Biological and Spectral Studies of Newly Synthesized Triazole Schiff Bases and Their Si(IV), Sn(IV) Complexes

The Schiff bases HL1-3 have been prepared by the reaction of 5-bromothiophene-2-carboxaldehyde with 4-amino-5-mercapto-3-methyl/propyl/isopropyl-s-triazole, respectively. Organosilicon(IV) and organotin(IV) complexes of formulae (CH3)2MCl(L1-3), (CH3)2M(L1-3)2 were synthesized from the reaction of (CH3)2MCl2 and the Schiff bases in 1 : 1 and 1 : 2 molar ratio, where M = Si and Sn. The synthesized Schiff bases and their metal complexes have been characterized with the aid of various physicochemical techniques like elemental analyses, molar conductance, UV, IR, 1H, 13C, 29Si, and 119Sn NMR spectroscopy. Based on these studies, the trigonal bipyramidal and octahedral geometries have been proposed for these complexes. The ligands and their metal complexes have been screened in vitro against some bacteria and fungi.


Introduction
Recently, the research relating with metal complexes of heteronuclear Schiff bases has expanded enormously and now comprising their interesting aspects in coordination chemistry with a special emphasis in bioinorganic chemistry. A use of organosilicon and organotin compounds as reagents or intermediates in the inorganic synthesis has further strengthened their applications [1,2].
More-over, metal complexes of organosilicon(IV) and organotin(IV) halides with N, O, and S donor ligands have received much more consideration due to their industrial, environmental, and biological applications [3][4][5]. The N, O and S donor ligands have been used to enhance the biological activity of organosilicon and organotin derivatives [6]. Organosilicon(IV) complexes have been subjected of interest for their versatile applications in pharmaceutical and chemical industries. Organosilicon compounds of nitrogen and sulphur containing ligands are well known for their anticarcinogenic, antibacterial, antifungal, tuberculostatic, insecticidal, and acaricidal activities [7][8][9][10]. Generally, organosilicon complexes seem to owe their antitumor properties to the immune-defensive system of the organism [11]. Similarly, organotin compounds are the active components in a number of biocidal formulations in such diverse areas as fungicides, miticides, molluscicides, antifouling paints and surface disinfectants [12,13]. In addition, many organotin compounds have been tested for a large variety of tumor lines and found to be more effective than traditional heavy metal anticancer drugs [14,15]. Ahmad et al. have also screened some organotin compounds against tumor cells [16]. Prompted by these applications, few new organosilicon and organotin compounds have already been synthesized and screened for antibacterial and antifungal activities [17,18], and in continuation to this, in the present paper, the synthesis, characterization, and biological activities of new triazole Schiff bases and their organosilicon and organotin complexes have been carried out.

Analytical Methods and Physical Measurements.
Silicon and tin were determined gravimetrically as silicondioxide (SiO 2 ) and tindioxide (SnO 2 ). Melting points were determined on a capillary melting point apparatus. Molar conductance measurements of 10 −3 M solution of metal complexes in dry DMF were measured at room temperature (25 ± 1 • C) with a conductivity bridge type 305 Systronic model. Carbon, hydrogen, nitrogen and sulfur were estimated using elemental analyzer Heraeus Vario EL-III Carlo Erba 1108 at CDRI Lucknow. The electronic spectra of the ligands and their metal complexes were recorded in dry methanol, on a Systronics, Double-beam spectrophotometer 2203, in the range of 600-200 nm. The IR spectra of the ligands and metal complexes were recorded in nujol mulls/KBr pellets using BUCK scientific M5000 grating spectrophotometer in the range of 4000-350 cm −1 . Nuclear magnetic resonance spectra ( 1 H, 13 C) were recorded on BRUKER-300ACF and 29 Si and 119 Sn were recorded on BRUKER-400ACF spectrometer in DMSO-d 6 using tetramethylsilane (TMS) as an internal standard.

Synthesis of Metal Complexes.
To a weighed amount of dimethylsilicondichloride (Me 2 SiCl 2 ) and dimethyltindichloride (Me 2 SnCl 2 ) in ∼30 mL of dry methanol, was added the calculated amount of the sodium salt of the ligands in 1 : 1 and 1 : 2 molar ratios. The sodium salts of the ligands were prepared by dissolving the appropriate amount of the sodium metal and ligands in ∼30 mL dry methanol. The reaction mixture was refluxed for about 12 h and then allowed to cool at room temperature and removed the chlorine as sodium chloride. The excess of solvent was removed under reduced pressure by vacuum pump and the resulting solid was repeatedly washed with 5-10 mL dry cyclohexane and again dried under vacuum. The elemental analyses and physical properties of the complexes are reported in Table 1.

Results and Discussion
The reactions of Me 2 SiCl 2 and Me 2 SnCl 2 with the sodium salt of monobasic bidentate ligands in 1 : 1 and 1 : 2 molar      Table 2: IR-spectroscopic data (cm −1 ) of the ligands and their metal complexes.
Compound    showing that all 1 : 1 and 1 : 2 complexes are nonelectrolytic in nature Table 1.  respectively, and indicating the coordination of azomethine nitrogen atom to the metal atom [16]. In addition to this, the three medium intensity bands at 244 nm, 240 nm, and 260 nm due to π-π * transition in the ligands remain unchanged or show a minor change in the spectra of metal complexes [17].

IR Spectra.
In the IR spectra of the ligands, a broad band in the region of 3117-3094 cm −1 due to ν(N-H) [13] and a band at ∼1120 cm −1 due to ν(C=S) [21], is assigned to ν(C-S) and which indicates the complexation of ligands through S-atom with the metal atom. The metal sulphur bond formation is further supported by a band at ∼452 cm −1 and ∼426 cm −1 for ν(Si-S) [24] and ν(Sn-S) [25] in the spectra of organosilicon and organotin complexes, respectively. A sharp and strong band in the region of 1582-1597 cm −1 for ν(N=CH) [26] in case of ligands, was shifted to a higher wavelength number and appears in the region of 1628-1674 cm −1 in the spectra of metal complexes, indicating the coordination of ligands through azomethine nitrogen to the metal atom. The metal nitrogen bond was further supported by the presence of a band at about ∼535 cm −1 for ν(Sn-N) [27] and ∼575 cm −1 for ν (Si-N) [28]. A strong band in the region of 425-378 cm −1 was assigned to ν(M-Cl) [29]. The IR-spectral data of the ligands and their metal complexes are listed in Table 2. 3.3. 1 H NMR Spectra. The 1 H NMR spectra of the ligands show the -SH proton signal at δ 10.47 (s), δ 13.75 (s), and δ 11.10 (s) ppm for HL 1 , HL 2 , and HL 3 , respectively [26] (Figure 2). The disappearance of the signal due to -SH proton in the spectra of metal complexes indicates the deprotonation of the thiol group and supports the coordination of ligand through sulphur atom to the metal atom. A signal at δ 11.72 (s), 10.91 (s), and 10.63 (s) ppm was observed due to azomethine proton in the spectra of free ligands HL 1 , HL 2 and HL 3 , respectively, which moves upfield in the 1 H NMR spectra of metal complexes [13], indicates the bonding through the azomethine nitrogen atom to the central metal atom (Figure 3). The aromatic protons of the thiophene moiety in the ligands appear as two doublets, which remain more or less unchanged in the 1 Table 3.
The additional signals in the region δ 0.3-1.5 ppm are also observed in the spectra of complexes due to CH 3 -M group.
3.4. 13 C NMR Spectra. The 13 C NMR spectral data of ligands HL 1 , HL 2 , and HL 3 , and their corresponding 1 : 1 and 1 : 2 metal complexes [17,18] have been reported in Table 4. The signal due to the carbon atom attached to the azomethine group in the ligands HL 1 , HL 2 , and HL 3 appear at δ 166.42 ppm, δ 162.23 ppm, and δ 160.79 ppm, respectively. However, in the spectra of the corresponding metal complexes, the shift in the 13 C resonance indicate the coordination of nitrogen atom of azomethine group with the central atom in 1 : 1 and 1 : 2 metal complexes. Moreover, the shifting of the 13 C resonance of triazole which is attached to sulphur atom in the spectra of 1 : 1 and 1 : 2 metal complexes compared to the free ligands indicates the coordination through sulphur atom with the central metal atom. The new signal due to the methyl groups attached to the metal atom in the spectra of metal complexes has also been reported in Table 4.

29
Si and 119 Sn NMR Spectra. The value of δ 29 Si and δ 119 Sn indicates the coordination number of the central metal atom in the corresponding complexes [30], and generally (Figure 4), 29 Si and 119 Sn chemical shifts move to lower frequency with increasing coordination number of the metal atoms. The spectrum shows in each case only a sharp singlet indicating the formation of single species. 29 Si and 119 Sn NMR spectra of

Biological Activities
The bactericidal and fungicidal activities of the free ligands and their metal complexes against various gram positive and gram negative bacteria and fungi are reported in Tables 5, 6, and 7.  [31,32]. All the microbial cultures were adjusted to 0.5 McFarland standards, which is visually comparable to a microbial suspension of approximately 1.5 × 10 8 cfu/mL. 20 mL of Mueller Hinton Agar medium was poured into each petri plate and plates were swabbed with 100 μL inocula of the test microorganisms and kept for 15 min for adsorption. Using sterile cork borer of 8 mm diameter, wells were bored into the seeded agar plates, and these were loaded with a 100 μL volume with concentration of 4.0 mg/mL of each compound reconstituted in the DMSO. All the plates were incubated at 37 • C for 24 hrs. Antibacterial activity of each synthetic compound was evaluated by measuring the zone of growth inhibition against the test organisms with zone reader (Hi Antibiotic zone scale). DMSO was used as a negative control, whereas Ciprofloxacin was used as positive control. This procedure was performed in three replicate plates for each organism.   1 to the tube 10 and excess broth (100 μL) was discarded from the test tube no. 10. To each tube, 100 μL of standard inoculum (1.5 × 10 8 cfu/mL) was added. Ciprofloxacin was used as control. Turbidity was observed after incubating the inoculated tubes at 37 • C for 24 hrs.

4.3.
In Vitro Antifungal Activity. The ligands and their metal complexes were also screened for their antifungal activity against two fungi, namely, A. niger and A. flavus, the ear pathogens isolated from the patients of Kurukshetra [34], by poison food technique [35]. The moulds were grown on Sabouraud dextrose agar (SDA) at 25 • C for 7 days and used as inocula. The 15 mL of molten SDA (45 • C) was poisoned by the addition of 100 μL volume of each compound having concentration of 4.0 mg/mL reconstituted in the DMSO, poured into a sterile petri plate and allowed it to solidify at room temperature. The solidified poisoned agar plates were inoculated at the center with fungal plugs (8 mm diameter) obtained from the colony margins and incubated at 25 • C for 7 days. DMSO was used as the negative control whereas Fluconazole was used as the positive control. The experiments were performed in triplicates. Diameter of fungal colonies was measured and expressed as percent mycelial inhibition by applying the formula.
Percent inhibition of mycelial growth = dc − dt dc × 100 , (2) where dc is the average diameter of fungal colony in negative control sets and dt is the average diameter fungal colony in experimental sets.

Observations.
The antibacterial data reveals that the complexes are superior compared to the free ligands. The free ligands and their metal complexes are active against Grampositive bacteria (Staphylococcus aureus and Bacillus subtilis) and inactive against gram negative bacteria (Escherichia coli and Pseudomonas aeruginosa). Among the synthesized compounds tested compounds, Me 2 SnCl(L 1 ) and Me 2 SiCl(L 2 ) show more antibacterial activity that is, near to standard drug (Ciprofloxacin) ( Table 5). In the series, the MIC of the compounds ranged between 28-128 μg/mL against Grampositive bacteria. Compound Me 2 SnCl(L 1 ) and Me 2 SiCl(L 2 ) show highest MIC of 28 μg/mL against S. aureus ( Table 6). The antifungal activity of compounds ( Figure 5) shows more than 50% inhibition of mycelia growth against Aspergillus niger and A. flavus (Table 7). Thus, it can be postulated that further studies of these complexes in this direction could lead to more interesting results.

Conclusion
Trigonal bipyramidal and octahedral geometries have been proposed for 1 : 1 and 1 : 2 organosilicon(IV) and organotin(IV) complexes with the help of various physicochemical studies like IR, UV, 1 H, 13 C, 29 Si, and 119 Sn NMR ( Figure 6). The free ligands, and their metal complexes were screened against various fungi and bacteria to access their potential as antimicrobial agents. The antimicrobial data reveals that the complexes are superior to the free ligands and their toxicity has increased as per the increase in Bioinorganic Chemistry and Applications 9 concentration. These compounds were found more potent inhibitor of fungal growth as compared to the bacterial culture.