Design and syntheses of functionalized ionic liquids (ILs) and their nanocomposites for biomedical applications

Abstract

Ionic liquids (ILs) are remarkable chemical compounds exhibiting various applications in vast areas of modern science. Owing to their highly tunable nature along with significant properties, ILs have become essential players in the fields of extraction, catalysis and synthesis, analytics, electrochemistry, biotechnology, materials science, and so forth. Imidazolium/Benzimidazolium-based ILs exhibit very high sensing ability and are extensively used for cation/anion recognition due to their inherently cationic and hydrogen bond donating properties. Apart from the physical and chemical features of ILs, their potent biological activity has attracted significant attention from biochemists and medicinal scientists. Literature reports reveals that the antimicrobial IL materials simply rely on the influence of the organic cations and effect of the anions, whereas the structural symmetry and their modifications through functional moieties are still rarely explored. Therefore, the present investigations were aimed at rational design and syntheses of variedly substituted imidazolium/benzimidazolium-based ILs to address these challenges. The benzimidazolium-based dipodal ionic liquids ILs 1-3, have been designed, synthesized and fully characterized by various spectroscopic techniques. The binding affinity of the synthesized ILs 1-3, was evaluated by UV-visible absorption and fluorescence emission spectroscopy which demonstrated that the ILs 2-3 showed the strong binding affinity towards Fe (III) metal ion and acts as iron chelator. Literature reports also revealed that iron plays an important role in the growth of the microorganism, thus keeping in mind these parameters, the ILs 2-3 were evaluated in vitro for their antibacterial activity by colony forming unit assay, broth microdilution and surface plating method. Results revealed that the obtained ILs 1-3 possess good activity, however, amongst these, IL-3 exhibit significant antibacterial activity. Further experiments explored the mechanism of action of the prepared synthetic siderophore IL-3 that possessing cationic part as well as iron chelating binding sites which initially target the negatively charged bacterial cell membrane and bind with extracellular iron which is present around the environment of the bacteria cell, and make it iron deficient for bacterial consumption that leads to the bacteria cell death. By taking cognizance of antibacterial potential of prepared IL-3, the development of the simple wound dressing gauze loaded with the synthesized IL-3, which were fully characterized and investigated for antibacterial potential. Results revealed that optimized IL-3 coating is significantly active against E. coli and S. aureus. Moreover, the IL-3-coated gauze was also evaluated for its hemostatic potential and results showed that the developed gauze significantly shorten the time period required for blood clotting in comparison to simple dressing gauze and pledget that are commonly used in clinics. Thus, the potential antibacterial and hemostatic results exhibited by developed IL-3-coated gauze make it useful tool that can be used in wards, ICU and particularly, by military persons in war front areas. Furthermore, a pyrimidine-imidazolium and dipodal thiourea based IL conjugate system (IL-4, IL-5 and IL-6) have been designed and synthesized. The synthesized IL-4, IL-5 and IL-6 were fully characterized by 1H NMR, 13C NMR and mass spectroscopic techniques. Furthermore, the prepared ILs-4-6 have been screened for their potential activity and found that IL-5 exhibit potent activity against the investigatory bacterial strains. Based on the design parameter and significant antibacterial activity of IL-5, the IL-5 was decorated onto the surfactant (SDS) and formation of the aggregates. The morphology and size distribution of obtained aggregates were characterized by atomic force microscopy, and dynamic light scattering analysis. Thus, the obtained IL-5-SDS aggregates were screened for their potential activity against examined bacterial strains via colony forming counting (CFU) assay. The minimum inhibitory concentrations (MICs) value was determined and found that the IL-5-SDS exhibits high potent activity against both Gram positive and Gram negative bacteria strains in comparison to the IL-5. These results showed that the enhanced antibacterial activity was achieved towards the investigated strains of bacteria which is due to the cooperative antibacterial effects of ionic liquids and surfactant. The mechanism of action of the IL-5-SDS involves the pierce and breakdown of the bacterial membrane, which leads to cell death as revealed by the AFM and SEM microscopy. Thus, the combining strategy of ionic liquids and surfactant provides the new pathway for preventing the bacterial associated infections Due to the significant antibacterial activity exhibited by ILs, further, it was decided to synthesize the ionic liquid functionalized multiwalled carbon nanotubes to enhance their activity and utilized in the biomedical fields. The prepared ionic liquid functionalized multiwalled carbon nanotubes were fully characterized with various techniques such as FTIR, PXRD, EDS, and SEM and also screened for their potential activity. The obtained results revealed the excellent antibacterial activity exhibited by IL-7d@MWCNT against Gram-positive (Staphylococcus aureus and MRSA) and Gram-negative (Escherichia coli) bacterial strains. The mechanism of action of IL-7d@MWCNT toward the bacterial cell revealed that the hydrophobic segment (long carbon chain) of the IL-7d@MWCNT composite might insert into the lipid membrane of bacteria, leading to the distorted cell membrane composition as observed by SEM and AFM results. Whereas the stronger binding interactions between the cationic moiety and DNA, as supported by DNA binding studies analysis, that may also increase the antibacterial efficiency. The interaction of investigatory IL-7d@MWCNT composite with DNA could be due to hydrophobic–hydrophobic interactions of the DNA bases with the MWCNT or through electrostatic interactions (between the positive cationic part and the negatively charged DNA phosphates). Furthermore, the developed material (IL-7d@MWCNT) is coated onto the surface of polyvinyl chloride (PVC) and possesses hydrophobicity determined through water contact angle (WCA) measurements; and showed long-term antibacterial efficiency against investigatory pathogenic bacterial strains. For biocompatibility assay, the obtained coated PVC material has also been evaluated for its cytotoxicity and results reveal no toxicity against viable cells. These all results are taken together, indicating that by coating with the developed material IL-7d@MWCNT, a robust self-sterilizing surface has achieved, which helps in maintaining a bacteria-free surface. Thus, keeping in mind the overwhelming importance of ILs, a focused on to further explore the N3-substituted varied alkyl chain length attached to benzimidazolium based ILs-8 (a-d). All the synthesized ILs-8 (a-d) were evaluated for their activity against E. coli and S.aureus and found to exhibited significant antibacterial activity. Amongst all the evaluated ILs-8 (a-d), the IL-8d containing long alkyl chain length (C12) was found to show significant inhibitory activity against the examined bacterial strains and thus, possesses potent antibacterial activity as evidenced by antibacterial studies. Further, the IL-8d was utilized for preparing ionic liquid@metal nanocomposites IL-8d@M; (M=Ag, Cu and Au) with different metal ions such as silver (Ag), gold (Au) and copper (Cu), and characterized by AFM, FTIR, EDS, PXRD and DLS. In particular, the IL-8d@Ag exhibit most potent activity against both E. coli and S. aureus bacterial strains with MIC = 12 ± 2 and 08 ± 2 μg/mL, respectively. The mechanism of action for antibacterial activity of IL-8d@Ag nanocomposites was investigated through generation of 1O2 (ROS), whereas the morphology of treated pathogenic bacteria was examined through AFM and SEM analysis. Furthermore, to utilize this developed material IL-8d@Ag in biomedical applications, the prepared ionic liquid material was fabricated onto a microstructured aluminum (Al) substrate with hierarchically arranged functionalities, and the modified surface was characterized and evaluated for their antibacterial activity. Moreover, the hydrophobicity of the material coated onto the Al substrate was also measured by static water contact angle measurement, which reveals its improved hydrophobic character. Thus, the developed hierarchical hydrophobic coating material possessing long-term antibacterial activity on an Al substrate that minimize the wetting by biological secretions and also prevent the substrate from corrosion. In another study, the ZnO nanosheets have been prepared by greener and cost-effective synthesis via the hydrothermal method, and these prepared ZnO nanosheets were further functionalized with eco- friendly ionic liquid (IL-8d). Thus, the sustainable approach was established to synthesize the ZnO nanosheets. The functionalization of ZnO nanosheets by IL-8d was fully examined by advanced spectroscopic and microscopic techniques, such as Fourier-transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM), Powder X-ray diffraction (PXRD), energy-dispersive X-ray spectroscopy (EDS), and thermo-gravimetric (TGA) analysis. Preparation of ionic liquid-functionalized ZnO nanosheets (IL-8d@ZnO) showed self-organized with layered-sheet arrangements caused by intercalation of IL-8d onto the surface of ZnO nanosheets as revealed by the SEM. The design of the IL-8d comprised an acid moiety for functionalization onto the surface of ZnO and whereas the hydrophobicity was tuned through the incorporation of a long alkyl chain. Further, both Gram-positive and Gram-negative pathogenic bacteria were tested in developed IL-8d@ZnO material for their potential antibacterial activity. The antibacterial efficacy of IL-8d@ZnO was evaluated by CFU and MIC tests, and concluded that it has a strong influence on antibacterial activity against tested bacteria’s. Although, the potent activity was observed towards the Gram-positive skin-specific Staphylococcus aureus bacteria strain. Moreover, the mechanism of killing against bacteria have been explored, and the cytotoxicity towards mammalian cells was determined and found to be nontoxic. For the developed material utilization in biomedical fields, the obtained material IL-8d@ZnO was further fabricated on the surface of cotton fibre, their surface morphology was achieved by SEM and the potent activity of IL-8d-ZnO@treated cotton fibre was evaluated by zone of inhibition assay (ZOI). In addition, the stability of the IL-8d-ZnO@treated cotton fibre was evaluated by FTIR and EDS spectroscopy.