Bactericidal Efficiency of Silver Nanoparticles Deposited onto Radio

Jul 12, 2010 - The potential application of low-temperature radio frequency (RF) plasma for ... discharge at atmospheric pressure improves the loading...
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Ind. Eng. Chem. Res. 2010, 49, 7287–7293

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Bactericidal Efficiency of Silver Nanoparticles Deposited onto Radio Frequency Plasma Pretreated Polyester Fabrics Vesna Ilic´,† Zoran Sˇaponjic´,‡ Vesna Vodnik,‡ Sasˇa Lazovic´,§ Suzana Dimitrijevic´,| Petar Jovancˇic´,† Jovan M. Nedeljkovic´,‡ and Maja Radetic´*,† Textile Engineering Department, Faculty of Technology and Metallurgy, UniVersity of Belgrade, KarnegijeVa 4, 11120 Belgrade, Serbia, Vincˇa Institute of Nuclear Sciences, P. O. Box 522, 11001 Belgrade, Serbia, Institute of Physics, PregreVica 118, 11080 Zemun, Serbia, and Department of Bioengineering and Biotechnology, Faculty of Technology and Metallurgy, UniVersity of Belgrade, KarnegijeVa 4, 11120 Belgrade, Serbia

The potential application of low-temperature radio frequency (RF) plasma for fiber surface activation in order to enhance the binding efficiency of colloidal silver nanoparticles onto the polyester fabrics and improve the stability of antibacterial effects was studied. Antibacterial activity and laundering durability were tested against gram-negative bacterium Escherichia coli and gram-positive bacterium Staphylococcus aureus. Plasma treatment positively affected the loading of silver nanoparticles as well as antibacterial activity and laundering durability of these textile nanocomposite materials. In spite of good laundering durability after five washing cycles, it was found that silver leached from the fabric into the bath during washing. Released silver from the washing effluent was efficiently removed by recycled wool-based nonwoven sorbent modified with hydrogen peroxide and biopolymer alginate. 1. Introduction Textile materials have been recognized as media that can easily support the growth of different microbes.1 Hence, the growing production of advanced medical, protective, and hygiene textiles requires efficient antimicrobial finishing. A wide range of antimicrobial agents have been employed so far in the antimicrobial finishing of textile materials: metals and metal compounds, quaternary ammonium salts, poly(hexamethylene biguanide), triclosan, chitosan, dyes, regenerable N-halamine compounds, and peroxyacids.1 Relatively poor efficiency and/ or high toxicity made most of them unsuitable for long-term use. However, silver in different forms exhibits outstanding antimicrobial activity with low toxic impact to mammalian cells.2 It is a powerful biocide for more than 650 various microbes.3 In particular silver nitrate has been widely used as an antimicrobial agent.4,5 Despite its excellent antimicrobial properties, silver nitrate is not convenient for the treatment of textile materials as it stains to black-brown when exposed to air and light, due to uncontrolled reduction processes.6 On the other hand, the deposition of engineered silver nanoparticles (Ag NPs) onto textile materials can provide an adequate level of antimicrobial efficiency without considerable color change.3 Recently developed simple procedures for synthesis of Ag NPs and their high antimicrobial efficiency make them a viable substitute to conventional antimicrobial agents. Consequently, the treatment of different textile materials with Ag NPs is receiving remarkable, not only scientific but also industrial, attention.2,7–11 Numerous methods have been developed for the loading of textile substrates with Ag NPs. In addition to the most commonly applied dip-coating methods,7,8,10,11 sonochemical coating using ultrasound irradiation12,13 as well as the sputter deposition14,15 of Ag NPs onto textile surfaces, were performed. * To whom correspondence should be addressed. Fax: +381 11 3370387. Tel.: +381 11 3303 857. E-mail: [email protected]. † Textile Engineering Department, University of Belgrade. ‡ Vincˇa Institute of Nuclear Sciences. § Institute of Physics. | Department of Bioengineering and Biotechnology, University of Belgrade.

Vigneshwaran et al. proposed in situ synthesis of Ag NPs on cotton fabrics where the aldehyde terminal of starch made possible the reduction of the silver nitrate to silver metal, simultaneously stabilizing the NPs on the fabric.9 Dura´n et al. reported that good antibacterial efficiency can be achieved using the Ag NPs produced by a fungal process on cotton fabrics.16 Ag NPs can be also efficiently incorporated into fibers by electrospinning.17 However, the latest trends are more oriented toward obtaining stable and durable nanocomposite textile materials with Ag NPs.2,18 Plasma activation of textile materials, in particular hydrophobic polyester (PES) fabrics, appears to be beneficial for Ag NPs loading from colloids.2,18 Conventional chemical treatments that can increase the surface energy of PES fibers, and, hence, improve their wettability and adhesion properties, are recognized as ecologically unacceptable as they require huge amounts of water and chemicals.19 Unlike them, plasma processing is dry, clean, simple, multifunctional, environmentally friendly, and an economically feasible treatment. Additionally, it is not time-consuming, and it provides superficial modification of a fiber surface, leaving the bulk properties unaltered. The desired surface chemistry, i.e., plasma functionalization, can be achieved by adequate control of plasma parameters (treatment time, power, gas type, pressure, and gas flow). It has been shown that pretreatment of PES fabrics by corona discharge at atmospheric pressure improves the loading of Ag NPs, providing the enhanced antimicrobial activity and laundering durability.20–22 The main advantage of corona systems is that they operate at atmospheric pressure. Although corona systems principally meet the demands of the textile industry from the standpoint of speed and width, generated type of plasma cannot provide the desired spectrum of surface functionalizations on textile materials.23 Plasma particles cannot penetrate deeply into yarns, and, hence, achieved effects are short-lived. Additionally, the thickness of the textile materials is limited due to small interelectrode spacing.23 However, lowpressure devices, in particular radio frequency (RF) powered plasma sources, allow easier control of properties and provide

10.1021/ie1001313  2010 American Chemical Society Published on Web 07/12/2010

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a greater stability and uniformity at the cost of a more complex handling of fabric through the vacuum system.24–26 Therefore, the first part of this study discusses the potential application of low-temperature air RF plasma for fiber surface activation that can facilitate the deposition of colloidal Ag NPs onto the PES fabrics and, thus, enhance their antibacterial properties. Antibacterial activity and laundering durability were tested against gram-negative bacterium Escherichia coli (E. coli) and grampositive bacterium Staphylococcus aureus (S. aureus). In spite of growing commercialization of Ag NPs in general, little is known about the environmental impact of the products containing these species.27 It is well-established that ionic silver is very toxic to aquatic organisms, and its concentration in water is strictly regulated by water quality criteria.28,29 In contrast, data corresponding to toxicity and exposure of Ag NPs are still lacking.28 To our knowledge, there are only a few studies on silver release during washing of textile materials and possible treatments of these effluents.28–30 Benn and Westerhoff analyzed the form and amount of silver released from different sorts of commercial socks into water and its fate in wastewater treatment plants.28 This study clearly indicates that silver is released either in the form of NPs or as ions. Additionally, the amount and rate of silver release strongly depends on the sock type, suggesting that the manufacturing process may control silver release.28 Dura´n et al. studied the bioremediation process of Ag NPs released from cotton fabrics with the bacterium Chromobacterium Violaceum (C. Violaceum).29 This treatment based on biosorption seems to be efficient for removal of released Ag NPs in water. However, both studies reported the leaching of silver in ultrapure or tap water; i.e., the effect of washing agents on silver release from textile materials into the washing effluents was not addressed in their work. Geranio et al. followed the effect of pH, surfactants, and oxidizing agents on the amount and the form of released silver during washing from nine different fabrics with silver bound to the fiber surface or incorporated into the fiber.30 Again, they came to the same conclusion that the release of silver either in ionic or particulate form varies remarkably among the products (from less than 1 to 45%). However, particulate silver seems to be the predominant form of silver released under conditions relevant to washing. Hence, the second part of this study considers silver release from the PES fabrics during washing in the presence of washing agent and the possibility of silver removal by recycled woolbased nonwoven sorbent from washing effluent. Extensive research on potentials of this sorbent for removal of metal ions (Pb2+, Cu2+, and Zn2+), different dyes, and oils from water indicated its multifunctionality and high sorption efficiency.31–33 It is well-known that Ag+ is bound to the wool primarily via carboxylic groups.34 Thus, in order to introduce new carboxylic groups to the wool fiber surface, the recycled woolbased nonwoven sorbent was modified with hydrogen peroxide and biopolymer alginate. 2. Materials and Methods 2.1. Materials. 2.1.1. Treatment of PES Fabrics. Desized and bleached polyester (PES, 115 g/m2) fabrics were cleaned in a bath containing 0.50% nonionic washing agent Felosan RG-N (Bezema) at a liquor-to-fabric ratio of 50:1.20 After 15 min of washing at 50 °C, the fabrics were rinsed once with warm water (50 °C) for 3 min and three times (3 min) with cold water. The samples were dried at room temperature. Low-temperature plasma treatment of fabrics was carried out in capacitively coupled, radio frequency (13.56 MHz) air

induced plasma. This capacitively coupled plasma (CCP) reactor was previously used for treatment of polymers and different textile materials.24,26,35 The apparatus consisted of a constant RF power supply (Dressler Caesar 1010), matching box (Variomatch matching network), vacuum pump, chamber, gas supply with appropriate pressure gauges, current and voltage probes, digital oscilloscope, and a computer. To keep the reflected power at the minimum, the impedance was adjusted by tuning the matching box. With reduction of the reflected power, the power transmitted to the system increased and a stable operation was achieved. The chamber was cylindrical (37 cm in diameter, 50 cm in length) with a central electrode (14 mm in diameter) that was powered through the matching box. Plasma formed between the central electrode and the wall of the chamber that was grounded. The samples were placed on the platform at the bottom of the chamber. Such an asymmetric system was intentionally constructed in order to provide operating conditions under which the sheath potential is not too high36 but sufficient for optimum modification of different textile materials, avoiding their permanent damage. Preliminary studies indicated that plasma parameters which showed optimum effects in previous work seemed to be also adequate for PES fabrics. Therefore, the power applied to the CCP reactor was 100 W; treatment time was 2.5 min, while the pressure was maintained at a constant level of 0.27 mbar. AgNO3 (Kemika) and NaBH4 (Fluka) of p.a. grade were used without any further purification for the synthesis of colloidal Ag NPs. Briefly, 8.5 mg of AgNO3 was dissolved in 250 mL of water purged by argon for 30 min.37,38 Under vigorous stirring, reducing agent NaBH4 (125 mg) was added to the solution and left for 1 h in argon atmosphere. The concentration of Ag colloid was 50 ppm. A 1 g amount of PES fabric was immersed in 65 mL of colloid of Ag NPs for 5 min and dried at room temperature. After 5 min of curing at 100 °C, the samples were rinsed twice (5 min) with deionized water and dried at room temperature. To investigate the influence of the colloid concentration, the whole procedure was repeated on certain fabrics. 2.1.2. Treatment of Sorbent. The possibility of silver removal from the washing effluent by sorption was tested on the recycled wool-based nonwoven material (78/22 wool/ polyester). This sorbent was produced from second-hand military knitted pullovers. A procedure for the production of recycled wool-based nonwoven material is described elsewhere in detail.32 The recycled wool-based nonwoven material was treated with hydrogen peroxide and biopolymer alginate in order to improve its sorption properties. Hydrogen peroxide treatment (H2O2, 20 mL/L; Na4P2O7, 1.5 g/L; NH3(aq), 2.5 mL/L) was carried out in static conditions (without shaking). Samples were treated in the solution for 1 h (liquor ratio, 30:1) at 70 °C and pH 9.4, washed with water, and dried at room temperature. Low-viscosity sodium alginate (CHT-alginat NVS, Bezema) was used for the preparation of 0.5% alginate solution. Sodium alginate was dissolved in deionized water and stirred for 30 min. A 1 g amount of sorbent was dipped into 50 mL of freshly prepared 0.5% alginate solution for 10 min. After 10 min of curing at 100 °C, the fabrics were rinsed twice (5 min) with deionized water and dried at room temperature. 2.2. Methods. 2.2.1. Scanning Electron Microscopy. Fiber morphology was investigated by scanning electron microscopy (SEM, JEOL JSM 6460 LV). A golden layer was deposited on the samples before the analysis.

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2.2.2. Atomic Absorption Spectroscopy. The content of silver in the washing bath after each washing cycle and in the fabrics after the fifth washing cycle was determined by a PerkinElmer 403 atomic absorption spectrometer. 2.2.3. Antibacterial Efficiency. The antibacterial efficiency of PES fabrics was quantitatively assessed using a gram-negative bacterium E. coli ATCC 25922 and gram-positive bacterium S. aureus ATCC 25923. Bacterial inoculum was prepared in the Tryptone soya broth (Torlak, Serbia), which was used as a growing medium for bacteria, and potassium hydrogen phosphate buffer solution (pH 7.2) as a testing medium. Bacteria were cultivated in 3 mL of Tryptone soya broth at 37 °C and left overnight (late exponential stage of growth). Sterile potassium hydrogen phosphate buffer solution (70 mL) was added to a sterile Erlenmeyer flask (300 mL), which was then inoculated with 0.7 mL of bacterial inoculum. A 1 g amount of sterile fabric cut into small pieces (1 × 1 cm2) was placed in the flask and shaked for 1 h. Time zero counts were made by removing 1 mL aliquots from the inoculum which were diluted with physiological saline solution (8.5 g of NaCl in 1 L of water). A 0.1 mL aliquot of the solution was placed onto a Tryptone soya agar, and after 24 h of incubation at 37 °C, the zero time counts (initial number of bacterial colonies) of viable bacteria were made. Counts of 1 h each were made in accordance with the previously described procedure. The percentage of bacterial reduction (R, %) was calculated using the following equation: C0 - C × 100% R) C0

(1)

where C0 (CFU, colony forming units) is the number of bacterial colonies on the control fabric (untreated fabric without Ag) and C (CFU) is the number of bacterial colonies on the fabric loaded with Ag NPs.7,8,39 2.2.4. Laundering Durability. Laundering durability of antibacterial effects was evaluated after five washing cycles in Polycolor (Werner Mathis AG) laboratory beaker dyer at 45 rpm. The fabrics were washed in the bath containing 0.5% Felosan RG-N (Bezema) at a liquor-to-fabric ratio of 40:1. After 30 min of washing at 40 °C, the fabrics were rinsed once with warm water (40 °C) for 3 min and three times (3 min) with cold water. Afterward, the fabrics were dried at 70 °C.8 The percentage of bacterial reduction after five washing cycles was determined in accordance with eq 1. 2.2.5. Sorption of Silver. The effluents collected after the first two cycles of washing of all studied samples were mixed, and silver concentration was measured by atomic absorption spectroscopy (AAS). The measured pH value of the effluent was pH 4.5. Subsequently, 0.50 g of recycled wool-based nonwoven material was shaken in 25 mL of effluent for 3 and 24 h. Ag concentration after the sorption was also followed by AAS. 3. Results and Discussion 3.1. Characterization of PES Fabrics Modified by Plasma Treatment and Ag NPs. To enhance the interaction between hydrophilic colloidal Ag NPs and hydrophobic PES fibers, the surface of the substrate was modified by air RF plasma. Plasma induced morphological changes of PES fibers were analyzed by SEM. SEM images of untreated (UPES) and plasma treated PES (PPES) fibers are shown in Figure 1. Figure 1a reveals the smooth surface of the UPES fiber. The topography

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Figure 1. SEM images of UPES (a) and PPES fibers (b).

of the fiber was considerably altered after plasma treatment due to plasma etching (Figure 1b). Namely, energetic and highly reactive plasma species attacked the fiber surface and triggered the fiber ablation. Consequently, uneven cracks, pits, and striations running parallel to the fiber axis appeared, inducing the increase in fiber surface roughness.20,40,41 In addition to morphological changes, the chemical composition of the outer layers of the PES fibers was significantly altered. The XPS measurements in our previous study showed that air plasma treatment of PES fabrics resulted in an increase of the O/C ratio.42 The formation of new oxygen-containing groups on the fiber surface is suggested to be due to the presence of extremely reactive atomic oxygen species in discharge during the air plasma processing and/or post-plasma chemical reactions when the activated fiber surface reacts with environmental species.43–45 The rise of oxygen content ensures the improvement of PES fiber surface hydrophilicity and better accessibility of hydrophilic species. UPES and PPES fabrics were loaded once or twice with colloidal Ag NPs. For this purpose, uniform nearly spherical Ag NPs with an average diameter of approximately 10 nm, synthesized without using any stabilizer, were applied.20 The changes in fiber surface morphology after deposition of Ag NPs was also followed by SEM. SEM images of the PES fabrics loaded once and twice with Ag NPs (UPES + Ag and UPES + Ag × 2) as well as of the PPES fabrics loaded once and twice with Ag NPs (PPES + Ag and PPES + Ag × 2) are shown in Figure 2. Only a few almost spherical aggregates of Ag NPs with dimensions around 100 nm are observed on the surface of the UPES + Ag fiber (Figure 2a). On the contrary, a higher amount of smaller aggregates of Ag NPs with dimensions ranging from 40 to 70 nm was deposited on the surface of the UPES + Ag × 2 fibers (Figure 2b). As expected, plasma treatment positively affected the deposition of Ag NPs onto PES fibers (Figure 2c,d). Aggregates of Ag NPs (from 40 to 70 nm) were more uniformly distributed particularly over the surface of the PPES + Ag × 2 fibers (Figure 2d). Namely,

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Figure 2. SEM images of UPES fabrics loaded with Ag NPs once (a) and twice (b) and PPES fabrics loaded with Ag NPs once (c) and twice (d). Table 1. Antibacterial Efficiency of Ag Loaded UPES and PPES Fabrics

sample control UPES + Ag UPES + Ag × 2 PPES + Ag PPES + Ag × 2 control UPES + Ag UPES + Ag × 2 PPES + Ag PPES + Ag × 2

bacteria E. coli

S. aureus

initial no. of bacterial colonies (CFU)

no. of bacterial colonies on the fabric (CFU)

5.6 × 10

3.4 × 10