Study of Crystal Formation and Nitric Oxide (NO) Release Mechanism from S-Nitroso-N-acetylpenicillamine (SNAP)-Doped CarboSil Polymer Composites for Potential Antimicrobial Applications.

Stable and long-term nitric oxide (NO) releasing polymeric materials have many potential biomedical applications. Herein, we report the real-time observation of the crystallization process of the NO donor, S-nitroso-N-acetylpenicillamine (SNAP), within a thermoplastic silicone-polycarbonate-urethane biomedical polymer, CarboSil 20 80A. It is demonstrated that the NO release rate from this composite material is directly correlated with the surface area that the CarboSil polymer film is exposed to when in contact with aqueous solution. The decomposition of SNAP in solution (e.g. PBS, ethanol, THF, etc.) is a pseudo-first-order reaction proportional to the SNAP concentration. Further, catheters fabricated with this novel NO releasing composite material are shown to exhibit significant effects on preventing biofilm formation on catheter surface by Pseudomonas aeruginosa and Proteus mirabilis grown in CDC bioreactor over 14 days, with a 2 and 3 log-unit reduction in number of live bacteria on their surfaces, respectively. Therefore, the SNAP-CarboSil composite is a promising new material to develop antimicrobial catheters, as well as other biomedical devices.

Antimicrobial nitric oxide releasing surfaces based on S-nitroso-N-acetylpenicillamine impregnated polymers combined with submicron-textured surface topography.

A novel dual functioning antimicrobial CarboSil 20 80A polymer material that combines physical topographical surface modification and nitric oxide (NO) release is prepared and evaluated for its efficacy in reducing bacterial adhesion in vitro. The new biomaterial is created via a soft lithography two-stage replication process to induce submicron textures on its surface, followed by solvent impregnation with the NO donor, S-nitroso-N-acetylpenicillamine (SNAP), to obtain long-term (up to 38 d) NO release. The NO releasing textured polymer surface is evaluated against four bacteria commonly known to cause infections in hospital settings and the results demonstrate that the combined strategy enables a synergistic effect on reducing the bacterial adhesion of Staphylococcus epidermidis and Pseudomonas aeruginosa bacteria.

Reduction of Thrombosis and Bacterial Infection via Controlled Nitric Oxide (NO) Release from S-Nitroso-N-acetylpenicillamine (SNAP) Impregnated CarboSil Intravascular Catheters.

Nitric oxide (NO) has many important physiological functions, including its ability to inhibit platelet activation and serve as potent antimicrobial agent. The multiple roles of NO in vivo have led to great interest in the development of biomaterials that can deliver NO for specific biomedical applications. Herein, we report a simple solvent impregnation technique to incorporate a nontoxic NO donor, S-nitroso-N-acetylpenicillamine (SNAP), into a more biocompatible biomedical grade polymer, CarboSil 20 80A. The resulting polymer-crystal composite material yields a very stable, long-term NO release biomaterial. The SNAP impregnation process is carefully characterized and optimized, and it is shown that SNAP crystal formation occurs in the bulk of the polymer after solvent evaporation. LC-MS results demonstrate that more than 70% of NO release from this new composite material originates from the SNAP embedded CarboSil phase, and not from the SNAP species leaching out into the soaking solution. Catheters prepared with CarboSil and then impregnated with 15 wt % SNAP provide a controlled NO release over a 14 d period at physiologically relevant fluxes and are shown to significantly reduce long-term (14 day) bacterial biofilm formation against Staphylococcus epidermidis and Pseudonomas aeruginosa in a CDC bioreactor model. After 7 h of catheter implantation in the jugular veins of rabbit, the SNAP CarboSil catheters exhibit a 96% reduction in thrombus area (0.03 ± 0.01 cm2/catheter) compared to the controls (0.84 ± 0.19 cm2/catheter) (n = 3). These results suggest that SNAP impregnated CarboSil can become an attractive new biomaterial for use in preparing intravascular catheters and other implanted medical devices.

Inhibition of bacterial adhesion and biofilm formation by dual functional textured and nitric oxide releasing surfaces.

In separate prior studies, physical topographic surface modification or nitric oxide (NO) release has been demonstrated to each be an effective approach to inhibit and control bacterial adhesion and biofilm formation on polymeric surfaces. Such approaches can prevent biomaterial-associated infection without causing the antibiotic resistance of the strain. In this work, both techniques were successfully integrated and applied to a polyurethane (PU) biomaterial surface that bears ordered pillar topographies (400/400nm and 500/500nm patterns) at the top surface and a S-nitroso-N-acetylpenicillamine (SNAP, NO donor) doped sub-layer in the middle, via a soft lithography two-stage replication process. Upon placing the SNAP textured PU films into PBS at 37°C, the decomposition of SNAP within polymer film initiates NO release with a lifetime of up to 10days at flux levels >0.5×10-10molmin-1cm-2 for a textured polyurethane layer containing 15wt% SNAP. The textured surface reduces the accessible surface area and the opportunity of bacteria-surface interaction, while the NO release from the same surface further inhibits bacterial growth and biofilm formation. Such dual functionality surfaces are shown to provide a synergistic effect on inhibition of Staphylococcus epidermidis bacterial adhesion that is significantly greater than the inhibition of bacterial adhesion achieved by either single treatment approach alone. Longer term experiments to observe biofilm formation demonstrate that the SNAP doped-textured PU surface can inhibit the biofilm formation for >28d and provide a practical approach to improve the biocompatibility of current biomimetic biomaterials and thereby reduce the risk of pathogenic infection. STATEMENT OF SIGNIFICANCE: Microbial infection remains a significant barrier to development and implementation of advanced blood-contacting medical devices. Clearly, determining how to design and control material properties that can reduce microbial infection is a central question to biomaterial researchers. In separate prior studies, physical topographic surface modification or nitric oxide (NO) release has been demonstrated to each be an effective approach to inhibit and control bacterial adhesion and biofilm formation on polymeric surfaces. Such approaches can prevent biomaterial-associated infection without causing antibiotic resistance of the bacterial strain. However, efficiency of antimicrobial properties of each approach is still limited and far from sufficient for widespread clinical use. This work successfully integrates both techniques and applies them to a polyurethane (PU) biomaterial surface that bears dual functions, surface topographic modification and NO release. The former reduces the surface contact area and changes surface wettability, resulting in reduction of bacterial adhesion, and NO release further inhibits bacteria growth. Such dual functionalized surfaces provide a synergistic effect on inhibition of Staphylococcus epidermidis bacterial adhesion that is significantly greater than the inhibition of bacterial adhesion achieved by either single treatment approach alone. Furthermore, longer-term experiments demonstrate that the dual functionalized surfaces can inhibit biofilm formation for >28days. The success of this work provides a practical approach to improve the biocompatibility of current biomaterials and thereby reduce the risk of pathogenic infection.

Recent advances in thromboresistant and antimicrobial polymers for biomedical applications: just say yes to nitric oxide (NO).

Biomedical devices are essential for patient diagnosis and treatment; however, when blood comes in contact with foreign surfaces or homeostasis is disrupted, complications including thrombus formation and bacterial infections can interrupt device functionality, causing false readings and/or shorten device lifetime. Here, we review some of the current approaches for developing antithrombotic and antibacterial materials for biomedical applications. Special emphasis is given to materials that release or generate low levels of nitric oxide (NO). Nitric oxide is an endogenous gas molecule that can inhibit platelet activation as well as bacterial proliferation and adhesion. Various NO delivery vehicles have been developed to improve NO's therapeutic potential. In this review, we provide a summary of the NO releasing and NO generating polymeric materials developed to date, with a focus on the chemistry of different NO donors, the polymer preparation processes, and in vitro and in vivo applications of the two most promising types of NO donors studied thus far, N-diazeniumdiolates (NONOates) and S-nitrosothiols (RSNOs).

S-Nitroso-N-acetylpenicillamine (SNAP) Impregnated Silicone Foley Catheters: A Potential Biomaterial/Device To Prevent Catheter-Associated Urinary Tract Infections.

Urinary Foley catheters are utilized for management of hospitalized patients and are associated with high rates of urinary tract infections (UTIs). Nitric oxide (NO) potently inhibits microbial biofilm formation, which is the primary cause of catheter associated UTIs (CAUTIs). Herein, commercial silicone Foley catheters are impregnated via a solvent swelling method with S-nitroso-N-acetyl-D-penicillamine (SNAP), a synthetic NO donor that exhibits long-term NO release and stability when incorporated into low water-uptake polymers. The proposed catheters generate NO surface-fluxes >0.7 × 10-10 mol min-1 cm-2 for over one month under physiological conditions, with minimal SNAP leaching. These biomedical devices are demonstrated to significantly decrease formation of biofilm on the surface of the catheter tubings over 3, 7, and 14 day periods by microbial species (Staphylococcus epidermidis and Proteus mirabilis) commonly causing CAUTIs. Toxicity assessment demonstrates that the SNAP-impregnated catheters are fully biocompatible, as extracts of the catheter tubings score 0 on a 3-point grading scale using an accepted mouse fibroblast cell-line toxicity model. Consequently, SNAP-impregnated silicone Foley catheters can likely provide an efficient strategy to greatly reduce the occurrence of nosocomial CAUTIs.

Origin of Long-Term Storage Stability and Nitric Oxide Release Behavior of CarboSil Polymer Doped with S-Nitroso-N-acetyl-D-penicillamine.

The prolonged and localized delivery of nitric oxide (NO), a potent antithrombotic and antimicrobial agent, has many potential biomedical applications. In this work, the origin of the long-term storage stability and sustained NO release mechanism of S-nitroso-N-acetyl-D-penicillamine (SNAP)-doped CarboSil 20 80A polymer, a biomedical thermoplastic silicone-polycarbonate-urethane, is explored. Long-term (22 days) localized NO release is achieved by utilizing a cross-linked silicone rubber as topcoats, which can greatly reduce the amount of SNAP, NAP, and NAP disulfide leaching from the SNAP-doped CarboSil films, as measured by LC-MS. Raman spectroscopy and powder X-ray diffraction characterization of SNAP-doped CarboSil films demonstrate that a polymer-crystal composite is formed during the solvent evaporation process when SNAP exceeds its solubility in CarboSil (ca. 3.4-4.0 wt %). Further, when exceeding this solubility threshold, SNAP exists in an orthorhombic crystal form within the bulk of the polymer. The proposed mechanism of sustained NO release in SNAP-doped CarboSil is that the solubilized SNAP in the polymer matrix decomposes and releases NO, primarily in the water-rich regions near the polymer/solution interface, and the dissolved SNAP in the bulk polymeric phase becomes unsaturated, resulting in the dissolution of crystalline SNAP within the bulk of the polymer. This is a very slow process that ultimately leads to NO release at the physiological flux levels for >3 weeks. The increased stability of SNAP within CarboSil is attributed to the intermolecular hydrogen bonds between the SNAP molecules that crystallize. This crystallization also plays a key role in maintaining RSNO stability within the CarboSil polymer for >8 months at 37 °C (88.5% remains). Further, intravascular catheters fabricated with this new material are demonstrated to significantly decrease the formation of Staphylococcus aureus biofilm (a leading cause of nosocomial bloodstream infections) (in vitro) over a 7 day period, with 5 log units reduction of viable cell count on catheter surfaces. It is also shown that the NO release catheters can greatly reduce thrombus formation on the catheter surfaces during 7 h implantation in rabbit veins, when compared to the control catheters fabricated without SNAP. These results suggest that the SNAP-doped CarboSil system is a very attractive new composite material for creating long-term NO release medical devices with increased stability and biocompatibility.

Polymeric optical sensors for selective and sensitive nitrite detection using cobalt(III) corrole and rhodium(III) porphyrin as ionophores.

Cobalt(III) 5,10,15-tris(4-tert-butylphenyl) corrole with a triphenylphosphine axial ligand and rhodium(III) 5,10,15,20-tetra(p-tert-butylphenyl) porphyrin are incorporated into plasticized poly(vinyl chloride) films to fabricate nitrite-selective bulk optodes via absorbance measurements. The resulting films yield sensitive, fast and fully reversible response toward nitrite with significantly enhanced nitrite selectivity over other anions including lipophilic anions such as thiocyanate and perchlorate. The selectivity patterns differ greatly from the Hofmeister series based on anion lipophilicity and are consistent with selectivity obtained with potentiometric sensors based on the same ionophores. The optical nitrite sensors are shown to be useful for detecting rates of emission of nitric oxide (NO) from NO releasing polymers containing S-nitroso-N-acetyl-DL-penicillamine.