Modified Clays and Modified Clay–Based Antimicrobial Polymer Nanocomposites
MMT signifies a group of 2:1 phyllosilicate materials with the advantages of easy intercalation capabilities, easy availability, and low cost [140-144]. MMT is a unique ID layered silicate material that possesses a unique crystal structure. Chemically it is sodium calcium aluminum magnesium silicate hydroxide (Na,Ca)o.33(Al,Mg)2(Si401o)(OH)2-nH20. Due to a low layer charge on the platelets, a weak van der Waals force exists between the layers, which makes the cations in the interlayer exchangeable by simple cation exchange reactions. This leads to high intercalation capabilities in between the layers where cationic materials of our choice can be incorporated by cation exchange reactions and thus be used to suit the needs of various end use applications. For increasing the efficacy of dispersion of nanoclay within the polymer matrix, long-chain organic cations (alkylammonium, alkyl phosphonium, and alkyl imidazol(idin)ium are often intercalated into MMT with an aim to increase the hydrophobicity of the material. The hydrophobic interactions between the tethered ammonium surfactant intercalated inside the clay and the microbe cell cause alteration in the permeability of the cell membranes, leaking out low-molecular-weight metabolites and hampering healthy cell functions . Organomodified nanoclay offers a number of advantages, along with significantly increasing the mechanical properties of the neat polymer, and is commercially available.
As previously emphasized, the direct usage of NPs, though highly efficient against microbes, presents two problems. First, as NPs have a very high surface area, they tend to coalesce or form aggregates; thus stability is a problem. Second, as NPs are hydrophobic, large quantities are required for optimum results. Therefore, colloids and deposition of NPs on a suitable substrate have been attempted to overcome this deficiency. A popular approach utilizes the adsorption capacity of MMT where Na/Ca ions in the interlayer space are replaced by active ions/NPs. Thus, an NP-clay complex structure is obtained that possesses excellent antimicrobial activity without having the drawbacks associated with Ag NPs.
Synthesis of silver-exchanged montmorillonite (Ag-MMT) has been reported in a few papers [145-150] and its antimicrobial property studied for a variety of applications, ranging from antibiotics for animal feed to material for packaging applications. The preparation of Ag-MMT from MMT with the incorporation of Ag atoms is essentially an ion exchange reaction process and involves breaking down the individual tactoids of the MMT to reduce the thickness to the nanometer regime, achieved by a suitable mechanical attrition method like dry ball milling for 60 min. Further, an Ag precursor salt solution is added so that the clay platelets are homogeneously covered by Ag. Girase et al.  reported the synthesis of Ag- MMT via three synthesis methods: by reduction with sodium borohydride, by calcination at 400°C, and by UV irradiation. The after-treatments were carried out on three different samples after the ion exchange reaction. The authors reported that the largest particle sizes of Ag NPs precipitated on the clay structure were obtained in the case of calcination after treatment (20-25 nm), followed by sodium borohydride (NaBH4) reduction (10 nm), and the smallest particle sizes were obtained by the UV irradiation method (5-10 nm). Synthesis of Ag NPs attempted without the use of clay presented larger particle sizes (60-200 nm). Smaller particle sizes and uniformity in dispersion were obtained in the samples with clay support as the clay provided a surface for NPs to precipitate and acted as a nucleating agent. In samples where clay was absent, there was ample opportunity for Ag NPs to coalesce and form aggregates, resulting in larger particle sizes. Release tests showed that a metal-clay complex structure is much more stable than unsupported Ag NPs and exhibits a controlled release mechanism that lasts a longer time. The antimicrobial activity of the Ag-clay structure was also much higher—4 times that of Ag NPs— owing to the smaller size of the Ag NPs formed in samples with clay. Ag-MMT with smaller NP sizes can also be synthesized by modifying the starting raw material, that is, Na-MMT. Tian et al.  modified Na-MMT with three sulfur-containing amino acids separately. The amino acids were 1-cystine, 1-cysteine, and 1-methionine. This was followed by an ion exchange reaction for successful loading of Ag under microwave irradiation. Ag atoms were anchored to the sulfur atoms on the modified MMT, preventing Ag aggregation, thus reducing the particle size. The highest loading of Ag, of about 11 wt%, was obtained in the case of Na-MMT modified with 1-cystine, as confirmed by X-ray photoelectron spectroscopy (XPS) and Fourier-transform infrared spectroscopy (FTIR). Ag-MMT prepared by the same method, that is, ion exchange followed by UV photoreduction by Xu et al. , reported Ag content as determined by the Volhard method to be 6.4 wt%. The diameter of Ag NPs characterized by SEM was about 15-20 nm. The metal- clay complex structure helped in a sustained release profile. When tested against bacteria E. coli, the minimal inhibitoiy concentration (MIC) and sterilizing efficiency of Ag-MMT was determined to be around 100 x 10-6 and 100%, respectively. Ag-MMT was formed by the intercalation of Ag ions into MMT with reduction with borohydride and formaldehyde . Smaller particles with an even distribution were obtained in the case of borohydride reduction, but aggregates of Ag NPs, both large and small in size, were obtained with the use of formaldehyde as a reducing agent.
Transmission electron microscope (ТЕМ) micrographs revealed that both crystalline (cubic/hexagonal) and amorphous particles were formed on the surface of MMT. A comparison was made between MMTs calcined at 550 C for 3 h and MMTs grinded for 300 s . It was hypothesized that during thermal treatment or calcination, crystallization water is lost, which results in phase transformation, whereas mechanical attrition methods like grinding or ball-milling separate the individual tactoids, collapsing the interlayer structure and increasing the surface area. The grinded sample presented a lower MIC value than the annealed sample, but in the disk diffusion test, the thermally treated sample had a bigger zone of inhibition.
Ag NPs were deposited over MMT in an attempt to produce Ag-MMT in the presence and absence of sodium carbonate in two media—water and ethylene glycol . Ethylene glycol was chosen as an ecofriendly reducing agent. In samples where NaC03 was used, Ag carbonate NPs also formed and were responsible for smaller particle sizes of Ag NPs, as confirmed by ТЕМ studies. In samples where carbonate was not used, larger particles resulted, indicating the crucial role played by carbonate ions in retarding the growth of Ag NPs. Antimicrobial test against £ coli proved that the sample prepared in water media and having Ag2C03 NPs had the highest antimicrobial activity. Ag ions were also used to prepare other clay structures, such as a Ag-vermiculite hybrid structure . The mean size distribution of Ag-MMT when compared to Ag-vermiculite was much narrower. Ag-vermiculite is also a potent bacteria inhibitor and showed greater resistance against bacteria K. pneumonia and Pseudomonas aeruginosa. Ag-MMT has also been used to impart antibacterial activity to conventional sutures by the dipping and rolling method . It exhibited good blood and tissue biocompatibility and when tested against bacteria, it showed over 99% inhibition with Staphylococcus aureus and E. coli bacteria.
Copper-exchanged montmorillonite (Cu-MMT) has been prepared from calcium montmorillonite, Na-MMT, and acid-activated montmorillonite (AA-MMT), and their antibacterial activities have been compared . All three samples acted as potent antibacterial agents having antimicrobial activity ranging from 98.6% to 95.6%. The highest antibacterial activity, that is, 98.6%, was exhibited by Cu-MMT prepared from AA-MMT. No reduction in the antimicrobial property was observed after it was washed in saline for 24 h. Na- MMT was first modified by an anionic biosurfactant, which helps in better distribution of clay platelets in solution. The adsorption rate of MMT was improved owing to a decrease in the diffusion resistance. The antibacterial property of Cu-MMT prepared by the ion-exchange method using CuS04 has also been evaluated against £. coli and Salmonella choleraesuis in vitro . The MIC values of Cu-MMT against £. coli and Salmonella choleraesuis were 1024 mg/mL and 2048 mg/mL, respectively. This is due to the desorption of Cu ions from Cu-MMT and is estimated to be around 6.51-45.65 mg/mL by atomic absorption spectroscopy. ТЕМ studies revealed that the cell walls of the bacteria were structurally impaired, leading to the release of intracellular materials and enzymes.
In an interesting work, Malachova et al. described and compared the antimicrobial activity of Ag, Cu-MMT, and Zn-exchanged montmorillonite (Zn-MMT) against both bacteria and fungi . These were tested against bacteria £. coli and white rot fungi Pycnoporus cinnabarinus and Pleurotus ostreatus. Against E. coli, the highest antibacterial activity was exhibited by Ag-MMT, while the antibacterial activities of Cu-MMT and Zn-MMT were more or less the same. This order of bacterial inhibition is comparable to the activity exhibited by free ions of Ag, Cu, and Zn. Ag ions are the most effective in inhibiting the bacteria £. coli owing to their reaction with the thiol groups present in the cytoplasm. But the results against fungi are different from that against bacteria. Against Pycnoporus cinnabarinus, Zn-MMT showed the highest antifungal activity, followed by Cu-MMT and Ag-MMT. Against Pleurotus ostreatus, the highest antifungal activity was demonstrated by Cu- MMT, followed by Zn-MMT and Ag-MMT.