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J Nanopart Res (2010) 12:15311551 DOI 10.

1007/s11051-010-9900-y

REVIEW PAPER

A review of the antibacterial effects of silver nanomaterials and potential implications for human health and the environment
Catalina Marambio-Jones Eric M. V. Hoek

Received: 29 July 2009 / Accepted: 6 March 2010 / Published online: 23 March 2010 Springer Science+Business Media B.V. 2010

Abstract Here, we present a review of the antibacterial effects of silver nanomaterials, including proposed antibacterial mechanisms and possible toxicity to higher organisms. For purpose of this review, silver nanomaterials include silver nanoparticles, stabilized silver salts, silverdendrimer, polymer and metal oxide composites, and silver-impregnated zeolite and activated carbon materials. While there is some evidence that silver nanoparticles can directly damage bacteria cell membranes, silver nanomaterials appear to exert bacteriocidal activity predominantly through release of silver ions followed (individually or in combination) by increased membrane permeability, loss of the proton motive force, inducing de-energization of the cells and efux of phosphate, leakage of cellular content, and disruption DNA replication. Eukaryotic cells could be similarly impacted by most of these mechanisms and, indeed, a small but growing body of literature supports this concern. Most antimicrobial studies are performed in simple aquatic media or cell culture media without proper characterization of silver nanomaterial stability (aggregation, dissolution, and re-precipitation). Silver nanoparticle stability is governed by particle size, shape, and capping agents as

well as solution pH, ionic strength, specic ions and ligands, and organic macromoleculesall of which inuence silver nanoparticle stability and bioavailability. Although none of the studies reviewed denitively proved any immediate impacts to human health or the environment by a silver nanomaterial containing product, the entirety of the science reviewed suggests some caution and further research are warranted given the already widespread and rapidly growing use of silver nanomaterials. Keywords Silver Nanoparticle Antimicrobial Antibacterial Nanotechnology Nanotoxicology Safety EHS

Introduction The broad-spectrum antimicrobial properties of silver encourage its use in biomedical applications, water and air purication, food production, cosmetics, clothing, and numerous household products. With the rapid development of nanotechnology, applications have been extended further and now silver is the engineered nanomaterial most commonly used in consumer products (Rejeski 2009). Clothing, respirators, household water lters, contraconceptives, antibacterial sprays, cosmetics, detergent, dietary supplements, cutting boards, sox, shoes, cell phones, laptop keyboards, and childrens toys are among the

C. Marambio-Jones E. M. V. Hoek (&) Department of Civil and Environmental Engineering, California NanoSystems Institute, University of California, Los Angeles, 5732G Boelter Hall, PO Box 951593, Los Angeles, CA 90095-1593, USA e-mail: emvhoek@ucla.edu

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retail products that purportedly exploit the antimicrobial properties of silver nanomaterials. Different forms of silver nanomaterials already in such products include: metallic silver nanoparticles (Arora et al. 2008; Chi et al. 2009; Choi et al. 2008; Hwang et al. 2008; Kim et al. 2007, 2008a, b; Kvitek et al. 2008; Lok et al. 2006; Raf et al. 2008; Schrand et al. 2008; Sondi and Salopek-Sondi 2004; Vertelov et al. 2008), silver chloride particles (Choi et al. 2008), silver-impregnated zeolite powders and activated carbon materials (Cowan et al. 2003; Inoue et al. 2002; Yoon et al. 2008a, b), dendrimersilver complexes and composites (Balogh et al. 2001; Lesniak et al. 2005; Zhang et al. 2008), polymersilver nanoparticle composites (Bajpai et al. 2007; Damm and Munstedt 2008; Damm et al. 2008; Hlidek et al. 2008; Jin et al. 2007; Kim et al. 2009a, b; Kvitek et al. 2008; Naidu et al. 2008; Nita 2008; Sambhy and Sen 2008; Sanpui et al. 2008; Xu et al. 2006), silver-titanium dioxide composite nanopowders (Yeo and Kang 2008), and silver nanoparticles coated onto polymers like polyurethane (Jain and Pradeep 2005). While all of these forms of silver exert antimicrobial activity to some extent through release of silver ions, silver nanoparticles might exhibit additional antimicrobial capabilities not exerted by bulk or ionic silver (Chen and Schluesener 2008). Already, silver nanoparticles have been shown to be effective biocides against: (a) bacteria such as Escherichia coli, Staphylococcus aureus, Staphylococcus epidermis, Leuconostoc mesenteroides, Bacillus subtilis, Klebsiella mobilis, and Klebsiella pneumonia among others (Benn and Westerhoff 2008; Chen and Chiang 2008; Falletta et al. 2008; Hernandez-Sierra et al. 2008; Ingle et al. 2008; Jung et al. 2009; Kim 2007; Kim et al. 2007, 2009a, b; Kvitek et al. 2008; Raf et al. 2008; Ruparelia et al. 2008; Smetana et al. 2008; Sondi and Salopek-Sondi 2004; Vertelov et al. 2008; Yang et al. 2009; Yoon et al. 2008a, b); (b) fungi such as Aspergillus niger, Candida albicans, Saccharomyces cerevisia, Trichophyton mentagrophytes, and Penicillium citrinum (Kim et al. 2007, 2008a, b, 2009a, b; Roe et al. 2008; Vertelov et al. 2008; Zhang et al. 2008); and (c) virii such as Hepatitis B, HIV-1, syncytial virus (Elechiguerra et al. 2005; Lu et al. 2008; Sun et al. 2008; Zodrow et al. 2009). Hybrid silver nanocomposites with dendrimers and polymers have been

shown effective for S. aureus, Pseudomonas aeruginosa, E. coli, B. subtilis, and K. mobilis (Balogh et al. 2001; Zhang et al. 2008). Furthermore, silver loaded in nanoporous materials such as silverexchanged zeolites exhibit antibacterial effects for Pseudomonas putida, E. coli, B. subtilis, S. aureus, and P. aeruginosa (Cowan et al. 2003; Inoue et al. 2002; Lind et al. 2009; McDonnell et al. 2005). Despite the vast number of papers touting the benecial antimicrobial effects of silver nanomaterials, a relatively modest number of studies have attempted to elucidate the mechanisms by which silver nanomaterials exert this antimicrobial activity. As a result, the mechanisms are not widely understood or agreed upon. For bacteria, commonly proposed mechanisms in the literature begin with the release of silver ions (Hwang et al. 2008; Smetana et al. 2008) followed by generation of reactive oxygen species (ROS) (Hwang et al. 2008; Kim et al. 2007) and cell membrane damage (Choi et al. 2008; Raf et al. 2008; Smetana et al. 2008; Sondi and Salopek-Sondi 2004), but there are many contradictory ndings reported. The more widespread our use of silver nanomaterials becomes the more widespread will become the potential for human and ecosystem exposure. Silver nanoparticles may be released to the environment from discharges at the point of production, from erosion of engineered materials in household products (e.g., antibacterial coatings and silver-impregnated water lters), and from washing or disposal of silver-containing products (Benn and Westerhoff 2008). Silver released to both natural and engineered systems will likely impact the lowest trophic levels rst, i.e., bacteria. However, little is known about trophic transfer of silver and impacts to higher organisms. Indeed, silver nanoparticles have already been proven toxic to both aerobic and anaerobic bacteria isolated from wastewater treatment plants (Choi and Hu 2008), which we speculate could lead to severe disruption of this critical environmental infrastructure if the load of silver into wastewater treatment plants increases signicantly. Given the vast number of products leveraging the benets of silver, it seems prudent to assess the potential human and ecosystem hazards associated with its increased utilization. The main routes of human exposure would be the respiratory system, gastrointestinal system, and skin, which are interfaces

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between the internal systems of the human body and the external environment (Chen and Schluesener 2008). For example, silver nanomaterials may enter through the respiratory tract due to inhalation of dust or fumes containing silver nanomaterials at the point of manufacture, it may be ingested from water, childrens toys, or food containers treated with silver, or it may penetrate the skin via silver-containing textiles and cosmetics. Additionally, other potential entryways could include the female genital tract (due to incorporation of silver nanoparticles into numerous female hygienic products), via systemic administration as it is used for some imaging and therapeutic purposes (Chen and Schluesener 2008; Schrand et al. 2008; West and Halas 2003), or by incorporation into medical implants, catheters, and wound dressings (Furno et al. 2004; Galiano et al. 2008; Maneerung et al. 2008; Roe et al. 2008). In addition to broad-spectrum antimicrobial effects, silver nanoparticles have produced toxic effects in higher cell lines like zebra sh, clams, rats, and humans (Arora et al. 2008, 2009; Asharani et al. 2008; Braydich-Stolle et al. 2005; Hsin et al. 2008; Hussain et al. 2005; Kim et al. 2008a, b; Sung et al. 2008; Yeo and Kang 2008; Yeo and Yoon 2009). Evidence in rodents shows that after entering into the body silver nanoparticles can accumulate and, in some cases, damage tissues such as the liver, lungs, and olfactory bulbs, or penetrate the blood brain barrier (Arora et al. 2009; Braydich-Stolle et al. 2005; Hussain et al. 2005; Sung et al. 2008). A study in human cells concluded that silver can be genotoxic (Asharani et al. 2009). Additionally, a release of ionic silver led to the sterility of Macoma balthica clams in the San Francisco Bay during the 1980s (Brown et al. 2003). If silver nanomaterials exhibit similar or stronger reactivity, the impacts of this isolated event in San Francisco may foreshadow potential ecosystem impacts of silver nanomaterials. The various forms of silver nanomaterials are among the most promising antimicrobial agents being developed from nanotechnology, but the preliminary evidence of effects on higher organisms alerts us to remain cautious of its widespread utilization. This cautiousness demands additional research to determine how to safely design, use, and dispose products containing silver nanomaterials without creating new risks to humans or the environment. Consequently, the goal of this article is to provide a critical review

of the state-of-knowledge about silver nanomaterial antibacterial effects with insights toward better understanding potential implications for human health and the environment.

Typical forms of silver nanomaterials Herein, the term silver nanomaterials refers to any silver-containing materials with enhanced activity due to their nanoscale features. In some cases, commercial products containing metallic silver nanoparticles in the range of 550 nm or ionic silver are given the name nanosilver (Panyala et al. 2008). Silver nanoparticles are nanoscale clusters of metallic silver atoms, Ag0, engineered for some practical purposemost typically antimicrobial and sterile applications. The most common method of producing silver nanoparticles is chemical reduction of a silver salt dissolved in water with a reducing compound such as NaBH4, citrate, glucose, hydrazine, and ascorbate (Gulrajani et al. 2008; Martinez-Castanon et al. 2009; Panacek et al. 2006; Pillai and Kamat 2004). Strong reductants lead to small monodisperse particles, while generating larger sizes can be difcult to control. Weaker reductants produce slower reduction reactions, but the nanoparticles obtained tend to be more polydisperse in size. In order to generate silver nanoparticles with controlled sizes, a two-step method is usually utilized. In this method, nuclei particles are prepared using a strong reducing agent and they are enlarged by a weak reducing agent (Schneider et al. 1994; Shirtcliffe et al. 1999). Since reducing agents for silver nanoparticle synthesis are often considered toxic or hazardous, the use of green synthesis methods is becoming a priority (Panacek et al. 2006). A recent review of green synthesis methods for silver nanoparticles discussed the use of polysaccharides, polyphenols, Tollens agent, irradiation, biological reduction, and polyoxometalate (Sharma et al. 2009). Polysaccharides and polyphenols are typically used as capping agents during silver nanoparticle synthesis, but they may also contribute to reduction of silver ions through as yet poorly understood mechanisms. For polysaccharides, the reduction of silver may be linked to the oxidation of aldehyde groups to carboxylic acid groups (Manzi and van

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Halbeek 1999). Typical polysaccharides used are glucose, starch, and heparin (Batabyal et al. 2007; Huang and Yang 2004; Manno et al. 2008; Singh et al. 2009; Venediktov and Padokhin 2008). In the Tollens method, a silver ammoniacal solution is reduced by an aldehyde forming silver nanoparticles. This method can be altered (i.e., modied Tollens method) by reducing Ag? using saccharides in the presence of ammonia resulting in lms with nanoparticles sizes ranging from 50 to 200 nm and silver hydrosols ranging from 20 to 50 nm (Panacek et al. 2006; Saito et al. 2003; Yu and Yam 2004). Silver nanoparticles can also be synthesized by irradiating silver salts solutions containing reducing and capping agents. Different sources of irradiation have been used such as laser, microwave, ionization radiation, and radiolysis (Abid et al. 2002; Ha et al. 2006; Li et al. 2006; Long et al. 2007; Mahapatra et al. 2007; Pillai and Kamat 2004; Sharma et al. 2007, 2008; Yanagihara et al. 2001; Yin et al. 2004; Zeng et al. 2007). Biological methods involve the production of silver nanoparticles utilizing extracts from bio-organisms as reductant, capping agents or both (Li et al. 2007; Sanghi and Verma 2009). Such extracts can include proteins, amino acids, polysaccharides, and vitamins (Eby et al. 2009; Sharma et al. 2009). Plant extracts such as apiin (a glucoside compound) and leaf extract from magnolia, Persimmon, geranium, and Pine leaf have also been used as reducing agents of Ag? to produce silver nanoparticles (Kasthuri et al. 2009; Shankar et al. 2003; Song and Kim 2009). Additionally, silver nanoparticles can be synthesized by several microorganisms such as the bacterial strains Bacillus licheniformis, K. pneumonia, and fungi strains such as Verticillium and Fusarium oxysporum, Aspergillus avus (Ahmad et al. 2003; Kalishwaralal et al. 2008; Mokhtari et al. 2009; Mukherjee et al. 2001; Senapati et al. 2004; Vigneshwaran et al. 2007). It is not clear if these microbes are impacted (favorably or unfavorably) by exposure to engineered forms of nano-scaled silver, but their seemingly favorable interactions with silver suggest resistance may be fairly widespread. Another procedure utilized to synthesize silver nanoparticles is the solvated metal atom dispersion (SMAD) method (Stoeva et al. 2002). In this method, a metal is co-vaporized with a solvent onto a liquid nitrogen cooled surface, as liquid nitrogen is removed

the metal atoms and solvent warms causing the aggregation of metal atoms. SMAD can be performed in conjunction with digestive ripening, in this way the nanoparticles resulting from SMAD method are further rened by heating them in inert atmosphere in the presence of selected ligands that encourage the particles to reach a narrow size range. As a result, monodisperse spherical particles are obtained (Smetana et al. 2005, 2008). The use of silver ions as antimicrobial agents is limited by the solubility of silver ions in biological and environmental media containing Cl-, because AgCl has a very low solubility and rapidly precipitates out of solution. In some cases, silver salts are stabilized with hyperbranched polymers or dendrimers that act as nanoreactors, wherein silver ions are initially complexed with a specic moiety in the polymeric structure and then reduced to form silver nanoparticles within the polymeric matrix (Fig. 1) (Lesniak et al. 2005; Zhang et al. 2008). Dendrimersilver complexes prevent silver ions from precipitating and keep silver dispersed in the media long enough to be delivered where it is desired (Balogh et al. 2001; Lesniak et al. 2005; Zhang et al. 2008). Silver ions can also be stabilized in zeolite channels (Fig. 2) or deposited in activated carbon bers (Inoue et al. 2002; Ogden et al. 1999; Pal et al. 2009). Composites of silver coatings over titanium dioxide nanoparticles are used in products such as baby bottles and blood-clotting agents to produce antibacterial activity (Yeo and Kang 2008). Other hybrid silver nanomaterials may include silver nanoparticles coated onto polyurethane and silvermagnetite composite nanoparticles (Fe3O4@Ag); both of these hybrids are utilized for water disinfection (Gong et al. 2007; Jain and Pradeep 2005). One of the challenges of using silver (or any) nanoparticles for water treatment is recovering the particles after the treatment process. Silvermagnetite nanoparticles offer the potential advantage of being removed by a magnet, avoiding release to the environment, and making possible direct reuse without additional separation processes. For example, in related research, magnetite particles proved effective for removal of arsenic from water (Mayo et al. 2007; Yavuz et al. 2006). However, for silver materials, an additional concern is controlling the release of metal ions into the nal produce water.

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J Nanopart Res (2010) 12:15311551 Fig. 1 General formation scheme of PAMAM dendrimer complexes and nanocomposites. PAMAM structural subunits: core = ethylenediamine, branching site = N\, chains connecting the branching sites = CH2CH2CONHCH2CH2. Terminal groups on the surface are marked as CH2CH2COZ. Silver ions (represented by M?) can be pre-organize and subsequently contained in the form of solubilized and stabilize, high-surface area silver domains. Redrawn based on (Balogh et al. 2001) Fig. 2 Silver ions stabilized in zeolite channels and ionic exchange of silver ions with other cations in the media. Left picture shows zeolite type A and right picture shows zeolite type X. Adapted from (Auerbach 2003)

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Evidence of silver toxicity in microbes and higher organisms Here, toxicity refers to any deleterious effects on an organism upon exposure to silver. Obviously, if the practical intent is to disinfect or sterilize a specic type of organism, then toxicity may be interpreted as a positive outcome (e.g., antibacterial, antiviral, etc.). However, if the same material exerts unintended or undesired impacts to other organisms, then such toxicity may be interpreted as a potential hazard.

Evidence of toxicity to bacteria Table 1 presents a concise summary of silver nanomaterial antibacterial studies. Kim et al. reported that 13.4-nm silver nanoparticles prepared by reduction of silver nitrate with sodium borohydride show minimum inhibitory concentration (MIC) against E. coli below 6.6 nM and above 33 nM for S. aureus (Kim et al. 2007). In another study, 16-nm silver nanoparticles generated by gas condensation were able to completely inhibit colony forming units (CFU) ability of E. coli at 60 lg/mL (Raf et al. 2008).

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1536 Table 1 Bactericidal activity of nano-scaled silver and silver loaded in zeolite Silver form Silver nanoparticles/ nano-sized silver powders Size data 13.4 nmb Bacterial strain E. coli, S. aureus Key aspects

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References (Kim et al. 2007)

Minimal inhibition concentration against E. coli was lower than 6.6 nM and higher than 33 nM for S. aureus Complete inhibition of CFU ability at 60 lg/mL CFU reduced by 4 to 5 log units Minimal inhibition concentration from 1.69 to 13.5 lg/mL

16 nm 1 lm 26 nm

E. coli E. coli, S. aureus Standard strains and strains isolated from clinical material E. coli, S. aureus L. mesenteroides B. subtilis

(Raf et al. 2008) (Smetana et al. 2008) (Kvitek et al. 2008)

10 nma Silver nanoparticles applied as aerosol Silver nanoparticles stabilized in hyperbranched polymers Silverdendrimer complexes and nanocomposites 14.1710 nmc

Growth inhibition achieved at 5 lg/mL Growth inhibition achieved at 2.5 lg/mL 76% CFU reduction by applying silver nanoparticles aerosol on B. subtilis aerosol Microbial activity increases as silver content in polymer decreases since decrease in silver nanoparticle size Antimicrobial activity was comparable or higher to those of silver nitrate solutions. The activity and solubility did not decrease even in presence of sulfate or chloride ions CFU reduced by 7 log units in 5 min Minimal bactericidal concentration of 78 lg/mL (as Ag?) for bacteria grown is LuriaBertani broth Minimal bactericidal concentration of 39 lg/mL (as Ag?) for bacteria grown is Tryptic Soy broth
c

(Vertelov et al. 2008) (Yoon et al. 2008a, b) (Zhang et al. 2008)

1.47.1 nmb

E. coli, S. aureus, B. subtilis, K. mobilis S. aureus, P. aeruginosa, E. coli

(Balogh et al. 2001)

Silver zeolite Zeolite containing silver and zinc

E. coli E. coli

(Inoue et al. 2002) (Cowan et al. 2003)

E. coli, S. aureus, P. aeruginosa

Note: Size measured by

dynamic light scattering,

TEM images, and

scanning mobility particle sizer

Commercially available silver nanopowders at a concentration of 300 lg/mL and SMAD-produced silver nanoparticles at a concentration of 30 lg/mL were able to reduce (after 10 min contact time) colony forming units of E. coli and S. aureus from 2 9 104 CFU/mL to 0 and \20, respectively (Smetana et al. 2008). Additionally, MICs ranging from 13.5 to 1.69 lg/mL were reported for bacterial strains such as S. aureus CCM 3953, Enterococcus faecalis CCM 4224, E. coli CCM 3954, and P. aeruginosa CCM 3955, and for strains isolated from human clinical material like P. aeruginosa, methicillin-susceptible S. epidermidis, methicillin-resistant S. epidermidis, methicillin-resistant

S. aureus, vancomycin-resistant Enterococcus faecium, and K. pneumonia when exposed to 26-nm silver nanoparticles prepared by the reduction of [Ag(NH3)2]? with D-maltose (Kvitek et al. 2008). Silver nanoparticles also produced 76% reduction of B. subtilis CFU after applying silver nanoparticles with a size distribution from 14 to 710 nm in an aerosol form (Yoon et al. 2008a, b). Likewise, silver nanoparticles are effective against E. coli, S. aureus, and L. mesenteroides. Also, 10-nm Myramistin stabilized silver nanoparticles inhibit growth of E. coli and S. aureus at 2.5 lg/mL and L. mesenteroides at 5 lg/mL (Vertelov et al. 2008).

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Dendrimersilver nanocomposites have also been proven effective antibacterials. For example, poly(amidoamine) dendrimersilver composites have been used against S. aureus, P. aeruginosa, E. coli, B. subtilis, and K. mobilis (Balogh et al. 2001; Zhang et al. 2008). Additionally, silver ions loaded in zeolites elicit antibacterial properties. Two recent studies demonstrated 7 log reduction in CFUs for E. coli, from an initial concentration of 107 CFU/cm3, after 5 min of contact time with 333.3 lg/mL of Ag-loaded

zeolite (Inoue et al. 2002), and MICs of 78 lgAg?/mL for E. coli and 39 lgAg?/mL for S. aureus and P. aeruginosa, plus some bactericidal activity against Listeria monocytogenes (Cowan et al. 2003). Evidence of toxicity to other microorganisms Silver nanoparticles also inactivate fungi, virii, and algae (Table 2). For example, silver nanoparticles of sizes ranging from 1.4 to 7.1 nm and stabilized in

Table 2 Summary of silver nanoparticles toxicity to other microorganisms Strain Fungi A. niger Myramistin stabilized silver nanoparticles Silver nanoparticles stabilized in hyper branched polymers S. cerevisiae T. mentagrophytes C. Albanicas Myramistin stabilized silver nanoparticles Silver nanoparticles Silver nanoparticles 10a 1.47.1b MIC were found to be 5 mg/L Formation of inhibition zones around silver nanoparticles inoculated spots in agar plates MIC were found to be 5 mg/L IC80 between 1 and 4 mg/L Silver nanoparticles inhibited micelial formation, which is responsible for pathogenicity (Vertelov et al. 2008) (Zhang et al. 2008) Silver nanoparticles Size (nm) Key aspects Reference

10a 3b 3b

(Vertelov et al. 2008) (Kim et al. 2008a, b) (Kim et al. 2008a, b)

Silver nanoparticles

3b

Antifungal activity may be exerted (Kim et al. 2009a, b) by cell membrane structure disruption and inhibition of normal budding process Catheter coated with silver nanoparticles inhibited growth and biolm formation. MIC estimated between 6.6 nM and 13.2 nM Formation of inhibition zones around silver nanoparticles inoculated spots in agar plates Inhibition of virus replication (Roe et al. 2008)

Silver nanoparticles coated on plastic catheters Yeast (isolated from bovine mastitis) P. citrinum Silver nanoparticles

318b

13.4b

(Kim et al. 2007)

Silver nanoparticles stabilized in hyper branched polymers Silver nanoparticles Silver nanoparticles

1.47.1b

(Zhang et al. 2008)

Viruses Hepatitis B virus HIV-1 10b 16.19 8.68b (Lu et al. 2008) Only 110 nm nanoparticles (Elechiguerra et al. 2005) attached to virus restraining virus from attaching to host cells. 44% inhibition of Syncitial virus infection 10200a EC50 for the photosynthetic yield was found in 0.35 mg/L of total silver content after 1 h of exposure (Sun et al. 2008)

Syncitial virus Algae C. reinhardtii

Silver nanoparticles

Silver nanoparticles

(Navarro et al. 2008)

Note: Size measured by

dynamic light scattering and

TEM images

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hyperbranched polymers, and silver nanoparticles stabilized with Myramistin (size 10 nm) inhibited the growth of A. niger (Tomsic et al. 2009; Vertelov et al. 2008; Zhang et al. 2008); the same Myramistin stabilized particles were found toxic toward S. cerevisiae showing a MIC of 5 mg/L. Further, 3-nm silver nanoparticles showed IC80 values from 1 to 4 lg/mL against T. mentagrophytes and 2 to 4 lg/mL for C. albanicans (Kim et al. 2008a, b); while in other study, it was reported that the antifungal activity of silver nanoparticles against C. albanicans could be exerted by cell membrane structure disruption leading to reproduction inhibition (Kim et al. 2009a, b). Additionally, silver nanoparticles (sizes ranging from 3 to 18 nm) coated in catheters were able to inhibit growth of C. albicans. Although, in this case, no analysis of the antifungal molecular mechanism was done, the authors speculate that silver ions (Ag?) released from the matrix were the antifungal agents (Kim et al. 2008a, b, 2009a, b; Roe et al. 2008). Furthermore, silver nanoparticles suppress yeast growth and show MIC between 6.6 and 13.2 nM (Kim et al. 2007). Silver nanoparticles of sizes from 1.4 to 7.1 nm and stabilized in hyperbranched polymers (HPAMAMN(CH3)2/AgNPs composite) inhibit P. citrinum growth (Zhang et al. 2008). Although information about toxicity for algae is limited, silver nanoparticles reduced the photosynthetic yield of Chlamydomonas reinhardtii; in this case, the observed toxicity was attributed to Ag? ions (Navarro et al. 2008). Evidence of virus inactivation is also reported in literature. For example, silver nanoparticles of 10 nm

are able to inhibit hepatitis B virus replication (Lu et al. 2008). Additionally, PVP-coated silver nanoparticles in the range 110 nm attach to HIV-1 virus, inhibiting the virus from attaching to host cells (Elechiguerra et al. 2005). In other study, PVP-coated silver nanoparticles reduced respiratory syncytial virus infection by 44% (Sun et al. 2008). Polysulfone ultraltration membranes impregnated with silver nanoparticles of sizes ranging from 1 to 70 nm showed enhanced virus removal, thus improving water disinfection via low pressure membrane ltration (Zodrow et al. 2009).

Evidence of toxicity for mammalian cells Signicant evidence has been reported in relation to the toxicity of silver nanoparticles to higher organisms. It has been shown toxic to sh such as zebrash (Asharani et al. 2008; Yeo and Kang 2008; Yeo and Yoon 2009), Diptera species such as Drosophila melanogaster, known asfruit y (Ahamed et al. 2010) and different mammalian cell lines of mice (Braydich-Stolle et al. 2005; Hussain et al. 2005), rats (Kim et al. 2008a, b; Sung et al. 2008), and also humans (Asharani et al. 2009; Braydich-Stolle et al. 2005; Hsin et al. 2008; Hussain et al. 2005). This review presents only a few, brief examples of silver nanomaterial toxicity for mammalian cells (Table 3). More detailed, focused reviews on this topic are available elsewhere (Chen and Schluesener 2008; Panyala et al. 2008).

Table 3 Evidence of nano-scaled silver toxicity for mammalian cells Target cell/organism Rat lung cells Sprague-Dawley rats Mouse stem cells Rat liver cells Human brosarcoma and human skin/carcinoma Mouse broblast Human colon cancer Human glioblastoma Human broblast Key aspects Reduction in lung function and inammatory lesions Silver nanoparticles accumulation in olfactory bulb and subsequent translocation to the brain Cell leakage and reduction of mitochondrial function Cell leakage and reduction of mitochondrial function Oxidative stress. Low doses produced apoptosis and higher dose necrosis 50 lg/mL induced apoptosis to 43.4% of cells 100 lg/mL produced necrosis to 40.2% of cells Silver nanoparticles were found cytotoxic, genotoxic and antiproliferative Silver nanoparticles were found cytotoxic, genotoxic and antiproliferative (Asharani et al. 2009) (Asharani et al. 2009) Reference (Sung et al. 2008) (Kim et al. 2008a, b) (Braydich-Stolle et al. 2005) (Hussain et al. 2005) (Arora et al. 2008) (Arora et al. 2009)

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Evidence of silver nanoparticle toxicity for mammalian cells was presented in the in vivo studies performed by Sung et al. (2008) and Kim et al. (2008a, b). In the former, a 90-day inhalation study in rats showed that silver nanoparticles reduce lung function and produce inammatory lesions in the lungs. In the later, silver nanoparticles accumulated in the olfactory bulbs of Sprague-Dawley rats and also accumulated in the brain. Evidence from in vitro studies is also available in the literature. For example, silver nanoparticles have been shown to reduce mitochondrial function and to increase membrane leakage of mouse spermatogonial stem cell and rat liver cells (Braydich-Stolle et al. 2005; Hussain et al. 2005). Studies performed on human brosarcoma and human skin/carcinoma cells with silver nanoparticles used in a topical antimicrobial agent concluded that in the presence of the nanoparticles the cellular levels of glutathione are reduced, indicating oxidative stress, that results in cellular damage and lipid peroxidation (Arora et al. 2008). However, in the study performed by Arora et al. the dose required to induce apoptosis (0.781.56 lg/mL) was much smaller than that required to produce necrosis (12.5 lg/mL) in both cell types. Therefore, the authors concluded that, after the required in vivo studies, it would be possible to dene a safe range for the application of silver nanoparticles as a topical antimicrobial agent. Similar differences of the required concentration to cause apoptosis or necrosis were found in a second study by the same authors in mouse broblasts and liver cells (Arora et al. 2009). In this second article, it was suggested that although silver nanoparticles may enter into the cells, the cellular antioxidant mechanisms would limit oxidative stress. Mechanistic studies of silver nanoparticle toxicity in mammalian cells have considered mouse broblast and human colon cancer cells (Hsin et al. 2008). In this study, silver nanoparticle doses of 50 lg/mL induced apoptosis to 43.4% of broblast cells, while 100 lg/mL produced necrosis to 40.2% of the cancer cells. The authors concluded that the apoptotic mechanisms in brosblast cells are a mitochondrial mediated pathway including the generation of ROS in the cell, which activate the apoptosis regulators JNK and p53 proteins inducing protein Bax to migrate to the surface of the mitochondria. That subsequently induces cytochrome C release from mitochondria and cleavage of PARP. Additionally, in a study done by

Asharani, a possible mechanism of toxicity to human cells was proposed (Asharani et al. 2009). Silver nanoparticles would affect the mitochondrial respiratory chain, causing ROS generation and affecting the production of ATP, which subsequently leads to DNA damage. In this study, the authors also concluded that even and small dose of Ag-NP (silver nanoparticles) has the potential to cause toxicity and that silver nanoparticles are cytotoxic, genotoxic, and antiproliferative, being as toxic for human glioblastoma as for normal human broblasts cells.

Mechanisms of silvers antibacterial properties Although the mechanisms behind the activity of nano-scaled silver on bacteria are not yet fully elucidated, the three most common mechanisms of toxicity proposed to date are: (1) uptake of free silver ions followed by disruption of ATP production and DNA replication, (2) silver nanoparticle and silver ion generation of ROS, and (3) silver nanoparticle direct damage to cell membranes. The various observed and hypothesized interactions between silver nanomaterials and bacteria cells are conceptually illustrated in Fig. 3.

Fig. 3 Diagram summarizing nano-scaled silver interaction with bacterial cells. Nano-scaled silver may (1) release silver ions and generate ROS; (2) interact with membrane proteins affecting their correct function; (3) accumulate in the cell membrane affecting membrane permeability; and (4) enter into the cell where it can generate ROS, release silver ions, and affect DNA. Generated ROS may also affect DNA, cell membrane, and membrane proteins, and silver ion release will likely affect DNA and membrane proteins. Similar pictures have been published in (Damm et al. 2008; Neal 2008)

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Free silver ion uptake Silver nanoparticles have been reported to dissolve generating silver ions and it is thought that in vivo this release would be product of reactions of silver nanoparticles with H2O2 (Asharani et al. 2009). Asharani has proposed the following reaction as a possible mechanism for silver nanoparticle oxidative dissolution. 2Ag H2 O2 2H ! 2Ag 2H2 O E0 0:17 V: Asharani suggests that in eukaryotic cells this reaction could occur in the mitochondria, where exists an important concentration of H?. Similarly, we hypothesized that a similar mechanism could occur in the bacterial cell membrane where proton motive force takes place. Another possible mechanism for the oxidative dissolution of silver nanoparticles has been reported by Choi et al., in this case silver is oxidized in the presence of oxygen. Choi et al. speculated that the observed changed of color of their silver nanoparticles suspensions, over a week period, would be attributed to this mechanism. 4Ag O2 2H2 O $ 4Ag 4OH : The amount of free silver ion measured in this case was approximately 2.2% of the total silver content in the silver nanoparticle suspension (Choi et al. 2008). In other article, a 0.1% content of the total silver in partially oxidized silver nanoparticles suspensions was attributed to silver ions (Lok et al. 2007). Ionic silver has known antibacterial properties; thus, it is expected that eluted ions from silver nanoparticles are responsible for at least a part of their antibacterial properties. At sub-micromolar concentrations, Ag? interacts with enzymes of the respiratory chain reaction such as NADH dehydrogenase resulting in the uncoupling of respiration from ATP synthesis. Silver ions also bind with transport proteins leading to proton leakage, inducing collapse of the proton motive force (Dibrov et al. 2002; Holt and Bard 2005; Lok et al. 2006). Silver inhibits the uptake of phosphate and causes the efux of intracellular phosphate (Schreurs and Rosenberg 1982). The interaction with respiratory and transport proteins is due to the high afnity of Ag? with thiol groups present in the cysteine residues of those

proteins (Holt and Bard 2005; Liau et al. 1997; Petering 1976). Additionally, it has been reported that Ag? increases DNA mutation frequencies during polymerase chain reactions (Yang et al. 2009). Bacterial cells exposed to milli-molar Ag? doses suffer morphological changes such as cytoplasm shrinkage and detachment of cell wall membrane, DNA condensation and localization in an electron-light region in the center of the cell, and cell membrane degradation allowing leakage of intracellular contents (Feng et al. 2000; Jung et al. 2008). Physiological changes occur together with the morphological changes. Bacterial cells enter an active, but non-culturable state in which physiological levels can be measured but bacteria are not able to growth and replicate. Several studies have linked the toxicity of silver nanoparticles to the release of silver ions. For example, Smetana et al. observed that silver ions eroded from high-surface area silver powders prepared by SMAD method interacted and destroyed bacterial cells (Smetana et al. 2008). In the same study, a second preparation of silver nanoparticles using water-soluble ligands was used to obtain silver nanoparticles with higher surface area to improve their antibacterial efcacy. However, the second preparation of silver nanoparticles showed lower toxicity toward bacterial cells than uncoated powders, suggesting the ligands prevented silver ion erosion; thus, diminishing the resulting toxicity. Another possible reason for this result is that the surface coatings prevented adhesion of silver nanoparticles to the bacterial cell surface, but the authors did not explore this option. Hwang et al. observed that Ag? induced the same effect in bioluminescence bacteria sensitive to membrane protein damage and slightly less effect in a strain sensitive to superoxides compared to silver nanoparticles (Hwang et al. 2008). The authors suggested that silver nanoparticles produce silver ions that move inside the cell producing ROS through redox reactions with oxygen. In other research, bacterial activity of activated carbon ber supported silver was attributed to the synergistic action of silver ions, superoxides, and hydrogen peroxide (Le Pape et al. 2004). Generation of reactive oxygen species Reactive oxygen species (ROS) are natural byproducts of the metabolism of respiring organisms. While

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small levels can be controlled by the antioxidant defenses of the cells such glutathione/glutathione disulde (GSH/GSSG) ratio, excess ROS production may produce oxidative stress (Nel et al. 2006). The additional generation of free radicals can attack membrane lipids and lead to a breakdown of membrane and mitochondrial function or cause DNA damage (Mendis et al. 2005). Metals can act as catalysts and generate ROS in the presence of dissolved oxygen (Stohs and Bagchi 1995). In this context, silver nanoparticles may catalyze reactions with oxygen leading to excess free radical production. Studies done in eukaryotic cells suggest that silver nanoparticles inhibit the antioxidant defense by interacting directly with GSH, binding GSH reductase or other GSH maintenance enzymes (Carlson et al. 2008). This could decrease the GSH/GSSG ratio and, subsequently, increase ROS in the cell. Silver ions eluted from nano-scaled silver or chemisorbed on its surface may also be responsible for the generation of ROS by serving as electron acceptor. In bacterial cells, silver ions would likely induce the generation of ROS by impairing the respiratory chain enzymes through direct interactions with thiol groups in these enzymes or the superoxideradical scavenging enzymes such as superoxide dismutases (Park et al. 2009). ROS generation from silver nanoparticles and silver ions may also be induced photocatalytically; however, no correlations have been reported between bacterial toxicity and photocatalytic ROS concentration (Choi and Hu 2008). Evidence of toxicity related to ROS generated from silver nanoparticles and silver ions, released from or chemisorbed on its surface, has been shown previously. Kim et al. determined the existence of free radicals from silver nanoparticles by means of spin resonance measurements (Kim et al. 2007). They observed that silver nanoparticles and silver nitrate toxicity was abolished in the presence of an antioxidant, these results leaded them to suggest that the antimicrobial mechanisms of silver nanoparticles against S. aureus and E. coli was related to the formation of free radicals from the surface of silver nanoparticles and subsequent free radical induced membrane damage. Bactericidal activity of silver ions loaded in nanoporous materials such as zeolites has also been related to the generation of ROS. Using ROS

scavengers, it was determined that superoxide anions, hydrogen peroxide, hydroxyl radicals, and singlet oxygen contributed to the antibacterial activity against E. coli (Inoue et al. 2002). In another article, the bactericidal activity of activated carbon ber supported silver occurred only in the presence of oxygen and could not be explained by the silver ions eluted from the ber (Yoon et al. 2008a, b). These observations together with a study of gene expression related to oxidative stress lead the authors to suggest that the strong bactericidal activity of this materials resulted from the combined effects of silver ions, superoxides ions, and hydrogen peroxide (Le Pape et al. 2004). A study of the toxicity effects of different silver forms on nitrifying bacteria showed that not only Ag? and AgCl, but also silver nanoparticles are able to induce intracellular ROS generation. Moreover, the bacterial inhibition of each of these silver species correlates with their individual generation of intracellular ROS concentration. However, at the same level of intracellular ROS concentration, silver nanoparticles appeared more toxic than the other species indicating that other factors besides the generation of intracellular ROS played a role in the overall toxicity (Choi et al. 2008). In other research, a panel of recombinant bioluminescent bacteria was used to test different pathways of silver nanoparticles toxicity such as oxidative stress, DNA damage and protein or membrane damage in bacterial strains. The addition of silver nanoparticles to growing bacterial cultures leaded to the production of superoxide radicals, but not of hydroxyl radicals. The authors conclude that toxicity occurs via membrane protein damage and oxidative stress, but not DNA damage (Hwang et al. 2008). Direct cell membrane damage Silver nanoparticles interact with the bacterial membrane and are able to penetrate inside the cell. Transmission electron microscopy data show that silver nanoparticles adhere to and penetrate into E. coli cells and also are able to induce the formation of pits in the cell membrane (Choi et al. 2008; Raf et al. 2008; Sondi and Salopek-Sondi 2004). Silver nanoparticles have been observed within E. coli cells albeit at sizes much smaller than the original particles; moreover, silver nanoparticles with oxidized surfaces

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induce the formation of huge holes in E. coli surfaces after the interaction and large portions of the cellular content seemed to be eaten away (Smetana et al. 2008). Silver nanoparticle accumulation on the cell membrane and uptake within the cell has also been reported for other bacteria such as V. cholera, P. aeruginosa, and S. typhus. In these cases, only nanoparticles smaller than 10 nm attached to bacteria cell membranes or where observed inside the bacteria (Morones et al. 2005). However, in other research silver nanoparticles with sizes up to 80 nm were transported through the inner and outer membrane of P. aeruginosa (Xu et al. 2004). Silver nanoparticle composites and silver nanoparticles stabilized with surfactants are also thought to interact with cell membranes. Myramistin capped silver nanoparticles showed notable antibacterial activity against gram-positive and -negative bacteria, it was speculated that Myramistin anchors to the cell wall and weakens it allowing penetration of silver nanoparticles inside the cells (Vertelov et al. 2008). Composites of HPAMAMN(CH3)2 silver nanoparticle are thought to catch bacteria by ionic interactions between the cationic polymer and the negative cell membrane, followed by release the silver nanoparticles (Zhang et al. 2008). The detailed mechanism by which silver nanoparticles interact with cytoplasmic membranes and are able to penetrate inside cells is not fully determined. One hypothesis is that the interaction between nanoparticles and bacterial cells are due to electrostatic attraction between negatively charged cell membranes and positively charged nanoparticles (Raf et al. 2008). However, this mechanism does not likely explain the adhesion and uptake of negatively charged silver nanoparticles. It has been also proposed that the preferential sites of interaction for silver nanoparticles and membrane cells might be sulfur containing proteinsin a similar way as silver interacts with thiol groups of respiratory chain proteins and transport proteins, interfering with their proper function (Morones et al. 2005). This seems more likely than electrostatic attraction because evidence of protein membrane damage on bacteria after silver nanoparticle exposure has been demonstrated with bioluminescence bacteria (Hwang et al. 2008). Another proposed mechanism of E. coli membrane damage by silver nanoparticles relates to metal

depletion, i.e., the formation of irregular-shaped pits in the outer membrane and change in membrane permeability by the progressive release of LPS molecules and membrane proteins (Amro et al. 2000; Sondi and Salopek-Sondi 2004). This may be fairly general for gram-negative bacteria. It has been reported that extrusion pump systems such as MexAB-OprM system of P. aeruginosa could also play an important role in controlling the accumulation of silver nanoparticles in living cells (Xu et al. 2004). The same pump systems are responsible for the silver resistance of several bacteria (see Mechanisms of silver resistance section). Despite the mechanism of interaction involved, it is evident that silver nanoparticles attached to bacterial cell membranes increase permeability and disturb respiration. Proteomic data show the accumulation of envelope protein precursors in E. coli cells after exposure to silver nanoparticles (Lok et al. 2006). Energy from ATP and proton motive force is required in order to newly synthesize envelope proteins to be translocated to the membrane, therefore cytoplasmic accumulation of protein precursors suggests dissipation of proton motive force and depletion of intracellular levels of ATP. In a similar way, silver ions have also been linked to the collapse of proton motive force and destabilization of the cell membrane, but the concentrations at which this occur are much higher than the ones of silver nanoparticles (micromolar vs. nanomolar) (Dibrov et al. 2002; Lok et al. 2006). Mechanisms of silver resistance Bacterial strains resistant to specic toxicants are naturally selected in environments where these agents are present (Gupta and Silver 1998). In this way, the widespread use of silver, whether in nano or bulk form, could lead to selection of bacterial communities exhibiting silver resistance. Since most commercial uses of silver and silver nanoparticles relate to ghting infection, widespread silver resistance is a growing concern. In fact, it has been suggested that it already occurs extensively, but it not recognized due to a lack of testing for silver resistance (Silver et al. 2006). For example, 10% of the enteric bacteria tested randomly in a hospital in Chicago, IL, showed genes for Ag? resistance (Silver 2003). Silver resistance in bacteria (and other heavy metals) is often encoded by plasmids genes, as for

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example the Salmonella plasmid pMG101, within the IncHI incompatibility group, as well as in other ve plasmids in the same group (Gupta et al. 2001). Although resistance is mainly encoded by plasmids, bacterial chromosomes have also been reported to encode Ag? resistance genes (sil) in strains such as E. coli K-12 and O157:H7 (Gupta et al. 2001; Silver et al. 2006). The plasmid pMG101 has been studied in detail (Gupta et al. 1999; Silver 2003; Silver et al. 2006), this plasmid has nine genes, whose products have been identied to be proteins responsible for heavy metal resistance. Here, the resistance is achieved by two efux systems SilCBA and SilP acting in combination with two periplasmic binding proteins SilE and SilF. The complex SilCBA is constituted by three proteins SilC in the outer membrane, SilA in the inner membrane and SilB that links SilA and SilC. This complex acts as an antiporter that transports Ag? out of the cell by pumping a proton inside. In turn, SilC is a P-type ATPase that transports Ag? from the cytoplasm to the perisplasm. These two efux pumps work jointly with proteins SilE and SilF, which bind to Ag?. SilF is thought to act as a chaperone by taking one Ag? ion from its release site in SilP to its uptake site in SilCBA while SilE binds 5 Ag? ions to 10 histidines preventing silver ions entrance to the cytoplasm (Gupta et al. 1999). Limited evidence has been reported of the resistance shown by silver-resistant strains to silver nanoparticles. However, in one study, it was reported that albumin-stabilized oxidized silver nanoparticles were unable to inhibit growth of 116 AgNO3R and J53 (pMG101) silver-resistant strains even when apply at a concentration of up to 80 nM (Lok et al. 2007). Therefore, it seems that silver resistance may also be of concern for the efcacy of silver nanoparticles use as antibacterial agent. Bacterial resistance and sensitivity to silver is affected by environmental conditions such as the presence of halides in the media due to mainly the variation of silver bioavailability experienced at different concentrations of Cl- and other halides such as Br- and I- (Gupta et al. 1998). For moderate Cl- concentration, silver is precipitated as AgCl, which decreases silver bioavailability and increases bacterial silver resistance, as observed for E. coli J53 (pMG101). Nevertheless, silver bioavailability rises when ionic complexes such as AgCl2-, AgCl32- are formed as product of a higher halide concentration.

This increase in bioavailability increases sensitivity of the silver-resistant and silver-sensitive bacteria. Similar, behavior is also observed for silver in the presence of other halides (Gupta et al. 1998). Further insights about potential toxicity to higher organisms Silver nanomaterials exhibit outstanding antibacterial properties that give rise to many potentially benecial applications. However, misuses of silver nanoparticles as a bactericide may also result in unintended exposure to higher organisms, including humans. This is of concern because silver nanoparticles appear to have toxic effects in higher organisms (Evidence of silver toxicity in microbes and higher organisms section). While eukaryotic and prokaryotic cells are different in many ways, understanding the fundamental mechanisms governing silver nanoparticle toxicity in bacterial cells may shed light on the potential effects of silver nanoparticles on important organelles such as mitochondria. Mitochondria are believed to have evolved from bacteria (i.e., the serial primary endosymbiosis theory) and they share several characteristics (Kutschera 2009). For example, the inner membrane of a mitochondrion and the prokaryotic cell membrane are structurally very similar. Respiration occurs in bacterial cells similarly as in the mitochondria of eukaryotes, including electron transport, ATP synthesis, and proton motive force. Likewise mitochondrial DNA, the bacterial DNA are analogous, and the division of the mitochondria is similar to ssion of prokaryotes (Slonczewski and Foster 2009). Based on these similarities, it is hypothesized that silver nanoparticles may affect mitochondria in higher organisms via similar mechanisms as in bacterial cellsassuming they can be internalized by the cell.

Factors affecting the toxicity of silver nanomaterials Several factors have been reported to inuence silver nanoparticle toxicity like particle size, shape, crystallinity, surface chemistry, capping agents, as well, as for environmental factors such as pH, ionic strength, and the presence of ligands, divalent cations, and macromolecules (Carlson et al. 2008;

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Choi and Hu 2008; Kvitek et al. 2008; Lok et al. 2007; Morones et al. 2005; Pal et al. 2007; Smetana et al. 2008). Many publications have shown sizedependent toxicities of silver nanoparticles (Carlson et al. 2008; Choi and Hu 2008; Martinez-Castanon et al. 2008; Morones et al. 2005). As particle size decreases, the specic surface area increases leaving a higher number of atoms exposed on the surface available for redox, photochemical, and biochemical reactions in addition to physicochemical interactions with cells. As discussed previously, one of the key mechanisms for silver nanomaterials to exert biocidal activity is through the release of silver ions. As the rate of ion release, in general, is proportional to particle surface area, nanoparticles can release ions more rapidly than larger particles and macroscopic materials. In effect, chemisorbed silver ions would be the cause of the antibacterial activity observed by Lok et al., who reported that only partially oxidized silver nanoparticles exhibit antibacterial activities, while zero valent silver nanoparticles do not (Lok et al. 2007). This surface area effect also inuences RO generation. For example, at the same silver concentration, silver nanoparticles of 15 nm generated higher levels of ROS in macrophages than 30 and 50 nm particles (Carlson et al. 2008). High atom densities at h111i facets increased the toxicity of silver nanoparticles to several bacterial strains; the increased toxicity was the result of the higher reactivity presented by the h111i facets (Morones et al. 2005). Silver nanoparticle shape may also be a factor. Truncated triangular nanoplates exert stronger antibacterial activity than sphericaland rod-shaped silver nanoparticles because they contain more h111i facets; thus, they would be more reactive (Pal et al. 2007). Hence, the antibacterial properties of silver nanoparticles are related to both size (i.e., reactive surface area) and crystallinity (i.e., surface reactivity). Stability of silver nanoparticles also inuences toxicity since the formation of aggregates tends to decrease biocidal activity (Kvitek et al. 2008; Shrivastava et al. 2007; Teeguarden et al. 2007). Different surfactants and polymers (e.g., sodium dodecyl sulfate, polyoxyethylene-sorbitan monooleate, polyvinylpyrrolidone, Na?-carrying poly(glutamic acid), hydroxyl functionalized ionic liquids and hydroxyl functionalized cationic surfactants) have been used to

stabilize silver nanoparticle dispersions and enhance biocidal activity (Dorjnamjin et al. 2008; Kvitek et al. 2008; Yu 2007). However, some ligand-capped silver nanoparticlesalthough highly stable and monodisperse in suspensionwere less bioactive because the capping agent hindered release of silver ions (Smetana et al. 2008). Environmental conditions such as pH, ionic strength, presence of complexing agents, and natural organic matter, also affect the toxicity of silver nanoparticles. High salt concentrations and pH values close to the isoelectric point promote nanoparticle aggregation by screening electrical double layer repulsion (Nel et al. 2009). However, water chemistry also governs silver dissolution and/or re-precipitation through various possible redox and precipitation reactions (Lok et al. 2007). Dissolved Ag? ions are sparingly soluble in the presence of various ligands such as chloride (pK = 9.75), sulfate (pK = 4.9), sulde (pK = 49), hydroxide (pK = 7.8) and dissolved organic carbon (pK [ 7.5) (Choi et al. 2008; Gao et al. 2009). The released Ag? can also form Ag0 -containing clusters through light or chemical reduction (Morones et al. 2005). Therefore, nanomaterials in aqueous suspensions must be considered in a continuous state of ux where the apparent speciation is controlled by the aquatic media pH, redox potential, ionic composition, and exposure to light. Previously, cysteine ligands and chloride were used to scavenge or precipitate eluted silver ions from silver nanoparticles; thus, diminishing their toxicity (Navarro et al. 2008). Sulde ligands have also been used to reduce the inhibition of silver nanoparticles to nitrifying bacteria (Choi et al. 2009). More recently, it was reported that halide ions act as precipitating agents and profoundly affect the bioavailability of Ag? in unexpected ways (Silver 2003). For example, at low chloride concentrations, soluble Ag? binds to cell membranes impacting respiration and producing other toxics effects (Gupta et al. 1998). As chloride concentration increases, silver becomes less bioavailable because the solubility of AgCl is very low. However, further increase in halide concentration results in the formation of water-soluble anionic complexes in the form of AgCl2- and AgCl32-. These anionic complexes are more bioavailable and increase silver toxicity to both silver-sensitive and silver-resistant bacteria. Other halides such as Brand I- have similar effects, but the concentrations at

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which ionic complexes form depends on their respective solubility limits. Finally, aerobic versus anaerobic conditions are important when assessing the antibacterial properties of silver loaded in zeolite or activated carbon bers (Inoue et al. 2002; Le Pape et al. 2002; 2004). Dissolved oxygen is important in these cases because ROS contributes signicantly to bactericidal activity. Dissolved organic carbon (DOC) seems also to inuence the toxicity of silver nanoparticles, specifically, toxicity may decrease for aquatic invertebrates as the concentration DOC increases (Gao et al. 2009). A concise summary of the factors discussed above is presented in Table 4. Since water chemistry dramatically impacts the toxicity for virtually all forms of silver nanomaterials, it may be difcult to predict environmental impacts using laboratory data produced in relatively simple aquatic media or media that is not very representative of natural aquatic chemistries. This presents a major hurdle to science-based policy development for environmental protection.

Summary and future research In summary, silver nanomaterials exhibit broadspectrum biocidal activity toward bacteria, fungi, viruses, and algae. This motivates its use in a large number of biomedical and environmental applications as well as a growing list of consumer products.
Table 4 Factors affecting nano-scaled silver toxicity Factor Particle size Tendency Smaller particles sizes tend to enhance antibacterial properties Higher stability produces a higher antibacterial properties

However, if the amount of nano-scaled silver entering sewers becomes higher than the tolerable levels for microbial communities in wastewater treatment plants, critical environmental infrastructure might be impacted. Further, there is mounting evidence that silver nanoparticles exhibit an array of cytotoxic and genotoxic effects in higher organisms. This raises concern about possible impacts to higher organisms including humans. The antibacterial mechanisms of silver nanomaterials are not fully elucidated, but the prevailing paradigm suggests various combinations of: (1) silver ion release followed by cellular uptake and a cascade of intracellular reactions, (2) extracellular and intracellular generations of ROS, and (3) direct interactions between nano-scaled silver and cell membranes. At sub-micromolar concentrations, silver ions are internalized and react with thiol groups of cellular proteins, which lead to uncoupling of ATP synthesis from respiration, loss of proton motive force, and interference with the phosphate efux system. At milli-molar levels, silver nanoparticles induce detachment of the cell wall membrane from the cytoplasm, possible release of intracellular content, DNA condensation and loss of replication ability. ROS produce oxidative stress resulting in membrane lipid and DNA damage. Finally, silver nanoparticles increase cell membrane permeability and, subsequently, penetrate inside cells to induce any one or the entire cascade of effects just described.

Possible explanation As size decrease, there is lager number of atoms on the surface available to interact with bacteria or to release a higher amount of silver ions Non-stable nanoparticles will tend to form aggregates thus surface area will be reduced and the density of atoms available on the surface will be lower (Kvitek et al. 2008; Shrivastava et al. 2007; Teeguarden et al. 2007) \111[ facets would contain larger atom densities thus more atoms available for interaction (Morones et al. 2005) Since water chemistry affects particle suspension/ solubility, particle size distribution, as well as, bacterial ability to face environmental stresses, water chemistry will affect the interaction between nano-scaled silver and bacteria thus inuencing the resulting toxicity

Particle stability

Particle shape

Particles with shapes containing more \111[ facets like triangular particles tend to have strongest antibacterial properties Depending in a case to case base

Water chemistry

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J Nanopart Res (2010) 12:15311551 Ahmad A, Mukherjee P, Senapati S, Mandal D, Khan M, Kumar R, Sastry M (2003) Extracellular biosynthesis of silver nanoparticles using the fungus Fusarium oxysporum. Colloids Surf B 28:313318. doi:10.1002/cbic.2007 00592 Amro N, Kotra L, Wadu-Mesthrige K, Bulychev A, Mobashery S, Liu G (2000) High-resolution atomic force microscopy studies of the Escherichia coli outer membrane: structural basis for permeability. Langmuir 16:27892796. doi:10.1021/la9910 13x Arora S, Jain J, Rajwade J, Paknikar K (2008) Cellular responses induced by silver nanoparticles: in vitro studies. Toxicol Lett 179:93100. doi:10.1016/j.toxlet.2008.04.009 Arora S, Jain J, Rajwade JM, Paknikar KM (2009) Interactions of silver nanoparticles with primary mouse broblasts and liver cells. Toxicol Appl Pharmacol 236:310318. doi:10.1016/ j.taap.2009.02.020 Asharani P, Wu Y, Gong Z, Valiyaveettil S (2008) Toxicity of silver nanoparticles in zebrash models. Nanotechnology 19:18. doi:10.1088/0957-4484/19/25/255102 Asharani PV, Mun GLK, Hande MP, Valiyaveettil S (2009) Cytotoxicity and genotoxicity of silver nanoparticles in human cells. ACS Nano 3:279290. doi:10.1021/nn800 596w Auerbach SM (2003) Zeolite science and technology. Marcel Dekker, New York Bajpai S, Mohan Y, Bajpai M, Tankhiwale R, Thomas V (2007) Synthesis of polymer stabilized silver and gold nanostructures. J Nanosci Nanotechnol 7:29943010. doi: 10.1166/jnn.2007.911 Balogh L, Swanson D, Tomalia D, Hagnauer G, McManus A (2001) Dendrimersilver complexes and nanocomposites as antimicrobial agents. Nano Lett 1:1821. doi:10.1021/ nl005502p Batabyal S, Basu C, Das A, Sanyal G (2007) Green chemical synthesis of silver nanowires and microbers using starch. J Biobased Mater Bioenergy 1:143147. doi:10.1166/jbmb. 2007.016 Benn T, Westerhoff P (2008) Nanoparticle silver released into water from commercially available sock fabrics. Environ Sci Technol 42:41334139. doi:10.1021/es7032718 Braydich-Stolle L, Hussain S, Schlager J, Hofmann M (2005) In vitro cytotoxicity of nanoparticles in mammalian germline stem cells. Toxicol Sci 88:412419. doi:10.1093/toxsci/ k256 Brown C, Parchaso F, Thompson J, Luoma S (2003) Assessing toxicant effects in a complex estuary: a case study of effects of silver on reproduction in the bivalve, Potamocorbula amurensis, in San Francisco Bay. Hum Ecol Risk Assess 9:95119. doi:10.1080/713609854 Carlson C, Hussain SM, Schrand AM, Braydich-Stolle LK, Hess KL, Jones RL, Schlager JJ (2008) Unique cellular interaction of silver nanoparticles: size-dependent generation of reactive oxygen species. J Phys Chem B 112: 1360813619. doi:10.1021/jp712087m Chen C, Chiang C (2008) Preparation of cotton bers with antibacterial silver nanoparticles. Mater Lett 62:36073609. doi: 10.1016/j.matlet.2008.04.008 Chen X, Schluesener H (2008) Nanosilver: a nanoproduct in medical application. Toxicol Lett 176:112. doi:10.1016/ j.toxlet.2007.10.004

Silver nanoparticle toxicity is inuenced by intrinsic nanoparticle features like size, shape, chemistry, crystallinity, and capping agents, but also by the aquatic chemistry through such factors as solution pH, redox state, ionic strength, and ionic composition. Most of the toxicity data presented in the literature are obtained in relatively simple media like distilled water or cell culture media, which do not reect the aquatic environment inside living organisms or the natural environment. Hence, the surface chemistry, reactivity, and state of dispersion achieved in the laboratory may not be relevant for assessing behavior in real systems. In natural waters, the wide variation of pH, ionic strength, ionic composition, and natural organic matter will induce widely varying aggregation states of silver nanoparticles; thus, resulting in widely varying antimicrobial activities and toxicities. In addition, most toxicity data are obtained by dispersing silver nanomaterials in or on nutrient rich growth media; however, oligotrophic conditions prevail in most natural environmental systems. Microorganisms in these conditions may be more or less vulnerable to silver than predicted by laboratory studies. Although signicant progress has been made to elucidate the mechanisms of silver nanomaterial toxicity, further research is required to fully understand the processes involved and to safely exploit the tremendous antimicrobial properties of silver without jeopardizing human health, critical infrastructure, and the environment. Future in vivo, in vitro, and environmental studies should consider more systematically the various effects of aquatic chemistry on nano-scaled silver fate, transport, and toxicity.
Acknowledgments Financial support for this research was provided by the University of California Toxic Substances Research and Training Program: Lead Campus Program in Nanotoxicology.

References
Abid J, Wark A, Brevet P, Girault H (2002) Preparation of silver nanoparticles in solution from a silver salt by laser irradiation. Chem Commun 792793. doi: 10.1039/ b200272h Ahamed M, Posgai R, Gorey TJ, Nielsen M, Hussain SM, Rowe JJ (2010) Silver nanoparticles induced heat shock protein 70, oxidative stress and apoptosis in Drosophila melanogaster. Toxicol Appl Pharmacol 242:263269. doi: 10.1016/j.taap.2009.10.016

123

J Nanopart Res (2010) 12:15311551 Chi Z, Liu R, Zhao L, Qin P, Pan X, Sun F, Hao X (2009) A new strategy to probe the genotoxicity of silver nanoparticles combined with cetylpyridine bromide. Spectrochim Acta A 72:577581. doi:10.1016/j.saa.2008.10.044 Choi O, Hu Z (2008) Size dependent and reactive oxygen species related nanosilver toxicity to nitrifying bacteria. Environ Sci Technol 42:45834588. doi:10.1021/es703238h Choi O, Deng K, Kim N, Ross L, Surampalli R, Hu Z (2008) The inhibitory effects of silver nanoparticles, silver ions, and silver chloride colloids on microbial growth. Water Res 42:30663074. doi:10.1016/j.watres.2008.02.021 Choi O, Cleuenger T, Deng B, Surampalli R, Ross L, Hu Z (2009) Role of sulde and ligand strength in controlling nanosilver toxicity. Water Res 43:18791886. doi:10.1016/ j.watres.2009.01.029 Cowan M, Abshire K, Houk S, Evans S (2003) Antimicrobial efcacy of a silver-zeolite matrix coating on stainless steel. J Ind Microbiol Biotechnol 30:102106. doi:10.1007/ s10295-002-0022-0 Damm C, Munstedt H (2008) Kinetic aspects of the silver ion release from antimicrobial polyamide/silver nanocomposites. Appl Phys A 91:479486. doi:10.1007/s00339008-4434-1 Damm C, Munstedt H, Rosch A (2008) The antimicrobial efcacy of polyamide 6/silver-nano- and microcomposites. Mater Chem Phys 108:6166. doi:10.1016/j.matchemphys. 2007.09.002 Dibrov P, Dzioba J, Gosink K, Hase C (2002) Chemiosmotic mechanism of antimicrobial activity of Ag? in Vibrio cholerae. Antimicrob Agents Chemother 46:26682670. doi:10.1128/AAC.46.8.2668-2670.2002 Dorjnamjin D, Ariunaa M, Shim Y (2008) Synthesis of silver nanoparticles using hydroxyl functionalized ionic liquids and their antimicrobial activity. Int J Mol Sci 9:807819. doi:10.3390/ijms9050807 Eby D, Schaeublin N, Farrington K, Hussain S, Johnson G (2009) Lysozyme catalyzes the formation of antimicrobial silver nanoparticles. ACS Nano 3:984994. doi:10.1021/ nn900079e Elechiguerra J, Burt J, Morones J, Camacho-Bragado A, Gao X, Lara H, Yacaman M (2005) Interaction of silver nanoparticles with HIV-1. J Nanobiotechnol 3:6. doi:10.1186/ 1477-3155-3-6 Falletta E, Bonini M, Fratini E, Lo Nostro A, Pesavento G, Becheri A, Lo Nostro P, Canton P, Baglioni P (2008) Clusters of poly(acrylates) and silver nanoparticles: structure and applications for antimicrobial fabrics. J Phys Chem C 112:1175811766. doi:10.1021/jp8035814 Feng Q, Wu J, Chen G, Cui F, Kim T, Kim J (2000) A mechanistic study of the antibacterial effect of silver ions on Escherichia coli and Staphylococcus aureus. J Biomed Mater Res 52:662668 Furno F, Morley KS, Wong B, Sharp BL, Arnold PL, Howdle SM, Bayston R, Brown PD, Winship PD, Reid HJ (2004) Silver nanoparticles and polymeric medical devices: a new approach to prevention of infection? J Antimicrob Chemother 54:10191024. doi:10.1093/Jac/Dkh478 Galiano K, Pleifer C, Engelhardt K, Brossner G, Lackner P, Huck C, Lass-Florl C, Obwegeser A (2008) Silver segregation and bacterial growth of intraventricular catheters impregnated with silver nanoparticles in cerebrospinal

1547 uid drainages. Neurol Res 30:285287. doi:10.1179/ 016164107x229902 Gao J, Youn S, Hovsepyan A, Llaneza VL, Wang Y, Bitton G, Bonzongo JCJ (2009) Dispersion and toxicity of selected manufactured nanomaterials in Natural River water samples: effects of water chemical composition. Environ Sci Technol 43:33223328. doi:10.1021/es803315v Gong P, Li HM, He XX, Wang KM, Hu JB, Tan WH, Zhang SC, Yang XH (2007) Preparation and antibacterial activity of Fe3O4@Ag nanoparticles. Nanotechnology 18. doi: 10.1088/0957-4484/18/28/285604 Gulrajani M, Gupta D, Periyasamy S, Muthu S (2008) Preparation and application of silver nanoparticles on silk for imparting antimicrobial properties. J Appl Polym Sci 108: 614623. doi:10.1002/app.27584 Gupta A, Silver S (1998) Molecular geneticssilver as a biocide: Will resistance become a problem? Nat Biotechnol 16: 888. doi: 10.1038/nbt1098-888 Gupta A, Maynes M, Silver S (1998) Effects of halides on plasmid-mediated silver resistance in Escherichia coli. Appl Environ Microbiol 64:50425045 Gupta A, Matsui K, Lo JF, Silver S (1999) Molecular basis for resistance to silver cations in Salmonella. Nat Med 5:183 188. doi:10.1038/5545 Gupta A, Phung LT, Taylor DE, Silver S (2001) Diversity of silver resistance genes in IncH incompatibility group plasmids. Microbiology 147:33933402 Ha H, Xu W, An J, Li D, Zhao B (2006) A simple method to synthesize triangular silver nanoparticles by light irradiation. Spectrochim Acta A 64:956960. doi:10.1016/ j.saa.2005.09.004 Hernandez-Sierra J, Ruiz F, Pena D, Martinez-Gutierrez F, Martinez A, Guillen A, Tapia-Perez H, Castanon G (2008) The antimicrobial sensitivity of Streptococcus mutants to nanoparticles of silver, zinc oxide, and gold. Nanomed Nanotechnol 4:237240. doi:10.1016/j.nano. 2008.04.005 Hlidek P, Biederman H, Choukourov A, Slavinska D (2008) Behavior of polymeric matrices containing silver inclusions. 1Review of adsorption and oxidation of hydrocarbons on silver surfaces/interfaces as witnessed by FTIR spectroscopy. Plasma Process Polym 5:807824. doi: 10.1002/ppap.200800083 Holt K, Bard A (2005) Interaction of silver(I) ions with the respiratory chain of Escherichia coli: an electrochemical and scanning electrochemical microscopy study of the antimicrobial mechanism of micromolar Ag. Biochemistry 44:1321413223. doi:10.1021/bi0508542 Hsin Y, Chena C, Huang S, Shih T, Lai P, Chueh P (2008) The apoptotic effect of nanosilver is mediated by a ROS- and JNK-dependent mechanism involving the mitochondrial pathway in NIH3T3 cells. Toxicol Lett 179:130139. doi: 10.1016/j.toxlet.2008.04.015 Huang H, Yang X (2004) Synthesis of polysaccharide-stabilized gold and silver nanoparticles: a green method. Carbohydr Res 339:26272631. doi:10.1016/j.carres.2004.08.005 Hussain S, Hess K, Gearhart J, Geiss K, Schlager J (2005) In vitro toxicity of nanoparticles in BRL 3A rat liver cells. Toxicol In Vitro 19:975983. doi:10.1016/j.tiv.2005.06.034 Hwang E, Lee J, Chae Y, Kim Y, Kim B, Sang B, Gu M (2008) Analysis of the toxic mode of action of silver nanoparticles

123

1548 using stress-specic bioluminescent bacteria. Small 4:746 750. doi:10.1002/smll.200700954 Ingle A, Gade A, Pierrat S, Sonnichsen C, Rai M (2008) Mycosynthesis of silver nanoparticles using the fungus Fusarium acuminatum and its activity against some human pathogenic bacteria. Curr Nanosci 4:141144. doi: 10.2174/157341308784340804 Inoue Y, Hoshino M, Takahashi H, Noguchi T, Murata T, Kanzaki Y, Hamashima H, Sasatsu M (2002) Bactericidal activity of Ag-zeolite mediated by reactive oxygen species under aerated conditions. J Inorg Biochem 92:3742. doi:10.1016/S0162-0134(02)00489-0 Jain P, Pradeep T (2005) Potential of silver nanoparticle-coated polyurethane foam as an antibacterial water lter. Biotechnol Bioeng 90:5963. doi:10.1002/bit.20368 Jin W, Jeon H, Kim J, Youk J (2007) A study on the preparation of poly(vinyl alcohol) nanobers containing silver nanoparticles. Synthetic Met 157:454459. doi:10.1016/j.synthmet. 2007.05.011 Jung W, Koo H, Kim K, Shin S, Kim S, Park Y (2008) Antibacterial activity and mechanism of action of the silver ion in Staphylococcus aureus and Escherichia coli. Appl Environ Microbiol 74:21712178. doi:10.1128/AEM.02001-07 Jung R, Kim Y, Kim H, Jin H (2009) Antimicrobial properties of hydrated cellulose membranes with silver nanoparticles. J Biomater Sci Polym Ed 20:311324. doi:10.1163/ 156856209X412182 Kalishwaralal K, Deepak V, Ramkumarpandian S, Nellaiah H, Sangiliyandi G (2008) Extracellular biosynthesis of silver nanoparticles by the culture supernatant of Bacillus licheniformis. Mater Lett 62:44114413. doi:10.1016/j.matlet. 2008.06.051 Kasthuri J, Veerapandian S, Rajendiran N (2009) Biological synthesis of silver and gold nanoparticles using apiin as reducing agent. Colloids Surf B 68:5560. doi:10.1016/ j.colsurfb.2008.09.021 Kim J (2007) Antibacterial activity of Ag? ion-containing silver nanoparticles prepared using the alcohol reduction method. J Ind Eng Chem 13:718722 Kim J, Kuk E, Yu K, Kim J, Park S, Lee H, Kim S, Park Y, Park Y, Hwang C, Kim Y, Lee Y, Jeong D, Cho M (2007) Antimicrobial effects of silver nanoparticles. Nanomed Nanotechnol 3:95101. doi:10.1016/j.nano.2006.12.001 Kim K, Sung W, Moon S, Choi J, Kim J, Lee D (2008a) Antifungal effect of silver nanoparticles on dermatophytes. J Microbiol Biotechnol 18:14821484 Kim Y, Kim J, Cho H, Rha D, Kim J, Park J, Choi B, Lim R, Chang H, Chung Y, Kwon I, Jeong J, Han B, Yu I (2008b) Twenty-eight-day oral toxicity, genotoxicity, and genderrelated tissue distribution of silver nanoparticles in Sprague-Dawley rats. Inhal Toxicol 20:575583. doi:10.1080/ 08958370701874663 Kim J, Lee J, Kwon S, Jeong S (2009a) Preparation of biodegradable polymer/silver nanoparticles composite and its antibacterial efcacy. J Nanosci Nanotechnol 9:10981102. doi:10.1166/jnn.2009.C096 Kim K, Sung W, Suh B, Moon S, Choi J, Kim J, Lee D (2009b) Antifungal activity and mode of action of silver nanoparticles on Candida albicans. Biometals 22:235242. doi:10.1007/s10534-008-9159-2

J Nanopart Res (2010) 12:15311551 Kutschera U (2009) Symbiogenesis, natural selection, and the dynamic Earth. Theory Biosci 128:191203. doi:10.1007/ s12064-009-0065-0 Kvitek L, Panacek A, Soukupova J, Kolar M, Vecerova R, Prucek R, Holecova M, Zboril R (2008) Effect of surfactants and polymers on stability and antibacterial activity of silver nanoparticles (NPs). J Phys Chem C 112: 58255834. doi:10.1021/jp711616v Le Pape H, Solano-Serena F, Contini P, Devillers C, Maftah A, Leprat P (2002) Evaluation of the anti-microbial properties of an activated carbon bre supporting silver using a dynamic method. Carbon 40:2954. doi:10.1016/S0008-6223(02) 00246-4 Le Pape H, Solano-Serena F, Contini P, Devillers C, Maftah A, Leprat P (2004) Involvement of reactive oxygen species in the bactericidal activity of activated carbon bre supporting silver bactericidal activity of ACF(Ag) mediated by ROS. J Inorg Biochem 98:10541060. doi:10.1016/j.jinorgbio. 2004.02.025 Lesniak W, Bielinska A, Sun K, Janczak K, Shi X, Baker J, Balogh L (2005) Silver/dendrimer nanocomposites as biomarkers: fabrication, characterization, in vitro toxicity, and intracellular detection. Nano Lett 5:21232130. doi: 10.1021/nl051077u Li Y, Kim Y, Lee E, Cai W, Cho S (2006) Synthesis of silver nanoparticles by electron irradiation of silver acetate. Nucl Instrum Methods B 251:425428. doi:10.1016/ j.nimb.2006.06.019 Li S, Shen Y, Xie A, Yu X, Qiu L, Zhang L, Zhang Q (2007) Green synthesis of silver nanoparticles using Capsicum annuum L. extract. Green Chem 9:852858. doi:10.1039/ b615357g Liau S, Read D, Pugh W, Furr J, Russell A (1997) Interaction of silver nitrate with readily identiable groups: relationship to the antibacterial action of silver ions. Lett Appl Microbiol 25:279283. doi:10.1046/j.1472-765X.1997. 00219.x Lind ML, Jeong BH, Subramani A, Huang XF, Hoek EMV (2009) Effect of mobile cation on zeolite-polyamide thin lm nanocomposite membranes. J Mater Res 24:1624 1631. doi:10.1557/Jmr.2009.0189 Lok C, Ho C, Chen R, He Q, Yu W, Sun H, Tam P, Chiu J, Che C (2006) Proteomic analysis of the mode of antibacterial action of silver nanoparticles. J Proteome Res 5:916924. doi:10.1021/pr0504079 Lok C, Ho C, Chen R, He Q, Yu W, Sun H, Tam P, Chiu J, Che C (2007) Silver nanoparticles: partial oxidation and antibacterial activities. J Biol Inorg Chem 12:527534. doi: 10.1007/s00775-007-0208-z Long D, Wu G, Chen S (2007) Preparation of oligochitosan stabilized silver nanoparticles by gamma irradiation. Radiat Phys chem 76:11261131. doi:10.1016/j.radphyschem. 2006.11.001 Lu L, Sun R, Chen R, Hui C, Ho C, Luk J, Lau G, Che C (2008) Silver nanoparticles inhibit hepatitis B virus replication. Antivir Ther 13:253262 Mahapatra S, Bogle K, Dhole S, Bhoraskar V (2007) Synthesis of gold and silver nanoparticles by electron irradiation at 5 15 keV energy. Nanotechnology 18. doi: 10.1088/09574484/18/13/135602

123

J Nanopart Res (2010) 12:15311551 Maneerung T, Tokura S, Rujiravanit R (2008) Impregnation of silver nanoparticles into bacterial cellulose for antimicrobial wound dressing. Carbohydr Polym 72:4351. doi: 10.1016/j.carbpol.2007.07.025 Manno D, Filippo E, Di Giulio M, Serra A (2008) Synthesis and characterization of starch-stabilized Ag nanostructures for sensors applications. J Non-Cryst Solids 354: 55155520. doi:10.1016/j.jnoncrysol.2008.04.059 Manzi AE, van Halbeek H (1999) Saccharide structure and nomenclature. In: Varki A, Cummings R, Esko J, Freeze H, Hart G, Marth J (eds) Essentials of glycobiology, 1st edn. Cold Spring Harbor Laboratory Press, New York Martinez-Castanon GA, Nino-Martinez N, Martinez-Gutierrez F, Martinez-Mendoza JR, Ruiz F (2008) Synthesis and antibacterial activity of silver nanoparticles with different sizes. J Nanopart Res 10:13431348. doi:10.1007/s11051008-9428-6 Martinez-Castanon G, Nino-Martinez N, Loyola-Rodriguez J, Patino-Marin N, Martinez-Mendoza J, Ruiz F (2009) Synthesis of silver particles with different sizes and morphologies. Mater Lett 63:12661268. doi:10.1016/j.matlet. 2009.02.061 Mayo JT, Yavuz C, Yean S, Cong L, Shipley H, Yu W, Falkner J, Kan A, Tomson M, Colvin VL (2007) The effect of nanocrystalline magnetite size on arsenic removal. Sci Technol Adv Mater 8:7175. doi:10.1016/j.stam.2006.10. 005 McDonnell AMP, Beving D, Wang AJ, Chen W, Yan YS (2005) Hydrophilic and antimicrobial zeolite coatings for gravity-independent water separation. Adv Funct Mater 15:336340. doi:10.1002/adfm.200400183 Mendis E, Rajapakse N, Byun H, Kim S (2005) Investigation of jumbo squid (Dosidicus gigas) skin gelatin peptides for their in vitro antioxidant effects. Life Sci 77:21662178. doi:10.1016/j.lfs.2005.03.016 Mokhtari N, Daneshpajouh S, Seyedbagheri S, Atashdehghan R, Abdi K, Sarkar S, Minaian S, Shahverdi H, Shahverdi A (2009) Biological synthesis of very small silver nanoparticles by culture supernatant of Klebsiella pneumonia: the effects of visible-light irradiation and the liquid mixing process. Mater Res Bull 44:14151421. doi:10.1016/ j.materresbull.2008.11.021 Morones J, Elechiguerra J, Camacho A, Holt K, Kouri J, Ramirez J, Yacaman M (2005) The bactericidal effect of silver nanoparticles. Nanotechnology 16:23462353. doi: 10.1088/0957-4484/16/10/059 Mukherjee P, Ahmad A, Mandal D, Senapati S, Sainkar S, Khan M, Parishcha R, Ajaykumar P, Alam M, Kumar R, Sastry M (2001) Fungus-mediated synthesis of silver nanoparticles and their immobilization in the mycelial matrix: a novel biological approach to nanoparticle synthesis. Nano Lett 1:515519. doi:10.1021/nl0155274 Naidu B, Park J, Kim S, Park S, Lee E, Yoon K, Lee S, Lee J, Gal Y, Jin S (2008) Novel hybrid polymer photovoltaics made by generating silver nanoparticles in polymer: fullerene bulk-heterojunction structures. Sol Energy Mater Sol Cells 92:397401. doi:10.1016/j.solmat.2007.09.017 Navarro E, Piccapietra F, Wagner B, Marconi F, Kaegi R, Odzak N, Sigg L, Behra R (2008) Toxicity of Silver Nanoparticles to Chlamydomonas reinhardtii. Environ Sci Technol 42:89598964. doi:10.1021/es801785m

1549 Neal AL (2008) What can be inferred from bacterium-nanoparticle interactions about the potential consequences of environmental exposure to nanoparticles? Ecotoxicology 17:362371. doi:10.1007/s10646-008-0217-x Nel A, Xia T, Madler L, Li N (2006) Toxic potential of materials at the nanolevel. Science 311:622627. doi: 10.1126/science.1114397 Nel AE, Madler L, Velegol D, Xia T, Hoek EMV, Somasundaran P, Klaessig F, Castranova V, Thompson M (2009) Understanding biophysicochemical interactions at the nanobio interface. Nat Mater 8:543557. doi:10.1038/ nmat2442 Nita T (2008) Synthesis of antimicrobial polymer composition and in vitro drugs release study. e-Polymers Ogden J, Bogdanchikova N, Corker J, Petranovskii V (1999) Structure of silver clusters embedded in erionite channels. Eur Phys J D 9:605608. doi:10.1007/s100530050509 Pal S, Tak Y, Song J (2007) Does the antibacterial activity of silver nanoparticles depend on the shape of the nanoparticle? A study of the gram-negative bacterium Escherichia coli. Appl Environ Microbiol 73:17121720. doi:10.1128/ AEM.02218-06 Pal S, Tak YK, Joardar J, Kim W, Lee JE, Han MS, Song JM (2009) Nanocrystalline silver supported on activated carbon matrix from hydrosol: antibacterial mechanism under prolonged incubation conditions. J Nanosci Nanotechnol 9:20922103. doi:10.1166/jnn.2009.427 Panacek A, Kvitek L, Prucek R, Kolar M, Vecerova R, Pizurova N, Sharma V, Nevecna T, Zboril R (2006) Silver colloid nanoparticles: synthesis, characterization, and their antibacterial activity. J Phys Chem B 110:16248 16253. doi:10.1021/jp063826h ndez EM, Havel J (2008) Silver or silver a-Me Panyala NR, Pen nanoparticles: a hazardous threat to the environment and human health? J Appl Biomed 6:117129 Park H, Kim J, Kim J, Lee J, Hahn J, Gu M, Yoon J (2009) Silver-ion-mediated reactive oxygen species generation affecting bactericidal activity. Water Res 43:10271032. doi:10.1016/j.watres.2008.12.002 Petering H (1976) Pharmacology and toxicology of heavymetals-silver. Pharmacol Ther A 1:127130. doi:10.1016/ 0362-5478(76)90002-4 Pillai Z, Kamat P (2004) What factors control the size and shape of silver nanoparticles in the citrate ion reduction method? J Phys Chem B 108:945951. doi:10.1021/jp037018r Raf M, Hussain F, Bhatti T, Akhter J, Hameed A, Hasan M (2008) Antibacterial characterization of silver nanoparticles against E. coli ATCC-15224. J Mater Sci Technol 24:192196 Rejeski D (2009) Nanotechnology and consumer products. http://www.nanotechproject.org/publications/archive/ nanotechnology_consumer_products/. Accessed 22 February 2010 Roe D, Karandikar B, Bonn-Savage N, Gibbins B, Roullet J (2008) Antimicrobial surface functionalization of plastic catheters by silver nanoparticles. J Antimicrob Chemother 61:869876. doi:10.1093/jac/dkn034 Ruparelia J, Chatteriee A, Duttagupta S, Mukherji S (2008) Strain specicity in antimicrobial activity of silver and copper nanoparticles. Acta Biomater 4:707716. doi: 10.1016/j.actbio.2007.11.006

123

1550 Saito Y, Wang J, Batchelder D, Smith D (2003) Simple chemical method for forming silver surfaces with controlled grain sizes for surface plasmon experiments. Langmuir 19:68576861. doi:10.1021/la0301240 Sambhy V, Sen A (2008) Novel process of incorporating silver biocides into polymers. Chim Oggi 26:1618 Sanghi R, Verma P (2009) Biomimetic synthesis and characterisation of protein capped silver nanoparticles. Bioresour Technol 100:501504. doi:10.1016/j.biortech.2008. 05.048 Sanpui P, Murugadoss A, Prasad P, Ghosh S, Chattopadhyay A (2008) The antibacterial properties of a novel chitosanAg-nanoparticle composite. Int J Food Microbiol 124: 142146. doi:10.1016/j.ijfoodmicro.2008.03.004 Schneider S, Halbig P, Grau H, Nickel U (1994) Reproducible preparation of silver sols with uniform particle-size for application in surface-enhanced raman-spectroscopy. Photochem Photobiol 60:605610. doi:10.1111/j.17511097.1994.tb05156.x Schrand A, Braydich-Stolle L, Schlager J, Dai L, Hussain S (2008) Can silver nanoparticles be useful as potential biological labels? Nanotechnology 19. doi: 10.1088/09574484/19/23/235104 Schreurs W, Rosenberg H (1982) Effect of silver ions on transport and retention of phosphate by Escherichia coli. J Bacteriol 152:713 Senapati S, Mandal D, Ahmad A, Khan M, Sastry M, Kumar R (2004) Fungus mediated synthesis of silver nanoparticles: a novel biological approach. Indian J Phys A 78A:101105 Shankar SS, Ahmad A, Sastry M (2003) Geranium leaf assisted biosynthesis of silver nanoparticles. Biotechnol Progr 19: 16271631. doi:10.1021/Bp034070w Sharma S, Thakur M, Deb M (2007) Synthesis of silver nanoparticles using N-1, N-2-diphenylbenzamidine by microwave irradiation method. J Exp Nanosci 2:251256. doi:10.1080/17458080701753744 Sharma S, Thakur M, Deb MK (2008) Preparation of silver nanoparticles by microwave irradiation. Curr Nanosci 4: 138140 Sharma VK, Yngard RA, Lin Y (2009) Silver nanoparticles: green synthesis and their antimicrobial activities. Adv Colloid Interface Sci 145:8396. doi:10.1016/j.cis.2008. 09.002 Shirtcliffe N, Nickel U, Schneider S (1999) Reproducible preparation of silver sols with small particle size using borohydride reduction: for use as nuclei for preparation of larger particles. J Colloid Interface Sci 211:122129. doi: 10.1006/jcis.1998.5980 Shrivastava S, Bera T, Roy A, Singh G, Ramachandrarao P, Dash D (2007) Characterization of enhanced antibacterial effects of novel silver nanoparticles. Nanotechnology 18. doi: 10.1088/0957-4484/18/22/225103 Silver S (2003) Bacterial silver resistance: molecular biology and uses and misuses of silver compounds. FEMS Microbiol Rev 27:341353. doi:10.1016/S0168-6445(03) 00047-0 Silver S, Phung LT, Silver G (2006) Silver as biocides in burn and wound dressings and bacterial resistance to silver compounds. J Ind Microbiol Biotechnol 33:627634. doi: 10.1007/s10295-006-0139-7

J Nanopart Res (2010) 12:15311551 Singh M, Sinha I, Mandal R (2009) Role of pH in the green synthesis of silver nanoparticles. Mater Lett 63:425427. doi:10.1016/j.matlet.2008.10.067 Slonczewski J, Foster J (2009) Microbiology: an evolving science. W. W. Norton & Co, New York Smetana A, Klabunde K, Sorensen C (2005) Synthesis of spherical silver nanoparticles by digestive ripening, stabilization with various agents, and their 3-D and 2-D superlattice formation. J Colloid Interface Sci 284:521526. doi:10.1016/j.jcis.2004.10.038 Smetana A, Klabunde K, Marchin G, Sorensen C (2008) Biocidal activity of nanocrystalline silver powders and particles. Langmuir 24:74577464. doi:10.1021/la800091y Sondi I, Salopek-Sondi B (2004) Silver nanoparticles as antimicrobial agent: a case study on E. coli as a model for Gram-negative bacteria. J Colloid Interface Sci 275:177 182. doi:10.1016/j.jcis.2004.02.012 Song J, Kim B (2009) Rapid biological synthesis of silver nanoparticles using plant leaf extracts. Bioproc Biosyst Eng 32:7984. doi:10.1007/s00449-008-0224-6 Stoeva S, Klabunde K, Sorensen C, Dragieva I (2002) Gram-scale synthesis of monodisperse gold colloids by the solvated metal atom dispersion method and digestive ripening and their organization into two- and three-dimensional structures. J Am Chem Soc 124:23052311. doi:10.1021/ ja012076g Stohs SJ, Bagchi D (1995) Oxidative mechanisms in the toxicity of metal-ions. Free Radic Biol Med 18:321336. doi: 10.1016/0891-5849(94)00159-H Sun L, Singh A, Vig K, Pillai S, Singh S (2008) Silver nanoparticles inhibit replication of respiratory syncytial virus. J Biomed Nanotechnol 4:149158. doi:10.1166/jbn.2008. 012 Sung J, Ji J, Yoon J, Kim D, Song M, Jeong J, Han B, Han J, Chung Y, Kim J, Kim T, Chang H, Lee E, Lee J, Yu I (2008) Lung function changes in Sprague-Dawley rats after prolonged inhalation exposure to silver nanoparticles. Inhal Toxicol 20:567574. doi:10.1080/089583707 01874671 Teeguarden J, Hinderliter P, Orr G, Thrall B, Pounds J (2007) Particokinetics in vitro: dosimetry considerations for in vitro nanoparticle toxicity assessments. Toxicol Sci 95: 300312. doi:10.1093/toxsci/k165 Tomsic B, Simoncic B, Orel B, Zerjav M, Schroers H, Simoncic A, Samardzija Z (2009) Antimicrobial activity of AgCl embedded in a silica matrix on cotton fabric. Carbohydr Polym 75:618626. doi:10.1016/j.carbpol.2008.09. 013 Venediktov E, Padokhin V (2008) Synthesis of silver nanoclusters in starch aqueous solutions. Russ J Appl Chem 81:20402042. doi:10.1134/S1070427208110323 Vertelov G, Krutyakov Y, Efremenkova O, Olenin A, Lisichkin G (2008) A versatile synthesis of highly bactericidal Myramistin stabilized silver nanoparticles. Nanotechnology 19. doi: 10.1088/0957-4484/19/35/355707 Vigneshwaran N, Ashtaputre N, Varadarajan P, Nachane R, Paralikar K, Balasubramanya R (2007) Biological synthesis of silver nanoparticles using the fungus Aspergillus avus. Mater Lett 61:14131418. doi:10.1016/j.matlet.2006. 07.042

123

J Nanopart Res (2010) 12:15311551 West JL, Halas NJ (2003) Engineered nanomaterials for biophotonics applications: improving sensing, imaging, and therapeutics. Annu Rev Biomed Eng 5:285292. doi: 10.1146/annurev.bioeng.5.011303.120723 Xu X, Brownlow W, Kyriacou S, Wan Q, Viola J (2004) Realtime probing of membrane transport in living microbial cells using single nanoparticle optics and living cell imaging. Biochemistry 43:1040010413. doi:10.1021/bi036231a Xu X, Yang Q, Wang Y, Yu H, Chen X, Jing X (2006) Biodegradable electrospun poly(L-lactide) bers containing antibacterial silver nanoparticles. Eur Phys J D 42:2081 2087. doi:10.1016/j.eurpolymj.2006.03.032 Yanagihara N, Tanaka Y, Okamoto H (2001) Formation of silver nanoparticles in poly(methyl methacrylate) by UV irradiation. Chem Lett 30:796797 Yang W, Shen C, Ji Q, An H, Wang J, Liu Q, Zhang Z (2009) Food storage material silver nanoparticles interfere with DNA replication delity and bind with DNA. Nanotechnology 20. doi: 10.1088/0957-4484/20/8/085102 Yavuz CT, Mayo JT, Yu WW, Prakash A, Falkner JC, Yean S, Cong L, Shipley HJ, Kan A, Tomson M, Natelson D, Colvin VL (2006) Low-eld magnetic separation of monodisperse Fe3O4. Nanocrystals 314:964967. doi:10.1126/ science.1131475 Yeo M, Kang M (2008) Effects of nanometer sized silver materials on biological toxicity during zebrash embryogenesis. Bull Korean Chem Soc 29:11791184 Yeo M, Yoon J (2009) Comparison of the effects of nano-silver antibacterial coatings and silver ions on zebrash embryogenesis. Mol Cell Toxicol 5:2331 Yin H, Yamamoto T, Wada Y, Yanagida S (2004) Large-scale and size-controlled synthesis of silver nanoparticles under

1551 microwave irradiation. Mater Chem Phys 83:6670. doi: 10.1016/j.matchemphys.2003.09.006 Yoon K, Byeon J, Park C, Hwang J (2008a) Antimicrobial effect of silver particles on bacterial contamination of activated carbon bers. Environ Sci Technol 42:12511255. doi: 10.1021/es0720199 Yoon K, Byeon J, Park J, Ji J, Bae G, Hwang J (2008b) Antimicrobial characteristics of silver aerosol nanoparticles against Bacillus subtilis bioaerosols. Environ Eng Sci 25:289293. doi:10.1089/ees.2007.0003 Yu D (2007) Formation of colloidal silver nanoparticles stabilized by Na?-poly(gamma-glutamic acid)-silver nitrate complex via chemical reduction process. Colloids Surf B 59:171178. doi:10.1016/j.colsurfb.2007.05.007 Yu D, Yam V (2004) Controlled synthesis of monodisperse silver nanocubes in water. J Am Chem Soc 126:13200 13201. doi:10.1021/ja046037r Zeng H, Zhao C, Qiu J, Yang Y, Chen G (2007) Preparation and optical properties of silver nanoparticles induced by a femtosecond laser irradiation. J Cryst Growth 300:519 522. doi:10.1016/j.jcrysgro.2006.11.308 Zhang Y, Peng H, Huang W, Zhou Y, Yan D (2008) Facile preparation and characterization of highly antimicrobial colloid Ag or Au nanoparticles. J Colloid Interface Sci 325: 371376. doi:10.1016/j.jcis.2008.05.063 Zodrow K, Brunet L, Mahendra S, Li D, Zhang A, Li QL, Alvarez PJJ (2009) Polysulfone ultraltration membranes impregnated with silver nanoparticles show improved biofouling resistance and virus removal. Water Res 43: 715723. doi:10.1016/j.watres.2008.11.014

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