(M-499) Enhanced Antibacterial Efficacy Against Antibiotic-Resistant Bacteria via Nitric Oxide-Releasing Ampicillin Polymer Substrates

Introduction:: Despite decades of advancements in biomaterials and medical device surface design, indwelling device-related infection remains a significant contributing factor to healthcare-associated infections. Nitric oxide (NO) is a small molecule gasotransmitter produced by macrophages and has a wide range of biological roles in a dose-dependent manner, including antibacterial characteristics. Among NO donors, S-nitroso-N-acetylpenicillamine (SNAP), a synthetic tertiary RSNO, has been studied extensively due to its stability.

Herein, we report the fabrication of medical-grade polymers incorporated with a dual-action S-nitroso-N-acetylpenicillamine-functionalized ampicillin (SNAPicillin) conjugated through a solvent evaporation process. The resulting SNAPicillin-modified materials act as broad-spectrum antibacterial surfaces that improve the administration efficacy of conventional antibiotics through the targeted release of both NO and ampicillin.


Materials and Methods:: Conjugate of ampicillin with the NO donor SNAP was prepared through the aminolysis of a self-protected N-acetyl penicillamine thiolactone. Polymers containing 10 wt% SNAPicillin were prepared using a solvent evaporation method. The polymer surfaces were characterized by scanning electron microscopy and static contact angle measurements. The NO release profile and diffusion of SNAPicillin from polymers were quantified using a chemiluminescence-based nitric oxide analyzer and UV-vis spectroscopy. A standard 24 h leachate exposure procedure was used to quantify the cytotoxicity of the films against mouse fibroblasts to assess the biocompatibility of these NO-releasing materials. Finally, the antimicrobial efficacy of SNAPicillin blended ChronoSil and SG80A was evaluated through a 24 h viable bacterial adhesion assay against β-lactam antibiotic-resistant Escherichia coli (E. coli), methicillin-resistant Staphylococcus aureus (MRSA), and Staphylococcus aureus (S. aureus).


Results, Conclusions, and Discussions:: The majority of the SNAPicillin reservoir was preserved for SNAPicillin-incorporated polymers after 24 h of incubation in the physiological environment (37 °C), indicating their longer-term NO release ability and therapeutic window for antibacterial effects compared to other tested polymers. SNAPicillin-SG80A has the highest total NO released of 8.61 ± 1.04 × 10^-9 mol cm^-2 within 24 h. Our results suggested that the intrapolymeric decomposition of SNAPicillin and its subsequent bactericidal activities can be tuned by using polymer with different water uptake and SNAPicillin wt% composition formulation.

SNAPicillin film leachates did not elicit a cytotoxic effect toward 3T3 mouse fibroblast cells, supporting the material's biocompatibility in vitro. Furthermore, the complementary effect of SNAPicillin-ChronoSil and SNAPicillin-SG80A's prolonged and stable SNAPicillin release can help prevent undesired clinical infection post-implantation of polymeric medical devices. The SNAPicillin-incorporated polymers were able to reduce the viability of adhered bacteria in culture, as shown in Figure 1. These results indicate that the complementary effects of NO-release along with ampicillin in SNAPicillin eluting polymers can be used to enhance the properties of commonly infected medical device surfaces for antibacterial purposes.


The incorporation of SNAPicillin within polymer films/coatings provides targeted delivery of NO and ampicillin, avoiding complications from systemic administration of antibiotics ranging from gastrointestinal symptoms to antibiotic-resistance development. For future studies, SNAPicillin-incorporated materials offer the potential to improve the antibacterial functionality of biomedical devices for many applications.




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