Antimicrobial activity of essential oils and metals and their interaction with layered silicates

Iglesias, David Perez (2018) Antimicrobial activity of essential oils and metals and their interaction with layered silicates. Masters by Research thesis (MPhil), Manchester Metropolitan University.

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Abstract

Essential Oils (EOs) and metal ions have been utilised and investigated for antimicrobial applications for centuries; antimicrobial resistance has led to renewed interest in such antimicrobials. This study assesses their use as broad-spectrum antimicrobial agents. The essential oils (EOs) investigated were; Rosewood oil (RO), Clove Leaf Oil (CLO), Orange Oil (OO), Myrtle Oil (MO) and Manuka Oil (MNO). The metal ions studied were; Silver (Ag), Palladium (Pd) and Platinum (Pt). The ultimate aim was to encapsulate the best performing antimicrobial agents in a layered silicate controlled release substrate for use in various polymer-based formulations. The first stage of the study was targeted at assessment of the antimicrobial efficacy of these agents, both individually and in combination (to establish synergistic effects), against Staphylococcus aureus (gram-positive) and Pseudomonas aeruginosa (gram-negative). Antimicrobial efficacy was monitored using Zone of Inhibition, micro-dilution and checkerboard micro-broth dilution methods. Zone of Inhibition (ZoI) proved to be a reliable qualitative method for assessment of the antimicrobial activity of metal ions. Nutrient Agar (NA) was found to be the best growth media for higher metal concentrations (>100 mg/L) and Mueller-Hinton (MH) worked better at lower ion concentrations (<100 mg/L). However, ZoI tests (using wells, solid diffusion or vapour diffusion) were ineffective for assessment of the antimicrobial efficacy of EOs. On an individual basis, all the metal ions (Ag, Pd and Pt) gave a Minimum Inhibitory Concentration (MIC) of 25 mg/L, however, all metal ions, apart from Ag, gave Minimum Bactericidal Concentration (MBC) values higher than 25 mg/L. Of the individual EOs, MNO gave the highest level of antimicrobial performance (MIC of 2.5% (v/v)); no MBC values were recorded for the any of the EOs at the maximum concentration investigated (20%(v/v)). Blends of Ag (50 mg/L) + RO (20%(v/v)) and Ag (50 mg/L) + MNO (5%(v/v)) at a 1:2 ratio shared the highest level of antimicrobial performance, in terms of both MIC and MBC. The next stage of the study involved investigation of incorporation of silver ions and the best performing EO (MNO), plus RO and CLO, due to their relatively simple compositions, into unmodified (sodium) montmorillonite and a range of organically modified montmorillonites (o-MMT). The sodium montmorillonite used was (Cloisite Na+, Rockwood Additives) and the organically modified montmorillonites were (ranked in order of increasing gallery polarity); Cloisite 15A < Cloisite 20A < Cloisite 30B < Claytone APA < Tixogel VXZ (from Rockwood additives and BYK). Silver ions were incorporated in to Cloisite Na+ only, via ion exchange. The adsorption of EOs was monitored using Thermal Gravimetric Analysis (TGA), X-ray diffraction (XRD) and Fourier Transform Infrared Spectroscopy (FTIR). The TGA data enabled determination of the amount of EO adsorption, after taking account of the amount of organic modifier in cases of o-MMT. The XRD data provided insight into the effect of EO adsorption on the stacking uniformity of the MMT platelets. Finally, FTIR data provided insight into oxidation of the EOs and supporting data to verify EO – organic modifier interactions that led to increases in MMT platelet stacking uniformity. O-MMTs with benzyl and hydrogenated tallow functionality (i.e., Claytone APA and Tixogel VZ) provided the best EO adsorption capacity with levels of g EO / 100g o-MMT being achieved. In several cases interaction between the EO components and the organic modifiers led to increased MMT platelet stacking uniformity this effect tended to be most pronounced with o-MMTs containing dehydrogenated tallow functional organic modifiers (i.e., Cloisites 15A and 20A). The CAPA loaded RO and MNO showed antimicrobial activity against S. aureus. The adsorption (exchange) of silver ions into Cloisite Na+ was monitored using Energy dispersive X ray spectroscopy (EDX), Atomic Absorption Spectroscopy (AAS) and XRD. The highest level of silver incorporated was 10.4 wt.%. EDX was the most reliable method for determining the amount of silver adsorbed as it was carried out on washed and unwashed samples, the former provided the most reliable estimate of the amount of silver ions actually incorporated into the MMT galleries, rather than simply being adsorbed on the external surfaces. The figure of 10.4 wt.% was obtained from a washed sample. XRD data showed that treatment of the Na-MMT platelets with Ag+ aqueous solution, followed by drying led to a substantial disruption of stacking disorder. The Ag+ intercalated MMT showed antimicrobial activity against P. aeruginosa.