Synthesis, Characterization, Biological Evaluation, and In Silico Studies of Imidazolium-, Pyridinium-, and Ammonium-Based Ionic Liquids Containing n-Butyl Side Chains

Ionic liquids (ILs) have emerged as active pharmaceutical ingredients because of their excellent antibacterial and biological activities. Herein, we used the green-chemistry-synthesis procedure, also known as the metathesis method, to develop three series of ionic liquids using 1-methyl-3-butyl imidazolium, butyl pyridinium, and diethyldibutylammonium as cations, and bromide (Br−), methanesulfonate (CH3SO3−), bis(trifluoromethanesulfonyl)imide (NTf2−), dichloroacetate (CHCl2CO2−), tetrafluoroborate (BF4−), and hydrogen sulfate (HSO4−) as anions. Spectroscopic methods were used to validate the structures of the lab-synthesized ILs. We performed an agar well diffusion assay by using pathogenic bacteria that cause various infections (Escherichia coli; Enterobacter aerogenes; Klebsiella pneumoniae; Proteus vulgaris; Pseudomonas aeruginosa; Streptococcus pneumoniae; Streptococcus pyogenes) to scrutinize the in vitro antibacterial activity of the ILs. It was established that the nature and unique combination of the cations and anions were responsible for the antibacterial activity of the ILs. Among the tested ionic liquids, the imidazolium cation and NTf2− and HSO4− anions exhibited the highest antibacterial activity. The antibacterial potential was further investigated by in silico studies, and it was observed that bis(trifluoromethanesulfonyl)imide (NTf2−) containing imidazolium and pyridinium ionic liquids showed the maximum inhibition against the targeted bacterial strains and could be utilized in antibiotics. These antibacterial activities float the ILs as a promising alternative to the existing antibiotics and antiseptics.


Introduction
The development of robust broad-spectrum antibacterial drugs to treat common skin and soft tissue infections (SSTIs) is a major challenge in the healthcare community. Various chemical synthetic products have been reported to combat pathogenic bacteria [1]. The continuous emergence of antibiotic resistance makes it so that the development of new antibiotics requires time [2]. SSTIs affect workers, families, companies, and countries by compromising human health. The different species of pathogenic bacteria that cause SSTIs are: Escherichia coli (E. coli); Klebsiella aerogene/Enterobacter aerogenes (K. aerogene); Klebsiella pneumoniae (K. pneumoniae); Proteus vulgaris (P. vulgaris); Pseudomonas aeruginosa (P. aeruginosa), namely, Streptococcus pyogenes (S. pyogenes) and Streptococcus pneumoniae (S. pneumoniae). Gram-positive cocci cause one out of three skin-infection wounds, while Gram-negative bacteria, such as E. coli and K. pneumoniae, are under observation and alert by the World Health Organization because of their increasing resistance against Table 1. Library of ionic liquids synthesized in lab.

Imidazolium-Based ILs
Pyridinium-Based ILs Quaternary Ammonium-Based ILs and leading to the cell bursting. Quaternary ammonium ions, especially, have been reported to change the outer zeta potential of Gram-negative bacteria, and this changes the cellular function. ILs have also been reported to affect the diffusion, fluidity, and permeability of cellular membranes [24]. Therefore, in this study, we used the most common pathogenic bacteria (i.e., five strains of Gram-negative bacteria and two strains of Gram-positive bacteria) with our library of compounds to evaluate the antimicrobial potential of the synthesized ILs. Moreover, we also carried out the modeling of the ionic-liquid antimicrobial activity through FTIR spectral-data results [25]. Herein, we report the synthetic procedures and in vitro antibacterial studies of these synthesized ILs in detail. and leading to the cell bursting. Quaternary ammonium ions, especially, have been reported to change the outer zeta potential of Gram-negative bacteria, and this changes the cellular function. ILs have also been reported to affect the diffusion, fluidity, and permeability of cellular membranes [24]. Therefore, in this study, we used the most common pathogenic bacteria (i.e., five strains of Gram-negative bacteria and two strains of Gram-positive bacteria) with our library of compounds to evaluate the antimicrobial potential of the synthesized ILs. Moreover, we also carried out the modeling of the ionic-liquid antimicrobial activity through FTIR spectral-data results [25]. Herein, we report the synthetic procedures and in vitro antibacterial studies of these synthesized ILs in detail. properties, have the potential to interact with the cellular membrane, producing lipids and leading to the cell bursting. Quaternary ammonium ions, especially, have been reported to change the outer zeta potential of Gram-negative bacteria, and this changes the cellular function. ILs have also been reported to affect the diffusion, fluidity, and permeability of cellular membranes [24]. Therefore, in this study, we used the most common pathogenic bacteria (i.e., five strains of Gram-negative bacteria and two strains of Gram-positive bacteria) with our library of compounds to evaluate the antimicrobial potential of the synthesized ILs. Moreover, we also carried out the modeling of the ionic-liquid antimicrobial activity through FTIR spectral-data results [25]. Herein, we report the synthetic procedures and in vitro antibacterial studies of these synthesized ILs in detail.
The interaction of ILs and cells (prokaryotic or eukaryotic) is the major question to be answered before using ILs for antimicrobial activity. ILs, because of their amphiphilic properties, have the potential to interact with the cellular membrane, producing lipids and leading to the cell bursting. Quaternary ammonium ions, especially, have been reported to change the outer zeta potential of Gram-negative bacteria, and this changes the cellular function. ILs have also been reported to affect the diffusion, fluidity, and permeability of cellular membranes [24]. Therefore, in this study, we used the most common pathogenic bacteria (i.e., five strains of Gram-negative bacteria and two strains of Gram-positive bacteria) with our library of compounds to evaluate the antimicrobial potential of the synthesized ILs. Moreover, we also carried out the modeling of the ionic-liquid antimicrobial activity through FTIR spectral-data results [25]. Herein, we report the synthetic procedures and in vitro antibacterial studies of these synthesized ILs in detail.

Characterization Techniques
Thin-layer chromatography (TLC) was carried out to check the progress of the reaction. Analytical TLC was performed using Merck-prepared plates (silica gel 60 F-254 on aluminum). The solvent system used to check the completion of the ion-exchange reaction was DCM:MeOH:CH 3 COOH (80:20:5). Spectroscopic methods were used for the chemical-structure elucidation. Infrared spectra were obtained on a Bruker platinum ATR model Alpha spectrophotometer (Germany). A total of 20 mg of sample was scanned from the 4000 to 400 cm −1 wave numbers. 1 HNMR spectra were recorded for all samples at room temperature in deuterated dimethyl sulfoxide (DMSO-d6) on a 400 MHz Bruker AV400 spectrometer (Bruker Corporation, Billerica, MA, USA), with a concentration of 20 mg/mL −1 . An agar well diffusion assay was carried out to evaluate the antibacterial potential of the prepared samples against pathogenic bacteria. A total of 6 different species/strains of bacteria were tested against 18 ionic-liquid samples. Fresh bacterial culture (24 h) of five Gram-negative strains (Escherichia coli; Klebsiella aerogene/Enterobacter aerogenes; Klebsiella pneumoniae; Proteus vulgaris; Pseudomonas aeruginosa) and two Grampositive strains (Streptococcus pyogenes and Streptococcus pneumoniae) were used in this experiment. Each bacterial culture was freshly prepared in nutrient broth to the logarithmic phase at 37 • C in an orbital shaker from 16 to 18 h. A UV-visible spectrophotometer was used to check the optical density of the culture at 600 nm. When the optical density was between 0.58 and 0.6, working solutions of a 10 6 -10 5 CFU/mL bacterial concentration from these bacterial cultures were prepared for the antibacterial studies. Samples of equal concentrations and densities were prepared in sterile distilled water. Sterile Petri dishes were inoculated with 100 µL of each equalized bacterial sample to create a uniform lawn on each Petri plate. Using a sterilized borer, 3 mm wells were made at equal distances. Levofloxacin as the positive control and distilled water as the negative control were used in equal volume-by-volume ratios on every plate. About 25 µL of each sample solution was poured into each well and was then incubated for 24 h at 37 • C. Each sample was tested in triplicate.

Synthesis
The three different starting ionic liquids synthesized were 1-butyl-3-methylimidazolium bromide, butylpyridinium bromide, and diethyldibutylammonium bromide, which were synthesized by the metathesis method. The 1-methyl-3-butylimidazolium bromide and butylpyridinium bromide were synthesized according to the literature [26]. Their synthesis methods are briefly described in Section 2.3.
Two anions (sodium methanesulfonate (Na [CH 3 SO 3 ]) and sodium dichloroacetate (Na [CHCl 2 CO 2 ])) were synthesized in a lab. Their synthesis methods are briefly described in Section 2.4. 1-methyl imidazole (5 g, 60 mmol) was reacted with butyl bromide (8.36 g, 61 mmol) in acetonitrile (60 mL) to obtain 1-methyl-3-butylimidazolium bromide (Scheme 1), and the reaction mixture was refluxed and stirred for 48 h to obtain the maximum yield of the product. The completion of the reaction was confirmed with the help of TLC, after the reaction completion solvent was evaporated with the help of a rotary evaporator, and the reactants were decanted. Then, the product was dried in vacuum oven for 12 h at 50 • C. The product obtained was a yellow oily liquid with a 92% yield.
Samples of equal concentrations and densities were prepared in sterile distilled water. Sterile Petri dishes were inoculated with 100 µL of each equalized bacterial sample to create a uniform lawn on each Petri plate. Using a sterilized borer, 3 mm wells were made at equal distances. Levofloxacin as the positive control and distilled water as the negative control were used in equal volume-by-volume ratios on every plate. About 25 µL of each sample solution was poured into each well and was then incubated for 24 h at 37 °C. Each sample was tested in triplicate.

Synthesis
The three different starting ionic liquids synthesized were 1-butyl-3-methylimidazolium bromide, butylpyridinium bromide, and diethyldibutylammonium bromide, which were synthesized by the metathesis method. The 1-methyl-3-butylimidazolium bromide and butylpyridinium bromide were synthesized according to the literature [26]. Their synthesis methods are briefly described in Section 2.3.

Synthesis of Bromide-Based Ionic Liquids
Br) Synthesis 1-methyl imidazole (5 g, 60 mmol) was reacted with butyl bromide (8.36 g, 61 mmol) in acetonitrile (60 mL) to obtain 1-methyl-3-butylimidazolium bromide (Scheme 1), and the reaction mixture was refluxed and stirred for 48 h to obtain the maximum yield of the product. The completion of the reaction was confirmed with the help of TLC, after the reaction completion solvent was evaporated with the help of a rotary evaporator, and the reactants were decanted. Then, the product was dried in vacuum oven for 12 h at 50 °C. The product obtained was a yellow oily liquid with a 92% yield.

Butyl Pyridinium Bromide ([C 4 py] Br) Synthesis
Pyridine (5 g, 63.3 mmol) was reacted with butyl bromide (8.66 g, 63.3 mmol) in acetonitrile (60 mL) to obtain butyl pyridinium bromide. The reaction mixture was refluxed and stirred for 48 h to obtain the maximum yield of the product (Scheme 1). After confirmation of the reaction completion with the help of TLC, the reaction was stopped, the solvent was rotary evaporated, and the reactants were decanted to obtain the brown-colored liquid product. The product was further dried in a vacuum oven for 12 h at 50 • C, and the yield was 93%.

Diethyl Dibutyl Ammonium Bromide ([N 2,2,4,4 ] Br) Synthesis
Diethyl dibutyl ammonium bromide was synthesized by reacting diethylamine (2.5 g, 34 mmol) with 2 molar equivalents of bromobutane (9.36 g, 68 mmol) in acetonitrile (60 mL). A total of 3 molar equivalents (10.8 g, 102 mmol) of sodium carbonate (Na 2 CO 3 ) were taken as the base. The reaction mixture containing reactants, solvent, and base was evacuated with the help of a vacuum pump to remove air, and it was then stirred and refluxed under a nitrogen atmosphere at 90 • C for 24 h to obtain the required product (Scheme 1). The completion of the reaction was confirmed with TLC, the solvent was rotary evaporated, the unreacted material was decanted, the base was filtered to obtain pure product, and the pure crystalline product was obtained and dried in a vacuum oven for 12 h at 50 • C; the yield was 93%.

Synthesis of Butyl Imidazolium-Based Ionic Liquids
1-butyl-3-methylimidazolium methanesulfonate was synthesized with the help of a metathesis reaction of 1-butyl-3-methylimidazolium bromide (2.41 g, 11 mmol) with sodium methanesulfonate (1.3 g, 10 mmol). The reaction mixture was stirred overnight at room temperature in methanol (Scheme 2). Sodium bromide was removed from the product with the help of a solvent extraction followed by filtration. Chloroform and ethyl-acetate were used for the solvent extraction of the product from sodium bromide. Several fractions of these solvents containing the product were collected through filtration, and then the solvent was rotary evaporated to obtain a pure product, which was further completely dried in a vacuum oven overnight at 50 • C; the product yield was 78%.

3-Butyl-1-Methylimidazolium Bis(Trifluoromethanesulfonyl)Imide ([C 4 mim] [Tf 2 N]) Synthesis
A solution of LiTf 2 N (0.313 g, 1 mmol) in 2-neck round-bottom flasks was evacuated with the help of a vacuum pump, an inert atmosphere was created in the flask, and then 3-butyl-1-methylimidazolium bromide (0.3 g, 1 mmol) solution was added to the flask. Two reactants were stirred continuously overnight under an inert atmosphere in methanol (60 mL) (Scheme 2). After the product formation, the methanol was evaporated, and the lithium bromide (LiBr) was separated with the help of solvent extraction and filtration, for which chloroform was used, and the chloroform was rotary evaporated. The product formed was dried in a vacuum oven, which resulted in a 78% yield.

1-Methyl-3-Butylimidazolium Tetrafluoroborate ([C 4 mim] [BF 4 ]) Synthesis
Solutions of [C 4 mim] Br (0.59 g, 2.6 mmol) and Na [BF 4 ] (0.3 g, 2.7 mmol) in acetonitrile (60 mL) were kept at room temperature for 12 h in an inert atmosphere (Scheme 2). The NaBr was separated as residue on filter paper and the product was passed as filtrate, and the filtrate was dried through the rotary evaporator to obtain the product, while the remaining NaBr was further removed with the solvent-extraction technique, the solvent was evaporated, and the product was vacuum-dried. Butyl pyridinium bromide (0.9 g, 4 mmol) and sodium methanesulfonate (0.5 g, 4.2 mmol) were stirred overnight at room temperature in methanol (80 mL) to obtain butyl pyridinium methanesulfonate (Scheme 3). After reaction completion, the solvent was rotary evaporated and sodium bromide was solvent-extracted with the help of ethyl-acetate and chloroform. Several fractions of solvent were collected through filtration that contained the product and remaining sodium bromide on filter paper. Fractions of solvent were evaporated, and the product was vacuum-dried.

Butyl Pyridinium Bis(Trifluoromethanesulfonyl)Imide ([C 4 py] [Tf 2 N]) Synthesis
Li [Tf 2 N] (0.3 g, 1 mmol) solution in methanol (30 mL) was formed in 2-neck RB flasks, and it was then evacuated, and an inert atmosphere was created. A solution of [C 4 py] Br (0.39 g, 1.8 mmol) in methanol (30 mL) was added and stirred overnight to form [C 4 py] [Tf 2 N] (Scheme 3). The methanol was evaporated, the LiBr with solvent extraction and filtration was separated from the product, the solvent was rotary evaporated to obtain the pure product, and the product was further dried in a vacuum oven at 40 • C overnight. ] solution and stirred overnight at room temperature (Scheme 3). After product formation, NaBr was separated from the solvent through filtration, and the solvent was rotary evaporated, and further NaBr was separated with solvent extraction followed by filtration. The product was completely dried in a vacuum oven overnight.

H]) Synthesis
Diethyl dibutyl ammonium bromide (1 g, 3.7 mmol) solution in methanol (40 mL) was added to a sodium methanesulfonate (0.44 g, 3.6 mmol) methanol solution (40 mL) and stirred overnight at room temperature to obtain diethyl dibutyl ammonium methanesulfonate (Scheme 4). Methanol was evaporated, sodium bromide was solvent-extracted, solvents were rotary evaporated, and product was completely dried in a vacuum oven. The NaBr was filtered, the solvent was dried, the pure product was obtained using the solvent-extraction technique, and the product was then dried completely.  4). Solvent was rotary evaporated, and KBr was separated through solvent extraction followed by filtration. The product was vacuum-dried to obtain a completely dried product.

Density Functional Theory
All eighteen ionic liquids were modeled and optimized using Gaussian 09 [27]. The geometry optimizations were performed using the functional B3LYP/6-31g level of the density-functional-theory methods [28]. For validation purposes, the frequencies [29] were calculated for the optimized geometries at the same energy level.

Molecular-Docking Studies
For molecular docking, optimized ionic structures were used. The imidazolium-, pyridinium-, and quaternary ammonium-based ionic liquids were docked against six strains of Gram-positive and Gram-negative bacteria (E. coli (PDB ID: 5A924) [30]; K. pneumonia (PDB ID: 6MGX6); E. aerogenes (PDB ID: 5KID5) [31]; P. aeruginosa (PDB ID: 4GZB8) [32]; P. vulgaris (PDB ID: 1HZO7) [33]; S. pneumoniae (PDB ID: 1RPS9) [34]) using PyRx software. As per the requirements, before docking, energy minimization was performed for all proteins where water molecules and protonations were removed. Docking was performed by maintaining default settings for all parameters. Each docked pair of protein and ligand generated nine different poses, out of which the pose having the highest value of binding affinity was chosen.

Results and Discussion
Three different cations (1-butyl-3-methylimidazolium bromide, butylpyridinium bromide, and diethyldibutylammonium bromide) were synthesized by the metathesis method [35], which is also known as the green method of synthesis. The 1-methyl-3-butylimidazolium bromide and butylpyridinium bromide were synthesized according to the literature [11]. Three different series of the ionic liquids were prepared: imidazolium-based, butylpyridinium-based, and ammonium-based ionic liquids. All the samples were characterized by spectroscopic data, and their in vivo antibacterial potentials were carried out.
The IR spectrum of the butyl pyridinium methane sulfonate S=O stretching vibration at 1042 cm −1 shows the exchange of the methane sulfonate anion. The FTIR for the butyl pyridinium bis These IR spectra confirmed the successful synthesis of the new imidazolium-, butyl pyridinium bromide-, and ammonium-based ionic liquids. Furthermore, these ILs were analyzed by NMR spectroscopy.

HNMR Analysis of Ionic Liquids
The 1 HNMRs of the ILs were recorded to understand the structure of the synthesis products by evaluating the environments of the protons in the ILs. The HNMRs of the imidazolium-based ionic liquids showed characteristic chemical-shift values of the 1-methyl-3-butylimidazolium bromide [44] cation, as mentioned in Section 2.7. Further changing the bromide ions in the imidazolium-based ILs had almost similar HNMR spectra, having a small shifting of the chemical-shift values for 3-butyl-1-methylimidazolium bis(trifluoromethanesulfonyl)imide and 3-butyl-1-methylimidazolium tetrafluoroborate, while some new chemical-shift values were also observed, which were as follows: 1-butyl-3-methylimidazolium methanesulfonate CH 3 hydrogens appeared as singlets at 2.84 ppm; 3-butyl-1-methylimidazolium dichloroacetate had 6.32 ppm singlets for -CH-Cl; for the 1-methyl-3-butylimidazolium hydrogen sulfate, a small singlet appeared at 8.5 ppm [45].
Butyl pyridinium bromide has the characteristic 1 HNMR [46], as presented in Section 2.7. In the case of the butylpyridinium bromide methanesulfonate, singlets of the methyl group of methanesulfonate appeared at 2.84 ppm, while CH-Cl of the butylpyridinium dichloroacetate was observed at 6.35 ppm, and the hydrogen of the OH group of the sulfonic part in the butyl pyridinium hydrogen sulfate had much less intensity (around 8.45 ppm), while there was only a small change in the chemical-shift values of the butylpyridinium bis(trifluoromethanesulfonyl)imide. Butylpyridinium tetrafluoroborate was found, which may be due to the close vicinity of the electronegative atoms.
The newly synthesized diethyldibutylammonium bromide, as shown in the HNMR peaks, is depicted in Section 2.7. Further modification into the other quaternary ammoniumbased ILs had the following additions in the quaternary ammonium spectra. The diethyl dibutyl ammonium methanesulfonate had a singlet at 2.84 ppm that corresponded to the CH 3 present in the methanesulfonate, and the diethyldibutylammonium dichloroacetate had a very strong downshift singlet (CH-Cl) appearing at 6.35 ppm because of the electronegative atom (Cl) attached to it. A very small singlet of OH of hydrogen sulfate in the diethyldibutylammonium hydrogen sulfate appeared downshifted at 8.53 ppm. In the cases of the diethyldibutylammonium bis(trifluoromethanesulfonyl)imide and diethyldibutylammonium tetrafluoroborate, no new chemical-shift values were observed, but the chemical-shift values appeared downshifted because of the electronegativity of the fluorine atom.
The NMR spectroscopic data corroborates the FTIR data and strongly confirms the synthesis of the new ILs, and especially the new series of quaternary ammonium-based ionic liquids that were successfully synthesized. Wang J et al. have also reported similar 1 HNMR chemical-shift values for ILs [45].

Antibacterial Activities of Ionic Liquids
Encouraged by our work [11,25], we evaluated the antibacterial potential of the newly synthesized ILs. We evaluated the in vitro antibacterial activity by the agar well diffusion method, and the zones of inhibition were measured in 16 hr time, as shown in Figure 1. All the experiments were performed in triplicate, and the results are shown as average values in Figure 1. Levofloxacin as the control was found to be effective against all the bacterial strains of the five Gram-negative bacterial stains (Escherichia coli; Klebsiella aerogene/Enterobacter aerogenes; Klebsiella pneumoniae; Proteus vulgaris; Pseudomonas aeruginosa) and two Gram-positive strains (Streptococcus pyogenes and Streptococcus pneumoniae). All the imidazolium-based ionic liquids showed the highest antibacterial activity against all the Gram-negative bacteria and Gram-positive bacteria (Figure 1a) [47]. All the butylpyridinium-based ionic liquids showed antibacterial activity against only one Gram-negative bacterium (Enterobacter aerogenes) (Figure 1b) [48], while no activity was found by any of the other butylpyridinium-based ionic liquids, which might be because of the small alkyl chain used in the case of our ILs, while longer chains have been found to be effective against bacteria elsewhere [49]. The ammonium-based ionic liquids (Figure 1c) diethyldibutylammonium tetrafluoroborate and diethyldibutylammonium hydrogen sulfate showed effectiveness against two Gram-negative bacteria (Proteus vulgaris and Klebsiella pneumoniae), while no activity was found against the other strains. These results are quite convincing and promising because ILs with butyl chains are showing positive and better antibacterial effects against pathogenic Gram-negative bacteria to ILs [18,50,51]. Our results are also in accordance with the PLS data calculated by using FTIR [25]. Borkowski, A et al. has reported that the E coli bacterial cultures become adapted and start growing in the presence of tetradecyltrimethylammonium theophyllinate (quaternary ammonium ILs) because quaternary ammonium changes the lipid-membrane structure and protein patterns responsible for bacterial cell death. Bacterial adaptation was responsible for showing antagonistic action with other bactericides [24]. Only imidazolium-based ILs were effective against the Gram-positive bacteria streptococcus pyogenes and streptococcus pneumoniae, while pyridinium and quaternary ammonium were found to be ineffective against Gram-positive bacteria. This can be explained by the fact that the antibacterial potential is dependent on the unique combination of the cations and anions in the ILs. It was also found that the anions here, such as BF 4 − and HSO 4 − , also change the antibacterial potential of quaternary ammonium ILs against Proteus vulgaris and Klebsiella pneumoniae.

In Silico Antibacterial Activity
The in silico antibacterial activity could be analyzed by calculating the protein-ligand binding energy. The minimum energy value shows a strong protein-ligand binding energy, which means strong inhibition. We performed in silico analyses on all 18 ionic liquids with the Beta Lactamase protein of six bacteria: E. coli (PDB ID: 5A92); K. pneumonia (PDB ID: 6MGX); E. aerogenes (PDB ID: 5KID); P. aeruginosa (PDB ID: 4GZB); P. vulgaris (PDB ID: 1HZO); S. pneumoniae (PDB ID: 1RPS). The results are presented in Table 2. According to the computational analysis, the imidazolium-based ionic liquids showed binding-affinity values ranging from −0.8 to −7.9 kcal/mol for all the Gram-positive and Gram-negative bacterial strains analyzed. Comparing all six imidazolium-based ionic liquids showed that the bistrifluoromethane and sulfonylimide (Tf 2 N − ) anions containing ionic liquid showed the maximum inhibitions for all six bacterial strains, with the maximum for E. aerogenes (−7.9 kcal/mol), and the minimum for P. aeruginosa and S. pneumonia (both −5.9 kcal/mol) (Table 2, Figure 2).
Upon comparing the in silico-analysis results for the pyridinium-based ionic liquids, it was observed that the bistriflimide (Tf 2 N − ) anion containing ionic liquid showed the maximum inhibition for all six bacterial strains (from −7.1 kcal/mol for K. pneumonia to −8.5 kcal/mol for S. pneumonia) ( Table 2). It was also observed that the binding energies for the (Tf 2 N − ) anion containing pyridinium-based ionic liquid for E. aerogenes (−8.3 kcal/mol) and S. pneumonia (−8.5 kcal/mol) were similar, which shows that the (Tf 2 N − ) anion containing pyridinium-based ionic liquid had a similar effect on both strains.
Upon analyzing the molecular-docking results for the quaternary ammonium-based ionic liquids, it was observed that the dicholoroacetate anion containing ionic liquid showed the maximum inhibition for all six bacterial strains, with the maximum against E. aerogenes and S. pneumonia (−5.5 and −5.4 kcal/mol, respectively). For the other four bacterial strains, it showed the same binding energy (−4.6 kcal/mol) ( Table 2). From Figure 2 (S1-03-ILPs), we interpret that the imidazole sulfonylimide (Tf 2 N − ) docked with the Ser237, Ser 70, and Ser130 amino acids in the case of P. vulgaris, with Ser 315, Asn153, Ser65, and Tyr151 in the case of E. aerogenes, and with Asn220 and His 250 in the case of K. Paneumoniae were docked with the oxygen of (Tf 2 N − ) for the pyridiniumbased ILs against the bacteria E. aerogenes. Sulfonylimide (Tf 2 N − ) was found docked with Ser 237, as shown in Figure 2 (S2-03-ILP2).
The stabilities of the docked ligands with these bacteria were also observed through the binding interactions between S1-Tf 2 N and the amino acids present at the active sites. The docking of S1-Tf 2 N with E. aerogenes, P. vulgaris, and K. pneumoniae showed the stable hydrophilic interaction of the ligand with the neighboring amino acid residues; Tf 2 N forms the most stable hydrogen-bond interactions (binding affinity: −7.9 kcal/mol) with Ser65, Tyr151, Asn153, and Ser315 when docked with E. aerogenes; with P. vulgaris, the Tf 2 N showed interactions with Ser70, Ser130, and Ser237, whereas S1-Tf 2 N showed hydrogen bonding with Asn220 and His 250 when docked with K. pneumoniae. There were no ligand interactions observed with E. coli and P. aeruginosa. All these interactions indicated the partial positive nature of the binding sites, as in all cases, and the anionic part of the ionic liquids showed interaction with the surrounding amino acid residues.

Conclusions
Three different series of ILs using three cations (imidazolium, pyridinium, and quaternary ammonium) and six anions (bromide; methane sulfonate; bis(trifluoromethanesulfonyl) imide; dichloroacetate; tetrafluoroborate; hydrogen sulfate) are reported for the first time. Spectroscopic data were assessed for the structure confirmation of the ILs. The antimicro-bial potentials of the ILs were evaluated against the most common pathogenic bacteria that cause infections in wounds. Within 24 h of contact with the bacteria, we found that the imidazolium cation was effective against most of the bacterial strains (with all anions), while the pyridinium-based ILs with all anions were effective against Enterobacter aerogenes, the quaternary ammonium with BF 4 − was effective against Proteus vulgaris, and HSO 4 − added antibacterial potential to the quaternary ammonium against Proteus vulgaris and Klebsiella pneumoniae. Our studies are proving that ILs are a good choice in the pharmaceutical industry for antibiotics and antiseptics. According to the molecular-docking results, imidazoliumand pyridinium-based ionic liquids containing the bistriflimide (Tf 2 N − ) anion, whereas the quaternary ammonium-based ionic liquid containing the dichloroacetate (CHCl 2 CO 2 − ) anion showed the maximum inhibition against all six bacterial stains. Upon comparing the overall results, it was observed that the bistriflimide (Tf 2 N − ) containing pyridiniumand imidazolium-based ionic liquids could be a good choice to be used as antibiotics or antiseptics; however, if longer chains (octyl based) are taken, then this would enhance the antibacterial activity.