Development of Cu-Modified PVC and PU for Catalytic Generation

colloids and interfaces

Article Development of Cu-Modiﬁed PVC and PU for Catalytic Generation of Nitric Oxide

Liana Azizova 1,* , Santanu Ray 2 , Sergey Mikhalovsky 3 and Lyuba Mikhalovska 1

1 School of Pharmacy and Biomolecular Sciences, University of Brighton, Brighton BN2 4GJ, UK; [email protected] 2 Surface Analysis Laboratory, School of Environment and Technology, University of Brighton, Brighton BN2 4GJ, UK; [email protected] 3 ANAMAD Ltd., Science Innovation Park, Science Park Square, Falmer, Brighton BN1 9SB, UK; [email protected] * Correspondence: [email protected]; Tel.: +44-1273-64-1914

Received: 22 December 2018; Accepted: 4 March 2019; Published: 9 March 2019

Abstract: Nitric oxide (NO) generating surfaces are potentially promising for improving haemocompatibility of blood-contacting biomaterials. In the present report, Cu-modified poly(vinyl chloride) (PVC) and polyurethane (PU) were prepared via polydopamine (pDA)-assisted chelation. The copper content on the PVC and PU modified surfaces, assessed by inductively coupled plasma - optical emission spectrometry (ICP-OES), were about 3.86 and 6.04 nmol·cm−2, respectively. The Fourier-transform infrared spectroscopy (FTIR) and X-ray photoelectron spectroscopy (XPS) data suggest that copper is attached to the polymer surface through complex formation with pDA. The cumulative leaching of copper from modified PVC and PU during the five day incubation in phosphate buffered saline (PBS), measured by inductively coupled plasma mass spectrometry (ICP-MS), was about 50.7 ppb and 48 ppb, respectively which is within its physiological level. Modified polymers were tested for their ability to catalytically generate NO by decomposing of endogenous S-nitrosothiol (GSNO). The obtained data show that Cu-modified PVC and PU exhibited the capacity to generate physiological levels of NO which could be a foundation for developing new biocompatible materials with NO-based therapeutics.

Keywords: nitric oxide; S-nitrosothiol (GSNO); copper; polydopamine; poly(vinyl chloride) (PVC); polyurethane (PU)

1. Introduction Poly(vinyl chloride) (PVC) and polyurethanes (PU) are the most commonly used polymers for blood-contacting applications [1]. The foremost important requirement for these polymers is the prevention of coagulation and platelet activation [1]. However, despite the progress that has been achieved in improving haemocompatibility of implantable biomaterials their long-term clinical applications still poses a problem [2]. The surface modification of the commercialized polymers is an efficient and cost effective approach to improving their haemocompatibility in comparison to creating new biomaterials [3]. During the recent decades, a number of biomaterials which release nitric oxide (NO) from the surface have been developed. NO has various physiological functions in the human body such as antiplatelet, anti-inflammatory, vasodilation [4,5] and inhibition of biofilm formation properties [6–8]. Healthy endothelial cells produce and release endogenously a NO flux of 0.5 − 4.0 × 10−10 mol·cm−2·min−1 [4]. Therefore, surfaces which release/generate NO locally are potentially promising for improving thromboresistance of biomaterials. One such approach to designing NO releasing surfaces is incorporation of exogenous NO donor molecules [5] and covalent

Colloids Interfaces 2019, 3, 33; doi:10.3390/colloids3010033 www.mdpi.com/journal/colloids Colloids Interfaces 2019, 3, 33 2 of 13 Colloids Interfaces 2019, 3, x FOR PEER REVIEW 2 of 13 bindingdonors to of NOthe donorssurface toof the various surface materials. of various Howe materials.ver, biomaterials However, biomaterials with covalently with covalentlylinked or linkedembedded or embedded NO donors NO have donors several have limitations several limitations such as the such limited as the surface limited loading surface capacity loading of capacity the NO ofdonor, the NO low donor, NO donor low NO stability, donor and stability, formation and formation of potentially of potentially toxic decomposition toxic decomposition products products such as suchN-nitrosoamines. as N-nitrosoamines. The utility The of utilityNO-releasing of NO-releasing biomaterials biomaterials is limited to is short-term limited to short-termcontact biomedical contact biomedicaldevices [9–12]. devices S-nitrosothiols [9–12]. S-nitrosothiols (RSNO) are (RSNO)a class of are NO a class donors ofNO which donors are thought which are to thoughtserve as to a servereservoir as a and reservoir transporter and transporter of NO within of NO biological within biologicalsystems [13]. systems The catalytic [13]. The generation catalytic generation of NO from of NOendogenous from endogenous S-nitrosothiolsS-nitrosothiols at the blood/material at the blood/material interface was interface suggested was suggested as an alternative as an alternative approach approachfor the local for NO the delivery local NO to deliverythe surface to the[6,14 surface–18]. RSNOs [6,14– 18are]. natural RSNOs metabolites are natural existing metabolites in the existing tissues inand the blood. tissues Bearing and blood. in mind Bearing that RSNOs in mind are that continuously RSNOs are produced continuously in the produced blood at inconcentrations the blood at concentrationsranging from 10 ranging to 100 μ fromM, they 10 to could 100 µ beM, considered they could as be an considered inexhaustible as an source inexhaustible of NO in source vivo [19,20]. of NO inTherefore, vivo [19 ,20designing]. Therefore, NO-generating designing NO-generating surfaces could surfacesbe a potentially could be useful a potentially approach useful for long-term approach forcontact long-term medical contact devices. medical devices. 2+ ItIt has been been reported reported that that polymers polymers with with immobilized immobilized transition transition metal metal ions, ions, primarily primarily CuCu and 2+ andFe Fe [17,21],[17, 21copper], copper complexes complexes or orcopper-containing copper-containing metal metal–organic–organic frameworks frameworks [[6,8,146,8,14–1717]] catalytically decomposedecompose endogenous endogenousS-nitrosothiols S-nitrosothiols with with in situ in generationsitu generation of NO of (Scheme NO (Scheme1). Copper 1). ionsCopper catalysing ions catalysing RSNO decomposition RSNO decomposition are present are in thepresent blood in and the essential blood and for theessential normal for physiological the normal functionphysiological of the function body. of the body. Depending onon thethe substrate substrate surface surface chemistry, chemistry, different different methods methods of copperof copper immobilization immobilization on theon surfacethe surface have have been been proposed. proposed. Among Among them them is a is one-step a one-step incorporation incorporation of copper of copper on theon surfacethe surface via catecholamines.via catecholamines. Catecholamines Catecholamines are naturally are occurringnaturally aminesoccurring that functionamines asthat neurotransmitters function as andneurotransmitters hormones within and the hormones body. Dopamine within (DA) the (Scheme body. 1),Dopamine one of catecholamines, (DA) (Scheme has a1), half-life one ofof acatecholamines, few minutes when has a circulating half-life of ina few the minutes blood; it when is secreted circulating into urinein the afterblood; being it is secreted broken down into urine [22]. Byafter self-polymerization being broken down DA [22]. can By form self-polymerization a very thin layer DA of can polydopamine form a very thin (pDA) layer (Scheme of polydopamine1) capable of(pDA) attaching (Scheme to various1) capable surfaces of attaching [23]. pDAto various exhibits surf strongeraces [23]. adhesion pDA exhibits to the stronger hydrophobic adhesion surfaces to the andhydrophobic adheres lesssurfaces strongly and adheres to the hydrophilic less strongly surfaces to the hydrophilic [24]. The metal surfaces ions [24]. attach The to metal thesurface ions attach via formationto the surface of complexesvia formation with of complexes pDA quinone with and pDA catechol quinone units and catechol [25,26]. units pDA [25,26]. provides pDA a platformprovides for further immobilization of biomolecules containing −NH and −SH groups via Schiff base or a platform for further immobilization of biomolecules containing2 −NH and −SH groups via Schiff Michaelbase or Michael addition addition reactions reactions [27,28]. According[27,28]. According to literature, to literature, the structure the structure of pDA is of consists pDA is of consists a mixture of ofa mixture open-chain of open-chain dopamine dopamine units and differentunits and oligomers different witholigomers indole with units indole with differentunits with degrees different of saturationdegrees of (Schemesaturation2). (Scheme 2). Using the polydopamine-assisted chelationchelation withwith copper we aimed to prepare novel Cu-modified Cu-modiﬁed PVC and PUPU polymerspolymers ableable toto catalyticallycatalytically generate NO from thethe endogenousendogenous sourcessources atat physiologicalphysiological conditions.conditions.

SchemeScheme 1. Scheme 1. Scheme of S-nitrosoglutathione of S-nitrosoglutathione (GSNO) (GSNO) structure, structure, reaction reaction of GSNO of GSNOdecomposition decomposition by copper. by copper.

Colloids Interfaces 2019, 3, 33 3 of 13 Colloids Interfaces 2019, 3, x FOR PEER REVIEW 3 of 13

Scheme 2. Scheme of dopamine and polydopamine (pDA) structure [[26,29,30].26,29,30].

2. Materials and Methods 2.1. Materials 2.1. Materials Polyvinylchloride film (PVC, thickness 0.2 mm, Goodfellow Cambridge Ltd, Huntingdon, Polyvinylchloride film (PVC, thickness 0.2 mm, Goodfellow Cambridge Ltd, Huntingdon, UK), UK), polyurethane film (PU, thickness 80 µm, Delstar International Ltd, Brough, UK), polyurethane film (PU, thickness 80 μm, Delstar International Ltd, Brough, UK), phosphate-buffered phosphate-buffered saline tablets (PBS, MP Biomedicals, Illkirch, France), dopamine hydrochloride saline tablets (PBS, MP Biomedicals, Illkirch, France), dopamine hydrochloride (DA), (DA), tris(hydroxymethyl)aminomethane (Tris), S-nitrosoglutathione (GSNO), L-glutathione reduced tris(hydroxymethyl)aminomethane (Tris), S-nitrosoglutathione (GSNO), L-glutathione reduced (GSH), copper (II) chloride dihydrate (CuCl2·2H2O), sodium nitrite and sodium nitrate (Sigma-Aldrich, (GSH), copper (II) chloride dihydrate (CuCl ∙2HO), sodium nitrite and sodium nitrate (Sigma- St. Louis, MO, USA), and Cu solution (1000 ppm in 1 M HNO3 from Fisher Scientific, Hampton, NH, Aldrich, St. Louis, MO, USA), and Cu solution (1000 ppm in 1 M HNO from Fisher Scientific, USA) were used as received. All solutions were prepared with 18 MΩ cm−1 deionized (DI) water Hampton, NH, USA) were used as received. All solutions were prepared with 18 MΩ cm−1 deionized obtained using a Milli-Q system (Millipore Corp., Billerica, MA, USA). It showed negative results for (DI) water obtained using a Milli-Q system (Millipore Corp., Billerica, MA, USA). It showed negative the presence of Cu ions measured by inductively coupled plasma mass spectrometry (ICP-MS). results for the presence of Cu ions measured by inductively coupled plasma mass spectrometry (ICP- 2.2.MS). Method of Copper Immobilization

2.2. MethodPVC and of PUCopper films Immobilization were cut into round discs with d = 1.3 cm. Before modification, the polymer discs were cleaned by rinsing with ethanol, immersed in DI water and sonicated for 3 min, then rinsed PVC and PU films were cut into round discs with d = 1.3 cm. Before modification, the polymer with DI water, and soaked in Tris-HCl buffer solution (10 mM, pH 8.5) for 15 min prior to modification. discs were cleaned by rinsing with ethanol, immersed in DI water and sonicated for 3 min, then rinsed Copper (II) chloride solution was prepared by dissolving CuCl2·2H2O in a Tris-HCl buffer solution with DI water, and soaked in Tris-HCl buffer solution (10 mM, pH 8.5) for 15 min prior to (10 mM, pH 8.5) with stirring. To study the effect of DA:Cu2+ molar ratios, solutions with three different modification. Copper (II) chloride solution was prepared by dissolving CuCl ∙2H O in a Tris-HCl copper (II) concentrations, 3.5, 5.8 and 11.7 mM were made. Dopamine solution (2 g·L−1; 10.5 mM) buffer solution (10 mM, pH 8.5) with stirring. To study the effect of DA:Cu2+ molar ratios, solutions was prepared by dissolving dopamine hydrochloride in Tris-HCl buffer solution (10 mM, pH 8.5) with three different copper (II) concentrations, 3.5, 5.8 and 11.7 mM were made. Dopamine solution with stirring. A freshly prepared dopamine solution was always used because of self-polymerization (2 g ∙L−1; 10.5 mM) was prepared by dissolving dopamine hydrochloride in Tris-HCl buffer solution of dopamine in the buffer in aerobic conditions [31]. Pre-soaked in buffer, PVC and PU discs were (10 mM, pH 8.5) with stirring. A freshly prepared dopamine solution was always used because of immersed into fresh dopamine solution, to which immediately an equal volume of a copper salt self-polymerization of dopamine in the buffer in aerobic conditions [31]. Pre-soaked in buffer, PVC solution was gradually added with stirring and polymerization was conducted for 24 h at room and PU discs were immersed into fresh dopamine solution, to which immediately an equal volume temperature (RT) under continuous stirring in the dark. To determine the optimal DA:Cu2+ molar ratio, of a copper salt solution was gradually added with stirring and polymerization was conducted for the polymerization was carried out at three different ratios, 3.5:1, 1.8:1 and 0.9:1. The total volume of 24 h at room temperature (RT) under continuous stirring in the dark. To determine the optimal the solution was 1 mL per disc. After incubation, discs were sonicated in DI water for 5 min to remove DA:Cu2+ molar ratio, the polymerization was carried out at three different ratios, 3.5:1, 1.8:1 and 0.9:1. the non-attached pDA, then rinsed with DI water three times and air-dried. The obtained discs were The total volume of the solution was 1 mL per disc. After incubation, discs were sonicated in DI water denoted as PVC/pDA/Cu and PU/pDA/Cu. Polymers coated with pDA only without copper were for 5 min to remove the non-attached pDA, then rinsed with DI water three times and air-dried. The use as controls (PVC/pDA and PU/pDA). obtained discs were denoted as PVC/pDA/Cu and PU/pDA/Cu. Polymers coated with pDA only without copper were use as controls (PVC/pDA and PU/pDA).

Colloids Interfaces 2019, 3, 33 4 of 13

2.3. Fourier Transform Infrared Spectroscopy Method (FTIR) for Studying the Chemical Composition of Cu-Modified Polymers To confirm pDA binding to the polymer surface, the chemical structure of the coatings was studied with FTIR using a PerkinElmer Spectrum 65 FT-IR Spectrometer (PerkinElmer, Machelen, Belgium) in the 650–4000 cm−1 range; the spectra were obtained in a transmittance mode averaging eight scans with a resolution of 8 cm−1. Discs of Cu-modified polymers were placed on the sample holder and FTIR spectra were recorded.

2.4. Analysis of Copper Content by Inductively Coupled Plasma-Optical Emission Spectrometry (ICP-OES) A Perkin-Elmer Optima 2100 DV ICP-OES spectrometer was used for the copper content analysis. The experimental measurement conditions were as follows: wavelength 324.393 nm, sample flow rate 1.5 mL·min−1, sample uptake rate 1.0 mL·min−1, plasma gas flow rate 15 L·min−1, sample flow rate 1.5 mL·min−1, and operating power 1300 W. Discs of Cu-modified polymers were individually placed ◦ in 2 mL mixture of HCl:HNO3 (1:1 v/v, concentrated) and incubated at 50 C for 5 min. The obtained solution was cooled and filtered through a glass filter in order to separate from undissolved PU parts. The filtrate with concentrated acids was diluted with DI water to make 5 mL. Standards required for the calibration curve were prepared by serial dilution of the standard 1000 ppm copper solution. Blank and copper standard solutions were nebulized into plasma, where all components were vaporized. Blank and copper samples were dissociated and excited, and then the intensity of the radiation emitted by each sample was measured at a wavelength of 324.393 nm.

2.5. X-ray Photoelectron Spectroscopy (XPS) Analysis of the Chemical Composition of the Surface of Cu-Modified Polymeric Samples The surface chemical composition of Cu-modified polymers was analysed with XPS using an ESCALAB 250 Xi system (Thermo Scientific, Waltham, MA, USA) equipped with a monochromated Al Kα X-ray source and based at the Surface Analysis Laboratory, University of Brighton, UK. Uniform charge neutralization was provided by beams of low-energy (≤10 eV) Ar+ ions and low-energy electrons guided by the magnetic lens. The standard analysis spot of ca. 900 × 600 µm2 was defined by the microfocused X-ray source. Full survey scans (step size 1 eV, pass energy 200 eV, dwell time 50 ms and 5 scans) and narrow scans (step size 0.1 eV, pass energy 40 eV, dwell time 100 ms and 15 scans) of the Cu2p (binding energy, BE ~935 eV), N1s (BE ~399 eV), C1s (BE ~285 eV) and O1s (BE ~531 eV) regions were acquired from three separate areas on each sample. Data were transmission function corrected and analysed with Thermo Avantage Software (Version 5.952, Thermo Fisher Scientific) using the Smart background. Discs of Cu-modified polymers were glued on sample holder and placed in the instrument chamber.

2.6. Evaluation of Copper Leaching from Polymers Studied by Inductively Coupled Plasma-Mass Spectrometry (ICP-MS) Copper leaching from Cu-modified polymers was measured every 24 h for five days by ICP-MS 7900 (Agilent, Tokyo, Japan). The discs of Cu-modified polymers were individually placed into flat bottom vials, filled with 4 mL of PBS at RT and incubated for five days while shaking. Every 24 h polymeric discs were removed and placed into new vials containing 4 mL of fresh PBS. Then solutions from each incubation time (each vial) were transferred to the centrifuge flasks and 50 µL of concentrated nitric acid was added; then the resulting solutions were diluted up to 5 mL with PBS.

2.7. Nitric Oxide (NO) Release Measurements The NO release catalysed by Cu-modified polymers was examined using the Arrowstraight™ − − Nitric Oxide (NO2 /NO3 ) measurement system (Lazar Research Laboratories, Los Angeles, CA, USA) with micro ion-selective electrodes for independent measurement of both nitrite and nitrate ions. The GSNO stock solution of 500 µM in PBS was freshly prepared each time and its concentration Colloids Interfaces 2019, 3, 33 5 of 13 was verified spectrophotometrically by measuring the absorption of the SNO group at 335 nm using Colloids Interfaces 2019, 3, x FOR PEER REVIEW 5 of 13 an extinction coefficient of 922 M−1 cm−1 [28]. Stock solution of GSH in PBS (500 µM) was stored in averified fridge forspectrophotometrically few months. The by polymer measuring discs the wereabsorption placed of atthe the SNO bottom group at of 335 a 24-well nm using plate an and 0.5 mLextinction of PBS coefficient solution of was 922 added M−1 cm to−1 [28]. each Stock well solution to equilibrate of GSH polymersin PBS (500 for μM) 5 minwas stored in PBS in prior a to addingfridge the for reagents. few months. Afterwards, The polymer GSH discs and were GSNO placed stock at the solutions, bottom of botha 24-well of 500 plateµ M,and were 0.5 mL added of to eachPBS well solution (sample) was in added the volume to each to well get to the equilibrate final concentration polymers for of 5 both, min in GSH PBS and prior GSNO to adding of 100 theµ M in the resultingreagents. Afterwards, volume of 1GSH mL and solution. GSNO Polymers stock solutions, samples both were of 500 incubated μM, were in added these to solutions each well for 1 h at RT(sample) with shaking. in the volume After incubation,to get the final 200 concentrationµL of solution of both, from GSH each and well GSNO was injectedof 100 μM in in the the system resulting volume− of 1 mL− solution. Polymers samples were incubated in these solutions for 1 h at RT to measure NO2 and NO3 concentrations. The catalytic decomposition of GSNO and quantification with shaking. After incubation, 200 μL of solution from each well was injected− in the− system to of NO release was calculated by summing up the concentrations of NO and NO in the resulting measure NO and NO concentrations. The catalytic decomposition of GSNO2 and quantification3 of solutions. As a standard, the serial dilutions of 0.1 M sodium nitrite and 0.1 M sodium nitrate solutions NO release was calculated by summing up the concentrations of NO and NO in the resulting weresolutions. used. NO As measurements a standard, the were serial performed dilutions accordingof 0.1 M sodium to the manufacturer’snitrite and 0.1 M instructions sodium nitrate using the softwaresolutions provided. were Allused. measurements NO measurements were performedwere performed in triplicate. according During to the the NOmanufacturer’s measurements, all solutionsinstructions containing using the GSNOsoftware were provided. shielded All measurements from light by were foil asperformed light greatly in triplicate. decreases During the the half-life of GSNO.NO measurements, all solutions containing GSNO were shielded from light by foil as light greatly decreases the half-life of GSNO. 3. Results and Discussion 3. Results and Discussion 3.1. Chemical Structure of Cu-Modified Polymers 3.1. Chemical Structure of Cu-Modified Polymers A series of copper-modified polymers with different DA:Cu2+ molar ratios were prepared A series of copper-modified polymers with different DA:Cu molar ratios were prepared (Figure1). This was done to find the optimal concentration of copper on the surface of Cu-modified (Figure 1). This was done to find the optimal concentration of copper on the surface of Cu-modified polymers. Previously it was established that catecholamines, and in particular pDA, form stable polymers. Previously it was established that catecholamines, and in particular pDA, form stable metal-pDAmetal-pDA complexes complexes [32 [32]] and and the the molar molar ratioratio of of dopamine dopamine to tometal metal salt salt in the in thesolution solution should should be be betweenbetween 2:1 and2:1 and 3:1 3:1 to to ensure ensure full full incorporation incorporation of of metal metal into into the the pDA pDA [33]. [33 A]. higher A higher ratioratio of copper of copper ionsions in the in incubationthe incubation solution solution leads leads to to aggregationaggregation of of pDa pDa in in solution. solution. The The polymer polymer samples samples with with differentdifferent DA:Cu DA:2Cu+ molar molar ratios ratios were were preparedprepared (Figure (Figure 1).1).

FigureFigure 1. ( Left1. (Left)—Photographs)—Photographs of non-modified non-modiﬁed poly(vinyl poly(vinyl chloride) chloride) (PVC) (PVC) and polyurethane and polyurethane (PU) (PU) (upper(upper row) row) and and Cu-modiﬁed Cu-modified polymers polymers (lower (lower rows) rows) obtained obtained viavia incubationincubation of of polymers polymers in in in in Tris- Tris-HCl bufferHCl with buffer different with different DA:Cu 2DA:+ molarCu ratios;molar ratios; (Right (Right)—methods)—methods of analysis of analysis used used to determineto determine copper copper content and NO generation by the polymers. content and NO generation by the polymers.

TheThe results results of FTIR of FTIR analysis analysis of Cu-modified of Cu-modified samples samples are shown are shown in Figure in Figure2. Spectra 2. Spectra of unmodified of PVCunmodified and PU are PVC typical and PU of are these typical polymers of these as polymers reported as reported in literature. in literatur Theire. Their assignation assignation is given is in −1 given in the legend to Figure 2. In the spectrum of PVC/pDA at 3333 cm− 1and in the region between the legend to Figure−1 2. In the spectrum−1 of PVC/pDA−1 at 3333 cm and in the region between 1500–1600− cm1 new peaks at 1600 cm−1 and 1515 cm − appeared,1 as shown in Figure 2a. These peaks 1500–1600can be cmattributednew to peaks pDA atas 1600 reported cm inand [31]. 1515 Alth cmough theappeared, chemical as structure shown inof FigurepDA is2 stilla. These under peaks can bediscussion, attributed it is to generally pDA as accepted reported that in [31it contains]. Although indole the and/or chemical indoline structure units [29]. of pDA Accordingly, is still under discussion,these peaks it is were generally assigned accepted to the overlapping that it contains C=C indolestretching and/or vibrations indoline from unitsindole [ 29ring]. Accordingly,and, by these peaks were assigned to the overlapping C=C stretching vibrations from indole ring and, Colloids Interfaces 2019, 3, 33 6 of 13 by different authors, to N–H scissoring vibrations or C=N vibrations thus presenting evidence of −1 pDAColloids formation Interfaces 2019 [34]., 3, A x FOR peak PEER at 333REVIEW cm in the spectrum of Cu-modified PVC was attributed 6 of to 13 N–H vibrations. The peaks at 2867 cm−1, 2850 cm−1 and 2817 cm−1 in the FTIR spectrum of non-modified PVCdifferent were assigned authors, to N C–H–H scissoring stretching vibrations vibrations or (FigureC=N vibratio2a, linens 1).thus presenting evidence of pDA formation [34]. A peak at 333 cm−1 in the spectrum of Cu-modified PVC was attributed to N–H The appearance of these peaks confirmed the presence of pDA film on the PVC surface. In the vibrations. The peaks at 2867 cm−1, 2850 cm−1 and 2817 cm−1 in the FTIR spectrum of non-modified spectrum of Cu-modified PVC (Figure2a) the band at 1600 cm −1 shifted to higher wavenumbers PVC were assigned to C–H stretching vibrations (Figure 2a, line 1). appearing at 1674 cm−1 and its intensity was decreased. The band at 1515 cm−1 split into two bands The appearance of these peaks confirmed the presence of pDA film on the PVC surface. In the −1 −1 at 1493spectrum cm ofand Cu-modified 1562 cm PVC. Such (Figure split 2a) indicates the band metal at 1600 binding cm−1 withshifted polydopamine to higher wavenumbers [35]. The FTIR −1 −1 spectraappearing of Cu-modified at 1674 cm−1 and PU its are intensity shown was in Figure decreased.2b. The band peaks at centred1515 cm− at1 split 3333 into cm two bands, 2955 at cm , −1 −1 −1 −1 −1 28741493 cm cm−,1 1728and 1562 cm cm, 1701−1. Such cm split, 1531indicates cm metalare binding peaks originating with polydopamine from the [35]. PU. The Peaks FTIR at spectra 3333 cm , − − − 2954of cmCu-modified1, 2874 cm PU are1 are shown from in N–H Figure and 2b. C–H The peaks groups; centred the band at 3333 at cm 1728−1, 2955 cm cm1 is−1, due 2874 to cm the−1, 1728 carbonyl groupscm−1, in1701 urethane cm−1, 1531 bonds cm−1 (C=O);are peaks the originating band centred from the at 1701 PU. Peaks cm−1 atis 3333 attributed cm−1, 2954 to carbonylcm−1, 2874 groups cm−1 in ureaare bond; from theN–H 1531 and cmC–H−1 groups;band was the band assigned at 1728 to secondarycm−1 is due to amide the carbonyl (RCONHR’). groups The in urethane FTIR spectra bonds of PU modified(C=O); the by band PDA centred and Cu at did 1701 not cm provide−1 is attributed clear to evidence carbonyl of groups the presence in urea bond; of PDA the 1531 and cm copper−1 band on the modifiedwas assigned PU surface to secondary because amide PU matrix (RCONHR’). bands overlappedThe FTIR spectra with of PDA PU modified peaks which by PDA have and absorbance Cu did in thenot same provide region. clear evidence of the presence of PDA and copper on the modified PU surface because PU matrix bands overlapped with PDA peaks which have absorbance in the same region.

(b) (a)

(c)

FigureFigure 2. 2.(a )(a Infrared) Infrared spectra spectra ofof PVCPVC discs: discs: (1) (1) non-modi non-modified;fied; (2) (2) modified modified with with PDA PDA and and(3) modified (3) modified withwith Cu-PDA; Cu-PDA; (b ()b) Infrared Infrared spectraspectra of PU discs: discs: (1) (1) non- non-modified;modified; (2) (2)modified modified with with PDA PDAand (3) and (3) modified with Cu-PDA; (c) A diagram of appearing and shifting absorbance bands in Fourier- modified with Cu-PDA; (c) A diagram of appearing and shifting absorbance bands in Fourier-transform transform infrared spectroscopy (FTIR) spectra of PVC, PVC/PDA and PVC/PDA/Cu. infrared spectroscopy (FTIR) spectra of PVC, PVC/PDA and PVC/PDA/Cu.

Colloids Interfaces 2019, 3, 33 7 of 13

The characteristic peaks of pDA appeared in the FTIR spectra after modification confirms the successful modification by pDA. The shift peak at 1600 cm−1 to higher wavenumbers and split of the band at 1515 cm−1 into two bands at 1491 cm−1 and 1562 cm−1 indicates the formation of a Cu-PDA complex as observed previously for Fe-PDA and Zn-PDA complexes [8,36]. It was reported that the amount of 13 nmol of Cu2+ immobilized on a modified PU was sufficient to produce physiologically relevant levels of NO in 2 mL of 10 µM solution of GSNO/GSH in PBS [15]. The amount of copper in the coatings was estimated using ICP-OES. The results are shown in Table1. The highest copper content was found in the samples obtained at DA:Cu 2+ molar ratio 3.5:1, being 3.86 nmol cm−2 and 6.04 nmol cm−2 for Cu-modified PVC and PU, respectively. Subsequently, the samples obtained at DA:Cu2+ molar ratio 3.5:1 were chosen for further investigation.

Table 1. Copper content on the surface of PVC and PU modiﬁed by dopamine from solutions with different DA:Cu2+ molar ratios according to ICP-OES.

Sample Cu, nmol cm−2 DA: Cu2+ Molar Ratio PVC/pDA/Cu 3.86 ± 0.30 3.5:1 PVC/pDA/Cu 1.35 ± 0.28 1.8:1 PVC/pDA/Cu 0.96 ± 0.30 0.9:1 PU/pDA/Cu 6.04 ± 0.17 3.5:1 PU/pDA/Cu 1.97 ± 0.42 1.8:1 PU/pDA/Cu 1.4 ± 0.70 0.9:1

XPS analysis was performed to detect copper in the coating along with finding the valence states of individual elements to better understand about their chemical surroundings and the mechanism of attachment to the polymer surface. In Figure3, the narrow scan spectra of Cu2p, O1s, C1s and N1s of Cu-modified PVC surface are shown. The two peaks observed for the binding energy (BE) of copper with maxima at 955 eV and 935 eV can be attributed to Cu 2p1/2 orbital and Cu 2p3/2 orbitals respectively, and peaks around 943 and 964 eV are the satellite peaks, which are common to Cu2p XPS data. The position of the main Cu 2p3/2 peak ca. 933–934 eV and the satellite peak ca. 942–943 eV is typical of Cu(II) catecholate complexes [37]. Deconvoluted Cu2p3/2 narrow scan data for four representative systems are shown in Figure S1. Cu is present in both of its chemical states of + +2 Cu and Cu , with Cu2O, CuO and Cu(OH)2. All peaks are assigned with references under NIST database. The XPS data suggest that Cu-pDA was successfully immobilized on the surface of PVC and PU, which are consistent with the results of FTIR and ICP-OES studies. XPS of C1s and N1s also confirm the presence of C–N bonds, i.e., pDA and XPS of O1s indicate the presence of oxygen atoms in the state similar to that in Cu–O bonds. The presence of Cu(I) species in the surface layer is likely to be the result of a redox reaction between pDA and Cu(II). The atomic fraction of copper element in the surface coating of Cu-modified PVC is 1.5% (Table2), which is close to the reported 1.4% on polysulfone and 1.36% on polyvinydienefluoride ultrafiltration membranes [38]. In case of Cu modified PU, there is much less Cu detected than expected from other experimental results. It could be due to various reasons, but most possible reason could be that majority of the Cu is located below 10 nm of top surface layer, making it relatively unquantifiable using XPS and the other reason could be possible adventitious organic layer on the top. Colloids Interfaces 2019, 3, 33 8 of 13 Colloids Interfaces 2019, 3, x FOR PEER REVIEW 8 of 13

FigureFigure 3. XPS 3. XPS narrow narrow spectra spectra of of PVC/pDA/Cu PVC/pDA/Cu disc disc surface; surface; (a () aCu2p,) Cu2p, (b) ( bO1s,) O1s, (c) (C1sc) C1s and and (d) N1s. (d) N1s. ReferencesReferences to individual to individual peak peak assignments assignments are are shownshown in supporting information information table table SI-T1. SI-T1.

UsingUsing XPS, XPS, we we evaluated evaluated the the leachingleaching ofof copper from from Cu-modified Cu-modified polymers polymers to 100 to 100μM µM GSNO/GSHGSNO/GSH solution solution during during 1 1 h h (Table (Table2). 2). Copper Copper content content of of PVC/pDA/Cu PVC/pDA/Cu and and PU/pDA/Cu PU/pDA/Cu decreaseddecreased from from 1.5 1.5 to to 1.1 1.1 atomic atomic %% and from from 0.54 0.54 to to 0.44 0.44 atomic atomic %, respectively, %, respectively, after afterincubation incubation in in GSNO/GSHGSNO/GSH solution. solution. A decrease A decrease in surface in surface carbon, carbon, nitrogen nitrogen and oxygen and contents oxygen should contents be observed should be observedin case in ofcase the detachment of the detachment of PDA. The of PDA. XPS results The XPS show results that the show carbon that content, the carbon O/C and content, N/C atomic O/C and ratio of initial PVC/pDA/Cu are 72.4, 0.22 and 0.08, respectively, whereas the carbon content after N/C atomic ratio of initial PVC/pDA/Cu are 72.4, 0.22 and 0.08, respectively, whereas the carbon incubation in the 100 μM GSNO/GSH solution reduced to 67.8 atomic %, and the atomic ratios of O/C content after incubation in the 100 µM GSNO/GSH solution reduced to 67.8 atomic %, and the atomic and N/C of PVC/pDA/Cu increased to 0.27 and 0.10, which indicated that chemical reactions took ratiosplace of O/Cduring and incubation N/C of PVC/pDA/Cuof the sample in increasedGSNO/GSH. to 0.27The andsame 0.10, trend which similar indicated to PVC/pDA/Cu that chemical is reactionsobserved took for place PU/pDA/Cu. during incubationDespite the ofwidespread the sample application in GSNO/GSH. of pDA coatings The same for trendbiomaterials similar to PVC/pDA/Cudevelopment, is its observed structure for is PU/pDA/Cu. still under discussion. Despite It the is widespreadknown that applicationGSNO exhibits of pDA oxidizing coatings for biomaterialsproperties, so development, the increase in its the structure O/C ratio is could still under be explained discussion. by the It issurface known oxidation, that GSNO but exhibitsit is oxidizingdifficult properties, to determine so which the increase groups inwere the oxidized. O/C ratio Similarly, could bethe explainedreduction in by carbon the surface content oxidation, could but itbe is explained difficult toby determinethe surface whichoxidation, groups however were it oxidized. requires further Similarly, investigation. the reduction On the in carbonother hand, content couldthe be pDA-coated explained bymaterial the surface used in oxidation, the catalytic however oxidation it requires in the reaction further investigation.medium containing On the 30% other hand,aqueous the pDA-coated hydrogen peroxide material exhibited used in the good catalytic stability oxidation [39]. Moreover, in the reaction the Michael medium addition containing reaction 30% aqueousbetween hydrogen pDA and peroxide glutathione exhibited (GSH) goodcan occur stability due to [39 the]. Moreover,ability of pDA the to Michael react with addition amine- reaction and thiol-containing molecules, which could also explain the increase in O/C and N/C ratios. between pDA and glutathione (GSH) can occur due to the ability of pDA to react with amine- and thiol-containingTable 2. Atomic molecules, composition which couldof the Cu-modified also explain PV theC before increase and in after O/C incubation and N/C with ratios. 100 μM GSNO/L-glutathione reduced (GSH) solution during 1 h obtained by XPS. Table 2. Atomic composition of the Cu-modified PVC before and after incubation with 100 µM

GSNO/L-glutathione reduced (GSH)PVC/pDA/Cu solution during 1 h obtained byPU/pDA/Cu XPS. Elements Before After Before After N1s 5.6 PVC/pDA/Cu6.9 PU/pDA/Cu3.23 3.82 C1s 72.4 67.8 75.61 66.26 O1s Elements15.7 Before After18.5 Before20.14 After 25.08 Cl2p N1s4.4 5.6 6.94.7 3.230.11 3.82 1.75 Na1s C1s0.1 72.4 67.80.7 75.610.37 66.26 2.65 Cu2p3 O1s1.5 15.7 18.51.1 20.140.54 25.08 0.44 N/C ratio Cl2p 0.08 4.4 4.7 0.1 0.11 0.04 1.75 0.06 O/C ratio Na1s 0.22 0.1 0.7 0.27 0.37 0.27 2.65 0.38 Cu2p3 1.5 1.1 0.54 0.44 N/C ratio 0.08 0.1 0.04 0.06 O/C ratio 0.22 0.27 0.27 0.38 Colloids Interfaces 2019, 3, 33 9 of 13

Colloids Interfaces 2019,, 3,, xx FORFOR PEERPEER REVIEWREVIEW 9 9 of of 13 13 Leaching of Cu from Cu-modified samples to PBS during 5 days was evaluated. Cumulative · −2 leachingLeaching from of Cu-modified Cu from Cu-modified PVC for 5 dayssamples was to 1.51 PBS nmol duringcm 5 ,days for Cu-modifiedwas evaluated. PU Cumulative was equal · −2 toleachingleaching 1.41 nmol fromfromcm Cu-modifiedCu-modified(Figure 4PVCPVC). That forfor is55 daysdays about waswas 39% 1.511.51 of nmolnmol Cu content··cm−2,, forfor on Cu-modifiedCu-modified Cu-modified PUPU PVC waswas and equalequal 23% toto 1.411.41 on Cu-modifiednmol··cm−−22 (Figure(Figure PU. 4).4). ThatThat isis aboutabout 39%39% ofof CuCu contencontent on Cu-modified PVC and 23% on Cu-modified PU.

Figure 4.4. ICP-MS datadata ofof coppercopper leachedleached fromfrom thethe surfacesurface ofof Cu-modiﬁedCu-modified polymerspolymers duringduring 55 days.days.

TheThe normalnormal levellevel of free coppercopper in the human blood serum is 1.6 1.6–2.4–2.4 μµM or 100 100–150–150 ppb [40]. [40]. TheThe highesthighest measuredmeasured Cu Cu leaching leaching level level from from samples samples was was recorded recorded at at 50.7 50.7 ppb ppb in in ﬁve five days days (Table (Table3), which3), which is well is well below below cytotoxic cytotoxic concentrations concentrations for for mammalian mammalian cells. cells. It was It was reported reported that that the the viability viability of mammalianof mammalian cells cells was was not not affected affected until until the the copper coppe concentrationr concentration reached reached 1000 1000 ppb ppb[ [41].41]. OurOur resultsresults suggestsuggest thatthat coppercopper leachingleaching fromfrom thethe samplessamples shouldshould posepose nono riskrisk toto health.health.

Table 3.3. CumulativeCumulative CuCu leachingleaching fromfrom thethe samplessamples duringduring 55 daysdays determineddetermined byby ICP-MSICP-MS data.data.

SampleSample 55 Days, Days, ppb ppb 5 Days, 5 Days, % % from from Original Original PVC/pDA/CuPVC/pDA/Cu (3.5:1) (3.5:1) 50.7 50.7± ±14.5 14.5 39 39 PU/pDA/CuPU/pDA/Cu (3.5:1) (3.5:1) 48.1 48.1± ±11.6 11.6 23 23

3.2.3.2. MeasurementMeasurement of of NO NO Generation Generation Catalyzed Catalyzed by Modified by Modified Materials Materials + AccordingAccording toto XPSXPS datadata coppercopper onon thethe surfacesurface ofof Cu-modifiedCu-modifiedCu-modified polymerspolymers existsexistsexists inin thethe form of CuCu2 + + andand CuCu.. TheThe actual actual catalystcatalyst ofofof GSNOGSNOGSNO decompositiondecompositiondecomposition isisis CuCu... However, However, thethethe catalyticcatalyticcatalytic mechanismmechanismmechanism + + includesincludesincludes CuCuCu redox-cycleredox-cycleredox-cycle betweenbetweenbetween thethethe CuCu2 andandand CuCu.. The The NONO generatinggeneratinggenerating mechanismmechanismmechanism involvesinvolvesinvolves thethethe + decompositiondecomposition of GSNOGSNO byby CuCu ions, ions, ions, whichwhichwhich partiallypartiallypartially existexist inin thethethe samplesamplesample surfacesurfacesurface andand alsoalsoalso formformform bybyby reductionreduction ofof CuCu2+ ionsionsions bybyby glutathioneglutathioneglutathione (GSH)(GSH)(GSH) (Figure(Figure(Figure5 5)5))[ [35].[35].35]. TheTheThe tracestracestraces amountsamountsamounts ofofof CuCu+ affectaffectaffect thethethe catalysis.catalysis. TheseThese reactionsreactions proceedsproceeds untiluntil GSNOGSNO depletiondepletion andand itsits conversionconversion toto disulfide.disulfide.

FigureFigure 5.5. TheThe schematicschematic presentationpresentation of of the the mechanism mechanism of of nitric nitricnitric oxide oxideoxide (NO) (NO)(NO) generation generationgeneration from fromfrom GSNO GSNOGSNO in theinin thethe presence presencepresence of GSHofof GSHGSH in phosphate-bufferedinin phphosphate-buffered saline saline (PBS). (PBS).

NO generation ability of Cu-containing samples was studied using GSNO as the nitric oxide donor, because GSNO is a compound relatively stable to self-decomposition and has long half-life.

Colloids Interfaces 2019, 3, 33 10 of 13 Colloids Interfaces 2019, 3, x FOR PEER REVIEW 10 of 13

TheColloids NO Interfaces release generation 2019 from, 3,ability xS FOR-nitrosoglutathione PEER of Cu-containing REVIEW catalyzed samples by was Cu-polymer studiedusing samples GSNO was asfound the nitricto be within 10oxide of 13 donor,the range because (0.43− GSNO1.4) × 10 is−10 a mol compound cm min relatively (Figure stable 6). These to self-decomposition data are within the and physiological has long half-life. level of The NO release from S-nitrosoglutathione catalyzed by Cu-polymer samples was found to be within The NO release from S-nitrosoglutathione catalyzed by−10 Cu-polymer samples was found to be within NO release by endothelial cells, which is (0.5−4) × 10 mol cm min [4]. thethe range range (0.43 (0.43−1.4)1.4) × ×1010−10 −mol10 mol cm ·cm min−2·min (Figure−1 (Figure 6). These6). These data are data within are within the physiological the physiological level of levelNO ofrelease NO releaseby endothelial by endothelial cells, which cells, whichis (0.5− is4) (0.5× 10−−104) mol× cm10−10 minmol·cm [4].−2 ·min−1 [4].

FigureFigure 6.6. NONO generationgeneration afterafter incubationincubation ofofCu-modified Cu-modifiedpolymers polymerswith with100 100µ μMM GSNO/GSH GSNO/GSH inin thethe PBS during 1 h. PBSFigure during 6. NO 1 h. generation after incubation of Cu-modified polymers with 100 μM GSNO/GSH in the PBS during 1 h. ItIt waswas reportedreported thatthat GSNOGSNO isis alsoalso ableable toto releaserelease NONO inin thethe presencepresence ofof GSHGSH withoutwithout Cu [[42],42], but both endogenous and synthetic RSNOs are prone to catalytic decomposition by copper ions but bothIt was endogenous reported and that synthetic GSNO is RSNOs also able are to prone release to catalytic NO in the decomposition presence of byGSH copper without ions Cu [17 ,[42],32]. [17,32]. We have shown that Cu-pDA modified PVC and PU can catalyse NO generation in the Webut have both shown endogenous that Cu-pDA and synthetic modified RSNOs PVC and are PUprone can catalyseto catalytic NO decomposition generation in theby presencecopper ions of presence of GSNO. The highest NO generation rate was exhibited by the Cu-modified PU which GSNO.[17,32].The We highesthave shown NO generation that Cu-pDA rate modified was exhibited PVC and by thePU Cu-modifiedcan catalyse PUNO whichgeneration correlated in the correlated with higher copper content in the polymer in comparison with Cu-modified PVC withpresence higher of copperGSNO. contentThe highest in the NO polymer generation in comparison rate was exhibited with Cu-modified by the Cu-modified PVC according PU which to according to ICP-OES analysis. ICP-OEScorrelated analysis. with higher copper content in the polymer in comparison with Cu-modified PVC According to the kinetics of NO generation, about 35% of GSNO was catalytically decomposed accordingAccording to ICP-OES to the kinetics analysis. of NO generation, about 35% of GSNO was catalytically decomposed by by PVC/pDA/Cu and 25% by PU/pDA/Cu within the first hour (Figure 7). PVC/pDA/CuAccording and to the 25% kinetics by PU/pDA/Cu of NO generation, within the about first 35% hour of (Figure GSNO7 ).was catalytically decomposed by PVC/pDA/Cu and 25% by PU/pDA/Cu within the first hour (Figure 7).

µ FigureFigure 7. 7. NONO generation generation after after incubation incubation of Cu-modified of Cu-modiﬁed polymers polymers with with100 μ 100M GSNO/GSHM GSNO/GSH in the PBS. in the PBS. Figure 7. NO generation after incubation of Cu-modified polymers with 100 μM GSNO/GSH in the PBS.

Colloids Interfaces 2019, 3, 33 11 of 13

4. Conclusions Cu-modified PVC and PU polymers for catalytic NO generation were produced using a one-step preparation method. The presence of Cu on both modified polymers was proved by ICP-OES and XPS data. According to ICP-OES measurements, the amount of Cu species on the surface of Cu-modified PU was higher than on the Cu-modified PVC. FTIR spectra showed the characteristic peaks of pDA and Cu complexes with pDA on the modified PVC polymer surface. The FTIR data did not give clear evidence of polydopamine presence and Cu complex formation on Cu-modified PU surface because of overlapping of the characteristic absorbance bands of the coatings with strong bands from PU matrix. It was found that a small amount of copper can leach from the modified polymer surfaces during five days, however it is within its physiological level in blood. The NO release test showed that Cu-modified PVC and PU polymers are capable of providing the physiological level of NO at physiological pH. The obtained data suggest that Cu-modified polymers potentially can produce nitric oxide in the blood from endogenous S-nitrosothiols. Taking into account the catalytic mechanism of NO generation, this process could continue long enough while in contact with blood to secure the long-term haemocompatibility of medical implants.

Supplementary Materials: The following are available online at http://www.mdpi.com/2504-5377/3/1/33/s1, Figure S1: Cu2p3/2 narrow scan XPS data of four samples, Cu modified PVC and PU with PDA, before and after incubation. Corresponding peaks are assigned following NIST database. Author Contributions: Conceptualization, L.M. and S.M.; Methodology, L.A. and L.M.; Investigation, L.A., S.R. and L.M.; Writing—Original Draft Preparation, L.A.; Writing—Review & Editing, L.M. and S.M.; Visualization, L.A. and S.R.; Supervision, L.M.; Project Administration, L.A.; Funding Acquisition, L.M. and S.M. Funding: This research was funded by the Marie Skłodowska-Curie Action of the European Union H2020-MSCA-IF-2016, grant number 749207. Acknowledgments: This work is supported by the Marie Skłodowska-Curie Action of the European Union (H2020-MSCA-IF-2016, grant agreement No. 749207). The authors acknowledge the DelStar Company for donating PU films and Peter Lyons and the Magdalena Grove at School of Environment and Technology, University of Brighton, for ICP-OES and ICP-MS measurements. Conflicts of Interest: The authors declare no conflict of interest.

References

1. Dumitriu, S. Polymeric Biomaterials, Revised and Expanded; CRC Press: Boca Raton, FL, USA, 2001. 2. Bajpai, A.K.; Bajpai, J.; Saini, R.K.; Agrawal, P.; Tiwari, A. Smart Biomaterial Devices: Polymers in Biomedical Sciences; CRC Press: Boca Raton, FL, USA, 2016. 3. Yang, Y.; Qi, P.K.; Yang, Z.L.; Huang, N. Nitric oxide based strategies for applications of biomedical devices. Biosurf. Biotribol. 2015, 1, 177–201. [CrossRef] 4. de Mel, A.; Murad, F.; Seifalian, A.M. Nitric oxide: A guardian for vascular grafts? Chem. Rev. 2011, 111, 5742–5767. [CrossRef][PubMed] 5. Naghavi, N.; Mel, A.D.; Alavijeh, O.S.; Cousins, B.G.; Seifalian, A.M. Nitric oxide donors for cardiovascular implant applications. Small 2013, 9, 22–35. [CrossRef][PubMed] 6. Pant, J.; Goudie, M.J.; Hopkins, S.P.; Brisbois, E.J.; Handa, H. Tunable nitric oxide release from s-nitroso-n-acetylpenicillamine via catalytic copper nanoparticles for biomedical applications. ACS Appl. Mater. Interfaces 2017, 9, 15254–15264. [CrossRef] 7. Wo, Y.; Brisbois, E.J.; Bartlett, R.H.; Meyerhoff, M.E. Recent advances in thromboresistant and antimicrobial polymers for biomedical applications: Just say yes to nitric oxide (no). Biomater. Sci. 2016, 4, 1161–1183. [CrossRef][PubMed] 8. Wu, H.; Ang, J.M.; Kong, J.; Zhao, C.; Du, Y.; Lu, X. One-pot synthesis of polydopamine–zn complex antifouling coatings on membranes for ultraﬁltration under harsh conditions. RSC Advances 2016, 6, 103390–103398. [CrossRef] 9. Damodaran, V.B.; Joslin, J.M.; Wold, K.A.; Lantvit, S.M.; Reynolds, M.M. S-nitrosated biodegradable polymers for biomedical applications: Synthesis, characterization and impact of thiol structure on the physicochemical properties. J. Mater. Chem. 2012, 22, 5990–6001. [CrossRef] Colloids Interfaces 2019, 3, 33 12 of 13

10. Riccio, D.A.; Schoenfisch, M.H. Nitric oxide release: Part I. Macromolecular scaffolds. Chem. Soc. Rev. 2012, 41, 3731–3741. [CrossRef] 11. Seabra, A.B.; Marcato, P.D.; de Paula, L.B.; Durán, N. New strategy for controlled release of nitric oxide. J. Nano Res. 2012, 20, 61–67. [CrossRef] 12. Kim, J.; Saravanakumar, G.; Choi, H.W.; Park, D.; Kim, W.J. A platform for nitric oxide delivery. J. Mater. Chem. B 2014, 2, 341–356. [CrossRef] 13. Dicks, A.P.; Swift, H.R.; Williams, D.L.H.; Butler, A.R.; Al-Sa’doni, H.H.; Cox, B.G. Identification of cu+ as the effective reagent in nitric oxide formation from s-nitrosothiols (rsno). J. Chem. Soc. Perkin Trans. 2 1996, 481–487. [CrossRef] 14. Puiu, S.C.; Zhou, Z.; White, C.C.; Neubauer, L.J.; Zhang, Z.; Lange, L.E.; Mansfield, J.A.; Meyerhoff , M.E.; Reynolds, M.M. Metal ion-mediated nitric oxide generation from polyurethanes via covalently linked copper(ii)-cyclen moieties. J. Biomed. Mater. Res. Part B Appl. Biomater. 2009, 91, 203–212. [CrossRef] [PubMed] 15. Hwang, S.; Meyerhoff, M.E. Polyurethane with tethered copper(ii)–cyclen complex: Preparation, characterization and catalytic generation of nitric oxide from s-nitrosothiols. Biomaterials 2008, 29, 2443–2452. [CrossRef][PubMed] 16. Luo, R.; Liu, Y.; Yao, H.; Jiang, L.; Wang, J.; Weng, Y.; Zhao, A.; Huang, N. Copper-incorporated collagen/catechol film for in situ generation of nitric oxide. ACS Biomater. Sci. Eng. 2015, 1, 771–779. [CrossRef] 17. Neufeld, M.J.; Lutzke, A.; Jones, W.M.; Reynolds, M.M. Nitric oxide generation from endogenous substrates using metal–organic frameworks: Inclusion within poly(vinyl alcohol) membranes to investigate reactivity and therapeutic potential. ACS Appl. Mater. Interfaces 2017, 9, 35628–35641. [CrossRef][PubMed] 18. Oh, B.K.; Meyerhoff, M.E. Spontaneous catalytic generation of nitric oxide from s-nitrosothiols at the surface of polymer films doped with lipophilic copper(ii) complex. J. Am. Chem. Soc. 2003, 125, 9552–9553. [CrossRef] 19. Nagababu, E.; Rifkind, J.M. Routes for formation of s-nitrosothiols in blood. Cell Biochem. Biophys. 2013, 67, 385–398. [CrossRef] 20. Zhang, Y.; Hogg, N. S-nitrosothiols: Cellular formation and transport. Free Radic. Biol. Med. 2005, 38, 831–838. [CrossRef] 21. Vanin, A.F.; Papina, A.A.; Serezhenkov, V.A.; Koppenol, W.H. The mechanisms of s-nitrosothiol decomposition catalyzed by iron. Nitric Oxide 2004, 10, 60–73. [CrossRef] 22. Peaston, R.T.; Weinkove, C. Measurement of catecholamines and their metabolites. Ann. Clin. Biochem. 2004, 41, 17–38. [CrossRef] 23. Liu, Y.; Ai, K.; Lu, L. Polydopamine and its derivative materials: Synthesis and promising applications in energy, environmental, and biomedical fields. Chem. Rev. 2014, 114, 5057–5115. [CrossRef][PubMed] 24. Zhang, C.; Gong, L.; Xiang, L.; Du, Y.; Hu, W.; Zeng, H.; Xu, Z.-K. Deposition and adhesion of polydopamine on the surfaces of varying wettability. ACS Appl. Mater. Interfaces 2017, 9, 30943–30950. [CrossRef][PubMed] 25. Mrówczyński,R.; Markiewicz, R.; Liebscher, J. Chemistry of polydopamine analogues. Polym. Int. 2016, 65, 1288–1299. [CrossRef] 26. Saiz-Poseu, J.; Mancebo-Aracil, J.; Nador, F.; Busqué, F.; Ruiz-Molina, D. The chemistry behind catechol-based adhesion. Angew. Chem. Int. Ed. 2019, 58, 696–714. [CrossRef] 27. Ding, Y.H.; Floren, M.; Tan, W. Mussel-inspired polydopamine for bio-surface functionalization. Biosurf. Biotribol. 2016, 2, 121–136. [CrossRef] 28. Huang, N.; Zhang, S.; Yang, L.; Liu, M.; Li, H.; Zhang, Y.; Yao, S. Multifunctional electrochemical platforms based on the michael addition/schiff base reaction of polydopamine modified reduced graphene oxide: Construction and application. ACS Appl. Mater. Interfaces 2015, 7, 17935–17946. [CrossRef] 29. Liebscher, J.; Mrówczyński,R.; Scheidt, H.A.; Filip, C.; Hădade, N.D.; Turcu, R.; Bende, A.; Beck, S. Structure of polydopamine: A never-ending story? Langmuir 2013, 29, 10539–10548. [CrossRef] 30. Ryu, J.H.; Messersmith, P.B.; Lee, H. Polydopamine surface chemistry: A decade of discovery. ACS Appl. Mater. Interfaces 2018, 10, 7523–7540. [CrossRef] 31. Dreyer, D.R.; Miller, D.J.; Freeman, B.D.; Paul, D.R.; Bielawski, C.W. Elucidating the structure of poly(dopamine). Langmuir 2012, 28, 6428–6435. [CrossRef] Colloids Interfaces 2019, 3, 33 13 of 13

32. Szpoganicz, B.; Gidanian, S.; Kong, P.; Farmer, P. Metal binding by melanins: Studies of colloidal dihydroxyindole-melanin, and its complexation by cu(ii) and zn(ii) ions. J. Inorg. Biochem. 2002, 89, 45–53. [CrossRef] 33. Wang, Z.; Xie, Y.; Li, Y.; Huang, Y.; Parent, L.R.; Ditri, T.; Zang, N.; Rinehart, J.D.; Gianneschi, N.C. Tunable, metal-loaded polydopamine nanoparticles analyzed by magnetometry. Chem. Mater. 2017, 29, 8195–8201. [CrossRef] 34. Zangmeister, R.A.; Morris, T.A.; Tarlov, M.J. Characterization of polydopamine thin films deposited at short times by autoxidation of dopamine. Langmuir 2013, 29, 8619–8628. [CrossRef] 35. Zhang, C.; Biggs, T.D.; Devarie-Baez, N.O.; Shuang, S.; Dong, C.; Xian, M. S-nitrosothiols: Chemistry and reactions. Chem. Commun. 2017, 53, 11266–11277. [CrossRef][PubMed] 36. Sever, M.J.; Weisser, J.T.; Monahan, J.; Srinivasan, S.; Wilker, J.J. Metal-mediated cross-linking in the generation of a marine-mussel adhesive. Angew. Chem. Int. Ed. 2004, 43, 448–450. [CrossRef][PubMed] 37. Brown, D.G.; Hughes, W.J.; Knerr, G. Electronic and x-ray photoelectron spectra of copper catecholate complexes. Inorg. Chim. Acta 1980, 46, 123–126. [CrossRef] 38. Liu, Z.; Hu, Y.; Liu, C.; Zhou, Z. Surface-independent one-pot chelation of copper ions onto filtration membranes to provide antibacterial properties. Chem. Commun. 2016, 52, 12245–12248. [CrossRef][PubMed] 39. Ye, W.; Shi, X.; Su, J.; Chen, Y.; Fu, J.; Zhao, X.; Zhou, F.; Wang, C.; Xue, D. One-step reduction and functionalization protocol to synthesize polydopamine wrapping ag/graphene hybrid for efficient oxidation of hydroquinone to benzoquinone. Appl. Catal. B Environ. 2014, 160–161, 400–407. [CrossRef] 40. Brewer, G.J. Wilson’s disease. In Harrison’s Principles of Internal Medicine, 18th ed.; Longo, D., Fauci, A., Kasper, D., Hauser, S., Jameson, J., Loscalzo, J., Eds.; McGraw-Hill Education: New York, NY, USA, 2011; pp. 3188–3190. 41. Valodkar, M.; Rathore, P.S.; Jadeja, R.N.; Thounaojam, M.; Devkar, R.V.; Thakore, S. Cytotoxicity evaluation and antimicrobial studies of starch capped water soluble copper nanoparticles. J. Hazard. Mater. 2012, 201–202, 244–249. [CrossRef] 42. Singh, S.P.; Wishnok, J.S.; Keshive, M.; Deen, W.M.; Tannenbaum, S.R. The chemistry of the s-nitrosoglutathione/glutathione system. Proc. Natl. Acad. Sci. USA 1996, 93, 14428–14433. [CrossRef]

© 2019 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).