Corresponding Author:
M. A. Neelakantan
Chemistry Research Centre, National Engineering College, K. R. Nagar, Kovilpatti-628 503
E-mail: [email protected]
Date of Submission 4 August 2009
Date of Revision 1 December 2009
Date of Acceptance 20 February 2010
Indian J Pharm Sci, 2010, 72 (2): 216-222

Abstract

Some new mixed ligand complexes (1-5) of type ML'B (M(II)=Mn(II), Co(II), Ni(II), Cu(II) and Zn(II); HL'= o-vanillidene-2-aminobenzothiazole; B= 1,10-phenanthroline) and Schiff base metal complexes of types (ML 2 ") (6-10) and (M 2 L") (11-15) (HL"= o-vanillidene-2-amino-N-(2-pyridyl)-benzene sulfonamide) were synthesized and characterized by elemental analysis and spectral (IR, 1 H NMR and 13 C NMR) studies. The free ligands and their metal complexes have been screened for their in vitro biological activities against bacteria, fungi and yeast. The metal complexes show more potent activities compared with Schiff base ligands.

Keywords

Antibacterial activities, antifungal activities, mixed ligand complexes, o-vanillin, schiff base, spectral studies

Metal complexes of N and S chelating ligands have attracted considerable attention because of their interesting physicochemical properties and pronounced biological activities. The N and S atoms play a key role in the coordination of metals at the active sites of numerous metallobimolecules. o-vanillin is a natural aldehyde found in Andropogen nardus. It is used to treat bellyaches and also used in spicery[1]. Schiff bases containing o-vanillin possesses antifungal, antibacterial properties[2] and it acts as a weak inhibitor of tyrosinase, display both antimutagenic and comutagenic properties in Escherichia coli[3]. Heterocycles containing thiazole ring is present in a number of pharmacologically and biologically active compounds. Compounds containing benzothiazole and sulphonamide derivatives were used as antifungal[4,5], antiinfl ammatory[6], antiHIV[7], anticancer[8], anticarbonic anhydrase[9], diuretic, hypoglycaemic[10], antithyroid[11], antimalarial and in therapeutic fields. In view of the pronounced biological activities of these compounds, we report herein the synthesis and characterization of (i) mixed ligand complexes of Mn(II), Co(II), Ni(II), Cu(II) and Zn(II) derived from 1,10–phenanthroline and o-vanillidene-2-aminobenzothiazole (HL') and (ii) Schiff base metal complexes of o-vanillidene-2-amino-N-(2-pyridyl)-benzene sulfonamide (HL"). The biological screening of free ligands and their complexes against different bacteria, fungi and yeast are reported.

Materials and Methods

All the chemicals and solvents used were of AR grade (Merck, Mumbai, India) except o-vanillin (Fluka, Switzerland) and 2-amino-N-(2-pyridyl)- benzene sulfonamide (Sigma, USA). Melting points of all the compounds were determined in open glass capillaries and are uncorrected. The purity of the Schiff bases was ascertained by TLC on Silica gel–G plates and spots were visualized by using iodine vapours. The elemental analysis was performed using a Thermo Finnigan Flash EA 1112 CHNS analyzer at Central Electro Chemical Research Institute (CECRI), Karaikudi, India. The conductometric measurements of the complexes were carried out in DMSO solution using Systronics 611 conductivity bridge. Vibrational spectra were recorded using KBr pellets on FT-IR Shimadzu 8400S spectrophotometer, in the region 4000 – 400 cm-1 range. The 1H NMR and 13C NMR of Schiff base ligands and their diamagnetic zinc complexes in DMSO-d6 were recorded on Perkin Elmer R-32 spectrometer using Tetramethylsilane as internal standard at IIT, Chennai, India.

Biological evaluation

The newly synthesized ligands and their metal complexes were screened in vitro for their antibacterial activity against bacteria: Escherichia coli, Pseudomonas aeruginosa, Salmonella Typhi and Vibrio parahaemolyticus by well diffusion method[12] using agar nutrient. The antifungal activities were tested against fungus: Aspergillus Niger, Penicillium, Trichoderma virida and yeast: Saccharomyces cerevisiae by well diffusion method using potato dextrose agar as the medium. Ampicillin and nystatin are used as control for bacteria and fungi, respectively. The suspension of each microorganism was added to a sterile agar medium, then poured into sterile Petri plates and left to solidifi cation. The well was dug in the agar media using sterile metallic borer in each plate. The test solution (3×10-3 M) was prepared by dissolving the compounds in DMSO and the well was fi lled with the test solution using micropipette. The plates were incubated for 24 h in the case of bacteria and 72 h for fungi at 35°. The extracts were subjected to further assay with a series on time basis (24, 48 and 72 h). During this period, the test solution was diffused and affected the growth of the inoculated microorganisms. Activity was determined by measuring the diameter of the zone showing complete inhibition (mm). Growth of inhibition was compared with the control. The zone of inhibition is given as the average of three independent determinations.

Synthesis of mixed ligand complexes (1-5)

The Schiff base (HL') was synthesized[13] by the condensation of 20 ml of o-vanillin (0.030 g/10 mmol) with 2-amino benzothiazole (0.030 g/10 mmol) in ethanol (1:1 molar ratio). Ten millilitres of Schiff base (0.028 g/10 mmol) and 1,10-phenanthroline (0.020 g/10 mmol) in ethanol was added drop wise to 10 ml of 10 mmol metal salts in hot ethanol (0.020g of MnCl2.4H2O, 0.024g of CoCl2.6H2O, 0.025 g of Ni(CH3COO)2.4H2O, 0.020 g of Cu(CH3COO)2.H2O and 0.022 g of Zn(CH3COO)2. 2H2O). The reaction mixture was refluxed for 3 h on a water bath and the volume of the solution was reduced to half of its original volume. The solid compound (fig. 1) obtained was fi ltered off, washed with water, diethyl ether and dried in vacuum over CaCl2. The color, elemental analysis data, molar conductivity and melting point of Schiff base (HL') and its mixed ligand complexes (1-5) are given in Table 1.

Figure

Figure 1: Structure of the complexes.
(a) Mixed ligand complexes (1-5), (b) Schiff base complexes (6-10) and (c) Schiff base complexes (11-15).

Compound Color Mol. formulaa Mol. Wt. Yield (%) Λm (Ω-1 cm2 mol-1) Mp (0)
HL' Reddish yellow C15H12N2O2S 284.00 65 6.78 180
HL" Orange C19H17N3O4S 383.42 94 4.23 198
1 Pale yellow MnC27H21O3N4SCl 571.74 67 11.38 205
2 Green CoC27H21O3N4SCl 575.73 62 17.39 145
3 Light brown NiC29H24O5N4S 599.04 69 14.22 169
4 Dark brown CuC29H24O5N4S 603.90 67 9.42 202
5 Yellow ZnC29H24O5N4S 605.76 65 6.78 130
6 Brown MnC38H34O9N6S2Cl 896.43 54 5.50 210
7 Lignt Brown CoC38H34O9N6S2Cl 899.88 64 8.10 >300
8 Lignt Green NiC40H37O11N6S2 900.22 59 3.00 285
9 Black CuC40H37O11N6S2 904.93 63 5.13 290
10 Dark yellow ZnC40H37O11N6S2 906.86 57 3.40 220
11 Light brown Mn2C19H19O6N3SCl2 718.27 58 9.80 225
12 Brown Co2C19H19O6N3SCl2 725.17 51 13.90 >300
13 Dark Green Ni2C23H25O10N3S 725.85 68 12.90 293
14 Black Cu2C23H25O10N3S 663.28 83 10.80 320
15 Light yellow Zn2C23H25O10N3S 739.13 56 12.40 254

Table 1: Physio-chemical properties of schiff base and metal complexes

Compound 1, yield: 67%, mp: 205°, IR (KBr) cm-1: 1639 (C=N, azomethine), 1557 (C=N, thiazole ring), 1217 (C–O, phenolic), 746 (C–S–C, thiazole ring), 569 (Mn–N), 424 (Mn–O), 3396 and 856 (H2O molecule). Compound 2, yield: 62%, mp: 145°, IR (KBr) cm-1: 1618 (C=N, azomethine), 1558 (C=N, thiazole ring), 1215 (C–O, phenolic), 747 (C–S–C, thiazole ring), 572 (Co–N), 430 (Co–O), 3387, 848 (H2O molecule). Compound 3, yield: 69%, mp: 169°, IR (KBr) cm-1: 1624 (C=N, azomethine), 1558 (C=N, thiazole ring), 1207 (C–O, phenolic), 747 (C–S–C, thiazole ring), 567 (Ni–N), 420 (Ni–O), 3387, 850 (H2O molecule). Compound 4, yield: 67%, mp: 199o, IR (KBr) cm-1: 1606 (C=N, azomethine), 1560 (C=N, thiazole ring), 1217 (C–O, phenolic), 748 (C–S–C, thiazole ring), 540 (Cu–N), 422 (Cu–O), 3380 and 852 (H2O molecule). Compound 5, yield: 65%, mp: 130°, IR (KBr) cm-1: 1641 (C=N, azomethine), 1561 (C=N, thiazole ring), 1213 (C–O, phenolic), 748 (C–S–C, thiazole ring), 563 (Zn–N), 437 (Zn– O), 3321, 852 (H2O molecule), 1H NMR (TMS, DMSO-d6) d ppm: 7.67 (s, 1H, HC=N, azomethine), 6.80–7.40 (d, 4H, Ar–H), 6.90 (s, 1H, HC=N, thiazole ring), 3.70 (s, 3H, OCH3), 1.80 (s, 3H, OOCCH3).

Synthesis of Schiff base

The Schiff base (HL") was prepared by refl uxing a mixture of equimolar quantities of o-vanillin (10 ml, 0.300 g/10 mmol) and 2-amino-N-(2-pyridyl)-benzene sulfonamide (10 ml, 0.490 g/10 mmol, Scheme 1) in hot ethanol. After 8 h of refluxing, the reaction mixture was kept at room temperature overnight and the orange colored product was fi ltered, washed with distilled water, diethyl ether and recrystallized from the same solvent. Yield: 85%, mp: 198°, IR (KBr) cm-1: 1631 (C=N, azomethine), 1269 (C–O, phenolic), 1361, 1138 (SO2), 3246 (N–H, sulfonamide group), 1025 (–HC=N, pyridine ring nitrogen), 1H NMR (TMS, DMSO-d6) d ppm: 9.92 (s, 1H, HC=N, azomethine), 12.82 (s, –OH), 7.60–8.00 (d, 4H, Ar – H), 8.40–8.60 (s, pyridine –HC=N), 11.04 (s, NH), 3.48 (s, 3H, OCH3), 2.30 (s, 3H, CH3), 5.72 (s, 2H, SO2NH group), 13C NMR (TMS, DMSO-d6) d ppm: 164.66 (HC=N, azomethine), 155.29 (–C–OH, phenolic), 151.44 (HC=N, pyridine ring), 142.04–140.36 (C=C, pyridine ring), 148.51 (C–O–C), 121.69–117.96 (Ar, C=C), 56.24 (OCH3).

Figure

Scheme 1: Synthesis of Schiff base (HL' and HL") ligands.

Synthesis of metal complexes (6-15)

Ten millilitres of metal salts (10 mmol, 0.02 g of MnCl2.4H2O, 0.024 g of CoCl2.6H2O, 0.025 g of Ni(CH3COO)2.4H2O, 0.02 g of Cu(CH3COO)2. H2O and 0.022 g of Zn(CH3COO)2. 2H2O) in hot ethanol was added to 10 ml of the Schiff base ligand (0.038 g/10 mmol) in ethanol (1:1). 10 ml of metal salts (20 mmol, 0.04 g of MnCl2.4H2O, 0.048 g of CoCl2.6H2O, 0.05 g of Ni(CH3COO)2.4H2O, 0.04 g of Cu(CH3COO)2. H2O and 0.044 g of Zn(CH3COO)2. 2H2O) in hot ethanol was added to 10 ml of the Schiff base ligand (0.038 g/10 mmol) in ethanol (2:1). The mixture was refl uxed for 8 h on water bath. The volume of the solution was reduced to one-third of its original volume and left overnight. The precipitate was fi ltered off, washed with 50% (v/v) ethanol-water mixture, diethyl ether and dried over fused CaCl2 in a vacuum desiccator. The elemental analysis, molar conductivity value, color and melting point of Schiff base (HL") and its metal(II) complexes (6-15) are given in Table 1.

Compound 6, yield: 54%, mp: 210°, IR (KBr) cm-1: 1599 (C=N, azomethine), 1293 (C–O, phenolic), 1362, 1130 (SO2), 3248 (N–H, sulfonamide group), 1026 (–HC=N, pyridine ring nitrogen), 594 (Mn–N), 521 (Mn–O), 3398, 976, 774 (–OH, H2O molecule). Compound 7, yield: 64%, mp: >300°, IR (KBr) cm-1: 1602 (C=N, azomethine), 1272 (C–O, phenolic), 1364, 1132 (SO2), 3244 (N–H, sulfonamide group), 1030 (C=N, pyridine ring nitrogen), 604 (Co–N), 520 (Co–O), 3405, 976, 759 (–OH, H2O molecule). Compound 8, yield: 59%, mp: 285°, IR (KBr) cm-1: 1610 (C=N, azomethine), 1286 (C–O, phenolic), 1364, 1135 (SO2), 3246 (N–H, sulfonamide group), 1029 (C=N, pyridine ring nitrogen), 588 (Ni–N), 503 (Ni–O), 3398, 968, 767 (–OH, H2O molecule), 1616,1402 (–COO, acetate group). Compound 9, yield: 63%, mp: 290°, IR (KBr) cm-1: 1616 (C=N, azomethine), 1291 (C–O, phenolic), 1362, 1138 (SO2), 3248 (N–H, sulfonamide group), 1027 (C=N, pyridine ring nitrogen), 569 (Cu–N), 513 (Cu–O), 3415, 980, 775 (–OH, H2O molecule), 1626, 1412 (–COO, acetate group). Compound 10, yield: 57%, mp: 220°, IR (KBr) cm-1: 1598 (C=N, azomethine), 1278 (C–O, phenolic), 1363, 1136 (SO2), 3247 (N–H, sulfonamide group), 1026 (C=N, pyridine ring nitrogen), 596 (Zn–N), 507 (Zn–O), 3382, 962, 769 (–OH, H2O molecule), 1631,1408 (–COO, acetate group). 1H NMR (TMS, DMSO-d6) δ ppm: 10.12 (s, 1H, HC=N, azomethine), 7.65–7.97 (d, 4H, Ar–H), 8.38–8.61 (s, pyridine, –HC=N), 11.00 (s, NH), 3.46 (s, 3H, OCH3), 2.31 (s, 3H, CH3), 5.70 (s, 2H, SO2NH group), 4.73 (s, 2H, H2O molecule), 13C NMR (TMS, DMSO-d6) δ ppm: 169.42 (HC=N, azomethine), 150.94 (HC=N, pyridine ring), 141.98–140.52 (C=C pyridine ring C), 147.91 (C–O–C), 122.76–118.72 (Ar, C=C), 55.82 (OCH3).

Compound 11, yield: 58%, mp: 225°, IR (KBr) cm-1: 1615 (C=N, azomethine), 1279 (C–O, phenolic), 1006 (C=N, pyridine ring nitrogen), 589 (Mn–N), 518 (Mn–O), 3403, 987, 779 (–OH, H2O molecule). Compound 12, yield: 51%, mp: >300°, IR (KBr) cm-1: 1599 (C=N, azomethine), 1286 (C–O, phenolic), 1013 (C=N, pyridine ring nitrogen), 586 (Co–N), 516 (Co–O), 3394, 997, 765 (–OH, H2O molecule). Compound 13, yield: 68%, mp: 293°, IR (KBr) cm-1: 1605 (C=N, azomethine), 1298 (C–O, phenolic), 1350, 1009, 669 (C=N, pyridine ring nitrogen), 605 (Ni–N), 513 (Ni–O), 3379, 986, 776 (–OH, H2O molecule), 1631, 1405 (–COO, acetate group). Compound 14, yield: 83%, mp: 320°, IR (KBr) cm-1: 1620 (C=N, azomethine), 1287 (C–O, phenolic), 1012 (C=N, pyridine ring nitrogen), 605 (Cu–N), 517 (Cu–O), 3415, 980, 775 (–OH, H2O molecule), 1632, 1408 (– COO, acetate group). Compound 15, yield: 56%, mp: 254°, IR (KBr) cm-1: 1612 (C=N, azomethine), 1304 (C–O, phenolic), 1006 (C=N, pyridine ring nitrogen), 579 (Zn–N), 510 (Zn–O), 3410, 989, 787 (–OH, H2O molecule), 1623, 1408 (–COO, acetate group). 1H NMR (TMS, DMSO-d6) d ppm: 10.03 (s, 1H, HC=N, azomethine), 7.64–7.99 (d, 4H, Ar–H), 8.23–8.54 (s, pyridine, –HC=N), 3.40 (s, 3H, OCH3), 2.28 (s, 3H, CH3), 11.86 (s, 2H, SO2NH group), 4.67 (s, 2H, H2O molecule), 13C NMR (TMS, DMSO-d6) d ppm: 172.26 (–HC=N, azomethine), 164.49 (HC=N, pyridine ring), 140.19 – 138.25 (C=C pyridine ring C), 148.17 (C–O–C), 124.27 – 119.58 (Ar, C= C), 56.15 (OCH3).

Results and Discussion

The zone of inhibition of the complexes against the growth of bacteria and fungi were given in Table 2 and Table 3, respectively. A representative graph is given in fig 2. From the table it is evident that the complexes (1-5) and (11-15) have higher antibacterial activity than the complexes (6-10). The complexes (1-5) and (6-10) shows higher antifungal activity than the complexes (11-15). In general, the synthesized metal complexes have higher biological activities compared to the free ligands. The increased inhibition activity of the metal complexes can be explained on the basis of Tweedy’s chelation theory[14]. In metal complexes, on chelation the polarity of the metal ion will be reduced to a greater extent due to the overlap of the ligand orbital and partial sharing of the positive charge of the metal ion with donor groups. Further, it increases the delocalization of p- electrons over the whole chelate ring. The large ring size of 1,10-phenanthroline moiety makes the complexes more lipophillic[15]. This increased lipophillicity enhances the penetration of the metal complexes into lipid membranes and block the metal binding sites in the enzymes[16]. Metal complexes also disturb the respiration process of the cell and thus block the synthesis of proteins, which restricts further growth of the organisms. This enhancement in inhibiting the growth of bacteria and fungi can also be explained on the basis of their structure. The azomethine linkage and hetero aromatic moiety in the synthesized complexes exhibit extensive biological activities[17,18] due to increased liposolubility of the molecules in crossing cell membrane of the microorganism. The presence of electron donor group (-OCH3) in the complexes also plays a role in enhancing the inhibition activity. The antibacterial activity is found to be in the order; Control>(1-5)>(11-15)>(6- 10)>HL'>HL". The antifungal activity is found to be in the order; Control>(6-10)>(1-5)>(11-15)>HL'>HL".

Figure

Figure 2: Biological activities of (a) mixed ligand complex (b) Schiff base complexes.
(a) Mixed ligand complex (3) at 24, 48 and 72 h; (b) Schiff base complexes (HL", 6-10 and 11-15) at 24 h by well diffusion method (zone formation in mm); B-Salmonella typhi, C-Pseudomonas aeruginosa, D-Escherichia coli, E-Vibrio parahaemolyticus, F-Aspergillus niger, G–Penicillium, H–Trichoderma virida and I-Saccrharomycies species.

Compound Diameter of inhibition zone in mm for different microbial species
Salmonella typhi Pseudomonas aeruginosa Escherichia coli Vibrio parahaemolyticus
24 h 48 h 72 h 24 h 48 h 72 h 24 h 48 h 72 h 24 h 48 h 72 h
HL' 7 8 8 8 9 9 10 11 12 8 9 9
HL" 6 7 7 7 8 8 - - - 7 8 8
1 9 10 10 9 11 12 9 11 11 9 10 13
2 - - - 14 15 16 - - - 8 8 11
3 9 9 11 22 23 23 20 22 22 8 9 9
4 - 9 10 8 10 10 - - - 10 10 10
5 9 10 10 - - - 12 12 14 10 11 12
6 7 7 8 - - - - - - 6 7 9
7 7 7 8 - - - - - - 6 7 9
8 - - - - - - - - - 5 10 10
9 9 8 7 - - - - - - 7 8 9
10 - - - 6 7 7 - - - 12 13 14
11 6 8 8 - - - 6 7 8 8 9 9
12 - - - 8 9 9 9 9 10 - - -
13 15 15 16 6 7 7 10 12 13 10 10 11
14 - - - 7 8 9 - - - 8 8 9
15 14 15 17 11 12 12 11 14 14 9 11 12
Control 11 14 16 32 34 34 16 17 18 31 33 34

Table 2: Bacterial activities of the schiff base and metal complexes.

Compound Diameter of inhibition zone in mm for different microbial species
Aspergillusniger Penicillium Trichodermavirida Saccharomyces cerevisiae
24 h 48 h 72 h 24 h 48 h 72 h 24 h 48 h 72 h 24 h 48 h 72 h
HL' 5 6 6 5 6 6 8 8 9 8 9 9
HL" 6 6 6 - 6 8 6 7 7 4 5 6
1 - - - 9 9 10 13 14 14 12 12 13
2 9 9 10 9 10 11 - - - 8 8 9
3 - 9 9 8 8 9 9 9 9 10 10 11
4 10 11 11 9 10 10 12 14 14 9 10 10
5 - 10 10 - 9 10 12 12 13 10 11 12
6 10 12 13 10 11 12 8 9 9 8 11 12
7 12 14 16 15 16 16 8 8 8 6 7 8
8 10 12 12 - - - 4 4 8 6 8 10
9 7 9 13 - - - 4 6 6 - - -
10 7 8 16 13 15 16 6 10 10 6 8 8
11 8 8 10 6 8 8 4 4 4 - - -
12 - - - 7 8 8 - - - 6 6 6
13 9 12 12 10 10 11 4 4 4 - - -
14 8 8 9 - - - - - - 9 9 10
15 12 13 14 16 18 19 8 12 12 8 8 8
Control 14 15 16 15 18 18 20 22 23 25 28 28

Table 3: Fungal activities of the schiff base and metal complexes

Acknowledgements

The authors wish to thank the Management, Director and Principal of National Engineering College, Kovilpatti for their keen interest and constant encouragement throughout this investigation.

References