Page 1 of 36 

# Proposed Decision Document for the Registration of HeiQ AGS-20 as a

Materials Preservative in Textiles

August 12, 2010

Environmental Protection Agency Office of Pesticide Programs

Antimicrobials DivisionPage 2 of 36 TABLE OF CONTENTS

I. SUMMARY OF EPA’S PROPOSED ACTION .................................................................. 3

II. BACKGROUND ................................................................................................................. .... 4 A. Regulatory History.......................................................................................................... .... 4 B. FIFRA Scientific Advisory Panel (SAP) Meeting.............................................................. 5

III. PRODUCT CHARACTERIZATION................................................................................... 5

IV. HUMAN HEALTH RISK ASSESSMENT........................................................................... 8 A. Toxicology .......................................................................................................................... 8 1. AGS-20 Nanosilver-Silica Composite ........................................................................... 8

2. Nanosilver Particles........................................................................................................ 9

3. Silver Ion .................................................................................................................. .... 11 B. Human Exposure Scenarios and Data Submitted ............................................................. 11 1. Consumer Exposure ..................................................................................................... 11 2. Occupational Exposure................................................................................................. 12

C. Assessment of Potential Risks to Consumers and Workers.............................................. 14

1. Consumer Risk Assessment ......................................................................................... 15 2. Occupational Risk Assessment .................................................................................... 18

V. ENVIRONMENTAL RISK ASSESSMENT...................................................................... 20 A. Environmental Fate and Transport.................................................................................... 20 B. Environmental Toxicity .................................................................................................... 23 C. Environmental Exposure Assessment............................................................................... 24

VI. PROPOSED REGULATORY ACTION............................................................................ 26 A. Legal Framework............................................................................................................. . 26 B. Findings and Proposed Decision....................................................................................... 27 C. Basis for Conditional Registration.................................................................................... 27

1. Data Generation............................................................................................................ 2 7

2. Public Interest............................................................................................................. .. 28

3. No Unreasonable Adverse Effects ............................................................................... 31

References: ............................................................................................... ...34

Appendix A: Data Requirements for the Registration of HeiQ AGS-20Page 3 of 36 I. SUMMARY OF EPA’S PROPOSED ACTION

EPA is announcing and seeking public comment on its proposed decision to conditionally register a pesticide product c ontaining nanosilver as an active ingredient. The product is the subject of an application submitted by HeiQ Ma terials Ag (“HeiQ”) in September 2008. The product is named “HeiQ AGS-20,” and the nanosilver active ingredient is intended for use as a preservative in textile products. Generally speak ing, with respect to pesticides, EPA views a

“nanoscale material” as an active or inert ingredient and any component parts thereof intentionally produced to have at least one dimension that me asures between approximately 1

and 100 nanometers.

EPA proposes to grant a conditiona l registration to this product under section 3(c)(7)(C) of the Federal Insecticide, Fungicide and Rodenticid e Act (FIFRA). The Agency’s basis for the conditional registration is as follows:

1) Insufficient time has elapsed for the generation of data since the requirement for that data was imposed; 2) Use of the pesticide is in the public interest; and 3) Use of the pesticide during the period that th e newly required data is being developed and reviewed by the Agency will not cau se unreasonable adverse effects.

Until very recently, EPA had not reached a position w ith regard to which types of data would be needed to evaluate the potentia l risks to human and the environm ent of AGS-20. This was due in large part to the need to understand and appl y the advice provided in the report from the consultation with the FIFRA Scientific Advisory Panel. EPA has determined that more extensive product chemistry, toxicology, exposure, and envir onmental data are necessary. Because the list of data requirements is being finalized with today’s action, insufficient time has elapsed for HeiQ to have generated the data. Therefore, the Ag ency is proposing to re quire these studies as a condition of registration, allowing sufficient time for the studies to be conducted and for the Agency to review them. Ultimately, the Agency will use these data to determine whether the ingredient can be registered under FIFRA section 3(c)(5).

The Agency has determined that the benefits expected from this proposed pesticide product, together with other considerations, justify the conclusion that granting a conditional registration is in the public interest. EPA believes HeiQ’s pro duct offers potential benefits in terms of both conservation of the environment, through less us e of silver, and cons umer benefits, through prolonged efficacy. Moreover, considerations relati ng to market equity and international trade and promoting innovation lend further support to a determination that a conditional registration is in the public interest. Specifically, some curre nt registrants of silver-based antimicrobial products have recently reported th at their products contain nanoscal e silver material. While EPA approved these registrations wit hout knowledge that these products contained nanoscale silver and without specifically assessing any potential risks that might be associated with the specific nanoparticles contained in those products, th ey are on the market. Arguably, this unfairly disadvantages HeiQ because HeiQ’s competitors ar e in the market ahead of them. None of the nanosilver products have submitted the types of datathat the Agency is proposing to require as aPage 4 of 36 condition for the registration of HeiQ’s nanosil ver product. The Agency believes that in connection with this newly requi red data, treating all of the re gistered nanosilver products equally is in the public interest. In light of this, and in concer t with the Agency’s request to HeiQ for additional data as part of the proposed conditional registration, the Agency intends to require that similar data be developed to support the continue d registration of these other nanoscale products as well.

With respect to the third conclusion, EPA conducted a screening leve l risk assessment of risks to workers, consumers and the environment associ ated with the use of AGS-20 as a materials preservative. EPA also reviewed pertinent litera ture on silver and nanosilver toxicity and fate. Although the available data are limited, the EPA wa s able to complete a screening level risk assessment that indicated, with the exception of wo rker exposures, that the proposed use pattern for AGS-20 will likely lead to low levels of human and environmental exposure. Therefore despite considerable unc ertainty about the risk assessment, EPA concludes that the proposed registration will pose relatively little risk to c onsumers and the environment. As a result, EPA believes that the use of AGS-20 during the peri od of the proposed conditio nal registration would not pose unreasonable adverse effects on the environm ent. To mitigate potential worker risks, EPA is requiring label language and engineeri ng controls to reduce potential exposures.

As a condition of registration, EPA is requiring the company to conduct a number of studies, based on a tiered approach, which will allow the Ag ency to further identify and characterize any potential risks that may be associated with the continued use of HeiQ AG S-20. EPA is including a time limitation of four years on th e registration to ensure the st udies are completed in a timely fashion. The time duration of four years was chosen to allow time for protocol reviews prior to initiation of the studies, comple tion of the studies and Agency review of the studies following completion. The Agency will evaluate these data as they are submitted during the period of the conditional registration to conf irm the product will not cause unr easonable adverse effects to

human health and the environment.

II. BACKGROUND

A. Regulatory History

In September 2008, HeiQ submitted an application for registration of a new antimicrobial pesticide product named HeiQ AGS-20, which is a silver-based product that is proposed for use as a materials preservative additive to coatin gs, polymers and textiles. HeiQ’s application originally claimed that HeiQ AGS-20 is simila r to other currently registered silver-based antimicrobial pesticide products. The company later amended its application to limit the proposed use of HeiQ AGS-20 for treatment onl y of textiles. EPA reviewed the product chemistry data for HeiQ AGS-20 and determined that the nanosilver active ingredient in the product differed from currently re gistered silver-based antimicr obial products. (EPA has not previously registered any pesticide products cont aining as an ingredient nanoscale silver-silica composite structures similar to AGS-20.) Theref ore, EPA reclassified the application under the PRIA (Pesticide Registration Improvement Act) as one involving a “New Active Ingredient Registration.” EPA published a Notice in the Fe deral Register announcing receipt of the HeiQPage 5 of 36 application. See [Federal Register Vol. 75, No. 61, March 31, 2010, page 16109] EPA received comments on this notice, which are located in th e docket for this action. EPA plans to address these comments in conjunction with comment s on this proposed decision document.

B. FIFRA Scientific Advisory Panel (SAP) Meeting

In November, 2009 the Agency convened a meeti ng of the FIFRA Scientific Advisory Panel (SAP) to address a number of questions associat ed with assessing the hazard of and exposure to

nanosilver and other nanoscale metal-base d pesticides. The SAP released its report in January

2010 ( http://www.epa.gov/scipoly/sap/mee tings/2009/november/110309ameetingminutes.pdf ).

In general, the SAP advised that the toxicity of nanosilver particles coul d differ from and might be higher than bulk silver or th e silver ion and that nanosilvers with different physicochemical properties may behave differently in biological systems. In addition, the SAP commented that not enough literature is available to draw any fi rm conclusions regarding human (occupational or consumer) exposures. Potentially, three major rout es exist for human expos ure to nanoparticles: oral, inhalation, and dermal. Only a few studies are known that addr ess the toxicity of nanosilver to humans from exposure by these routes. Nor is there much information on the level of human exposure to nanosilver by these routes, for eith er workers or consumer s using products treated

with AGS-20. The same situation ex ists for environmental fate and transport. For nanosilver, the concentration in the environment, exposure path ways, bioavailability, toxicity and potential impact on ecological systems are s till not quantified. Furthermore, little or no information on the

fate of nanosilver in soils and sediments is found. As a result, the SAP recommended a case-bycase approach to hazard and exposure assessm ent (i.e., product by product). The SAP also advised that existing requirements may have to be adjusted to obtain data appropriate to assess the fate, degradation, metabolism, mobility, di ssipation and accumulation of nanomaterials.

The SAP report further suggested that existing information on conve ntional silver would not be sufficient in assessing potential nanosilver risk s. The SAP recommended that the Agency treat nanosilver differently from its conventional silver counterpart in evaluating proposed nanosilver product applications (in terms of both data requi rements and the conduct of risk assessments). Moreover, the Panel recommended that EPA requi re additional data on the physical chemistry, exposure potential, and the potential hazar d to human health and the environment.

III. PRODUCT CHARACTERIZATION

HeiQ AGS-20 is a silver-silica nanocomposite ma terial that contains silver nanoparticles imbedded in a matrix of amorphous silicon dioxide (SiO 2 ). The SiO 2 fine structure consists of aggregate matrix particles with an average diameter of approximately 1 micron. Each silica particle contains many small silver metal partic les with a typical diameter between 1 and 10 nm (Egger et al., 2009).

HeiQ proposes to formulate its product as a powder which would be used as a materials preservative to treat woven, knitted or non-wove n textiles. The textile s will be treated by application of HeiQ AGS-20 eith er as a surface coating or by in corporation into the startingPage 6 of 36 materials (i.e. the masterbatch) prior to textile manufacture. The types of textiles include those made from natural and synthetic fibers and whic h are used to manufact ure indoor use articles such as sheets, blankets, towels, napkins, outer wear, sportswear, sleepwear, undergarmets, socks and hosiery and outdoor use ar ticles such as sailcloth, tarps, tents and awnings.

EPA reviewed the following guideline studies subm itted by HeiQ and found the data acceptable. (The numbers in parentheses preceding the study name refer to the EPA guidelines for conducting that kind of study.)

• (830.1000, 1620, 1650, 1670): Description of Produc tion Process, Description of Formulating Process, Discussion on Formation of Impurities

• (830.1700) Preliminary Analysis , five batch analysis

• (830.6302): Color

• (830.6303): Physical State

• (830.6304: Odor

• (830.7000) pH

• (830.7200): Melting Point

• (830.7300): Density

NOTE: EPA’s data requirements re gulations often require other type s of data on the physical or chemical properties of a product. EPA has determin ed however, that these types of data were not needed for an evaluation of AGS-20. Specifica lly, EPA proposes not to require the following types of data:

• (830.6314): Oxidizing/Reducing characteristics . Not required as the nanocomposite contains no oxidizing or reducing agent.

• (830.6315): Flammability. Not required as th e product does not contain combustible liquid

• (830.6316): Explodability. Not required, the pr oduct is not potentially explosive

• (830.7100): Viscosity. Not requir ed. Product is not a liquid.

• (830.7370): Dissociation constant. No t required. Material is no t expected to dissociate.

• (830.7550/7560/7570):Partition Coefficient (Log K

ow ). Not required. The product is a

solid.

• (830.7220/7950): Boiling Point, Vapor Pressure . Not required. The product is a solid.

HeiQ submitted data to address the following gui delines, but the Agency has determined that

additional information is needed as follows:

• (830.1550): Product Identity and Composition. Th e submitted data are acceptable, but EPA is requiring additional information a bout the nanocomposite (see Appendix A).Page 7 of 36 • (830.1800): Enforcement Method. The submitted me thod is based on analysis of total

silver. Need the method to include high resolution images of the active.

• (830.7840): Water Solubility. The submitted data for pH 7 are acceptable, but additional data are required for pH 5 and 9.

• (830.1750): Certified limits. The certified lim its of the nanocomposite are not accurately

identified.

• (830.6317): Storage Stability. A Storage Stab ility study was submitted but it was not acceptable because it used an accelerated met hod. The Agency requires a 1-year study be submitted to support this guideline. The study is currently being conducted by HeiQ.

• (830.6320): Corrosion Characteristics. A Co rrosion Characteristics study was submitted but it was not acceptable because it used an ac celerated method. The Agency requires a 1-year study be submitted to support this guideline. The study is currently being conducted by HeiQ.

• (830.7520): Size and Size Distribut ion. Some particle size in formation is given in a published paper (Egger et al., 2009) however, th is information is incomplete. Additional data must be submitted that adequately char acterizes the size distribution of the AGS-20 nanosilver-silica composite and the nanosilver particles in the composite. (See Appendix A).

• (Non-guideline): Surface Area. Some surface area information is given in a published paper (Egger et al., 2009) however this informa tion is incomplete. Additional data must be submitted that adequately characterize s the surface area of the AGS-20 nanosilversilica composite and the nanosilver particles in the composite. Currently there is no Agency guideline available for this type of st udy, however, it is anticipated that this study could be accomplished using methods repor ted in the literatu re. (See Appendix A).

Additionally, the Agency is requiring the following product chemistry data

• (830.1900): Submittal of Samples

• (830.6313): Stability to normal and elevated te mperatures, and metals and metal ions. Also need to test stability to sunlight, detergents, and salinity

.

• (830.7050): UV-vis absorption spectra . This study is a reliable identification method for nanoproducts and one that the SAP recommended.Page 8 of 36 IV. HUMAN HEALTH RISK ASSESSMENT

HeiQ has submitted a number of studies and ot her information to support its application for registration, including informati on relevant to assessing the toxi city of and exposure to their product. In addition, EPA has also reviewed da ta and information from the public literature.

This section discusses EPA’s asse ssment of the potential risks to human health from the use of

AGS-20. First the section addresses information c oncerning toxicity of the different substances to which people may be exposed. Then the secti on addresses potential le vels of exposure and finally it concludes with a quantitative assessmen t of the risks for different exposed populations

and a characterization of the unc ertainties associated with the assessment. As part of these discussions, additional data that EPA proposes to require in order to reduce its uncertainty with respect to assessing the risks of HeiQ AGS-20 are identified.

A. Toxicology

Based on the proposed use pattern, EPA anticipates humans could be exposed to the following

substances: AGS-20 nanosilver-silic a composite particles, silver nanoparticles that break away from the composite particle and silver ions released from the treated textiles.

1. AGS-20 Nanosilver-Silica Composite

Acute Toxicology

HeiQ submitted data from the battery of six requir ed acute toxicity studies, and these studies are

listed in Table 1. The AGS-20 nanosilver silica co mposite exhibited low acute toxicity by all routes of exposure.

Table 1 - Acute Toxicity Profile for HeiQ AGS-20

Study Toxicity Category Status Acute Oral Toxicity III Acceptable Acute Dermal Toxicity III Acceptable Acute Inhalation Toxicity III Acceptable

Primary Eye Irritation III Acceptable Primary Skin Irritation IV Acceptable

Dermal Sensitization Non-Sensitizer Acceptable

It is important to note that th ese acute studies do not characteri ze the hazard of AGS-20 resulting from longer periods of exposure. These studie s also do not evaluate a number of potential endpoints from acute exposure and they are conduc ted only with non-pregnant, adult animals.

Subchronic/Chronic Toxicology

Based on its assertion that it s product was similar to curren tly registered, silver-based antimicrobial products and could rely on data su pporting those registrations, HeiQ did not submit any subchronic or chronic toxicity data on AGS -20. For a non-food use pesticide, such as aPage 9 of 36 material preservative like AGS -20, EPA requires a subchronic toxicity study, a developmental toxicity study, and a battery of mutagenicity studies.

EPA proposes to require HeiQ to conduct studie s to identify the subchr onic hazard of the nanocomposite. EPA will use these data to conf irm that the screening level occupational and

residential risk assessments for AGS-20 descri bed elsewhere in this document provide an

accurate assessment of the risks.

Based on the proposed use pattern for AGS-20 and the possibility of occupational and residential exposures, the Agency has determined that r oute-specific subchronic inhalation and dermal toxicity studies, a combined repeated-dos e oral toxicity study with reproduction and developmental toxicity screening test, and a battery of genetic t oxicity studies are required as

Tier 1 studies to assess the subc hronic/chronic risks of this pr oduct to adults and children. Completion of these studies with the AGS-20 nanosilver -silica composite as the test material will provide the kinds of data typi cally required for a pesticide co ntaining a new active ingredient intended for use as a materials preservative. Additional toxicity studies (Tier 2) may be required based upon the results of the Tier 1 test resu lts and the residential exposure studies (see Appendix A).

2. Nanosilver Particles

In addition to potential exposure to the AGS -20 nanosilver-silica composite, humans may be exposed to silver nanoparticles that might break away from the nanosilver-silica composite. HeiQ did not conduct any research on the toxico logical effects of nanos cale particles that may break away from AGS-20 by the relevant routes of exposure (inhalation, oral, and dermal). However, there are studies in the open scientific literature that investigate the toxicological effects of other nanosilver nanoparticles by th ese routes. It is unknown if AGS-20 silver nanoparticles would behave similarly (toxicologically ) to these nanosilvers us ed in the literature studies because of potential differences in phys icochemical properties (e.g. size, surface area, surface modifications). The FIFR A Scientific Advisory Panel (S AP) report specifically noted issues with regard to size: “for nanosilver spec ifically, the literature does suggest that silver nanoparticles in the range of a bout 1 to 20 nm do possess the gr eatest quantum properties, are most optically active and may have the potential to induce the greatest toxi cities” Further, citing the literature, the SAP suggested “biological activ ity and penetration thro ugh biological barriers depends on particle size”. However, in the ab sence of product-specific data and as further explained below, these studies do provide some insight into what kinds of hazards may be anticipated for any released AGS-20 silver nanoparticles.

HeiQ submitted a literature study (Sung et al ., 2009) to support the assessment of subchronic/chronic toxicity of AGS-20 by the inhalation route of exposure. This study showed toxic effects in the liver (bil e-duct hyperplasia) and lungs (c hronic alveolar inflammation and macrophage accumulation in the lungs of males and fe males, and erythrocyte aggregation) of rats after inhalation of pure nano-sized (18-19 nm) s ilver particles for 13 w eeks at the high-dose level. The Agency considers these effects adverse. Also, significant incr eases in the amount of silver nanoparticles in tissues, such as lungs , liver, olfactory bulb, br ain, kidneys, and blood, was also reported, particularly in the lungs. Fema les had two to three times more silver accumulationPage 10 of 36 in their kidneys than males. Based on th e toxic effects observed, a No-Observed-AdverseEffect-Level (NOAEL) of 133 μg/m 3 (1.4 x 10 6 particle/cm 3

, the mid-level dose tested) was determined. This study indicates to the Agen cy that, if the AGS-20 nanocomposite releases silver nanoparticles which become airborne, and if such nanopartic les display toxicity similar to the nanoparticles used in this st udy, inhalation of sufficient quantitie s of silver na noparticles that break away from AGS-20 may result in adverse health effects.

There is also a study in the open sc ientific literature th at investigates toxic ity of nanosilver via the oral route (Kim, et al, 2008). The reported fi ndings after 28 days of repeated administration of 60 nm silver nanoparticles were liver effects (dilation of the central ve in, bile-duct hyperplasia and increased foci), a coagulative effect on peri pheral blood, and an increa se in serum alkaline phosphatase (ALP) and cholesterol. A dose-dependent increase in nanosilver distribution in many tissues (liver, kidneys, stomach, brain, lung s, testes, and blood), with a two-fold higher accumulation in the kidneys of female rats when compared with male rats across all dose groups, was also reported. A NOAEL of 30 mg/kg/ day (lowest dose level), based on the observed increase in alkaline phosphatase (ALP) and cholesterol at 300 mg/kg/day (mid dose level), was determined. This study indicates to the Agen cy that, if the AGS-20 nanocomposite releases silver nanoparticles which are ingested, and if su ch nanoparticles display t oxicity similar to the nanoparticles used in this study, ingestion of suffi cient quantities of s ilver nanoparticles that break away from AGS-20 may result in adverse health effects.

There are no available subchronic dermal toxicity studies on nanosilver particles in animals. In the absence of dermal toxicity studies, the Ag ency normally uses extrapolation from another route of exposure (usually oral) combined with a dermal penetration study to determine the fraction of a topically applied dos e that is available for system ic absorption (i .e. the dermal absorption factor, or DAF). The Agency typically uses in vivo studies to cal culate a DAF. However, the Agency will consider in vitro studies when in vivo data are also provided that show

the in vitro study is predictive. An in vitro dermal penetration study w ith nanosilver is available in the peer reviewed literature (Larese, et al, 200 9) that utilized intact and damaged human skin. This study found limited penetration in intact sk in (about 0.6%) and increased penetration in damaged skin (3.3%). No data were provided to confirm the predictivity of in vivo results. This study indicates to the Agency that , if they behave similarly to the nanoparticles used in this study, any silver nanopartic les that break away from AGS-20 may be absorbed to some degree through intact skin and that absorption may be greater th rough damaged skin.

The Agency considers the potential for adverse systemic effects of nanosilvers via dermal exposure to be low. There is a single publishe d report of a burn patien t with nanosilver-coated wound dressing, who developed clin ical signs of argyria and el evated serum liver enzymes indicative of liver toxicity along with elevated silver concentrations in blood and urine (Trop, 2006). However, another study in 30 patients reported no adverse effects associated with the use of these dressings in the tr eatment of burns (Viachou et al., 2007). Systemic exposures to nanosilver would be expected to be much greater in burn patients than in normal subjects, since the dermal barrier is compromised.Page 11 of 36 3. Silver Ion

Humans may also be exposed to silver ions that would be released by AGS-20. Conventional silver, and the silver ions it releases, are pesticid es. The SAP concluded that the hazards of silver ions would be the same, whether they came fr om conventional silver or from nanosilver particles. With respect to silver ions, the Agency notes that safe exposure levels for silver have been established by several regulatory agenci es including FDA, OSHA, and EPA based on the common endpoint argyria and using the same human studies. Argyria is a bl ue-gray discoloration

of the skin and is not considered as being of toxicological concern.

B. Human Exposure Scenarios and Data Submitted

EPA expects both consumers and wo rkers are likely to be exposed as a result of the use of AGS20. Consumer exposures are likely to occur during the following scenarios :

1. Inhalation exposure during laundry dr ying of AGS-20 treated fabrics;

2. Dermal exposure while wearing AGS-20 treated fabrics; and

3. Incidental oral exposure from mouthing or sucking on AGS-20 treated fabrics.

Occupational dermal and inhalation exposures ar e likely to occur during the addition of the AGS-20 powder at textile manuf acturing and treatment facilitie s and during subsequent work activities involving the treated textile products. The above exposures could be to AGS-20 composite particles, silver nanopa rticles that break away from th e composite particle, and silver

ions released from the treated textiles.

1. Consumer Exposure

Textile Leaching Studies Submitted

A textile leaching study (MRID 477287-01) was submitted by HeiQ. The study measured the release of silver nanoparticles from textiles treated with their proposed product, AGS-20. The testing was conducted by agitating fabric sample s in room temperature ultrapure water for 24 hours. The wash water was analyzed for silver ions using an ion specific electrode (ISE) and for particles using scanning electron microscopy (S EM) and energy dispersive x-ray spectroscopy (EDX). According to HeiQ, no ionic silver was detected using ISE and no silver particles were detected using SEM/EDX. There were other par ticles detected, but they were confirmed to be non-silver using EDX.

EPA thinks this study is not suffi cient to support HeiQ’s conclusion that no ionic silver and no silver particles were released fr om AGS-20. The ISE method that wa s used to detect silver ions had a detection limit of 160 ppb which is much hi gher than other available methods such as Induction Coupled Plasma ( ICP), which has a detection limit of 0.2 ppb. The magnification used for SEM, which was at the micron scale, was insuffi cient to detect the possible presence of silver nanoparticles that are 1-10 nm in diameter. It is also not known if the si lver nanoparticles would have aggregated and become vi sible at lower magnifications. At best, this study supports thePage 12 of 36 limited conclusions that no micron sized AGS-20 com posite particles were released and that the release of ionic silver did not exceed 160 ppb.

In addition to the study just mentioned above, HeiQ submitted a literature study (Geranio, 2009) that used their product, HeiQ AGS-20, in its te sting. During this study, 9 textiles treated with silver, including two treated with HeiQ AGS-20, were machine washed using an International Standards Organization (ISO) wash-test method to determine the effect of pH, surfactants and bleaching agents. The HeiQ samples were polyester fabrics treated with ei ther a surface coating (NP-PES-SURF) or by incorporat ion into the polyester fiber during manufacturing (NP-PES). Samples of the wash water were analyzed for s ilver ions before and after filtration using 0.45 micron filters and 30 kDa membranes (~5 nm) to separate large and small particulate silver from

silver ions.

As shown in Table 2, the amount of silver rel eased was greater for the surface coating treated fabric which released thirty percent of its silver content during the 1 st wash as compared to incorporated fabric which released 1.3 percent. The analysis of the filtered and unfiltered samples indicated that for the surface coated fabric, 80 percent of th e released silver was composed of particulate >450 nm in size, while for the incorporated fabr ic, roughly 50 percent or more of the silver was composed of particulat es >450 nm in size. The study suggests that the

mechanical stresses related to the ISO wash-t est method, which uses steel balls, were a major cause of particulate release.

Table 2 - Silver Released During Washing of Polyester Fabric Treated with AGS-20

Silver Released (ug/g)# Designation in

Geranio et al. 2009 Designation

in MRID

477287-01 Treatment Type Silver

Content

μg/g* 1st

Wash 2nd

Wash Bleach Cycle

NP-PES-SURF

NP-PES Sample #2 Sample #3 Surface Coating Incorporation 29

99 10.1

1.3 N/A

0.35 N/A

2.7

* Units are in μg silver / g of textile # Units are in μg silver release / g silver in textile

EPA has concluded that the HeiQ treated textile s, particularly when treated with a surface coating, released silver in the form of coarse part iculate, but it is not k nown if this particulate consists of nanosilver particles in fibers or coating fragments, aggr egates of silver nanoparticles, or precipitates of ionic silver. EPA has de termined that an add itional leaching study, which includes electron microscopy to char acterize the particulate, is n eeded. The characterization is

needed to determine if na noparticles could be released from the particulate.

2. Occupational Exposure

Occupational Exposure Studies Submitted

HeiQ submitted a research report that was published in peer reviewed literature (Demou et al., 2008). The purposes of the study that is the subjec t of the research repor t were to: (1) Measure occupational inhalation exposures during the p ilot production of AGS-20 and determine whatPage 13 of 36 control measures are needed to minimize these exposures and (2) Measure the penetration of AGS-20 nanosilver particles through commerc ially available respirator filters.

Occupational Inhalation Exposure Study

Inhalation exposures to particle s were measured during the pilo t scale production of AGS-20. The process monitored was a small scale test vers ion of the full scale process that will be used for the production of AGS-20. Airborne partic ulate concentrations were measured using nonspecific direct reading instruments such as C ondensation Particle Counters (CPC), a Dust Trak TM

aerosol monitor and a Scanning Mobili ty Particle Sizer (SMPS). Although these instruments cannot identify the composition of the particles detected, they are often used to identify and characterize emissions sources in workplaces wher e nanoparticles might be present. In this study, temporal and spatial analysis of particle conc entrations and sizes was performed during AGS-20 production, maintenance and handling. Samples taken during this test occurred at the end of the process, therefore generating th e highest exposure measurement. The results indicated that the highest particle concentrations occurred during production and that these c oncentrations were an order of magnitude above the background particle levels. (Note - heating system combustion byproducts, vehicle exhaust, and electric motors are background sources of nanoparticles.) It was reported by the authors that the production rate was related to both th e profile and magnitude of the airborne particle concentration. It wa s determined that particle re-suspension was not

relevant because airborne concentrations did no t increase when the equipment was operating and production was not occurring. Manual cleaning of the reac tor with a vacuum cleaner was also a major source of particle emission. Mechanical handling of the powder before packaging and transferring of powder caused little or no increase in particle concentrations.

Respirator Filter Penetration

The penetration of particles th at are generated during the prod uction of AGS-20 through four types of respirator filters from two different manufacturers was measured in a filter chamber. Three of these respirator filters were certified to the EN 143:2000 P3 standard, which indicates that they provide the highest level of protecti on while one filter was certified to the EN 149:2001 P2 standard which indicates a lower level of pr otection. Particle laden air from the production process was drawn into the inlet end of the chamber across the resp irator filter and out the outlet

end. Particle concentration measurements we re taken on each side of the filter using two

Condensation Particle Counters. The initial chamber flowrate wa s set at one of two levels (1.0

m 3 /hr and 2.15 m 3 /hr) to represent the inhalation rates for adults engaged in light and moderate

activities. These breathi ng rates correspond to 6.7 liters per minute (LPM) for light activities

and 36 LPM for moderate activities. The result s of the testing indicate d that retention was 99.89 percent or greater for the P3 re spirators and 96.66 percent for the P2 respirator. These results are

consistent with other studies cited in the Natio nal Institute for Occupational Safety and Health Document Approaches to Safe Nanotechnology (NIOSH, 2009) that have reported very high retention rates for respirator filt ers challenged with nanoparticles.Page 14 of 36 Conclusions Regarding the Occupational Exposure Study

The submitted occupational exposure study discu ssed above provides some useful information regarding the exposure potential of AGS-20 but it does not provide quantitative exposure data. EPA proposes to require HeiQ to conduct an in door applicator study to quantify the exposure potential of AGS-20. EPA will us e these data to confirm that the screening level occupational

risk assessments for AGS-20 described elsewh ere in this document provide an accurate

assessment of the risks.

C. Assessment of Potential Risks to Consumers and Workers

EPA’s Margin of Exposure (MOE) Approach to Assess Risk

EPA uses a Margin of Exposure (MOE) approach to assess risk in which a calculated MOE is compared to a target MOE. If the calculated MOE is greater than the target MOE, then EPA

does not have a risk concern. If the calculated MOE is less than the target MOE, EPA does have a risk concern. In this case, mitigation measures such engineering controls and/or personal protective equipment (PPE) are employed until the calculated MOE exceeds the target MOE

. These relationships are summarized below:

MOE CALC > MOE TARGET = Risk is not of concern and mitigation is not required.

MOE CALC < MOE TARGET = Risk is of concern and mitigation is required.

The Target MOE is determined by uncertainty factors. These commonly include a 10X intraspecies uncertainty factor (to account for variation within the human population) and a 10X interspecies uncertainty factor (to account for differences between animals and humans) as well as any database uncertainty factors (up to 10X ). Thus, the Target MOE commonly ranges from

100-1000.

The calculated MOE is determined by dividing the toxicological point of departure (POD) by the estimated dose to which humans will be expos ed. The POD is determined by the dose at the most sensitive endpoint in the most sensitive species for relevant duration and routes of exposure. Commonly, this is a NOAEL from a la boratory animal toxicity study. When a routespecific study (such as a dermal toxicity study ) is not available, a NOAEL from a study using a different route of exposure may be used and the do se adjusted (such as ap plication of a dermal absorption factor to a NOAEL from an oral toxicity study).

MOE CALC =POD (mg/kg/day) ÷ Dose (mg/kg/day)

The Agency’s MOE approach uses mass-based me trics, both for determining the POD and for calculating exposure. The Agen cy is aware of the ongoing deba te within the scientific community that metrics other than mass (such as particle number or surface area) may be more suitable for assessing nanoparticle risks and therefore acknowledge s the potential for limitations

of mass-based risk estimates.Page 15 of 36 Uncertainty Factors and Target MOE for AGS20

The Agency has determined that the Target MO E for AGS 20 is 1000 for all routes of exposure (oral, dermal, and inhalation) and all populations exposed (occupa tional and residential). The Target MOE includes the standard safety factors of a 10X for intraspecies variation and 10X for interspecies extrapolation as we ll as a 10X database uncertainty factor to account for the limited toxicological database for nanosilver.

1. Consumer Risk Assessment

There is the potential for dermal exposures to nanosilver by children an d adults when textiles

treated with AGS-20 are worn and for incidental oral exposures by children when these textiles are put in the mouth. Because th ere are no oral or dermal toxi cology studies available for AGS20 or the nanoparticles that might break away from textiles treated with AGS-20, these exposures were assessed using a 28 day oral toxicity study on 60 nm silver nanoparticles that is reported in the literature (Kim et al., 2007) as a surrogate. For an oral exposure by a toddler, the

POD is the NOAEL from the Kim et al. oral toxicity study (30 mg/kg/day). For dermal exposures, the POD is the NOAEL from the oral toxicity study, divided by a dermal absorption factor (which was estimated from a lite rature study, as described below).

Dermal Exposure Calculation

The dermal exposure was calculated using the following formula:

Exposure = Application Rate * Cloth Density * Su rface Area Exposed * Transfer Efficiency

Where:

• The application rate is 1 00 ppm silver when AGS-20 is incorporated during fiber production and 30 ppm silver when AGS-20 is applied as a coating. These application rates are based on the HeiQ and Gera nio et al. leaching studies.

• The cloth density is 10 mg/cm 2 based on the density of mixed cotton and synthetics. This value is a standard assumption used in O PP risk assessments and was taken from the

HERA Guidance Document Met hodology (AISE/CEFIC, 2005)

• The surface area exposed is 5,700 cm 2 /day which is the median surface area of clothing contacting the skin of a 3-year-old toddler. Th is value is a standard assumption used in

OPP risk assessments and was derived fr om the Child Specific Exposure Factors Handbook (EPA, 2008).

• The clothing-to-skin transfer efficiency factors of 3.1 and 10.1 percent from the HeiQ leaching study are based on a detection limit of 160 ug/liter, a washwater sample volume of 38 ml, a fabric sample weight of 2 grams and a silver content of either 100 ppm or 30 ppm, respectively.Page 16 of 36 Dermal Dose Calculation

The dermal dose was calculated from the derm al exposure using the following formula to account for dermal absorption:

Dose = [Exposure (mg/day)* Dermal Absorp tion Factor (1%)] / Body Weight (kg)

Where:

• The dermal absorption factor is estimated at 1.0 percent in intact skin, based on an in

vitro dermal penetration study published in the peer-reviewed literature (Larese, et al. 2009) in which 0.46 μg/cm 2 nanosilver was found in the rece ptor fluid after application of 70 μg/cm 2 to human skin in diffusion cells (0.46/70 x 100% = 0.59%, rounded up to 1.0%).

• The body weight of a toddler (3 years old) is 15 kg which is a standard assumption from the Exposure Factors Handbook (EPA, 1997).

Dermal MOEs

The MOEs for dermal exposures were calcula ted from the dermal dose using the NOAEL of 30 mg/kg/day from Kim et al., 2007 as the POD. These MOEs are listed in Table 3 and range from 79,000 to 270,000 depending upon the application rate and transfer factor used. These MOEs all exceed the target MOE of 1000 which i ndicates that the risks are not of concern. Although these MOEs were calculated for children, they are protective of adults, since ratio of skin surface area to the body weight is gr eater for children than for adults.

Table 3 - Dermal MOEs for Toddlers Exposed to AGS-20 Treated Textiles

Application

Rate Cloth Density
(mg/cm 2 ) Surface

Area Exposed
(cm 2 /day) Cloth to Skin
Transfer Efficiency Exposure
(mg/day) Dose
(mg/kg/day) MOE
(mg/kg/day)

<3.1% (HeiQ A ) <0.18 0.012 250,000 100 ppm 10 5,700 1.1% (Geranio, No Bleach B ) 0.063 0.0042 710,000

<10.1% (HeiQ A ) <0.17 0.011 270,000 30 ppm 10 5,700 33% (Geranio, No Bleach C ) 0.56 0.038 79,000

A. Based on an LOD of 160 ug/lite r from the HeiQ leaching study. B. Based on Geranio study data for Sample #3 (Incorporated, 1 st Wash, No Bleach) C. Based on Geranio study data for Sample #2 (Surface Coating, 1 st Wash, No Bleach)

Incidental Oral Exposure Calculations

Incidental oral exposures were calculate d using the following general formula:

Exposure = Application Rate * Cloth Density * Surface Area Mouthed * Saliva Extraction EfficiencyPage 17 of 36 Where:

• The textile contains 100 ppm s ilver when AGS-20 is incorpor ated during fiber production and 30 ppm silver when AGS-20 is applied as a coating.

• The cloth density is 10 mg/cm 2 based on the density of mixed cotton and synthetics.

• The surface area of fabric that is mo uthed per day is assumed to be 100 cm 2 (~16 in. 2 ) which is a standard assumption The saliva extraction factors for mouthing fabric are based on the resu lts of the HeiQ and Geranio leaching studies and are the same factors used for the dermal exposure calculations.

Incidental Oral Dose Calculation

The incidental oral dose was calculated from th e incidental oral expos ure using the following

formula:

Dose = Exposure (mg/day) / Body Weight (15 kg)

Incidental Oral MOEs

The MOEs for incidental oral exposures were calc ulated from the incidental oral dose using the NOAEL of 30 mg/kg/day from Kim et al., 2007 as the POD. The estimated MOEs for incidental oral exposures are listed in Table 4 a nd range from 45,000 to 410,000 depending upon the application rate and saliva extract ion factor used. These MOEs all exceed the target MOE of

1000 which indicates that the risks are not of concern.

Table 4 - Incidental Oral MOEs for To ddlers Exposed to AGS-20 Treated Textiles

Application

Rate Cloth Density

(mg/cm 2 ) Surface Area

Mouthed (cm 2 /day) Saliva Extraction Efficiency Exposure

(mg/day) Dose

(mg/kg/day) MOE

(mg/kg/day)

<3.1% (HeiQ leaching study A ) <0.0031 <0.00021 140,000 100 ppm 10 100 1.1% (Geranio, No Bleach B ) 0.0011 0.000073 410,000

<10.1% (HeiQ leaching study A ) <0.0030 <0.00020 150,000 30 ppm 10 100 33% (Geranio, No Bleach C ) 0.0099 0.00066 45,000

A. Based on an LOD of 160 ug/lite r from the HeiQ leaching study B. Based on Geranio study data for Sample #3 (Incorporated, 1 st Wash, No Bleach) C. Based on Geranio study data for Sample #2 (Surface Coating, 1 st Wash, No Bleach)

Conclusions Regarding Risks to Consumers

The risk calculations indicate that the MOEs for dermal and incidental or al exposures range from 45,000 to 71,000 which exceed the target MOE of 1000. The exposure portion of the calculations is based on the existing leaching studie s, the use of which is conservative because of uncertainties surrounding the det ection limits, TEM magnification, and identity of the particulate fraction. If these uncertainties can be addressed by future studies and if leaching is confirmed as lower than these conservative estimates, expos ures would be lower and the MOEs would bePage 18 of 36 higher, reflecting even greater confidence that consumer exposure is unlikely to pose a significant risk. The point of departure used wa s based on an oral toxi city study that was done using different silver nanopartic les, which have a larger partic le size of 60 nm, and a dermal absorption factor of 1 percent from an in vitro study. The Agency rec ognizes the uncertainties regarding extrapolation from an or al route to a dermal route with nanoparticles, the use of an oral study with different sized nanoparticles than t hose that may be released from AGS-20, and the

use of an in vitro dermal absorption study in which the predictivity for in vivo dermal absorption

has not been confirmed.

2. Occupational Risk Assessment

Inhalation Exposures

The occupational handler inhalati on exposures that occur during the mixing and loading of AGS20 powder during textile treatment were asse ssed using the Pesticid e Handlers Exposure Database (PHED) unit exposure values from the Wettable Powder, Open Mixing and Loading Scenario (EPA, 1998). These values are an appr oximation as the AGS-20 powder has a particle size of approximately one micron while wettable powder pesticide formulations have a high proportion of particles that are le ss than 5 microns (Matthews, 2000). To account for the use of full face respiratory protection as suggested in the Risk Assessment of AGS-20 (Submitted by HeiQ as MRID 479344-02) the unit exposure was divided by a protection factor of 50. This factor is listed in the Approached to Safe Nanotechnology (NIOSH, 2009).

The MOEs were calculated as shown in Table 5 by comparing the inhalation exposure to the point of departure of 133 ug/m 3 , which is the NOAEL from a 90 day inhalation toxicity study (Sung, 2009). The MOE is 8 when no respirators are wo rn and it is 430 when full face respirators

are worn. Both of these MOEs indicate a risk of concern because they are less than the Target MOE of 1000. It is important to note, however, that these MOEs are not precise estimates of risk because AGS-20 has a smaller particle size than the wettable powders that were used for the

PHED studies and it is not known if the test mate rial used in the inhalation toxicity study has the same properties as AGS-20.

Table 5 – Inhalation MOEs for AGS-20 Occupational Handlers

Scenario Amount of

Textile Treated per day A Application

Rate B Amount of

ai handled

per day Unit Exposure C
(ug/m 3 /lb ai) Exposure F
(ug/m 3 ) MOE G,H

7.8 D 16 8 Mix/Load AGS-20 Powder During

Textile Treatment 20,000 lb 0.01% silver by weight 2 lb

0.16 E 0.31 430

A. Standard OPP assumption for textile treatment. B. Rate for incorporation treatment included in studies submitted by HeiQ. C. PHED unit exposure data converted to air concentration units based on the mean 8 hour TWA. D. Assuming no respirator is worn. E. Assuming a full face respirator is worn which provides a protection factor of 50. F. Exposure = Amount ai Handled * Unit Exposure G. MOE = NOAEL / Exposure where the NOAEL is 133 ug/m 3 from Sung, 2009. H. The target MOE is 1000 which includes uncertainty fact ors of 10X for interspecies variation, 10X for intraspecies extrapolation and 10X for database uncertainty. MO Es that are less than 1000 indicate risks of concern.Page 19 of 36 Dermal Exposures

Because there are no dermal toxicology studies ava ilable, route to route extrapolation from the oral toxicity study was used to calculate the risks from dermal exposures, as for toddlers (above). As shown in Table 6, the MOE for dermal expos ures is 610,000 which ex ceeds the target MOE

of 1000 and indicates that the risk is not of concern. It should be noted, however, that this MOE is calculated on the assumption that the work ers would wear protective gloves while handling

AGS-20.

Table 6 – Dermal MOEs for AG S-20 Occupational Handlers

Scenario Amount of ai

added or

handled A Unit Exposure B
(mg/lb ai) Exposure C
(mg/day) Dose D
(mg/kg/day) MOE E

Mix/Load AGS-20 Powder During Textile Treatment 2 lb 0.17 0.34 0.0049 610,000

A. Based on the same assumptions that were us ed for the inhalation MOE (see previous table) B. PHED unit exposure data for wettable powder assuming gloves are worn. C. Dermal Exposure = Amount ai Handled * Dermal Unit Exposure D. Dermal Dose = [Dermal Exposure * Dermal Absorption Factor (1%)]/ Body Weight (70 kg) E. Dermal MOE = NOAEL/Dermal Dose where the NOAEL is 30 mg/kg/day

Conclusions Regarding Occupational Risks

The material to which the workers will be exposed depends upon the condition of the AGS-20 powder. If the powder remains intact during hand ling and silver nanoparticles do not break away from the aggregate, workers would be exposed only to micron sized particles which are similar to wettable powder. Although not conclusive, th e submitted literature study suggests that this might be the case because the levels of nanopartic les did not greatly incr ease during the handling of the finished product, which would be expected if the silver nanoparticles broke away from the silica matrix. This finding will be confirmed when the required additional applicator indoor exposure study is completed.

The inhalation MOEs of 8 and 430 indicate risks of concern because they are less than the Target MOE of 1000. These MOEs, however, are not preci se estimates of risk because it is not known if the test material used in the 90 day inhalati on toxicity study has the same properties as AGS20. The dermal MOE of 610,000 indicates that risks for dermal exposure to silver nanoparticles

are not of concern even when the uncertainties are considered. Give n that the calculated

inhalation risks are of co ncern and given the uncertainties in the unit exposures and toxicological endpoints used in the inhalation e xposure calculations, occupationa l handler inhalation exposures should be minimized by using engineering controls such as closed system loading or local exhaust ventilation. The use of engineering cont rols as a primary method of reducing worker exposure to nanoparticles is discussed in Appr oaches to Safe Nanotechnology (NIOSH, 2009). In addition, as a secondary method of exposure control, EPA proposes to require workers to wear full face respirators with high efficiency filte r cartridges (i.e. P100); these respirators will provide a protection factor of 50 (a ssuming that they are properly fitted). The use of gloves and Tyvek suits to minimize dermal exposure will al so be required given the uncertainties in the

dermal risk estimates.Page 20 of 36 Given the engineering control a nd PPE requirements, the Agency anticipates that the inhalation MOEs will be greater than 1000 and the risks wi ll not be of concern. The efficacy of the engineering controls such lo cal exhaust ventilation is depe ndent upon their specific design characteristic but in general it can be assumed th at local exhaust without enclosure will result in a 3 to 10 fold reduction in exposure (AIHA, 2006, Marquart, 2008, Burge ss, 1995) while local

exhaust with enclosure will result in a 30 to 100 fold reduction in exposure (Marquart, 2008, AIHA, 2006). With respect to closed system loading, the use of cont ainment without local

exhaust ventilation can result in a 3X reduction in exposure (Marquart, 2008) and the use of water soluble packaging can result in a 180X reducti on in exposure (EPA, 1998).

V. ENVIRONMENTAL RISK ASSESSMENT

Based on the proposed use pattern of HeiQ AGS-20 in treated textiles, the AGS-20 nanocomposite and nanosilver particles in singular or aggregate forms may enter the environment through washing or disposal processes. In addition, silver ions will be released from AGS-20 treated textiles or from particles that br eak away from the treated textiles. Wastewater from washing cycles may contaminate streams and surface waters causing adverse effect to aquatic species. Solid waste (biosolids) remove d during wastewater treatment may be deposited on agricultural fields where it c ould adversely impact terrestrial an imals and plants. When treated articles like fabric are disposed of by land filling or incinera tion, the nanocomposite, nanosilver, or an aggregate of nanosilver may contaminate th e environment. Therefore, the characterization and identification of the nanocomposite, nanosilver , and its aggregates in each environmental media are critical factors in assessing the envi ronmental fate and transport of HeiQ AGS-20.

This section discusses the envi ronmental fate of silver na noparticles then discusses the environmental hazards posed by silver and nanosilver particles and conclude s with a quantitative assessment of the risks. As part of these discussions, additional da ta that EPA proposes to require in order to reduce its uncertainty with respec t to assessing the risk s of HeiQ AGS-20 are

identified.

A. Environmental Fate and Transport

HeiQ has not conducted any studies to characte rize the environmental fate of HeiQ AGS-20. However, HeiQ has submitted a number of literature studies to address the issues outlined above. The Agency has reviewed these studies and has noted a number of limitations with respect to their applicability to support a complete assessment of the risk of AGS-20 to the environment.

a. “Nanoparticle Silver Released into Wa ter From Commercially Available Sock Fabrics”, Benn and Westerhoff, 2008. This study attempts to characterize the effects washing can have on various types of nanosilv er-treated textiles like socks. The work was conducted on pre-leached and post- leached socks. Silver was observed to be

released from the treated items. The st udy conditions, however, do not reflect typical washing scenarios and it is possible that sa mple preparation methods may have impacted the integrity of the study. Moreover, the analytical met hods may not have of been adequate to identify all silver forms released from the treated items.Page 21 of 36 b. “Nitrification Inhibition by Silver Nanoparticles”, Choi and Hu, 2009. This study

monitors the effect that 15 nm nanosilver particles have on the ni trification process-- oxidation of ammonia by certain bacteria in so ils. Inhibitory eff ects of the evaluated nanosilver on nitrification proce ss were greater than that obse rved for colloidal silver and silver ions. This study indicates that nanosilv er in the size range of 15 nm is toxic to some microbes. However, it is unclear if the 15 nm nanosilver par ticles used in the study represent the size of the nanosilver particles pres ent in HeiQ AGS-20.

c. “The speciation of Silver Nanoparticles in Antimicrobials Fabric Before and after Exposure to a Hypochlorite/detergent Solution”, Impellitteri et al., 2009. The study monitors one type of sock treated with a nanosilver-based product in the presence of

bleach and sodium chloride. The treated items are shown to be stable in the presence of sodium chloride; however, in the presence of bleach elemental silver was shown to

oxidize and convert into insoluble silver chlo ride. This result suppor ts the likelihood that nanosilver products will end up in waste wate r streams or surface water as insoluble silver chloride when exposed to oxidizing agents.

d. “Behavior of Silver Nano-Textiles during Washing”, Geranio et al., 2009. In addition to the washing machine testi ng discussed in the residentia l exposure section, this study included dissolution studies on silver nanopa rticles (NPs) and immersion testing on AGS20 treated polyester textiles. The effect of pH, surfactants and oxidizing agents was

evaluated. The results for the dissolution studie s show that little disso lution of silver NPs occurs under conditions relevant to washing (pH 10) with dissolved concentrations 10 times lower than at pH 7. However, bleaching agents such as hydrogen peroxide or peracetic acid (PAA - formed by the perbor ate/TAED bleach system) can greatly

accelerate the dissolution of silver NPs. Th e results of the textile immersion testing indicated that for the polyeste r with incorporated AGS-20, the silver ion concentration in

the immersion water remained below the lim it of detection of 10.7 ug/liter during the entire experiment, and for the polyester tr eated with AGS-20 in a surface coating the silver ion concentration rose to 107 ug/liter du ring the first 50 minutes and continued on a slight rise until the end of the experiment at 240 minutes. The rate of increase was not affected by the addition of 0.1 M PAA and was only slightly affected by the addition of 0.5M PAA. The analysis of the immersion water with and without filtration indicated

that most of the silver released from the su rface treated fabric was ionic while most of the silver from the incorporated fabric was particulate >450nm.

The Agency concludes that these studies are re levant and help with understanding the behavior of silver nanoparticle s and silver nanopartic le-treated items under various environmental conditions. However, the studies do not necessarily represent He iQ AGS-20 and are insufficient to fully characterize the environmental fate and transport for HeiQ AGS-20 and the silver moieties (nanoparticles, silver ions, nanocom posites, etc.) resulting from HeiQ AGS-20.

To this end, the Agency is requiring additional en vironmental fate and transport studies. This requirement is based on a tiered approach as shown in Figure 1. This tiered approach consists of two tiers (Tier 1 and Tier 2) and is based on a co nceptual exposure model that takes into accountPage 22 of 36 the proposed use of HeiQ AGS-20. Tier 1 st udies will give preliminary data on potential environmental exposures. These studies must be conducted with HeiQ AGS-20. Tier 2 studies will provide quantitative data that can be used in the risk assessment conducted by the EPA for HeiQ AGS-20. These studies must be conducte d on HeiQ AGS-20 as well as all materials

released from it as determined fr om Tier 1 studies. Therefore, if needed, Tier 2 studies should be conducted following Tier 1 studies. The Agency is requiring a Tier 1 diss olution study for HeiQ AGS-20 and based on the results of this study, additional Tier 2 studies including a Fish Bioconcentration Factor (BCF) test and Adsorpti on/Desorption (batch Equilibrium) test may be required. Data justifications are provided in Appendix A.

Figure 1 - Test Material and Tiered Approach for HeiQ AGS-20Page 23 of 36 B. Environmental Toxicity

EPA has considerable data on the environmental hazards posed by the release of silver ions from conventional silver-based products. For environmen tal concerns, the ecological fate database is defined; ecological toxicity data indicate that silver is toxic to fish, aquatic invertebrates and estuarine organisms. Because of use patterns for silver, however, EPA does not expect

unreasonable adverse effects to the environment fr om the uses. Silver is currently being reevaluated through the Agency’s Re gistration Review program.

In contrast, the information available to assess th e potential hazards of nanosilver to non-target

wildlife is limited. The information below summar izes some of the available ecotoxicity data relating to nanosilver f ound in the literature.

a. “Toxicity of Silver Nanoparticles to Chlamydomanas reinhardtii”, Navaro et al., 2008. This short-term toxicity study measured photosynthesis of Chlamydomonas reinhardtii , a freshwater algae, in a suspension of carbona te coated silver na noparticles (Ag NPs; approximately 10-200 nm with a mean partic le size around 25 nm) and silver nitrate (AgNO 3 ). The toxicity (EC 50 ) was higher for AgNO 3 compared to AgNPs, based on total

silver concentration. When the EC 50 were calculated based on the concentration of the silver ion (Ag + ) present in each solution before inc ubation, the toxicity of the AgNPs was higher than AgNO 3 . The authors also showed that the amount of bioavailable Ag + is directly related to its observed toxicity; supporting the free ion activity model for Ag +
toxicity. However, the observed increase in toxicity of AgNP (EC 50 based on Ag +
concentration) to Chlamydomonas reinhardtii cannot be explained solely by the concentration of Ag + present in each solution before incubation.

b. “In Vivo Imaging of Transport and Biocompa tibility of Single Silver Nanoparticles in Early Development of Zebrafish Embryos” Lee et al., 2007. This study shows that citrate coated silver nanoparticles (AgNPs; 5– 46 nm) were transported into and out of Zebrafish embryos. Individual AgNPs we re observed inside embryos at every developmental stage in normally develope d, deformed, and dead zebrafish. AgNPs toxicity was shown to be dose-dependent w ith a critical co ncentration of 0.19 nM (0.021 ug/liter). At lower AgNP concentrations of 0.08 nM (0.0086 ug/liter) only a few dead and deformed zebrafish were observed; however, as AgNP concentrations increased, the frequency of deformed and dead zebrafish increased. This study indicates that the incidence of abnormalities in the zebrafish likely depends on the rates of passive diffusion of the silver nanopartic les, as well as accumulation of silver nanoparticles in the embryos.

c. “Toxicity of Silver Nanoparticles in Zebrafish Models” Asharani et al, 2000. Zebrafish embryos were incubated with Bovine Seru m Albumin “BSA” or potato starch coated silver nanoparticles (AgNPs) to determine dele terious effects and dist ribution patterns. A dose-depended increase in mortality and hatching delay was observed for embryos incubated with AgNPs. Defects observed included bent and tw isted notochord, in

addition to accumulation of blood in the bl ood vessels near the tail, low heart rate, pericardial edema, and degeneration of body parts. Silver nanoparticles were foundPage 24 of 36 distributed in the brain, heart, yolk and blood of embryos. No developmental defects were observed for embryos incubated with silver ion (Ag + ) or the capping agents but the concentrations used are not directly comparab le to the AgNP concentrations used in the study.

d. “The Effects of Silver Nanoparticles on Fathead Minnow” Laban et al., 2010. This study shows that commercial silver na noparticles (AgNPs; 20– 280 nm) are rapidly taken-up by fathead minnow during incubation. Exposure of fathead minnow embryos to AgNPs induced a variety of developmental abnormalities. Defects include absence of air sac, pericardial and yolk sac edema, hemorrh ages, and lordosis. The study also reports that sonication increases the relative toxicity of AgNPs 10-fold. The authors attribute this

increase to an increase in the bioavailabil ity of the AgNPs and not the amount of silver released from the AgNPs. Based on the c oncentration of dissol ved silver (silver

concentration determined after filtration through a 0.02 μ m membrane), the authors demonstrate that silver nitrate (AgNO 3 ) is more toxic than the AgNPs.

It is difficult to compare these studies because of the differences in the experimental conditions (e.g. water or growth media chemistry, nanopartic le size and capping agent variability, and taxa)

and the differences and uncertainti es in the methods that were used to quantify the dosages. In general, these studies indicate th at exposure to silver nanopartic les can result in developmental defects and bioaccumulation of silver nanoparticles in aquatic organisms. It is also difficult to pinpoint the toxicity threshold for nanoparticles in comparison to ionic silver. Although limited ecotoxicity data are available for nanosilver, the ex isting data seem to indicate that its effects are

different and/or more severe than for silver . Given this preliminary indication, the Agency requires additional data to assess the hazards resu lting to fish, aquatic invertebrates, birds and algae resulting from exposures to AGS-20 nanosilver.

C. Environmental Exposure Assessment

Screening Level Assessment Us ing the Down-the-Drain Model

The concentration of ionic silverin surface water resulting from the proposed use of HeiQ AGS20 was calculated using the Down the Drain Mo dule of the Exposure and Fate Assessment Screening Tool (E-FAST model, version 2) . The following input values were used:

• Production Volume: 4500 kg/year. The valu e was derived assuming that 300 million people (U.S. population) purchase one t-shirt treated with HeiQ AGS-20 each year. Each t-shirt weighs 150 grams and cont ains 100 ppm by weight silver.

• Release Days: 356 days per year. This assume s that each t-shirt treated with HeiQ AGS20 releases all its silver as silver ions over the course of one year.

• Waste Water Removal: 88%. This value is based on a study of Publicly Owned Treatment Works (POTWs) published by the US EPA Office of Water (US EPA, 2003).

• Stream Dilution Factor: 1.0X or 20.1X. Thes e values are the 10t h and 50th percentile

values for the dilution that o ccurs during one day of lowest stream flow over a ten year period (1Q10).Page 25 of 36 • The toxicity value: An acute LC 50 of 0.19 ug/L for Daphia magna (MRID 000545-96). This value is protective of other organisms which are less sensitive to silver.

• Level of Concern for the RQ: The level of concern (LOC) is 0.05 for listed (i.e. endangered or threatened) aquatic orga nisms and 0.5 for non-listed organisms.

The down-the-drain modeling results are shown in Table 7 and these results were divided by the LC50 of 0.19 ug/L for Daphia magna to obtain risk quotients (RQs). The RQ of 0.068 at the

worst case stream dilution factor of 1.0X indicates a risk concer n for listed species; however, it does not indicate a risk of concern for non-listed sp ecies. It is important to note that the stream

dilution factor of 1.0X means that the stream is fed entirely by the waste water treatment plant

and has no other source of water. The RQ of 0.0034 at the 50 th percentile stream dilution factor

of 20.1X is not of concern for eith er listed or non- listed species.

Table 7 - Down-the-Drain Estimates of Silver from AGS-20 in Surface Water.

Waste Water

Treatment

Removal A Stream

Dilution Factor Surface Water Silver Concentration (ug/liter) Risk Quotient D RQ Exceeds LOC?*

1.0X B 0.013 0.068 Yes, for listed species 88% 20.1X C 0.00065 0.0034 No

* The LOC is 0.05 (listed species) a nd 0.5(non-listed species). RQs th at exceed the LOC are of concern

Notes for Table 7 A. Silver removed from wastewater during treatment be fore discharge to a water body (e.g. lake, river etc.). B. 10% Percentile dilution factor for 1Q10 stream flow. C. 50% Percentile dilution factor for 1Q10 stream flow. D. RQ = Surface Water Concentration / LC 50 for Daphnia magna (0.19 ug/liter)

Refined Assessment Using the Biotic Ligand Model

The Biotic Ligand Model (BLM) Version 2.2.3 (Hydoqua l Inc., 2007) was used to refine the risk quotients obtained from the down the drain screen ing level assessment. This model uses water chemistry data from 811 surface water sites in the United States to calculate RQs that incorporate the biotic ligand into a chemical equilibri um framework that includes aqueous metal complexation, the relation between free metal io n concentrations and toxicity. The following values were used as inputs:

• Silver surface water concentra tion: 0.013 ug/liter. This is th e highest concentration that

was obtained from the down the drain model.

• Nitrate Concentration: 10 mg/liter. This is the National Recommended Water Quality Criteria for Non-Priority Pollutants (EPA, 2009a ). A value of 45 mg/liter was also tested in the model and did change the resultant RQs.

• Dapnia magna was selected as the target organism because it is the most sensitive species.

The RQs obtained from the BLM are shown in Table 8. The RQs range from 0.0024 to 0.021 depending upon which of the 811 sites is used by the BLM in the calculat ions. All of these RQs

are less than 0.05 which means that the risk s are not of concern for listed species.Page 26 of 36 Table 8 – Summary of BLM RQs for S ilver from AGS-20 in Surface Water.

Silver Surface

Water

Concentration Nitrate

Concentration Minimum RQ* Average RQ* Maximum RQ* RQ Exceeds

LOC?*

0.013 ug/liter 10 mg/liter 0.0024 0.012 0.021 No

* The LOC is 0.05 (listed species) a nd 0.5(non-listed species). RQs th at exceed the LOC are of concern

Neither the Down the Drain Mode l or the BLM can distinguish be tween silver nanoparticles or silver ions and for the purposes of this assessment, it is assumed that all of the silver is released from AGS-20 as ionic silver. In addition, this assessment only considers silver that could be released by AGS-20 and does not include the ot her known sources of silver which will all

contribute to the environmental load ing of silver. Lastly, it is impor tant to note that the toxicity

values are based on ionic silver a nd it is not known if th ey are applicable to silver nanoparticles.

VI. PROPOSED REGULATORY ACTION

A. Legal Framework

The Federal Insecticide, Fungi cide, and Rodenticide Act (FIF RA) section 3(c)(7)(C) provides

that:

“The Administrator may conditiona lly register a pesticide contai ning an active ingredient not contained in any currently re gistered pesticide for a peri od reasonably sufficient for the generation and submission of required data (whi ch are lacking because a period reasonably sufficient for generation of the data has not el apsed since the Administ rator first imposed the data requirement) on the condition that by the en d of such period the Administrator receives

such data and the data do not meet or exceed ri sk criteria enumerated in regulations issued under this Act, and on such other conditions as the Administrator may prescribe. A conditional registration under this subparagraph shall be granted only if the Administrator determines that use of the pesticide duri ng such period will not cause any unreasonable

adverse effect on the environment and that use of the pesticide is in the public interest.”

Therefore the statute requires EPA to make three findings in order to grant a conditional registration for a pesticide product containing a new active ingredient:

1. Insufficient time has elapsed for the generation of data since the requirement for that data was imposed;

2. Use of the pesticide is in the public interest; and

3. Use of the pesticide during the period that th e newly required data is being developed and reviewed by the Agency will not cau se unreasonable adverse effects.Page 27 of 36 B. Findings and Proposed Decision

The Agency is proposing to issue a conditional, four year time limited registration for HeiQ AGS-20 in accordance with FIFRA section 3( c)(7)(C) and to require as a condition of registration that the company provide data from a number of studies that will allow the Agency to more confidently characterize the potential risks associated w ith this material. Specific data requirements are outlined in Appendix A. Although th ese data requirements ar e specific to HeiQ AGS-20, they form a starting point for identifying the types of data the Agency will require for other nanomaterial-based products on a case-by-case basis.

In order to mitigate occupational risks, label language is required to reduce potential exposures. Specific label revisions to the labeling as currently proposed will include the following:

1. The application rates must be limited to 100 ppm silver by weight.

2. Engineering controls such as closed system load ing or local exhaust ventilation must be used when handling AGS-20 powder.

3. The following PPE must be worn when handling AGS-20 powder.

• Gloves that are chemically resistant to all of the components of the textile fiber master batch or coating formulations to which the AGS-20 powder is added.

• Coveralls that cover the arms, legs and torso.

• NIOSH Certified Full face resp irators with P100 or equi valent filter cartridges.

C. Basis for Conditional Registration

The Agency’s basis for the conditional registration is as follows:

1. Data Generation

HeiQ submitted its registration application th inking its product was similar to currently registered silver-based antimicrobial pesticid e products and provided all of the required productspecific data for this type of application. EP A typically does not require generation of any additional generic data to support such an applic ation; applicants fulfill the applicable generic data requirements by citing previously submitted data. Over a year after HeiQ submitted its application the Agency made a final determinat ion that HeiQ’s product contained nanosilver, and

that EPA would treat nanosilver as a new activ e ingredient. Throughout the application review process, EPA and HeiQ have discussed what data might address the Agency’s concerns. Although HeiQ has attempted to provide EPA with da ta in an effort to address Agency questions and concerns, until very recently, EPA had not reach ed a position with regard to which types of data would be required. This was due in larg e part to the need to understand and apply the advice provided in the report from the consultation with the FIFRA Scientific Advisory Panel. As a result, EPA has determined that insufficien t time has elapsed from the point at which EPA determined and informed HeiQ of the data requi rements needed to assess HeiQ’s application for HeiQ to have generated the data.Page 28 of 36 EPA conducted a screening level ri sk assessment of risks to human health and the environment

associated with the use of AGS-20 as a material s preservative. However, the majority of the studies used for the screening level risk assessm ent were not conducted with AGS-20; rather the studies were conducted using other silver nanomat erials. As indicated in the 2010 SAP report, nanomaterials such as HeiQ AGS-20 are physi cally and chemically different from single

molecules or bulk materials of the same substa nce, and these differences may impart unique properties that ultimately affect the potential ri sks of the nanoscale material. Therefore, EPA determined that more extensive product chemistr y, toxicology, exposure, and environmental data are necessary to confirm that the screening level risk assessments for the AGS-20 nanosilversilica composite provide an accurate assessment of the risks. Because this list of data requirements are only being finalized with today’ s action, HeiQ has not yet been able to conduct these studies. Therefore, the Agency is proposi ng to require these stud ies as a condition of registration, allowing sufficient time for the stud ies to be conducted and for the Agency to review them. Ultimately, the Ag ency will use these data to dete rmine whether the ingredient can be registered under FIFRA section 3(c)(5).

As discussed above, a listing of th e studies that are needed for the registration of HeiQ AGS-20 under FIFRA section 3(c)(5), are in Appendix A.

2. Public Interest

As required under FIFRA section 3(c)(7)(C), the Agency has de termined that the benefits expected from this proposed pest icide product, together with ot her considerati ons, justify the conclusion that granting a conditi onal registration is in the pu blic interest. HeiQ provided information to the Agency regarding the ec onomic benefits offered by its technology when compared to other options. EPA agrees that this information on conservation of the environment

and consumer benefits contribute s to the public interest. Moreove r, considerations relating to market equity and international trade and promoting innovation lend further support to a determination that a conditional registration is in the public interest. These latter two points are

discussed in more detail below:

Conservation of the Environment

EPA has already registered a number of silver-bas ed antimicrobial products for use as materials preservatives. All antimicrobial silver-based pesticide products act vi a the release of a low

concentration of silver ions that then interact with bacteria. Commonly, regardless of the silver additive type, upon contact with moisture, silver ions are released fr om the additive, and subsequently from the object treated with the ad ditive. The antimicrobial potency of a silver additive is therefore directly related to the potential for releasing silver ions. The release potential differs between various s ilver materials. As the size of silver particles decreases (from micro-size silver to nano-size silver), the potential for releasing silver ions increases, due to the increasing unit of surface area (i.e ., availability of ions for rel ease) per unit mass of silver.

Specifically, most antimicrobial silver-based pes ticide products currently contain a silver salt, [e.g., AgCl or AgNO 3 ]. Compared to the amount of silver in HeiQ’s product, most currentlyPage 29 of 36 registered silver-based materials preservatives require larger amounts of silver to be added to an article in order to provide a sufficient lifetime of activity for antimicrobial treatment. Therefore the overall potential environmental exposure to silv er resulting from the lower-volume use of the HeiQ product should be smaller than from a compar able use of currently re gistered silver-based pesticides.

Consumer Benefits

A nanosilver materials preservative should maintain its efficacy longer th an other silver active ingredients due to an expected gr adual and controlled release of silver from the nano material as opposed to the rapid release of for example, silv er from the zeolite structure and the immediate

dissolution of the silver salt. While other s ilver active ingredients are more effective in applications where high silver concentrations are required imme diately, their effects are only short-lived. In contrast, the Agency believes th at AGS-20 will allow slow and controlled release of silver ions, resulting in mo re prolonged antimicrobial activity as described in the Efficacy Review posted in the docket for today’s acti on (EPA, 2009). Data provided to the Agency by HeiQ supports this theory and will be confirme d by the required leaching study. Thus, consumers purchasing textile treated with nanosilver should receive a durabl e antimicrobial protection with

less silver than other conventi onal silvers on the market.

Market Equity and International Trade

The regulatory framework established in FI FRA generally reflects a policy of providing equitable or fair consideration of pesticides that are similarly-situated, so that competing products are given the same opportunity to enter the marketplace. Evidence that FIFRA intends to protect similar interests equally includes the Section 3(c)(5) bar on denying a pesticide registration because it is not essen tial. Further, the authority to conditionally register pesticides was added to FIFRA in 1978 to address percei ved unequal treatment in registering similar pesticides. See S. Rep. No. 95-334 (1977). Previous to the addition of this authority there was a “needlessly anti-competitive” “double standard” where “some producers enjoy[ed] an advantage over those who wish[ed] to enter the market by virt ue of the date of their registrations.” Id. Moreover, due to improvements in scientific knowledge which over time lead to new data requirements, an anomaly exists such that the Agency “often knows more about the safety of a new pesticide which is excluded from registration and use until all data are in than [the Agency] does for a previously registered pe sticide that stays on the market.” Id. By adding the authority to register a new pesticide while imposing cond itions to deal with mi ssing data, Congress sought

to eliminate these barriers and intended to leve l the playing field between already registered and new pesticides. See id.

Specifically, Section 3(c)(7)(A) of FIFRA provide s that the Agency may conditionally register pesticides that are identical or substantially simila r to currently registered pesticides or pesticides that differ only in ways that w ould not significantly increase th e risk of unreasonable adverse

effects on the environment.

Recently, some current registrants of silver-based antimicrobial pr oducts have reported that four of their products contain nanoscale silver material . On the surface, these products appear to bePage 30 of 36 somewhat similar in composition and use to th e product HeiQ is proposing, although not similar enough to warrant considering the HeiQ appl ication under the FIFRA section 3(c)((7)(A) standard. At least one of the currently register ed products identified as containing nanosilver has been on the market for many years. While EPA approved these registrations without knowledge that these products contained nanoscale silver and without specifically assessing any potential risks that might be associated with the specifi c nanoparticles contained in those products, they are on the market. Arguably, this unfairly disadvantages HeiQ because HeiQ’s competitors are in

the market ahead of them.

While the HeiQ nanosilver is a new active ingred ient and thus does not meet the criteria of Section 3(c)(7)(A), it is similarly situated beca use it is a nanosilver active ingredient, and EPA now has information indicating that there are so me nanosilver products al ready registered. None of the nanosilver products have submitted the types of data that the Agency is proposing to require as a condition for the re gistration of HeiQ’s nanosilver product. The Agency believes

that in connection with this ne wly required data, treati ng all of the registered nanosilver products equally is in the public interest. In light of this, and in conc ert with the Agency’s request to HeiQ for additional data as part of the proposed conditional registration, the Agency intends to require that similar data be developed to support the continue d registration of these other nanoscale products as well.

Finally, the AGS-20 nanosilver com posite product has been approve d for sale in Switzerland and is produced by HeiQ, a Swiss based company. EPA does not want to create an unintentional disadvantage for any registrant. EPA attempts to provide fairness in its regulatory activities so that companies making comparable products r eceive equal treatment and have the same opportunities to market their products.

Innovation

EPA sees the emergence of nanotechnology as o ffering potential benefits for society in many different fields, including pest control products. The use of nanotechnol ogy in pesticide products may allow for more effective targeting of pests a nd use of smaller quantities of pesticide. These could contribute to improved human and environm ental safety and could lower pest control costs. Therefore EPA seeks to encourage i nnovative work to real ize these benefits.

Developing new technologies can, however, be expensive and unpredictable, and meeting regulatory requirements can add costs and delay the marketin g of new products. Overly burdensome regulatory requirement s and / or unnecessarily lengt hy regulatory reviews could discourage technology providers from pursuing the development of beneficial new applications of nanotechnology in the field of pesticides. Consequently, EPA thinks it is appropriate to consider the potential impacts of its regulatory process and decisi ons on the incentive to pesticide developers to pursue the app lication of nanotechnology.

In the case of the HeiQ application, EPA’s c onditional registration of HeiQ’s product and the imposition of additional data requirements is reasona ble in that it will ensure EPA receives the

information it needs to confirm its screening level assessment of the safety of AGS-20, whilePage 31 of 36 allowing an innovative product to reach the mark et already being served by competitors in a reasonable period of time.

3. No Unreasonable Adverse Effects

As noted in section III, EPA lacks information to conduct a complete a ssessment of the potential

risks to human health and the environment associated with the use of AGS-20 as a materials preservative. Based on available informa tion, however, EPA conducted a screening level

assessment of risks to human health and the envi ronment associated with the use of AGS-20 as a materials preservative. As a resu lt, EPA believes that the likely risks from the use of AGS-20 during the period of the proposed conditional regi stration appear to be small and the overall exposure from such use is lower than exposures resulting from existing uses of nanosilver. At the same time, the use of AGS-20 would provide benefits both from the decreased release of silver to the environment and from likely exte nded antimicrobial efficacy compared to other silver-based antimicrobial products, as described in the Public Interest section above. After weighing the potential risks and benefits, EPA concludes that the propos ed registration would

not cause unreasonable adverse effects on the environment during the period of the proposed conditional registration. This conclu sion is based on the following findings:

Risks to Human Health

The risk to humans depends on both the toxicity of a substance and th e level and duration of exposure to a substance. As discussed above, human s could be exposed to silver ions, to AGS-20 (the nanosilver-silica com posite), and to nanosilver particles. With respect to silver ions, the Agency notes that safe exposure levels for silv er have been establishe d by several regulatory agencies including FDA, OSHA, and EPA based on the common endpoint argyria and using the same human studies. Argyria is a blue-gray discoloration of the sk in and is not considered as being of toxicological concern. The Agency has completed a risk assessment (EPA, 1993) of

conventional silver and silver salt s, and the risks are acceptable. In light of the estimates of low dermal and oral risks, findings from the textil e leaching studies, as well as the absence of any incidents of argyria associated with the use of currently marketed nanosilver consumer products, the Agency concludes that human exposure to si lver ions from the use of AGS-20 and textiles treated with AGS-20 is not likely to a pproach a level that causes argyria.

With respect to AGS-20, the acute toxicity of AGS-20 nanosilver silica composite is low by all routes of exposure and the textile leaching studie s indicate that exposure to AGS-20 will be low.

Because there are no subchronic or chronic oral or dermal toxicity studies available for AGS-20 or on the nanoparticles that might break away from textiles treated with AGS-20, these exposures were estimated using a study on silver na noparticles that is report ed in the literature (Kim et al., 2007) as a surrogate. The risk calcul ations indicate that th e MOEs for dermal and incidental oral exposures exceed the target MO E of 1000 by at least an order of magnitude. The Agency recognizes the multiple uncertainties regarding these estimates. Even so, the Agency considers it unlikely that either AGS-20 or na nosilver particles that break away from AGS-20 would exhibit significantl y greater toxicity compared to the nanosilver materials for which data are available, such that they would pose a risk of concern during the period of conditional registration. Whatever occupational risks may exist will be significantly reduced through thePage 32 of 36 required use of engineering c ontrols and PPE. Thus, EPA c oncludes that neither acute occupational nor acute residential exposure woul d pose a risk of concern during the period of conditional registration.

With respect to nanosilver and nanoscale particle s of the silver-silica composite, EPA lacks data to assess completely either the toxicity or the human exposure to such materials. The available leaching data indicate, however, that release of these substances is generally limited. To the extent that these materials might be released during the acute t oxicity studies with AGS-20, they did not cause significant acute toxic effects.

Apart from the studies specifically considered in the screening level assessment of the HeiQ product, EPA has considered information on othe r nanosilver products. The Agency expects that the overall exposure from use of AGS-20 woul d be lower than the corresponding exposures resulting from existing uses of nanosilver. The Project on Emerging Nanotechnologies (PEN) has compiled an inventory of all consumer produc ts reported by their ma nufacturers to contain

nanomaterials. Based on the information PE N has compiled, nanosilver is the most common nanoscale material currently used in consumer products. Approximately a quarter of the identified consumer nanomaterial products are claimed to contain nanosilver. Nanosilver products are sold in the U.S. and in foreign mark ets such as Asia, Europe , Australia, and South

America. As discussed in the Market Equity and Internationa l Trade section, while EPA cannot confirm the assertion, four regist ered antimicrobial products have recently been identified by the registrants as containing nanosilv er. In addition, FDA has appr oved the marketing of several medical devices containing nanosilver. Specifica lly, FDA has authorized the sale of bandages

treated with nanosilver to contro l post-operative infections in surgical wounds and the sale of catheters and other devices, wh ich may come in contact with human tissues, treated with nanosilver to control the growth of microorganisms.

While EPA does not have data to quantify relativ e exposure to nanosilver from these products vs. HeiQ’s product, EPA thinks it is reasonable to expect that nanosilv er exposure resulting from the multiple consumer products and FDA-approved uses is likely to be considerably higher than the residential exposure that would result fr om wearing treated te xtile products.

In sum, the current presence of nanosilver products on the market, potentially including registered antimicrobial products, allows the Agen cy to anticipate that conditionally registering the HeiQ product will not add sufficient increm ental human exposure to nanosilver that would cause significant adverse effects while the re quired additional data are being generated.

Risks to the Environment

With regard to environmental exposure, the Agency does not have enough information to determine whether exposure from the HeiQ product is to silver ions, silver nanoparticles, the silica-silver composite or any combination of the three, and to what levels the environment is exposed. The ecological fate database for silver is defined; silver is toxic to fish, aquatic invertebrates and estuarine organisms. Because of use patterns for registered silver products, however, EPA does not expect unreasonable advers e effects to the environment from silver based antimicrobial products.Page 33 of 36 EPA estimates, derived from down-the-drain modeli ng, of the concentrations of silver resulting from the proposed use of HeiQ AGS-20, do not ex ceed the Agency’s estimate of the highest

concentration of ionic silver in surface water to which an aquatic community can be exposed briefly without resulting in an unacceptable effect. These estimat es do not take into account any potential higher toxicity of silv er nanoparticles than ionic silver. Nonetheless, the available leaching data indicate that environmental rel ease will be generally limited, and would involve relatively smaller masses of silver than other si lver-based antimicrobial products. Moreover, the Agency expects that the overall environmenta l exposure from use of AGS-20 would be lower than the corresponding exposures resulting from existing uses of nanosilver. For the same reasons as discussed above, EPA expects condit ionally registering the HeiQ product will not add sufficient incremental environmental exposure to nanosilver that would cau se significant adverse environmental effects while the required additional data are being generated.Page 34 of 36 References:

AIHA, 2006. A Strategy for Assessing a nd Managing Occupational Exposures, 3 rd Edition

. Ignacio, J.S.. and Bullock, W.H., American Industrial Hygiene Association, 2006. (see page 270).

AISE/CEFIC, 2005. Human & Environmental Risk Assessment (HERA) on Ingredients of Household Cleaning Products, Guidance Document Methodology, Internati onal Association for Soaps, Detergents and Maintenance Products (AISE) and the European Chemical Industry Council (CEFIC), February 2005.

Asharani P.V. et al., 2008. Toxicity of Silver Nanoparticles in Zebrafish Models. Nanotechnology 19(255102): 8pp.

Benn, Troy M. and Westerhoff, Paul, 2008. Nanoparticle Silver Released into Water from Commercially Available Sock Fabrics. Environmental Science and Technology. 42:4133-9. (MRID 477575-01).

Burgess, W., 1995, Recognition of Health Hazards in Industry, 2 nd Edition , Published by John Wiley and Sons, Inc., 1995 (see page 289)

Choi, O. K. and Hu, Z. Q. 2009. Nitrification In hibition by Silver Nanopart icles. Water Science Technology-WST59.9/2009. (MRID 477953-01)

Demou, E., Peter, P. and Hellweg, S. 2008. Exposur e to Manufactured Nanostructured Particles

in an Industrial Pilot Plant. Annuals Occupational Hygiene, 52:695-06, published online 17 October 2008. (MRID 477575-02)

Egger et al., 2009. Antimicrobial Pr operties of a Novel Silver-S ilica Nanocomposite Material. Applied and Environmental Microbiology, May 2009, p. 2973-2976.

Geranio et al., 2009. The Behavior of Silver Nanoparticles During Washing. Environmental Science and Technology. Published online September 24, 2009.

HydroQual Inc., 2007. The Biotic Lingand Model Windows Interface, Version 2.2.3 Users

Guide and Reference Manual , HydroQual Inc., June 2007

Impellitteri, CA, Tolaymat, TM and Sch eckel, KG. 2009. The Speciation of Silver Nanoparticles in Antimicrobials Fabric Before and After Exposure to a Hypochlorite/Detergent Solution. J. Environ Qual 38: 1528-30. (MRID 477953-02)

Kim, YS, Kim, JS and Cho, HS, et al., 2007. Twen ty-Eight-Day Oral Toxicity, Genotoxicity, and

Gender-Related Tissue Distribution of Silver Nanoparticles in Sprague -Dawley Rats. Korea Environment and Merchandise Testing Inst itute, Songdo-dong, Yeonsu-gu, Incheon, Korea. Inhalation Toxicology. 10:575-83.Page 35 of 36 Laban, L., Nies, R., and Turco, J ., et al., 2010. The Effects of S ilver Nanoparticles on Fathead Minnow (Pimephales promelas) Embryos). Ecotoxicology. 19:185-95.

Larese, F.F., D'Agostin, F., Crosera, M., et al., 2009. Human Skin Penetration of Silver Nanoparticles through Intact a nd Damaged Skin. Toxicology. 255(1-2):33-7. Epub 2008 Oct 9.

Lee, K.J., Nallathamby P.D., and Browning, L.M. 2007. In Vivo Imaging of Transport and Biocompatibility of Single Silver Nanoparticles in Early Development of Zebrafish Embryos. ACS NANO, Published online September 28, 2007.

Marquart et al., 2008. ‘Stoffenmanager’, a Web Based Control Banding Tool Using an Exposure Process Model. Annuals of Occupational Hygiene, Volume 52, No. 6 pp. 429-441, 27 June 2008

Matthews, GA. 2000. Pesticid e Application Methods 3 rd Edition , Blackwell Science, 2000

Navarro, E. et al., 2008. Toxicity of Silver Nanoparticles to Chlamydomonas reinhardtii. Environmental Science and Technology. 42:8959-64.

NIOSH, 2009 Approaches to Safe Nanotechnology: Managing the Safety and Health Concerns Associated with Engineered Nanomater ials. DHHS (NIOSH) Publication No. 2009-125

Sung, J.H., Ji, J.H. and Park, J. D., et al., 2009. Subchronic Inhalation Toxicity of Silver Nanoparticles. Toxicological Sciences. 108:452-61. (MRID 477575-05)

Trop et al., 2006. Silver-Coated Dr essing Acticoat Caused Rais ed Liver Enzymes and Argyrilike Symptoms in Burn Patient. J. Trauma, Injury, Infection and Critical Care. 60:648-52.

US EPA, 1993. RED Fact Sheet for Silver , EPA-738-F-93-005. EPA Office of Pesticide Programs, June 1993.

U.S. EPA. 1997. Exposure Factors Handbook. Vol. I- III. Office of Research and Development. Washington, D.C. EPA/600/P-95/002Fa.

U.S. EPA, 1998. PHED Surrogate Exposure Guid e, EPA Office of Pesticide Programs, August

1998

U.S. EPA, 2003. Development Document for the Final Effluent Limitations Guidelines and

Standards for the Metal Produc ts and Machinery Point S ource Category. EPA-821-B-03-001.

U.S. EPA. 2008. Child-Specific Exposure Factors Handbook. National Center for Environmental Assessment. Washingt on, DC; EPA/600/R-06/096F. September 2008.

U.S. EPA. 2009a. National Recommended Wate r Quality Criteria, Office of Water, 2009.Page 36 of 36 U.S. EPA, 2009b. Literature Review of EPA File Symbol 85249-R, HeiQ AGS-20, Memorandum from Tajah Blackburn to Ma rshall Swindell, DP 366658, July 14, 2009.

Viachou et al, The Safety of Nanocrystalline Si lver Dressings on Burns: A Study of Systemic Silver Absorption. Burns 33(8) pp 979-85, December 2007Appendix A - Page 1 of 13 

# Appendix A – Data Requirements for the Registration of HeiQ AGS-20 as a

Materials Preservative in TextilesAppendix A - Page 2 of 13 1.0 Introduction

HeiQ AGS-20 is a novel silver-silica nanocomposite material (Egger et al., 2009) that contains silver nanoparticles embedded in a matrix of amorphous silica which prevents agglomeration of the silver particles. This material will be formulated as a powder which will be used as a materials preservative to treat textiles by application as a surface coating or by incorporation into the starting materials (i.e. the masterbatch) prior to fiber manufacture. The human and environmental exposures that result from the use of AGS-20 and the use and disposal of textiles treated with AGS-20 will largely be a function of what materials leach or break away from the treated textile during use and disposal. The materials could include:

• Micron sized particles of the nanosilver - silica composite,

• Silver nanoparticles that break away from the composite and

• Silver ions released from the silver nanoparticles.

2.0 AGS-20 Product Use and Exposure Pathways

Manufacture - AGS-20 will be manufactured using a proprietary process where the starting materials are dissolved in a solvent and subjected to flame spray pyrolysis. The resulting material consists of silver nanoparticles embedded within a matrix of amorphous silica as shown in Figure 1. The silver nanoparticles have a typical diameter of 1 to 10 nm and the silver-silica composite material has an average diameter of approximately one micron (Egger et al., 2009). As is the case with all pesticides, the worker exposures that might occur during the manufacture of AGS-20 are outside the purview of FIFRA a nd fall under the jurisdiction of the Occupational Safety and Health Administration OSHA.

Application - It is proposed that AGS-20 will be formulated as a powder. This powder will be used to treat textiles either by application as a polymeric coating or by incorporation into the fiber during manufacture. Occupational handler exposures will occur during the addition of the AGS-20 powder at end user textile manufacturing and treatment facilities and during subsequent work activities involving the treated textile products. The highest exposures will occur as workers pour the powder into coating mix tanks for textile surface treatment or into blending hoppers for melt spinning textile fibers. Both dermal and inhalation exposures are anticipated. Therefore, personal protective equipment and/or engineering controls will be needed.

Consumer Use - The AGS-20 treated textiles will be used to manufacture consumer items such as clothing and it is intended that the AGS-20 treatme nt will provide a slow release of silver ions during the life of the treated item. These items will be purchased and used by consumers in a manner that is similar to untreated clothing with the exception that they may be washed less often due to the reduced growth of odor-causing bacteria. Exposures to humans and the environment can occur if the particles leach or break away from the treated textile. The particles could consist of either silver-silica composite particles, silver nanoparticles that break away from the composite or silver ions.Appendix A - Page 3 of 13 The following consumer exposure scenarios may be possible for AGS-20 based on the proposed

use pattern:

• Dermal and incidental oral exposures from wearing and mouthing treated clothing.

• Inhalation exposures during machine drying.

Exposures to environment can also occur if the particles are released into the washwater.

Figure 1 – Product Use Analysis for HeiQ AGS-20Appendix A - Page 4 of 13 3.0 Data Needed to Assess the Risks of Exposure to HeiQ AGS-20

The nanosilver in HeiQ AGS-20 is physically and chemically different than single molecules of elemental silver or bulk silver, and this difference may impart unique properties to the nanomaterials, affecting structure, function, and ultimately their environmental fate. For this reason, the data necessary for the Agency to make the required statutory safety finding for this product are provided in this document. The following factors were considered in the development of these data requirements:

• AGS-20 is a new active ingredient and is therefore subject to the data requirements for the registration of antimicrobial pesticides that are detailed in 40 CFR Part 161. These requirements include studies on physical and chemical characteristics, residue chemistry, environmental fate, toxicology, reentry protection, spray drift, wildlife and aquatic organisms, plant protection, non target insects and product performance.

• Although some studies, such as those dealing with physical and chemical characteristics are required for all use patterns, many of the data requirements are conditional based on the potential for exposure. Information provided by HeiQ and information from the literature was used to tailor the requirements to the proposed use pattern based upon the anticipated exposures.

• Additional studies in the area of physical and chemical characterization that are not

included in 40 CFR Part 161 are needed because AGS-20 contains nanosilver. These studies are needed because nanosized materials have unique and new characteristics which are not present in the bulk or conventional materials. These characteristics have been recognized in the FIFRA SAP Report (SAP Minutes No. 2010-01) and by the MINChar Initiative (ISO TC229 WG3/PG5 Project).

In addition, the following recommendations from the FIFRA SAP report were considered in the development of the data requirements for AGS-20:

• Both “nano-sized particles of silver (Ag)” as well as “ionic silver (Ag + )” contribute to toxic effects of silver nanoparticles. The rate of silver ion production, as well as the distribution of silver nanoparticles in tissues and the environment, may differ substantially between silver nanomaterials and other forms of silver, as nanomaterials can deliver ions directly to specific tissues, cell membranes or inside cells – places where other forms of silver compounds cannot reach. Therefore, the hazard profile of silver nanomaterials may differ from other forms of silver.

• Particle size substantially impact particles properties, such as rate and concentration of silver ion release, reactivity and catalytic efficiency, plasmon resonance, and quantum effects. Smaller sized-particles are more easily taken up by organisms and are distributed more widely. Other physicochemical properties, such as shape, surface area, surface charge or coating, are also likely to impact biological response and environmental fate.Appendix A - Page 5 of 13 • The Panel “disagreed that nanosilver applied to a substrate will permanently bind with the substrate”. It is “especially challenging to determine that there is no release of nanomaterials from a substrate”, under current state of science and available measurement standards. The Panel suggested that the Agency require tests that simulate realistic use of products and potential nanosilver release along with quantitative life cycle analysis and risk assessment.

A listing of the studies that are needed for the re gistration of HeiQ AGS-20 is included in Tables 1 and 2. The studies included in Table 1 are considered to be Tier 1 meaning that their need is

not based on the results of other studies. The studies listed in Table 2 are considered to be Tier 2 because they may or may not be required depending upon the results of the Tier 1 studies. In the Tier 1 studies the test material will be the AGS-20 composite, however, for some of the Tier 2 studies, the test material will be dependent upon the results of the Tier 1 leaching and dissolution studies. These studies may indicate that the silver nanoparticles, in addition to the AGS-20 composite, should be tested in the Tier 2 studies. A diagram that outlines this approach is included in Figure 2.

Figure 2 - Test Material and Tiered Approach for HeiQ AGS-20Appendix A - Page 6 of 13 4.0 Submission of Protocols

Because the guidelines are based on conventional chemicals used in industry and agriculture, they provide only general guidance on study design issues that might arise when testing nanomaterials. Due to the unique properties of nanomaterials, the Agency is requiring HeiQ to submit protocols for the majority of the studies as noted in this Appendix. Once approved, the protocols will be made available on the EPA website. Although these data requirements are specific to HeiQ AGS-20, they form a starting point for identifying the types of data the Agency will require for other nanomaterial-based products on a case-by-case basis.

5.0 References:

1. Mortensen LJ, Oberdörster G, Pentland AP, et al. (2008) In vivo skin penetration of quantum dot nanoparticles in the murine model: the effect of UVR. Nano Lett. 2008 Sep;8 (9):2779-87. Epub 2008 Aug 8.

2. Wu J, Liu W, Xue C, et al. (2009) Toxi city and penetration of TiO2 nanoparticles in hairless mice and porcine skin after subchronic dermal exposure. Toxicol Lett. 2009 Dec 1; 191(1):1-8. Epub 2009 Jun 6.

3. Larese FF, D'Agostin F, Crosera M, et al. (2009) Human skin penetration of silver nanoparticles through intact and damaged sk in. Toxicology. 2009 Jan 8;255(1-2):33-7. Epub

2008 Oct 9.

4. Geranio et al., 2009. The Behavior of Silver Nanoparticles during Washing. Environmental Science and Technology. Epub2009 Sept 24.Appendix A - Page 7 of 13 This Page Intentionally Left BlankAppendix A - Page 8 of 13Table 1 - Summary of Tier 1 Required Data for HeiQ AGS-20OSCPP Data Requirement (Note 1)Guideline Number: Study TitleStudyStatusData TriggerTestMaterialCommentsProduct Chemistry830.1550: Product Identity and Composition Incomplete AGS-20The topology of the nanocomposite needs to be fully described.Information is needed on how the silica and nanosilver are boundand on the distribution of nanosilver in the silica matrix (uniformor heterogeneous).830.1750: Certified Limits Incomplete AGS-20The submitted study only provides data on the average amount ofnanosilver present in AGS-20. Data are also needed on the upperand lower limits of nanosilver present in AGS-20.830.1800: Enforcement Method Incomplete AGS-20Submitted method is based on analysis of total silver. Needmethod to include high resolution images of the active.830.1900: Submittal of SamplesNo Data AGS-20830.6313: Stability to Normal and ElevatedTemperatures, and Metals/Metal IonsNo dataAGS-20Also need to test stability to sunlight, detergents, and salinity.The results of these tests will dictate the test substance for theTier 2 studies.830.6317: Storage Stability830.6320: Corrosion CharacteristicsNotAcceptableAGS-20Originally submitted studies were not acceptable because theyused an accelerated method. HeiQ is currently conducting oneyear studies.830.7050: UV-Visible Light Adsorption No data AGS-20UV-Vis data will be used to confirm that the material is in factnano-sized. It may also provide data on the range of particlesizes of the nanosilver particles in AGS-20.830.7520: Particle Size, Fiber Length, andDiameter (size) DistributionIncompleteThese studies are required forantimicrobial pesticidesper 40 CFR 161.AGS-20No Guideline: Surface Area DeterminationNo data AGS-20A literature study (Egger et al., 2009) provides some particle sizeand surface area data; however, it does not have the qualitycontrol information and documentation that is required for aproduct chemistry study.830.7840: SolubilityIncomplete 40 CFR 161 AGS-20Submitted data for pH 7. Need data for pH 5 and 9.Appendix A - Page 9 of 13Table 1 - Summary of Tier 1 Required Data for HeiQ AGS-20OSCPP Data Requirement (Note 1)Guideline Number: Study TitleStudyStatusData TriggerTestMaterialCommentsEnvironmental FateNo Guideline: Dissolution Kinetics Study No dataNeeded to assessenvironmental exposure fromwash water.AGS-20This is a fundamental study on the persistence of AGS-20 in theenvironment and the extent of silver ion or nanoparticle release.This test will dictate the test substance to be used for the Tier 2other studies.Environmental Effects850.1010: Acute Toxicity FreshwaterInvertebrates.850.1075: Freshwater Fish Toxicity850.2100: Avian Oral Toxicity850.5400: Algal Toxicity, Tier 2No dataNeeded to assessenvironmental exposure fromwash water. These studies arealso conditionally required per40 CFR 161.490.AGS-20The proposed use of AGS-20 may lead to surface watercontamination that could impact fish, invertebrates, avian, andalgal communities. These studies will address concernsregarding the potential risks of AGS-20 and will provide toxicityendpoints applicable to ecological risk assessment.Human Exposure875.1200 and 875.1400Applicator, Indoor ExposureSupplementalNeeded to assess occupationalhandler exposure duringpouring of powder.AGS-20This study will provide EPA with information useful forevaluating the form, route, and level of exposure experienced byworkers who handle AGS-20 in the process of treating textiles.The exposure data will be used in conjunction with thetoxicology endpoints to assess the risk of applicator exposure toAGS-20. This risk assessment will be used to determine whatmitigation measures will be needed to prevent adverse healthaffects in exposed workers.No Guideline: Textile Leaching StudySupplementalNeeded to assess human andenvironmental exposure.This test will determine whattest substance should be usedfor other studies.AGS-20TreatedTextilesA leaching study is needed to determine what, if any, materialsare released from AGS-20 treated textiles under conditions ofuse. HeiQ has submitted both a leaching study they conductedand a leaching study from the literature (Geranio, 2009). HeiQclaims that these studies indicatethat no particles were released.EPA has reviewed these studies and determined they do notsupport HeiQ claims of no particle release. An additional,follow-up leaching study is needed and this study could be donein the manner of Geranio et al. with the addition of electronmicroscopy to characterize the particulate.Appendix A - Page 10 of 13Table 1 - Summary of Tier 1 Required Data for HeiQ AGS-20OSCPP Data Requirement (Note 1)Guideline Number: Study TitleStudyStatusData TriggerTestMaterialCommentsToxicology870.3465 90-Day Inhalation Toxicity (Rat) No DataInhalation toxicity studies areconditionally required per 40CFR 161.340. In this case, thetrigger for this requirement isthe potential for inhalationexposure to AGS-20 basedupon its formulation as apowder and its use in textiles.AGS-20It is proposed that HeiQ AGS-20 will be formulated as a powder.Occupational inhalation exposures will occur during the handlingof this powder during textile treatment and during manufacturingof clothing from the treated textiles. Inhalation toxicity data areneeded to assess the risks of these exposures.The inhalation study is a route-specific study and is moreappropriate than a subchronic oral toxicity study. Inhalationstudies of shorter duration (e.g. acute inhalation studies orrepeated exposure inhalation studies less than 90 days) would notbe sufficient to identify health effects such as pulmonary fibrosis,which has been observed with other nanoparticles, and whichtakes several months to develop.870.3200 21-Day Dermal Toxicity (Rat) No DataDermal toxicity studies areconditionally required per 40CFR 161.340. In this case, thetrigger for this requirement isthe potential for dermalexposure to AGS-20 basedupon its formulation as apowder and its use in textiles.AGS-20Occupational dermal exposures could occur during the handlingof this powder during textile treatment and during manufacturingof clothing from the treated textiles. Consumer dermal exposurescould occur while wearing treated clothing. A 21 day dermaltoxicity study is needed to assess the risks of these exposures.The dermal toxicity study is route-specific, and is moreappropriate than a subchronic toxicity study via another route(e.g. oral). The use of studies via other routes requires dermalpenetration studies to estimate dermal absorption.870.3650 Combined Repeat Dose ToxicityStudy with Reproduction/DevelopmentalToxicity Screening TestNo DataOral developmental andreproductive studies areconditionally required per 40CFR 161.340. In this case, thetrigger for this requirement isthe potential for incidental oralexposure to AGS-20 basedupon its use in textiles.AGS-20There is potential for children’s incidental oral exposure to AGS20 during the wearing and mouthing of treated clothing. Thecombined repeated-dose toxicity study with thereproduction/developmental toxicity screening test will provideinitial information of potential developmental, reproductive, andother hazard concerns of AGS-20. In addition, the study mayalso provide a toxicity endpointapplicable to a risk assessmentfor oral incidental exposure.Genetic Toxicity Tests870.5100, 5300, 5375, 5385, 5395No DataThese studies are required per40 CFR 161.340.AGS-20Genetic toxicity tests are used toscreen chemicals for mutagenicor carcinogenic potential. The data from these tests will be usedto determine if AGS-20 is a potential mutagen or carcinogen.Appendix A - Page 11 of 13Table 2 - Summary of Tier 2 Required Data for HeiQ AGS-20OSCPP Data Requirement (Note 1)Guideline Number: Study TitleStudyStatusData TriggerTestMaterialCommentsProduct Chemistry830.7050: UV-Visible Light Adsorption830.7520: Particle Size, Fiber Length, andDiameter (size) DistributionNo Guideline: Surface Area Determination830.7840: SolubilityNo Guideline: Zeta Potential and SurfaceCharge DeterminationNo dataThese studies are needed ifsilver nanoparticles are releasedduring the Tier 1 stability,dissolution or leaching studies.SilverNanoparticlesEnvironmental Fate850.1730: Fish BCF No dataSeeNote 2The proposed AGS-20 use pattern may lead tobioconcentration/bioaccumation ofAGS-20, or constituents of it,which may leach or break away from AGS-20 treated items. Openliterature studies indicate that nanometals are not excreted by biotaand can accumulate. Bioconcentration studies will assess thepotential that the test substance will accumulate in fish andtherefore its potential to impact the food chain.835.1230: Adsorption/Desorption(Batch Equilibrium)No dataAGS-20If the material is removed during waste water treatment, it may bedeposited on land through the deposition of sewer sludge on land(i.e. land farming). If the material is not removed during wastewater treatment it may be released into aquatic environments andmay bind to sediment. For this reason, EPA is requestingadsorption/desorption equilibrium studies to be conducted withAGS-20. This information will be used to assess the sorption of atest material on soil/sediment and to estimate exposure.No Guideline: MITI Glucose No DataThese studies are needed ifAGS-20 or silver nanoparticlesare released during the Tier 1stability, dissolution, orleaching studies.SeeNote 2Silver from industrial processes (e.g., film processing) has beenshown to reduce microbial activity in waste water treatmentsystems. The purpose of the study isto assess the impact of AGS20 or its constituents on microbial activity in waste water treatment.EPA will use this information to determine adverse effects onmicrobial communities in waste water treatment.Appendix A - Page 12 of 13Table 2 - Summary of Tier 2 Required Data for HeiQ AGS-20OSCPP Data Requirement (Note 1)Guideline Number: Study TitleStudyStatusData TriggerTestMaterialCommentsEnvironmental Effects850.1010: Acute Toxicity FreshwaterInvertebrates850.1075: Freshwater Fish Toxicity850.2100: Avian Oral Toxicity850.5400 Algal Toxicity, Tier 2No dataThese studies are needed ifsilver nanoparticles are releasedduring stability, dissolution, orleaching studies.SilverNanoparticlesThe proposed use of AGS-20 may lead to the release of silvernanoparticles into surface water. This could impact fish,invertebrate, avian, and algal communities. These studies willaddress concerns regarding the potential risks of silvernanoparticles from AGS-20 and will provide a toxicity endpointapplicable to ecological risk assessment.Human Exposure –No Tier 2 Studies are RequiredToxicology870.3465 90-Day Inhalation Toxicity (Rat)No DataThis study is needed if silvernanoparticles are released fromAGS-20 powder during theoccupational exposure study.SilverNanoparticlesThe inhalation study is a route-specific study and is moreappropriate than a subchronic oraltoxicity study. Inhalation studiesof shorter duration (e.g. acute inhalation studies or repeatedexposure inhalation studies less than 90 days) would not besufficient to identify health effects such as pulmonary fibrosis,which has been observed with other nanoparticles, and which takesseveral months to develop.870.3200 21-Day Dermal Toxicity (Rat)No DataThis study is needed if silvernanoparticles are released fromAGS-20 powder during theoccupational exposure study, orif silver nanoparticles arereleased during the textileleaching study.SilverNanoparticlesOccupational dermal exposures to silver nanoparticles could occurwhile handling AGS-20 powder and consumer dermal exposurescould occur while wearing treated clothing. A 21 day dermaltoxicity study is needed to assessthe risks of these exposures. Thedermal toxicity study is route-specific and is more appropriate thana subchronic toxicity study via another route (e.g. oral). The use ofstudies via other routes requires dermal penetration studies toestimate dermal absorption.Although the weight-of-evidence presently indicates thatnanoparticles do not penetrate intact human skin, the possibilitycannot be completely ruled out. There is scientific evidenceindicating that, under certain conditions (e.g., formulation, size,and/or composition), nanoparticles are able to penetrate skin and beabsorbed into the body (Mortensen et al, 2008; Wu et al., 2009; andLarese et al, 2009). Due to concerns about human exposures fromintact, or more likely, abraded skin (which is common), a dermaltoxicity study on silver nanoparticles is required to assess thehazards of dermal exposure.Appendix A - Page 13 of 13Table 2 - Summary of Tier 2 Required Data for HeiQ AGS-20OSCPP Data Requirement (Note 1)Guideline Number: Study TitleStudyStatusData TriggerTestMaterialComments870.3650 Combined Repeat Dose Tox Studywith Repro/Dev. Tox Screening TestNo DataSilverNanoparticlesThere is potential for children’s incidental oral exposure to silvernanoparticles during the wearing andmouthing of treated clothing.The combined repeated-dose toxicity study with thereproduction/developmental toxicity screening test will provideinitial information of potential developmental, reproductive, andother hazard concerns of the silver nanoparticles. In addition, thestudy may also provide a toxicity endpoint applicable to riskassessment for oral incidental exposure.Genetic Toxicity Tests870.5100, 5300, 5375, 5385, 5395No DataThese studies are needed ifsilver nanoparticles are releasedduring the leaching studies.SilverNanoparticlesThe data from these studies will be used to determine if silvernanoparticles are a potentially mutagenetic or carcinogenetic.Note 1:These guidelines only provide general guidance. Protocols should be submitted prior to conducting these studies.Note 2:The test material should include materials that are released during the stability, dissolution/dispersability, or textile leaching studies. These materials include theAGS-20 Composite or Silver Nanoparticles released from the AGS-20 Composite.