Research Journal of Chemical Sciences ________________________________________ ISSN 2231-606X Vol. 1(5), 80-84, Aug. (2011) Res.J.Chem.Sci. International Science Congress Association 80 Further Biologically Active Derivatives of 1, 3-DiketonesGeorge Mulongo, Jolocam Mbabazi*, P. Nnamuyomba and G.B. MpangoDepartment of Chemistry, Gulu University, P.O. Box 166, Gulu, UGANDA Department of Chemistry, Makerere University, P.O. Box 7062, Kampala, UGANDA Available online at: www.isca.in (Received 20th July 2011, revised 26th July 2011, accepted 1st Augest 2011)Abstract This study presents the synthesis and characterisation of new compounds of antimicrobial activity by the coupling of aromatic aldehydes with 5,5-dimethylcyclohexan-1,3-dione (dimedone). The products were refluxed with N–benzyl-N-phenylhydrazine in acetic acid. With the help of micro- and IR-spectral analysis, the molecular structures of the synthesised products were determined. These were ascertained using H NMR at 60MHz and TMS as internal standard. Biological activity of the derivatives against gram-positive Cocci and Bacilli as well as gram-negative Bacilli was tested and found to vary widely from inactive to highly active, which could prove to be of practical pharmaceutical application. Key words:Aromatic aldehydes, dimedone, coupling derivatives, Antimicrobial activity.IntroductionPursuant upon an earlier communication in which we reported some new biologically active compounds from 1,3-diketones using aromatic amines, the present paper reports on further derivatives with aromatic aldehydes. These compounds have been found to exhibit similar biological activity as their aromatic amine counterparts. The chemistry2-6 and ready availability of cyclohexane-1,3-diones7-11 render it a suitable starting material12,13 for the synthesis of organic compounds such as oxozolidinones with known antibacterial activity14-16, and phenylbutazone which is quite effective in the treatment of the pain associated with rheumatoid arthritis and Tietze’s syndrome18-20 . The aim of the present study was to create new derivatives of 1, 3-diketones using aromatic aldehydes and -benzyl--phenylhydrazine, consequent upon which their biologically active properties would be investigated and established. Material and Methods The chemicals and solvents used in this study were obtained from Merck, Fluka and Sigma (Aldrich). They were of reagent grade and required little further purification. An open-tube capillary method was employed to determine melting points in which are quoted uncorrected. Thin layer chromatography (TLC) was of invaluable help in controlling the purity of the compounds. A Mattson 5000 FTIR spectrophotometer (USA) was used to record the IR spectra in KBr pellets. Also, a Perkin-Elmer Instrument (200B, USA) was used to estimate the C, H, N, Cl constitutional data of the derivatives. The H NMR spectra of the compounds were measured in CDCland DMSO-d solution on a DRX – 300MHz spectrometer (Bruker, UK) with TMS as internal standard. Synthesis of 2-arylidene-1, 3-diones (2): Requisite amounts of aromatic aldehyde (0.002 mol) and 5, 5-dimethylcyclohexan-1, 3-dione, dimedone (0.28 g, 0.002 mol) were fused at 150C for 30 minutes (see Scheme) in a dry pear-shaped 50 ml flask in an oil-bath. The residue was cooled and shaken with benzene. The resulting solid product was filtered off, re-crystallised and the melting point taken. + C O O H3C H3C R2 R1 R3 R4 R5 OHC fuse, 150 H3C R1 R2 R3 R4 R5 O O C H3C H3C R1 R2 R3 R4 R5 N O N(CH)C C6H5 Scheme-1An illustrated reaction pathway for the synthesis of antimicrobial products using dimedone (1), aromatic aldehydes and N-benzyl-N-phenyl hydrazine 3 i) R=R=R=R5=H, R=N (CH (2a) (3a) ii) R=R=R=R=H, R=OH (2b) (3b) iii) R1==R=R=H, R=Cl (2c) (3c) iv) R=R=R=H, R=NO, R=Cl (2d) (3d) v) R=R=R=H, R=Cl, R=NO (2e) (3e) Synthesis of 2-arylidene-1, 3-diketone derivatives (3): An appropriate quantity of 2- arylidenecyclohexan-1, 3- Research Journal of Chemical Sciences _______________________________________________________ ISSN 2231-606X Vol. 1(5), 80-84, Aug. (2011) Res.J.Chem.Sci. International Science Congress Association 81 dione (see Scheme, 0.002 mol) was dissolved in a mixture of 50% acetic acid (25 ml) and -benzyl-phenylhydrazine (0.002 mol, 0.396 g, 0.4 ml), refluxed for 3 to 4 hours, left to cool and the products re-crystallised. The melting points and percentage yields were determined. Biological activity tests21-22: The required amounts of liquid agar media were poured into sterile petri-dishes to a depth of 3 to 4 mm. After solidifying, the liquid media test organism was spread over the solidified agar media and incubated in the petri-dish at 37C for 24 hours to facilitate the growth of the micro-organisms. With the help of a sterile rod, a hole was made on the medium and poured on the known (1000 µg/ml concentration) test solution in that hole. The biological activity of the derivatives was evaluated by determining the average diameter of the inhibition zone (figure-1). Figure-1 IR spectrum for Compound 3a (see Scheme) Figure-2 Confirmatory H nmr spectrum for Compound 3a Research Journal of Chemical Sciences _______________________________________________________ ISSN 2231-606X Vol. 1(5), 80-84, Aug. (2011) Res.J.Chem.Sci. International Science Congress Association 82 Results and DiscussionCompound (2a) was obtained as orange crystals in 70% yield, m.p. 197C, C1721NO (RMM, 267.35); IR, max in cm-1: 3039, 1520 (CH-aromatic), 2960-2633, 1448, 1369 (CH, -N(CH)), 811(=CH). Compound (2b) was obtained as yellow crystals in 62% yield, m.p. 246.0C, C1516(RMM, 244.28). Compound (2c) was obtained as yellow crystals in 52.3% yield, m.p. 185.0C, C1515 (RMM, 257.27). Compound (2d) was obtained as yellow crystals in 66.0% yield, m.p. 218.0C, C1516 (RMM, 273.28). Compound (2e) was obtained as light yellow crystals in 64.0% yield, m.p. 207.0C, C1516 (RMM, 228.28); IR, max in cm-1: 3058 (=CH, aromatic), 2961-2568, 1449, 1372 (CH), 776, 693 (=CH). Derivatives of 2-arylidene-1,3-diketones were obtained in fairly quantitative yields. The products (3a – 3e) were isolated, re-crystallised (from mixtures of methanol and water) and afforded in 40-60% yields. Compound (3a), figure 1, was obtained as yellow crystals in 40.0% yield, m.p. 170.0C; IR, max in cm-1: 3050, 1601, 1493 (aromatic), 2953, 2851, 1447, 1343 (CH), 692, 816 (=CH). Anal. calcd for C3033O (RMM, 451.58): C, 79.73; H, 7.37; N, 9.31; Found: C, 79.65; H, 7.45; N, 9.11%. H NMR spectra, figure 2, at 60 MHz (units ppm) showed: 1.083 and 0.984 (s, 3H, CH), 8.14 (s, 5H, =CHar), 7.236 (s, 4H, =CHar), 7.414 (s, 2H, =CHar), 4.33 (s, 2H, CH), 7.447 (s,2H, =CHar), 7.416 (s, 2H, =CH), 3.0 (s, 3H, CH), 6.859 (s, 2H, =CHar), 6.934 (s, 2H, =CHar), 6.876 (s, H, =CHar). Compound (3b) was obtained as red crystals in 65.0% yield, m.p. 217.0C; IR, max in cm-1: 3400-3208 (OH), 3068, 1599, 1490 (aromatic), 2959, 2876, 1456, 1366 (CH), 1635 (�C=O). Anal. calcd. for 2828O (RMM, 408.52): C, 82.17; H, 6.91; N, 6.86; Found: C, 82.41; H, 6.77; N, 6.67%. H NMR spectra at 60MHz ( units ppm) showed: 1.103 and 1.024 (s, 3H, CH), 7.984 (s, 5H, =CHar), 7.152 (s, 4H, =CHar), 7.295 (s, 2H, =CHar), 4.329 (s, 2H, CH), 7.507 (s,2H, =CHar), 7.392 (s, 2H, =CH), 3.214 (s, H, OH), 6.901 (s, 2H, =CHar), 6.788 (s, 2H, =CHar), 6.674 (s, H, =CHar). Compound (3c) was obtained as orange crystals in 66.5% yield, m.p. 238.0C; IR, max in cm-1: 2960, 2874, 1460, 1354 (CH), 1667 (�C=O), 1660, 1495 (aromatic). Anal. calcd. for 2827 (RMM, 453.52): C, 74.15; H, 6.00; N, 9.26; Found: C, 74.44; H, 6.12; N, 9.35%. Compound (3d) was obtained as light orange crystals in 40.0% yield, m.p. 172.0C; IR, max in cm-1: 2955, 2872, 1452, 1373 (CH), 1643 (�C=O), 753, 697 (=CH). Anal. calcd. for 2827 (RMM, 453.52): C, 74.15; H, 6.00; N, 9.27; Found: C, 73.81; H, 5.97; N, 9.10%. Compound (3e) was obtained as orange crystals in 60.0% yield, m.p. 202.0C; IR, max in cm-1: 3456-3247 (NH), 3063, 3030, 1626, 1495 (aromatic), 1662 (�C=O), 2957, 2872, 1465, 1362 (CH), 745, 697 (=CH). Anal. calcd. for C2828O (RMM, 394.508): C, 85.24; H, 7.15; N, 3.55; Found: C, 85.03; H, 6.94; N, 3.41%. H NMR spectra at 60MHz ( units ppm) showed: 1.066 and 0.974 (s, 3H, CH), 8.027 (s, 5H, =CHar), 7.035 (s, 4H, =CHar), 7.398 (s, 2H, =CHar), 4.242 (s, 2H, CH), 7.357 (s,2H, =CHar), 7.219 (s, 2H, =CH), 7.149 (s, 2H, =CHar), 7.240 (s, 2H, =CHar), 6.689 (s, H, =CHar). Table 1 Collective data showing the spectra of antimicrobial activity of the compounds (3a – 3e) at 100µg/ml concentration level against micro-organisms used Test strains of micro-organisms Test Compounds 3a 3b 3c 3d 3e A)Gram-Positive Cocci1.Staphylococcus aureus 2.Staphylococcus epidermis 3.Sarcina lutea B)Gram- Positive Bacilli1.Bacillus permal 2.Bacillus subtilis C)Gram-Negative Bacilli1.Aerobacterium klebsiella 2.Bacillus Arizona 3.Bacillus proteus 4.Bacillus pseudomonas 5.Escherichia coli 6.Salmonella paratyphi A 7.Salmonella paratyphi B 8.Salmonella paratyphi C 9.Shigella flexneri 10.Shigella sonnei - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - + + + + + + + + + + + + + + + ± ± + + ± ± + + + + ± + + ± + + + + + + + + + + + + + + + N.B. (+) no growth (High activity), (±) Moderate growth (moderate activity), (-) High growth (inactive)Biological Screening: The 2-arylidene-1,3-diketone derivatives () were examined in vitro against bacterial species which included gram-positive Cocci, gram-positive Bacilli and gram-negative Bacilli. The photographs provided (figure 3) are representative of the major observations made. Tables 1 and 2 show the spectral data of the synthesised biologically active compounds (3a – 3e) at 100 and 1000 g/ml concentrations, respectively, against the micro-organisms. Research Journal of Chemical Sciences _______________________________________________________ ISSN 2231-606X Vol. 1(5), 80-84, Aug. (2011) Res.J.Chem.Sci. International Science Congress Association 83 Table 2 Collective data showing the spectra of antimicrobial activity of the compounds (3a – 3e) at 1000µg/ml concentration level against micro-organisms used Test strains of micro-organisms Test Compounds 3a 3b 3c 3d 3e D)Gram-Positive Cocci4.Staphylococcus aureus 5.Staphylococcus epidermis 6.Sarcina lutea E)Gram- Positive Bacilli3.Bacillus permal 4.Bacillus subtilis F)Gram-Negative Bacilli11.Aerobacterium klebsiella 12.Bacillus Arizona 13.Bacillus proteus 14.Bacillus pseudomonas 15.Escherichia coli 16.Salmonella paratyphi A 17.Salmonella paratyphi B 18.Salmonella paratyphi C 19.Shigella flexneri 20.Shigella sonnei - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - + + + + + + + + + + + + + + + ± ± + + ± ± ± ± + + + + ± ± + + + + + + + + + + + + + + + N.B. (+) no growth (High activity), (±) Moderate growth (moderate activity), (-) High growth (inactive The results presented in table 1 suggested that compounds 3a and 3b were inactive (at 100 g /ml) against all the tested micro-organisms. This is attributed to the presence of the methylsubstituted amino (-N (CH) and hydroxyl (-OH) groups. The reactivity of the amino group might be severely hampered by the steric hindrance arising as a result of the bulkiness of the substituent at that position of the aromatic ring4,5 Compounds 3c and 3e exhibited high antimicrobial activity against all the test micro-organisms most probably due to the presence of the nitro (-NO) group that is dependent of the electronic richness of the nitrogen atoms4,5. Compound (3d) showed high antimicrobial activity against Sarcina lutea, Bacillus permal, Bacillus arizona, Bacillus proteus, Bacillus pseudomonas, Escherichia coli, Salmonella paratypi B and Salmonella paratypi C. It however exhibited moderate biological activity towards Staphylococcus aureus, Staphylococcus epidermis, Bacillus subtilis, Aerobacterium klebsiella, Salmonella paratypi A, Shigella flexneri and Shigella sonnei. When concentrations were increased to 1000 g/ml, there was little change in the antimicrobial activity of the compounds 3a, 3b, 3c and 3e (table 2) against all the micro-organisms under investigation. The antimicrobial activity of compound 3d was obstructed on a few micro-organisms such as Bacillus Permal, Bacillus arizona, Bacillus proteus and Salmonella paratypi C.Conclusion Dimedone has been shown to be an adaptable source of a number of biologically active compounds that might be of use in pharmaceutical research. AcknowledgementThe authors wish to express their gratitude to Department of Chemistry, Makerere University (Uganda) for laboratory facilities. Our sincere thanks also go to Dr A. Metwally, formerly at Makerere University and presently at Faculty of Science, Mansoura University (Egypt) for carrying out the elemental, IR and NMR analyses. References1.Mulongo George, Mbabazi Jolocam, Odongkara B., Twinomuhwezi H. and Mpango G.B., New biologically active compounds from 1,3-diketones, Res. J. Chem. Sci., , 102 (2011) 2.Dioxon K. and Greenhill J.V., Enaminones, J. Chem. Soc., Perkin, (II), 277 (1977) 3.Guan-Wu W. and Chun-Bao M., Environmentally Benign One-Pot Multi-Component Approaches to the Synthesis of Novel Unsymmetrical 4-Arylacridinediones, Green Chem.,, 1080 (2006) 4.James L.D., Richard A.G. and Surya K.De., An efficient one-pot synthesis of polyhydroquinoline derivatives through the Hantzsch four component condensation, J. Mol. Cat. A: Chemical, 256, 309 2006) 5.Majumdar K.C. and Samanta S.K., Aza- claisein rearrangement: Synthesis of dimedone-annaleted unusual heterocycles, Tetrahedron, 57, 4955 (2001) 6.Majumdar K.C. and Samanta S.K., sulfoxide- claisein rearrangement: Synthesis of dimedone-annaleted unusual heterocycles, Tetrahedron, 58, 22, 4551 2002) 7.French H. S. and Holden M. E. T., Absorption Spectra of Certain , -Unsaturated Ketones, including Benzal Compounds, J. Amer. Chem. Soc.,67, 1239 (1945) Research Journal of Chemical Sciences _______________________________________________________ ISSN 2231-606X Vol. 1(5), 80-84, Aug. (2011) Res.J.Chem.Sci. International Science Congress Association 84 8.Schwarzenbach G. and Wittwer C., Uber das Keto-Enol-Gleichgewicht bei cyclischen -Diketonen, Helv. Chim. Acta, 30, 663 (1947) 9.Conroy H., Picrotoxin. II, The Skeleton of Picrotoxinin The Total Synthesis of dl-Picrotoxadiene, J. Amer. Chem. Soc., 74, 3046 (1952) 10.Meek E.G., Turnbull J.H. and Wilson W., Alicyclic compound. Part II. The preparation of cyclohexane-1,3-diones and their enol ethers, J. Chem. Soc., 811 1953) 11.Shriner R.L. and Todd H.R., 1,3-Cyclohexadiene-5,5-dimethyl, Org. Synth., II, 200 (1943) 12.Frank R.L. and Hall H.K., Monocyclic Terpenes from Cyclic 1,3-Diketones, J. Chem. Soc., 72, 1645 (1950) 13.Pal B.C., Dehydration of -Phenylethylcyclohexanol-3, J. Amer. Chem. Soc., 11, 3397 (1955) 14.Cui Y., Dang Y., Yang Y. and Ji R., Synthesis of Novel Oxazolidinone Derivatives for Antibacterial Investigation, Current Sci., 83, 531 (2005) 15.Jae-Min H., Sung-Ho Y. and Kang-Yeoun J., Synthesis of Oxazolidinone Phosphonate Derivatives-Part II, Bull. Korean Chem. Soc.,28, 821 (2007) 16.Yeong W.J., Weon B.I., Jae K.R., Mi Ja Shim, Woo B. K. and Eung C. C., Synthesis and Antibacterial activity of oxazolidinones Containing Pyridine Substituted with Heteroaromatic Ring, Bioorg. Med. Chem., 12, 5909 (2004) 17.Ae N.P., Hye Y. K., Hyun J.J., Bo H.K., Yong S.C., Kyung II C., Jung H. C. and Hun Y.K., Synthesis and In Vitro Activity of New Oxazolidinone Antibacterial Agents having Substituted Isoxazoles, Bioorg. Med. Chem. Lett., , (18), 2679 (1999) 18.Dewse C.D. and Potter C.G., Inhibitory Effect of Phenylbutazone and Oxyphenylbutazone on DNA Synthesis in Normal Human Bone Marrow, J. Pharm. Pharmac. 27, 523 (1975) 19.Grevsten S. and Johansson H., Phenylbutazone in the Treatment of Acute Lumbago-Sciatica, Z. Rheumatol, 34, 444 (1975) 20.Erkki J.V., Phenylbutazone in the Treatment of Tietze’s syndrome, Ann. Rheum. Dis., 26, 133 (1967) 21.Chitra M., Shyamala D.C.S. and Sukumar E., Antibacterial Activity of Embelin, Filotropia, 74, 401 (2003)22.Manjudar S.H., Chakra G.S. and Kulkarni K.S., Medicinal Potential of Semecarpus anacardium Nut., J. Herb. Med. Toxicol. , 9 (2008)Figure-3