Synthesis, Characterization and Antimicrobial Evaluation of Ru(II) and Co(III) Complexes of Phenylene-1,2-bis( iminoflavone) Derivatives

Ashok Kumar Singh1*, Suresh Kumar Patel2 and Asif Jafri3

1 & 2 Department of chemistry, University of Lucknow, Lucknow - 226007 , INDIA

3 Department of Zoology, University of Lucknow, Lucknow - 226007, INDIA

* Correspondence: E-mail: skp0505@gmail.com

(Received 03 Feb, 2018; Accepted 23 Mar, 2019; Published 28 Mar, 2019 )

ABSTRACT: Bioactive ruthenium and cobalt scaffolds of molecular formula [Ru(PPh3)2L1-4] and [Co(H 2O)Cl L1-4] {where L1-4 represents the tetradentate ligands (E)-4-((2-((E)-(3-hydroxy-2-(4-substitutedphenyl)-4a,8a-dihydro-4H-chromen-4-ylidene)amino)phenyl)imino)-2-(4-substituted phenyl)-4H-chromen-3-ol (phenylenebis(iminoflavone)} have been synthesized from Ru(II) precursor [Ru(PPh3)4 Cl2] and CoCl2.3H2O and phenylenebis(iminoflavone). Tetradentate ligands of flavone (phenylenebis(iminoflavone)) have been derived by the condensation of 3-hydroxy-2-(4-substitutedphenyl)-4H-chromen-4-one with o-phenylenediamine. These ligands and scaffolds were characterized by elemental analysis, NMR, IR, ESI-MS, UV-Visible spectrometry, and conductometric measurement. These ligands and scaffolds were screened the in vitro toxicity for antimicrobial activity against the growth of bacterial species Staphylococcus aureus, Bacillus subtilis, Escherichia coli, Salmonella typhi and fungal species Candida albicans and Aspergillus flavus.

Keywords: Tetradentate ligands; Ru(II) and Co(III) scaffolds; spectrometry; conductometric and magnetic moment; antimicrobial activity.

INTRODUCTION: Flavonoids, a ubiquitous heterocyclic naturally occurring compounds are found in fruit, vegetables, nuts, seeds, stems, flowers, tea, wine, Propolis and honey.1 & 2 From earlier these compounds have been used as medicine to treat human diseases as they possess wide range of biological activity such as antioxidant3-5, antifungal6, antibacterial7, antiviral activity 8. Increase in resistance power of Microbes towards antimicrobial agents has become a great problem of world as microbes have adapted to new environments more than people. Microbes are changing new properties to resist drug treatments that were once effecting at destroying them and generate new type of disease as jeeka virus disease.9 Thus survival of human being against microorganism with ingenious tactics has challenged. Therefore, it is more difficult to treat the pathogenic viruses, bacteria, fungi, and protozoa with the existing drugs. In order to remove resistance against antimicrobial drugs and make them more effective other mode of action of drugs must be developed.9 It has shown that metal complexes of flavonoids show wide range of biological activities. 10 & 11 Flavonoids itself chelates with metals such as Fe, Cu, and Zn and show enhance activity in vivo and provide a new route to design new metal drugs active towards various diseases related to biological role of metals.12 Flavone complexescis-dichlorobis(3-aminoflavone)platinum,13cis-dichloro bis(3-imino-2-R-O-flavanone) ruthenium(II)(R= CH 3 or CH2CH3),14 cis-dichloro(3-nitrosoflavone)(3-hydroxyiminoflavan-one)ruthenium(II) 15 and dichloro(pcym)(6- or 7-aminoflavone) ruthenium(II), (pcym is ?6-p-cymene)16 exhibit cytotoxic activity. It has been reported that heteroatom containing flavone has antimicrobial activity. Certain flavones show antifungal activity against plant pathogen which explored against fungal pathogens causing infection in human.17 Synthetic biflavones, amentoflavone, isocryptomerin, ginkgetin, bilobetin had shown good antifungal activity.18-20 Natural products containing imine group play important role towards its biological activity.21 Metal complexes containing ligands having imine group exhibit stupendous chemical and biological significance. 22 Along with ruthenium certain Co(III) complexes also showed antimicrobial activity.21

Here in this paper we have synthesized the phenylenebis(iminoflavone) ligands of 3-hydroxy-2-(4-substitutedphenyl)-4H-chromen-4-one and their ruthenium(II) and cobalt(III) complexes in order to enhance the antimicrobial activity. The synthesized phenylenebis(iminoflavone) ligands and their ruthenium complexes were characterized and screened for antibacterial activity against Staphylococcus aureus, Bacillus subtilis, Escherichia coli, Salmonella typhi and antifungal activity against Aspergillus flavus,Candida albicans by using Ciprofloxacin and Fluconazole as a standard drug respectively.

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Scheme 1: Synthesis of ligands and their Ru(II) and Co(III) complexes.

MATERIAL AND METHODS: All chemicals used were purchased from sigma Aldrich. The solvents used were purified by reported method.24 Double distilled water has been used wherever necessary. [Ru(PPh3)4Cl2] was synthesized from RuCl3.H 2O by reported procedure.25 The purity of the compound was monitored by TLC (CHCl3/CH3OH, 9:1), using silica gel plate (Merck). Melting points were determined with SSU melting point apparatus. Euro Vector E 3000 Elemental Analyzer was used for elemental analysis. UV visible spectra were recorded on a double beam UV -Vis near IR Labtronics LT-2900 instrument. IR spectra (KBr discs) were recorded on Agilent Cary 360 FTIR spectrometer. 1H NMR, 13C NMR and 31P NMR spectra were recorded on Brucker Advance 400 MHz FT NMR spectrometer. ESI mass spectrum was recorded on Waters UPLC-TQD Mass spectrometer.

Antimicrobial evaluation: The organisms were used in this study are Staphylococcus aureus, Bacillus subtilis, Escherichia coli, Salmonella typhi (bacteria), Candida albicans and Aspergillus flavus (fungi). All the synthesized ligands (L1-L4) and complexes 4I (1-8) has been screened for their in vitro inhibitory activity (antibacterial and antifungal activities) against strains of microorganisms for determination of minimum inhibitory concentrations (MICs) in µg/mL by micro dilution method.26 Ciprofloxacin and Fluconazole are used as standard drugs.

Synthesis of ligands-phenylenebis(iminoflavone): The phenylenebis(iminoflavone) ligands were synthesized by reported procedure with some minor modifications.27 To the 10 mmol solution of a 3-hydroxy-4-substituted flavone derivative (Sub.= 0.268g (-OCH3), 0.254g (-OH), 0.364g (-Cl), 0.281g (-N(CH3) 2) in methanol, o- Phenylenediamine (0.541g, 5 mmol) dissolved in methanol (20 mL) and few drops of glacial acetic acid were added. The reaction mixture was refluxed for 7 hours at 70-80 C. The resulting solution was cooled to room temperature, and then poured into ice with constant stirring. The precipitate thus obtained was filtered and washed with 10% ethanol and diethyl ether. The crude products were purified by column chromatography on silica gel. Petroleum ether/ethyl acetate (95/5%) used as eluent. The ligand was recrystallized with hot ethanol and dried in vacuo over anhydrous CaCl2.

Synthesis of Ru(II) complexes (4I-1 to 4I-4) : The Ru(II) complexes were prepared by reported procedure with minor modification.28 To the 0.10 mmol (0.0994 g) solution of [RuCl 2(PPh3)4] in 20 mL benzene, 0.1 mmol of ligand (L1-0.610 g, L2-0.582 g, L3-0.619 g, L4-0.637 g) added. The mixture was refluxed for 5h in round bottom flask and then volume was reduced up to 3 mL. The product was separated by the addition of small quantity of petroleum ether at 60 -80 . The complex was then filtered, washed and recrystallized from dichloromethane and petroleum ether mixture in 1:3 volume ratio respectively.

Synthesis of Co(III) complexes (4I-5 to 4I-8) : Further, the synthesized phenylenebis(iminoflavone) ligands were interacted with the CoCl2.6H2O. The Co(III) complexes were synthesized by reported procedure with slight modification.29 To the warm 1 mmol methanolic solution (20mL) of ligand (L1=0.610 g, L2=0.588 g, L3=0.619 g, L4=0.636 g,), 1 mmol (0.238 g) methanolic solution (15 mL) of CoCl2.6H 2O was added and refluxed for 3 hour by adding 1 mmol of sodium acetate. On cooling the reaction mixture at room temperature, the precipitate was formed, which was filtered, washed with 10% ethanol and ether and dried in vacuum over anhydrous CaCl2. The complexes were recrystallized in hot ethanol. During the synthesis performed under aerobic conditions, the colour changes from violet red to dark brown which suggest the occurrence of oxidation process induced by the oxygen in air (CoII?CoIII).

(E)-4-((2-((E)-(3-hydroxy-2-(4-methoxyphenyl)-4a, 8a-dihydro-4H-chromen-4-ylidene)amino)phenyl) imino)-2-(4-methoxyphenyl)-4H-chromen-3-ol (L1):

Yield: 78% (0.281g), Colour: Brown, m.p.: 147°C, IR (KBr, cm-1): 3410(OH), 2812(CH), 1609(C=N), 1582(C=C), 1260(C-O), 1198(C-N). 1H NMR(400MHz, DMSO-d6): d(ppm) 3.47(6H, s), 6.79(4H, d, J=8.28Hz), 6.69-6.84(8H, m), 7.02-7.19(4H, m,), 7.75(4H, d, J = 8.68 Hz), 9.56(2H, s), 13C NMR (100MHz, DMSO-d6, d (ppm): 55.45, 55.45, 113.74, 113.74, 114.43, 114.43, 114.43, 114.43, 118.67, 118.67, 120.52, 120.52, 124.80, 124.80 124.24, 124.24, 128.07, 128.07, 129.27, 129.27, 131.34, 131.34, 131.71, 131.71, 131.71, 131.71, 136.22, 136.22, 154.36, 154.36, 155.67, 155.67, 155.81, 155.81, 159.58, 159.58, 160.40, 160.40, Elemental analysis (%) Calc.: C, 74.99; H, 4.64; N, 4.60; Observed: C, 75.02; H, 4.67; N, 4.59; ESI-MS: : 609.62 (Observed), 608.64 (Calculated). M. F.: C38H 28N2O6, UV-vis.: ?max in nm (in DMSO, 2×10-4M), 256, ( - *transition), 360(n- * transition).

(E)-4-((2-((E)-(3-hydroxy-2-(4-hydroxyphenyl)-4a, 8a-dihydro-4H-chromen-4-ylidene)amino)phenyl) imino)-2-(4-hydroxyphenyl)-4H-chromen-3-ol (L2):

Yield: 62% (0.22g), Colour: Reddish brown, m.p.: 210°C, IR (KBr, cm -1): 3523(OH), 2906(CH), 1606(C=N), 1573(C=C), 1256(C-O), 1208(C-N), 1H NMR (400MHz, DMSO-d6): d(ppm) 6.98(4H, d, J = 8.3, Hz), 7.06(4H, d, J = 8.3, Hz), 7.28(4H, m), 7.19-7.29(8H, m), 9.11(2H, s), 9.43(2H, s), 13C NMR (100MHz, DMSO-d6): d (ppm): 113.74, 113.74, 115.74, 115.74, 115.74, 115.74, 118.67, 118.67, 120.50, 120.50, 124.24, 124.24, 124.80, 124.80, 128, 128, 129.27, 129.27, 131.32, 131.32, 131.41, 131.41, 131.41, 131.41, 136.24, 136.24, 154.36, 154.36, 155.65, 155.65, 155.82, 155.82, 157.83, 157.83, 159.58, 159.58, Elemental analysis (%) Calc.: C, 74.22; H, 4.50; N, 4.81; Observed: C, 74.26; H, 4.52; N, 4.79; ESI-MS: : 583.58(Observed), 582.60(Calculated). M. F. C36H26N 2O6, UV-vis. ?max in nm (in DMSO, 2×10-4M), 271, ( - * transitions), 410 (n- * transition).

(E)-2-(4-chlorophenyl)-4-((2-((E)-(2-(4-chlorophe nyl)-3-hydroxy-4a,8a-dihydro-4H-chromen-4-yli dene)amino)phenyl)imino)-4H-chromen-3-ol (L3):

Yield: 68% (0.25g), Colour: Brownish black, m.p.: 121°C, IR (KBr, cm -1): 3569(OH), 2869(CH), 1618(C=N), 1564(C=C), 1286(C-O), 1180(C-N), 723(C-Cl). 1H NMR (400MHz, DMSO-d6): d (ppm): 7.19(4H, d, J = 8.4Hz), 7.14(4H, d, J = 8.4Hz), 7.00-7.10(8H, m), 7.29(4H, m), 9.62(2H, s), 13C (100MHz, DMSO-d6) NMR: d(ppm): 113.72, 113.72, 118.66, 118.66, 120.52, 120.52, 124.24, 124.24, 124.80, 124.80, 126.50, 126.50, 126.50, 126.50, 128, 128, 128.94, 128.94, 128.94, 128.94, 129.25, 129.25, 131.34, 131.34, 135.66, 135.66, 136.22, 136.22, 154.35, 154.35, 155.66, 155.66, 155.80, 155.80, 159.62, 159.62; Elemental analysis (%) Calc.: C, 69.80; H, 3.90; Cl, 11.45; N, 4.52; Observed: C, 69.82; H, 3.91; Cl, 11.44; N, 4.51; ESI-MS: : 620.48 (Observed), 619.49 (Calculated), M. F.: C36H24Cl2N2O4, UV-vis. ? max in nm (in DMSO, 2×10-4M), 262( - * transitions), 385(n- * transition).

(E)-2-(4-(dimethylamino)phenyl)-4-((2-((E)-(2-(4-(dimethylamino)phenyl)-3-hydroxy-4a,8a-dihydro-4H-chromen-4-ylidene)amino)phenyl)imino)-4H-chromen-3-ol (L4):

Yield: 73% (0.27g), Colour: Brown, m.p.: 135°C, IR (KBr, cm-1): 3462(OH), 2879(CH), 1607(C=N), 1576(C=C), 1278(C-O), 1214(C-N), 1H NMR (400MHz, DMSO-d6): d(ppm): 2.88(12H, s), 6.93(4H, d, J = 7.5 Hz), 6.99-7.17(8H, m), 7.24(4H, m), 7.19-7.26(4H, m), 9.50(2H, s), 13C NMR (100MHz, DMSO-d6 ): d (ppm): 40.30, 40.30, 40.30, 40.30, 113.74, 113.74, 113.82, 113.82 ,113.82, 113.82, 118.66, 118.66,120.50, 120.50, 124.23, 124.23, 126.4, 126.40, 126.40, 126.40, 128, 128, 124.80, 124.80, 129.26, 129.26, 136.23, 136.23, 131.34, 131.34, 151.42, 151.42, 154.36, 154.36, 155.64, 155.64, 155.80, 155.80, 159.60, 159.60, Elemental analysis (%) Calc.: C, 75.45; H, 5.70; N, 8.80; Observed: C, 75.54; H, 5.74; N, 8.83; ESI-MS: : 637.74 (Observed), 636.74 (Calculated), M. F. C40H36N 4O4 UV-vis. ?max in nm (in DMSO, 2×10-4M), 252( - * transitions), 356(n- * transition).

Ruthenium(II) complexes of L1 ligands: (4I-1):

Yield 71%, Colour: Brown; m. p.: 248oC, IR (KBr, ?cm-1): 2833(CH), 1527(C=N), 1544(C=C), 1296(C-O), 1232(C-N), 1435(Ru-P), 1162(P-Ph), 556(Ru-O), 492(Ru -N), 1H-NMR (DMSO-d6, 400 MHz, 25oC, TMS): d (ppm): 3.47(6H, s), 7.17(PPh 3, m), 7.00-9.80(m, Ar-H), 31P NMR (DMSO-d6, 400 MHz, 25oC, TMS): d(ppm): 49.82(PPh3, s), Elemental analysis (%) Calc.: C, 72.13; H, 4.58; N, 2.27; P, 5.03; Observed: C, 72.43; H, 4.66; N, 2.26; P, 5.00; Ru, 8.21; For ESI-MS: : 1233.24(Observed), 1232.26 (Calculated). M. F.: C74H56N2O6P2Ru, UV-vis. ? max (nm) (in DMSO, 2×10-4M) 254( - * transitions), 485(n- * transition).

Ruthenium(II) complexes of L2 ligands: (4I-2):

Yield: 65%, Colour: Blackish brown; m. p.: 282oC, IR (KBr, ?cm -1): 2837(CH), 1538(C=N), 1629(C=C), 1302(C-O), 1248(C-N), 1418(Ru-P), 1178(P-Ph), 550(Ru-O), 490(Ru-N), 1H-NMR (DMSO-d6, 400 MHz, 25oC, TMS): d (ppm): 7.15(PPh 3, m), 9.11(2H, s), 6.86-8.96(Ar-H, m), 31P NMR (DMSO-d6, 400 MHz, 25oC, TMS), d (ppm): 49.90(PPh3, s), Elemental analysis (%) Calc.: C, 71.81; H, 4.35; N, 2.33; P, 5.14; Ru, 8.39, Observed: C, 71.86; H, 4.38; N, 2.31; P, 5.12; Ru, 8.40; For ESI-MS: 1205.20(Observed), 1204.21(Calculated), M. F.: C 72H52N2O6P2Ru, UV-vis.: ?max in nm (in DMSO, 2×10-4M), 262( - * transitions), 505(n- * transition).

Ruthenium(II) complexes of L3 ligands: (4I-3):

Yield: 66%, Colour: Brown; m. p.: 228oC, IR (KBr, ?cm -1): 3486 (OH), 3033(CH), 1542(C=N), 1624(C=C), 1314(C-O), 1228(C-N), 1408(Ru-P), 1182(P-Ph), 548(Ru-O), 488(Ru-N), 1H-NMR (DMSO-d6, 400 MHz, 25oC, TMS), d (ppm): 7.19(PPh3, m), 7.46-9.23(m, Ar-H), 31P NMR (DMSO-d6, 400 MHz, 25oC, TMS), d (ppm): 49.86(PPh 3, s), Elemental analysis (%) Calc.: C, 69.68; H, 4.06; Cl, 5.71; N, 2.26; P, 4.99; Ru, 8.14; Observed: C, 69.70; H, 4.16; Cl, 5.70; N, 2.25; P, 4.97; Ru, 8.15; For ESI-MS: 1242.08(Observed), 1241.10 (Calculated). M. F.: C72H50Cl2N2O4P2Ru, UV-vis.: ?max in nm (in DMSO, 2×10-4M), 264( - * transitions), 507(n- * transition).

Ruthenium(II) complexes of L4 ligands: (4I-4):

Yield: 68%, Colour: Brown, m. p.: 232oC, IR (KBr, ?cm -1): 3021(CH), 1530(C=N), 1654(C=C), 1296(C-O), 1254(C-N), 1426(Ru-P), 1170(P-Ph), 558(Ru-O), 502(Ru-N), 1H-NMR (DMSO- d6, 400 MHz, 25oC, TMS), d (ppm): 2.88(6H, s), 3.96(12H, s), 7.14(PPh3, m), 7.16-9.86(m, Ar-H), 31P NMR (DMSO-d6, 400 MHz, 25oC, TMS), d (ppm): 49.02(PPh3, s), Elemental analysis (%) Calc.: C, 72.54; H, 4.97; N, 4.45; P, 4.92; Ru, 8.03; Observed: C, 72.58; H, 4.99; N, 4.46; P, 4.90; Ru, 8.04; For ESI-MS: : 1259.34(Observed), 1258.35(Calculated), M. F.: C 76H62N4O4P2Ru, UV-vis.: ?max in nm (in DMSO, 2×10-4M),266( - * transitions), 482(n- * transition).

Cobalt(III) complexes of L1 ligands: (4I-5):

Yield: 65%, Colour: Yellow brown; m. p.: 262oC, IR (KBr, ?cm -1): 3545(OH, broad), 798(OH), 2890(CH), 1526(C=N), 1564(C=C), 1293(C-S), 1297(C-O), 348(Co-Cl), 456(Co-N), 524(Co-O), 1H-NMR (DMSO-d6, 400 MHz, 25oC, TMS): d (ppm). 3.95(3H, s), 6.96-8.76(m, Ar-H); Elemental analysis (%) Calc.: C, 63.48; H, 3.93; Cl, 4.93; Co, 8.20; N, 3.90; Observed: C, 63.50; H, 3.97; Cl, 4.92; Co, 8.21; N, 3.89; For ESI-MS: : 720.00(Observed), 719.02(Calculated), M. F.: C38H28ClCoN2O7, UV-vis.: ?max in nm (in DMSO, 2×10-4M), 265( - * transition), 385(n- * transition), 505(n- * transition).

Cobalt(III) complexes of L2 ligands: (4I-6):

Yield: 64%, Colour: Reddish Brown; m. p. >300oC. IR (KBr, ?cm -1): 3547(OH), 777(OH), 2990(CH), 1526(C=N), 1614(C=C), 1281(C-S), 1307(C-O), 354(Co-Cl), 454(Co-N), 545Co-O), 1H-NMR (DMSO-d6, 400 MHz, 25oC, TMS), d (ppm): 9.78(s, OH), 6.96-8.93(m, Ar-H), Elemental analysis (%) Calc.: C, 62.58; H, 3.50; Cl, 5.13; Co, 8.53; N, 4.05; Observed: C, 62.61; H, 3.55; Cl, 5.15; Co, 8.54; N, 4.04; For ESI-MS: : 691.95(Observed), 690.97(Calculated) .M. F.: C36H24ClCoN2O7, UV-vis .: ?max in nm (in DMSO, 2×10-4M), 269 ( - * transitions), 392(n- * transition), 546(n- * transition).

Cobalt(III) complexes of L3 ligands: (4I-7):

Yield: 62%, Colour: Blackish brown, m. p. >300oC. IR (KBr, ?cm-1): 3507(OH, broad), 796(OH), 2870(CH), 1539(C=N), 1627(C=C), 1274(C-S), 1316 (C-O), 363(Co-Cl), 450(Co-N), 525(Co-O),1H-NMR (DMSO-d6, 400 MHz, 25oC, TMS), d (ppm): 6.72-8.82(m, Ar-H), Elemental analysis (%) Calc.: C, 59.40; H, 3.05; Cl, 14.61; Co, 8.10; N, 3.85; Observed: C, 59.42; H, 3.12; Cl, 14.60; Co, 8.12; N, 3.84; For ESI-MS: : 728.85(Observed), 727.86(Calculated), M. F.: C36H22Cl3CoN2O5, UV-vis.: ?max in nm (in DMSO, 2×10-4M), 277( - * transitions), 396(n- * transition), 555(d-d transition).

Cobalt(III) complexes of L4 ligands: (4I-8):

Yield: 68%, Colour: Yellow brown; m. p. >360oC, IR (KBr, ?cm -1): 3545(OH, broad), 786(OH), 2890(CH), 1529(C=N), 1564(C=C), 1302(C-S), 1297(C-O), 334(Co-Cl), 460(Co-N), 510Co-O), 1H-NMR (DMSO-d6, 400 MHz, 25oC, TMS), d (ppm): 3.95(3H, s), 6.96-8.76(m, Ar-H), Elemental analysis (%) Calc.: C, 64.48; H, 4.60; Cl, 4.76; Co, 7.91; N, 7.52; Observed: C, 64.51; H, 4.64; Cl, 4.75; Co, 7.92; N, 7.50; For ESI-MS: : 746.10(Observed), 745.11(Calculated), M. F.: C40H34ClCoN4O5, UV-vis.: ?max in nm (in DMSO, 2×10-4M), 260( - * transition), 385(n- * transition), 505(n- * transition).

RESULTS AND DISCUSSION:

Spectroscopic studies:

FT-IR Spectra: The vibrational frequency of phenylenebis(iminoflavone) was compared with those of complexes in order to infer the coordination mode. The frequencies of ligands were observed at 3410-3580 cm-1, 1605-1625cm-1, 1250-1300cm-1, and 1175-1225cm-1 for(O-H) phenolic, (C=N), (C-O) phenolic, and (C-N) respectively. In all the complexes, the C=N band is shifted to lower frequency, 1565-1580 cm-1, indicated that the azomethine N-atom of phenylenebis(iminoflavone) had coordinated to the metal ion.31 Further, in all complexes, the C-O stretching vibration appears at higher frequency 1280-1320cm-1 inferred, coordination occurred through the phenolic O-atom. The disappearance of band due to (O-H) in the complexes occurred during co-ordination shows de-protonation of phenolic protons. However, the new bands appeared in the metal complexes in the region 545-560 cm-1, 488-504 cm-1 , 510-525 cm-1 and 450-460 cm-1 are attributed to (Ru-O), (Ru-N), (Co-O) and (Co-N) respectively.32 & 33 In complexes 4I(1-4) at 1430-1440 cm-1, 1165-1182 cm-1 and 1085-1090 cm-1 appeared due to (Ru-P) and (P-Ph), respectively.34 On the basis of vibrational bands it is inferred that the ligands are behaving as a dibasic tetradentate ligands. The broad band in the region 3200-3500 cm -1 and two weaker bands in the region 750-800 cm-1 due to (O-H) rocking and wagging mode of vibration respectively, indicated the presence of coordinated water molecule in complexes (4I-5 to 4I-8).36

1 HNMR Spectra: The 1HNMR spectra of ligands and complexes were recorded in DMSO –d6 and the chemical shift d (ppm) is referenced to internal TMS. The protons of phenylenbis(iminoflavones) appeared in the region d (ppm) d 3.20-3.80, d 2.2-2.9, 6.90-9.98 (broad), and d 8.90-9.83 of methoxy (-OCH3, L 1, 4I-1, 4I-5), N-methyl (NCH3, L4, 4I-4, 4I-8), aromatic (Ar-H) and phenolic (OH), protons respectively. The downfield shift of aromatic protons in the complexes from d 6.90-9.98 to d 6.25-8.63, shows metal ion coordinated to the ligand. Furthermore, the disappearance of the phenolic proton in the complexes shows that the de-protonation occurred during complexation of phenolic O-atom with metal.

31 P NMR Spectra: The 31P NMR spectra of complexes (4I-1 to 4I-4) shows singlet at 25.12, 24.46, 25.04 and 24.36 ppm confirm the presence of triphenylphosphine group with magnetically equivalent phosphorous atom.

Electronic Spectra: The electronic spectral data of ligands and complexes were recorded in 2×10 -4M solution of DMSO. Bands of electronic spectra of Ru(II) complexes showed 2-3 bands in region 256-520nm. The complexes are diamagnetic, which showed ruthenium in (II) oxidation state and t62g configuration in octahedral environment. The ground state is 1A1g arising from the t62g configuration in an octahedral environment. The excited states corresponding to thet2g5, eg1 configuration are 3 T1g, 3T2g, 1T1g and1T2g. The possible transitions are 1A1g ?3T1g, 1A1g ?1T2g, 1A1g ? 1T1g and 1A1g ? 1T2g .in order of increasing energy. The bands in the region 460-520 nm are due 1A1g ? 1T1g transition36 on the basis of low extinction coefficient and other high intensity band in the region 250-270 nm due to charge transfer transition arising from the excitation of electrons from the metal t2g level to the unfilled molecular orbitals derived from the level of legands.37

Electronic spectra of Co(III) complexes (4I-5 to 4I-8) exhibited three absorption bands in the region 250-280 nm, 380-400 nm and 505-560 nm due to - *, n- * and d-d transitions. The absorption bands of ligands due to - * and n- * transitions shifted at higher wavelength in complexes and a new band observed in region 505-560 nm due tod-d transitions. The d-d bands originate from the 1A 1g?1T1g transition for the distorted octahedral Co(III) ion in complexes.38 & 40

Conductometric and magnetic moment Measurement: The molar conductivity of complexes was measured in DMF (dimethylformamide) by digital TDS-Conductivity meter. All complexes of Ru(II) and Co(III) showed molar conductivity in the range 2.6-5.6 O-1 cm 2 mol-1 which shown all complexes are non electrolyte.39 Also the magnetic susceptibility measurement shown all complexes are of diamagnetic (µ = 0.0 BM)

Biological Evaluation: The antimicrobial activities of the synthesized compounds have been screened in vitro, as growth inhibiting agents. The antifungal and antibacterial screening were carried out using micro dilution Method against some strains of bacteria and fungi like S. aureus, B. subtilis,( Gram positive bacteria) E. coli, S. typhi (Gram negative bacteria) and A. flavus, and C. albicans respectively. The MIC values of synthesized compounds were obtained compared with standard drug ciprofloxacin and fluconazole. The MIC values of the ligand and its complexes are given in Tables 1, 2, 3 and 4, respectively. The growth inhibitory activity of ligands and their complexes are shown in figure 1, 2, 3 and 4 respectively. The antimicrobial evaluation demonstrated that the complexes 4I-4 and 4I-8 showed good activity against different microorganism in comparison to free ligand. This gradual enhancing antimicrobial activity of the metal complexes, compared with that of Schiff bases, is conceivably owing to modification in structure due to coordination, and chelating tends to build metal complexes act as more influential antimicrobial agents, thus inhibiting the development of the microorganisms. The Overtone’s principle and chelation theory illustrates the enhanced antimicrobial effect as the chelation have a tendency to make the ligand a more powerful and potent antimicrobial agent. On chelation, the polarity of the metal ion will be reduced to a better range due to the overlap of the ligand orbital and partial sharing of the positive charge of the metal ion with donor groups. It is responsible for enhancing delocalization of p-electrons over the whole chelate ring and increases the penetration of the complexes into lipid membranes blocking of the metal binding sites in the enzymes of pathogens. In this way we can say that, metal complexes are more effective than the ligands because metal complexes may act as a vehicle for activation of ligands as a principle of cytotoxic species.

Table 1: In vitro antibacterial activity of (E)-2-(4-(substituted)phenyl)-4-((2-substitutedphenyl) imino)-4H-chromen-3-ol.

Ligands

Minimum inhibitory concentration

S. aureus

B. subtilis

E. coli

S. typhi

L1

>200

>100

50

25

L2

>100

>200

>200

>200

L3

100

50

>100

50

L4

6.5

75

7.5

6.5

DMSO

00

00

00

00

Ciprofloxacin

12.5

12.5

12.5

12.5

img2

Figure 1: Bardiagram of antibacterial evaluation of ligands L1 to L4.

Table 2: In vitro antibacterial activity of Ru(II) and Co(III) complexes of (E)-4-((2-((E)-(3-hydroxy-2-(4-substitutedphenyl)-4a,8a-dihydro-4H-chromen-4-ylidene)amino)phenyl)imino)-2-(4-substitutedphenyl)-4H-chromen-3-ol.

Complexes

Minimum inhibitory concentration

S. aureus

B. subtilis

E. coli

S. typhi

4I-1

>200

>100

75

50

4I-2

>100

>200

>200

>200

4I-3

100

50

>100

75

4I-4

10.5

100

9.5

8.5

4I-5

100

75

50

50

4I-6

75

>100

100

>200

4I-7

25

50

75

100

4I-8

8.5

9.5

100

10.0

DMSO

00

00

00

00

Ciprofloxacin

12.5

12.5

12.5

12.5

img2

Figure 2: Bardiagram of antibacterial evaluation of Ru(II) and Co(III) complexex 41(1-8).

Table 3: In vitro antifungal activity of (E)-2-(4-(substituted)phenyl)-4-((2-substituted phenyl)imino)-4H-chromen-3-ol.

Complexes

Minimum inhibitory concentration

A. flavus

C. albicans

L1

100

>100

L2

50

25

L3

>100

75

L4

3.75

75

DMSO

00

00

Fluconazole

12.5

12.5

img2

Figure 3: Bardiagram of antifungal evaluation of ligands L1 to L4.

Table 4: In vitro antifungal activity of Ru(II) and Co(III) complexes of (E)-4-((2-((E)-(3-hydroxy-2-(4-substitutedphenyl)-4a,8a-dihydro-4H-chromen-4-ylidene)amino)phenyl)imino)-2-(4-substituted phenyl)-4H-chromen-3-ol.

Complexes

Minimum inhibitory concentration

A. flavus

C. albicans

4I-1

>200

>100

4I-2

75

50

4I-3

>100

25

4I-4

6.75

100

4I-5

100

>100

4I-6

50

75

4I-7

25

>200

4I-8

8.5

7.75

DMSO

00

00

Fluconazole

12.5

12.5

img2

Figure 4: Bardiagram of Ru(II) and Co(III) complexes 41(1-8).

CONCLUSION: Tetradentate phenylenebis(iminoflavone) ligands 2F(1-4) and their Ru(II) and Co (III) complexes 4I(1-8) were synthesized and characterized by using physiochemical and spectroscopic techniques and these suggest the low spin octahedral geometry of Ru(II) and Co(III) complexes. The antimicrobial activities of complexes and its ligands were evaluated against some Gram-positive and some Gram-negative bacteria as well as fungi. Among Ru(II) and Co (III) complexes 4I-1and 4I-8 shows good antibacterial and antifungal activities. The antimicrobial activities of complexes exhibited better antimicrobial properties and showed enhanced inhibitory activities as compared to the ligands.

ACKNOWLEDGEMENT : The authors are sincerely thankful to the Head, Department of Chemistry, University of Lucknow, Lucknow, for providing necessary laboratory facilities and Director, SAIF, CDRI, for providing spectral data and department of Zoology, University of Lucknow for biological data of the compounds.

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