Carbenes are typically neutral compounds R₂C: derived from the parent methylene (H₂C:) which has a divalent carbon atom with just six valence electrons arising from four bonding electrons in the two R – C bonds and two nonbonding electrons remaining at the center of the carbene¹. NHCs is considered to be singlet carbenes, unlike Fisher and Schröckcarbenes, These are typically prepared by utilizing a strong base like NaOH or KOH for deprotonation of azolium salts². Carbene production is carried out by retaining electron pairs on the carbon atom by deprotonation. A σ orbital is occupied by a pair of electrons, P-orbital is still usable for receiving the electronic pair of neighboring nitrogen atoms. The lone pair of the carbene is given to metal for forming Metal−Carbon bond³. NHCshave been widely used in organometallic and inorganic chemistry due to their strong metal coordination properties.

There are various methods that can be used to synthesis NHC ligands and to attach substituents and functional groups to the ligands, and this area has been reviewed extensively.^4-6. The explanation for the biological effect of silver is that it interferes with the electronic transport mechanism of the cell and interacts with the membrane and the thiol group in the biological enzymes of the bacteria⁷. Young and his colleagues (In 2004) made a capsule of two silver-NHC complexes as an antimicrobial medication for bacteria, including Escherichia coli, where this study verified the biological function of the silver complexes that exceeded the silver nitrate. Palladium (II) NHC complexes aren't very common in the medicine, Yet it were also investigated as a potential solution to the treatment of many cancers that do not lead to existing cancer therapies Ray and colleagues researched the effectiveness of these compounds against colon cancer (adenocarcinoma) in 2007, and the results proved that it is much stronger than cis-platin⁸. These complexes demonstrated antimicrobial activity, especially against Escherichia coli, when recently tested.

2. EXPERIMENTAL:

2.1 General considerations:

All chemicals and solvents, were of the highest analytical grade and used as supplied from commercial sources. Nuclear magnetic resonance (NMR) spectra were recorded using Bruker 400 MHz spectrometers at ambient temperature. 'H NMR peaks are labeled as singlet (s), doublet (d), triplet (t) and multiplet (m), chemical shifts were referenced with respect to solvent signals. The infrared spectra were recorded with FT-IR spectrophotometer (FTIR- 8400s, Bruker). UV-Visible spectrometer double beem were assigned on (Shimadzu UV 1650 PC) and The elemental analysis (CHN) were carried out on (PerkinElmer series II, 2400 microanalyzer).

2.2 Synthesis of 1-(2-((4-chlorophenyl)amino)-2-oxoethyl)-3-(4-cyanobenzyl)-1H-benzo[d]imidazol-3-ium chloride (A)

(2g, 0.0085mol) 4-(1H-benzimidazol-1-ylmethyl) benzonitrilein 10mL of dioxane was placed in a 50mL round bottom flask. (1.75g , 0.0085 mol) of2-chloro-N-(4-chlorophenyl) acetamidewas added onto the mixture. The mixture was refluxed at 90^oC for 24 hours. After the completion of reaction, the solvent was evaporated then recrystallized using methanol, 2.8g (75% yield ) as a white powder (m.p = 288-290℃). FT-IR cm^-1: 3244 , 3045, 2979, 2861, 2227, 1669, 1598, 1490

¹H NMR (400MHz,DMSO) ppm : 10.98(NHCO), 10.00(benzimi H2’), 7.96, 8.25 (C-Hbenzimidazole), 7.68 , 7.24(Ar-H), 7.96 , 7.41(ArCN-H), 5.63((benzylic-CH₂), 6.06(CH₂-ArCN).Anal. Calc. for C₂₃H₁₈Cl₂N₄OC: 63.17; H: 4.15; N: 12.81. Found: C: 63.09, H: 4.09, N: 12.75.

2.2 Synthesis of Silver(I)-NHC Complex:

2.2.1 Synthesis of bis(1-(2-((4-chlorophenyl)amino)-2-oxoethyl)-3-(4-cyanob- enzyl)-2,3-dihydro-1H-benzo[d]imidazol-2-yl)silver chloride(B)

Silver oxide (0.5g, 0.002mol) was added to a solution of compound (A) (0.8g , 0.002mol) in 20mL methanol . The mixture was stirred for 10 hr in glassware, covered by aluminum foil. After the black suspension was filtered through the celite to remove the excess Ag₂O, the solvent was removed to give the product as a white solid 0.8g (61 % yield) (m.p = 223-225 ℃). FT-IR cm^-1 :3260, 3036, 2956, 2848, 2228, 1685,1608, 1490 ¹H NMR (400MHz,DMSO) ppm: 10.78 (NHCO),7.39, 7.23, 8.46 (C-Hbenzimidazole), 7.30 , 7.09 (Ar-H), 7.64, 7.55 (ArCN-H),4.56 ((benzylic-CH₂), 5.22 (CH₂-ArCN). Anal. Calc. for [C₄₆H₃₄AgCl₂N₈O₂]Cl, C: 58.46; H: 3.63; N: 11.86. Found: C: 58.34, H: 3.52, N: 11.83.

2.4 Synthesis ofbis-NHC palladium(II) complex

2.4.1 Preparation of Bis (acetonitrile) dichloropalladium (II) (C)

Palladium chloride (0.9g,0.005mol) was suspension with acetonitrile (30mL) and heated under reflux at 90℃ for 1 hr. The mixture was allowed to cool down and the solvent was evaporated⁸⁹ to give complex (C) as orange-reddish 0.87 g (96.66% yield ) (m.p = 142-144℃) .

2.4.2 Synthesis of bis(1-(4-cyanobenzyl)-3-(2-((2,5-dimethylphenyl)amino)-2-oxoethyl)-2,3-dihydro-1H-benzo[d]imidazol-2-yl)palladium(II) dichloride (D)

Palladium complex (C) (0.04g, 0.0001mol) was dissolved in methanol (7.5mL) and the silver complex (B) ( 0.16g ,0.0001mol ) was dissolved in methanol (7.5 mL) then the complex ( C ) solution was added to the complex (B) solution, drop by drop, then the mixture was stirred for 4 hours at room temperature . The product was filtered by using celite, the solution is left todrytoobtain0.11 g (84 % yield) as a pale-Brown solids (m.p = 183-185℃).FT-IR cm^-1 : 3017,2957 ,2848,2226,1663,1616,1497¹H NMR (400MHz,DMSO) ppm : 9.61,9.97 (NHCO),7.91, 7.73, 8.06 (C-Hbenzimidazole), 7.39,7.40 , 7.20,7.26 (Ar-H), 7.65, 7.67, 7.87,7,89 (ArCNH),4.29,4.37 (benzylic-CH₂), 5.60, 5.97 (CH₂-ArCN).Anal. Calc. for C₄₆H₃₄Cl₄N₈O₂Pd C: 56.43; H: 3.50; N: 11.45. Found: C: 56.22, H: 3.42, N: 11.12.

2.5 Antibacterial activity test:

Ligand (A) and complexes (B and D) were screened for antibacterial activity against Staphylococcus aureus and Escherichia coli in Muller Hinton agar by measuring the inhibition zone in (6mm). Azithromycin: AZ(200,400) µg\ml was the chosen standdard drug for antibacterial activity. Each bacteria isolate was inoculated on to the Muller- Hinton Agar [sterilize in autoclave] by dipping a cotton swab in to the suspension and streaking over the surface of the agar plates. Then, in the solidified medium, four holes were made (6 mm). These holes were filled with (0.5 ml) of the prepared compounds (200,400) µg/ mL of the compound dissolved in 1 mL of DMSO solvent). These plates were incubated at 37 °C and measured of zone inhibition after 48 hours.

3. RESULTS AND DISCUSSION:

3.1 Synthesis:

1-(2-((4-chlorophenyl)amino)-2-oxoethyl)-3-(4-cyanobenzyl)-1H-benzo[d]imidazol-3-ium chloride (A) was prepared by the reaction 4-(1H-benzimidazol-1-ylmethyl) benzonitrilewith 2-chloro-N-(4-chlorophenyl) acetamide. The Ag-NHC complex was synthesized in analogous to the reported procedure by Wang and Lin⁹. The benzimidazolium salts (A) was treated with silver oxidein methanol to form the desired Ag-NHC complex (B). The palladium complex (C) were prepared via the transmetallationof corresponding silver complex (B) by treating these complex with Pd(CH₃CN)₂Cl₂ complex with methanol solvent. This method is described by Wang and Lin⁹, Scheme (3.1) showed the preparation of compound (A,B and D).

Scheme (3.1) preparation of compounds (A,B and D)

3.2 Identifications via all techniques of ligand and complexes:

3.2.1 UV-Vis Study:

The spectra of ligand were recorded in methanol. The (A) spectrum showed two distinguishable bands at 240 and 280 nm these bands can be indicating to π-π* and n-π* for the benzimidazole ring Figure (3.1). In addition to the absorption peaks 253, 268nm due to electronic transitions in ligands, the novel Ag-NHC complex (B) showed a new absorption peaks at 344 nm, related to MLCT (metal-ligand charge transfer). This complex couldn't show any d-d transition due to its d¹⁰configuration because of d- d transitions are forbidden by the Laporte selection rule, which confirms the absence of any (d-d) transitions and absence of visible region absorptions in their electronic spectra. The difference in absorption peaks and the appearance of new absorption peaks confirm the complexity process figures (3.2) .Pd (II)- NHC complex spectrum of (D) show three bands in 356, 540 and 684nm attributed to ѵ2 , ѵ3 and π- π* transitions, which are referenced to square planar stereochemistryFigure (3.3).

3.2.2 FTIR Study:

The FT-IR spectrum of ligand (A), was gave the following peaks which was assigned as following, a band at 3244 cm^-1 due to (N-H) stretching vibration respectively, 2227 cm^-1 due to (C≡N) group respectively, 1669 cm^-1 due to (C=0) group respectively, 1598 cm^-1 due to (C=N) Figure (3.4).The FTIR spectra of complex (B), Figure (3.5) showed the following characteristic peaks; bands at 3260cm^-1 for the (N-H) stretching vibration respectively, the bands 1667 cm^-1assigned to for δ(C=O), The peak (1608) cm^-1 for the (C=N) of benzimidazolium ring was shifted in the ligands. This shifting may be caused by the contribution of the back bonding of Ag electrons. Thus the bond order of C=N would be enhanced after complexation.The FT-IR spectrum of complex (D)Figure (3.6) showed the following characteristic peaks: bands at 3261 cm^-1 for the (N-H) stretching vibration respectively the bands at 1668 cm^-1 attributed to δ(C=O) stretching vibration,

3.2.3 NMR study:

The ¹H NMR for (A) in d₆-DMSO showed the singlet peak at (10.98) ppm was assigned to amide proton (NHCO). The signal of benzimidazolium proton H2΄ appeared at 10.00 ppm as a singlet peak. In addition, the -CH₂ proton peaks bonding to the Ar-CN ring and the benzylic -CH₂ has been observed as triplets at (6.06, 5.66) ppm.The aromatic proton peaks of Ar-CN ring have observed as multiplited at 7.96, 7.41 ppm. The aromatic proton peaks of aryl ring have observed as multiple at 7.68 , 7.24 ppm.The aromatic proton peaks of benzimidazole have observed as multiple between (7.68-8.25). Figure (3.7).The ¹H NMR for ( B,D ) in d₆-DMSO showed the singlet peaks at (10.78, 9.61) ppm were assigned to amide proton (NHCO) respectively. In addition, the - CH₂ proton peaks bonding to the Ar-CN ring and the benzylic -CH₂ has been observed as triplets at 5.22, 4.56 and 5.97, 4.29 ppm. The aromatic proton peaks of Ar-CN ring have observed as multiple at 7.64, 7.55 and 7.67, 7.87 ppm. The aromatic proton peaks of aryl ring have observed as multiple at 7.30, 7.09 and 7.39, 7.20 ppm. The aromatic proton peaks of benzimidazole have observed as multiple between 8.46 - 7.39 and 8.06 - 7.73 ppm respectively.The successful coordination of carbon carbene to (B, D) complexes via de-protonation of C2 is the absence of characteristic singlet protons 10.00 ppm compared with (A) ligand spectra¹⁰ Figures (3.8) and (3.9) respectively. The ¹H NMR for (D) show two sets of closely spaced signals. These spectra are assigned two sets of resonances to two isomers, which indicate the existence of an inseparable isomeric mixture for most of the complex that attributable to inter conversion between cis and trans conformers which is relatively fast on the NMR timescale^11-14.

3.2.4 The Antibacterial Activity Study:

In general, in comparison with azithromycin, all the substituted benzimidazolium salts and respective Ag(I) and Pd(II) complexes showed an activity against the tested bacteria. A representative picture of zone of inhibition is shown in figures 3.10, 3.11 and Table (3.1). According to the tabulated results,, the antibacterial activity of (B) is the highest inhibition against the tested bacteria even more than the azithromycin.A moderate activity for the complex (D), low activity for the ligand (A)hase been shown. For the bacteria Escherichia coli, the results were almost similar to that of S.aureusas depicted in Figures 3.12, 3.13 . Once again, complex(B) showed the highest activity against the tested bacteria and moderate for the complex (D). The ligand (A) showed low antibacterial activity. Similarly, the sensitivity of the gram positive bacteria increases as the concentration of the complex suspensions increases.All other compound showed different values at the concentrations 200 and 400 µg L^-¹.

Table (3.1) Antibacterial activities of compounds A, B, and D against S. aureus and E. coli

Compound	*S.aureusas Inhibition zone (mm)*		*E,coli* Inhibition zone (mm)
	200µg ml^-¹	400µg ml^-¹	200µg ml^-¹	400µg ml^-¹
A	8	12	8	12
B	14	20	14	20
D	11	14	11	14
AZ	15	18	15	18

Figure (3.10) Antibacterial activities of compounds (A, B and D) against Staphylococcus aureus

Figure (3.11) Antibacterial activities of compounds (A, B and D) against Escherichia coli

4. CONCLUSION:

In conclusion, a new unsymmetrical ligand (A) of replaced benzimidazolium salts was preperated. At 90oC, benzimidazole reacts to formed benzimidazolium salts with aromatic substituents. This approach demonstrated good yield. The 1H NMR spectra showed good signalsfor the prepared benzimidazolium salt. A new Ag(I)-and Pd(II)-NHC complexes were prepared from prepared benzamidazolium salt . These complexes (B,D) were characterized by ¹HNMR spectroscopy, FT-IR, visible UV spectroscopy, C.H.N elemental micro analysis and melting point. Based on an in situ technique by reacting Ag₂O with precursor salt of NHC in suitable solvents such as methanol, dioxane and dichloromethane, these compounds were prepared. Pd(II)-NHC complex (D) was synthesized in quantitative yields under a mild conditions by using transmetallation method of corresponding of Ag(I)-NHC complex as a carbene transfer ligand .Been discussing the ability of the NHC salts and their respective Ag (I), and Pd (II) complexes to kill bacteria. benzimidazolium ligands, Ag (I) and Pd (II) -NHC complexes (A, B, and D) showed acceptable cytotoxicity.

5. REFERENCES:

1. H. V. Huynh, The Organometallic Chemistry of N-heterocyclic Carbenes. 2017.

2. M. Fèvre, J. Pinaud, Y. Gnanou, J. Vignolle, and D. Taton, “N-Heterocyclic carbenes (NHCs) as organocatalysts and structural components in metal-free polymer synthesis,” Chem. Soc. Rev., vol. 42, no. 5, pp. 2142–2172, 2013, doi: 10.1039/c2cs35383k.

3. T. Cantat, N. Mézailles, A. Auffrant, and P. Le Floch, “Bis-phosphorus stabilised carbene complexes,” Journal of the Chemical Society. Dalton Transactions. 2008, doi: 10.1039/b717600g.

4. W. A. Herrmann, “N-heterocyclic carbenes: A new concept in organometallic catalysis,” Angewandte Chemie - International Edition. 2002, doi: 10.1002/1521-3773(20020415)41:8<1290::AID-ANIE1290>3.0.CO;2-Y.

5. T. Weskamp, V. P. W. Bhm, and W. A. Herrmann, “synthesis *,” vol. 600, pp. 12–22, 2000.

6. C. M. Crudden and D. P. Allen, “Stability and reactivity of N-heterocyclic carbene complexes,” Coord. Chem. Rev., vol. 248, no. 21–24, pp. 2247–2273, 2004, doi: 10.1016/j.ccr.2004.05.013.

7. N. C. Kasuga, A. Sugie, and K. Nomiya, “Syntheses, structures and antimicrobial activities of water-soluble silver(I)-oxygen bonding complexes with chiral and racemic camphanic acid (Hca) ligands,” Dalt. Trans., 2004, doi: 10.1039/b411859f.

8. S. Ray et al., “Anticancer and antimicrobial metallopharmaceutical agents based on palladium, gold, and silver N-heterocyclic carbene complexes,” J. Am. Chem. Soc., vol. 129, no. 48, pp. 15042–15053, 2007, doi: 10.1021/ja075889z.

9. X. Wang, S. Liu, and G.-X. Jin, “ Preparation, Structure, and Olefin Polymerization Behavior of Functionalized Nickel(II) N-Heterocyclic Carbene Complexes † ,” Organometallics, 2004, doi: 10.1021/om049467z.

10. J. C. Garrison and W. J. Youngs, “Ag(I) N-heterocyclic carbene complexes: Synthesis, structure, and application,” Chemical Reviews. 2005, doi: 10.1021/cr050004s.

11. M. V. Baker et al., “Cationic, linear Au(I) N-heterocyclic carbene complexes: Synthesis, structure and anti-mitochondrial activity,” Dalt. Trans., 2006, doi: 10.1039/b602560a.

12. A. A. Danopoulos, S. Winston, and M. B. Hursthouse, “C-H activation with N-heterocyclic carbene complexes of iridium and rhodium,” J. Chem. Soc. Dalt. Trans., 2002, doi: 10.1039/b205617h.

13. A. M. Magill et al., “Palladium(II) complexes containing mono-, bi- and tridentate carbene ligands. Synthesis, characterisation and application as catalysts in C-C coupling reactions,” J. Organomet. Chem., 2001, doi: 10.1016/S0022-328X(00)00720-8.

14. R. A. Haque, M. Z. Ghdhayeb, A. W. Salman, S. Budagumpi, M. B. Khadeer Ahamed, and A. M. S. Abdul Majid, “Ag(I)-N-heterocyclic carbene complexes of N-allyl substituted imidazol-2-ylidenes with ortho-, meta- and para-xylyl spacers: Synthesis, crystal structures and in vitro anticancer studies,” Inorg. Chem. Commun., vol. 22, pp. 113–119, 2012, doi: 10.1016/j.inoche.2012.05.037.

Received on 28.10.2020 Modified on 10.12.2020

Research J. Pharm. and Tech 2021; 14(10):5366-5370.

DOI: 10.52711/0974-360X.2021.00936