Hybrid Antifouling and Antimicrobial Coatings ... - ACS Publications

Subscriber access provided by UNIV OF NEBRASKA - LINCOLN

Article

Hybrid Antifouling and Antimicrobial Coatings Prepared by Electroless Codeposition of Fluoropolymer and Cationic Silica Nanoparticles on Stainless Steel: Efficacy Against L. monocytogenes Kang Huang, Juhong Chen, Sam R. Nugen, and Julie M. Goddard ACS Appl. Mater. Interfaces, Just Accepted Manuscript • DOI: 10.1021/acsami.6b04187 • Publication Date (Web): 06 Jun 2016 Downloaded from http://pubs.acs.org on June 12, 2016

Just Accepted “Just Accepted” manuscripts have been peer-reviewed and accepted for publication. They are posted online prior to technical editing, formatting for publication and author proofing. The American Chemical Society provides “Just Accepted” as a free service to the research community to expedite the dissemination of scientific material as soon as possible after acceptance. “Just Accepted” manuscripts appear in full in PDF format accompanied by an HTML abstract. “Just Accepted” manuscripts have been fully peer reviewed, but should not be considered the official version of record. They are accessible to all readers and citable by the Digital Object Identifier (DOI®). “Just Accepted” is an optional service offered to authors. Therefore, the “Just Accepted” Web site may not include all articles that will be published in the journal. After a manuscript is technically edited and formatted, it will be removed from the “Just Accepted” Web site and published as an ASAP article. Note that technical editing may introduce minor changes to the manuscript text and/or graphics which could affect content, and all legal disclaimers and ethical guidelines that apply to the journal pertain. ACS cannot be held responsible for errors or consequences arising from the use of information contained in these “Just Accepted” manuscripts.

ACS Applied Materials & Interfaces is published by the American Chemical Society. 1155 Sixteenth Street N.W., Washington, DC 20036 Published by American Chemical Society. Copyright © American Chemical Society. However, no copyright claim is made to original U.S. Government works, or works produced by employees of any Commonwealth realm Crown government in the course of their duties.

Page 1 of 32

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

ACS Applied Materials & Interfaces

Hybrid Antifouling and Antimicrobial Coatings Prepared by Electroless Codeposition of Fluoropolymer and Cationic Silica Nanoparticles on Stainless Steel: Efficacy Against L. monocytogenes Kang Huang1, Juhong Chen1, Sam R. Nugen1,2, and Julie M. Goddard*1,2 1

Department of Food Science, University of Massachusetts, 102 Holdsworth Way, Amherst, MA, USA 01003

2

Current affiliation: Department of Food Science, Cornell University, Ithaca, NY USA 14853

KEYWORDS: fluorinated nanoparticle, silica nanoparticle, electroless nickel plating, antifouling, antimicrobial

ACS Paragon Plus Environment


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

1. Abstract Controlling formation, establishment, and proliferation of microbial biofilms on surfaces is critical for ensuring public safety.

Herein, we report on the synthesis of antimicrobial

nanoparticles and their co-deposition along with fluorinated nanoparticles during electroless nickel plating of stainless steel. Plating bath composition is optimized to ensure sufficiently low surface energy to resist fouling and microbial adhesion as well as exert significant (>99.99% reduction) antimicrobial activity against Listeria monocytogenes. The resulting coatings present hybrid antifouling and antimicrobial character, can be applied onto stainless steel, and do not rely on leaching or migration of the antimicrobial nanoparticles to be effective. Such coatings can support reducing public health issues related to microbial crosscontamination in areas such as food processing, hospitals, and water purification. 2. Introduction Establishment of microbial biofilms on surfaces (e.g. biomedical implants, food processing equipment, water purification systems) remains a significant challenge to public health. Strategies for biofilm removal rely on chemical disinfectants which present an environmental burden, are often ineffective, and may promote development of resistant organisms. Researchers have therefore explored surface modification methods to prevent surface protein conditioning and subsequent microbial attachment (antifouling), inactivate adhered microorganisms (antimicrobial), or facilitate biofilm removal (self-cleaning). While there have been significant technological advances in the development of surfaces with either antifouling or antimicrobial properties, it has been established that an ideal antimicrobial coating should exhibit hybrid antifouling and antimicrobial character.1 Several groups have reported on co-grafting of polymer brushes with antimicrobial


Page 2 of 32

Page 3 of 32

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


peptides or enzymes.2-4 While effective in conformally coating a range of material types, polymer brush type coatings may not be robust against thermal, chemical, or mechanical stresses typical in cleaning and disinfection protocols.5

Electroless nickel plating is an

autocatalytic process for direct coating of ferrous metals (e.g. steel), providing a hard coating with high wear and corrosion resistance, good weldability, and electrical conductivity.6-7 A unique attribute of electroless plating is that particles dispersed in the plating bath will become part of the coating, enabling introduction of functional character to the coated surface depending on the nature of the included particles. Co-deposition of polytetrafluoroethylene (PTFE) nanoparticles during electroless nickel-phosphorus plating has resulted in a nonfouling Ni-PTFE coating on stainless steel with efficacy in reducing accumulation of complex foulant as well as adhered microorganisms.8-13 Inclusion of silver nitrate in the plating bath has resulted in Ni-Ag-PTFE coatings with demonstrated antimicrobial and nonfouling character,14-15 yet in addition to cost and toxicity concerns, there are inherent limitations to the coating lifetime because the metal ions must migrate from the coatings to be effective. An opportunity therefore remains for synthesis of low cost, non-leaching antimicrobial nanoparticles suitable for co-deposition during Ni-PTFE plating. Polyethylenimine (PEI) (and its alkyl derivatives) is a cationic polymer which has been explored to impart antimicrobial character as a coating on planar solid supports and porous silica.16-18 In other work, cross-linked polyethylenimine nanoparticles were embedded in polymer coatings and composite resins with demonstrated antimicrobial efficacy.19-20 We hypothesized that PEIfunctionalized silica nanoparticles would have potent antimicrobial efficacy and could be codeposited during electroless nickel plating. In this work, we report on an electroless nickel plating technique in which antifouling



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 4 of 32

and antimicrobial nanoparticles are codeposited onto stainless steel in order to provide a surface which not only prevents protein conditioning and subsequent microbial attachment but is capable of inactivating any adhered microorganisms.

PEI-functionalized silica

nanoparticles were synthesized and co-deposited with commercial PTFE nanoparticles during electroless nickel plating of stainless steel coupons. The antimicrobial nanoparticles were characterized for size, morphology, charge, and chemistry; nanocomposite coatings were characterized for wettability, surface energy, morphology and chemistry. The antimicrobial and anti-biofilm forming character of the coated steel surfaces was demonstrated against Listeria monocytogenes. 3. Experimental 3.1. Materials Stainless steel sheets (316, 2B finish, medium hardness [Rockwell B93], with thickness of 0.61 ± 0.07 mm) were purchased from McMaster-Carr (Robbinsville, NJ, USA). Pure ethyl alcohol (>99.5%), tetraethyl orthosilicate (TEOS) (99.999%), divinyl sulfone (DVS), (3Aminopropyl)triethoxysilane (APTES) (99%), branched polyethylenimine (PEI) (average Mw ~25,000 Da), sodium hypochlorite (5%), ninhydrin reagent, ethanolamine (≥99.0%), and sodium dodecyl sulfate (SDS) were purchased from Sigma Aldrich Co. (St. Louis, MO, USA) and were used without further purification. Ammonium hydroxide (29.7%), 2-Propanol, nickel (II) chloride, and hydrochloric acid (HCl) were purchased from Fisher Scientific (Waltham, MA, USA). Components for a high-phosphorous electroless nickel plating bath and PTFE nanoparticles were purchased from MacDermid Inc. (Denver, CO, USA). Tryptic soy broth (TSB), tryptic soy agar (TSA), and neutralizing buffer were from Difco, Becton Dickinson (Sparks, MD, USA).


Page 5 of 32

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Table 1: Abbreviations used to describe nanoparticle synthesis and coating compositions. Abbreviation

Description

Ni

Nickel

NPs

nanoparticles

PTFE NPs

polytetrafluoroethylene nanoparticles

SiO2 NPs

silica nanoparticles

SiO2@APTES NPs

amine-functionalized silica nanoparticles

SiO2@DVS NPs

Vinyl sulfone-functionalized silica nanoparticles

SiO2@PEI NPs

Polyethylenimine decorated silica nanoparticles (in solution)

SiNP

+

Polyethylenimine decorated silica nanoparticles (in coating)

3.2. Synthesis of SiO2@PEI nanoparticles For clarity in interpreting nanoparticle synthesis illustrated in Figure 1, as well as results and discussions, a list of abbreviations is provided in Table 1. Silica nanoparticles were prepared according to the modified Stöber method.21-22 The synthesis process of the PEI-functionalized silica nanoparticles was accomplished using the previously published method in which PEI is linked to silica nanoparticles by divinyl sulfone.23 A mixture of 34.82 ml of deionized water, 3.25 ml of ammonium hydroxide (29.7 %), and 100 ml of ethanol was stirred at 600 rpm in a round-bottom flask with a stopper and stabilized in a water bath at 40 ºC, after which 6.20 ml of TEOS was added at an injection rate of 5 ml/min. The mixture was allowed to react for 2 h, after which the resulting silica nanoparticles (SiO2 NPs) were washed twice by ethanol and suspended in 100 ml of anhydrous ethanol.

To prepare amine functionalized silica

nanoparticles (SiO2@APTES NPs), 2 ml of APTES was introduced dropwise to the SiO2 suspension with the stirring rate of 600 rpm. The reaction was conducted at room temperature for 20 h. The resulting SiO2@APTES NPs were collected, washed twice by ethanol, and stored in ethanol at 4 ºC. Immediately before further use, the SiO2@APTES NPs were transferred into isopropanol. To prepare vinyl sulfone-functionalized silica nanoparticles (SiO2@DVS NPs), 28 µl of divinyl sulfone (DVS) was added to a sonicated dispersion of



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

SiO2@APTES NPs in isopropanol. After a reaction period of 2 h at room temperature, the resulting SiO2@DVS NPs were washed for one cycle and redispersed in isopropanol. Finally, 25 kDa branched PEI solution (280 mg in 2 ml isopropanol) was introduced into the asprepared and sonicated SiO2@DVS NPs. The mixture was sonicated for 5 min, and then allowed to stir overnight. After the reaction was finished, the excess PEI was washed by two cycles using ethanol. The resulting PEI decorated silica nanoparticles (SiO2@PEI NPs) were redispersed in ethanol and stored at 4 ºC for further use.

Fig. 1 Schematic diagram of bactericidal properties of Ni-PTFE-SiNP+ modified coatings (A) and synthesis process of SiO2@PEI NPs (B). 3.3.Characterization of SiO2@PEI nanoparticles The size and surface charge (ζ-potential) of the synthesized nanoparticles at each stage were measured using Dynamic Light Scattering (DLS, Nano-ZS, Malvern Instruments, Worcestershire, UK) after dilution in DI water. The refractive index of silica used in the light ACS Paragon Plus Environment

Page 6 of 32

Page 7 of 32

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


scattering calculation was 1.4585.24 Three measurements were performed for each nanoparticle suspension, and three independent syntheses for each kind of particles were analyzed. Surface chemistry of synthesized nanoparticles at each stage was further characterized by X-ray photoelectron spectroscopy (XPS). Briefly, 1 ml of suspension (with 0.15 g of nanoparticles) was deposited on a 1 × 1 cm2 piece of cleaned stainless steel coupon, and airdried at room temperature. The nanoparticles were analyzed using a Physical Electronics Quantum 2000 (Physical Electronics, Chanhassen, MN). A single 100 µm spot was analyzed from every coupon at an angle of 45 º relative to the coupons’ surface with Al Kα excitation. Survey scans were collected at a pass energy of 187.85 eV. The reported XPS results are representative spectra of three scans on each kind of nanoparticles prepared on three independent days. The morphology and structure of the nanoparticles were characterized by Transmission electron microscopy (TEM) using a JEOL JEM-2200FX transmission electron microscope (Akishima, Tokyo, Japan), equipped with an Oxford 80 mm2 X-max energy dispersive X-ray spectrometer (EDX, Oxford Instruments, Abingdon, UK) to analyze the chemical composition of nanoparticles. 3.4. Preparation of antifouling and antimicrobial Ni-PTFE-SiNP+ coating An autocatalytic, electroless nickel plating process was used to introduce an antifouling and antimicrobial coating onto stainless steel surfaces based on a previously reported method with slight modification.8 This electroless nickel plating procedure results in a coating thickness of approximately 10 µm, which is controllable based on plating time.8 Briefly, coupons of 316 stainless steel were subjected to alkali cleaning (85 ºC, 5 min), alkali electro-cleaning (85 ºC,



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 8 of 32

2 min), acid pickle (room temperature, 2 min), and Wood’s nickel strike (room temperature, 5 min). The resulting cleaned, Wood’s striked coupons were submerged in the electroless nickel plating bath (85 ºC, 60 min). After that, as-prepared coupons were submerged in the modified electroless nickel plating bath with antifouling (PTFE) and/or antimicrobial (SiNP+) nanoparticles for another 60 min at 85 ºC. The modified electroless plating bath was prepared by dispersing various concentrations of polyethylenimine decorated silica nanoparticles (dubbed SiO2@PEI NPs when in solution and SiNP+ when in coating) into the plating solution with 0.025 g/L of surfactant (SDS) using sonication for 5 min. Compositions of the plating baths for each variant are defined in Table 2, with native (clean stainless steel), NiPTFE (plated steel including only PTFE nanoparticles), and Ni-SiNP+ (plated steel including only SiO2@PEI NPs) serving as controls.

Table 2 Components of electroless nickel plating bath No. 1 2 3 4 5 6 7

Sample Native Ni-PTFE Ni-SiNP+ (0.20 g/L) Ni-PTFE-SiNP+ (0.05 g/L) Ni-PTFE-SiNP+ (0.10 g/L) Ni-PTFE-SiNP+ (0.15 g/L) Ni-PTFE-SiNP+ (0.20 g/L)

PTFE NPs (ml/l) 5 5 5 5 5

SiO2@PEI NPs (g/L) 0.20 0.05 0.10 0.15 0.20

SDS (g/L) 0.025 0.025 0.025 0.025 0.025 0.025

3.5.Characterization of native and Ni-PTFE-SiNP+ modified stainless steel Contact angle and surface energy of native and coated stainless steel coupons were quantified using the dynamic contact angle method measured on a Kruss DSA100 Drop Shape Analyzer (Hamburg, Germany) recording both the advancing (θA) and the receding contact angles (θR). Three measurements were performed at three separate areas for each test coupon, and three individual coupons for each surface were analyzed. Surface energies were quantified using a


Page 9 of 32

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Neumann model as described in previously published work.25-26 As a complementary source of coating characterization, coupons of native and modified stainless steel were analyzed using X-Ray photoelectron spectroscopy, on a Physical Electronics Quantum 2000 (Physical Electronics, Chanhassen, MN, USA). A single 100 µm spot was analyzed from every coupon at an angle of 45 º relative to the coupons’ surface with Al Kα excitation. Survey scans were collected at a pass energy of 187.85 eV, and highresolution spectra of Ni 2p, P 2p, C 1s, O 1s, F 1s, Si 2p, and N 1s were collected at a pass energy of 46.95 eV. Atomic percentages and high-resolution spectra were analyzed with Multipak software 6.1A (Physical Electronics). Data are reported as average ± standard deviation of four scanning points (from two distinct regions on each of two independent samples). Morphology, topography, and nanoparticle chemistry and distribution within coatings were characterized using scanning electron microscopy-Energy dispersive X-ray (SEM-EDX) analysis with an FEI SEM Magellan (Hillsboro, OR, USA) equipped with an Oxford 80 mm2 X-max energy dispersive X-ray spectrometer (Oxford Instruments, Abingdon, UK). SEM analysis was performed with a current of 13 pA and voltage of 10 kV. EDX spectra were collected at 30º angle, 20 kV accelerating voltage and a 20 mm working distance. Surface topographies of native and modified stainless steel coupons were further analyzed with a Dimension 3000 atomic force microscope (Digital Instruments, Santa Barbara, CA, USA) under tapping mode. Surface roughnesses were calculated using NanoScope Analysis 1.40 (Bruker Corporation, Billerica, MA, USA). The primary amine content of the stainless steel coupons was characterized using the ninhydrin assay. Briefly, coupons (1 × 1 cm2) were immersed in ninhydrin reagent (2 ml of



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

0.2% w/v in 0.1 M buffer phosphate, pH = 9.0), and the mixture was heated in a boiling water bath for 15 min. After cooling to room temperature, absorbance intensities were measured at 570 nm and primary amine content (in nmol/cm2) of coupons were calculated by comparison to a standard curve of ethanolamine. 3.6. Antimicrobial activity of native and modified stainless steel The antimicrobial activities of each treatment identified in Table 2 (native stainless steel, coating containing only PTFE NPs, coating containing only SiO2@PEI NPs, as well as coatings containing PTFE and different concentrations of SiO2@PEI NPs) were evaluated. Bacterial cultures of L. monocytogenes Scott A (provided by Dr. Martin Wiedmann, Food Science Department, Cornell University, Ithaca, NY, USA) were prepared by streaking a loopful of stock culture kept at -80 ºC in TSB with 25% glycerol onto TSA. After incubation at 37 ºC for 24 h, a single colony was inoculated in 9 ml of TSB and incubated overnight for 14 h at 37 ºC. A 1% dilution of the overnight broth was prepared with fresh and sterile TSB and incubated for 4 h at 37 ºC. The bacterial suspension with approximately 6 log CFU/ml was obtained by preparing a 0.2% dilution of broth with sterile DI water. A volume of 1 ml of bacterial suspension and a volume of 1 ml of DI water was added in a test tube with one 1 × 1 cm2 coupon, and incubated for 2 h with rotation (60 rpm) at 30 ºC. Native SS was used as negative control. After 2 h of incubation, the bacterial suspensions of each treatment were serially diluted using PBS. A volume of 100 µl of each dilution was inoculated onto TSA plates and incubated at 37 ºC until enumeration. 3.7. Biofilm formation assay The ability of the composite antimicrobial and antifouling coatings to resist biofilm formation was performed using a modified CDC biofilm reactor as reported by our previous work.13 The


Page 10 of 32

Page 11 of 32

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


biofilm formation assay was performed on native, Ni-PTFE-modified, Ni-SiNP+ (0.10 g/L) modified, and Ni-PTFE-SiNP+ (0.10 g/L) modified stainless steel coupons to characterize the influence of each nanoparticle type on biofilm formation. After assembly of the sterile native and modified stainless steel coupons in the reactor system, the whole reactor (including coupons and tubing) was autoclaved. A 1: 100 dilution was prepared from the overnight bacterial suspension with sterile M9 broth (prepared according to ATCC medium #2511 M9 Minimal Agar/Broth) and transferred into the sterile vessel. The CDC biofilm reactor was operated in batch phase for the first 24 h at 30 ºC to allow for biofilm establishment, after which fresh M9 broth was continuously pumped into the reactor at a flow rate of 1 ml/min for another 48 h at 30 ºC using a Master Flex peristaltic pump. The rotating baffle was stirring at a constant rate of 60 rpm during the whole process. After 48 h of biofilm production (72 h total exposure), the pump was stopped, the coupons were collected and submitted to the procedure described just below to enumerate the strongly adherent/biofilm cells. Bacterial enumeration of the biofilm was tested using a modified bead vortexing method reported by previous researchers.27 After two rinses in deionized water to remove loosely adsorbed cells, each coupon was transferred to a sterile test tube containing 10 ml of PBS and 3 g of sterile glass beads (diameter 425–600 µm). After vortexing at full speed for 1 min in the tube by a vortex mixer (Fisher Scientific, Waltham, MA, USA), quantification of bacteria was performed by serial dilution and duplicate spread plating on TSA plates. To ensure that the bacterial enumeration method used to quantify the biofilm formation was effective in completely removing any adhered cells, rigorous electron microscopy was performed after antimicrobial activity and biofilm formation assays using a JEOL JCM-6000 NeoScope Benchtop SEM (Peabody, MA). Treated coupons were treated with 3%



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 12 of 32

glutaraldehyde in PBS for at least 4 h to fix any microorganisms. Afterward, the samples were washed twice with sterile water, air-dried at 37 ºC, sputter-coated with gold for 30 s, and finally imaged at an acceleration voltage of 10 kV and a magnification of 2000×. The images presented in this report are representative of 9 electron micrographs acquired at locations uniformly distributed across a grillage-type pattern on each of triplicate samples. 3.8.Statistical analysis Statistical analysis was performed by GraphPad Prism software V.5.04 (Graphpad Software, Inc., La Jolla, CA). To analyze the difference between multiple groups on each data set, oneway analysis of variance (ANOVA, P < 0.05) was performed for surface characterization, evaluation of antimicrobial property, and biofilm formation assay.

4. Results and Discussion 4.1. Characterization of synthesized SiO2@PEI nanoparticles The antimicrobial nanoparticles (SiO2@PEI NPs) were characterized for size, surface charge, surface chemistry, and morphology at each stage of their syntheses (Figure 2, Supplemental Figure S1). Dynamic light scattering revealed that as prepared SiO2 NPs had a mean diameter of 125.03 ± 18.12 nm, indicating a uniform size distribution (Figure 2a).

Surface

functionalization by introducing an aminated silane (APTES) followed by vinyl sulfone (DVS) and polyethylenimine (PEI) increased mean diameters by dimensions expected given the size of the ligands (130.87 ± 21.50, 137.37 ± 37.11, and 180.33 ± 40.32 nm, respectively). Dynamic light scattering was further used to characterize changes in surface charge at each stage during the surface functionalization (Figure 2b). SiO2 NPs presented negative zeta potential values, as expected from the hydroxyl group on the surface. SiO2@APTES NPs were less negatively charged due to the introduction of the positively charged primary amines ACS Paragon Plus Environment

Page 13 of 32

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


in the APTES, and SiO2@DVS NPs became again more negatively charged after the introduction of the negatively charged vinyl sulfone group. Immobilization of the branched polyethylenimine effectively introduced sufficient primary amines to result in a net positive charge on the nanoparticles. The positive charge (>30 mV) ensured sufficient repelling force among nanoparticles, enabling a stable dispersion of nanoparticles during storage and plating. XPS survey scans were performed on dried particles to characterize surface chemistry (Figure 2c), revealing characteristic peaks of Si 2s and Si 2p centered at 153 eV and 102 eV in all particles, with introduction of C 1s peaks for surface modified silica nanoparticles and introduction of S 2s and S 2p peaks in the SiO2@DVS NPs. SiO2@PEI NPs presented an additional peak at N 1s centered at 400 eV, further indicating the successful introduction of PEI at or very near the particle surface.

TEM images revealed uniform particle size,

distribution, and morphology both as prepared SiO2 NPs and in the surface functionalized SiO2@PEI NPs (Figure 2d). TEM-EDX mapping of SiO2@PEI NPs supported results of XPS, indicating the presence of Si, O, S, and N uniformly across the particle surface.



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Fig. 2. Characterization of synthesized SiO2@PEI NPs by dynamic light scattering for size (A) and zeta potential (B), XPS survey scan (C), TEM images of nanoparticles (D), and TEMEDX image of SiO2@PEI NPs used for the elemental mapping of Si, O, S, and N (E). Error bars represent SD of n = 3 values. 4.2.Wettability and surface energy of native and Ni-PTFE-SiNP+ modified stainless steel The influence of nanoparticle inclusion and plating bath composition on wettability and fouling of the modified stainless steel surfaces identified in Table 2 were characterized using contact angle and surface energy determinations (Figure 3). As indicated in Table 1, for clarity in data presentation, SiO2@PEI NPs are abbreviated as SiNP+ after incorporation into


Page 14 of 32

Page 15 of 32

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


the coating. Coating of PTFE nanoparticles alone (no SiNP+) resulted in a significant increase in hydrophobicity with both advancing and receding contact angles significantly higher than those of native stainless steel. The Ni-PTFE composite coating further exhibited a significant reduction in surface energy (from >30 mN/m in native steel to 22 mN/m in Ni-PTFE coated steel), in agreement with prior reports on the antifouling character of Ni-PTFE coatings.25 Coating of SiNP+ alone (no PTFE nanoparticles) resulted in a noted decrease in hydrophobicity and corresponding increase in surface energy, compared to native stainless steel, as expected due to the introduction of highly hydrophilic primary amine decorated silica nanoparticles. Codeposition of PTFE and SiNP+ in the nickel matrix enabled retention of hydrophobic, low surface energy character in the coated stainless steel. As the concentration of SiO2@PEI NPs increased, the hydrophobicity of the modified steel decreased, as expected due to the hydrophilic property of the PEI. The surface energy of Ni-PTFE-SiNP+ modified surfaces with increasing concentrations of SiO2@PEI NPs ranged from 20.72 ± 1.06 to 28.74 ± 0.66 mN/m, which all fall within the range defined by Baier (20-30 mN/m) as presenting antifouling character.28 In contrast, the surface energy of coating with SiNP+ alone (no PTFE nanoparticles) presented a surface energy greater than that of native stainless steel, 44.86 ± 0.53 mN/m. The retention of low surface energy (and corresponding antifouling character) in all Ni-PTFE-SiNP+ variants despite introduction of cationic antimicrobial nanoparticles is a unique property of the reported antifouling, antimicrobial coatings, and is attributed to the codeposition of PTFE with SiO2@PEI NPs. These results are significant as introduction of cationic antimicrobials to surfaces can result in fouling by biological fluid components (thus inhibiting their antimicrobial efficacy) as well as promoting adhesion of microorganisms due to electrostatic interactions.5,29-31



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Fig. 3 Contact angle (A) and surface energy (B) of native SS, Ni-PTFE SS, Ni-SiNP+ SS, and Ni-PTFE-SiNP+ SS with different concentrations of PEI-functionalized SiO2 NPs. Dash lines represent the advancing and receding contact angles and critical surface energy of native stainless steel coupons. Values represent means ± standard deviations of n=9 determinations (three determinations of each of three independently prepared samples). 4.3.Morphology and surface chemistry of native and Ni-PTFE-SiNP+ modified stainless steel Electron microscopy was performed to characterize the uniformity of dispersion of PTFE and SiO2@PEI NPs within the plated coating on stainless steel. Native stainless steel presented ACS Paragon Plus Environment

Page 16 of 32

Page 17 of 32

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


regular cracks and crevices typical of 2B finished 316 SS (Figure 4A). Inclusion of PTFE NPs alone (no SiO2@PEI NPs) resulted in a uniform deposition, indicated by dark spots of ~200 nm diameter, in agreement with prior reports.25 Introduction of SiO2@PEI NPs to the Ni-PTFE coating resulted in a uniform embedding of both particle types within the coating, evidenced by presence of both darker (from PTFE NPs) and lighter (from SiO2@PEI NPs) gray spheres, uniform within the nickel plated coating. SEM-EDX analysis was performed on Ni-PTFE coatings containing 0.05 g/L of SiO2@PEI NPs (Figure 4I), and presented characteristic peaks of F and Si, providing evidence of both NPs within the coating. Higher concentrations of SiO2@PEI NPs (0.20 g/L) in the plating bath resulted in coatings with poor integrity: without PTFE NPs (Figure 4C) the SiO2@PEI NPs did not embed well, and the electroless nickel matrix was not uniform. Similarly, Ni-PTFE-SiNP+ coatings prepared with 0.20 g/L SiO2@PEI NPs tended to delaminate. It is known that the microenvironment around nanoparticles can differ from that of the bulk environment, resulting in local differences in pH value and ionic strength, which can influence chemical processes such as electroless plating. It is therefore possible that concentrations exceeding 0.15 g/L, the cationic nanoparticles had a greater influence on both local and bulk pH values of the plating solution, thus inhibiting effective and uniform electroless nickel plating.



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Fig. 4 Representative SEM images of native (A), Ni-PTFE modified (B), Ni-SiNP+ modified (C), and Ni-PTFE-SiNP+ modified coatings with different concentrations of SiO2@PEI NPs (D-G). EDX spectrum of Ni-PTFE-SiNP+ containing 0.05 g/L SiO2@PEI NPs (H).

X-ray photoelectron spectroscopy was performed to characterize the influence of plating bath composition on surface chemistry (Table 3, Supplemental Figure S2). Ni-PTFESiNP+ samples presented lower fluorine content than samples prepared with Ni-PTFE alone. Further, fluorine content decreased (and nitrogen content increased) with increasing SiO2@PEI NP content with the exception of 0.20 g/L SiO2@PEI NPs, again supporting the likelihood that high concentrations of SiO2@PEI NPs yield irregular, unstable coatings. Decreasing fluorine and increasing nitrogen content are indicative of the introduction of SiO2@PEI NPs in the nickel matrix.


Page 18 of 32

Page 19 of 32

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Table 3. Atomic composition of the top ∼5 nm of native and modified stainless steel as determined by X-Ray photoelectron spectroscopy. Different superscript letters within the same column indicate statistical significance at (P < 0.05) with error bars representing SD of n = 4 values (2 regions of each of 2 independent samples). Sample

% Atomic composition Ni

P

C

O

F

Si

N

SS

--

--

46.1 ± 1.9 a

49.6 ± 1.7 a

--

4.3 ± 1.1 a

--

Ni-PTFE

9.7 ± 1.0 a

1.8 ± 1.0 a

32.2 ± 2.4 bd

15.2 ± 0.9 b

40.0 ± 3.2 a

1.1 ± 0.3 b

--

17.2 ± 2.3 b

8.2 ± 1.1 b

33.8 ± 0.7 b

34.1 ± 1.8 c

--

1.7 ± 0.9 b

5.0 ± 0.6 a

20.0 ± 1.8 c

2.3 ± 0.9 a

18.4 ± 3.8 c

19.6 ± 1.5 d

37.4 ± 0.6 a

1.8 ± 1.4 b

0.6 ± 0.5 b

20.2 ± 2.4 c

2.9 ± 1.1 a

17.6 ± 1.6 c

26.9 ± 2.4 e

29.2 ± 2.4 b

1.5 ± 0.5 b

1.9 ± 0.2 c

11.9 ± 2.7 d

1.2 ± 0.6 a

30.8 ± 2.9 bd

34.3 ± 1.7 c

18.8 ± 1.5 c

0.6 ± 0.8 b

2.5 ± 1.0 c

12.8 ± 1.5 d

2.1 ± 1.2 a

29.0 ± 2.4 d

25.2 ± 6.0 e

28.3 ± 6.6 b

1.0 ± 0.9 b

1.6 ± 1.0 c

Ni-SiNP+ (0.20 g/L) Ni-PTFE-SiNP+ (0.05 g/L) Ni-PTFE-SiNP+ (0.10 g/L) Ni-PTFE-SiNP+ (0.15 g/L) Ni-PTFE-SiNP+ (0.20 g/L)

Atomic force microscopy was performed to characterize nanoscale surface topography, uniformity of nanoparticle deposition within composite, and surface roughness (Figure 5). Roughness values were similar among all coating treatments, and remained lower than that of native stainless steel. AFM micrographs supported electron microscopy results suggesting that as SiO2@PEI NP content in the plating bath increased to 0.20 g/L, the embedding behavior was degraded, with evidence of nanoparticle leaching from the coating. These results suggested that there is a critical nanoparticle concentration in the plating bath, beyond which particles do not embed well. Height profile analysis of SiO2@PEI NPs after embedding in the coating (Fig. 5H) confirmed that nanoparticles were embedded within the



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

coating rather than deposited on the surface, with heights of 20-80 nm protrusion above the plane of the composite and diameters of 200-250 nm.

Fig. 5 Representative AFM images of SS (A), Ni-PTFE (B), Ni-SiNP+ (C), Ni-PTFE-SiNP+ 0.05 g/l (D), Ni-PTFE-SiNP+ 0.19 g/l (E), Ni-PTFE-SiNP+ 0.15 g/l (F), and Ni-PTFE-SiNP+ 0.20 g/l (G) modified stainless steel surfaces. Surface roughness of native and modified stainless steel (H), in which (a) – (e) represent the height of nanoparticles baring on the different surfaces. Values of surface roughness reported are averages of 3 area scans on each ACS Paragon Plus Environment

Page 20 of 32

Page 21 of 32

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


of 2 independent coupons (n = 6) with standard deviation.

Primary amine content of SiO2@PEI NPs and nanocomposite coatings was characterized using ninhydrin reagent. PEI decorated silica nanoparticles presented 134.21 ± 3.7 nmol amines per mg nanoparticles, and imparted significant primary amine content to the surface of the coated stainless steel. Raw absorbance spectra for representative samples of ninhydrin reagent after exposure to stainless steel (SS), Ni-PTFE, Ni-SiNP+ (containing 0.20 g/L SiO2@PEI NPs in the plating bath), and all Ni-PTFE-SiNP+ variants (containing 0.050.20 g/L SiO2@PEI NPs in the plating bath) are reported in Figure 6A. Native stainless steel presented no detectable amines, as expected. Ni-PTFE coated stainless steel presented a more strongly pronounced sloped baseline spectrum, likely a result of interaction of nickel with the ninhydrin reagent. Indeed, it has been reported that divalent metal cations can enhance the signal intensity of ninhydrin reagent.32-33 To account for this phenomenon, Ni-SiNP+ and NiPTFE-SiNP+ primary amine contents were determined by comparison to the Ni-PTFE control in Figure 6B. Ni-SiNP+ coatings (prepared with 0.20 g/L SiO2@PEI NPs and no PTFE NPs) presented the highest amine content, at 128.80 ± 1.7 nmol/cm2. As expected the primary amine content of Ni-PTFE-SiNP+ coatings increased as SiO2@PEI NPs content increased from 0.05 to 0.15 g/L in the plating bath. However, the primary amine content of Ni-PTFESiNP+ (0.20 g/L) SS was lower than that of Ni-PTFE-SiNP+ (0.15 g/L) SS, again supporting the conclusion that at too high an SiO2@PEI NP content, nanoparticles do not effectively embed in the coating.



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Fig. 6 Determination of primary amine content of native and coated stainless steel. (A) Representative absorbance spectra. (B) Primary amine content quantified at 570 nm by comparison to a Ni-PTFE control. Values with the same letter (a-d) are not significantly different (P > 0.05) with error bars representing the SD of n = 6 values.

Fig. 7.

Antimicrobial efficacy of native and modified stainless steel against Listeria

monocytogenes.

Values represent means ± standard deviations of n=6 determinations

(duplicate analysis on each of three independent experiments), with different letters indicating significant differences at P

Hybrid Antifouling and Antimicrobial Coatings ... - ACS Publications

Recommend Documents