3, (CH₃)₂CH); 5.47 (1H, dd, J = 12.9, J = 2.8, 8-CH); 6.51 (1H, s, H Ar); 7.30 (2H, d, J = 8.2, H Ar); 7.38 (2H, d, J = 8.1, H Ar); 8.56 (1H, s, H Ar). ¹³C NMR spectrum, δ, ppm: 23.8; 26.6; 27.0; 33.9; 44.2; 48.7; 79.9; 80.5; 105.4; 115.1, 115.8, 126.3; 126.9; 128.4; 135.4; 149.9; 165.3; 166.7; 190.1; 190.4. Mass spectrum, m/z: 365 [M+H]⁺. Found, %: C 75.77; H 6.60. C₂₃H₂₄O₄. Calculated, %: C 75.80; H 6.64.

Synthesis of aurones 6a–h (General method). A mixture of chalcone 4a–h (10 mmol), mercuric acetate (35 mg, 11.0 mmol), and pyridine (2 ml) was taken up in a quartz tube which was inserted into a screw-capped Teflon vial and then subjected to microwave irradiation at 320 W for 2–4 min. After completion of the reaction (as indicated by TLC), the reaction mixture was poured into ice-cold water and extracted with dichloromethane (2×30 ml). The organic phase was dried over Na₂SO₄ and purified by column chromatography (eluent n-hexane–AcOEt, 9:1).

2-Benzylidene-7,7-dimethyl-6,7-dihydro-5 H -furo[3,2- g ]-chromene-3,5(2 H )-dione (6a). IR spectrum, ν, cm^–1: 2928, 1671 (C=O), 1605, 1326, 1242, 1138. ¹H NMR spectrum, δ, ppm (J, Hz): 1.44 (6H, s, 2CH₃); 2.71 (2H, s, CH₂); 6.73 (1H, s, H Ar); 6.78 (1H, s, CH); 7.32–7.41 (3H, m, H Ar); 7.81 (2H, d, J = 7.3, H Ar); 8.35 (1H, s, H Ar). 13C NMR spectrum, δ, ppm: 26.7; 48.5; 87.5; 102.8; 112.8; 113.9; 115.0; 127.9; 128.5; 128.6; 130.0; 132.3; 146.7; 168.3; 171.2; 182.7; 190.9. Mass spectrum, m/z: 321 [M+H]⁺. Found, %: C 74.90; H 5.00. C₂₀H₁₆O₄. Calculated, %: C 74.99; H 5.03.

2-(4-Fluorobenzylidene)-7,7-dimethyl-6,7-dihydro-5 H -furo[3,2- g ]chromene-3,5(2 H )-dione (6b). IR spectrum, ν, cm^–1: 2935, 1698 (C=O), 1604, 1330, 1240, 1150. ¹H NMR spectrum, δ, ppm (J, Hz): 1.45 (6H, s, 2CH₃); 2.72 (2H, s, CH₂); 6.72 (1H, s, H Ar); 6.79 (1H, s, CH); 7.29 (2H, d, J = 8.5, H Ar); 7.54 (2H, d, J = 8.5, H Ar); 8.36 (1H, s, H Ar). ¹³C NMR spectrum, δ, ppm: 26.8; 48.5; 81.2; 101.2; 111.6; 115.9; 116.0; 116.3; 117.4; 125.4; 128.2; 133.4; 146.9; 162.2; 164.7; 166.6; 169.6; 182.5; 190.4. Mass spectrum, m/z: 339 [M+H]⁺. Found, %: C 70.95; H 4.40. C₂₀H₁₅FO₄. Calculated, %: C 71.00; H 4.47.

2-(4-Chlorobenzylidene)-7,7-dimethyl-6,7-dihydro-5 H -furo[3,2- g ]chromene-3,5(2 H )-dione (6c). IR spectrum, ν, cm^–1: 2925, 1699 (C=O), 1605, 1328, 1238, 1137. ¹H NMR spectrum, δ, ppm (J, Hz): 1.45 (6H, s, 2CH₃); 2.72 (2H, s, CH₂); 6.72 (1H, s) and 6.73 (1H, s, CH, H Ar); 7.35 (2H, d, J = 8.5, H Ar); 7.74 (2H, d, J = 8.5, H Ar); 8.35 (1H, s, H Ar). ¹³C NMR spectrum, δ, ppm: 26.7; 48.3; 87.3; 102.3; 111.6; 113.2; 115.7; 128.7; 129.0; 130.0; 130.4; 133.5; 146.9; 168.1; 170.2; 182.6; 190.9. Mass spectrum, m/z: 355 [M+H]⁺. Found, %: C 67.67; H 4.20. C₂₀H₁₅ClO₄. Calculated, %: C 67.71; H 4.26.

2-(4-Bromobenzylidene)-7,7-dimethyl-6,7-dihydro-5 H -furo[3,2- g ]chromene-3,5(2 H )-dione (6d). IR spectrum, ν, cm^–1: 2964, 1700 (C=O), 1607, 1250, 1016. ¹H NMR spectrum, δ, ppm (J, Hz): 1.45 (6H, s, 2CH₃); 2.72 (2H, s, CH₂); 6.70 (1H, s, H Ar); 6.72 (1H, s, CH); 7.49–7.53 (2H, m, H Ar); 7.64–7.67 (2H, m, H Ar); 8.35 (1H, s, H Ar). ¹³C NMR spectrum, δ, ppm: 26.7; 48.3; 87.3; 102.8; 111.6; 113.6; 115.0; 122.3; 128.7; 130.0; 131.5; 146.9; 168.3; 171.2; 182.8; 191.2. Mass spectrum, m/z: 399 [M(79Br)+H]⁺. Found, %: C 60.10; H 3.72. C₂₀H₁₅BrO₄. Calculated, %: C 60.17; H 3.79.

7,7-Dimethyl-2-(4-methylbenzylidene)-6,7-dihydro-5 H -furo[3,2- g ]chromene-3,5(2 H )-dione (6e). IR spectrum, ν, cm^–1: 2968, 1705, 1601, 1254, 1011. ¹H NMR spectrum, δ, ppm (J, Hz): 1.52 (6H, s, 2CH₃); 2.41 (3H, s, CH₃); 2.78 (2H, s, CH₂); 6.80 (1H, s, H Ar); 6.85 (1H, s, CH); 7.27–7.28 (2H, m, H Ar); 7.78 (2H, d, J = 7.8, H Ar); 8.42 (1H, s, H Ar). ¹³C NMR spectrum, δ, ppm: 21.3; 26.7; 48.3; 87.3; 102.8; 112.4; 113.9; 115.0; 128.5; 128.9; 129.3; 130.0; 137.6; 146.9; 168.2; 171.2; 182.6; 191.2. Mass spectrum, m/z: 335 [M+H]⁺. Found, %: C 75.40; H 5.40. C₂₁H₁₈O₄. Calculated, %: C 75.43; H 5.43.

2-(4-Methoxybenzylidene)-7,7-dimethyl-6,7-dihydro-5 H -furo[3,2- g ]chromene-3,5(2 H )-dione (6f). IR spectrum, ν, cm–1: 2969, 1701 (C=O), 1607, 1253, 1020. ¹H NMR spectrum, δ, ppm ( J, Hz): 1.52 (6H, s, 2CH₃); 2.78 (2H, s, CH₂); 3.88 (3H, s, OCH₃), 6.78 (1H, s, H Ar); 6.84 (1H, s, CH); 6.97 (2H, d, J = 8.6, H Ar); 7.84 (2H, d, J = 8.6, H Ar); 8.41 (1H, s, H Ar). ¹³C NMR spectrum, δ, ppm: 26.7; 48.3; 55.8; 87.3; 102.1; 112.4; 113.8; 114.2; 115.0; 124.6; 131.3; 132.8; 146.9; 159.8; 168.5; 171.2; 182.6; 190.7. Mass spectrum, m/z: 351 [M+H]⁺. Found, %: C 71.92; H 5.13. C₂₁H₁₈O₅. Calculated, %: C 71.99; H 5.18.

2-(4-Isopropylbenzylidene)-7,7-dimethyl-6,7-dihydro-5 H -furo[3,2- g ]chromene-3,5(2 H )-dione (6g). IR spectrum, ν, cm^–1: 2962, 1702 (C=O), 1606, 1353, 1138. ¹H NMR spectrum, δ, ppm (J, Hz): 1.28 (6H, d, J = 6.8, (CH ₃)₂CH); 1.52 (6H, s, 2CH₃); 2.78 (2H, s, CH₂); 2.96 (1H, sept, J = 6.8, (CH₃)₂CH); 6.79 (1H, s, H Ar); 6.85 (1H, s, CH); 7.32 (2H, d, J = 7.9, H Ar); 7.82 (2H, d, J = 7.9, H Ar); 8.42 (1H, s, H Ar). 13C NMR spectrum, δ, ppm: 23.3; 26.7; 33.2; 48.3; 86.5; 102.6; 112.3; 113.9; 115.0; 126.0; 128.3; 129.5; 130.4; 147.0; 147.6; 168.4; 171.5; 181.0; 190.8. Mass spectrum, m/z: 363 [M+H]⁺. Found, %: C 76.18; H 6.09. C₂₃H₂₂O₄. Calculated, %: C 76.22; H 6.12.

7,7-Dimethyl-2-(naphthalen-1-ylmethylidene)-6,7-dihydro-5 H -furo[3,2- g ]chromene-3,5(2 H )-dione (6h). IR spectrum, ν, cm^–1: 2986, 1709 (C=O), 1610, 1386, 1025. ¹H NMR spectrum, δ, ppm (J, Hz): 1.44 (6H, s, 2CH₃); 2.71 (2H, s, CH₂); 6.73 (1H, s, CH); 7.51–7.53 (3H, m, H Ar); 7.60 (1H, s, H Ar); 7.83 (2H, t, J = 9.1, H Ar); 8.21 (1H, d, J = 8.4, H Ar); 8.30 (1H, d, J = 7.3, H Ar); 8.38 (1H, s, H Ar). ¹³C NMR spectrum, δ, ppm: 26.7; 48.3; 87.4; 102.6; 112.8; 112.9; 113.9; 115.0; 122.9; 124.0; 126.0; 126.3; 126.9; 128.8; 130.0; 132.0; 133.5; 135.6; 147.2; 169.5; 171.2; 186.5; 191.8. Mass spectrum, m/z: 371 [M+H]⁺. Found, %: C 77.80; H 4.85. C₂₄H₁₈O₄. Calculated, %: C 77.82; H 4.90.

Biological assay. For the antibacterial activity test, the cultures were grown in nutrient agar media and subcultured for log phase cultures in a liquid nutrient broth medium for zone of inhibition test and further subcultured onto media in Petri plates for the experimental purposes. The broth cultures were diluted with sterilized saline to bring the final size of inoculum to 10⁵–10⁶ CFU/ml. The compounds were diluted in acetone, DMSO, and diethyl ether for biological assays. Among the three solvents, diethyl ether was taken as the best one. The bacterial cultures were placed on the media and incubated at 37°C for 24–48 h along with the diluted compounds introduced through discs dipped and placed over the nutrient media. The bacterial growth inhibition on the media was expressed as zone of inhibition in mm.

For the antifungal activity test, the synthesized compounds were dissolved in diethyl ether (10 ml) before mixing with Potato Dextrose Agar medium (PDA, 90 ml). The final concentration of compounds in the medium was maintained to be 500 μg/ml. The cultures of fungi were incubated in PDA at 25±1°C for 48–72 h to get long mycelium for antifungal assay. The mycelia disc of approximately 0.45 cm in diameter was cut from the PDA medium with a sterilized inoculation needle and inoculated in the center of PDA plate. The inoculated plates were incubated at 27±1°C for 3 days. Diethyl ether in sterilized distilled water was taken as blank control, while hymexazol was used as positive control. The growth of the fungal colonies was measured on the third day and the data were statistically analyzed.

The Supplementary information file containing ¹H NMR spectra of compounds 5a–e,g, 6a,c–h and ¹³C NMR spectra of compounds 5a,b is available at http://springerlink.bibliotecabuap.elogim.com/journal/10593.

The authors are thankful to the Head of Department of Chemistry, Osmania University and JNTU, Hyderabad, India, and the Managing Director, Richmond Vivek Laboratories, Hyderabad, India, for providing laboratory facilities to carry out the research work. We also are thankful to the Director of Central Facilities for Research and Development, Osmania University for providing spectral analysis facilities.