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This study presents the utilization of a novel, highly lipophilic nitric oxide (NO) donor molecule, S-nitroso-1-adamantanethiol (SNAT), for developing an NO-emitting polymer surface aimed at preventing thrombus formation and bacterial infection in extracorporeal circuits (ECCs). S-nitroso-1-adamantanethiol, a tertiary nitrosothiol-bearing adamantane species, was synthesized, characterized, and used to impregnate polyvinyl chloride (PVC) tubing for subsequent in vivo evaluation. The impregnation process with SNAT preserved the original mechanical strength of the PVC. In vitro assessments revealed sustained NO release from the SNAT-impregnated PVC tubing (iSNAT), surpassing or matching endothelial NO release levels for up to 42 days. The initial NO release remained stable even after 1 year of storage at -20°C. The compatibility of iSNAT with various sterilization techniques (OPA Plus, hydrogen peroxide, EtO) was tested. Acute in vivo experiments in a rabbit model demonstrated significantly reduced thrombus formation in iSNAT ECCs compared with controls, indicating the feasibility of iSNAT to mitigate coagulation system activation and potentially eliminate the need for systemic anticoagulation. Moreover, iSNAT showed substantial inhibition of microbial biofilm formation, highlighting its dual functionality. These findings underscore the promising utility of iSNAT for long-term ECC applications, offering a multifaceted approach to enhancing biocompatibility and minimizing complications., Competing Interests: Disclosure: O.L.C., R.G., R.H.B., S.P.S., G.L., and M.E.M. are inventors on US Provisional Patent PCT/US2024/031413 which is related to this work. The other authors have no conflicts of interest to report., (Copyright © ASAIO 2024.)

S-Nitroso-N-acetylpenicillamine (SNAP) is among the most common nitric oxide (NO)-donor molecules and its solid-state photolytic decomposition has potential for inhaled nitric oxide (iNO) therapy. The photochemical NO release kinetics and mechanism were investigated by exposing solid-state SNAP to a narrow-band LED as a function of nominal wavelength and intensity of incident light. The photolytic efficiency, decomposition products, and the photolytic pathways of the SNAP were examined. The maximum light penetration depth through the solid layer of SNAP was determined by an optical microscope and found to be within 100-200 μm, depending on the wavelength of light. The photolysis of solid-state SNAP to generate NO along with the stable thiyl (RS·) radical was confirmed using Electron Spin Resonance (ESR) spectroscopy. The fate of the RS· radical in the solid phase was studied both in the presence and absence of O 2 using NMR, IR, ESR, and UPLC-MS. The changes in the morphology of SNAP due to its photolysis were examined using PXRD and SEM. The stable thiyl radical formed from the photolysis of solid SNAP was found to be reactive with another adjacent thiyl radical to form a disulfide (RSSR) or with oxygen to form various sulfonyl and sulfonyl peroxyl radicals {RS(O) x O·, x = 0 to 7}. However, the thiyl radical did not recombine with NO to reform the SNAP. From the PXRD data, it was found that the SNAP loses its crystallinity by generating the NO after photolysis. The initial release of NO during photolysis was increased with increased intensity of light, whereas the maximum light penetration depth was unaffected by light intensity. The knowledge gained about the photochemical reactions of SNAP may provide important insight in designing portable photoinduced NO-releasing devices for iNO therapy., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Background: Clinical translation of the extracorporeal artificial placenta (AP) is impeded by the high risk for intracranial hemorrhage in extremely premature newborns. The Nitric Oxide Surface Anticoagulation (NOSA) system is a novel non-thrombogenic extracorporeal circuit. This study aims to test the NOSA system in the AP without systemic anticoagulation., Methods: Ten extremely premature lambs were delivered and connected to the AP. For the NOSA group, the circuit was coated with DBHD-N 2 O 2 /argatroban, 100 ppm nitric oxide was blended into the sweep gas, and no systemic anticoagulation was given. For the Heparin control group, a non-coated circuit was used and systemic anticoagulation was administered., Results: Animals survived 6.8 ± 0.6 days with normal hemodynamics and gas exchange. Neither group had any hemorrhagic or thrombotic complications. ACT (194 ± 53 vs. 261 ± 86 s; p < 0.001) and aPTT (39 ± 7 vs. 69 ± 23 s; p < 0.001) were significantly lower in the NOSA group than the Heparin group. Platelet and leukocyte activation did not differ significantly from baseline in the NOSA group. Methemoglobin was 3.2 ± 1.1% in the NOSA group compared to 1.6 ± 0.6% in the Heparin group (p < 0.001)., Conclusions: The AP with the NOSA system successfully supported extremely premature lambs for 7 days without significant bleeding or thrombosis., Impact: The Nitric Oxide Surface Anticoagulation (NOSA) system provides effective circuit-based anticoagulation in a fetal sheep model of the extracorporeal artificial placenta (AP) for 7 days. The NOSA system is the first non-thrombogenic circuit to consistently obviate the need for systemic anticoagulation in an extracorporeal circuit for up to 7 days. The NOSA system may allow the AP to be implemented clinically without systemic anticoagulation, thus greatly reducing the intracranial hemorrhage risk for extremely low gestational age newborns. The NOSA system could potentially be applied to any form of extracorporeal life support to reduce or avoid systemic anticoagulation., (© 2023. The Author(s), under exclusive licence to the International Pediatric Research Foundation, Inc.)

Currently, the controlled release of nitric oxide (NO) plays a crucial role in various biomedical applications. However, injectable NO-releasing materials remain an underexplored research field to date. In this study, via the incorporation of S-nitroso-N-acetyl-penicillamine (SNAP) as an NO donor, a family of NO-releasing injectable hydrogels was synthesized through the in situ cross-linking between sodium alginate and calcium ion induced by D-(+)-gluconate δ-lactone as an initiator. Initially, the organic functional groups and the corresponding morphologies of the resulting injectable hydrogels were characterized by IR and SEM spectroscopies, respectively. The NO release times of hydrogels with different SNAP loading amounts could reach up to 36-47 h. Due to the release of NO, the highest antibacterial rates of these injectable hydrogels against Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus) were up to 95 %, respectively. Furthermore, the matrix of these hydrogels demonstrated great water absorption ability, swelling behavior, and degradation performance. Finally, we expect that these NO-releasing injectable hydrogels could have great potential applications various biomedical material fields., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier B.V.)

Electrocatalytic nitric oxide (NO) generation from nitrite (NO 2 - ) within a single lumen of a dual-lumen catheter using Cu II -ligand (Cu II -L) mediators have been successful at demonstrating NO's potent antimicrobial and antithrombotic properties to reduce bacterial counts and mitigate clotting under low oxygen conditions (e.g., venous blood). Under more aerobic conditions, the O 2 sensitivity of the Cu(II)-ligand catalysts and the reaction of O 2 (highly soluble in the catheter material) with the NO diffusing through the outer walls of the catheters results in a large decreases in NO fluxes from the surfaces of the catheters, reducing the utility of this approach. Herein, we describe a new more O 2 -tolerant Cu II -L catalyst, [Cu(BEPA-EtSO 3 )(OTf)], as well as a potentially useful immobilized glucose oxidase enzyme-coating approach that greatly reduces the NO reactivity with oxygen as the NO partitions and diffuses through the catheter material. Results from this work demonstrate that very effective NO fluxes (>1*10 -10 mol min -1 cm -2 ) from a single-lumen silicone rubber catheter can be achieved in the presence of up to 10% O 2 saturated solutions., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Patients with chronic rhinosinusitis (CRS) often show persistent colonization by bacteria in the form of biofilms which are resistant to antibiotic treatment. One of the most commonly isolated bacteria in CRS is Staphylococcus aureus ( S. aureus ). Nitric oxide (NO) is a potent antimicrobial agent and disperses biofilms efficiently. We hypothesized that S -nitrosoglutathione (GSNO), an endogenous NO carrier/donor, synergizes with gentamicin to disperse and reduce the bacterial biofilm density. We prepared GSNO formulations which are stable up to 12 months at room temperature and show the maximum amount of NO release within 1 h. We examined the effects of this GSNO formulation on the S. aureus biofilm established on the apical surface of the mucociliary-differentiated airway epithelial cell cultures regenerated from airway basal (stem) cells from cystic fibrosis (CF) and CRS patients. We demonstrate that for CF cells, which are defective in producing NO, treatment with GSNO at 100 μM increased the NO levels on the apical surface and reduced the biofilm bacterial density by 2 log units without stimulating pro-inflammatory effects or inducing epithelial cell death. In combination with gentamicin, GSNO further enhanced the killing of biofilm bacteria. Compared to placebo, GSNO significantly increased the ciliary beat frequency (CBF) in both infected and uninfected CF cell cultures. The combination of GSNO and gentamicin also reduced the bacterial density of biofilms grown on sinonasal epithelial cells from CRS patients and improved the CBF. These findings demonstrate that GSNO in combination with gentamicin may effectively reduce the density of biofilm bacteria in CRS patients. GSNO treatment may also enhance the mucociliary clearance by improving the CBF., Competing Interests: The authors declare the following competing financial interest(s): AW and GM are employees of the NOTA Labs MEM serves as Chief Technology Officer for NOTA Labs MZ, MBH, and US are scientific officers for NOTA Labs, (© 2022 The Authors. Published by American Chemical Society.)

Over the past 30 years, the significance of nitric oxide (NO) has become increasingly apparent in mammalian physiology. It is biosynthesized by three isoforms of nitric oxide synthases (NOS): neuronal (nNOS), endothelial (eNOS), and inducible (iNOS). Neuronal and eNOS both produce low levels of NO (nM) as a signaling agent and vasodilator, respectively. Inducible (iNOS) is present in activated macrophages at sites of infection to generate acutely toxic (μM) levels of NO as part of the mammalian immune defense mechanism. These discoveries have led to numerous animal and clinical studies to evaluate the potential therapeutic utility of NO in various medical operations/treatments, primarily using NO gas (via gas-cylinders) as the NO source. In this review, we focus specifically on recent advances in the electrochemical generation of NO (E-NOgen) as an alternative means to generate NO from cheap and inert sources, and the fabrication and testing of biomedical devices that utilize E-NOgen to controllably generate NO for medical applications.

Although there are many techniques to detect pathogenic bacteria, most of them are only suited for in vitro diagnostics. We report a urinary catheter-based colorimetric sensor for potential on-body detection of E. coli , the most prevalent bacterial species in urinary tract infections associated with the use of urinary catheters. In urine, indole is secreted by E. coli and reacts with a nitrosating agent incorporated in a silicone catheter. A red dimeric product, indoxyl red, is generated within silicone rubber to allow for color-based indole sensing with high sensitivity, linearity, and specificity. This reaction is initiated by the nitrosation reaction of indole at its C-3 position via reagents like sodium nitrite or S -nitroso- N -acetyl-penicillamine under aerobic conditions. The generated 3-nitrosoindole undergoes tautomerization, dimerization, and deoximation to form indoxyl red with high absorbance at 537 nm. In contrast to other indole sensors, the presented method can be applied in real catheters to detect indole and E. coli in biofluids such as urine. The is because (1) S -nitroso- N -acetyl-penicillamine, the nitrosating agent, can be impregnated into silicone elastomers, (2) indole from urine is extracted into silicone due to its hydrophobicity, and (3) the high acidity and oxygen solubility of silicone facilitates the sensing reaction within the silicone matrix. This silicone-based colorimetric sensor clearly differentiates E. coli below and above 10 5 CFU/mL, which is the threshold concentration of bacteriuria. We expect that early diagnosis of urinary tract infections using the naked eye is possible by functionalizing an exposed section of urinary catheters with the proposed molecular probe.

Droplet microfluidic optodes, or "droptodes", have emerged as a powerful technology for rapid detection of small ions in complex matrices. While using segmented aqueous phases provides the benefits of sample isolation, the influence of the liquid nature of the oil carrier phase has not yet been explored. In this paper, we examine the influence of microfluidic parameters on droptode efficiency, using potassium-sensitive droptodes as a model system. We found that while changing flow rates on device does not change droptode performance, both channel geometry and droplet size significantly impact droptode efficiency. Specifically, enhanced mixing of the droplets leads to faster equilibration on device and lowers limits of detection by about one order of magnitude. We also found that increasing the size of the sample droplet, at the expense of the size of the oil carrier/sensing phase, leads to higher sensitivity in the linear region of the droptode. These easily manipulated properties will allow one device to potentially be adapted for several different applications, based upon the type and concentration range of measurement required.

Clotting, anticoagulation, platelet consumption, and poor platelet function are major factors in clinical extracorporeal circulation (ECC). We have shown that nitric oxide-releasing (NOReL) coatings prevent thrombosis in a rabbit model of ECC without systemic anticoagulation. Nitric oxide-releasing prevents platelet adhesion and activation, resulting in preserved platelet count and function. Previous work has shown that activated platelets form platelet-derived microparticles (PMPs). These experiments were designed to determine if PMPs can identify platelet function during ECC. The objective of this study is to investigate the effects of NOReL on platelet activation and PMP formation during ECC. Uncoated ECCs, including with and without systemic heparin, and NOReL-coated ECCs, including DBHD/N2O2 and argatroban (AG)/DBHD/N2O2-coated ECCs without systemic heparin, were tested in a 4-hour rabbit thrombogenicity model. Before and after ECC exposure, platelets were stimulated with collagen, and PMPs were measured using flow cytometry. The uncoated ECCs clotted within the first hour, while the NOReL-coated ECCs circulated for 4 hours. During pre-ECC blood exposure, platelets stimulated with collagen produced PMPs. With post-ECC exposure, platelets from uncoated circuits generated less PMPs than baseline (mean ± SDs: 23246 ± 3611 baseline vs. 1300 ± 523 uncoated post circuit, p = 0.018) when stimulated with collagen. However, platelets from the AG/DBHD/N2O2-coated ECCs generated a greater number of PMPs as baseline values (23246 ± 3611 baseline vs. 37040 ± 3263 AG/DBHD/N2O2 post 4 hours circuit, p = 0.023). Blood exposure during ECC results in platelet activation and clotting in uncoated ECCs. The remaining circulating platelets have lost function, as demonstrated by the low PMP formation in response to collagen. AG/DBHD/N2O2-coated ECCs prevented significant platelet activation and clotting, while DBHD/N2O2 trended towards prevention of platelet activation. In addition, function of the circulating platelets was preserved, as demonstrated by PMP formation in response to collagen. These results indicate that PMPs may be an important measure of platelet activation during ECC. Platelet-derived microparticles may provide a simplified way to measure platelet function during clinical ECC., Competing Interests: Disclosure: The authors have no conflicts of interest to report., (Copyright © ASAIO 2020.)

In this letter, the innate ability of nitric oxide (NO) to inhibit platelet activation/adhesion/thrombus formation is employed to improve the hemocompatibility and in vivo accuracy of an intravascular (IV) potentiometric P CO 2 (partial pressure of carbon dioxide) sensor. The catheter-type sensor is fabricated by impregnating a segment of dual lumen silicone tubing with a proton ionophore, plasticizer, and lipophilic cation-exchanger. Subsequent filling of bicarbonate and strong buffer solutions and placement of Ag/AgCl reference electrode wires within each lumen, respectively, enables measurement of the membrane potential difference across the inner wall of the tube, with this potential changing as a function of the logarithm of sample P CO 2 . The dual lumen device is further encapsulated within a S -nitroso- N -acetyl-DL-penicillamine (SNAP)-doped silicone tube that releases physiological levels of NO. The NO releasing sensor exhibits near-Nernstian sensitivity toward P CO 2 (slope = 59.31 ± 0.78 mV/decade) and low drift rates (<2 mV/24 h after initial equilibration). In vivo evaluation of the NO releasing sensors, performed in the arteries and veins of anesthetized pigs for 20 h, shows enhanced accuracy (vs non-NO releasing sensors) when benchmarked to measurements of discrete blood samples made with a commercial blood gas analyzer. The accurate, continuous monitoring of blood P CO 2 levels achieved with this new IV NO releasing P CO 2 sensor configuration could help better manage hospitalized patients in critical care units.

The use of tunneled dialysis catheters (TDCs) for patients in need of hemodialysis treatments (HDs) causes a significant number of bloodstream infections (BSIs), with very few viable preventative/treatment methods. Use of antibiotics is relatively ineffective due to the development of multidrug-resistant bacterial strains and the inability to penetrate bacterial biofilms. Nitric oxide (NO) is an endogenous gas molecule that has broad-spectrum antimicrobial/antibiofilm activity. In this study, the potential of creating a NO-releasing insert device that is attached onto the hub region cap of TDCs and locally releases NO within the TDC hub is evaluated for its antimicrobial/antibiofilm effectiveness. The NO-releasing insert contains the natural NO donor S -nitrosoglutathione (GSNO), along with zinc oxide (ZnO) nanoparticles to accelerate NO release from the GSNO, within a short silicone tube that is sealed at both ends and attached to the catheter cap. An in vitro 3-d-long antimicrobial study using catheter hubs yielded >6.6 log reductions of both Pseudomonas aeruginosa and Staphylococcus aureus for the NO-releasing insert device compared to controls. Two 14-d-long sheep studies demonstrated that the NO-releasing insert devices are exceptionally potent at preventing bacteria/biofilm growth on the inner lumen walls of TDCs compared to controls that have no preventative treatment devices as well as implanted TDCs that have commercially available chlorhexidine-treated insert devices placed within the hub regions.

A novel strategy to functionalize transparent flexible plastic films with an optical ion-sensing layer using an inkjet-printing technology is described. The hydrophobic sensing chemicals that include a sodium ionophore, a lipophilic proton chromoionophore, and a lipophilic ion-exchanger are co-deposited onto substrates such as transparent polyester film sheets in the absence of any plasticizer and/or hydrophobic polymer matrix. The inkjet-printing process enables the formation of optode films with nanoscale thickness/roughness that readily facilitate interfacing with aqueous samples. Using a smartphone detector, the colorimetric response of the optodes is shown to reach 95% of equilibrium values within 100 s in response to different concentrations of sodium ions, which is more rapid than traditional ion-selective optodes based on plasticized PVC films as the sensing layer. The new optodes also exhibit high selectivity to Na + over interfering ions including K + , Ca 2+ , and Mg 2+ . Chemical leaching experiments show that the highly hydrophobic optode components remain in place on the plastic substrate surface. Hence, excellent sensor stability and fully reversible optical responses are obtained, which is essential for potential continuous monitoring applications. Further testing of the sensors with undiluted human sweat samples is shown to yield accurate values for sodium concentrations. Therefore, the use of plasticizer-free ion-selective optode nanolayers that enable highly selective ion sensing on a clear plastic support is likely to expand the range of available chemical sensors suited for preparing wearable real-time sweat analysis devices.

The blood compatibility of various intravascular (IV) devices (e.g., catheters, sensors, etc.) is compromised by activation of platelets that can cause thrombus formation and device failure. Such devices also carry a high risk of microbial infection. Recently, nitric oxide (NO) releasing polymers/devices have been proposed to reduce these clinical problems. CD47, a ubiquitously expressed transmembrane protein with proven anti-inflammation/anti-platelet properties when immobilized on polymeric surfaces, is a good candidate to complement NO release in both effectiveness and longevity. In this work, we successfully appended CD47 peptides (pepCD47) to the surface of biomedical grade polyurethane (PU) copolymers. SIRPα binding and THP-1 cell attachment experiments strongly suggested that the pepCD47 retains its biological properties when bound to PU films. In spite of the potentially high reactivity of NO toward various amino acid residues in CD47, the efficacy of surface-immobilized pepCD47 to prevent inflammatory cell attachment was not inhibited after being subjected to a high flux of NO for three days, demonstrating excellent compatibility of the two species. We further constructed a CD47 surface immobilized silicone tubing filled with NO releasing S-nitrosoglutathione/ascorbic acid (GSNO/AA) solution for synergistic biocompatibility evaluation. Via an ex vivo Chandler loop model, we demonstrate for the first time that NO release and CD47 modification could function synergistically at the blood/material interface and produce greatly enhanced anti-inflammatory/anti-platelet effects. This concept should be readily implementable to create a new generation of thromboresistant/antimicrobial implantable devices., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Constant therapeutic gas phase nitric oxide (NO) delivery is achieved from S-nitrosothiol (RSNO) type NO donor doped silicone rubber films using feedback-controlled photolysis. For photo-release of the NO gas, the intensity of the LED light source is controlled via a PID (proportional-integral-derivative) controller implemented on a microcontroller. The NO concentration within the emitted gas phase is monitored continuously with a commercial amperometric NO gas sensor. NO release was accurately adjustable up to 10 ppm across a broad range of setpoints with response times of roughly 1 min or less. When NO is generated into an air recipient stream, lower NO yields and a comparable level of toxic nitrogen dioxide (NO 2 ) formation is observed. However, NO gas generated into an N 2 recipient gas stream can be blended into pure O 2 with very low NO 2 formation. Following scale-up, this technology could be used for point-of-care gas phase NO generation as an alternative for currently used gas cylinder technology for treatment of health conditions where inhaled NO is beneficial, such as pulmonary hypertension, hypoxemia, and cystic fibrosis., (Copyright © 2019. Published by Elsevier B.V.)

When blood from a patient is circulated through extracorporeal circuits (ECCs), such as in cardiopulmonary bypass or extracorporeal life support, platelets in the blood are activated and form a thrombus. This is prevented clinically with a range of different systemic anticoagulation agents (e.g., heparin); however, this increases a patient's risk of hemorrhage. Previous work with nitric oxide (NO) releasing materials using the combined diazeniumdiolated diamine, N - N -di- N '-butyl-1,6-hexanediamine (DBHD), and a polymer-linked thrombin inhibitor, argatroban (AG), showed significant nonthrombogenicity in ECCs using a 4 h rabbit model. Herein, we evaluated if diazeniumdiolated N - N -di- N '-propyl-1,6-hexanediamine (DPHDN 2 O 2 ), which has a slightly lower degree of lipophilicity compared to DBHDN 2 O 2 , would provide similar nonthrombogenicity as the AG/DBHDN 2 O 2 -polymer-coated circuits. While DPHDN 2 O 2 releases NO at a higher flux rate than DBHDN 2 O 2 when coated (within CarboSil polymer) on the inner wall of polyvinyl chloride tubing, neither coated circuit significantly affected animal hemodynamics. Both diazeniumdiolated diamines, in combination with immobilized AG or alone, significantly reduced thrombus formation similarly in the 4 h rabbit model (vs uncoated control): AG/DBHDN 2 O 2 : 0.12 ± 0.03 cm 2 ; DBHDN 2 O 2 : 2.57 ± 0.82 cm 2 ; AG/DPHDN 2 O 2 : 0.68 ± 0.22 cm 2 ; DPHDN 2 O 2 : 1.87 + 1.26 cm 2 ; uncoated control: 6.95 ± 0.82 cm 2 . AG/DPHDN 2 O 2 was no different than AG/DBHDN 2 O in preserving platelet count and function. In addition, AG did not leach into the systemic circulation as the total clotting times were insignificantly different from the baseline values (AG/DPHDN 2 O 2 : 12.7 + 0.5 s ( n = 3); AG/DBHDN 2 O 2 : 12.3 + 0.7 s ( n = 3); baseline: 13.9 + 0.3 s ( n = 13)). Based on these results, both DPHDN 2 O 2 and DPHDN 2 O 2 are good candidates as NO donor molecules for creating nonthrombogenic polymer coatings for ECCs.

Implantable medical devices are an integral part of primary/critical care. However, these devices carry a high risk for blood clots, caused by platelet aggregation on a foreign body surface. This study focuses on the development of a simplified approach to create nitric oxide (NO) releasing intravascular electrochemical oxygen (O 2 ) sensors with increased biocompatibility and analytical accuracy. The implantable sensors are prepared by embedding S-nitroso-N-acetylpenacillamine (SNAP) as the NO donor molecule in the walls of the catheter type sensors. The SNAP-impregnated catheters were prepared by swelling silicone rubber tubing in a tetrahydrofuran solution containing SNAP. Control and SNAP-impregnated catheters were used to fabricate the Clark-style amperometric PO 2 sensors. The SNAP-impregnated sensors release NO under physiological conditions for 18 d as measured by chemiluminescence. The analytical response of the SNAP-impregnated sensors was evaluated in vitro and in vivo. Rabbit and swine models (with sensors placed in both veins and arteries) were used to evaluate the effects on thrombus formation and analytical in vivo PO 2 sensing performance. The SNAP-impregnated PO 2 sensors were found to more accurately measure PO 2 levels in blood continuously (over 7 and 20 h animal experiments) with significantly reduced thrombus formation (as compared to controls) on their surfaces., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Nitrite is a naturally occurring species present in various food samples and also present in our bodies as a product of nitric oxide (NO) oxidation. Considering the ubiquity of nitrite, its determination is of great importance in both biological and food samples. Herein, a very facile indirect method of nitrite determination in meat samples via selective reduction to nitric oxide (NO) is presented. The resulting gaseous product is quantified via portable and cost-effective electrochemical sensors. Both a novel laboratory prepared Pt-Nafion based NO sensor and a commercially available amperometric NO sensor are compared. Excellent correlations between the nitrite amount found in tested samples using both of the electrochemical sensors and a reference chemiluminescence method are demonstrated (r = 0.997 and r = 0.999 for Pt-Nafion based and commercially available NO-B4 electrochemical sensors, respectively, n = 12). Moreover, the slope of the linear regression curves are very close to unity for the comparison of the three systems tested. The amperometric sensors compared within this work exhibit good precision and accuracy and are shown to be an attractive alternative to the costly chemiluminescence detection method for accurately determining nitrite levels in food samples., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Intravascular (IV) catheters are essential devices in the hospital that are used to monitor a patient's blood and for administering drugs or nutrients. However, IV catheters are also prone to blood clotting at the point of insertion and infection by formation of robust bacterial biofilms on their surface. Nitric oxide (NO) is ideally suited to counteract both of these problems, due to its antimicrobial properties and its ability to inhibit platelet activation/aggregation. One way to equip catheters with NO releasing properties is by electrocatalytic nitrite reduction to NO by copper complexes in a multi-lumen configuration. In this work, we systematically investigate six closely related Cu(II) BMPA- and BEPA-carboxylate complexes (BMPA = bis-(2-methylpyridyl)amine); BEPA = bis-(2-ethylpyridyl)amine), using carboxylate groups of different chain lengths. The corresponding Cu(II) complexes were characterized using UV-Vis, EPR spectroscopy, and X-ray crystallography. Using detailed cyclic voltammetry (CV) and bulk electrocatalyic studies (with real-time NO quantification), in aqueous buffer, pH 7.4, we are able to derive clear reactivity relations between the ligand structures of the complexes, their Faradaic efficiencies for NO generation, their turnover frequencies (TOFs), and their redox potentials. Our results show that the complex [Cu(BEPA-Bu)](OAc) is the best catalyst with a high Faradaic efficiency over large nitrite concentration ranges and the expected best tolerance to oxygen levels. For this species, the more positive redox potential suppresses NO disproportionation, which is a major Achilles heel of the (faster) catalysts with the more negative reduction potentials.

Currently, most antimicrobial topical treatments utilize antibiotics to prevent or treat infection at a wound site. However, with the ongoing evolution of multi-drug resistant bacterial strains, there is a high demand for alternative antimicrobial treatments. Nitric oxide (NO) is an endogenous gas molecule with potent antimicrobial activity, which is effective against a wide variety of bacterial strains. In this study, the potential for creating NO releasing creams containing the naturally occurring NO carrier, S-nitrosoglutathione (GSNO), are characterized and evaluated. GSNO is shown to have prolonged stability (>300 days) when mixed and stored within Vaseline at 24 °C. Further, enhanced proliferation of NO from GSNO using zinc oxide nanoparticles (ZnO) is demonstrated. Triggering NO release from the GSNO/Vaseline mixture using a commercial zinc oxide-containing cream exhibits first-order NO release kinetics with the highest %NO release over the first 6 h. Significant killing effects against S. aureus, S. epidermidis, and P. aeruginosa are demonstrated for the GSNO/Vaseline/ZnO cream mixtures in a proportional manner dependent upon the concentration of GSNO in the final mixture., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Droplet microfluidics is an enabling platform for high-throughput screens, single-cell studies, low-volume chemical diagnostics, and microscale material syntheses. Analytical methods for real-time and in situ detection of chemicals in the droplets will benefit these applications, but they remain limited. Reported herein is a novel heterogeneous chemical sensing strategy based on functionalization of the oil phase with rationally combined sensing reagents. Sub-nanoliter oil segments containing pH-sensitive fluorophores, ionophores, and ion-exchangers enable highly selective and rapid fluorescence detection of physiologically important electrolytes (K + , Na + , and Cl - ) and polyions (protamine) in sub-nanoliter aqueous droplets. Electrolyte analysis in whole blood is demonstrated without suffering from optical interference from the sample matrix. Moreover, an oil phase doped with an aza-BODIPY dye allows indication of H 2 O 2 in the aqueous droplets, exemplifying sensing of targets beyond ionic species., (© 2019 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

The light induced nitric oxide (NO) release properties of S-nitroso-N-acetylpenicillamine (SNAP) and S-nitrosoglutathione (GSNO) NO donors doped within polydimethylsiloxane (PDMS) films (PDMS-SNAP and PDMS-GSNO respectively) for potential inhaled NO (iNO) applications is examined. To achieve photolytic release of gas phase NO from the PDMS-SNAP and PDMS-GSNO films, narrow-band LED light sources are employed and the NO concentration in a N 2 sweep gas above the film is monitored with an electrochemical NO sensor. The NO release kinetics using LED sources with different nominal wavelengths and optical power densities are reported. The effect of the NO donor loading within the PDMS films is also examined. The NO release levels can be controlled by the LED triggered release from the NO donor-doped silicone rubber films in order to generate therapeutic levels in a sweep gas for suitable durations potentially useful for iNO therapy. Hence this work may lay the groundwork for future development of a highly portable iNO system for treatment of patients with pulmonary hypertension, hypoxemia, and cystic fibrosis., (Copyright © 2019. Published by Elsevier Inc.)

Nitric oxide (NO) releasing polymers have been widely applied as biomaterials for a variety of biomedical implants and devices. However, the chemical leaching of NO donors and their byproduct species is almost always observed during the application of polymers doped with NO donors, unless the donor is covalently linked to the polymer. Herein, we report the first NO releasing poly(vinylidene fluoride-co-hexafluoropropylene) (PVDF-HFP) fluorinated copolymer prepared by incorporating a fluorinated S-nitrosothiol as the NO donor. Under physiological conditions, the resulting polymeric films can release NO for 16 days. Importantly, due to both fluorine-fluorine and electrostatic charge interactions between the fluorinated NO donor and the PVDF-HFP copolymer, the total chemical leaching of the fluorinated NO donor and its disulfide product after 9 day was only 0.6% (mol%) of the initial amount of NO donor loaded into the film. These new NO release PVDF-HFP films exhibit antimicrobial and anti-biofilm activities against both Gram positive S. aureus and Gram negative P. aeruginosa strains. The NO-releasing PVDF-HFP polymer can also be coated on Teflon tubing to release NO under physiological conditions for extended time periods. This NO-releasing PVDF-HFP copolymer with greatly reduced chemical leaching could help enhance the biocompatibility and antimicrobial activity of various biomedical devices. STATEMENT OF SIGNIFICANCE: Fluoropolymers have been widely used in creating various biomedical implants and devices. However, nitric oxide (NO) release fluoropolymers have not been well studied to date. Additionally, in the application of biomaterials doped with NO donors, a significant amount of NO donors and their byproducts almost always leach into aqueous environment. We now report an NO releasing poly(vinylidene fluoride-co-hexafluoropropylene) (PVDF-HFP) fluoropolymer by incorporating a new fluorinated S-nitrosothiol. The NO release can last for 16 days under physiological conditions. The total chemical leaching was determined to be only 0.6 mol% of the initial S-nitrosothiol loaded. As expected, significant antimicrobial/anti-biofilm activities of the NO release PVDF-HFP film were observed against Gram positive S. aureus and Gram negative P. aeruginosa bacterial strains., (Copyright © 2019 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

A dual functional polyurethane (PU) film that mimics aspects of blood vessel inner surfaces by combining surface texturing and nitric oxide (NO) release was fabricated through a soft lithography two-stage replication process. The fabrication of submicron textures on the polymer surface was followed by solvent impregnation with the NO donor, S-nitroso-N-acetylpenicillamine (SNAP). An in vitro plasma coagulation assay showed that the biomimetic surface significantly increased the plasma coagulation time and also exhibited reduced platelet adhesion and activation, thereby reducing the risk of blood coagulation and thrombosis. A contact activation assay for coagulation factor XII (FXII) demonstrated that both NO release and surface texturing also reduced FXII contact activation, which contributes to the inhibition of plasma coagulation. The biomimetic surface was also evaluated for bacterial adhesion in plasma and results demonstrate that this combined strategy enables a synergistic effect to reduce bacterial adhesion of Staphylococcus epidermidis, Staphylococcus aureus, and Pseudomonas aeruginosa microorganisms. The results strongly suggest that the biomimetic modification with surface texturing and NO release provides an effective approach to improve the biocompatibility of polymeric materials in combating thrombosis and microbial infection. STATEMENT OF SIGNIFICANCE: (1) Developed a dual functional polyurethane (PU) film that mimics blood vessel inner surface by combining surface texturing and nitric oxide (NO) release for combatting biomaterial associated thrombosis and microbial infection. (2) Studied the blood coagulation response and bacterial adhesion to such biomimetic PU surfaces, and demonstrated that the combination of surface texturing and NO release synergistically reduced the platelet adhesion and bacterial adhesion in plasma, providing an effective approach to improve the biocompatibility of biomaterials used in blood-contacting medical devices. (3) The NO releasing surface significantly inhibits the plasma coagulation via the reduction of contact activation of FXII, indicating the multifunctional roles of NO in improving the biocompatibility of biomaterials in blood-contacting medical devices., (Copyright © 2018 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

A series of new fluoro S-nitrosothiols is reported as potential nitric oxide (NO) donors. A three-step synthesis and the NO releasing kinetic profiles of these species are presented. The stoichiometric release of NO, with the clean formation of corresponding disulfides, confirms that these new species can facilitate their application as NO donors for various applications including creating novel antimicrobial and thromboresistant fluoropolymer-based medical devices.

Semi-crystalline thermoplastics are an important class of biomaterials with applications in creating extracorporeal and implantable medical devices. In situ release of nitric oxide (NO) from medical devices can enhance their performance via NO's potent anti-thrombotic, bactericidal, anti-inflammatory, and angiogenic activity. However, NO-releasing semi-crystalline thermoplastic systems are limited and the relationship between polymer crystallinity and NO release profile is unknown. In this paper, the functionalization of poly(ether-block-amide) (PEBA), Nylon 12, and polyurethane tubes, as examples of semi-crystalline polymers, with the NO donor S-nitroso-N-acetylpenicillamine (SNAP) within, is demonstrated via a polymer swelling method. The degree of crystallinity of the polymer plays a crucial role in both SNAP impregnation and NO release. Nylon 12, which has a relatively high degree of crystallinity, exhibits an unprecedented NO release duration of over 5 months at a low NO level, while PEBA tubing exhibits NO release over days to weeks. As a new biomedical application of NO, the NO-releasing PEBA tubing is examined as a cannula for continuous subcutaneous insulin infusion. The released NO is shown to enhance insulin absorption into the bloodstream probably by suppressing the tissue inflammatory response, and thereby could benefit insulin pump therapy for diabetes management.

Fluorinated polymers are widely used as biomaterials in various biomedical implant and device applications. However, thrombogenicity, surface-induced inflammation, and risk of microbial infection remain key issues that can limit their use. In this work, we describe the first nitric oxide (NO) releasing fluorinated polymer, in which a new fluorinated NO donor, S -nitroso- N -pentafluoropropionylpenicillamine (C 2 F 5 -SNAP), is incorporated within the polyvinylidene fluoride (PVDF) tubing. The synthesis, decomposition kinetics, and NO-release characteristics of the C 2 F 5 -SNAP species are described in detail. Then, using a simple solvent swelling method, we demonstrate that C 2 F 5 -SNAP can readily be doped into PVDF tubing. The resulting tubing can release NO for 11 days under physiological conditions, with an NO flux > 0.5 × 10 -10 mol/cm 2 ·min over the first 7 days. Due to fluorous-fluorous interactions, the leaching of the fluorinated NO donor and its decomposed products is shown to be very low (less than 5 nmol/mg, total). Further, the new NO-releasing PVDF tubing exhibits significant antimicrobial activity (compared to undoped PVDF tubing) against both gram positive and negative S. aureus and P. aeruginosa bacterial strains over a 7 d test period. This new NO-releasing fluorinated polymer is likely to have the potential to improve the biocompatibility and antimicrobial activity of various biomedical devices., Competing Interests: Conflicts of interest The authors declare no competing financial interest.

Nitric oxide (NO) levels in exhaled breath are a non-invasive marker that can be used to diagnose various respiratory diseases and monitor a patient's response to given therapies. A portable and inexpensive device that can enable selective NO concentration measurements in exhaled breath samples is needed. Herein, the performance of an amperometric Pt-Nafion-based gas phase sensor for detection of NO in exhaled human nasal breath is examined. Enhanced selectivity over carbon monoxide and ammonia is achieved via an in-line zinc oxide-based filter. Exhaled nasal NO levels measured in 21 human samples with the sensor are shown to correlate well with those obtained using a chemiluminescence reference method (R 2 = 0.9836).

Further studies aimed at examining the activity of different Cu(II)-ligand complexes to serve as electron-transfer mediators to prepare novel antimicrobial/thromboresistant nitric oxide (NO)-releasing intravenous catheters are reported. In these devices, the NO release can be modulated by applying different potentials or currents to reduce the Cu(II)-complexes to Cu(I) species which then reduce nitrite ions into NO (g) within a lumen of the catheter. Four different ligands are compared with respect to NO generation efficiency and stability over time using both single- and dual-lumen silicone rubber catheters: N-propanoate- N, N-bis(2-pyridylethyl)amine (BEPA-Pr), N-propanoate- N, N-bis(2-pyridylmethyl)amine (BMPA-Pr), 1,4,7-trimethyl-1,4,7-triazacyclononane (Me 3 TACN), and tris(2-pyridylmethyl)amine (TPMA). Of these, the Cu(II)BEPA-Pr and Cu(II)Me 3 TACN complexes provide biomedically useful NO fluxes from the surface of the catheters, >2 × 10 -10 mol·min -1 ·cm -2 , under conditions mimicking the bloodstream environment. Cu(II)Me 3 TACN exhibits the best stability over time with a steady and continuous NO release observed for 8 d under a nitrogen atmosphere. Antimicrobial experiments conducted over 5 d with NO-releasing catheters turned "on" electrochemically for only 3 or 6 h each day revealed >2 logarithmic units in reduction of bacterial biofilm attached to the catheter surfaces. The use of optimal Cu(II)-ligand complexes within a lumen reservoir along with high levels of nitrite ions can potentially provide an effective method of preventing/decreasing the rate of infections caused by intravascular catheters.

A universal method for the detection, quantification, and characterization of polyquaterniums (PQs) in a simple background electrolyte solution and in more complex recreational swimming pool water samples is presented. This method involves the application of polycation-sensitive ion-selective optodes (ISOs) prepared by inkjet printing dinonylnaphthalenesulfonic acid (H + DNNS - ) and chromoionophore I directly onto Whatman TM qualitative filter paper. No plasticizer or added polymer matrix is required for the fabrication of the sensing layer which is coated on the cellulose fibers of the filter paper. PQ-6, PQ-2, PQ-10, and poly(2-methacryloxyethyltrimethylammonium) chloride (PMETAC) are used as model PQ species for direct optical detection at ppm levels. We further demonstrate that PQ-6 can be detected in recreational swimming pool water samples using this new type of sensor, and that detection of polyanions is also possible using an indirect detection method. Lastly, to circumvent the challenge of polyion-sensitive ISOs exhibiting a pH dependence, the sensors were soaked in buffer and dried to provide local buffering for applied liquid samples and an optical signal independent of sample pH.

Previous work in a 4 h rabbit thrombogenicity model has shown that a nitric oxide- (NO) generating polymer extracorporeal circuits (ECC) with infusion of S- nitroso- N -acetyl-penicillamine (SNAP) preserved platelets eventhough platelets were activated as shown by an increase in the glycoprotein, p-selectin. The platelet preservation mechanism was shown to be due to a changing fibrinogen structure leading to attenuation of platelet aggregation. Understanding the effects that SNAP, another RSNO, S- nitroso-glutathione (GSNO) as well as the non-RSNO, sodium nitroprusside (SNP), may have on human fibrinogen polymerization, this in vitro study evaluated the released NO effects on the thrombin-mediated fibrin formation and fibrinogen structure. Thrombin-induced fibrin formation at 300 μM SNAP (50 + 11% of baseline) was significantly reduced compared to SNAP's parent, N-acetyl-penicillamine (NAP) (95 + 13%) after 1 h of RSNO exposure. GSNO, its parent, glutathione (GSH) and 1000 ppm NO gas did not attenuate the thrombin-mediated fibrin formation. SNAP, NAP and SNP exposure for 1 h, however, did not decrease thrombin activity by directly inhibiting thrombin itself. Changes in fibrinogen conformation as measured by intrinsic tryptophan fluorescence significantly decreased in the 300 μM SNAP (38057 + 1196 mean fluorescence intensity (MFI) and SNP (368617 + 541 MFI) groups versus the NAP control (47937 + 1196 MFI). However, infused 1000 ppm NO gas had no direct effect on the ITF after 1 h incubation at 37°C. High performance liquid chromatography (HPLC) showed that fibrinogen degradation by 0.03 U/ml thrombin was concentration-dependently reduced after 1 h with SNAP but not with NAP or SNP. Western blotting showed RSNOs, SNAP, NAP and the non-RSNO, SNP-incubated fibrinogen solutions showed that the percent level of the Aγ dimer to total Aγ dimer + γ monomer was significantly reduced in the case of the SNAP group when compared to SNP group. These results suggest that NO donors such as SNAP and SNP induce fibrinogen conformational changes by potentially nitrosating fibrinogen tyrosine residues. These NO-mediated fibrinogen changes induced via NO donors may provide another mechanism of NO for improving thromboresistance in ECC.

Over the last three decades, there has been extensive interest in developing in vivo chemical sensors that can provide real-time measurements of blood gases (oxygen, carbon dioxide, and pH), glucose/lactate, and potentially other critical care analytes in the blood of hospitalized patients. However, clot formation with intravascular sensors and foreign body response toward sensors implanted subcutaneously can cause inaccurate analytical results. Further, the risk of bacterial infection from any sensor implanted in the human body is another major concern. To solve these issues, the release of an endogenous gas molecule, nitric oxide (NO), from the surface of such sensors has been investigated owing to NO's ability to inhibit platelet activation/adhesion, foreign body response and bacterial growth. This paper summarizes the importance of NO's therapeutic potential for this application and reviews the publications to date that report on the analytical performance of NO release sensors in laboratory testing and/or during in vivo testing.

The first paper-based polyion-sensitive optodes are reported. Dinonylnaphthalene sulfonic acid (a cation exchanger) and chromoionophore I (a lipophilic optical pH indicator) are printed on filter paper in the absence of any plasticizer and/or additional hydrophobic polymeric phase. The resulting optodes exhibit sensitive colorimetric response to polycations such as protamine but not to small inorganic cations because only polycations are able to form cooperative ion pairs with dinonylnaphthalenesulfonate adsorbed to the cellulose paper. The color change of the optode is recorded via an iPhone camera and analyzed by an iPhone App. The protamine-sensing optode platform is used to indirectly detect protease activity (trypsin) based on proteolytic digestion of protamine, and polyanions (pentosan polysulfate and heparin) based on the strong binding reaction of polyanions with protamine. The indirect sensing system is further simplified on a multilayer membrane device that consists of an optode paper site modified with buffer to prevent optode dependence on sample pH, and an underlying cellulose acetate filter membrane coated with protamine to eliminate addition of the indicator polycation into the sample. The detection of pentosan polysulfate concentrations in an undiluted urine sample is successfully demonstrated via this approach. Lastly, it is shown that plasticizer-free polyanion-sensitive optodes based on an adsorbed layer of quaternary ammonium type anion exchanger and a phenolic azo type proton chromoionophore can also be fabricated directly on cellulose paper strips.

A new portable gas phase nitric oxide (NO) generator is described for potential applications in inhaled NO (INO) therapy and during cardiopulmonary bypass (CPB) surgery. In this system, NO is produced at the surface of a large-area mesh working electrode by electrochemical reduction of nitrite ions in the presence of a soluble copper(II)-ligand electron transfer mediator complex. The NO generated is then transported into gas phase by either direct purging with nitrogen/air or via circulating the electrolyte/nitrite solution through a gas extraction silicone fiber-based membrane-dialyzer assembly. Gas phase NO concentrations can be tuned in the range of 5-1000 ppm (parts per million by volume for gaseous species), in proportion to a constant cathodic current applied between the working and counter electrodes. This new NO generation process has the advantages of rapid production times (5 min to steady-state), high Faraday NO production efficiency (ca. 93%), excellent stability, and very low cost when using air as the carrier gas for NO (in the membrane dialyzer configuration), enabling the development of potentially portable INO devices. In this initial work, the new system is examined for the effectiveness of gaseous NO to reduce the systemic inflammatory response (SIR) during CPB, where 500 ppm of NO added to the sweep gas of the oxygenator or to the cardiotomy suction air in a CPB system is shown to prevent activation of white blood cells (granulocytes and monocytes) during extracorporeal circulation with cardiotomy suction conducted with five pigs.

The detection of four different polyquaterniums (PQs) using a fully reversible potentiometric polyion sensor in three different detection modes is described. The polyion sensing "pulstrodes" serve as the detector for direct dose-response experiments, beaker titrations, and in a flow-injection analysis (FIA) system. Direct polycation response toward PQ-2, PQ-6, PQ-10, and poly(2-methacryloxyethyltrimethylammonium) chloride (PMETAC) yields characteristic information about each PQ species (e.g., relative charge densities, etc.) via syringe pump addition of each PQ species to a background electrolyte solution. Quantitative titrations are performed using a syringe pump to deliver heparin as the polyanion titrant to quantify all four PQs at μg/mL levels. Both the direct and indirect methods incorporate the use of a three-electrode system including counter, double junction reference, and working electrodes. The working electrode possesses a plasticized poly(vinyl chloride) (PVC) membrane containing the neutral lipophilic salt of dinonylnaphthalenesulfonate (DNNS - ) tridodecylmethylammonium (TDMA + ). Further, the titration method is shown to be useful to quantify PQ-6 levels in recreational swimming pool water collected in Ann Arbor, MI. Finally, a FIA system equipped with a pulstrode detector is used to demonstrate the ability to potentially quantify PQ levels via a more streamlined and semiautomated testing platform.

The compatibility of nitric oxide (NO) release coatings with implantable enzymatic glucose sensors based on osmium (III/II) mediated electrochemical detection is examined for the first time. NO-releasing osmium-mediated glucose sensors are prepared using a S-nitrosothiol impregnated outer tubing and are tested in vitro in both phosphate buffer (pH 7.4) and whole porcine blood. It is demonstrated that after 3 days of continuous NO release at or above physiological levels, there are no negative effects on the osmium mediated electrochemical currents. Indeed, such sensors maintain their functionality, sensitivity, and accuracy for detecting glucose levels in blood. The results suggest that improved performance of both intravascular and, potentially, subcutaneous Os(III/II) mediated glucose sensors may be realized by taking advantage of NO's well-known anticlotting, anti-inflammatory, and antimicrobial properties.

A general anion-sensing platform is reported based on a portable and cost-effective ion-selective optode and a smartphone detector equipped with a color analysis app. In contrast to traditional anion-selective optodes using a hydrophobic polymer and/or plasticizer to dissolve hydrophobic sensing elements, the new optode relies on hydrophilic cellulose paper. The anion ionophore and a lipophilic pH indicator are inkjet-printed and adsorbed on paper and form a "dry" hydrophobic sensing layer. Porous cellulose sheets also allow the sensing site to be modified with dried buffer that prevents any sample pH dependence of the observed color change. A highly selective fluoride optode using an Al III -porphyrin ionophore is examined as an initial example of this new anion sensing platform for measurements of fluoride levels in drinking water samples. Apart from Lewis acid-base recognition, hydrogen bonding recognition is also compatible with this sensing platform., (© 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

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.

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.

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 -10 molmin -1 cm -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., (Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

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 cm 2 /catheter) compared to the controls (0.84 ± 0.19 cm 2 /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.

Two facile, robust, and universal methods by which various polymeric quaternary ammonium salts (polyquaterniums (PQs)) can be quantified and characterized using simple potentiometric polymeric membrane polyion-sensitive electrodes as detectors are described. The two methods are (a) direct detection with polycation sensitive membrane electrodes based on the sodium salt of dinonylnaphthalenesulfonate (NaDNNS), and (b) indirect detection using polyanion sensors based on tridodecylmethylammonium chloride (TDMAC) and dextran sulfate (DS) as a titrant to complex the various polyquaternary species (four different PQs: PQ-2, PQ-6, PQ-10, and poly(2-methacryloxyethyltrimethylammonium) chloride (PMETAC)). Direct detection yields information regarding the charge density of the polycationic species. For the titration method, a series of polyanion sensors doped with TDMAC are used to follow a potentiometric titration of a PQ species using a syringe pump to deliver the titrant. This indirect detection method is more reliable and yields limits of detection in the ppm range for the four PQs examined. The titration method is further explored for detecting excess levels of PQ-6, a common flocculating agent for municipal water supply systems, within the purified water emitted by the Ann Arbor, MI, drinking water treatment plant.

Blood-contacting devices, such as intravascular catheters, suffer from challenges related to thrombus formation and infection. Nitric oxide (NO) is an endogenous antiplatelet and antimicrobial agent. Exogenous release of NO from various polymer matrices has been shown to reduce thrombosis and infection of/on implantable medical devices. However, the clinical applications of such materials have been hindered due to factors such as NO donor leaching and thermal instability. In this study, a novel approach is demonstrated in which one lumen of commercial dual lumen catheters is dedicated to the NO release chemistry, allowing the other lumen to be available for clinical vascular access. A composite consisting of poly(ethylene glycol) (PEG) and S-nitroso-N-acetylpenicillamine (SNAP) is used to fill the NO-releasing lumen of commercial 7 French silicone catheters. Physiological levels of NO are released from the SNAP-PEG catheters for up to 14 d, as measured by chemiluminescence NO analyzer (in PBS buffer at 37 °C). PEG facilitates the NO release from SNAP within the lumen by increasing the water absorption and slowly dissolving the solid SNAP-PEG composite. In a CDC biofilm bioreactor, the SNAP-PEG catheters are found to reduce >97% bacterial adhesion as compared to the PEG controls for single bacterial species including E. coli and S. aureus. SNAP-PEG and PEG control catheters were implanted in rabbit veins for 7 h (single lumen) and 11 d (dual lumen) to evaluate their hemocompatibility properties. Significant reductions in thrombus formation on the SNAP-PEG vs PEG controls were observed, with ca. 85% reduction for 7 h single lumen catheters and ca. 55% reduction for 11 d dual lumen catheters.

Unlabelled: Two major problems with implanted catheters are clotting and infection. Nitric oxide (NO) is an endogenous vasodilator as well as natural inhibitor of platelet adhesion/activation and an antimicrobial agent, and NO-releasing polymers are expected to have similar properties. Here, NO-releasing central venous catheters (CVCs) are fabricated using Elast-eon™ E2As polymer with both diazeniumdiolated dibutylhexanediamine (DBHD/NONO) and poly(lactic-co-glycolic acid) (PLGA) additives, where the NO release can be modulated and optimized via the hydrolysis rate of the PLGA. It is observed that using a 10% w/w additive of a PLGA with ester end group provides the most controlled NO release from the CVCs over a 14d period. The optimized DBHD/NONO-based catheters are non-hemolytic (hemolytic index of 0%) and noncytotoxic (grade 0). After 9d of catheter implantation in the jugular veins of rabbits, the NO-releasing CVCs have a significantly reduced thrombus area (7 times smaller) and a 95% reduction in bacterial adhesion. These results show the promise of DBHD/NONO-based NO releasing materials as a solution to achieve extended NO release for longer term prevention of clotting and infection associated with intravascular catheters., Statement of Significance: Clotting and infection are significant complications associated with central venous catheters (CVCs). While nitric oxide (NO) releasing materials have been shown to reduce platelet activation and bacterial infection in vitro and in short-term animal models, longer-term success of NO-releasing materials to further study their clinical potential has not been extensively evaluated to date. In this study, we evaluate diazeniumdiolate based NO-releasing CVCs over a 9d period in a rabbit model. The explanted NO-releasing CVCs were found to have significantly reduced thrombus area and bacterial adhesion. These NO-releasing coatings can improve the hemocompatibility and bactericidal activity of intravascular catheters, as well as other medical devices (e.g., urinary catheters, vascular grafts)., (Copyright © 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

Nitric oxide (NO) releasing polymers are promising in improving the biocompatibility of medical devices. Polyurethanes are commonly used to prepare/fabricate many devices (e.g., catheters); however, the transport properties of NO within different polyurethanes are less studied, creating a gap in the rational design of new NO releasing devices involving polyurethane materials. Herein, we study the diffusion and partitioning of NO in different biomedical polyurethanes via the time-lag method. The diffusion of NO is positively correlated with the PDMS content within the polyurethanes, which can be rationalized by effective media theory considering various microphase morphologies. Using catheters as a model device, the effect of these transport properties on the NO release profiles and the distribution around an asymmetric dual lumen catheter are simulated using finite element analysis and validated experimentally. This method can be readily applied in studying other NO release medical devices with different configurations.

Herein, we report a novel design and the antimicrobial efficacy of a flexible nitric oxide (NO) releasing patch for potential wound healing applications. The compact sized polydimethylsiloxane (PDMS) planar patch generates NO via electrochemical reduction of nitrite ions mediated by a copper(II)-ligand catalyst using a portable power system and an internal gold coated stainless steel mesh working electrode. Patches are fabricated via soft lithography and 3-D printing. The devices can continuously release NO over 4 days and exhibit potent bactericidal effects on both Escherichia coli and Staphylococcus aureus . The device may provide an effective, safe, and less costly alternative for treating chronic wounds.

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).

Fast eradication of mature biofilms is the 'holy grail' in the clinical management of device-related infections. Endogenous nitric oxide (NO) produced by macrophages plays an important role in host defense against intracellular pathogens, and NO is a promising agent in preventing biofilms formation in vitro. However, the rate of delivery of NO by various NO donors (e.g., diazeniumdiolates, S-nitrosothiols, etc.) is difficult to control, which hinders fundamental studies aimed at understanding the role of NO in biofilm control. In this study, by using a novel precisely controlled electrochemical NO releasing catheter device, we examine the effect of physiological levels of NO on eradicating mature Pseudomonas aeruginosa biofilm (7 days), as well as the potential application of the combination of NO with antimicrobial agents. It is shown that physiological levels of NO exhibit mixed effects of killing bacteria and dispersing ambient biofilm. The overall biofilm-eradicating effect of NO is quite efficient in a dose-dependent manner over a 3 h period of NO treatment. Moreover, NO also greatly enhances the efficacy of antimicrobial agents, including human beta-defensin 2 (BD-2) and several antibiotics, in eradicating biofilm and its detached cells, which otherwise exhibited high recalcitrance to these antimicrobial agents. The electrochemical NO release technology offers a powerful tool in evaluating the role of NO in biofilm control as well as a promising approach when combined with antimicrobial agents to treat biofilm-associated infections in hospital settings, especially infections resulting from intravascular catheters.

Polymeric quaternary ammonium salts (polyquaterniums) have found increasing use in industrial and cosmetic applications in recent years. More specifically, polyquaternium-10 (PQ-10) is routinely used in cosmetic applications as a conditioner in personal care product formulations. Herein, we demonstrate the use of potentiometric polyion-sensitive polymeric membrane-based electrodes to quantify PQ-10 levels. Mixtures containing both PQ-10 and sodium lauryl sulfate (SLS) are used as model samples to illustrate this new method. SLS is often present in cosmetic samples that contain PQ-10 (e.g., shampoos, etc.) and this surfactant species interferes with the polyion sensor detection chemistry. However, it is shown here that SLS can be readily separated from the PQ-10/SLS mixture by use of an anion-exchange resin and that the PQ-10 can then be titrated with dextran sulphate (DS). This titration is monitored by potentiometric polyanion sensors to provide equivalence points that are directly proportional to PQ-10 concentrations.