Hans Joerg Griesser oral presentation (OB4-Thu4-1-2)
ToF-SIMS Characterisation of Antimicrobial Coatings
1 FII - University of South Australia, Mawson Lakes Boulevard, SA 5095 Adelaide, Australia
2 School of Agriculture, Food & WIne - University of Adelaide, Waite Campus, 5001 Adelaide, Australia
The extensive literature on antimicrobial coatings contains results that can be surprising or contradictory. For example, there is considerable interest in covalently grafting antimicrobial molecules onto biomaterials surfaces, but few studies have rigorously investigated whether observed biological effects are indeed due to covalently grafted surface layers or, alternatively, arise from drug molecules that were physisorbed or diffused into a polymeric material, and then exerted antimicrobial effectiveness as solvated molecules diffusing into the microbial test environment. Our research groups have focused in a number of studies on questions surrounding covalently anchored graft coatings comprising antimicrobially active molecules (low molecular weight, 300-1200 Da) or antimicrobial polymers, and ToF-SIMS is the most important, and most revealing, analytical method for probing the detailed nature of intended antimicrobial coatings.
With appropriately designed controls, ToF-SIMS enables probing for the presence of bioactive molecules that may not be covalently grafted and can interfere in biological tests, which would lead to incorrect conclusions that a covalently attached layer of molecules is active. We have used a range of antimicrobial molecules, both commercial and experimental, for various designed experiments with various grafted layers. The molecules were immobilised onto plasma polymer interlayers with suitable reactive surface groups. Before and after various extensive washing protocols. Coatings were characterised by ToF-SIMS and by studying the antimicrobial activity of samples via cell attachment and biofilm formation assays.
Our results show that physisorption of antimicrobials occurs to substantial extents onto many substrates - even in the presence of reactive surface groups and catalysts, such as carbodiimide. Much larger amounts of molecules can also be absorbed diffusively into polymer substrates, again bypassing reactive surface groups. Surface analysis data before and after various washing procedures clearly demonstrate the presence of both covalently grafted and non-covalent molecules. When placed into bioassays, the latter molecules diffuse into solution and interfere with microbes. This can lead to erroneous interpretation that a covalently grafted layer has activity, as we demonstrated using a negative control, levofloxacin, which has an intracellular target and thus no activity when covalently grafted, yet diffusing molecules gave rise to activity. ToF-SIMS analyses in combination with extensive washing protocols – including severe conditions such as SDS and high temperature – are needed to ascertain covalent grafting.