B 41 - 2010-11 (1. samling) - Endeligt svar på spørgsmål 1: Spm. om at oversende dokumentation for udtalelsen i artiklen Den Ukendte Bakteriedræber i Fyns Amts Avis den 23.marts.2011, 2. Sektion, side 1, hvor ministeren er citeret for at sige, at der ikke er noget der indikerer at nanoprodukter er så farlige i sig selv, at der er grund til at undgå dem og at det ikke giver mening at bruge forsigtighedsprincippet, for eksempel i forhold til nanosølv, til miljøministeren

ABSTRACT:Nanosilver is one nanomaterial that is currently under a lot of scrutiny. Much of the discussion is based on theassumption that nanosilver is something new that has not been seen until recently and that the advances in nanotechnology openedcompletely new application areas for silver. However, we show in this analysis that nanosilver in the form of colloidal silver has beenused for more than 100 years and has been registered as a biocidal material in the United States since 1954. Fifty-three percent of theEPA-registered biocidal silver products likely contain nanosilver. Most of these nanosilver applications are silver-impregnated waterﬁlters,algicides, and antimicrobial additives that do not claim to contain nanoparticles. Many human health standards for silver arebased on an analysis of argyria occurrence (discoloration of the skin, a cosmetic condition) from the 1930s and include studies thatconsidered nanosilver materials. The environmental standards on the other hand are based on ionic silver and may need to be re-evaluated based on recentﬁndingsthat most silver in the environment, regardless of the original silver form, is present in the form ofsmall clusters or nanoparticles. The implications of this analysis for policy of nanosilver is that it would be a mistake for regulators toignore the accumulated knowledge of our scientiﬁc and regulatory heritage in a bid to declare nanosilver materials as new chemicals,with unknown properties and automatically harmful simply on the basis of a change in nomenclature to the term“nano”.

’INTRODUCTIONThe potential adverse eﬀects of nanoparticles on humans andthe environment currently receive a lot of attention both inacademia and with regulators.1,2A lot of the discussion is centeredon the asserted assumption that nanoparticles are somethingfundamentally“new”and thus cannot be compared to conventionalchemicals or bulk materials. Nanosilver is one of the nanomaterialsthat is under the most scrutiny today3-5and its release and eﬀectsare studied widely.6-9Although changes in nomenclature over thedecades have created confusion among scientists and policymakers, it is undeniable that products containing nanoscale silverparticles have been commercially available for over 100 years andwere used in applications as diverse as pigments, photographics,wound treatment, conductive/antistatic composites, catalysts,and as a biocide. With this long and diverse history of use it isclear that an extraordinary amount of research into the chemistryof nanoscale silver has been conducted over the past 120 years;it should be noted that most research, until very recently, did notuse“nano”nomenclature.In this analysis we critically examine with respect to nanosilverthree important assertions often made when discussing riskassessment of silver nanoparticles:(1) Nanosilver is new and exhibits unique physical andchemical properties compared to“conventional”silver(e.g., macroscale“bulk”silver).(2) Nanosilver has been used for only a few years and theenvironment and humans have never been exposed tonanosilver before.(3) Existing risk assessments of silver have been based on adata set derived from conventional silver materials, sothey do not apply to nanosilver.r2011 American Chemical Society

’ANTIMICROBIAL BIOCIDESAntimicrobial biocides are commonly used to prevent thegrowth of bacteria on surfaces and within materials and aretypically added in small quantities to many applications to makeit more diﬃcult for bacteria to grow on the treated object. Bio-cidal functionality can be achieved by employing either organicor inorganic active agents. Compounds such as quaternaryammonium and chlorinated phenols are examples of two widelyused organic chemical biocidal agents. Inorganic active agents aregenerally based on metals such as silver and copper. Silver hasfound a growing presence in many applications due to a desire toshift away from organic chemical agents toward additives, whichcan be used in much lower concentrations in a wider variety ofproducts including applications such as plastics where high-temperature processing is not feasible for organic compounds.Examples of applications are bacteriostatic waterﬁltersforhousehold use10or swimming pool algicides.11To meet thediversity of application types, many diﬀerent forms of silvercompounds have been developed to service this market.Whereas biocidal action derives from interaction of silverions with bacteria, silver additives are diﬀerentiated primarily bythe way the silver ions are stored in the product. Common silverproducts range from additives that store and release discretesilver ions held within a ceramic (e.g., zeolite) or glass matrix,through to products that store silver ions as silver salts (e.g.,silver chloride) or elemental silver (e.g., nanoscale silver metal).Received:September 30, 2010Accepted:December 16, 2010Revised:December 13, 2010Published:January 10, 20111177dx.doi.org/10.1021/es103316q|Environ. Sci. Technol.2011, 45, 1177–1183

Upon contact with moisture, silver ions are released from theadditive and the object treated with the additive. The biocidalpotency of a silver additive is therefore directly related to thepotential for releasing silver ions.12,13However, in the case offree nanosilver particles the interactions can potentially bemore complex, and catalytic reactions on the particle surfacethat can change as a function of size and shape of the nano-particles can complicate the system.14It should be noted,however, that most commercial applications of nanosilverinvolve embedding the particles within a matrix such as a plasticor a coating.There are two clear“bookends”for illustrating the extremes ofthe potential for silver ion release from silver substances: namelysilver sulﬁde (highly insoluble, hence a low potential for silver ionrelease) and silver nitrate (completely soluble, maximum poten-tial for silver ion release). The release potential of diﬀerent silvermaterials can be distributed between the silver sulﬁde and silvernitrate extremes (Figure 1). Materials that store discrete silverions in a matrix show a high potential for releasing silver ions,only marginally less than silver nitrate. Silver salts such as silverchloride show a lower release potential than the ion-basedmaterials and so are positioned further from silver nitrate. Atthe other extreme, bulk silver metal (e.g., silver ingot) releasessilver ions to a small extent and so has a potential closer to thesilver sulﬁde extreme. As the size of silver metal is decreased frombulk through to micrometer-sized particles through to nanosizedparticles, the potential for releasing silver ions increases becauseof increasing surface availability per mass of silver and becauseboth the solubility and dissolution kinetics of silver may vary as afunction of size as silver metal size decreases. Therefore thepotential for releasing silver ions increases and so the behaviormoves away from the silver sulﬁde bookend toward silver nitrate.It is important to note that while the tendency for higher silverion release improves with smaller silver particle size, the silversalts and silver-ion materials still show higher potential andantimicrobial activity than the nanosized silver metal materials.12

’HISTORY OF NANOSILVER PRODUCTION AND USEOne of the standard deﬁnitions of nanotechnology encom-passes“researchand technology development at the atomic,molecular, or macromolecular levels using a length scale ofapproximately 1-100 nm in any dimension; the creation anduse of structures, devices, and systems that have novel propertiesand functions because of their small size, and the ability to controlor manipulate matter on an atomic scale.”15The unintentionalformation of nanoparticles thus would not fall under this deﬁni-tion of an engineered nanoparticle. It is estimated that todayabout 320 tons/year of nanosilver are produced and usedworldwide16(data on its historical production are not available).Now what about theﬁrstreport of nanosilver? Over 120 yearsago, in 1889, M. C. Lea reported the synthesis of a citrate-stabilized silver colloid.17The average diameter for the particlesobtained by this method is between 7 and 9 nm.18Their size inthe nanoscale and the stabilization by citrate are identical torecent reports about nanosilver formation using silver nitrate andcitrate, e.g., refs 19 and 20. Also the stabilization of nanosilverusing proteins has been described as early as 1902.21Under thename“Collargol”such a kind of nanosilver has been manufac-tured commercially since 1897 and has been used for medicalapplications.22Collargol has a mean particle size of 10 nm23andas early as 1907 its diameter was determined to be in thenanorange.24Other nanosilver preparations were also inventedin the next decades, for example the gelatin stabilized silvernanoparticles patented by Moudry in 1953 with 2-20 nmdiameter25and silver nanoparticle impregnated carbon with adiameter of silver particles below 25 nm.26It is important to notethat the inventors of nanosilver formulations understood decadesago that the viability of the technology required nanoscale silver,e.g., by the following statement from a patent:“forpropereﬃciency, the silver must be dispersed as particles of colloidalsize less than 250 Å [less than 25 nm] in crystallite size”.26Whereas it is true for many other engineered nanomaterials thatthey are novel, e.g., for fullerenes and carbon nanotubes, this is1178dx.doi.org/10.1021/es103316q |Environ.Sci. Technol.2011, 45,1177–1183

clearly not the case for nanosilver. This long history of rationalfabrication and use of colloidal nanosilver has resulted in a lot ofresearch and knowledge about these nanoparticles over the last100 years, even if this research is not reported under“nano”terminology.The nanosilver formulations mentioned in the precedingsection have not only been used by scientists and described inthe patent literature, but have consistently found their way intothe market. In the early part of the 20th century, the commercialsale of medicinal nanoscale silver colloids, known under diﬀerenttrade names such as Collargol, Argyrol, and Protargol, began andover a 50-year period their use became widespread. Thesenanosilver products were sold as over-the-counter medicationsand also used by medical doctors to treat various diseases such assyphilis and other bacterial infections.27

’REGISTRATION OF NANOSILVER PRODUCTS IN THEUNITED STATESApart from these medical applications, many biocidal nano-silver products were developed and registered in the UnitedStates. Information on EPA-registered silver products for the last60 years is contained in the NPIRS database (National PesticideInformation Retrieval System, http://ppis.ceris.purdue.edu/npublic.htm). We searched and evaluated this database for allregistered biocides referring to silver as the active substance, andregistered labels relating to each product were reviewed for theform of the silver and the company name. Additional searches inpublic patent literature and general Internet searches speciﬁc tothe company and product were also used to evaluate the natureof each registered product. Based on these reviews the list ofregistered silver products was divided intoﬁvecategories, fromsurely containing“nano”to not likely containing“nano”(Table 1).Of the biocidal silver products, 53% likely or surely containnanosilver but only 7% are advertised as containing nanoparti-cles. For the majority of the registered products only a combinedevaluation of patents and knowledge on manufacturing tech-niques and materials science can inform about the nature of thesilver present in the product. Figure 2 shows the frequency ofEPA-registered silver products over the years. Theﬁrstbiocidalsilver product registered in the U.S. under the Federal Insecti-cide, Fungicide, and Rodenticide Act (FIFRA) in 1954 wasAlgaedyn, a nanosilver product based on the patent by Moudry25that is still used today as an algicide in residential swimmingpools. After the establishment of EPA in 1970 all silver registra-tions in the next 23 years until 1993 were for nanosilver (colloidalsilver) or for silver nanocomposites. Theﬁrstnon-nanosilverproduct was registered in 1994. It is also apparent that thenumber of“non-nano”silver products has risen substantially inthe last 10 years, necessitating also new scientiﬁc studies onpotential ecotoxicological eﬀects of these“conventional”products.

There are at least three categories of EPA-registered productsthat employ elemental silver particles with particle sizes less than100 nm: (a) silver biocidal additives; (b) silver-impregnatedwaterﬁlters;(c) silver algicides and disinfectants. Several exam-ples of each category are identiﬁed below.Silver Biocidal Additives.EPA has registered numerousbiocidal additives based on elemental silver particles. Table 2contains some examples of currently registered biocidal additiveproducts that contain metallic (elemental) silver with very smallparticle size (<100 nm), including Additive SSB (EPA reg.83587-3, company NanoHorizons), MicroSilver BG-R (EPAreg. 84146-1, company Bio-Gate), and HyGate 4000 (EPA reg.70404-10, company BASF, formerly Ciba Corp.). These silverbiocides are typically used in plastic and textile applicationswhere the silver is effectively contained within polymersubstrates.Silver-Impregnated Water Filters.EPA has registered mul-tiple silver-impregnated water filters since the 1970s. Bacterio-static water filters are generally based on activated carbon orceramics that are impregnated with metallic (elemental) silverexhibiting a very small particle size (<100 nm). It should be notedthat impregnating carbon and ceramic materials with metals iswidely recognized as a standard technique for the synthesis ofnanoscale metal particles. In particular, wet impregnation meth-ods have been employed in production of nanostructuredindustrial catalysts for decades. Numerous peer-reviewed scien-tific literature publications clearly establish that impregnationmethods lead to nanoscale metal particles (e.g., nanosilver)supported on the impregnated substrate (e.g., carbon).28,29Silver-impregnated waterﬁlterscurrently registered underFIFRA employ impregnation-based manufacturing methods to1179dx.doi.org/10.1021/es103316q |Environ.Sci. Technol.2011, 45,1177–1183

US Patent 3,374,608,“SilverImpregnated Carbon” (1968), Pittsburgh Activated Carbon Company, now owned by Calgon (“activatedcarbonimpregnated with a metallic silver having a crystallite size ofnot over 250 A[25 nm].”...“[T]he silver must be dispersed as particles ofcolloidal size (less than250 A”)(emphasis added).bUS Patent 6,165,358,“WaterPuriﬁer for a Spa” (2000). Zodiac Pool Care, Inc.“puriﬁcationmaterials are described, forexample, in U.S. Pat. No. 5,352,369”.US Patent 5,352,369,“Methodof Treating Water” (1994). Fountainhead Technologies Inc.:“theelemental silverpreferably includes at least 2% of silver crystals having crystal sizes between approximately3 nmand10 nm”(emphasis added).c“Theseengineered silverparticles currently vary in size between about10-50 nm in diameter”.(April 26, 2005) William D. Moeller, President, American Biotech LaboratoriesTestimony on Malaria before the U.S. House of Representatives, International Relations Committee, Subcommittee on Africa, Global Human Rights,and International Operations. [http://www.foreignaﬀairs.house.gov/archives/109/20915.pdf] (p.38)].dSNWG“Evaluationof Hazard and ExposureAssociated with Nanosilver and Other Nanometal Oxide Pesticide Products”, Presentation to Scientiﬁc Advisory Panel (November 4, 2009). Docket ID:EPA-HQ-OPP-2009-0683-0165. [http://www.regulations.gov/search/Regs/contentStreamer?objectId=0900006480a52512&disposition=attachment-&contentType=pdf].eCiba Specialty Chemicals (Now BASF)“CibaSpecialty Chemicals forms marketing cooperation with Bio-Gate for silver anti-microbial technology” (December 14, 2005, Basel, Switzerland) [http://cibasc.com/index/med-index.htm?reference=41794&checkSum=C441-84952B5155A13ECC5E419C8F7310] Noteﬁgurecaption“Scanningelectron microscopy showing the high porosity HyGate 4000 powder:primaryparticle size 50-200 nm...”(emphasis added).

achieve the registered metallic silver form,26,30,31in many caseswith the clear intention to produce nanoscale silver. An exampleis given in one patent application from 1994:“theelemental silverpreferably includes at least 2% of silver crystals having crystal sizesbetween approximately 3 nm and 10 nm”.31Silver-impregnatedwaterﬁlterscontain nano-silver particles supported on theﬁltermatrix structure.To verify the presence of nanoscale Ag in waterﬁlters,commer-cially availableﬁlterswere disassembled and the carbon pellets werecrushed and investigated with TEM equipped with a HAADF (HighAngle Annular Dark Field) detector that is sensitive toward heavyelements (Figure 3). Details of the analysis can be found in theSupporting Information. Bright nanoparticles are visible distributedon matrix particles having sizes ranging from a few nm to about 100nm. EDX (energy dispersive X-ray spectroscopy) analysis showedthat the bright particles contain Ag whereas spectra of the light graymatrix background have no detectable Ag signal. These analysesclearly prove that during manufacturing of the silver-impregnatedwaterﬁltersAg nanoparticles are formed and that all silver is presentin nanoparticulate form in theseﬁlters.However, it remains to beinvestigated what the fate of this nanosilver is during use of the waterﬁlters,whether there is only dissolution and release of ionic silver orwhether there is also release of particulate silver.Silver-impregnated waterﬁltershave been safely used fordomestic water applications such as drinking water and swim-ming poolﬁltersfor decades. No reports about any health orenvironmental eﬀects have been reported, although the absenceof such reports does not mean that no eﬀects occurred. However,the many decades long use of these EPA registered nanosilvercontaining products in numerous households presents a uniqueopportunity for epidemiologists to investigate eﬀects of extendeduse of nanosilver on human health.Silver Algicides and Disinfectants.Silver algicides anddisinfectants have been FIFRA registered as biocides since

1954. Colloidal nanosilver algicides are based on elemental silverparticles maintained in a stabilized solution, containing silver invery small particle size (e.g., <100 nm). Table 2 contains someexamples of currently registered biocidal products includingSilver Algaedyn (EPA reg. 68161-1, Pool Products PackagingCorp), Nu-Clo Silvercide (EPA reg. 7124-101, Alden Leeds Inc.)and ASAP-AGX (EPA reg. 73499-1, American Biotech Labora-tories). Figure 4 shows a TEM micrograph of Algaedyn and thesize distribution of the particles determined by image analysis(>3500 particles, details of the analysis can be found in theSupporting Information), proving that Algaedyn contains silvernanoparticles with diameters between 2 and 20 nm.It should be noted that algicide applications have been usedsafely in high-exposure, direct water contact, and down-the-drainapplications such as swimming pool disinfection for decadeswithout any known damaging impact on humans or the environ-ment. The more than 50-year use of these nanosilver productspresents a unique opportunity for environmental scientists tostudy the eﬀects of the discharge of nanosilver into sewersystems, wastewater treatment plants, and natural waters, espe-cially in residential areas with a lot of swimming pools. Alsoepidemiologists could study populations of home-owners usingsilver-based algicides and compare with those using otherbiocides to get data on a population that has been exposed tonanomaterials for decades.

argyria were documented during this period. Argyria is a condi-tion characterized by a bluish-gray discoloration of the skin.33The toxicity of silver is considered to be relatively low3and toxiceﬀects on humans other than argyria are only observed at veryhigh concentrations (e.g., acute oral LD50 for rats is higher than1600 mg kg-1d-1).3,33Gaul and Staud in 193534listed 43 cases ofargyria of which 27 (63%) were caused by Collargol or Argyrol, thusoriginating from the medical use of colloidal nanosilver. Theother cases were caused by silver chloride or silver iodide. Asigniﬁcant portion of the historical toxicological research on theeﬀects of silver on humans can thus be considered early examples of“nanotoxicology”;predatingwhat is currently considered to benanotoxicology35by more than 80 years.In 1939 Hill and Pillsbury36collected the available literatureon silver and colloidal nanosilver toxicology and derived expo-sure limits based on the threshold value above which develop-ment of argyria can be expected. This threshold value was foundto be the intake of 0.9 g of silver over the whole lifetime. Themodern drinking water standard of 100μg/Lfor silver is basedon this value37and thus includes data on nanoscale-silver. Only avery few studies are available that describe toxicity of bulk silveras opposed to nanosilver or dissolved silver. However, for somestandards there is a distinction between metallic and ionic silver;for instance, the American Conference of Governmental Indus-trial Hygienists has established separate threshold limit values formetallic silver (0.1 mg/m3) and soluble compounds of silver(0.01 mg/m3).33We can thus state that with relation to human

toxicology and the legal standards related to occupational andconsumer health, the toxicity of nanosilver has been taken intoaccount and thus the existing standards are suﬃcient to protectconsumers also from novel nanosilver, at least in forms equiva-lent to those available in the 1930s.If we turn our attention to environmental risk assessment ofsilver we see a diﬀerent picture. The data about eﬀects of silver onenvironmental organisms were almost exclusively obtained usingdissolved silver.38Most research on nonmammalian species wasbased on the use of dissolved silver and only recently wereecotoxicological studies with nano-Ag published, e.g., refs 8, 40,and 41. However, it has been questioned if data obtained usingionic silver should be used to derive threshold values in theenvironment.42Silver in natural waters is typically associatedwith the particulate and colloidal fraction43and is thus to someextent naturally present as nanoparticles and metal-sulﬁdeclusters.44Furthermore it should be noted that the latest researchindicates that under real-life environmental exposure conditionsnanosilver is rapidly converted to silver sulﬁde, resulting inmaterial forms that give no measurable impact on wastewatertreatment plants and are much less toxic than ionic silver.44-46The conversion to silver sulﬁde highlights the importance ofaccounting for speciation and passivation of silver materials byubiquitous environmental species under real-life conditions inreal-world risk assessment of nanosilver materials.5Because many of the aquatic species are several orders ofmagnitude more sensitive to silver than mammals and humans1181dx.doi.org/10.1021/es103316q |Environ.Sci. Technol.2011, 45,1177–1183

record of demonstrated safe use together with a long period offormal and successful regulatory oversight. The use of very highdoses of colloidal nanosilver at the beginning of the 20th centuryhas sparked a vast amount of research on the toxicology ofnanosilver, resulting in theﬁrstexposure limits for silver andsubsequent regulations on its use. Clearly nanosilver is a materialthat does notﬁtthe paradigm of a“new”chemical with new andunknown risks. To consider otherwise is to confuse nomencla-ture (nano) instead of considering the material itself.Regulators rightly state that policy needs to be made on thebasis of sound science. Aﬁrstprinciple of science is thatassumptions need to be tested. For example nanosilver isassumed to be a new material because of the term“nano”.However, on close inspection nanosilver materials have a longhistory of relatively safe and regulated use. Historical perspectivealso shows us that nanosilver has been intentionally manufac-tured and adopted commercially across a wide spectrum of every-day applications for decades. For example, EPA-registered silvernanoparticles have been safely used in down-the-drain and high-volume water-contact applications (e.g., swimming pool algicidesand drinking waterﬁltersystems) bringing beneﬁt to millions ofconsumers over a period of 50 years. On balance, a substantialamount is known about silver and silver nanoparticles and thathistorical experience of use and exposure actually points to thesematerials being relatively safe. While there are naturally topicswhere there is ample opportunity to improve scientiﬁc under-standing about nanosilver (e.g., with respect to its environmentalbehavior and eﬀects), it would be a mistake for regulators toignore the accumulated knowledge of our scientiﬁc and regula-tory heritage in a mistaken bid to declare nanosilver materials asnew chemicals, with unknown properties and automaticallyharmful simply on the basis of a change in nomenclature to theterm“nano”.However, this does also not mean that nanosilvershould be treated as harmless without testing its eﬀect, but ratherthat it is suﬃcient to apply the already strict and coherent riskassessment framework for other silver-containing materials andproducts that often have a shorter history of regulated use.

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