Functionalization of ethylene vinyl acetate with antimicrobial chlorhexidine hexametaphosphate nanoparticles

Abstract

Ethylene vinyl acetate (EVA) is in widespread use as a polymeric biomaterial with diverse applications such as intravitreal devices, catheters, artificial organs, and mouthguards. Many biomaterials are inherently prone to bacterial colonization, as the human body is host to a vast array of microbes. This can lead to infection at the biomaterial's site of implantation or application. In this study, EVA was coated with chlorhexidine (CHX) hexametaphosphate (HMP) nanoparticles (NPs) precipitated using two different reagent concentrations: CHX-HMP-5 (5 mM CHX and HMP) and CHX-HMP-0.5 (0.5 mM CHX and HMP). Data gathered using dynamic light scattering, transmission electron microscopy, and atomic force microscopy indicated that the NPs were polydisperse, ~40-80 nm in diameter, and aggregated in solution to form clusters of ~140-200 nm and some much larger aggregates of 4-5 μM. CHX-HMP-5 formed large deposits on the polymer surface discernible using scanning electron microscopy, whereas CHX-HMP-0.5 did not. Soluble CHX was released by CHX-HMP-5 NP-coated surfaces over the experimental period of 56 days. CHX-HMP-5 NPs prevented growth of methicillin-resistant Staphylococcus aureus when applied to the polymer surfaces, and also inhibited or prevented growth of Pseudomonas aeruginosa with greater efficacy when the NP suspension was not rinsed from the polymer surface, providing a greater NP coverage. This approach may provide a useful means to treat medical devices fabricated from EVA to render them resistant to colonization by pathogenic microorganisms.

Keywords: EVA; biomaterial; polymer.

Figure 1

Transmission electron micrographs of CHX-HMP-5 (A) and CHX-HMP-0.5 (B) nanoparticles. Note: Scale bar 200 nm. Abbreviations: CHX, chlorhexidine; HMP, hexametaphosphate.

Figure 2

Dynamic light scattering data showing size distributions of CHX-HMP-5 (A) and CHX-HMP-0.5 (B) NPs, where CHX-HMP-X_1,2,3 indicate measurements in triplicate for concentration X. Abbreviations: CHX, chlorhexidine; HMP, hexametaphosphate; NPs, nanoparticles.

Figure 3

Zeta potential data showing charge distribution of CHX-HMP-5 (A) and CHX-HMP-0.5 (B) NPs where CHX-HMP-X_1,2,3 indicate measurements in triplicate for concentration X. Abbreviations: CHX, chlorhexidine; HMP, hexametaphosphate; NPs, nanoparticles.

Figure 4

Atomic force microscopy images and line plots of a single CHX-HMP NP (A, C) and an aggregate of NPs (B, D). In (D), the black line corresponds to profile 1 in (B), the red line to profile 2, and the green line to profile 3, each showing a feature thought to be a nanoparticle or small aggregate. The textured background in (B) is thought to be the polymer surface. Abbreviations: CHX, chlorhexidine; HMP, hexametaphosphate; NP, nanoparticle.

Figure 5

Scanning electron micrographs showing CHX-HMP nanoparticles on EVA polymer. (A, B) Control (no nanoparticles); (C, D) CHX-HMP-0.5 nanoparticles; (E, F) CHX-HMP-5 nanoparticles. The untreated EVA had a textured appearance (A, B). The CHX-HMP-0.5 surface was not clearly different from the control (C, D); there was no evidence of aggregations of nanoparticles as with the other material specimens. The CHX-HMP-5 surface displayed deposits of the porous aggregate coating much of the polymer surface (E, F). Abbreviations: CHX, chlorhexidine; HMP, hexametaphosphate; EVA, ethylene vinyl acetate.

Figure 6

Chlorhexidine release from nanoparticle-functionalized EVA polymer expressed in μmoles CHX released per unit surface area of specimen as a function of time. The EVA functionalized with CHX-HMP-5 nanoparticles showed a sustained release of soluble CHX which was still ongoing at the end of the experimental period. There was a lower release from the CHX-HMP-0.5 specimens but this was almost all within the first day or two; there was little or no release after this time. The control specimens treated with 25 μM aqueous CHX solution did not show a release of CHX. Abbreviations: CHX, chlorhexidine; HMP, hexametaphosphate; EVA, ethylene vinyl acetate.