Draft Decision Document 

Proposed Conditional Registration of Nanosilva as a Materials 

Preservative in Textiles and Plastics 

August 27, 2013 

Environmental Protection Agency 

Office of Pesticide Programs 

Antimicrobials Division 
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TABLE OF CONTENTS 

EXECUTIVE SUMMARY .............................................................................................................IV 

I. REGULATORY DECISION SUMMARY ................................................................................1

II. BACKGROUND .........................................................................................................................2

2.1 REGULATORY HISTORY .......................................................................................................................................2

2.2 FIFRA SCIENTIFIC ADVISORY PANEL (SAP) MEETING .......................................................................................2

III. PRODUCT DESCRIPTION, TESTING, AND USE .............................................................4

3.1 PRODUCT DESCRIPTION .......................................................................................................................................4

3.2 PRODUCT USE......................................................................................................................................................4

3.3 PRODUCT TESTING AND WAIVERS .......................................................................................................................4

IV. HAZARD CHARACTERIZATION OF NANOSILVER .....................................................8

4.1 ACUTE HUMAN TOXICOLOGY OF NANOSILVA .....................................................................................................8

4.2 SUBCHRONIC AND CHRONIC TOXICOLOGICAL STUDIES AVAILABLE FOR NANOSILVER.......................................9

4.2.1 Oral ............................................................................................................................................................. 9

4.2.2 Inhalation .................................................................................................................................................. 10

4.2.3 Dermal ...................................................................................................................................................... 12

4.2.4 Reproductive and Developmental Toxicity of Nanosilver ........................................................................ 14

4.2.5 Mutagenicity of Nanosilver....................................................................................................................... 14

4.2.6 Silver Ions ................................................................................................................................................. 16

4.3 ABSORPTION, DISTRIBUTION, METABOLISM, AND EXCRETION (ADME) OF NANOSILVER ................................17

4.3.1 Injection of Nanosilver .............................................................................................................................. 17

4.3.2 Oral Administration of Nanosilver ............................................................................................................ 18

4.4 SAFETY FACTOR FOR INFANTS AND CHILDREN (FQPA SAFETY FACTOR) .........................................................19

4.5 ANTIMICROBIAL RESISTANCE............................................................................................................................20

4.6 POINT OF DEPARTURE SELECTIONS AND TARGET MARGIN OF EXPOSURE .........................................................20

4.6.1 Point of Departures for Nanosilva ............................................................................................................. 21

4.6.2 Target Margin of Exposure ....................................................................................................................... 24

4.6.3 Margin of Exposure .................................................................................................................................. 25

V. OCCUPATIONAL AND CONSUMER RISK ASSESSMENT ..............................................27

5.1 OCCUPATIONAL RISK ASSESSMENT FOR MIXING AND LOADING THE NANOSILVA LIQUID SUSPENSION............27

5.1.1 Occupational Exposure when Preparing the Master Batch Containing Nanosilva ................................... 28

5.1.2 Occupational Margins of Exposure ........................................................................................................... 29

5.1.3 Conclusions for Occupational Risk ........................................................................................................... 32

5.1.4 Occupational Exposure and Health Data Requirements ........................................................................... 32

5.2 CONSUMER RISK ASSESSMENT FOR PRODUCTS INCORPORATING NANOSILVA ..................................................33

5.2.1 Consumer Exposures to Plastics and Textiles Incorporating Nanosilva ................................................... 34

5.2.2 Consumer Margins of Exposure to Plastics Incorporating Nanosilva ....................................................... 40

5.2.3 Consumer Margins of Exposure to Textiles Incorporating Nanosilva ...................................................... 45

5.2.4 Consumer Aggregate Margins of Exposure to Plastics and Textiles Incorporating Nanosilver ............... 50

5.2.5 Conclusions for Consumer Risk ................................................................................................................ 52

5.2.6 Consumer Exposure and Health Data Requirements ................................................................................ 54
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VI. ENVIRONMENTAL RISK ASSESSMENT ..........................................................................55

6.1 ENVIRONMENTAL FATE .....................................................................................................................................55

6.1.1 Nanosilver ................................................................................................................................................. 55

6.1.2 Silver Ions ................................................................................................................................................. 56

6.1.3 Impacts to Wastewater Treatment/Septic Systems ................................................................................... 56

6.2 ENVIRONMENTAL TOXICITY ..............................................................................................................................57

6.2.1 Silver Ions ................................................................................................................................................. 57

6.2.2 Aquatic Toxicity of Nanosilver ................................................................................................................. 57

6.3 AQUATIC RISK ASSESSMENT .............................................................................................................................59

6.3.1 Aquatic Risk Quotient ............................................................................................................................... 60

6.3.2 Conclusions for Aquatic Risk ................................................................................................................... 62

6.4 ECOLOGICAL EFFECTS AND ENVIRONMENTAL FATE DATA REQUIREMENTS. ....................................................63

VII. REGULATORY ACTION ......................................................................................................64

7.1 LEGAL FRAMEWORK..........................................................................................................................................64

7.2 FINDINGS AND REGISTRATION DECISION...........................................................................................................65

7.3 BASIS FOR CONDITIONAL REGISTRATION ..........................................................................................................66

7.3.1 Data Generation ........................................................................................................................................ 66

7.3.2 Public Interest ........................................................................................................................................... 67

7.3.3 No Unreasonable Adverse Effects ............................................................................................................ 69

References .................................................................................................................................... 71 

LIST OF TABLES 

Table 1 – Product Testing for the Nanosilva Liquid Suspension ................................................... 5

Table 2 – Acute Toxicity Profile for the Nanosilva Liquid Suspension ......................................... 8

Table 3 – Toxicity Endpoints for Use in Nanosilver Risk Assessment ........................................ 23

Table 4 – Unit Exposures for Mixing and Loading of Liquids ..................................................... 28

Table 5 – Occupational Handlers Inhalation MOEs for the Nanosilver in the Nanosilva Liquid 

Suspension .................................................................................................................................... 30

Table 6 – Occupational Handlers Dermal MOEs for the Nanosilver in the Nanosilva Liquid 

Suspension .................................................................................................................................... 31

Table 7 – Concentration of Silver in Fluid Contacting the Nanosilva Containing Plastic Coupons 

at 40 degrees Celsius ..................................................................................................................... 35

Table 8 – Concentration of Silver Released from Shirts Incorporating Nanosilva during Modified 

ISO Colour Fastness Test ............................................................................................................. 38

Table 9 – Amount of Silver Released from Shirts Incorporating Nanosilva ................................ 39

Table 10 – Incidental Oral MOEs for Children Exposed to Plastic Toys Incorporating Nanosilva

....................................................................................................................................................... 41

Table 11 – Dermal MOEs for Children Exposed to Flooring Incorporating Nanosilva ............... 43

Table 12 – Incidental Oral MOEs for Children Exposed to Flooring Incorporating Nanosilva ... 45

Table 13 – Inhalation MOE for Drying Textiles Incorporating Nanosilva .................................. 46
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Table 14 – Dermal Exposure to Textiles Incorporating Nanosilva .............................................. 47

Table 15 – Dermal MOEs for Children Exposed to Textiles Incorporating Nanosilva ................ 48

Table 16 – Incidental Oral Exposure to Textiles Incorporating Nanosilva .................................. 49

Table 17 – Incidental Oral MOEs for Children Exposed to Textiles Incorporating Nanosilva ... 49

Table 18 – Aggregate MOEs for Children Exposed to Plastics and Textiles Incorporating 

Nanosilva ...................................................................................................................................... 51

Table 19 – Nanosilver Toxicity to Aquatic Species ..................................................................... 58

Table 20 – Risk Quotients for Nanosilver in Surface Water ........................................................ 61

Appendix A: Evaluation of Nanosilva LLC’s Data Waiver Requests 

Appendix B: Data Requirements, Enforceable Schedule, and Conditions of the Proposed 

Conditional Registration for Nanosilva 

 
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EXECUTIVE SUMMARY 

EPA is proposing to grant a conditional registration for the Nanosilva LLC product named 

“Nanosilva” with a product code of NSPW-L30SS (hereafter referred to as “Nanosilva”) under 

section 3(c)(7)(C) of the Federal Insecticide, Fungicide and Rodenticide Act (FIFRA). The 

Agency’s basis for the conditional registration is that: 

 Nanosilva contains nanosilver as an active ingredient and the nanosilver in Nanosilva is 

not in any currently registered pesticide; 

 Use of Nanosilva will not cause unreasonable adverse effects on the environment during 

the period when newly required data are being developed; 

 Insufficient time has elapsed for Nanosilva to generate and submit the newly required 

data; and 

 Use of Nanosilva is in the public interest. 

Nanosilva, which contains 1% nanosilver by weight, is the subject of an application submitted by 

Nanosilva LLC in August 2009 (MRID 47828900). Nanosilva LLC requested that the nanosilver 

in Nanosilva be registered as a new active ingredient because it was not an active ingredient in 

any currently registered pesticide product. Nanosilva is a silica-sulfur-nanosilver complex where 

the nanosilver active-ingredient is attached to crystalline silica via a thiolate bond. The diameter 

of the spherical silica core is 320 nm on average and the nanosilver particles that are attached to 

the silica core have mean diameters between 6.9 and 10.6 nm. Nanosilva is proposed to be 

incorporated into plastics and textiles to suppress the growth of bacteria, algae, fungus, mold and 

mildew, which cause odors, discoloration, stains, and deterioration. The plastics and textiles will 

contain less than 0.003% silver by weight. 

EPA is proposing to make a registration decision for non-food contact uses of Nanosilva 

incorporated into plastic films, sheets, slabs, and molded parts, and textiles. Consistent with this, 

EPA is proposing to register Nanosilva for indoor use articles such as automotive parts, floor 

coverings, footwear, sportswear, uniforms and outdoor use articles such as furniture, decking, 

carpeting, and house siding. 

EPA determined that workers, consumers, and the environment could be exposed to: 

1) Silver ions released from Nanosilva; 

2) the Nanosilva complex; and/or 

3) Nanosilver that might break away from the Nanosilva complex. 

In evaluating the risk from exposure to silver ions, EPA relied on the existing reregistration 

decision for silver which concluded that the human health or ecological risk from exposure to 

silver ions derived from plastics and textiles incorporating Nanosilva are not of concern. 

In evaluating the risk from short-term exposure to Nanosilva, Nanosilva LLC submitted results 

from acute animal-toxicity tests completed using high-level doses of Nanosilva showing that 
vi 

there were no mortalities or abnormalities in test animals after administration of Nanosilva by 

oral, dermal, and inhalation exposure routes. Nanosilva caused moderate irritation to the eyes of 

test animals, and was not a skin sensitizer. Based on these results, shipping containers filled with 

Nanosilva are required to carry a label stating “CAUTION” where contact with eyes or clothing 

should be avoided. Nanosilva LLC submitted a request to waive the required intermediate-term 

toxicity studies because silver at concentrations greater than the analytical detection limit were 

not found leaching from plastic coupons incorporating Nanosilva and because of the lack of 

toxicity noted in the acute animal-toxicity tests completed using high-level doses of Nanosilva. 

To evaluate occupational exposure to the nanosilver that might break away from the Nanosilva 

complex, EPA calculated the daily-dose to workers assuming that all the silver in Nanosilva was 

freely available as nanosilver. EPA believes that this assumption overestimate the daily-dose of 

nanosilver that a person could potentially receive when working with Nanosilva. To evaluate 

consumer exposure to the nanosilver that might break away from plastics and textiles 

incorporating Nanosilva, Nanosilva LLC submitted studies showing that silver at concentrations 

greater than the analytical detection limit were not found leaching from plastic coupons and 

shirts incorporating Nanosilva. Thus, there is little exposure to nanosilver from plastics and 

textiles incorporating Nanosilva. EPA assumed that silver leaching from plastic coupons and 

shirts incorporating Nanosilva was at one-half the analytical detection limit and was in the form 

of nanosilver as found in Nanosilva. 

There are no intermediate- or long-term human or environmental toxicity studies available for 

Nanosilva or for the nanosilver released from products incorporating Nanosilva. In the absence 

of intermediate-term toxicity studies, EPA evaluated the risk from occupational and consumer 

exposure using intermediate-term hazard data available in the scientific literature for nanosilver. 

Because of the potential difference between the nanosilver in the available toxicity data and the 

nanosilver in Nanosilva along with the lack of an acceptable developmental and reproductive 

toxicity study for nanosilver, the Agency used a maximum 10-fold database uncertainty factor 

when evaluating the risk from exposure to Nanosilva and the nanosilver released from products 

incorporating Nanosilva. 

Impact to the environment was based on ecotoxicity studies available in the scientific literature 

for nanosilver and environmental exposure from indoor use was assessed assuming that 300 

million people (U.S. population) each purchased one t-shirt containing Nanosilva and from 

outdoor use was assessed assuming that 1% of the projected yearly production of plastic lumber 

in the U.S. would contain Nanosilva. 

Using robust toxicity data on nanosilver, conservative occupational exposure assumptions, 

leaching data showing minimal consumer exposure to nanosilver, and maximum values for risk 

uncertainty factors, EPA is able to determine that for the period of conditional registration, there 

is a low probability of adverse risk to human health and the environment from plastics and 

textiles incorporating Nanosilva. Thus, the Agency concludes that use of Nanosilva will not 
vii 

cause unreasonable adverse effects on the environment during the period when newly required 

data are being developed. 

As a condition of the proposed registration, EPA is proposing to require that Nanosilva LLC 

conduct studies during the period of conditional registration to better characterize the nanosilver 

in Nanosilva. In addition, EPA is proposing to require the following inhalation route-specific 

studies: 

 90-Day Inhalation Toxicity (Rat) (OCSPP 870.3465) modified to include in vivo bone 

marrow assay and functional observational battery, motor activity and detailed 

neuropathology 

 Reproduction/Developmental Toxicity Screening Test (Modified OCSPP 870.3550/ 

OECD TG 421) 

Although there is no risk concern for workers who use close-system loading when mixing and 

loading the Nanosilva liquid suspension, EPA is proposing to require these tests to confirm the 

adequacy of the 10-fold database uncertainty factor, to reduce the uncertainties related to 

differences in the physical properties of the nanosilver, and because there are currently no 

acceptable studies on the reproductive and developmental toxicity for nanosilver. 

These studies must be completed within four years after issuing the registration. This time period 

was chosen to allow time for protocol reviews prior to initiation of the studies, completion of the 

studies, and Agency review of the study results. The Agency will evaluate these data as they are 

submitted to confirm the Agency’s determination that the use of Nanosilva will not cause 

unreasonable adverse effects to human health and the environment. If Nanosilva LLC fails to 

take appropriate steps to initiate the required studies, or if Nanosilva LLC fails to submit the 

protocols or data, EPA is proposing to issue a notice of intent to cancel Nanosilva’s registration 

under FIFRA section 6(e). In addition, the conditional registration for Nanosilva will 

automatically expire four years after being issued. Nanosilva LLC should request an amendment 

to remove the expiration date once they have submitted the required data if they wish to continue 

to sell and distribute Nanosilva in the United States. 

EPA believes that the use of Nanosilva is in the public interest. Use of Nanosilva may lead to 

less environmental loading of silver as compared to currently registered products with the same 

use patterns. In addition, Nanosilva appears to offer prolonged ability to suppress the growth of 

odor and stain causing bacteria as compared to currently registered products containing silver 

salts with the same use patterns. 
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I. REGULATORY DECISION SUMMARY 

EPA is proposing to grant a conditional registration for the Nanosilva LLC product named 

“Nanosilva” under section 3(c)(7)(C) of the Federal Insecticide, Fungicide and Rodenticide Act 

(FIFRA). Nanosilva, which contains 1% nanosilver by weight, is the subject of an application 

submitted by Nanosilva LLC in August 2009 (MRID 47828900). Nanosilva LLC requested that 

the nanosilver in Nanosilva be registered as a new active ingredient because it was not an active 

ingredient in any currently registered pesticide product. 

As described in the risk assessment sections of this document, EPA has determined that there is a 

low probability of adverse risk to human health and the environment from plastics and textiles 

incorporating Nanosilva during the period required for developing and submitting protocols for 

review, conducting the studies, and submitting the resulting data, as well as EPA’s review of the 

submitted data. Thus, the Agency concludes that use of Nanosilva will not cause unreasonable 

adverse effects on the environment during the period when newly required data are being 

developed. 

In August, 2009 Nanosilva LLC submitted acute toxicity studies required under 40 CFR part 

161, a study that determined the amount of silver leaching from plastic coupons incorporating 

Nanosilva, and a request to waive required studies. The waiver request was based on the low 

exposure to Nanosilva given that the product was proposed to be embedded in plastic and the 

submitted leaching study for a plastic incorporating Nanosilva indicated that silver was not 

released at concentrations above the analytical detection limit. In May 2013, Nanosilva LLC also 

submitted a study showing that silver was not released at concentrations above the analytical 

detection limit from textiles incorporating Nanosilva. EPA’s proposal is to grant the registration 

request with requirements for new data as terms and conditions on the Nanosilva registration. 

Nanosilva LLC, however, has not had sufficient time to generate and submit these newly 

required data. 

For a variety of reasons, EPA has determined that use of Nanosilva is in the public interest. EPA 

believes that Nanosilva LLC’s product may lead to less environmental loading of silver as 

compared to currently registered products containing silver salts with the same use patterns. In 

addition, Nanosilva appears to offer prolonged ability to suppress the growth of odor and stain 

causing bacteria as compared to currently registered products containing silver salts with the 

same use patterns. Moreover, because we can find that a section 3(c)(7)(C) registration is 

unlikely to cause unreasonable adverse effects on the environment during the time while 

Nanosilva LLC is generating the newly required data, allowing such registration to be granted 

before the newly-required data is generated allows the Nanosilva LLC product to compete with 

other like-situated products. Allowing Nanosilva LLC’s product on the market pending 

generation of data allows Nanosilva LLC to participate in the plastics and textile economy along 
2 

with the other registrants with like-situated products, and allows consumers of these products a 

new choice among the potential like-situated products. 

EPA intends to require Nanosilva LLC to conduct a number of studies as a condition of 

registration. These studies must be completed within a time duration of four years which was 

chosen to allow time for protocol reviews prior to initiation of the studies, completion of the 

studies, and Agency review of the study results. The Agency will evaluate these data as they are 

submitted during the period of the conditional registration to confirm the Agency’s risk 

assessment. If Nanosilva LLC fails to take appropriate steps to initiate the required studies, or if 

Nanosilva LLC fails to submit the protocols or data, EPA has the authority to issue a notice of 

intent to cancel Nanosilva LLC’s registration under FIFRA section 6(e). In addition, Nanosilva 

LLC’s conditional registration for Nanosilva will automatically expire four years after being 

issued. Nanosilva LLC will have to submit an application to obtain an unconditional registration 

if they wish to continue to sell and distribute Nanosilva in the United States. 

II. BACKGROUND 

2.1 Regulatory History 

In August 2009, Nanosilva LLC submitted an application for the registration of a new 

antimicrobial pesticide product named Nanosilva, a nanosilver-based product that was proposed 

for use as a materials preservative additive with no food contact uses. Nanosilva LLC’s 

application claimed that Nanosilva was a new active ingredient not contained in any currently 

registered silver-based pesticide products and should be given a registration under FIFRA section 

3(c)(5) (MRID 47828900). 

2.2 FIFRA Scientific Advisory Panel (SAP) Meeting 

In November, 2009 the Agency convened a meeting of the FIFRA Scientific Advisory Panel 

(SAP) to address a number of questions associated with assessing the hazard of and exposure to 

nanosilver and other nanoscale metal-based pesticides (FIFRA SAP, 2009). In general, the SAP 

advised that the toxicity of nanosilver could differ from and might be higher than other forms of 

silver (e.g., silver ions). 

The SAP was unsupportive of bridging among silver-based materials with different properties. 

However, the SAP indicated that bridging would be appropriate for materials of similar size and 

essentially identical physical properties and that bridging between silver ions released from 

nanosilver and the existing database for silver ions is feasible (FIFRA SAP, 2009). The SAP 

cautioned about extrapolating from one nanosilver formulation to another when assessing 

hazards because differences in particle formulation (e.g., coating and inert ingredients) are likely 

to affect biological activity, among other things. 
3 

The SAP commented that not enough literature is available to draw any firm conclusions 

regarding human (occupational or consumer) and environmental exposures to nanosilver under 

realistic use scenarios. Potentially, three major routes exist for human exposure to nanoparticles: 

oral, inhalation, and dermal. Only a few studies in rodents are known that investigate the toxicity

of nanosilver from exposure by these routes. Nor is there much information on the level of 

human exposure to nanosilver by these routes, for either workers or consumers using products 

containing nanosilver. The same situation exists for the environmental fate and transport of 

nanosilver. The ability to measure concentrations of nanosilver in the environment along with the 

environmental exposure pathways, bioavailability, toxicity, and potential impact of nanosilver on 

ecological systems are not well quantified. Furthermore, little or no information on the fate of 

nanosilver in soils and sediments was found. As a result, the SAP recommended a case-by-case 

approach to hazard and exposure assessment (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, dissipation, and accumulation of nanomaterials. 

The SAP report further suggested that existing information on conventional silver could be 

useful but would not necessarily be sufficient in assessing potential nanosilver risks. 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 

requirements and the conduct of risk assessments). Moreover, the Panel recommended that EPA 

require additional data on the physical chemistry, exposure potential, and the potential hazard to 

human health and the environment. 

Historically, EPA has considered applications for pesticide products that claim to be identical or 

substantially similar in composition to a registered product as so-called “me-too registrations” 

under FIFRA registration authorities. Until recently, EPA generally has not focused on the size 

or surface coating of an ingredient as attributes relevant to determining if the product in an 

application is identical or substantially similar in composition to a registered pesticide product. 

However, a nanoscale ingredient may have properties that are different from those of 

conventionally-scaled ingredients and properties that differ from the atoms or molecules from 

which the nanoscale ingredient is constructed. Therefore, a nanoscale ingredient may also have 

different environmental health and safety properties. Accordingly, for a product containing an 

ingredient that is a nanoscale version of a conventionally-sized active or inert ingredient 

contained in an already-registered product or a different nanoscale version of a nanoscale 

material that is an active or inert ingredient in an already registered pesticide product, EPA 

necessarily will need to assess the nanoscale material and may require additional data to make 

the requisite statutory findings. 
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III. PRODUCT DESCRIPTION, TESTING, AND USE

3.1 Product Description 

Nanosilva (NSPW-L30SS) is a liquid suspension containing silica-sulfur-nanosilver particulates 

where the nanosilver active-ingredient is attached to crystalline silica via a thiolate bond. The 

particles are formed by reacting silver nitrate and polyvinylpyrrolidone (PVP) with spherical 

silica particles that have been modified with thiol groups and are suspended in ethanol (Lee et al. 

2007). The diameter of the spherical silica core particles are 320 nm on average (Lee et al., 

2006). The nanosilver particles form with time and have an average diameter of 6.9 nm 

(minimum diameter of 3 nm and maximum diameter of 12 nm) after 10 minutes and 10.6 nm 

(minimum diameter of 5 nm and maximum diameter of 18 nm) after 2 hours (Lee et al., 2007). 

The liquid suspension contains 1% nanosilver by weight with average diameters between 6.9 and 

10.6 nm (minimum diameter of 3 nm and maximum diameter of 18 nm), where the nanosilver 

surface is coated with sulfur and PVP, and is attached to silica. 

The proposed labels states that Nanosilva (NSPW-L30SS), which contains 1% nanosilver by 

weight, is to be formulated into a polymeric intermediate known as a master batch. Nanosilva 

LLC proposed that the final product contain between 5 and 15% by weight of the polymeric 

intermediate or master batch that contains Nanosilva. Although not stated on the proposed label, 

the final product that a consumer may be exposed to will contain less than 0.003% nanosilver by 

weight. EPA is proposing that Nanosilva LLC state on the pesticide label that products 

incorporating Nanosilva contain a maximum of 0.003% nanosilver by weight. 

3.2 Product Use 

Nanosilva LLC is proposing that Nanosilva be incorporated into polymer and polymer based 

products to suppress the growth of bacteria, algae, fungus, mold and mildew, which cause odors, 

discoloration, stains, and deterioration. Nanosilva LLC proposed a wide range of non-food 

contact use categories for Nanosilva including house wares, building materials, bathroom 

fixtures and accessories, electronics and appliances, personal care products, automotive 

equipment, hospital and institutional facility equipment, sporting goods, and textiles. 

3.3 Product Testing and Waivers 

As an applicant for registration of a new active ingredient in 2009, Nanosilva LLC was required 

to submit all the applicable information and studies under 40 CFR part 161. Nanosilva LLC 

submitted results from testing of Nanosilva to determine the product identity and composition, 

physical and chemical properties, and acute toxicity as summarized in Table 1. Some of these 

tests were completed using standard EPA test guidelines1

 prior to consulting with EPA. Although 

 

1

 Available at http://www.epa.gov/ocspp/pubs/frs/home/guidelin.htm 
5 

these test results provided useful information for Nanosilva, the guidelines on which they are 

based have not been adapted generally for use with nanoscale particles. EPA anticipates that 

these guidelines will require revision going forward in terms of their application to nanoscale 

materials. As a result, it is recommended that future applicants for products containing nanoscale 

materials consult with the EPA prior to performing any tests. 

Table 1 – Product Testing for the Nanosilva Liquid Suspension 

Guideline 

Number 

Guideline Name Required under 40 

CFR 

Nanosilva LLC 

Submission 

(MRID) 

Product Identity and Composition 

830.1550 Product identity and composition 161.155 47828901 

830.1600 Description of materials to produce 

product 

161.160 47828902 

830.1620 Description of production process 161.162 47828903 

830.1650 Description of formulation process 161.165 47828904 

830.1670 Discussion of formulation of 

impurities 

161.167 47828905 

830.1700 Preliminary analysis 161.170 47828906 

830.1750 Certified limits 161.175 47828907 

830.1800 Enforcement analytical method 161.180 47828908 

Physical and Chemical Properties 

830.6302 Color 161.190 47828909 

830.6303 Physical state 161.190 47828910 

830.6304 Odor 161.190 47828911 

830.6317 Storage stability 161.190 47828912 

830.6320 Corrosion characteristics 161.190 47828912 

830.7000 pH 161.190 47828913 

830.7100 Viscosity 161.190 47828914 

830.7200 Melting point/melting range 161.190 

830.7220 Boiling Point/Boiling Range 47828915 

830.7300 Density 161.190 47828916 

830.7520 Particle size, fiber length, and 

diameter distribution 

Not Required, information 

in Lee et al., 2006 and 

2007 

830.7840 Water solubility 161.190 47828917 

Health Effects (Toxicology) 

870.1100 Acute Oral Toxicity 161.340 47828918 

870.1200 Acute Dermal Toxicity 161.340 47828919 

870.1300 Acute Inhalation Toxicity 161.340 47828920 

870.2400 Acute Eye Irritation 161.340 47828921 

870.2500 Acute Dermal Irritation 161.340 47828922 

870.2600 Skin Sensitization 161.340 47828923 
6 

In addition to the test results listed in Table 1, Nanosilva LLC submitted the following study to 

support the registration of Nanosilva: 

Plastic Leaching Study

 Leaching protocol for Nanosilva treated LLDPE polymer in food and food simulated 

matrices as functions of time, temperature and chemistry of the matrix with determined 

migration values (MRID 47828925) 

Nanosilva LLC also submitted a request to waive the following data: 

1. Hydrolysis (OPPTS 835.2120) 

2. Aquatic Invertebrate Acute Toxicity Test, Freshwater Daphnids (OPPTS 850.1010) 

3. Fish Acute Toxicity Test, Freshwater and Marine (OPPTS 850.1075) 

4. Avian Acute Oral Toxicity Test (OPPTS 850.2100) 

5. 90-Day Dermal Toxicity (OPPTS 870.3250) 

6. Prenatal Developmental Toxicity Study ( OPPTS 870.3700) 

7. Bacterial Reverse Mutation Test (OPPTS 870.5100) 

8. Detection of Gene Mutations in Somatic Cells in Culture (OPPTS 870.5300) 

9. In Vitro Mammalian Cytogenetics (OPPTS 870.5375) 

10. Mammalian Bone Marrow Chromosome Aberration Test (OPPTS 870.5385) 

11. Immunotoxicity (OPPTS 870.7800) 

Waivers for the above studies were requested because of the lack of acute toxicity noted in the 

six acute toxicity studies (MRID 47828918 through 47828923), because Nanosilva has a specific 

gravity greater than water, because Nanosilva has low solubility in water as determined by the 

water column generator method (MRID 47828917), and due to the non-leaching characteristic of 

Nanosilva because silver at concentrations greater than the analytical detection limit were not 

detected in leaching studies conducted with a plastic incorporating Nanosilva (MRID 47828925). 

As detailed in Appendix A, the data requirements listed above in items 1 through 11 are either 

waived or satisfied. In general, the data requirement is met or satisfied because: 

 certain guideline studies are not appropriate for nanosilver and where appropriate are 

being modified and required as terms and conditions on the registration (i.e., 

Reproduction/Developmental Toxicity Screening Test) 

 certain environmental data are satisfied because the environmental hazard labeling 

statement was determined using literature studies completed with nanosilver 

 Dermal guideline studies are not appropriate given there was no risk concern 

 in vitro mutagenicity study requirements are satisfied and have shown that nanosilver is 

possibly mutagenic 

 the immunotoxicity study is initially waived and only triggered after the potential 

immune toxicity of nanosilver is determined during the 90-day inhalation toxicity study 

required as a term and condition on the registration 
7 

In response to EPA’s review of the leaching and product chemistry studies, Nanosilva LLC 

subsequently submitted the following documents: 

Nanosilver Content of Plastic Coupons Containing Nanosilva

 Determination of Silver Content and Silver Recovery Rate for NSPW-L30SS (MRID 

48652901) 

Description of Production and Formulation Process

 Lee, J.M., Kim, D.W., Jun, Y.D., Oh, S.G. 2006. Preparation of silica-silver 

heterogeneous nanocomposite particles by one-pot preparation strategy using polyol 

process: Size-controlled immobilization of silver nanoparticles. Materials Research 

Bulletin 41:1407-1416.(MRID 48379901 and 48379903) 

 Lee, J.M., Kim, D.W., Kim, T.H., Oh, S.G. 2007. Facile route for preparation of silicasilver heterogeneous nanocomposite particles using alcohol reduction method. Materials 

Letters 61:558-1562. (MRID 48379902 and 48379904) 

Textile Leaching Study

 NanoSilva LLC prepared and submitted on August 20, 2012 a draft protocol titled “The 

Quantification and Characterization of Silver Released from Textiles Treated with 

NanoSilva (NSPW-L30) as a Results of Washing” 

 The Quantification and Characterization of Silver Released from Textiles Treated with 

NSPW-L30SS As a Result of Simulated Laundering Conditions. Study Number: 

110112.0001 Revision 2 (MRID 49045301) 
8 

IV. HAZARD CHARACTERIZATION OF NANOSILVER 

Nanosilver is a broad spectrum antimicrobial agent that works by releasing ionic silver but also 

exhibits particle-specific effects (Wang et al. 2013). In November, 2009 the Agency convened a 

meeting of the FIFRA Scientific Advisory Panel (SAP) to address a number of questions 

associated with assessing the hazard of and exposure to nanosilver and other nanoscale metalbased pesticides (FIFRA SAP, 2009). In general, the SAP advised that the toxicity of nanosilver 

could differ from and might be higher than other forms of silver (e.g., silver ions). The Panel 

agreed with EPA that particle size has a substantial impact on particle properties, including rate 

and concentration of silver ion release, where the effects of size are generally most observable 

for particles with dimensions below 20 nm and largely below 10 nm (2009 SAP, p. 6). In 

addition to size, other properties such as shape, charge, and surface coating are also likely to 

impact the biological response to nanosilver. 

4.1 Acute Human Toxicology of Nanosilva 

Nanosilva LLC submitted results from guideline acute animal-toxicity tests completed using 

high-level doses of a liquid suspension containing Nanosilva with 1% nanosilver by weight. As 

outlined in Table 2, there were no mortalities or abnormalities noted in test animals after 

administration of Nanosilva by oral, dermal, and inhalation routes at dose levels of up to 5,000 

mg/kg, and 2.05 mg/L, respectively; Nanosilva caused moderate to no irritation to skin or eyes at 

dose levels of up to 0.1 mL and 0.5 mL, respectively; and was not a skin sensitizer. According to 

the Agency’s Toxicity Category system, which is used for product labeling purposes, shipping 

containers filled with Nanosilva are required to carry a label stating “CAUTION” where contact 

with eyes or clothing should be avoided. 

Table 2 – Acute Toxicity Profile for the Nanosilva Liquid Suspension 

Study Toxicity Category 

Acute Oral Toxicity Category IV 

No mortality or abnormalities after dose of 5,000 mg/kg 

Acute Dermal Toxicity Category IV 

No mortality or abnormalities after dose of 5,000 mg/kg 

Acute Inhalation Toxicity Category IV 

No mortality or abnormalities after dose of 2.05 mg/L 

Acute Eye Irritation Category III 

Moderate to not irritating after dose of 0.1 mL 

Acute Dermal Irritation Category IV 

Mild or slight irritation after dose of 0.5 mL 

Skin Sensitization Not a sensitizer 
9 

4.2 Subchronic and Chronic Toxicological Studies Available for Nanosilver 

There are no repeated-dose subchronic or chronic toxicity studies available for Nanosilva or the 

nanosilver in Nanosilva. However, there are repeated-dose toxicity studies available in the 

scientific literature for nanosilver. Since nanosilver might be released from Nanosilva and 

articles incorporating Nanosilva, the Agency considers the scientific literature studies on 

nanosilver toxicity relevant. The Agency believes the studies described in the following sections 

are sufficient for assessing the risks from the use of Nanosilva during generation of the requisite 

data. 

4.2.1 Oral 

There are currently three studies in the scientific literature that investigate the oral toxicity of 

nanosilver in rats and two studies completed with mice. The first reported findings after 28 days 

of repeated administration of carboxymethyl cellulose-coated nanosilver with average diameter 

of 60 nm (minimum diameter of 53 nm and maximum diameter of 71 nm) to four-week old male 

and female Sprague-Dawley rats (n = 10 per dose) (Kim, et al, 2008). There were liver effects 

(dilation of the central vein, bile-duct hyperplasia and increased foci), a coagulative effect on 

peripheral blood, and an increase in serum alkaline phosphatase (ALP) and cholesterol. There 

was a dose-dependent increase in silver distribution in many tissues (liver, kidneys, stomach, 

brain, lungs, testes, and blood) and a two-fold higher accumulation in the kidneys of female rats 

when compared with male rats was also reported for all dose groups. A no observed adverse 

effect level (NOAEL) of 30 mg/kg/day (lowest dose level) was reported based on the observed 

liver effects and increase in alkaline phosphatase (ALP) and cholesterol at 300 mg/kg/day (mid 

dose level). The second study was performed by the same group using four-week old Fisher rats 

(n= 10 per dose) for 90 days (Kim et al., 2010). This study involved repeated administration of 

carboxymethyl cellulose-coated nanosilver with average diameter of 56 nm (minimum diameter 

of 25 nm and maximum diameter of 125 nm) to rats and reported similar findings as the 28-day 

study including gender-related distribution of silver in the kidneys and a reported NOAEL of 30 

mg/kg/day. However, intestinal pigmentation from exposure to nanosilver was reported, which 

was not observed in the 28 day study. 

The third study involved feeding nanosilver stabilized with polyvinylpyrrolidone with average 

diameter of 14 nm and silver acetate to four-week old male (n= 6 per dose) and female (n=10 per 

dose) Wistar Hannover Galas rats for 28 days (Hadrup et al. 2012a). Consistent to the studies 

above, investigators did not find increases in ALP or cholesterol for nanosilver at doses of up to 

9 mg/kg/day. They also reported no observable effects on the microbiological status of the rat’s 

gastrointestinal tract caused by ingesting nanosilver. However, Hadrup et al., (2012a) did report 

effects for silver acetate at a dose of 14 mg/kg/day including an increase in ALP, decrease in 

plasma urea, and lower thymus weights. Hadrup et al. (2012b) compared the neurotoxic effects, 

in vivo and in vitro, of polyvinylpyrrolidone stabilized nanosilver with average diameter of 14 

nm and silver acetate. Following 28 days of oral administration, nanosilver (4.5 and 9 
10 

mg/kg/day) and silver acetate (9 mg/kg/day) significantly increased the concentration of 

dopamine in the brains of Wistar female rats, while the brain concentration of 5-

hydroxytryptamine (5-HT) was increased only by nanosilver at a dose of 9 mg/kg/day. However, 

in the 14-day range-finding study, the brain dopamine concentration decreased in rats treated 

with nanosilver at doses of 2.25 and 4.5 mg/kg/day. Three solutions consisting of 1) nanosilver, 

2) an ionic silver solution obtained by filtering a nanosilver suspension, and 3) silver acetate 

were tested in neuronal-like PC12 cells in vitro. Nanosilver did not induce necrosis; however, 

cell viability was decreased and apoptosis (involving both the mitochondrial and the death 

receptor pathways) was found with all three solutions where silver acetate was relatively more 

potent. 

There is one study for the repeated administration of nanosilver with average diameter of 42 nm 

(minimum diameter of 25 nm and maximum diameter of 55 nm) to mice over 28 days (Park, et 

al., 2010). The study reported that, after oral administration of nanosilver at the dose levels of 

0.25 mg/kg/day, 0.5 mg/kg/day, or 1.0 mg/kg/day, the serum enzyme levels of alkaline 

phosphatase (ALP) and aspartate transaminase (AST) were significantly elevated in both male 

and female mice in the high dose group. The level of alanine transaminase (ALT) was also 

elevated in the high dose females. Histopathological analysis was performed in the high-dose 

groups and revealed that tissue change (i.e., slight cell infiltration) was observed in the cortex of 

the kidneys in both male and female mice, but no other histopathological changes were found in 

the portions of liver or small intestines that were examined. A NOAEL of 0.5 mg/kg/day was 

reported, based on the observed findings of elevated ALP, AST and ALT and the 

histopathological changes in the kidneys at the 1.0 mg/kg/day dose level. 

Liu et al. (2013) reported that nanosilver did not affect spatial cognition or hippocampal 

neurogenesis in adult male ICR mice (n=15 per dose, n=10 for control). Adult mice were 

administered nanosilver with average diameter of 51.4 nm by DLS and 26.3 nm (size range from 

10 to 70 nm) by TEM via intraperitoneal injection, at doses of 0, 10, 25, or 50 mg/kg, once a day 

in the morning for 7 consecutive days. Another group of mice received scopolamine (3mg/kg) as 

a positive control for the behavioral studies. Investigators used the Morris water maze (MWM) 

test for spatial cognition along with bromodeoxyuridine to detect proliferating cells. The test 

results showed that both reference memory and working memory were not impaired in 

nanosilver exposed groups, compared with the control group, and no differences were revealed 

in hippocampal progenitor proliferation, new born cell survival, or differentiation in nanosilver 

treatment groups. 

4.2.2 Inhalation 

The subchronic inhalation toxicity of nanosilver was determined in six-week old male and 

female Sprague-Dawley rats (n=10 per dose) exposed to nanosilver with average diameters 

between 18 to 19 nm (minimum diameter of 2 nm and maximum diameter of 65 nm) in a whole 
11 

body inhalation chamber for 13 weeks (90-day) using OECD test guideline 413 (Sung et al., 

2009). Male and femal rats exposed to the high-dose level of 515 μg/m3

 (3.03×106 particles/cm3) 

showed toxic effects in the liver (bile-duct hyperplasia) and lungs (chronic alveolar inflammation 

and macrophage accumulation in the lungs of males and females, and erythrocyte aggregation). 

A follow-on study was performed to determine lung-function recovery of five-week old male 

and female Sprague-Dawley rats (n= 17 male and 12 female per dose) after exposure to 

nanosilver with average diameters between 14 to 15 nm (minimum diameter of 4 nm and 

maximum diameter of 47 nm) in a whole body inhalation chamber for 12 weeks (Song et al., 

2012). The pathological findings in the lungs were very similar to those observed in Sung et al. 

(2009) even though there were slight differences in particle size and concentration. During the 

12-week exposure period, there was a decrease in lung function of male animals exposed to all 

doses of nanosilver including a decrease in tidal and minute volume, and peak expiratory flow. 

After a 12-week recovery period, the male animals in the middle- (117 μg/m3 or 1.41×106

particles/cm3

) and high-dose (381 μg/m3 or 3.24×106 particles/cm3) groups did not achieve 

complete recovery of lung function. Remarkably, no adverse lung function effects were observed 

for female animals. The decrease in lung function was matched by histopathological 

observations of mixed cell infiltrate perivascular and chronic alveolar inflammation in male 

animals but these effects were also observed in female animals even though they had no 

measurable decrease in lung function. Although female animals showed gradual clearance of 

silver from lung tissues and decreased inflammation after 12 weeks of recovery, persistent 

inflammation in lung tissues was observed in the high-dose male animals throughout the 12-

week recovery period. This lack of recovery indicates permanent or persistent organ damage. 

Significant increases in the amount of silver in tissues, such as lungs, liver, olfactory bulb, brain, 

kidneys, and blood, were also reported. Females had two to three times more silver accumulation 

in their kidneys than males. 

Liu et al. (2012) investigated the effects of nanosilver on hippocampal synaptic plasticity and 

spatial cognition in adult male Wistar rats (n=8 per dose). Uncoated nanosilver with average 

diameter of 244.5 nm (size range from 33 to 380 nm) by Dynamic Light Scattering (DLS) and 

with sizes from 50 to 100 nm by Transmission Electron Microscopy (TEM) was administered to 

rats by nasal drops at doses of 3 mg/kg and 30 mg/kg once every two days for 14 consecutive 

days. Spatial cognition as determined by the Morris water maze (MWM) test showed that 

animals receiving nanosilver exhibited lower learning ability and memory retention, which was 

more prominent in the high-dose group (30 mg/kg). This finding was also supported by impaired 

synaptic plasticity as determined by long-term potentiation (LTP) recording for these same 

animals. Further, the quantity of reactive oxygen species (ROS) in hippocampal homogenate was 

significantly increased in nanosilver treated groups, and edema, nuclear shrinking phenomenon, 

and necrobiosis were shown in hematoxylin and eosin (HE) stains of pyramidal neurons in the 

PP (perforant path) and DG (dentate gyrus) regions of hippocampus. 
12 

4.2.3 Dermal 

There are only two dermal toxicity studies available for nanosilver. One study was performed to 

determine liver, skin and spleen pathologies of five to six week old male guinea pigs (n=6) after 

exposure to nanosilver (concentrations 100 μg/ml, 1000 μg/ml, and 10,000 μg/ml) with a particle 

size of less than 100 nm based on daily rub exposure to aqueous solution using AgNO3 as a 

positive control (Korani et al, 2011). The nanosilver suspension was applied by daily rubbing to 

an area of 5 cm by 5 cm on the back of the animal with no wipe off or removal of chemical 

mentioned. The applied dose in mg/kg was not determined in the study. According to email 

contact with authors, animals were restrained for 5 hours after application of chemical, however, 

the method of restraint was not discussed. During the 13 week study, dose dependent and 

nanosilver specific effects were seen in the liver, the spleen and on the skin. Skin effects 

included decreased thickness of the epidermis and dermis, inflammation, increased levels of 

round cells, acidophilic cytoplasm in muscle fibers and increased levels of macrophages in the 

endomysium. Liver effects included overproduction of Kupffer cells and degeneration of 

hepatocytes in a dose dependent fashion and necrosis at the 10,000 μg/ml concentration. In the 

liver, red capsules were thinner, inflammation, accumulation of red blood cells, and white pulp 

atrophy along with hepatic cord destruction in a dose dependent fashion. It does not appear that 

gauze or other dressing was used to cover the dosed area. No hematology, clinical chemistry or 

urinalysis was performed. Only the liver, spleen and skin were examined histopathologically. 

Another study was performed to determine the effect of skin exposure to nanosilver (size = 20 

and 50 nm, concentration = 0.34, 3.4 and 34 μg/ml aqueous solution) in 2 female pigs (Samberg 

et al, 2010). A 500 μL nanosilver suspension was placed on one of 14 spots on the back of a a 

hair-clipped pig, allowed to air dry, then covered with a Hilltop chamber (occlusion pad) and 

secured with non irritating tape. This was followed by a body stocking covering the dorsum of 

each pig. Actual exposure was 0.6, 6, and 60 ng/mm2

 and total exposure was 0.17, 1.7 and 17 μg 

per dosing period. No gross pathological effects were noticed on the porcine skin. A 

concentration, but not particle size or washing state, dependent effect was seen on the dermal 

layers under microscopic investigation. Low dose effects were slight intra and intercellular 

epidermal edema. Intermediate effects were a more focal intra and intercellular epidermal edema 

alongside focal dermal and epidermal inflammation. The highest dose caused severe edema, with 

severe focal dermal inflammation, epidermal hyperplasia and parakeratosis. Precautions were 

taken to prevent oral dosing, including restraint/anesthesia and multiple covering layers. Only 

effects relating to the skin were examined. 

None of the above studies are an acceptable substitute for a dermal subchronic study or a dermal 

irritation study. In no case was a full gross or microscopic histopathology panel performed, even 

in the cases where dose dependence effects of nano-silver particles was seen. None of these 

studies used the same size particles, coating type or washing state, making it potentially difficult 

to generalize information from them onto other products. In Korani et al. (2011), the study 
13 

lacked any clinical chemistry, hematology or urinalysis of the guinea pigs. This study also could 

not draw NOAEL/LOAEL conclusions given that effects were seen at all levels of dosing. In 

Samberg et al. (2010), no systemic histopathology would have been possible, as multiple dose 

concentrations were tested on the same animal. This study would not suffice as a dermal 

irritant/sensitizer as there is no induction or recovery period. 

In addition, there is one published report on nanosilver that is available for use by the Agency 

concerning nanosilver used in wound dressings for patients suffering from burns. In this report, a 

burn patient using nanosilver coated wound dressing developed clinical signs of argyria and 

elevated serum liver enzymes indicative of liver toxicity along with elevated silver 

concentrations in blood and urine (Trop, 2006). This study indicates to the Agency that 

nanosilvers can be systemically absorbed when a large area of the skin barrier is severely 

compromised. 

In the absence of any such dermal toxicity studies, the Agency normally uses extrapolation from 

another route of exposure (usually oral). However, use of the oral endpoint for evaluating dermal 

toxicity requires knowledge regarding the amount absorbed through the skin. This information is 

typically provided by a dermal penetration study using whole animals (usually rats), which 

allows determination of the fraction of topically applied dose that is available for systemic 

absorption (i.e., the dermal absorption factor or DAF). There are in vitro techniques available 

that allow determination of dermal penetration of chemicals through isolated animal or human 

skin. However, the Agency does not rely on in vitro dermal absorption study data as the sole 

basis for deriving a DAF because standardized in vitro test methodology is not currently 

available. Without detailed, standardized methodology for in vitro absorption studies, the 

Agency has observed variation in test results among laboratories. As a result, the Agency 

assumes a default DAF of 100% when no in vivo data are available. Therefore, if in vivo dermal 

penetration studies are available, either those studies alone or in combination with in vitro 

studies are used for deriving a DAF of less than 100%. 

Here, there is no guideline or scientific literature study conducted in animals for the in vivo

dermal absorption of the nanosilver in Nanosilva or of nanosilver available to the Agency. 

However, there is a human clinical study, which is observational, that examined silver levels in 

serum and urine after application of burn wound cream containing silver sulfadiazine nanosilver 

(Wan et al., 1991). EPA used this information to derive a conservative DAF of 6.7% for 

nanosilver. A study completed by Brandt et al. (2012) demonstrated that nanosilver and silver 

sulfadiazine had similar skin absorption characteristics in mice after normalizing for silver dose. 

In addition an in vitro study with nanosilver in human skin is available in the scientific literature 

indicating that nanosilver penetration was very low for both intact and abraded skin at 0.00066% 

and 0.0033%, respectively (Larese et al., 2009). 
14 

4.2.4 Reproductive and Developmental Toxicity of Nanosilver 

There are studies showing significant, dose-dependent increases in the concentration of silver in 

the testes of rats after oral ingestion, inhalation, and injection of nanosilver (see Section 4.3); 

however, there are few studies available on the reproductive and developmental toxicity of 

nanosilver. 

There is an in vitro study investigating the toxicity of 15 nm nanosilver on spermatagonia 

isolated from 6-day old mouse testes and immortalized with SV40 large T antigen (BraydichStolle et al., 2005). In this transformed (i.e. immortalized) cell line, nanosilver and silver ions 

caused altered cellular morphology, decreased mitochondrial activity (as indicated by MTS 

assay), and increased apoptosis at doses up to 10 μg/ml; however, the affects from nanosilver 

were greater than observed for silver ions. The conservative interpretation of this data is that 

nanosilver that reaches the testes may be able to cause decreased fertility due to toxicity to 

spermagonia and this effect would be more severe for nanosilver than for silver ion. 

Austin et al. (2011) investigated the distribution of citrate coated nanosilver with diameters 

between 30 and 60 nm and silver nitrate in pregnant mice (n= 6 to 12 per dose) and developing 

embryos. Nanosilver suspensions and silver nitrate were administered by intravenous injection 

(i.v.) on gestation days (GD) 7, 8, and 9 at nanosilver concentrations of 0, 0.4, and 0.73 

mg/kg/day. Austin et al. (2011) reported a significant increase in nanosilver content as compared 

to silver nitrate treated animals in nearly all tissues; nanosilver accumulation was significantly 

higher in liver, spleen, lung, tail (injection site), visceral yolk sac, and endometrium. Nanosilver 

was identified in vesicles in endodermal cells of the visceral yolk sac. This study demonstrated 

that nanosilver distributed to major maternal organs and extra-embryonic tissues, but the authors 

stated that very little silver reached developing embryos and no adverse morphological effects on 

the developing embryos were observed. 

The teratogenicity potential of nanosilver in pregnant rats was investigated by Mahabady et al. 

(2012). Nanosilver of unknown size and surface coating was administered to pregnant rats via 

intraperitoneal injection (i.p.) on GD 8 and 9. Fetuses collected on GD 20 from animals that 

received nanosilver were reported to have reduced weight and length but no effect on the skeletal 

system as compared to animals treated with a saline control. Because Mahabady et al. (2012) did 

not report key information about the nanosilver particles size and surface coatings, it is not 

possible to compare these results to the nanosilver in any other study or in any nanosilver 

containing product. 

4.2.5 Mutagenicity of Nanosilver 

Currently, there are no studies in the scientific literature that investigate the potential of 

nanosilver to cause cancer (i.e., carcinogenicity). However, the potential of nanosilver to induce 

changes in genetic material (i.e., mutagenicity, genotoxicity) has been investigated in vitro using 
15 

traditional mutagenicity tests including the bacterial reverse mutation assay (i.e., Ames test), the 

mouse lymphoma forward mutation test, the mammalian cell chromosome aberration test in 

Chinese hamster ovary cells, and the mouse lymphoma Comet assay for oxidative damage. 

Additionally, nanosilver was investigated in an in vitro human lymphoblastoid micronucleus 

assay as well as and in an in vivo rat bone marrow micronucleus assay. Results from these assays 

are summarized below: 

In vitro reverse gene mutation assay in bacterial cells: The mutagenicity of nanosilver (average 

diameter of 10 nm) suspended in a 1% citric acid solution was determined at concentrations of 

up to 500 μg/plate in bacterial cells using the Ames test (Kim et al., 2012). Although cytotoxicity 

was observed at 31.25 μg/plate, nanosilver did not induce a mutagenic effect in the histidinerequiring strains of Salmonella typhimurium TA98, TA100, TA1538 and TA1537 or in 

tryptophan-requiring Escherichia coli WP2uvrA with or without the metabolic activation system 

(±S9). A similar result was obtained by Li et al.(2012) for nanosilver with an average diameter 

of 5 nm (size range from 4 to 12 nm) prepared in TEM for the primary particles and for particles 

with a diameter of 1608.7 ± 175.4 nm, prepared in culture media. In this test, there was no 

increase in revertant mutant colonies of the standard S. typhimurium tester strains, which 

included the strain used to detect oxidative damage (TA102) up to cytotoxic concentration 

(31.25-62.5 μg/plate -S9; 125-250 μg/plate +S9). However, Li et al. (2012) cautioned that 

because of cytotoxicity and the physical properties of nanosilver, this test system may lack the 

sensitivity required to detect the mutagenic action of the test material. 

In vitro forward gene mutation assay in mouse lymphoma L5178Y/TK+/- cells with a Comet 

Assay: The mutagenicity of nanosilver, which had an average diameter of 5 nm (size range from 

4 to 12 nm) prepared in TEM for the primary particles and a diameter of 1608.7 ± 175.4 nm in 

the culture media, was evaluated in mouse lymphoma L5178Y at the TK+/- locus and the modes 

of action was assessed using standard alkaline and enzyme-modified Comet assays with a gene 

expression analysis (Mei et al., 2012). Nanosilver induced dose-dependent cytotoxicity and 

mutagenicity with a marked increase in the mutation frequency at 4 and 5 μg/mL (<50% cell 

survival at ≥5 μg/mL) where nanosilver had a clastogenic mode of action. Subsequent testing 

revealed no evidence of DNA damage (Comet test) but oxidative damage (modified Comet test), 

confirmed by gene expression analysis, which showed an expression pattern consistent with 

production of reactive oxygen species (ROS). 

In Vitro Chromosome Aberration Test: The clastogenic effect of nanosilver (average diameter of 

10 nm) suspended in a 1% citric acid at concentrations of less than 31.25 μg/mL was determined 

in Chinese hamster ovary cells (CHO-k1) after 6- and 24-hour exposures ±S9 using OECD Test 

Guideline 473 (Kim et al. 2012). Nanosilver did not induce any statistically significant increase 

in the number of cells with chromosome aberration, polyploidy, or endoreduplication when 

compared with the control group at concentrations causing approximately 50% cytotoxicity. 
16 

In Vitro Mammalian Cell Micronucleus Test: An in vitro mammalian cell micronucleus test 

(OECD Test Guideline 487) was used to determine the genotoxicity of nanosilver, which had an 

average diameter of 5 nm (size range from 4 to 12 nm) by TEM for the primary particles and a 

diameter of 1608.7±175.4 nm prepared in culture media. Nanosilver at concentrations of up to 30 

μg/mL induced a significant and dose-related increase in micronuclei in human lymphoblastoid 

TK6 cells, indicating to the investigator that nanosilver has weakly positive genotoxic potential 

(Li et al, 2012). 

In Vivo Micronucleus Assay: In contrast to the above in vitro findings, oral administration of 

carboxymethyl cellulose-coated nanosilver with average diameter of 60 nm (minimum diameter 

of 53 nm and maximum diameter of 71 nm) caused overt toxicity but failed to induce an increase 

in micronucleated polychromatic erythrocytes (MN PCEs) in the bone marrow of male and 

female rats after 28 days of treatment at doses 0, 30, 300, and 1,000 mg/kg/day (Kim, et al, 

2008). This study, which was based on OECD Test Guideline 474, reported that there was no 

statistically significant treatment-related increase in MN PCEs when compared to the negative 

control. This study indicates that nanosilver is neither clastogenic nor aneugenic in vivo, 

although a limitation of this study is that no measurements were performed to determine if 

nanosilver reached the bone marrow. 

Overall the results suggest that of the in vitro tests discussed above, the data from the Ames and 

chromosome aberration assays indicate that nanosilver is not expected to be mutagenic while the 

mammalian cell micronucleus and mouse lymphoma with comet assay suggest that nanosilver 

may have mutagenic potential. This is consistent with a recent review article which stated that in 

vitro data point to possible mutagenic properties of nanosilver (Bartłomiejczyk et al., 2013). 

However, the lack of genotoxicity from the in vivo study suggests that the mutagenic potential 

observed in some of the in vitro studies may be intrinsic to nanoparticles but may not be 

expressed in whole animals or that nanosilver may be subjected to first-pass effects such as 

biliary elimination. At this time, there is inadequate information to assess mutagenic (and hence 

carcinogenic) potential of nanosilver due to differences in results between the in vitro studies and 

in vivo study, and the limitations of the only available in vivo study. 

4.2.6 Silver Ions 

Humans may also be exposed to silver ions that would be released by Nanosilva. Conventional 

silver, and the silver ions it releases, are defined as pesticides under FIFRA. The SAP concluded 

that the hazards of silver ions would be the same, whether they came from conventional silver or 

from silver nanoparticles. With respect to silver ions, the Agency evaluated the toxicity and 

exposure to silver ions and determined that unreasonable adverse effects from use of silver 

containing products are unlikely (U.S. EPA, 1993). 
17 

4.3 Absorption, Distribution, Metabolism, and Excretion (ADME) of 

Nanosilver 

Studies were completed using either injection or oral administration of nanosilver to laboratory 

animals to determine the absorption, distribution, metabolism, and excretion (ADME) of 

nanosilver in whole animals. Based on the studies described in the following sections the Agency 

believes biliary or fecal excretion of nanosilver is the primary elimination pathway. Silver is 

found primarily in the liver, spleen, and kidneys, but also in the thymus, brain, heart, lungs and 

testes of animals dosed with nanosilver, silver nitrate, and silver acetate, where the organs of 

animals dosed with silver nitrate and silver acetate contained greater amounts of silver than in 

animals dosed with nanosilver. Animals can clear silver from blood and most organs given 

enough time but retain silver in the testes and brain. Animals treated with silver nitrate, silver 

acetate, and nanosilver all contain silver granules with dimensions on the nanoscale. However, it 

is unclear if intact nanosilver is absorbed into tissues or if nanosilver dissolves into ionic silver 

before being absorbed into tissues and forming nanoscale granules. 

4.3.1 Injection of Nanosilver 

The translocation, distribution and accumulation of silver after a single subcutaneous injection at 

62.8 mg/kg of body weight of nanosilver with diameters between 50 and 100 nm and microsilver 

with diameters between 2,000 and 20,000 nm was determined in Wistar female rats (n = 30 per 

group) (Tang et al, 2009). The silver content of feces between 2 and 24 weeks after injection was 

significantly higher than in urine for both the nanosilver and microsilver injected animals, which 

suggests that both nanosilver and microsilver were eliminated through biliary excretion. 

Although there was no significant difference between the amount of silver at the injection site or 

in excrements after administration of nanosilver or microsilver, the amount of silver in organs 

was significantly greater for nanosilver. Animals injected with nanosilver were found to contain 

significantly more silver in the liver, kidney, spleen, brain, lung and blood than in animals 

injected with microsilver. Histopathological observations found that nanosilver injected animals 

contained elemental silver spheres that were absent from the microsilver treated animals. The 

elemental silver spheres were observed in different kinds of cells, such as renal tubular epithelial 

cells and hepatic cells. Moreover, these elemental silver spheres also induced blood-brain barrier 

(BBB) destruction and astrocyte swelling, and caused neuronal degeneration. 

The serum kinetics, tissue distribution, and excretion of silver after single injections of 0.5 mg/kg 

and 5 mg/kg of citrate coated nanosilver with average diameter of 7.9±0.95 nm was determined 

in SPF New Zealand White rabbits (n = 4) (Lee et al, 2012). There was no significant general 

toxicity reported in either the 0.5 or 5 mg/kg treatment groups. Accumulation of silver was 

observed in all the tested organs including liver, kidney, spleen, lung, brain, testes, and thymus, 

where the liver and spleen contained the greatest amount of silver. As with the rat study above, 

the amount of silver in feces between 1 and 28 days after injection was significantly greater than 
18 

in urine, which suggests biliary excretion of silver is the major route of elimination after 

injection of nanosilver. 

Blood kinetics, tissue distribution, and organ accumulation of silver after daily intravenous 

injections of between 23.8 and 27.6 mg/L over 5 consecutive days of nanosilver with average 

diameters of 20, 80, and 110 nm was determined in six week old male Wistar rats (n=21 in 

treatment groups; n= 2 in control) (Lankveld et al., 2010). The concentration of silver in blood 

rapidly decreased during the initial 10 minutes following both single and repeated injection of 

nanosilver and then remained stable for up to one hour after injection. Silver was distributed to 

all organs evaluated including the liver, lungs, spleen, brain, heart, kidneys and testes, regardless 

of the size of nanosilver injected. After injecting 20 nm diameter nanosilver, silver was found to 

be distributed mainly in the liver, followed by kidneys and spleen, whereas after injection of the 

80 and 110 nm diameter nanosilver, silver was distributed mainly to spleen followed by liver and 

lung. Thus, there was a size dependent tissue distribution. Repeated administration of nanosilver 

resulted in accumulation of silver in liver, lung and spleen, indicating that these organs may be 

potential target organs for toxicity after repeated exposure. 

4.3.2 Oral Administration of Nanosilver 

The organ distribution and cellular localization of silver was determined following 28 day 

repeated oral administration at 9.0 mg/kg/day of polyvinylpyrrolidone stabilized nanosilver with 

average diameter of 14 ± 4 nm and silver acetate to four week old female Wistar Hannover Galas 

rats (n = 9 for nanosilver and n = 7 for silver acetate) (Loeschner et al., 2011). Although the 

distribution of silver in organs for animals treated with nanosilver and silver acetate was similar, 

the concentration of silver in the organs treated with silver acetate was greater than it was for 

nanosilver treated animals. This was in agreement with the higher fecal excretion of nanosilver 

as compared to silver actetate. Besides the intestinal system, the largest silver concentrations 

were detected in the liver and kidneys; however, silver was also found in the lungs and brain. 

Remarkably, silver containing granules in the same size range as that of the administered 

nanosilver were observed in the ileum and kidney tissues of rats exposed to nanosilver and silver 

acetate. Using transmission electron microscopy (TEM), sulfur and selenium containing granules 

were detected in the ileum of animals exposed to nanosilver and silver acetate, and were mainly 

located in the basal lamina of the ileal epithelium and in lysosomes of macrophages within the 

lamina propria. The results of the present study demonstrate that the organ distribution and form 

of silver was similar when nanosilver or silver acetate were administered orally to rats. 

The toxicokinetics and tissue distribution of silver was determined following 28 day oral gavage 

at 90 mg/kg/day of nanosilver with average diameter of 17.7±3.3 nm by TEM, 

polyvinylpyrrolidone coated nanosilver with average diameter of 12.1±8.0 nm by TEM, and 9 

mg/kg/day of silver nitrate to six-week-old male pathogen free Sprague Dawley rats (n = 5 per 

group) (Van der Zande et al., 2012). Greater than 99% of the silver administered to the rats was 
19 

excreted in their feces indicating that only a small fraction of silver from nanosilver and silver 

nitrate was absorbed. After normalizing for gavage dose, the concentration of silver in the blood 

of the silver nitrate treated animals was significantly higher than for the nanosilver treated 

animals at all time points during exposure. This clearly illustrates a much higher uptake of silver 

when silver nitrate was administered as compared to nanosilver. One day after the oral gavage, a 

significant reduction in blood silver concentration was observed. One week after the oral gavage, 

the concentration of silver in blood was reduced to nondetectable levels indicating rapid 

clearance of silver from the blood for both nanosilver and silver nitrate treated animals. After 

normalizing for gavage dose, silver was observed in all examined organs with the highest levels 

in the liver and spleen where animals treated with silver nitrate had accumulated significantly 

more silver than in animals treated with nanosilver. Silver was cleared from most organs eight 

weeks after the final gavage, but remarkably not from the brain and testes, where between 94 and 

100% of the silver was still present in the brain compared to the amount one day after the last 

gavage of nanosilver and silver nitrate, respectively. Using single-particle inductively coupled 

plasma mass spectrometry; nanosilver was detected one day after the final gavage in the liver, 

spleen, lungs, and gastrointestinal contents of nanosilver gavaged rats. Remarkably, nanosilver 

was also detected in the liver, spleen, lungs, and gastrointestinal contents one day after the final 

gavage from silver nitrate treated rats demonstrating the in vivo formation of nanosilver from 

silver nitrate. Blood enzyme levels were not significantly different from untreated animals 

indicating that there was no acute hepatotoxicity observed. Also, there was no indication that 

nanosilver caused nonspecific immune responses based on immunotoxic responses. 

4.4 Safety Factor for Infants and Children (FQPA Safety Factor) 

Under the Food Quality Protection Act (FQPA), P.L. 104-170, which was enacted in 1996 to 

amend the Federal Insecticide, Fungicide, and Rodenticide Act (FIFRA) and the Federal Food, 

Drug and Cosmetic Act (FFDCA), the Agency was directed to "ensure that there is a reasonable 

certainty that no harm will result to infants and children" from aggregate exposure to a pesticide 

chemical residue. The law further states that in the case of threshold effects, for purposes of 

providing this reasonable certainty of no harm, "an additional tenfold margin of safety for the 

pesticide chemical residue and other sources of exposure shall be applied for infants and children 

to take into account potential pre- and post-natal toxicity and completeness of the data with 

respect to exposure and toxicity to infants and children. Notwithstanding such requirement for an 

additional margin of safety, the Administrator may use a different margin of safety for the 

pesticide residue only if, on the basis of reliable data, such margin will be safe for infants and 

children." 

There is no food use proposed for Nanosilva at this time, and therefore, no FFDCA aggregate 

exposure to Nanosilva residue is required. 
20 

4.5 Antimicrobial Resistance 

Silver is currently used as a broad spectrum antiobiotic in wound dressings. There is a concern 

that increasing use of silver, such as nanosilver for preserving plastics, may result in more 

bacteria developing resistance to silver and limit its use as an antibiotic agent for wound care 

(Gupta and Silver, 1998). In the wound care setting, a recent review by Chopra (2007) concluded 

that the threat of bacterial resistance to silver in the clinical setting is low. However, Chopra 

(2007) cautioned against use of wound dressings that release sublethal levels of silver over a 

long period of time allowing bacteria to develop resistance. 

In terms of environment, a recent study involved releasing 1 mg/L of nanosilver into microcosms 

containing estuary water overlying estuarine sediment cores (Bradford et al., 2009). The study 

found no impact to the microbial community over a 30 day monitoring period. Evidence for 

antibacterial resistance was also evaluated during this study and no increase in antibiotic 

resistance to the bacterial population in the sediment was found (Mühling et al. 2010). Wigginton 

et al. (2010) suggested that the lack of antimicrobial effect in the microcosm was expected given 

that bacterial proteins efficiently bind to nanosilver. 

The Agency concludes that while development of antibacterial resistance due to the use of 

nanosilver in Nanosilva might be possible, the likelihood is low that the levels used in products 

incorporating Nanosilva will lead to the development of silver resistant microbes who will 

spread throughout the environment and results in widespread bacterial resistance to silver. 

4.6 Point of Departure Selections and Target Margin of Exposure 

The currently available oral toxicity studies indicate that nanosilver causes liver and kidney 

toxicity in laboratory animals where silver is distributed to all organs and tissues with 

accumulation of silver in the brain and male animal testes. Inhalation toxicity studies also 

identified liver toxicity as well as lung effects including chronic alveolar inflammation. There 

were potential neurotoxic effects identified with increases in neurotransmitter concentrations and 

loss of spatial cognition; however, these same effects were not observed in a follow-on study. 

EPA believes that these neurotoxicity studies are inadequate in their assessment of behavioral 

effects and do not use optimal methods to evaluate the potential toxicity to the nervous tissue 

structure and function. The in vitro Ames and chromosome aberration assays indicate that 

nanosilver is not expected to be mutagenic while the mammalian cell micronucleus and mouse 

lymphoma with comet assay suggest that nanosilver may have mutagenic potential. However, the 

lack of genotoxicity from the in vivo study indicates that there is inadequate information to 

assess mutagenic (and hence carcinogenic) potential of nanosilver. Finally, there is not enough 

information on the reproductive and developmental toxicity for nanosilver at this time. 

Together, these studies indicate to the Agency that, if sufficient quantities of nanosilver break 

away from Nanosilva, and if such nanosilver displays toxicity similar to the nanosilver used in 
21 

the oral and inhalation route-specific studies, then route-specific exposure to Nanosilva derived 

nanosilver may result in adverse health effects. 

4.6.1 Point of Departures for Nanosilva 

The toxicological point of departure (POD) is the lower confidence bound on the lowest 

experimental dose from a dose response study that showed an effect. This dose is determined 

from dose-response data and marks the beginning of extrapolation to determine the risk 

associated with environmentally relevant human exposures. Commonly, this is a NOAEL from a 

laboratory animal toxicity study, which represents the dose at which no adverse affects were 

observed in laboratory animals. Oral or dermal subchronic and chronic toxicology studies are not 

available for Nanosilva or the nanosilver that might break away from articles incorporating 

Nanosilva. In place of these studies, the Agency is determining NOAELs and LOAELs from 

subchronic inhalation and oral toxicity studies found in the scientific literature for nanosilvers to 

evaluate the effects that could occur from exposure to the nanosilver present in Nanosilva. 

As stated in Section 2.2, the FIFRA SAP was unsupportive of bridging among silver-based 

materials with different properties. However, the SAP indicated that bridging would be 

appropriate for materials of similar size and essentially identical physical properties and that 

bridging between silver ions released from nanosilver and the existing database for silver ions is 

feasible (FIFRA SAP, 2009). The SAP cautioned about extrapolating from one nanosilver 

formulation to another when assessing hazards because differences in particle formulation (e.g., 

coating and inert ingredients) are likely to affect biological activity, among other things. 

The oral toxicity study by Hardrup et al. (2012a) used PVP coated nanosilver with average 

diameter of 14 nm, which is similar to the diameter and surface coating of the nanosilver in 

Nanosilva. However, EPA cannot determine a NOAEL or LOAEL from this study because 

histopathological patterns indicative of distinct adverse effects at the highest nanosilver dose of 9 

mg/kg/day were not identified. The oral toxicity study by Park et al. (2010) used uncoated 

nanosilver with an average diameter of 42 nm, which is different from the diameter and surface 

coating of the nanosilver in Nanosilva. In 2011, EPA used the NOAEL reported in the Park et al. 

(2010) study as the POD for evaluating the oral and dermal toxicity of the nanosilver in HeiQ 

AGS-20 (U.S. EPA, 2011a). EPA reviewed this study again as part of this assessment and now 

concludes that although the results of the Park et al. 2010 study showed evidence of changes in 

clinical chemistry, it lacks histological support for the effects used as the basis for the study 

NOAEL/LOAELs. In the absence of histopathological findings, the clinical chemistry changes 

observed are insufficient evidence of an adverse effect. 

The oral toxicity studies by Kim et al. (2008) and Kim et al. (2010) used CMC coated nanosilver 

with average diameter of 56 and 60 nm, respectively, which is different from the diameter and 

surface coating of the nanosilver in Nanosilva. These studies were stated as being completed 
22 

according to OECD guidelines and identified histopathological patterns in the liver that were 

indicative of distinct adverse effects. EPA has determined that the NOAEL is 30 mg/kg/day from 

the 28-day Kim et al. (2008) study based on significant increases in alkaline phosphatase and 

cholesterol, significant changes in hematology, and accompanied by histopathological evidences 

of liver toxicity (bile-duct hyperplasia around central vein, infiltration of inflammatory cells, and 

dilation of the central vein) at the LOAEL dose of 300 mg/kg/day. A NOAEL for the 90-day 

Kim et al. (2010) cannot be established because of adverse histological patterns evident at the 

lowest dose of 30 mg/kg/day. EPA has determined that the LOAEL from the 90-day Kim et al. 

(2010) study is 30 mg/kg/day based on histopathological evidences of liver toxicity (bile-duct 

hyperplasia with focal, multifocal, or lobular necrosis) in both males and females. 

The inhalation toxicity studies by Sung et al. (2009) and Song et al. (2012) used uncoated 

nanosilver with average diameters of 18 to 19 nm and 14 to 15 nm, respectively, which is similar 

to the diameter and but different than the surface coating of the nanosilver in Nanosilva. EPA has 

determined that the NOAEL is 133 μg/m3

 from the 90-day Sung et al. (2009) study based on the 

minimal bile-duct hyperplasia, perivascular mixed cell infiltrate and chronic alveolar 

inflammation and macrophage accumulation in the lungs in both male and female rats, and 

decreases in physiological measures of lung function (respiratory minute volume, tidal volume, 

and peak inspiratory flow) in the male rats. EPA has determined that the NOAEL is 49 μg/m3

from the 12-week Song et al. (2012) study based on adverse histopathological patterns in lung 

tissue observed at the 117 μg/m3

 dose. In 2011, EPA used the NOAEL reported in the Sung et al. 

(2009) study as the POD for evaluating the inhalation toxicity of the nanosilver in HeiQ AGS-20 

(U.S. EPA, 2011a). EPA compared the Sung et al. (2009) study to the Song et al. (2012) study 

and now concludes that the newly available Song et al. (2012) study is more appropriate for 

evaluating inhalation exposure to nanosilver. 

Based on the above analysis, EPA has determined that the NOAEL of 30 mg/kg/day from 28-day 

oral toxicity study by Kim et al., 2008 is the POD for short-term oral exposures (< 30 days) to 

the nanosilver in Nanosilva (Table 3). EPA has also determined that the LOAEL from the 90 day 

oral toxicity study by Kim et al., 2010 is the POD for intermediate-term oral exposures (1 to 6 

months) to the nanosilver in Nanosilva. EPA has determined that the NOAEL of 49 μg/m3

derived from the Song et al. (2012) study is the POD for short- and intermediate-term inhalation 

exposures to the nanosilver in Nanosilva. These studies were conducted using OECD guidelines 

and the effects are consistent across studies, and therefore the NOAELs/LOAELs are protective 

of effects seen in other studies. 
23 

Table 3 – Toxicity Endpoints for Use in Nanosilver Risk Assessment 

Dietary Risk Assessment – No Dietary Exposures 

Nondietary Risk Assessments 

Exposure 

Scenario 

Point of 

Departure (POD) 

Uncertainty 

Factor 

Target MOE 

for Risk 

Assessment 

Study 

Incidental Oral 

(short-term) 

NOAEL = 30 

mg/kg/day 

UFA = 10 

UFH = 10 

UFD = 10 

Target MOE = 

1000 Kim et al. (2008) 

Incidental Oral 

(Intermediateterm) 

LOAEL = 30 

mg/kg/day 

UFA = 10 

UFH = 10 

UFD = 10 

UFL = 3 

Target MOE = 

3000 Kim et al. (2010) 

Dermal 

(short -term) 

NOAEL = 30 

mg/kg/day 

DAF = 6.7% 

UFA = 10 

UFH = 10 

UFD = 10 

Target MOE = 

1000 Kim et al. (2008) 

Dermal 

(Intermediateterm) 

LOAEL = 30 

mg/kg/day 

DAF = 6.7% 

UFA = 10 

UFH = 10 

UFD = 10 

UFL = 3 

Target MOE = 

3000 Kim et al. (2010) 

Inhalation 

(short- and 

intermediate- term) 

NOAEL = 49 

μg/m3

UFA = 10 

UFH = 10 

UFD = 10 

Target MOE = 

1000 Song et al. (2012) 

Cancer (oral, 

dermal, inhalation) 

At this time there is inadequate information to assess carcinogenic potential of nanosilver due 

to differences in results between the in vitro studies and in vivo study, and the limitations of the 

only available in vivo study. 

There are no acceptable dermal toxicity studies on nanosilver available to the Agency. In the 

absence of any such dermal toxicity studies, the Agency normally uses extrapolation from 

another route of exposure (usually oral) and assumes a default dermal absorption factor (DAF) of 

100% when no in vivo data are available (see Section 4.2.3). The available human in vivo study 

indicating absorption of nanosilver is 6.7% and the in vitro data indicating absorption of 

nanosilver from intact and abraded human skin is substantially below 0.1% provide scientific 

support for setting a conservative DAF of 6.7% for the nanosilver that might break away from 

the Nanosilva. EPA has determined that the dermal toxicity for the nanosilver in Nanosilva will 

be evaluated using the oral POD of 30 mg/kg/day and a DAFs of 6.7%. In 2011, EPA used a 

DAF of 0.1% that was based on human clinical study by Moiemen et al., (2011) for evaluating 

the dermal toxicity of the nanosilver in HeiQ AGS-20 (U.S. EPA, 2011a). EPA reviewed this 

study again as part of this assessment and now concludes that the 0.1% DAF was an 

underestimate because it was based on the Moiemen et al. (2011) study which only reported 

silver concentrations in the blood of patients. The 6.7% DAF is based on the observational study 

by Wan et al. (1991) which reported silver concentration in serum and eliminated through urine. 
24 

There are currently no acceptable studies on the reproductive and developmental toxicity for 

nanosilver. However, there were dose-dependent increases in the concentration of silver in the 

testes of rats after oral ingestion, inhalation and injection of nanosilver and, in another study, 

nanosilver was distributed to major maternal organs and extra-embryonic tissues although no 

adverse morphological effects on the developing embryos were observed. Therefore, EPA 

believes that the oral NOAEL and LOAEL of 30 mg/kg/day and inhalation NOAEL of 49 μg/m3

are expected to be protective of developmental and reproductive effects of nanosilver. 

 

The study completed by Hadrup et al. (2012b), which reported significant increases in 

neurotransmitter concentrations (e.g., dopamine) after oral administration of nanosilver to rats at 

concentrations of up to 9 mg/kg/day, lacks histological support for determining 

NOAEL/LOAELs. The study by Liu et al. (2013) reported no effects on the spatial cognition or 

hippocampal activity of mice after injecting nanosilver into the body cavity (i.e., intraperitoneal 

injection) at concentrations of up to 50 mg/kg. Therefore, EPA believes that the oral NOAEL 

and LOAEL of 30 mg/kg/day are expected to be protective of neurotoxic effects of nanosilver. 

The effects on spatial cognition and hippocampal activity observed by Liu et al. (2012) after 

administering nasal drops containing nanosilver at concentrations of 3 and 30 mg/kg to rats 

suggest possible neurotoxic effects from the inhalation of nanosilver. However, EPA believes 

that the doses used in the Liu et al. (2012) study were greater than the maximum dose used in the 

inhalation toxicity study by Song et al. (2012). Therefore, EPA believes that the inhalation 

NOAEL of 49 μg/m3

 based on the Song et al. (2012) study is expected to be protective of 

neurotoxic effects of nanosilver. 

The available in vitro studies suggest that nanosilver may have mutagenic potential. However, 

the one available in vivo micronucleus assay performed on rats after oral administration of 

nanosilver at concentrations of up to 1,000 mg/kg/day (Kim, et al, 2008) indicated that 

nanosilver is neither clastogenic nor aneugenic in vivo; however, there was no positive proof that 

nanosilver reached the bone marrow in the micronucleus assay. Therefore, EPA believes that the 

oral short-term NOAEL and intermediate-term LOAEL of 30 mg/kg/day are expected to be 

protective of the potential mutagenic effects of nanosilver. 

4.6.2 Target Margin of Exposure 

The target margin of exposure (MOE) is based on uncertainty factors. There are two standard 

uncertainty factors that account for potential interspecies extrapolation and intraspecies variation. 

The first is a 10 fold uncertainty factor (UFA) assigned to account for extrapolation of laboratory 

animal data to humans (interspecies). The second is a 10 fold uncertainty factor (UFH) assigned 

to account for variations in susceptibility within the human population (intraspecies). In addition 

to the two standard uncertainty factors given above, there is a third uncertainty factor (UFD) 
25 

which accounts for the incomplete characterization of nanosilver toxicity (database). In this case, 

the Agency has determined that the inhalation and oral toxicity data for nanosilver is sufficient to 

asses the potential health effects caused by nanosilver released from Nanosilva (Table 3). 

However, as the SAP concluded, there is uncertainty on how particle parameters such as size, 

surface charge and coating affect the toxicity potential of nanosilver. In addition, the database is 

incomplete with respect to reproductive and developmental effects. Therefore, the Agency is 

using the maximum 10 fold database uncertainty factor to cover the missing information on 

reproductive and developmental effects and the potential for differences between the nanosilver 

in the available toxicity data (U.S. EPA, 2002). Thus, the target MOE is: 

Target MOE: 10 (UFA) × 10 (UFH) × 10 (UFD) = 1,000 

The target MOE of 1,000 is for evaluating short-term (< 30 days) and intermediate-term (1 to 6 

months) continuous daily inhalation exposures to Nanosilva because the inhalation POD was 

based on a 90 day duration study by Song et al. (2012). The target MOE of 1,000 is for 

evaluating the short-term exposure (< 30 days) continuous daily oral exposures to Nanosilva 

because the oral POD was based on a 28-day oral toxicity study by Kim et al., 2008. 

The oral POD that was based on the LOAEL from the 90 day oral toxicity study by Kim et al., 

2010 has an additional 3 fold uncertainty factor (UFL) to account for the lack of NOAEL. The 

target MOE for evaluating intermediate-term exposure (1 to 6 months) continuous daily oral 

exposures to Nanosilva is: 

Target MOE (intermediate-term oral): 10 (UFA) × 10 (UFH) × 10 (UFD) × 3(UFL) = 3,000 

The maximum recommended long-term target MOE is 3,000 (U.S. EPA, 2002), which has 

already been established for intermediate-term exposures. Therefore, a long-term target MOE 

cannot be determined from the existing toxicity database. 

4.6.3 Margin of Exposure 

The margin of exposure (MOE) is used to determine if exposure to a chemical can be expected to 

cause an adverse effect. The MOE is calculated by dividing the POD by the estimated daily dose 

to which humans will be exposed as expressed by the following: 

MOE = POD / Daily Dose 

After calculating a MOE from the POD and daily dose, EPA evaluates the risk from exposure to 

a pesticide by comparing the calculated MOE to a target MOE (U.S. EPA, 2002). If a calculated 

MOE is equal to or greater than a target MOE, EPA may conclude that exposure to the pesticide 

is unlikely to pose a risk concern and, therefore, will not cause unreasonable adverse effects for 
26 

that specific exposure scenario. If a calculated MOE is less than a target MOE, then EPA may 

have a risk concern. However, the MOE analysis is not the only factor the Agency uses when 

determining if there is a risk concern from exposure to a pesticide. The Agency also considers 

other scientific evidence in a weight of evidence evaluation such as the severity of toxic effects, 

the controls used to minimize exposures, and the population exposed to the pesticide. A risk 

concern, however, is not the equivalent of a determination that the potential risk constitutes an 

unreasonable adverse effect. Rather, where EPA finds a risk concern, EPA will generally: (1) 

require protective measures or use restrictions to mitigate the risk; (2) further refine its risk 

assessment analysis, particularly if conservative assumptions were used to produce the initial 

assessment; or (3) explicitly analyze any potential benefits of the pesticide to determine whether, 

on balance, those benefits outweigh the identified risk. 

The Agency’s MOE approach for nanosilver uses mass-based metrics, both for determining the 

POD and for calculating exposure. The Agency is aware of the ongoing debate within the 

scientific community that metrics other than mass (such as particle number or surface area) may 

be more suitable for assessing nanosilver risks and therefore acknowledges the potential for 

limitations of mass-based risk estimates. 

 
27 

V. OCCUPATIONAL AND CONSUMER RISK ASSESSMENT 

Based on the proposed use pattern of Nanosilva in plastics and textiles, EPA anticipates humans 

could be exposed to the following substances: 

1) Silver ions released from Nanosilva; 

2) Nanosilva complex; and/or 

3) Nanosilver that might break away from the Nanosilva complex. 

Because the levels of silver in plastics and textiles incorporating Nanosilva are lower than for 

other currently registered products which use silver ions and because the Agency determined that 

unreasonable adverse effects from use of silver containing products are unlikely (U.S. EPA, 

1993), EPA concluded that the risk from exposure to silver ions derived from plastic and textile 

products incorporating Nanosilva is not of concern. Nanosilva LLC has submitted a number of 

studies and other information to support its application for registration, including information 

relevant to assessing the toxicity of and exposure to Nanosilva. In addition, EPA has also 

reviewed data and information on nanosilver from the scientific literature. This section discusses 

EPA’s assessment of the potential risks to human health from the use of Nanosilva. The first 

section addresses potential levels of occupational exposure and risk to workers who handle the 

Nanosilva liquid suspension. The final sections address potential levels of consumer exposure 

and risk to children who come in contact with plastics and textiles incorporating Nanosilva. 

5.1 Occupational Risk Assessment for Mixing and Loading the Nanosilva 

Liquid Suspension 

EPA expects that occupational inhalation and dermal exposures to Nanosilva and the 

nanoparticles that might break away from Nanosilva are likely to occur during the following use 

scenarios:

1. Mixing and loading of Nanosilva during preparation of a master batch 

2. Mixing, loading, and applying the Nanosilva containing master batch during treatment of 

plastics and textiles 

3. Handling plastics and textiles incorporating Nanosilva 

EPA evaluates the risk of occupational exposures from mixing, loading, and applying or 

handling pesticide products. Although exposure to Nanosilva during subsequent work activities 

involving plastics and textiles incorporating Nanosilva can occur in occupational settings (items 

2 and 3), the risk to workers for these scenarios is unlikely to exceed the risk during mixing and 

loading Nanosilva during preparation of the master batch (item 1). Therefore, only the mixing 

and loading of Nanosilva during preparation of a master batch is evaluated in the subsequent 

sections. 
28 

5.1.1 Occupational Exposure when Preparing the Master Batch Containing Nanosilva 

A master batch consists of polymer pellets (i.e., plastic pellets) containing Nanosilva which are 

subsequently used in treating plastics (MRID 48652901). A master batch is prepared by blending 

a polymer and the Nanosilva liquid suspension using a mixer and compounding this mixture 

using an extruder. Workers may be exposed to Nanosilva and nanosilver particles that break 

away from the Nanosilva complex while loading the Nanosilva liquid suspension into the master 

batch mixer. 

Nanosilva LLC did not submit data or information quantifying the amount of Nanosilva that 

workers are exposed to during loading of the Nanosilva liquid suspension into a master batch 

mixer. To calculate worker exposure, the Agency is using the standard occupational handler unit 

exposure values for mixing and loading of liquids, as shown in Table 4, which are from the 

Antimicrobial Exposure Assessment Task Force II (AEATF II) liquid pour human exposure 

monitoring study (U.S. EPA, 2012a) and the Pesticide Handlers Exposure Database (PHED) 

(U.S. EPA, 1998). 

Table 4 – Unit Exposures for Mixing and Loading of Liquids 

Personal Protective Equipment (PPE) 

Level 

Exposure 

Route Unit Exposure Data Source 

No respirator Inhalation 0.21 (μg/m3/lb) AEATF II 

Engineering Control – Closed System 

Loading Inhalation 0.016 (μg/m3

/lb) PHED 

No gloves, use of long-sleeve shirt, long 

pants, shoes plus socks Dermal 10 (mg/lb)A AEATF II 

Gloves, long-sleeve shirt, long pants, 

shoes plus socks Dermal 1.2 (mg/lb)B

 AEATF II 

Engineering Control – Closed System 

Loading Dermal 0.0086 (mg/lb) PHED 

AArithmetic mean for conventional liquid pour scenario 

B

Estimated from the no gloves unit exposure using a glove protection factor of 10. 

The inhalation unit exposure for mixing and loading of liquids using engineering controls (closed 

system loading) as listed in PHED is 0.083 mg/lb. This value was calculated based on the air 

concentrations measured in workers’ breathing zone during the handling of a pesticide, the time 

that elapsed during the sampling, an assumed breathing rate of 29 liters per minute, and the 

amount of the pesticide that was handled during the sampling period. When the PHED unit 
29 

exposures were initially calculated, they were expressed in units of mg/lb for comparisons to 

PODs that were expressed in units of mg/kg/day. Because the POD for inhalation exposure to 

nanosilver is expressed as a daily-average air concentration of μg/m3

, the PHED unit exposure 

was converted to a daily-average air concentration with units of μg/m3 per lb of pesticide handled 

per day. This was done by dividing the air concentrations listed in the PHED by the amount of 

pesticide handled while accounting for the time elapsed during sampling and an eight hour work 

day. This yielded an inhalation unit exposure of 0.016 μg/m3 per lb of pesticide and it represents 

an eight hour time weighted average (Table 4). This unit exposure is the arithmetic mean of 27 

PHED monitoring events (formerly called replicates) that represent inhalation exposure during 

the mixing and loading of pesticide containing liquids while using closed-system loading. 

5.1.2 Occupational Margins of Exposure 

As detailed in the following sections, EPA evaluated the risk to workers who load the Nanosilva 

liquid suspension into a master batch mixer. 

Occupational Inhalation Daily-Exposure and MOEs

The inhalation exposure to the nanosilver in the Nanosilva liquid suspension was calculated 

using the following: 

Inhalation Daily Exposure = Amount of nanosilver in Nanosilva handled per day × 

Inhalation Unit Exposure 

Where: 

 Amount of nanosilver in Nanosilva handled per day is 1.2 lb/day. This value was 

calculated assuming that the Nanosilva liquid suspension was mixed into a low density 

polyethylene plastic using an extruder with the maximum screw diameter of 8 inches 

(Chanda and Roy, 2007) which yields a plastic production rate of 5,000 lb/hour (Flite 

Technology, 2012). Assuming an 8 hour work day yields a plastics production volume of 

40,000 lb per day at each treatment facility. The maximum amount of nanosilver applied 

to plastics or textiles is 0.003% by weight. Thus, the amount of nanosilver that is handled 

per day is 40,000 lb/day × 0.003% ÷ 100% = 1.2 lb/day. 

 The unit exposure for inhalation is 0.21 μg/m3

/lb when no respirator is worn and 0.016 

μg/m3

/lb when closed-system loading is used (see Table 4). 
30 

Table 5 – Occupational Handlers Inhalation MOEs for the Nanosilver in the Nanosilva 

Liquid Suspension 

Mix/Load 

Nanosilva 

Liquid 

Suspension

Amount of 

Plastics 

Treated per 

day 

(lb)

Application 

RateA

(by wieght)

Amount of 

Nanosilver 

Handled 

per day 

(lb)

Inhalation 

Unit 

Exposure

(μg/m3/lb)

Daily 

ExposureC

(μg/m3)

MOED

Without 

Respirator 40,000 0.003% 1.2 0.21 0.25 200

Closed System 

Loading 

(Without 

Respirator) 

40,000 0.003% 1.2 0.016B

 0.02 2,500 

A. Maximum amount of nanosilver in plastics incorporating Nanosilva 

B. PHED unit exposure value converted to air concentration units based on mean 8 hour TWA 

C. Daily Exposure = Amount nanosilver Handled × Unit Exposure 

D. MOE = POD / Daily Exposure (rounded to two significant digits) 

The MOEs shown in Table 5 were calculated by dividing the inhalation exposure for nanosilver 

by the POD of 49 μg/m3

, which is the NOAEL from a 90-day inhalation toxicity study for 

nanosilver (Song et al., 2012). This assumes that all the nanosilver in Nanosilva is freely 

available after inhalation and behaves like the nanosilver used in the Song et al. (2012) study 

even though the nanosilver in Nanosilva is attached to a silica core particle through a thiolate 

bond and is covered with sulfur and polyvinylpyrrolidone. When no respirator is worn the MOE 

is 200, which is based on the AEATF II unit exposure data, and is less than the target MOE of 

1,000 indicating the risk from short- and intermediate-term exposures is of concern. The MOE is 

2,500 for workers who use closed-system loading when mixing and loading the Nanosilva 

suspension, which is based on PHED unit exposure data, and is greater than the target MOE of 

1,000 indicating the risk from short- and intermediate-term exposures is not of concern. These 

MOEs indicate that the risk concern for inhalation exposures to the nanosilver in Nanosilva is 

mitigated when workers use closed-system loading during mixing and loading of the Nanosilva 

liquid suspension. 

Occupational Dermal Daily-Dose and MOEs

The dermal dose from exposure to the nanosilver in the Nanosilva liquid suspension was 

calculated using the following: 

Dermal Daily Dose = (Amount of nanosilver in Nanosilva handled per day × Dermal Unit Exposure 

× Dermal Absorption Factor) / Body Weight 

Where: 

 Amount of nanosilver in Nanosilva handled per day is 1.2 lb/day (Table 5). 
31 

 The dermal unit exposures are 10 mg/lb when no gloves are worn, 1.2 mg/lb when gloves 

are worn, and 0.0086 mg/lb when closed-system loading is used (Table 4). 

 The dermal absorption factor (DAF) for nanosilver is 6.7% (see Section 4.2.3). 

 The body weight of an adult is 80 kg (U.S. EPA, 2011b). 

Table 6 – Occupational Handlers Dermal MOEs for the Nanosilver in the Nanosilva Liquid 

Suspension 

Mix/Load 

Nanosilva 

Liquid 

Suspension 

Amount of 

Nanosilver 

Added or 

HandledA

(lb) 

Dermal 

Unit 

Exposure 

(mg/lb) 

ExposureB

(mg/day) 

Daily DoseC

(mg/kg/day) 

MOED

Without 

Gloves 1.2 10 12 0.010 3,000 

With Gloves 1.2 1.2 1.4 0.0012 25,000 

Closed System 

Loading 1.2 0.0086 0.01 0.0000086 >1,000,000 

A. Based on the same assumptions as those used for the inhalation MOE (see Table 5) 

B. Dermal Exposure = Amount of nanosilver Handled × Dermal Unit Exposure 

C. Dermal Dose = (Dermal Exposure × Dermal Absorption Factor)/ Body Weight 

D. Dermal MOE = POD / Daily Dose (rounded to two significant digits) 

The MOEs shown in Table 6 were calculated by dividing the dermal dose by the POD of 30 

mg/kg/day, which is the NOAEL from a 28-day oral toxicity study (Kim et al., 2008) and the 

LOAEL from a 90 day oral toxicity study (Kim et al., 2010). This assumes that all the nanosilver 

in Nanosilva is freely available after dermal contact and behaves like the nanosilver used in the 

Kim et al. (2008 and 2010) studies even though the nanosilver in Nanosilva is attached to a silica 

core particle through a thiolate bond and is covered with sulfur and polyvinylpyrrolidone. The 

MOEs for dermal exposure are 3,000 when no gloves are worn and 25,000 when gloves are 

worn, which are based on the AEATF II unit exposure data, and are both greater than the target 

MOE of 3,000 indicating that the risk from short- and intermediate-term occupational dermal 

exposures are not of concern. The MOE is greater than 1,000,000 when closed-system loading is 

used, which is based on the PHED unit exposure data, and is greater than greater than the target 

MOE of 3,000 indicating that the risk from short- and intermediate-term occupational dermal 

exposures are not of concern. 
32 

5.1.3 Conclusions for Occupational Risk 

The MOE for occupational inhalation exposure to the nanosilver in the Nanosilva liquid 

suspension is 2,500 if workers use close-system loading when mixing and loading the Nanosilva 

liquid suspension, which is greater than the target MOE of 1,000 indicating the risk from shortand intermediate-term inhalation exposures is not of concern. The MOE for dermal exposure to 

the nanosilver in the Nanosilva liquid suspension is greater than 1,000,000 if workers use closedsystem loading when mixing and loading the Nanosilva liquid suspension, which is greater than 

the target MOE of 3,000 indicating that the risk from short- and intermediate-term dermal 

exposures is not of concern. Therefore, EPA proposes to require that closed-system loading as 

defined in 40 CFR part 170.240(d)(4) be used when mixing and loading the Nanosilva liquid 

suspension. The proposal to require closed-system loading is based on unit exposure data from 

PHED. The MOEs calculated using unit exposure data from AEATF II were shown for 

comparison purposes only and were not relied on to determine that Nanosilva will not cause 

unreasonable adverse effects to human health. 

EPA does not typically consider long-term occupational exposures to materials preservatives 

used to treat plastics or textiles because application of these chemicals does not typically occur 

on a daily basis for more than 6 months. 

There are several uncertainties in the occupational risk assessment. The exposure analysis 

assumes that all the nanosilver in Nanosilva is freely available even though the nanosilver in 

Nanosilva is attached to a silica core particle through a thiolate bond and the nanosilver in 

Nanosilva is covered with sulfur and polyvinylpyrrolidone. The PODs used were based on 

inhalation and oral toxicity studies completed using nanosilver, not Nanosilva or the nanosilver 

that might break away from Nanosilva, and thus these PODs may not equal PODs determined 

from testing conducted with Nanosilva or the nanosilver found in Nanosilva. Because the 

nanosilver in the toxicity database is different from the nanosilver in Nanosilva and because the 

database is incomplete with respect to reproductive and developmental effects, the Agency used 

a maximum 10-fold database uncertainty factor when evaluating the risk from occupational 

exposure to Nanosilva. There are also uncertainties in extrapolating from effects observed after 

feeding test-animals nanosilver (i.e., oral route) to the effects that might be observed after 

applying nanosilver to the skin of test animals (i.e., dermal route). 

5.1.4 Occupational Exposure and Health Data Requirements 

EPA is not proposing to require that Nanosilva LLC conduct an indoor applicator study to 

quantify the unit exposure values during mixing and loading of the Nanosilva liquid suspension 

into the master batch mixer. This is because closed-system loading, such as lock and load 

containers, where the Nanosilva liquid suspension is transferred to a plastic extruder through 

piping minimizes occupational exposure to Nanosilva to the greatest extent possible. 
33 

Although there is no risk concern for workers who use close-system loading when mixing and 

loading the Nanosilva liquid suspension, EPA is proposing to require an inhalation route-specific 

subchronic test (OCSPP 870.3465) to confirm the Agency’s determination that the risks to 

workers who mix and load Nanosilva are not unreasonable. Also, because workers of child 

bearing age could handle Nanosilva during mixing and loading, EPA is proposing to require a 

modified reproduction/developmental toxicity screening test since this data is currently missing 

from the toxicity database. In sum, EPA is proposing to require these tests to confirm the 

adequacy of the 10 fold database uncertainty factor, to reduce the uncertainties related to 

differences in the physical properties of the nanosilver, and because there are currently no 

acceptable studies on the reproductive and developmental toxicity for nanosilver. 

 In summary, the following guideline studies are proposed to be required for Nanosilva: 

 90-Day Inhalation Toxicity (Rat) (OCSPP 870.3465) modified to include in vivo bone 

marrow assay and functional observational battery, motor activity and detailed 

neuropathology 

 Reproduction/Developmental Toxicity Screening Test (Modified OCSPP 870.3550/ 

OECD TG 421) 

5.2 Consumer Risk Assessment for Products Incorporating Nanosilva 

EPA expects consumer exposures to Nanosilva, the silver ions derived from the Nanosilva 

complex, and the nanoparticles that break away from the Nanosilva complex could potentially 

occur during the following use scenarios: 

1. Incidental oral exposure to plastics and textiles incorporating Nanosilva 

2. Dermal exposure to plastics and textiles incorporating Nanosilva 

Nanosilva is proposed to be mixed into polymer and polymer based products to suppress the 

growth of bacterial, algae, fungus, mold and mildew, which cause odors, discoloration, stains, 

and deterioration of plastics and textiles. Because plastics incorporating Nanosilva could be 

subsequently used to manufacture children’s toys and textiles worn by children, it is assumed 

that children will be exposed to products containing Nanosilva. 

While children younger than 6 months may potentially be exposed to plastics and textiles 

incorporating Nanosilva, it is believed that exposure for children older than 6 months will be 

equivalent, if not greater, due to behavioral and anatomical/physiological development; 

therefore, EPA assessed exposure to children older than 6 months. EPA’s recently revised its 

standard operating procedures (SOPs) for residential pesticide exposure assessments (U.S. EPA, 

2012b). Based on the combined quantitative and qualitative analysis of the index lifestage, the 

Agency has determined that the 1 < 2 year old lifestage represents the most appropriate index 
34 

lifestage for children for most exposure scenarios. In 2011, EPA evaluated the exposure to 

textiles containing the nanosilver based pesticide HeiQ AGS-20 using children in the 2 to 3 year 

old lifestage based on the Agency’s determination, at that time, that the 2 to 3 year old child was 

likely the most vulnerable subpopulation from chewing on and wearing textiles treated with 

HeiQ AGS-20 (U.S. EPA, 2011a). After issuing the registration decision for HeiQ AGS-20, 

questions were raised about the use of the 2 to 3 year old lifestage. In light of those questions and 

in light of the newly revised SOPs, in this case, EPA is evaluating exposure to children in the 6 

to < 12 month, the 1 to < 2 year, and 2 to <3 year old age range from plastic toys and flooring 

and textiles incorporating Nanosilva for transparency. 

5.2.1 Consumer Exposures to Plastics and Textiles Incorporating Nanosilva 

Leaching studies are required to determine the amount and form of silver that consumers will be 

exposed to when in contact with plastics and textiles incorporating nanosilver. These studies 

typically involve immersing products incorporating nanosilver in biological fluids such as 

simulated saliva solutions for extended periods of time at physiological temperatures (i.e., 98.6 

degrees Fahrenheit or 37 degrees Celsius) and measuring the amount and form of silver released 

to those fluids. Such “leaching studies” were submitted by Nanosilva LLC using food contact 

solutions for plastic coupons incorporating Nanosilva and using laundry detergent and simulated 

saliva for shirts incorporating Nanosilva. 

Plastics Leaching Study

Nanosilva LLC submitted a leaching study on the migration of Nanosilva from a linear lowdensity polyethylene (LLDPE) plastic (MRID 478289-25). The study was based on Food and 

Drug Administration methodology to determine the migration of chemicals from plastics which 

contact food. The leaching study involved 44 LLDPE coupons with 14 being ashed to determine 

silver content and 28 used in the leaching tests. The 28 plastic coupons were prepared with four 

Nanosilva master batch concentrations where seven coupons each contained 0, 2.5, 5, and 10%, 

respectively. These master batch concentrations corresponded to nanosilver concentrations of 0, 

5, 10, and 20 mg/kg (supplementary information provided on 17 December 2010). The coupons 

each had a surface area of approximately 112 cm2

, a volume of approximately 16.3 mL and an 

approximate mass of 15g. Each coupon was exposed to 50 mL of the fluids listed in Table 7 at 

temperatures of 40 and 100 degrees Celsius for periods of up to 240 hours (10 days). The amount 

of nanosilver present in the coupons was 0, 75, 150, and 300 μg, respectively. Given that 

coupons were in contact with 50 mL of fluid, the maximum silver concentrations that could be 

obtained from each coupon was 0, 1500, 3000, and 6000 μg/L, respectively. 

In general, the concentration of silver in the fluids contacting plastic incorporating Nanosilva 

was low with most values below the analytical detection limit. The analysis method, inductively 

coupled plasma atomic emission spectroscopy (ICP-AES), employed an acid digestion and thus 

the silver concentrations are for total silver content. The Nanosilva LLC leaching study reported 
35 

method detection limits that ranged from 0.8 to 2 μg/L, with the exception of the Oil trial where 

the detection limit was 240 μg/L for the times of less than 240 hours. However, because there 

was no method detection limit study provided to support these values, EPA is using the silver 

detection limit of 5 μg/L, which is the value reported as the estimated detection limit in U.S. 

EPA Method 6010C (U.S. EPA, 2007a) and in the Nanosilva LLC submitted leaching study 

(Table 1, Page 30). 

Table 7 – Concentration of Silver in Fluid Contacting the Nanosilva Containing Plastic 

Coupons at 40 degrees Celsius 

Fluid 

Maximum Silver 

Concentration Detected 

(μg/L)

Silver Matrix Spike 

Recovery (%) 

Low pH, pH 2.0 using 10% nitric acid 

in water 15 of 15 samples < 5 μg/L 93 to 110 

High pH, pH 8.0 using 0.1N sodium 

hydroxide in water 15 of 15 samples < 5 μg/L 100 

Alcohol, 10% ethanol in water 

14 of 15 samples < 5 μg/L 

(11 μg/L for the 5% coupon after 

24 hr) 

34 to 120 

Salt, 10% sodium phosphate 

monobasic in water 

13 of 15 samples < 5 μg/L 

(9.9 μg/L for the 10% coupon 

after 24 hr) 

80 to 174 

Oil, 10% olive oil in water 12 of 12 samples < 240 μg/L 

3 of 3 samples < 5 μg/L 5.7 to 7.8 

Salt and Sugar, 10% sodium phosphate 

monobasic and 10% sucrose in water 

30 of 30 samples < 5 μg/L 

(14 μg/L for the 0% coupon after 

168 hr at 40°C) 

13.6 to 92 

Salt and Sugar after scrubbing the 

plastic coupon 

 13 of 15 samples < 5 μg/L 

(27 μg/L for the 5% coupon after 

48 hr) 

3.6 to 6.8 

Given that EPA is evaluating human exposure to plastics incorporating Nanosilva which occur at 

physiological temperatures (i.e., 98.6 degrees Fahrenheit or 37 degrees Celsius), the leaching 

results for the trials conducted at 100 degrees Celsius are not addressed here. The 100 degrees 

Celsius conditions are more representative of microwaving plastics than of contact with humans. 

Silver was found above the detection limit during 4 out of the 105 coupon leaching trials 

conducted at 40 degrees Celsius where the maximum silver concentration of 27 μg/L was found 

after 48 hours in a plastic coupon containing 5% Nanosilva master batch (10 mg/kg nanosilver) 

(Table 7). A value of 27 μg/L represents 0.9% of the maximum concentration of silver that could 

have leached from the 5% Nanosilva containing coupon. However, there was no clear 
36 

correlation between Nanosilva content and the duration or temperature of each experimental 

trial. In one analysis, silver was found in the salt and sugar fluid after 168 hours at 40 degrees 

Celsius in a coupon that did not contain Nanosilva (0% nanosilver) suggesting that the detections 

of silver may have been due to contamination from sources of silver other than from Nanosilva. 

Interpretation of these results is further complicated by the wide range in silver matrix spike 

recoveries determined for the alcohol, salt, oil, and salt and sugar fluids. This range in silver 

matrix spike recoveries may indicate that nanosilver released from the coupons was not 

detectable by the analytical method. 

The average concentration of silver was 3.3 μg/L, as calculated for the trials conducted at 40 

degrees Celsius. This was based on the concentrations reported and assuming that all results 

reported as being below the detection limit had a value of one-half the detection limit (i.e., 2.5 

μg/L), which is consistent with how EPA assigns values to non-detect results when evaluating 

pesticide residues in food (US EPA, 2000). This average excludes the concentrations reported for 

the oil trial because the detection limit for this trial was reported as 240 μg/L. The average 

concentration of 3.3 μg/L was determined for conditions where the plastic coupon incorporating 

Nanosilva was in contact with fluids having low and high pH, high salt and sugar content, and 

after mechanical abrasion of plastic incorporating Nanosilva. Thus, this average concentration 

represents a range of conditions that are expected to occur during residential use of plastics 

incorporating Nanosilva. However, EPA is only evaluating risk from human contact with plastics 

incorporating Nanosilva, and therefore, only the results for low and high pH fluids, which are 

more representative of biological fluids, are included in this assessment. The concentration of 

silver in the low and high pH fluids were all below the analytical detection limit of 5 μg/L 

meaning that EPA used 2.5 μg/L as the concentration of silver released from plastics 

incorporating Nanosilva. Since this value is below the detection limit, the form of silver is 

unknown and was assumed to be in the form of nanosilver as found in Nanosilva. Using the 

assumed concentration of 2.5 μg/L equates to silver releases of 0.17, 0.08, 0.042% from the 

coupons containing 5, 10, and 20 mg/kg of nanosilver, respectively. Nanosilva LLC is proposing 

that plastic products incorporating Nanosilva contain a maximum of 0.003% nanosilver by 

weight or 30 mg/kg of nanosilver. Assuming that a leaching study conducted using a coupon 

containing 30 mg/kg of nanosilver would also result in silver releases below the detection limit 

would result in a silver release rate of 0.028% for a coupon containing 30 mg/kg of nanosilver. 

However, Nanosilva LLC did not submit leaching results for coupons containing 30 mg/kg of 

nanosilver. 

The leaching study submitted by Nanosilva LLC for plastic coupons incorporating Nanosilva 

was initially determined to be unacceptable by EPA because less than 0.8% of the silver in the 

plastic coupons was recovered during the ashing process, the wide range in silver matrix-spike 

recoveries, and the lack of validation data for the silver analysis. Nanosilva LLC subsequently 

completed an additional study to determine the silver content of identically prepared plastic 
37 

coupons (MRID 48652901). As per EPA recommendation, Nanosilva LLC employed the nondestructive Neutron Activation Analysis (NAA) to determine the silver content of the Nanosilva 

liquid suspension, a master batch containing 2% Nanosilva, and plastic coupons containing 5% 

of the Nanosilva master batch. Using NAA, Nanosilva LLC determined the amount of silver in 

coupons was within 99% of the amount added. 

The available leaching study for plastic coupons containing Nanosilva was conducted using a 

protocol that was not reviewed by EPA prior to conducting the study. The leaching study did not 

employ physiological fluids such as artificial saliva, however, the study was completed at the 

physiological temperature of 40 degrees Celsius. The coupons tested contained nanosilver at 

concentrations of 5, 10, and 20 mg/kg, which are less than the maximum nanosilver 

concentration of 30 mg/kg that Nanosilva LLC is proposing for the Nanosilva pesticide label. 

Even though these issues preclude EPA from accepting the leaching study as final, the study 

does provide a reasonable first estimate for the amount of silver transferred to the mouth while 

mouthing and transferred to skin while contacting plastics incorporating Nanosilva. 

Textile Leaching Study

Nanosilva LLC prepared and submitted a draft protocol titled “The Quantification and 

Characterization of Silver Released from Textiles Treated with Nanosilva (NSPW-L30) as a 

Results of Washing” with the stated purpose to quantify and characterize silver released from 

textiles incorporating Nanosilva during laundering. The protocol was based on methods 

developed by Geranio et al. (2009) and Lorenz et al. (2012) who modified the ISO Colour 

Fastness test (ISO, 1997) to determine the amount and form of silver released to the environment 

when washing textiles containing silver and nanosilver. Because EPA must also evaluate the 

release of nanosilver when children chew and mouth textiles incorporating Nanosilva, EPA 

recommended including an additional test using simulated human saliva. EPA does not 

anticipate dermal exposure will result in a risk concern for textiles incorporating Nanosilva, and 

therefore a separate leaching study involving simulated human sweat was not recommended. 

EPA will use the results of the ISO Colour Fastness test completed with simulated human saliva 

to evaluate the dermal exposure to nanosilver from textiles incorporating Nanosilva. 

Nanosilva LLC completed and submitted the modified ISO Colour Fastness test (MRID 

49045301) using shirts incorporating Nanosilva. The washing tests were conducted on shirts 

composed of polyethylene terephthalate (PET), which incorporated Nanosilva into the PET yarn 

for a final nanosilver content of 0.00262% or 26.2 mg/kg per shirt. For the detergent wash test, a 

section was cut from each of three shirts and washed separately in 150 mL of distilled water with 

commercial laundry detergent and 10 hard rubber balls with diameter of 10 mm for 30 minutes at 

40 degrees Celsius. The detergent wash was followed by two rinse cycles with deionized water. 

The concentration of silver in the wash and rinse water were determined after filtering samples 

through a 0.45 μm pore size filter using inductively coupled plasma mass spectrometry (ICP-
38 

MS). The concentration of silver retained on the filter was also determined by ICP-MS. These 

samples were acid digested prior to analysis and thus the silver concentrations are for total silver 

content and do not distinguish between silver ions, nanosilver, or the Nanosilva complex. 

Although the Nanosilva wash test study reported an ICP-MS detection method of 0.0094 μg/L 

for aqueous samples and 0.94 μg/kg for filter samples, EPA determined that the detection limit 

for aqueous samples was 10 μg/L and 1,000 μg/kg for solids samples. EPA determined these 

detection limits based on a statistical analysis of seven ICP-MS calibration curves and the 

analysis results from 100 μg/L quality control samples. The concentration of silver in the wash 

and rinse water from the detergent wash test for three shirt sections was below the analytical 

detection limit of 10 μg/L (Table 8). The concentration of silver retained on 0.45 μm pore size 

filters was also less than the analytical detection limit of 1,000 μg/kg. 

Table 8 – Concentration of Silver Released from Shirts Incorporating Nanosilva during 

Modified ISO Colour Fastness Test 

Number of 

Shirt Sections Wash Medium 

Concentration of Silver 

Wash/Rinse 

Water (μg/L) 

0.45 μm Pore Size Filter 

(Particles with Diameters 

>0.45 μm) 

(μg/kg) 

3 Distilled Water 

with Detergent < 10 < 1,000 

9 Simulated Human 

Saliva < 10 < 1,000 

A separate study was conducted using three sections from each of three shirts and washed 

separately in 150 mL of simulated human saliva with 10 hard rubber balls with diameter of 10 

mm for 45 minutes at 40 degrees Celsius. The concentration of silver in the simulated saliva was 

determined after filtering through a 0.45 μm pore size filter using ICP-MS where the 

concentration of silver in the saliva for nine shirt sections was below the analytical detection 

limit of 10 μg/L (Table 8). The concentration of silver retained on filters was also less than the 

analytical detection limit of 1,000 μg/kg. 

Given that none of the silver concentrations in detergent or saliva solutions were above 10 μg/L 

and none of the filters contained silver at a concentration above 1000 μg/kg, EPA evaluated the 

amount of silver released from textiles incorporating Nanosilva by replacing the non-detect 

values with half the detection limit, 5 μg/L for liquid samples and 500 μg/kg for solids samples 

(U.S. EPA, 2000). The amount of silver released from textiles was calculated based on the 

volume of detergent and rinse water, and saliva along with the mass of the 0.45 μm filters used 

for each test (Table 9). 
39 

Table 9 – Amount of Silver Released from Shirts Incorporating Nanosilva 

Wash 

Medium Volume/Mass Concentration Amount of Silver (μg) 

Total Silver Potentially 

Released 

(μg) % 

Distilled 

Water with 

Detergent 

190 mL 

wash/rinse 

water 

5 μg/LB

 0.95 

1.05 1.6 

0.2 g filtersA 500 μg/kgB 0.1 

Simulated 

Human Saliva 

150 mL Saliva 5 μg/L 0.75 

0.81 0.9 

0.12 g filter 500 μg/kg 0.06 

AThere was a 0.1 g filter used for the detergent wash and another 0.1 g filter for the rinse. 

B

 Set to half the detection limit 

The initial amount of silver in textiles washed with distilled water and detergent was 66.54 μg, 

therefore the amount of silver released during the detergent wash was: 

Silver released in detergent wash = ଵ.଴ହஜ௚

଺଺.ହସ ஜ௚ ൈ 100% ൌ 1.6%

The initial amount of silver in textiles washed with simulated human saliva was 94.9 μg, 

therefore the amount of silver released during the saliva wash was: 

Silver released in saliva wash = ଴.଼ଵஜ௚

ଽସ.ଽ ஜ௚ ൈ 100% ൌ 0.9%

The value of 1.6% will be used in evaluating releases to the environment from wash water and 

the value of 0.9% will be used in calculating oral and dermal exposures to textiles incorporating 

Nanosilva. Since these releases were determined using concentrations that were below the ICPMS detection limit, the form of silver is unknown. EPA will assume the form of silver is 

identical to the nanosilver present in Nanosilva in the absence of further information. 

The results of these studies demonstrate that PET shirts which incorporate Nanosilva at 26.2 

μg/kg of nanosilver do not release silver at concentrations above the analytical detection limit. 

The ISO Colour Fastness test is thought to represent aggressive washing conditions with one 

wash cycle representing up to five domestic or commercial laundering cycles when the multiple 

test is employed. The amount of silver released during one ISO Colour Fastness test is believed 
40 

to exceed the daily exposure to nanosilver from a treated textile because the ISO Colour Fastness 

test involves immersing the textile in water containing detergents or simulated human saliva and 

hard rubber balls followed by mechanical agitation for 30 to 45 minutes. Thus, results from 

studies which are based on the ISO Colour Fastness test will be used to determine the daily dose 

of nanosilver for children who chew and mouth, adults who wear, and workers who manufacture 

items from nanosilver treated textiles even though this likely overestimates the daily dose of 

nanosilver. 

5.2.2 Consumer Margins of Exposure to Plastics Incorporating Nanosilva 

EPA expects that consumers will be exposed to plastics incorporating Nanosilva by the routes of 

incidental oral and dermal exposures. As detailed in the following sections, EPA evaluated the 

risk to children, as the likely most vulnerable subpopulation, from mouthing toys, and mouthing 

and crawling on flooring incorporating Nanosilva. 

Consumer Incidental Oral Daily-Dose and MOE for Toys Incorporating Nanosilva

Incidental oral exposures to toys incorporating Nanosilva were calculated using the following: 

Incidental Oral Exposure = Surface Residue (mg/cm2

) × Saliva Extraction Efficiency (%) × 

Surface Area of Toy Mouthed per day (cm2

) 

Where: 

 Surface area of a toy mouthed per day is 800 cm2

 for 6 to < 12 month old children, 560 

cm

2

 for 1 to < 2 year old children, and 396 cm2 for 2 to < 3 year old children. These 

values were calculated assuming that a child mouths an area of 10 cm2

 each time they 

contact a toy, that 6 to < 12 month old children contact objects 20 times per hour, 1 to < 2 

year old children contact objects 14 times per hour, and 2 to < 3 year old children contact 

objects 9.9 times per hour (U.S. EPA, 2011a) for 4 hours per day. 

The concentration of silver detected in low and high pH fluids, which are more representative of 

biological fluids, contacting Nanosilva containing coupons represents both the surface residue 

and the saliva extraction efficiency. The concentration of silver in the low and high pH fluids 

were all below the analytical detection limit of 5 μg/L for coupons containing 5, 10, and 20 

mg/kg of Nanosilva. EPA is assuming that non-detectable concentrations of silver would be 

found in a similar test conducted with coupons containing 30 mg/kg of Nanosilva. EPA is using 

2.5 μg/L, which is one-half the detection limit of 5 μg/L, as the concentration of silver released 

from plastic incorporating Nanosilva. Using the concentration of silver, the volume of the 

leachate (50 mL), and the surface area of the test coupons, the amount of available nanosilver at 

the coupon surface is: 

Surface residue and saliva extraction: ଴.଴଴ଶହ ௠௚

௅

ൈ

ହ଴ ௠௅

ൈ

௅

ଵ଴଴଴ ௠௅

ൈ

ଵଵଶ ௖௠మ ൌ ଴.଴଴଴଴଴ଵଵଶ௠௚

௖௠మ
41 

The incidental oral exposure for toys incorporating Nanosilva is: 

Incidental oral exposure for 6 to < 12 month old children: ଵ.ଵଶൈଵ଴షల ௠௚ ௡௔௡௢௦௜௟௩௘௥

௖௠మ ൈ

଼଴଴ ௖௠మ

ௗ௔௬ ൌ

଴.଴଴଴଼ଽଷ ௠௚ ௡௔௡௢௦௜௟௩௘௥

ௗ௔௬

Incidental oral exposure for 1 to < 2 year old children: ଵ.ଵଶൈଵ଴షల ௠௚ ௡௔௡௢௦௜௟௩௘௥

௖௠మ ൈ

ହ଺଴ ௖௠మ

ௗ௔௬ ൌ

଴.଴଴଴଺ଶହ ௠௚ ௡௔௡௢௦௜௟௩௘௥

ௗ௔௬

Incidental oral exposure for 2 to < 3 year old children: ଵ.ଵଶൈଵ଴షల ௠௚ ௡௔௡௢௦௜௟௩௘௥

௖௠మ ൈ

ଷଽ଺ ௖௠మ

ௗ௔௬ ൌ

଴.଴଴଴ସସଶ ௠௚ ௡௔௡௢௦௜௟௩௘௥

ௗ௔௬

The incidental oral daily-dose was calculated from the incidental oral exposure using the 

following: 

Incidental Oral Daily Dose = Exposure / Body Weight 

Where: 

 Exposure is determined in the calculations above. 

 The body weight of a child with age between 6 and < 12 months is 9.2 kg, between 1 and 

< 2 years is 11.4 kg, between 2 and < 3 years is 13.8 (U.S. EPA, 2011b). 

Table 10 – Incidental Oral MOEs for Children Exposed to Plastic Toys Incorporating 

Nanosilva 

Age Range of Child ExposureA

(mg/day) 

Daily DoseB

(mg/kg/day) 

MOEC 

6 to < 12 months 8.93×10-4 9.7×10-5 310,000 

1 to < 2 years 6.25×10-4 5.5×10-5 550,000

2 to < 3 years 4.42×10-4 3.2×10-5 940,000

A. Exposure = Surface Residue (mg/cm2) × Saliva Extraction Efficiency × Surface Area of Toy Mouthed (cm2) 

B. Dose = [Exposure (mg/day)] / Body Weight (kg) 

C. MOE = POD / Daily Dose (rounded to two significant figures) 
42 

The MOEs in Table 10 are for incidental oral exposures and were calculated from the incidental 

oral dose using the POD of 30 mg/kg/day, which is the NOAEL from a 28-day oral toxicity 

study (Kim et al., 2008) and the LOAEL from a 90 day oral toxicity study (Kim et al., 2010). 

The MOEs for incidental oral exposures are 310,000 for 6 to < 12 month old children, 550,000 

for 1 to < 2 year old children, and 940,000 for 2 to <3 year old children, which are greater than 

the target MOE of 3,000 indicating that the risk for short- and intermediate-term exposure to 

children who mouth plastic toys containing Nanosilva is not of concern. 

Consumer Dermal Daily-Dose and MOEs to Flooring Incorporating Nanosilva

The dermal exposure to flooring incorporating Nanosilva was calculated using the following: 

Dermal Exposure = Surface Residue (mg/cm2) × Transfer factor from flooring to skin (%) × Body surface 

area in contact with floor (cm2

/day) 

Where the: 

 Body surface area contacting flooring is 4,500 cm2

/day for children between 6 and < 12 

months of age, 5,300 cm2/day for children between 1 and < 2 years of age, and for a 2 to 

< 3 year old is 6,100 cm2/day (U.S. EPA, 2011b). These are the total surface area of 

children and don’t account for the portion of child covered with clothing. 

The concentration of silver detected in fluids contacting Nanosilva containing coupons (Table 7) 

represents both the surface residue and the fraction of Nanosilva transferred from flooring to 

skin. The concentration of silver detected in low and high pH fluids, which are more 

representative of biological fluids, were all below the analytical detection limit of 5 μg/L for 

coupons containing 5, 10, and 20 mg/kg of Nanosilva. EPA is assuming that non-detectable 

concentrations of silver would be found in a similar test conducted with coupons containing 30 

mg/kg of Nanosilva. EPA is using 2.5 μg/L, which is one-half the detection limit of 5 μg/L, as 

the concentration of silver released from plastic incorporating Nanosilva. Using the 

concentration of silver, the volume of the leachate (50 mL), and the surface area of the test 

coupons, the amount of available nanosilver at the coupon surface is: 

Surface Residue × Transfer factor from flooring to skin: ଴.଴଴ଶହ ௠௚

௅

ൈ

ହ଴ ௠௅

ଵ଴଴଴ ௠௅

ൈ

ଵଵଶ ௖௠మ ൌ

଴.଴଴଴଴଴ଵଵଶ ௠௚

௖௠మ

The dermal exposure for flooring incorporating Nanosilva is: 

Dermal Exposure for 6 to < 12 month old children: 

ଵ.ଵଶൈଵ଴షల௠௚

௖௠మ ൈ

ସହ଴଴ ௖௠మ

ௗ௔௬

 ൌ

଴.଴଴ହ଴ଶ ௠௚ ௡௔௡௢௦௜௟௩௘௥

ௗ௔௬

Dermal Exposure for 1 to < 2 year old children: 

ଵ.ଵଶൈଵ଴షల௠௚

௖௠మ ൈ

ହଷ଴଴ ௖௠మ

ௗ௔௬

 ൌ

଴.଴଴ହଽଶ ௠௚ ௡௔௡௢௦௜௟௩௘௥

ௗ௔௬
43 

Dermal Exposure for 2 to < 3 year old children: 

ଵ.ଵଶൈଵ଴షల௠௚

௖௠మ ൈ

଺ଵ଴଴ ௖௠మ

ௗ௔௬

 ൌ

଴.଴଴଺଼ଵ ௠௚ ௡௔௡௢௦௜௟௩௘௥

ௗ௔௬

The dermal daily-dose was calculated from the dermal exposure using the following: 

Dermal Daily Dose = Exposure × Dermal absorption factor (%) / Body Weight 

Where: 

 Exposure is determined in the calculations above. 

 The DAF is 6.7% (see Section 4.2.3). 

 The body weight of a child with age between 6 and < 12 months is 9.2 kg, between 1 and 

< 2 years is 11.4 kg, between 2 and < 3 years is 13.8 (U.S. EPA, 2011b). 

Table 11 – Dermal MOEs for Children Exposed to Flooring Incorporating Nanosilva 

Age Range of Child 

Dermal 

ExposureA

(mg/day) 

Daily DoseB

(mg/kg/day) 

MOEC 

6 to < 12 months 5.02×10-3 3.7×10-5 820,000 

1 to < 2 years 5.92×10-3 3.5×10-5 860,000 

2 to < 3 years 6.81×10-3 3.3×10-5 910,000 

A. Exposure = Surface Residue (mg/cm2) × Floor to Skin Transfer Factor (%) × Skin Surface Area of Child 

(cm2

) 

B. Daily Dose = [Exposure (mg/day)] × Dermal absorption factor (%) / Body Weight (kg) 

C. MOE = POD / Daily Dose (rounded to two significant figures) 

The MOEs in Table 11 are for dermal exposures to flooring incorporating Nanosilva and were 

calculated from the dermal dose using the POD of 30 mg/kg/day, which is the NOAEL from a 

28-day oral toxicity study (Kim et al., 2008) and the LOAEL from a 90 day oral toxicity study 

(Kim et al., 2010). The MOEs for dermal exposures listed in Table 11 are 820,000 for 6 to < 12 

month old children, 860,000 for 1 to < 2 year old children, and 910,000 for 2 to < 3 year old 

children, which are greater than the target MOE of 3,000 indicating that the risk for short- and 

intermediate-term exposure to children who contact flooring incorporating Nanosilva is not of 

concern. 

Consumer Incidental Oral Daily-Dose and MOEs to Flooring Incorporating Nanosilva

The incidental oral exposure to nanosilver in flooring incorporating Nanosilva was calculated 

using the following: 
44 

Incidental Oral Exposure = Surface Residue (mg/cm2) × Transfer factor from flooring to skin (%) × 

Surface area of hand that contacts the floor and the child’s mouth (cm2

/event) × Frequency of hand-tomouth contacts (events/hr) × Saliva extraction efficiency (%) × Exposure time (hr/day) 

Where the: 

 Surface area of the hands that contact both the floor and the child’s mouth is 10 

cm2/event for children from 1 to < 2 years of age (U.S. EPA, 2012b). 

 Frequency of hand-to-mouth events is 19 events/hr for 6 to < 12 month old children, 20 

events/hr for 1 to < 2 year old children, and 13 events/hr for 2 to <3 year old children 

(U.S. EPA, 2011b). 

 Saliva extraction efficiency from hand to mouth is 50% (U.S. EPA, 2012b) 

 Exposure time is 101 minutes per day for 6 to < 12 month old children, 126 minutes per 

day for 1 to < 2 year old children, and 108 minutes/day for 2 to <3 year old children 

which are the mean times for “Doers” in kitchens and bathrooms (Table 16-15, U.S. 

EPA, 2011b) 

The concentration of silver detected in fluids contacting Nanosilva containing coupons (Table 7) 

represents both the surface residue and the fraction of Nanosilva transferred from flooring to 

skin. The concentration of silver detected in low and high pH fluids, which are more 

representative of biological fluids, were all below the analytical detection limit of 5 μg/L for 

coupons containing 5, 10, and 20 mg/kg of Nanosilva. EPA is assuming that non-detectable 

concentrations of silver would be found in a similar test conducted with coupons containing 30 

mg/kg of Nanosilva. EPA is using 2.5 μg/L, which is one-half the detection limit of 5 μg/L, as 

the concentration of silver released from plastic incorporating Nanosilva. Using the 

concentration of silver, the volume of the leachate (50 mL), and the surface area of the test 

coupons, the amount of available nanosilver at the coupon surface is: 

Surface Residue × Transfer factor from flooring to skin: ଴.଴଴ଶହ ௠௚

௅

ൈ

ହ଴ ௠௅

ൈ

௅

ଵ଴଴଴ ௠௅

ൈ

ଵଵଶ ௖௠మ ൌ ଴.଴଴଴଴଴ଵଵଶ ௠௚

௖௠మ

The incidental oral exposure for flooring incorporating Nanosilva is: 

Incidental Oral Exposure for 6 to < 12 month old children: ଵ.ଵଶൈଵ଴షల௠௚

௖௠మ ൈ

ଵ଴ ௖௠మ

௘௩௘௡௧

ൈ

ଵଽ ௘௩௘௡௧௦

௛௥

 ൈ

ହ଴%

ଵ଴଴%

ൈ

ଵ଴ଵ ௠௜௡

ௗ௔௬

 ൈ

௛௥

଺଴ ௠௜௡ ൌ ଴.଴଴଴ଵ଻଼ ௠௚ ௡௔௡௢௦௜௟௩௘௥ௗ௔௬

Incidental Oral Exposure for 1 to < 2 year old children: ଵ.ଵଶൈଵ଴షల௠௚

௖௠మ ൈ

ଵ଴ ௖௠మ

௘௩௘௡௧

ൈ

ଶ଴ ௘௩௘௡௧௦

௛௥

 ൈ

ହ଴%

ଵ଴଴%

ൈ

ଵଶ଺ ௠௜௡

ௗ௔௬

 ൈ

௛௥

଺଴ ௠௜௡ ൌ ଴.଴଴଴ଶଷସ ௠௚ ௡௔௡௢௦௜௟௩௘௥ௗ௔௬
45 

Incidental Oral Exposure for 2 to < 3 year old children: ଵ.ଵଶൈଵ଴షల௠௚

௖௠మ ൈ

ଵ଴ ௖௠మ

௘௩௘௡௧

ൈ

ଵଷ ௘௩௘௡௧௦

௛௥

 ൈ

ହ଴%

ଵ଴଴%

ൈ

ଵ଴଼ ௠௜௡

ௗ௔௬

 ൈ

௛௥

଺଴ ௠௜௡ ൌ ଴.଴଴଴ଵଷଵ ௠௚ ௡௔௡௢௦௜௟௩௘௥ௗ௔௬

The incidental oral daily-dose was calculated from the dermal exposure using the following: 

Incidental Oral Daily-Dose = Exposure / Body Weight 

Where: 

 Exposure is determined in the calculations above. 

 The body weight of a child with age between 6 and < 12 months is 9.2 kg, between 1 and 

< 2 years is 11.4 kg, between 2 and < 3 years is 13.8 (U.S. EPA, 2011b). 

Table 12 – Incidental Oral MOEs for Children Exposed to Flooring Incorporating 

Nanosilva

Age Range of Child Oral ExposureA

(mg/day) 

Daily DoseB

(mg/kg/day) 

MOEC 

6 to < 12 months 1.8×10-4 1.9×10-5 >1,000,000 

1 to < 2 years 2.3×10-4 2.1×10-5 >1,000,000 

2 to < 3 years 1.3×10-4 0.95×10-5 >1,000,000 

A. Exposure = Surface Residue (mg/cm2) × Transfer factor from flooring to skin (%) × Surface area of hand 

that contacts the floor and the child’s mouth (cm2

/event) × Frequency of hand-to-mouth contacts 

(events/hr) × Saliva extraction efficiency (%) × Exposure time (hr/day) 

B. Dose = [Exposure (mg/day)] / Body Weight (kg) 

C. MOE = POD / Daily Dose 

The MOEs in Table 12 are for incidental oral exposures to flooring incorporating Nanosilva and 

were calculated from the incidental oral dose using the POD of 30 mg/kg/day, which is the 

NOAEL from a 28-day oral toxicity study (Kim et al., 2008) and the LOAEL from a 90 day oral 

toxicity study (Kim et al., 2010). The MOEs for incidental oral exposures are greater than 

1,000,000 for 6 to < 12 month old, 1 to < 2 year old children, and 2 to < 3 year old children, 

which are greater than the target MOE of 3,000 indicating that the risk for short- and 

intermediate-term exposure to children who mouth their hands after contacting flooring 

incorporating Nanosilva is not of concern. 

5.2.3 Consumer Margins of Exposure to Textiles Incorporating Nanosilva 

EPA expects that consumers will be exposed to textiles incorporating Nanosilva by the routes of 

inhalation, dermal, and incidental oral exposures. 
46 

Consumer Inhalation Daily-Dose and MOE

EPA recognizes the potential for inhalation exposure to nanosilver during laundry drying of 

textiles incorporating Nanosilva. However, EPA lacks information on the release rate of 

nanosilver from textiles incorporating Nanosilva during laundry drying. While exposure may 

occur during laundry drying, EPA believes that when compared to exposure through dermal and 

oral contact with textiles incorporating Nanosilva, exposure during laundry drying will likely be 

of lower significance. To support this, an estimate of the inhalation dose that might occur from 

exposure to textiles incorporating Nanosilva during laundry drying was calculated using the 

following: 

Inhalation Daily Dose = Amount of Nanosilva in clothing handled per day × Unit Exposure 

Where: 

 Amount of Nanosilva in clothing handled per day assumed that one t-shirt containin 

Nanosilva was laundered where the amount of nanosilver in a t-shirt is equal to 150 g × 

30 mg/kg × kg ÷ 1000 g = 4.5 mg. 

 The unit exposure for inhalation of wettable powder is 7.8 μg/m3

/lb when no respirator is 

worn (U.S. EPA, 2011a). 

Table 13 – Inhalation MOE for Drying Textiles Incorporating Nanosilva

Scenario

Amount of 

Nanosilver 

Laundered per dayA

Unit ExposureB

(μg/m3/lb AI)

Daily DoseC

(μg/m3)

MOED

Unload Clothes Dryer 

Without Respirator 4.5 mg 7.8 0.000077 640,000

A. Amount of AI laundered per day = Amount of Nanosilva in t-shirt × Mass of t-shirt 

B. PHED unit exposure value converted to air concentration units based on mean 8 hour TWA 

C. Dose = Amount Nanosilver Handled × Unit Exposure 

D. MOE = POD / Daily Dose (rounded to two significant digits) 

The MOE shown in Table 13 was calculated by dividing the inhalation dose by the POD of 49 

μg/m3

, which is the NOAEL from a 90-day inhalation toxicity study for nanosilver (Song et al., 

2012). This assumes that Nanosilva released from textiles during drying behaves like nanosilver 

after inhalation. This analysis further assumes that all of the nanosilver from the textile 

incorporating Nanosilva becomes airborne during a single laundry drying event and this 

exposure would be similar to mixing and loading of wettable powders (U.S. EPA, 2011a). The 

MOE of 640,000 indicates that the risk for short- and intermediate-term exposure to laundering 

one textile incorporating Nanosilva per day is not of concern. Up to 640 t-shirts incorporating 

Nanosilva could be laundered per day and the calculated MOE would be greater than the target 
47 

MOE of 1,000, indicating that the risk for short- and intermediate-term exposure to drying 

textiles incorporating Nanosilva is not of concern. 

Consumer Dermal Daily-Dose and MOEs

The dermal exposure to textiles incorporating Nanosilva was calculated using the following: 

Dermal Exposure = Amount of Nanosilva in Textile × Cloth Density × Surface Area Exposed × Transfer 

Efficiency 

Where: 

 The textile incorporating Nanosilva contains 30 mg/kg nanosilver. 

 The cloth density is 10 mg/cm2

 based on the density of mixed cotton and synthetics. This 

value is a standard assumption used in OPP risk assessments and was taken from the 

HERA Guidance Document Methodology (AISE/CEFIC, 2005). 

 The total surface area for a 6 to < 12 month old is 4,500 cm2

/day, for a 1 to < 2 year old is 

5,300 cm2/day, and for a 2 to < 3 year old is 6,100 cm2/day (U.S. EPA, 2011b). 

 The cloth-to-skin transfer efficiency was based on the amount of silver released during 

the leaching study which was 0.9% (see Table 9). 

Table 14 – Dermal Exposure to Textiles Incorporating Nanosilva 

Age Range of 

Child 

Application 

Rate (mg/kg) 

Cloth 

Density

(mg/cm2)

Surface Area 

Exposed 

(cm2/day) 

Cloth-to-Skin 

Transfer 

Efficiency 

ExposureA

(mg/day) 

6 to <12 months 30 10 4,500 0.9% 0.0122 

1 to < 2 years 30 10 5,300 0.9% 0.0143 

2 to < 3 years 30 10 6,100 0.9% 0.0165 

A. Exposure = Application Rate × Cloth Density × Surface Area Exposed × Cloth-to-Skin Transfer Efficiency × 

Dermal Absorption Factor 

The dermal dose was calculated from the dermal exposure using the following: 

Dermal Daily Dose = Exposure × Dermal Absorption Factor / Body Weight 

Where: 

 Exposure is determined in the calculation above. 
48 

 The DAF is 6.7% (see Section 4.2.3). 

 The body weight of a child with age between 6 and < 12 months is 9.2 kg, between 1 and 

< 2 years is 11.4 kg, between 2 and < 3 years is 13.8 (U.S. EPA, 2011b). 

Table 15 – Dermal MOEs for Children Exposed to Textiles Incorporating Nanosilva

Age Range of Child Body Weight 

of Child (kg) 

Exposure 

(mg/day) 

Daily DoseA

(mg/kg/day) MOEB

6 to <12 months 9.2 0.012 8.8×10-5 340,000 

1 to < 2 years 11.4 0.014 8.4×10-5 360,000 

2 to < 3 years 13.8 0.016 8.0×10-5 370,000 

A. Dose = [Exposure (mg/day)] × DAF / Body Weight (kg) 

B. MOE = POD / Daily Dose (rounded to two significant figures) 

The MOEs for dermal exposures were calculated from the dermal dose using the POD of 30 

mg/kg/day, which is the NOAEL from a 28-day oral toxicity study (Kim et al., 2008) and the 

LOAEL from a 90 day oral toxicity study (Kim et al., 2010). The MOEs for dermal exposures 

are 340,000 for 6 to < 12 month old, 360,000 for 1 to < 2 year old children, and 370,000 for 2 to 

< 3 year old children, which are greater than the target MOE of 3,000 indicating that the risk for 

short- and intermediate-term exposure to children from wearing textiles incorporating Nanosilva 

is not of concern. 

Consumer Incidental Oral Daily-Dose and MOEs

Incidental oral exposures were calculated using the following: 

Incidental Oral Exposure = Amount of Nanosilva in Textile × Cloth Density × Surface Area Mouthed × 

Saliva Extraction Efficiency 

Where: 

 The textile incorporating Nanosilva contains 30 mg/kg nanosilver. 

 The cloth density is 10 mg/cm2

 based on the density of mixed cotton and synthetics. This 

value is a standard assumption used in OPP risk assessments and was taken from the 

HERA Guidance Document Methodology (AISE/CEFIC, 2005) 

 The surface area of fabric that is mouthed by a toddler per day is assumed to be 100 cm2 

(~16 in.2) which represents an estimate for the area of blanket or shirt sleeve. 
49 

 The nanosilver saliva extraction efficiencies for mouthing fabric are based on the results 

of the leaching study which was 0.9% (see Table 9). 

Table 16 – Incidental Oral Exposure to Textiles Incorporating Nanosilva

Application Rate 

(mg/kg) 

Cloth Density

(mg/cm2) 

Surface Area 

Mouthed 

(cm2/day) 

Saliva 

Extraction 

Efficiency 

ExposureA

(mg/day) 

30 10 100 0.9% 0.00027 

A. Exposure = Application Rate × Cloth Density × Surface Area Exposed × Saliva Extraction Efficiency 

The incidental oral daily-dose was calculated from the incidental oral exposure using the 

following: 

Incidental Oral Daily-Dose = Exposure / Body Weight 

Where: 

 Exposure is determined in the calculation above. 

 The body weight of a child with age between 6 and < 12 months is 9.2 kg, between 1 and 

< 2 years is 11.4 kg, between 2 and < 3 years is 13.8, which is for children who mouth 

textiles (U.S. EPA, 2011b). 

Table 17 – Incidental Oral MOEs for Children Exposed to Textiles Incorporating 

Nanosilva

Age Range of Child Body Weight 

of Child 

Exposure 

(mg/day) 

Daily DoseA

(mg/kg/day) MOEB

6 to <12 months 9.2 0.00027 2.9×10-5 >1,000,000 

1 to < 2 years 11.4 0.00027 2.4×10-5 >1,000,000 

2 to < 3 years 13.8 0.00027 2.0×10-5 >1,000,000 

A. Dose = [Exposure (mg/day)] / Body Weight (kg) 

B. MOE = POD / Daily Dose (rounded to two significant digits) 

The MOEs in Table 17 are for incidental oral exposures were calculated from the incidental oral 

dose using the POD of 30 mg/kg/day, which is the NOAEL from a 28-day oral toxicity study 

(Kim et al., 2008) and the LOAEL from a 90 day oral toxicity study (Kim et al., 2010). The 

MOEs for incidental oral exposures are greater than 1,000,000 for 6 to < 12 month old, 1 to < 2 
50 

year old, and 2 to < 3 year old children, which are greater than the target MOE of 3,000 

indicating that the risk for short- and intermediate-term exposure to children who mouth textiles 

incorporating Nanosilva is not of concern. 

5.2.4 Consumer Aggregate Margins of Exposure to Plastics and Textiles Incorporating 

Nanosilver 

In the Federal Food, Drug, and Cosmetic Act (FFDCA), Congress specified that, to establish an 

acceptable level of a given pesticide’s chemical residue that could be found in or on food 

products, EPA must determine that “there is a reasonable certainty that no harm will result from 

aggregate exposures to the pesticide chemical residue, including all anticipated dietary exposures 

and all other exposures for which there is reliable information.” Furthermore, when enacting this 

provision of the FFDCA, Congress also amended FIFRA’s definition of unreasonable adverse 

effects. Specifically, Congress redefined unreasonable adverse effects to include “a human 

dietary risk from residues that result from a use of a pesticide in or on any food inconsistent with 

the standard under” the FFDCA. In other words, Congress explicitly required the consideration 

of aggregate exposures for registration decisions under FIFRA for food-use pesticides, but chose 

not to similarly alter the statutory requirements for non food-use pesticides. 

For transparency, the following analysis is for children who are simultaneously exposed to 

nanosilver via the incidental oral and dermal routes of exposure while mouthing toys, contacting 

flooring, and mouthing textiles incorporating Nanosilva. Inhalation exposure to children during 

laundry drying of textiles incorporating Nanosilva is not anticipated and is not considered as part 

of the aggregate exposure. The aggregate daily dose was calculated by adding the daily oral and 

dermal doses using the following: 

Aggregate Daily Dose = Incidental Oral Dose to Toys Incorporating Nanosilva + Dermal 

Dose to Flooring Incorporating Nanosilva + Incidental Oral Dose to Flooring Incorporating 

Nanosilva + Dermal Dose to Textiles Incorporating Nanosilva + Incidental Oral Dose to 

Textiles Incorporating Nanoslva 

Where: 

 Incidental Oral Daily Dose to Toys Incorporating Nanosilva is from Table 10 

 Dermal Daily Dose to Flooring Incorporating Nanosilva is from Table 11 

 Incidental Oral Daily Dose to Flooring Incorporating Nanosilva is from Table 12 

 Dermal Daily Dose to Textiles Incorporating Nanosilva is from Table 15 

 Incidental Oral Daily Dose to Textiles Incorporating Nanosilva is from Table 17 

The oral and dermal daily doses can be summed because they are evaluated using the same POD 

of 30 mg/kg/day. The aggregate MOE in Table 18 for 6 and < 12 month old children is 110,000, 
51 

is 140,000 for children who are 1 and < 2 years, and 170,000 for 2 to <3 year old children, which 

indicates that the risk for short- and intermediate-term exposure to children is not of concern. 

Table 18 – Aggregate MOEs for Children Exposed to Plastics and Textiles Incorporating 

Nanosilva 

Age Range of Child 

Incidental 

Oral Dose to 

Toys 

Flooring Textiles 

Aggregate 

Dose 

Aggregate 

MOEA

Dermal Dose Incidental 

Oral Dose Dermal Dose Incidental Oral Dose 

6 to < 12 months 9.7×10-5 3.7×10-5 1.9×10-5 8.8×10-5 2.9×10-5 2.7×10-4 110,000 

1 to < 2 years 5.5×10-5 3.5×10-5 2.1×10-5 8.4×10-5 2.4×10-5 2.2×10-4 140,000 

2 to < 3 years 3.2×10-5 3.3×10-5 0.95×10-5 8.0×10-5 2.0×10-5 1.7×10-4 170,000 

A. Aggregate MOE = POD / Daily Dose (rounded to two significant digits) 

The assessment above is for the concurrent exposure to the nanosilver in Nanosilva via the 

incidental oral and dermal routes of exposure while mouthing toys and textiles, and contacting 

flooring incorporating Nanosilva. Aggregate assessments can also include other sources of 

exposure such as to Nanosilva in food and drinking water, and to other nanosilvers in the market 

place that are identical to the nanosilver in Nanosilver. There are no anticipated food exposures 

to Nanosilva since Nanosilva LLC has not supplied the necessary information to support a food 

contact application and the pesticide label for Nanosilva states that it is only for non-food contact 

use. Neither the nanosilver that might break away from Nanosilva or Nanosilva is anticipated to 

enter drinking water because these particulates will be removed by gravitational sedimentation 

and adsorption (see Section 5.1.1). 

EPA has determined that the only other pesticide which was knowingly registered as containing 

nanosilver is HeiQ AGS-20 (U.S. EPA, 2011a); although, the nanosilver in HeiQ AGS-20 does 

not have the exact same size range and surface coatings as the nanosilver in Nanosilva. The 

incidental oral and dermal MOEs for HeiQ AGS-20 were all greater than 1,100 and the 

incidental oral and dermal MOEs for Nanosilva are all greater than 310,000. None of the MOEs 

for HeiQ AGS-20 were less than the target MOEs for that assessment, which was based on 

conservative estimates expected to significantly overestimate exposures to the nanosilver in 

HeiQ AGS-20. Although EPA has not determined whether it’s appropriate to consider 

concurrent exposures to the nanosilver in Nanosilva and HeiQ AGS-20, the agency has 

concluded that even if exposures to these two active ingredients were to occur concurrently, they 

would not result in MOEs that would be of concern. In addition, EPA is currently revising the 

HeiQ AGS-20 risk assessment based on newly submitted leaching data and to be consistent with 

the risk assessment framework proposed in this decision document. EPA anticipates that the 
52 

calculated MOEs for incidental oral and dermal exposure to textiles containing HeiQ AGS-20 

will increase based on newly submitted data. 

As to exposure to other nanosilvers in the market place, an aggregate risk assessment involves 

the analysis of exposure to a single chemical by multiple pathways and routes of exposure that 

share a common toxic effect. For EPA to conduct such an assessment requires that the Agency 

possess sufficient information such as: 

 identification of toxicological endpoints for each exposure route and duration; 

 identification of potential exposures for each pathway (food, water, and/or residential); 

 reconciliation of durations and pathways of exposure with durations and pathways of 

health effects; 

 determination of which possible residential exposure scenarios are likely to occur 

together within a given time frame (co-occurrence); 

 determination of magnitude and duration of exposure for all exposure combinations; 

 determination of the appropriate technique (deterministic or probabilistic) for exposure 

assessment; 

 and determination of the appropriate risk metric to estimate aggregate risk. 

When EPA has the above information, or as in the case of triclosan where population-based 

biological monitoring data were available to assess the co-occurrence of uses, EPA develops an 

aggregate exposure assessment for residential uses of a single chemical. However, in the case of 

nanosilver, with the exception of the HeiQ products, EPA does not yet have adequate 

information on the composition of the nanosilver or potential nanosilver in other silver based 

pesticides to determine if all nanosilvers should be treated as a single chemical. 

5.2.5 Conclusions for Consumer Risk 

The MOEs calculated for the incidental oral exposure to nanosilver in plastic toys incorporating 

Nanosilva are 310,000 for 6 to < 12 month old children, 550,000 for 1 to < 2 year old children, 

and 940,000 for 2 to <3 year old children, which are greater than the target MOE of 3,000 

indicating that there is no risk concern for short- or intermediate-term exposure to children who 

mouth toys incorporating Nanosilva. The MOEs calculated for the dermal exposure to nanosilver 

in flooring incorporating Nanosilva are 820,000 for 6 to < 12 month old children, 860,000 for 1 

to < 2 year old children, and 910,000 for 2 to <3 year old children, which are greater than the 

target MOE of 3,000 indicating that there is no risk concern for short- or intermediate-term 

exposure to children who crawl on flooring incorporating Nanosilva. The MOEs calculated for 

the incidental oral exposure to nanosilver in flooring incorporating Nanosilva are greater than 

1,000,000 for 6 to < 12 month old children, 1 to < 2 year old children, and 2 to <3 year old 

children, which are greater than the target MOE of 3,000 indicating that there is no risk concern 

for short- and intermediate-term exposure to children who mouth flooring incorporating 
53 

Nanosilva. The MOEs calculated for the dermal exposure to nanosilver in textiles incorporating 

Nanosilva are 340,000 for 6 to < 12 month old children, 360,000 for 1 to < 2 year old children, 

and 370,000 for 2 to <3 year old children, which are greater than the target MOE of 3,000 

indicating that there is no risk concern for short- and intermediate-term exposure to children who 

wear textiles incorporating Nanosilva. The MOEs calculated for incidental oral exposures to 

nanosilver in textiles incorporating Nanosilva are greater than 1,000,000 for 6 to < 12 month old 

children, 1 to < 2 year old children, and 2 to < 3 year old children, which are greater than the 

target MOE of 3,000 indicating that there was no risk concern for short- and intermediate-term 

exposure to children who mouth and chew textiles incorporating Nanosilva. 

The aggregate MOEs for children who are, within the same short time frame, mouthing a toy, 

flooring, and a textile while wearing a textile and crawling on flooring all incorporating 

Nanosilva are greater than 100,000 for all lifestages, which indicates that the risk for short- and 

intermediate-term simultaneous exposure to plastics and textiles incorporating Nanosilva is not a 

concern. 

Although children’s short- and intermediate-term exposure durations to plastic toys and textiles 

treated with Nanosilva are expected, children are not anticipated to play with Nanosilva treated 

toys or contact Nanosilva treated textiles every day for a period of greater than six months, thus 

long-term exposures are unlikely. Nanosilva is proposed to be used in flooring meaning that 

long-term dermal and incidental oral exposures to children contacting flooring are possible. 

However, EPA does not believe that children will have continuous and daily exposure to flooring 

for a period of greater than 6 months. Even if these unlikely long-term exposures were to occur, 

the calculated MOEs for children’s dermal and incidental oral exposure to plastics and textiles 

incorporating Nanosilva are greater than 100,000 indicating that daily exposure to plastics and 

textiles incorporating Nanosilva for greater than 6 months are not likely to be of concern. 

There are several uncertainties in the consumer risk assessment. Because Nanosilva LLC 

completed leaching tests using plastic coupons containing nanosilver at concentrations of 5, 10, 

and 20 mg/kg, EPA assumed that non-detectable concentrations of silver would also be found if 

a leaching test was conducted with coupons containing nanosilver at 30 mg/kg. This assumption 

was reasonable because the concentration of silver in the textile leaching studies, which were 

conducted on textiles incorporating nanosilver at 30 mg/kg, was also below the analytical 

detection limit, making it unlikely that Nanosilver incorporated into plastic at 30 mg/kg will 

release nanosilver at detectable concentrations. The PODs used were based on inhalation and 

oral toxicity studies completed using nanosilver, not Nanosilva or the nanosilver that might break 

away from Nanosilva, and thus these PODs may not equal PODs determined from testing 

conducted with Nanosilva or the nanosilver found in Nanosilva. Because the nanosilver in the 

toxicity database is different from the nanosilver in Nanosilva and because the database is 

incomplete with respect to reproductive and developmental effects, the Agency used a maximum 
54 

10-fold database uncertainty factor when evaluating the risk from exposure to Nanosilva and the 

nanosilver that might be released from products incorporating Nanosilva. There are also 

uncertainties in extrapolating from effects observed after feeding test-animals nanosilver (i.e., 

oral route) to the effects that might be observed after applying nanosilver to the skin of test 

animals (i.e., dermal route). 

EPA is able to determine that for the period of conditional registration, there is a low probability 

of adverse risk to children from plastics and textiles incorporating Nanosilva. Thus, the Agency 

concludes that use of Nanosilva will not cause unreasonable adverse effects on the environment 

during the period when newly required data are being developed. 

5.2.6 Consumer Exposure and Health Data Requirements 

EPA has determined that an additional leaching study is required for plastics incorporating 

Nanosilva using biological fluids and physiological temperatures, which includes methods to 

determine the form of silver released. EPA has determined that the textile leaching study 

submitted by Nanosilva LLC is acceptable and no additional studies on textiles incorporating 

Nanosilva are required. 

EPA is proposing to require that Nanosilva LLC conduct plastic leaching tests as part of Tier I 

studies because the available study was completed using a protocol that was not reviewed by 

EPA prior to conducting the study, the coupons tested has less than the maximum nanosilver 

concentration of 30 mg/kg, and the study did not use physiological fluids. The newly required 

leaching study should determine the nature and quantity of silver released from plastics 

incorporating Nanosilva under conditions of use. EPA will evaluate the results from the plastic 

leaching test along with results from Tier I toxicity studies discussed in Section 5.1.4 using a 

weight of evidence approach based on physical and chemical properties, toxicity, and exposure. 

If the Agency identifies a concern, then the Agency may require additional toxicity data to assess 

potential adverse health outcomes in young children resulting from incidental oral exposure to 

Nanosilva-treated plastic toys (i.e., children chewing on toys). If additional toxicity data are 

needed, the study should evaluate peri- and post-natal exposure to Nanosilva. A more detailed 

description of these data requirements is provided in Appendix B. 
55 

VI. ENVIRONMENTAL RISK ASSESSMENT

EPA anticipates the following substances could enter the environment through leaching of 

plastics and textiles incorporating Nanosilva: 

1) Silver ions released from Nanosilva; 

2) Nanosilva complex; and/or 

3) Nanosilver that might break away from the Nanosilva complex. 

There are no studies available to characterize the environmental fate or ecotoxicity of Nanosilva. 

However, there are studies available in the scientific literature for nanosilver. Since nanosilver 

may be released from Nanosilva, the Agency has considered the scientific literature studies on 

nanosilver fate and ecotoxicity relevant to Nanosilva. The following sections cover the 

environmental fate of nanosilver, the environmental hazards posed by silver and nanosilver, and 

the potential risk to aquatic species from nanosilver. As part of these discussions, additional data 

that EPA is proposing to require in order to confirm its assessment of the environmental risks of 

Nanosilva are also identified. 

6.1 Environmental Fate 

Nanosilva LLC has not conducted any studies to characterize the environmental fate of 

Nanosilva or the other particles that could be released during leaching or disposal of plastics and 

textiles incorporating Nanosilva. In lieu of this information, the Agency is relying on studies 

available in the scientific literature, discussed in the following section, as the basis for 

determining the fate of nanosilver in the environment. 

6.1.1 Nanosilver 

The rate at which nanosilver transforms into ionic silver determines the length of time that these 

particles will reside in the environment. Although there are studies reporting that nanosilver will 

completely transform into ionic silver within six days after being dispersed into deionized water 

(Liu and Hurt, 2010), these results are only for one form of nanosilver and under conditions 

which are not representative of the environment. In the environment, nanosilver is likely to 

complex with naturally occurring anions such as chloride and sulfide or natural organic matter 

such as humic acids, which will significantly delay the rate at which nanosilver transforms into 

ionic silver. For example, Choi et al., (2009) provided spectroscopic evidence showing that 

nanosilver reacts with sulfide to produce stable silver-sulfide complexes, which were shown by 

Liu et al. (2010) and Levard et al. (2011) to have undetectable rates of nanosilver to ionic silver 

transformation. These stabilized nanosilver complexes are likely to partition to sediments, rather 

than remain suspended in water, due to gravitational settling and coagulation processes (see Page 

19 FIFRA SAP, 2009). Likewise, nanosilver is anticipated to partition to biosolids during 
56 

wastewater treatment but may also be released in the effluent. Thus, there is the potential for 

nanosilver to reside or persist in the environment for a significant period of time where these 

particles are most likely to be associated with sediments. 

6.1.2 Silver Ions 

Ionic silver typically has low concentrations in natural waters, in the nanogram per liter range, 

due to its reactivity with chloride, sulfides, and natural organic matter (Andren and Armstrong, 

1999). As with nanosilver, ionic silver is found in sediments and associated with biosolids in 

wastewater treatment plants. 

6.1.3 Impacts to Wastewater Treatment/Septic Systems 

There is the potential for nanosilver that might be released from plastics and textiles 

incorporating Nanosilva to reach publicly owned wastewater treatment and privately owned 

septic systems where they will most likely complex with sulfide and partition to biosolids (Kaegi 

et al., 2011). Once entrained in the biosolids, the nanosilver could serve as a long term source of 

ionic silver and could potentially adversely affect microorganisms that are vital to the wastewater 

treatment process. There are contradictory reports in the scientific literature regarding the impact 

of nanosilver on wastewater treatment systems. For example, nanosilver was reported to inhibit 

nitrification in the range of 50% (Choi and Hu, 2009a) to 84% (Choi and Hu, 2009b) based on a 

reduction in oxygen uptake rate in simulated wastewater sludge. However, Burkhardt et al. 

(2010) found no impact to nitrification at nanosilver dosages of 1 mg/L, the same dosage that 

Choi and Hu (2009a and 2009b) reported as inhibitory in municipal wastewater sludge. These 

two research groups are reporting different findings with the Burkhardt group suggesting little 

impact of nanosilver to nitrification and the Hu group suggesting that an impact to wastewater 

treatment systems from nanosilver is expected. A third group independently determined that 

nanosilver at concentrations from 0.5 to 1.5 mg/L had no detectable effect on the ability of the 

wastewater bacteria to biodegrade organic material, as measured by chemical oxygen demand 

(COD) (Wang et al., 2012a). More recent work by the Hu group reported that nanosilver at 

concentrations of up to 40 mg/L had negligible impact on anaerobic digestion and methanogenic 

organisms (Yang et al., 2012). 

While there are reports suggesting the potential for nanosilver to impact wastewater treatment 

operations, the Agency does not anticipate that registering Nanosilva will lead to negative 

impacts to wastewater treatment systems. This conclusion is based on the limited amount of 

silver released from plastics and textiles incorporating Nanosilva (see Section 5.2.1) and the 

small volume of nanosilver (i.e., < 1,123 kg/yr as estimated in Section 6.3.1) expected to be 

introduced into commerce from plastics and textiles incorporating Nanosilva. 
57 

6.2 Environmental Toxicity 

Nanosilva LLC has not conducted any studies to characterize the ecotoxicity of Nanosilva or the 

other particles that could be released during leaching or disposal of plastics incorporating 

Nanosilva. In lieu of this information, the Agency is relying on studies available in the scientific 

literature, discussed in the following section, as the basis for determining the ecotoxicity of 

nanosilver. 

6.2.1 Silver Ions 

EPA has considerable data on the environmental hazards posed by the release of silver ions from 

conventional silver-based products. The precious metal silver is a trace element found in the 

Earth's crust and is generally naturally present in surface waters in relatively low concentrations 

as compared to metals such as copper and zinc. However, it may become toxic to aquatic life at 

elevated concentrations. Thus, silver concentrations in natural environments, and its biological 

availability, are important. Naturally occurring concentrations of silver have been reported from 

about 0.0002 to just over 1 μg/L in freshwater systems (Campbell et al., 2002). Elevated 

concentrations of silver in surface waters have generally been associated with wastewater 

treatment plant effluent discharges (Bell and Kramer, 1999). Based on the most recent Agency 

assessment of silver, EPA does not expect unreasonable adverse effects to the environment from 

registered uses, including the materials preservative use pattern (U.S. EPA, 1993). 

6.2.2 Aquatic Toxicity of Nanosilver 

Although no tests with Nanosilva and aquatic organisms were submitted, there are studies in the 

scientific literature covering the toxicity of nanosilver to aquatic organisms (Table 19). These 

studies indicate to the Agency that, if sufficient quantities of nanosilver break away from 

Nanosilva and reach surface water, and if such nanoparticles display toxicity similar to the 

nanosilver used in these studies, then exposure of Nanosilva derived nanosilver may result in 

adverse effects to aquatic species. These results also indicate that, of the organisms tested, 

Daphnia magna is the most acutely sensitive to nanosilver. As with the toxicity for silver ions, 

the toxicity of nanosilver may depend on the ligands or counter ions (e.g., Ca2+) present in the 

test media. For example, the LC50 reported by Laban et al. (2010) was up to 27 times greater than 

the LC50 reported by Kennedy et al. (2010) for the same organism where this difference may 

have been caused by the greater amount of calcium carbonate present in the work by Laban et al. 

(2010). 

 
58 

Table 19 – Nanosilver Toxicity to Aquatic Species 

Aquatic Organisms 

(Study Citation) Toxicity

Silver Nitrate Nanosilver

Total Silver 

(μg/L)

Total Silver 

(μg/L)

Dissolved 

Silver (μg/L)

Water 

Characteristics

Freshwater Algae 

Raphidocelis 

subcapitata 

(Wang et al., 2012b) 

4.5-hr EC50A 31.3 929-120,000 Not reported 

Dutch Standard 

Water 

(demineralized 

water with sodium 

bicarbonate, 

potassium 

bicarbonate, 

calcium chloride, 

and magnesium 

sulfate and then 

aerated for 24 h at 

27 °C) 

Newborn Neonates of 

Freshwater 

Zooplankton 

Chydorus sphaericus

(Wang et al., 2012b) 

48-hr EC50 4.3 36.7-128.4 Not reported 

Freshwater Fish 

Embryos 

Danio rerio (Wang et 

al., 2012b) 

96-hr EC50 78.8 88.4-210.3 Not reported 

Freshwater Flea 

D. magna 

(Kennedy et al., 2010) 

96-hr LC50B

 1.2±0.5 1.8 to 97.0C 0.3 to 1.9C,D Moderately 

hard reconstituted 

water (MHRW), 

80 mg/L as calcium 

carbonate 

Freshwater Fish Larval 

Pimephales promelas 

(Kennedy et al., 2010) 

96-hr LC50 6.1±0.6 9.0 to 125.6C

 1.5 to 5.6C,D

Freshwater Flea 

D. magna 

(Hoheisel et al., 2012) 

48-hr LC50 0.98 4.3 to 30.4E

 Not reported 

Moderate ionic 

strength Lake 

Superior water 

Freshwater Flea 

D. magna 

(Asghari et al., 2012) 

48-hr LC50 0.23 2 to 187E

 Not reported OECD 202 Media 

Freshwater Flea 

D. magna 

(Hoheisel et al., 2012) 24-hr LC50 0.81 4.3 to 10.5 3.5 to 8.1 

Moderately 

hard reconstituted 

water (MHRW), 

96 mg/L as sodium 

bicarbonate 

Natural 

Bacterioplankton 

(Das et al., 2012) 48-hr EC50 Not reported 15 to 276 No reported 

EC50 values did 

not correlate with 

any of the basic 

limnological 

parameters 

Freshwater Fish 

Embryo 

Pimephales promelas 

(Laban et al., 2010) 

96-hr LC50 15 9,400 to 

10,600 40 

215 to 240 mg/L as 

calcium carbonate; 

pH 8.3 to 8.5 

Freshwater Fish 

Embryo 

Danio rerio 

(Wang et al., 2012b) 

96-hr LC50 77 to 82 72 to 223G 0.7 to 22.3H Not reported 
59 

A. EC50 (medium effect concentration): Concentration at which the response is halfway between the baseline and 

maximum after the specified exposure time. 

B. LC50 (median lethal concentration): Concentration required to kill half the members of a tested population after 

the specified exposure. 

C. Range for eight different nanosilver formulations. 

D. Expressed as the fraction of suspended nanosilver. 

E. For nanosilvers with diameters from 10 to 50 nm. 

F. Three nanosilver formulations 

G. Rage of three different nanosilver formulations 

H. Estimated 

Based on the studies listed in Table 19, the most sensitive aquatic species to nanosilver is 

Daphnia magna. The lowest 96-hr EC50 was 1.8 μg/L (Kennedy et al., 2010), which is 1.5 μg/L 

after subtracting the fraction of ionic silver present. This test was conducted in moderately hard 

reconstituted water (MHRW), which is a test medium representative of surface water in the 

United States. 

Although these studies are useful for indicating the concentration of nanosilver that might lead to 

effects in aquatic species, and to identify for this assessment an effect level for evaluating the 

ecotoxicity of nanosilver that could be released from plastics incorporating Nanosilva, the data 

represent short-term (e.g., 96 hr) acute exposures and do not characterize the effects from longerterm exposure to nanosilver. There is one long-term study on the effects of nanosilver on a 

freshwater fish (Oryzias latipes) that reported a no observed effect concentration (NOEC) of 100 

μg/L based on the lethal effect observed at the dose of 250 μg/L during a 14-day exposure period 

(Wu and Zhou, 2013). Non-lethal effects were reported for all nanosilver concentration tested 

(50 to 500 μg/L) including histological lesions in the fish tissues along with silver accumulation 

in the liver. 

6.3 Aquatic Risk Assessment 

EPA expects that Nanosilva, silver ions released from Nanosilva, and nanosilver that might 

break away from Nanosilva could enter the environment through the following scenarios: 

1. Leaching of plastics and textiles incorporating Nanosilva 

2. Application of wastewater treatment biosolids to agricultural fields 

Leaching of plastics and textiles incorporating Nanosilva is anticipated to be the primary route 

by which Nanosilva, silver ions, and nanosilver reach the environment. The silver released 

during leaching of plastics and textiles incorporating Nanosilva could be discharged to the 

sanitary sewer system leading to publically owned wastewater treatment and privately owned 

septic systems, also known as the down-the-drain discharge scenario. Once Nanosilva, silver 

ions, or nanosilver reach wastewater treatment and septic systems they will most likely complex 

with sulfide and partition to biosolids. However, some fraction of the silver compounds will 
60 

reach surface water and may potentially impact aquatic organisms. Leaching of building material 

such as plastic siding and decks, and outdoor furniture and carpeting incorporating Nanosilva 

will be discharged directly into the environment. 

As stated in Section 5.2.1, silver was not found above the analytical detection limit leaching from 

plastic coupons and shirts incorporating Nanosilva. EPA does not expect unreasonable adverse 

effects to the environment from registered uses of silver ions, including the materials 

preservative use pattern (U.S. EPA, 1993). To evaluate the impacts on surface water from 

leaching of Nanosilva from plastics and textiles, EPA is assuming that nanosilver as found in 

Nanosilva is the only silver compound released. 

6.3.1 Aquatic Risk Quotient 

EPA uses a Risk Quotient (RQ) approach to assess impacts to surface water, which is similar to 

the MOE used for the human health risk assessment. The RQ is used to compare toxicity from 

environmental exposure by dividing a point estimate of exposure by a point estimate of effects. 

This ratio is a simple, screening-level estimate that identifies high- or low-risk situations. In this 

method, the estimated environmental concentration is compared to an effect level, such as an 

LC50. After the RQ is calculated, it is compared to the Agency's Level of Concern (LOC). An 

LOC is a policy tool that the Agency uses to interpret the RQ and to analyze potential risk to 

non-target organisms and the need to consider regulatory action (U.S. EPA, 2011c). 

Indoor Use

 Mass of silver release per year: 1123 kg/year. The amount of silver released from textiles 

incorporating Nanosilva was derived assuming that 300 million people (U.S. population) 

purchase one t-shirt incorporating Nanosilva each year. Each t-shirt weighs 150 grams 

and contains 0.003% nanosilver by weight silver and releases 1.6% silver per wash (see 

Section 5.2.1). 

 Release Weeks: Each t-shirt is washed once per week for 52 weeks/yr. 

 Wastewater Removal Efficiency: Ranged from 85 to 99% based on the range provided by 

Blaser et al. (2008). Wang et al. (2012a) reported nanosilver removal of 88% with 

biomass present. 

Outdoor use

 Mass of silver release per year: 570 kg/year. The amount of silver released from plastics 

incorporating Nanosilva was derived assuming that 1% of the yearly plastic lumber 

production contains Nanosilva at 0.003%. Yearly plastic lumber production for 2006 was 

projected to be approximately 1900 million kg/yr (Stutz et al., 2003). This assumes that 

all the nanosilver in plastic incorporating Nanosilva is released during a year even though 
61 

results from the plastic leaching study indicate that little silver is released from plastic 

incorporating Nanosilva. 

 Release Days: 365 days per year. This assumes that each plastic incorporating Nanosilva 

releases all its silver as nanosilver over the course of one year. 

 Wastewater Removal Efficiency: 0% to represent direct release into aquatic environments 

from outdoor use of plastics incorporating Nanosilva (i.e., no removal of nanosilver in the 

run-off due to adsorption to soil). 

Down the Drain Model

The concentration of nanosilver in surface water resulting from the use of Nanosilva in plastics 

was calculated using the Down the Drain Module of the Exposure and Fate Assessment 

Screening Tool (E-FAST model, version 2). The following input values were used: 

 Stream Dilution Factor: 1.0 or 20.1. These values are the 10th and 50th percentile values 

for the dilution that occurs during one day of lowest stream flow over a ten year period 

(1Q10) (U.S. EPA, 2007b). 

 The toxicity value for nanosilver: 1.5 μg/L based on the 96-hr EC50 value for Daphnia 

magna. 

 Level of Concern for the RQ: The presumptive level of concern (LOC) is 0.05 for listed 

(i.e. endangered or threatened) aquatic organisms and 0.5 for non-listed organisms. 

Table 20 – Risk Quotients for Nanosilver in Surface Water 

Use Scenario 

Wastewater 

Treatment 

RemovalA 

Stream 

Dilution 

Factor 

Surface Water 

Silver 

Concentration 

(μg/L) 

Acute Risk 

QuotientD

Acute RQ 

Exceeds 

LOC?* 

Indoor Use 

(t-shirts) 85% 

1.0B

 0.004 0.003 No 

20.1C 0.0002 0.0001 No 

Outdoor Use 

(plastic lumber) 0%E

1.0 0.014 0.009 No 

20.1 0.001 0.0007 No 

*The LOC is 0.05 (listed species) and 0.5 (non-listed species). RQs that exceed the LOC are of concern 

A. Silver removed from wastewater during treatment before discharge to a water body (e.g. lake, river etc.). 

B. 10th Percentile dilution factor for 1Q10 stream flow. 

C. 50th Percentile dilution factor for 1Q10 stream flow. 

D. Acute RQ = Surface Water Concentration / LC50 for Daphnia magna (1.0 μg/liter) 

E. Assumes direct discharge to surface water with no retention in soil or sediment. 
62 

The down-the-drain modeling results are shown in Table 20 and these results were divided by 

the 96-hr EC50 of 1.5 μg/L for Daphnia magna to obtain acute risk quotients (RQs). The effect 

level used to calculate the acute RQs (i.e., 1.5 ug/L) was chosen to represent the most sensitive 

aquatic organism Daphnia magna and to represent conditions that are representative of surface 

water in the United States. The acute RQ of 0.009 at the worst case stream dilution factor of 1.0 

indicates that it is unlikely that the registration of Nanosilva as a plastics and textiles preservative 

will lead to adverse effects for listed or non-listed aquatic organisms during the four year period 

of conditional registration. However, this assessment does not consider long-term exposures or 

exposure to estuarine and marine species. 

6.3.2 Conclusions for Aquatic Risk 

The acute RQs for Daphnia magna exposed to nanosilver ranged from 0.0001 to 0.009, which 

are not of concern for listed or non-listed organisms because they were less than the acute LOC 

of 0.5 and 0.05, respectively. The effect level used to calculate the acute RQs (i.e., 1.5 μg/L) was 

chosen to represent the most sensitive aquatic organism Daphnia magna and to represent 

conditions of moderately hard reconstituted water (MHRW), which is a test medium 

representative of surface water in the United States. This analysis assumes that 1% of the 

projected yearly production of plastic lumber in the U.S. would contain Nanosilva at 0.003% 

resulting in a nanosilver production-volume of 570 kg/year. This may be an overestimate given 

that the total mass of silver distributed as a material preservative in the U.S. during 2009 was less 

than 6,800 kg, based on EPA confidential records. This analysis also assumes that all the 

nanosilver in plastic lumber incorporating Nanosilva would be released into the environment 

during a one year period when in fact the leaching study for plastic coupons did not find silver at 

concentrations above the analytical detection limit implying that significantly less silver would 

be released from plastic lumber incorporating Nanosilva (see Section 5.2.1). 

There are several uncertainties regarding the exposure portion of this evaluation. The production 

volume of Nanosilva is unknown since this product is not currently available in the U.S. Because 

of this, EPA made the assumption that 1% of all plastic lumber produced every year in the U.S. 

would contain 0.003% Nanosilva. The nanosilver that could be released from the lumber 

incorporating Nanosilva was assumed to directly enter the aquatic environment without any 

removal in soil or sediments. The rate at which nanosilver entered the sanitary sewer systems 

was assumed to be constant in the down-the-drain model, which does not account for variability 

in the distribution of plastic products incorporating Nanosilva among other factors. The 

ecotoxicity values shown in Table 19 are for acute effects and mortality, and there is only one 

study on the long-term exposure to nanosilver. Finally, this assessment only considers silver that 

could be released by Nanosilva and does not include other sources of silver which will contribute 

to the environmental loading of silver. 
63 

Given that all calculated RQs were below the presumptive LOC for listed species and the 

conservative assumptions used in calculating these RQs, EPA concludes, with a level of 

confidence acceptable for the period of conditional registration, that there is a low probability of 

adverse risk to the environment from plastics and textiles incorporating Nanosilva. 

6.4 Ecological Effects and Environmental Fate Data Requirements. 

Even though EPA does not anticipate adverse ecological effects from plastics and textiles 

incorporating Nanosilva during the period of conditional registration, EPA is requesting 

additional testing to confirm its conclusions. The testing is based on a tiered approach. Tier I 

studies will determine the nature and quantity of silver released from plastics incorporating 

Nanosilva and confirm that the nanosilver in Nanosilva is consistent with the risk from Table 19. 

The Tier I environmental fate testing categories include: 

 Product Characterization and Testing 

 Silver Release from Plastics: Form, Rate and Characteristics 

Completion of these Tier I studies will provide Nanosilva product characteristics and determine 

the substance or substances released during leaching of plastics incorporating Nanosilva. 

Information from these studies will be used to substantiate Nanosilva’s claim that no nanosilver 

is released from plastics incorporating Nanosilva during use. However, if nanosilver is found to 

be released during Tier I studies, and the RQ calculated using this release rate results in a risk 

concern or is within one order of magnitude of the LOC, then Tier II studies will be required. If 

necessary, the Tier II studies will provide quantitative toxicity data on the amounts and forms of 

silver released, which will then be used by EPA to re-run the environmental risk assessment for 

Nanosilva. The Tier II environmental fate and ecological effects testing categories include: 

 Product Characterization 

 Sorption/Desorption Characteristics 

 Bioaccumulation Characteristics 

 Impacts to Wastewater Treatment 

 Wastewater Treatment Removal Efficiency 

 Aquatic Plant testing 

 Acute Effects to Freshwater Fish 

 Chronic Effects to Freshwater Fish and Invertebrates 

 Acute and Chronic Effects to Estuarine/Marine Animals 

 Chronic Effects to Sediment Dwelling Organisms 

 Terrestrial Plant Toxicity 

A more detailed description of these data requirements is provided in Appendix B. 
64 

VII. REGULATORY ACTION 

A registration for a new active ingredient may be granted under either section 3(c)(5) or section 

3(c)(7)(C) of FIFRA. A FIFRA section 3(c)(7)(C) registration is appropriate where, as here, the 

data for registration of the new active ingredient are newly required or identified. As discussed 

more thoroughly below, because EPA has not reached a final decision with regard to which types 

of data would be further required, the data requirements for this registration are considered 

newly required. 

In addition, we believe granting the proposed conditional registration with the terms and 

conditions identified in Appendix B is particularly appropriate given EPA suspects that some 

already-registered like-situated products on the market contain nanosilver as the active 

ingredient. While EPA approved these registrations without knowledge that these products may 

contain nanoscale silver and without specifically assessing any potential risks that might be 

associated with any nanosilver contained in those products, they are nonetheless on the market. 

To avoid disparate treatment and consistent with the analytic decision framework in this 

document, EPA intends to seek similar data along with comparable terms on already registered 

products that are identified to contain nanosilver. As part of this effort, EPA opened the 

Nanosilver Registration Review (docket EPA-HQ-OPP-2011-0370) on July 6, 2012, which is the 

first step towards issuing a Data Call-In for products containing nanosilver. 

7.1 Legal Framework 

The Federal Insecticide, Fungicide, and Rodenticide Act (FIFRA) section 3(c)(7)(C) provides 

that: 

The Administrator may conditionally register a pesticide containing an active ingredient not 

contained in any currently registered pesticide for a period reasonably sufficient for the 

generation and submission of required data (which are lacking because a period reasonably 

sufficient for generation of the data has not elapsed since the Administrator first imposed the 

data requirement) on the condition that by the end of such period the Administrator receives 

such data and the data do not meet or exceed risk 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 during such period will not cause any unreasonable 

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

Therefore, EPA must make four findings in order to grant a section 3(c)(7)(C) conditional 

registration for a pesticide product containing a new active ingredient: 
65 

1) Nanosilva contains an active ingredient, silver nanoparticles also known as nanosilver, 

that is not an active ingredient in any currently registered pesticide (i.e., a “new” active 

ingredient); 

2) Use of Nanosilva will not cause unreasonable adverse effects on the environment during 

the period when newly required data are being developed; 

3) Insufficient time has elapsed for Nanosilva LLC to generate and submit the newly 

required data; and 

4) Use of Nanosilva is in the public interest. 

7.2 Findings and Registration Decision 

The Agency proposing to issue a conditional, four year registration for Nanosilva in accordance 

with FIFRA section 3(c)(7)(C) and is proposing to require as a condition of registration specified 

use conditions as well as proposing that the company provide data from a number of studies that 

will allow the Agency to confirm its current assessment of the risks. Specific data requirements 

are outlined in Appendix B. Although these data requirements are specific to Nanosilva, 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. 

Consistent with Nanosilva LLC’s application, label language is required to reduce potential 

exposures. The pesticide label must include the following: 

1. The application rate must be less than 30 ppm (mg/kg) or 0.003% silver on a weight basis for 

plastics and textiles which incorporate Nanosilva. 

2. Nanosilva may only be incorporated into linear low-density polyethylene (LLDPE) plastic 

and polyethylene terephthalate (PET) based textiles consistent with the materials used in the 

leaching studies. 

3. Nanosilva may only be used to preserve plastics and textiles and may not be used for 

drinking water or food contact uses. 

4. It is required that the following mitigation measures be employed when mixing and loading 

Nanosilva during treatment of articles: 

 Closed system loading of Nanosilva containing suspension. 

 NIOSH certified full-face respirators with P100 or equivalent filter cartridges 

immediately available for use in an emergency. 

 Gloves which are chemically resistant to all of the components of the Nanosilva liquid 

suspension. 

 A long-sleeve shirt, long pants, shoes plus socks. 

5. The label must have an environmental hazard statement consisting of the following: 
66 

“This pesticide is toxic to fish and aquatic invertebrates, and birds, and can’t be used 

for irrigation purposes.” 

“Do not discharge effluent containing this product into lakes, streams, ponds, estuaries, 

oceans, or other waters unless in accordance with the requirements of a National 

Pollutant Discharge Elimination System (NPDES) permit and the permitting authority 

has been notified in writing prior to discharge. Do not discharge effluent containing this 

product to sewer systems without previously notifying the local sewage treatment plant 

authority. For guidance contact your State Water Board or Regional Office of the 

EPA”. 

7.3 Basis for Conditional Registration 

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

7.3.1 Data Generation 

Nanosilva LLC submitted its registration application thinking its product was a new pesticide 

ingredient not contained in any currently registered silver-based pesticide product and should be 

given a registration under FIFRA section 3(c)(5). At the time of application for a new chemical, 

Nanosilva LLC was required to submit all the applicable information and studies under 40 CFR 

161. Although Nanosilva LLC submitted some of the required studies using guidelines not 

modified to be applicable for testing nanosilver, they also submitted a request to waive some of 

the data requirements. In addition, Nanosilva LLC completed a leaching study using a protocol 

that was modified to be applicable for nanosilver. Although Nanosilva LLC provided EPA with 

some data in an effort to address Agency questions and concerns, EPA had not reached a final 

decision with regard to which types of data would be further required for Nanosilva. This was 

due in large 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 

insufficient time has elapsed from the point at which EPA determined and informed Nanosilva 

LLC of the data requirements needed to assess Nanosilva LLC’s application for Nanosilva LLC 

to have generated the data. 

EPA is proposing to require the tests listed below to confirm the adequacy of the 10 fold 

database uncertainty factor, to reduce the uncertainties related to differences in the physical 

properties of the nanosilver, and because there are currently no acceptable studies on the 

reproductive and developmental toxicity for nanosilver. Because this list of data requirements are 

only being proposed with today’s action, Nanosilva LLC has not yet been able to conduct these 

studies. Therefore, the Agency is proposing to require these studies as a condition of registration, 

allowing sufficient time for the study protocols to be agreed upon, the studies to be conducted, 

and for the Agency to review them. The required studies include: 
67 

 90-Day Inhalation Toxicity (Rat) (OCSPP 870.3465) modified to include in vivo bone 

marrow assay and functional observational battery, motor activity and detailed 

neuropathology 

 Reproduction/Developmental Toxicity Screening Test (Modified OCSPP 870.3550/ 

OECD TG 421) 

 Product Characterization and Testing 

 Silver Release from Plastics: Form, Rate and Characteristics 

EPA will evaluate the results from the above studies using a weight of evidence approach based 

on physical and chemical properties, toxicity, and exposure. If the Agency identifies a concern, 

then one or a combination of the following tests will be triggered: 

 Peri- and post-natal exposure to Nanosilva 

 Product Characterization 

 Sorption/Desorption Characteristics 

 Bioaccumulation Characteristics 

 Impacts to Wastewater Treatment 

 Wastewater Treatment Removal Efficiency 

 Aquatic Plant testing 

 Acute Effects to Freshwater Fish 

 Chronic Effects to Freshwater Fish and Invertebrates 

 Acute and Chronic Effects to Estuarine/Marine Animals 

 Chronic Effects to Sediment Dwelling Organisms 

 Terrestrial Plant Toxicity 

As discussed above, a listing of the studies that are proposed to be required as terms and 

conditions of this registration of Nanosilva are in Appendix B. If Nanosilva LLC does not meet 

the conditions set forth in this Decision Document, EPA will issue a notice of intent to cancel 

Nanosilva LLC’s registration under section 6(e). In addition, Nanosilva LLC’s conditional 

registration for Nanosilva will automatically expire four years after being issued. Nanosilva LLC 

should request an amendment to remove the expiration date once they have submitted the 

required data if they wish to continue to sell and distribute Nanosilva in the United States. 

7.3.2 Public Interest 

As required under FIFRA section 3(c)(7)(C), for the reasons summarized below, the Agency 

believes granting a conditional registration with the terms and conditions specified is in the 

public interest. EPA believes that use of Nanosilva offers the potential for conservation of the 

environment and consumer benefits which contribute to the public interest. These points are 

discussed in more detail below: 
68 

Conservation of the Environment

EPA has already registered a number of silver-based pesticide products for use as materials 

preservatives. All silver-based pesticide products act via 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 from the object incorporating the additive. 

The antimicrobial potency of a silver additive is therefore directly related to the potential for 

releasing silver ions. The release potential differs between various silver materials. As the size of 

silver particles decreases (e.g., 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 release) per unit mass of silver. 

Specifically, most silver-based pesticide products currently contain a silver salt, [e.g., AgCl or 

AgNO3]. Compared to the amount of silver in Nanosilva LLC’s product, most currently 

registered silver-based materials preservatives incorporate larger amounts of silver. Therefore, 

the overall potential environmental loading of silver resulting from the lower-volume use of the 

Nanosilva product should be smaller than from a comparable use of currently registered silverbased pesticides. 

Consumer Benefits

A nanosilver materials preservative should maintain its ability to reduce the number of odor and 

stain causing bacteria longer than other silver active ingredients due to the expected gradual and 

controlled release of silver ions from nanosilver as opposed to the rapid release of silver ions 

from a zeolite structure or the immediate dissolution of the silver salt. In the case of Nanosilva, 

the release of silver ions from plastics and textiles incorporating Nanosilva are at concentrations 

below the analytical detection limit while these same plastics and textiles are reported to retain 

the ability to reduce the number of odor and stain causing bacteria. While it may be that other 

silver active ingredients are present at greater silver concentrations, it is expected that their 

effects on odor and stain causing bacteria are only short-lived due to the rapid release of silver 

ions. Thus, plastics or textiles incorporating nanosilver may have longer-term ability to reduce 

the number of odor and stain causing bacteria as compared to other similar products on the 

market that contain conventional silvers. 

Innovation

EPA sees the emergence of nanotechnology as offering potential benefits for society in many 

different fields, including pest control products. The use of nanotechnology in pesticide products 

may allow for more effective targeting of pests and use of smaller quantities of pesticide. These 
69 

could contribute to improved human and environmental safety and could lower pest control 

costs. Therefore, EPA seeks to encourage innovative work to realize these benefits. 

In the case of the Nanosilva LLC’s application, EPA’s proposed conditional registration of 

Nanosilva is in the public interest in that it will allow an innovative product to reach the market. 

7.3.3 No Unreasonable Adverse Effects 

In assessing the potential risks to human health and the environment associated with the 

distribution and use of Nanosilva as a materials preservative, EPA relied on data submitted by 

Nanosilva LLC and data in the public literature on nanosilver, and use of uncertainty factors and 

conservative assumptions. As a result of this analysis and taking into account the terms and 

conditions on this conditional registration, EPA believes that the likely risks from the use of 

Nanosilva during the period of the conditional registration are small. Moreover, EPA expects the 

overall quantity of silver used in plastics and textiles as a result of this conditional registration 

will be lower than the overall quantity of silver used in other materials preservative products 

containing conventional silver resulting in expected reductions in environmental loadings of 

silver and to humans. EPA concludes that the registration would not cause unreasonable adverse 

effects on the environment during the conditional period. This conclusion is based on the 

findings discussed in the following sections. 

Risks to Human Health

As discussed above, humans could be exposed to silver ions, to Nanosilva, and to nanosilver. 

With respect to silver ions, the Agency completed a risk assessment (U.S. EPA, 1993) for silver 

salts and determined that the risks from registered uses, including the materials preservative use 

pattern, were acceptable. The Agency concludes that human exposure to silver ions from the use 

of Nanosilva and consumer exposures to plastics and textiles incorporating Nanosilva is not 

unreasonable. 

With respect to Nanosilva, the acute toxicity of Nanosilva is low by all routes of exposure. 

Because there are no subchronic or chronic oral or dermal toxicity studies available for 

Nanosilva or on the nanosilver that might break away from products incorporating Nanosilva, the 

subchronic or chronic toxicities were estimated using studies on nanosilvers reported in the 

literature. The Agency relied on the results from studies submitted by Nanosilva LLC where the 

liquids in contact with plastic and shirts incorporating Nanosilva were analyzed for silver content 

to estimate exposure the amount of Nanosilva and nanosilver that might be released from plastics 

and textiles. The Agency recognized the uncertainties and incorporated multiple uncertainty 

factors and conservative assumptions when calculating the Margin of Exposures (MOEs) for 

estimating the risk to children from plastics and textiles incorporating Nanosilva. 
70 

Based on all of the foregoing, EPA concludes, with a level of confidence acceptable for the 

period of conditional registration, that there is a low probability of adverse risk to consumers 

from short- and intermediate-term exposure to toys, flooring, and textiles incorporating 

Nanosilva. Thus, the Agency concludes that use of Nanosilva will not cause unreasonable 

adverse effects on the environment during the period when newly required data are being 

developed. 

Risks to the Environment 

The environment could be exposed to silver ions, to Nanosilva, and to nanosilver. With respect 

to silver ions, the Agency completed a risk assessment (U.S. EPA, 1993) for silver salts and 

determined that the risks from registered uses, including the materials preservative use pattern, 

were acceptable. The Agency concludes that environmental exposure to silver ions from the use 

of Nanosilva and plastics and textiles incorporating Nanosilva is acceptable. 

With respect to the fate and ecotoxicity of Nanosilva, because there are no fate or ecotoxicity 

studies available for Nanosilva or the nanosilver that might break away from plastics or textiles 

incorporating Nanosilva, the fate and ecotoxicity were estimated using studies on nanosilvers 

reported in the scientific literature. Direct release from leaching of plastics and laundering of 

textiles incorporating Nanosilva were anticipated to be the primary route by which Nanosilva, 

silver ions, and nanosilver reach the environment. The Agency assessed impacts to surface 

waters assuming that nanosilver was the only compound released. Environmental exposure from 

indoor use was assessed assuming that 300 million people (U.S. population) each purchased one 

t-shirt containing 0.003% nanosilver where 1.6% of the silver in the t-shirt incorporating 

Nanosilva was released during each weekly washing over a one year period. Environmental 

exposure from outdoor use was assessed assuming that 1% of the projected yearly production of 

plastic lumber in the U.S. would contain Nanosilva at 0.003% where all the silver would be 

released into the environment as nanosilver during a one year period even though the leaching 

study for plastic coupons incorporating Nanosilva did not find silver at concentrations above the 

analytical detection limit. EPA estimates, derived from down-the-drain modeling, of the 

concentrations of nanosilver resulting from the use of Nanosilva do not exceed the Agency’s 

estimate of the highest concentration of nanosilver in surface water to which an aquatic 

community can be exposed without resulting in an unacceptable effect. Thus, the Agency 

concludes that use of Nanosilva will not cause unreasonable adverse effects on the environment 

during the period when newly required data are being developed. 
71 

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Critical Care 60:648-652. 

Van der Zande et al. 2012. Distribution, Elimination, and Toxicity of Silver Nanoparticles and 

Silver Ions in Rats after 28-Day Oral Exposure. ACS Nano DOI: 10.1021/nn302649p. 

U.S. EPA, 1993. RED Fact Sheet for Silver, EPA-738-F-93-005. EPA Office of Pesticide 

Programs, June 1993. 

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

1998, available at: http://www.epa.gov/pesticides/science/phedsur-expo-guide.pdf

U.S. EPA, 2000. Assigning Values to Non-Detected/Non-Quantified Pesticide Residues in 

Human Health Food Exposure Assessments. Available at: 

http://www.epa.gov/pesticides/trac/science/trac3b012.pdf 

U.S. EPA, 2001. General Principles For Performing Aggregate Exposure And Risk Assessments. 

November 28, 2001. Available at: http://www.epa.gov/opp00001/trac/science/aggregate.pdf
76 

U.S. EPA, 2002. A review of the Reference Dose and Reference Concentration Processes. 

EPA/630/P-02/002F. December. 

U.S. EPA. 2005. Guidance on Selecting Age Groups for Monitoring and Assessing Childhood 

Exposures to Environmental Contaminants. EPA/630/P-03/003F. November, 2005. 

U.S. EPA, 2007a. Method 6010C, Inductively Coupled Plasma-Atomic Emission Spectrometry. 

In EPA SW846: Test Methods for Evaluating Solid Waste, Physical/Chemical Methods. 

U.S. EPA, 2007b. Exposure and Fate Assessment Screening Tool (E-FAST), Version 2.0, 

Documentation Manual. October, 2007. Available at: 

http://www.epa.gov/oppt/exposure/pubs/efast2man.pdf 

U.S. EPA. 2008. Child-Specific Exposure Factors Handbook. National Center for Environmental 

Assessment. Washington, DC; EPA/600/R-06/096F. September. 

U.S. EPA. 2011a. Decision Document, Conditional Registration of HeiQ AGS-20 as a Materials 

Preservative in Textiles. December 1, 2011. 

U.S. EPA. 2011b. Exposure Factors Handbook: 2011 Edition. EPA/600/R-090/052F. September, 

2011. 

U.S. EPA, 2011c. Technical Overview of Ecological Risk Assessment, Risk Characterization. 

Risk Presumptions for Aquatic Animals. Available at: 

http://www.epa.gov/oppefed1/ecorisk_ders/toera_risk.htm

U.S. EPA, 2012a. Science Review of the AEATF II Liquid Pour Human Exposure Monitoring 

Study. October 9. Available at: http://www.epa.gov/hsrb/files/meeting-materials/nov-1-2012-

public-meeting/AEATF-Liquid-Pour-Study.pdf 

U.S. EPA, 2012b. Standard Operating Procedures for Residential Pesticide Exposure 

Assessment. Health Effects Division, Office of Pesticide Programs, Office of Chemical Safety 

and Pollution Prevention. 

Wan, A.T., Conyers, R.A.J., Coombs, C.J., et al. 1991. Determination of Silver in Blood, Urine, 

and Tissues of Volunteers and Burn Patients. Clinical Chemistry 37:1681-1687. 

Wang, Z., Liu, S.J., Ma, J. et al. 2013. Silver Nanoparticles Induced RNA Polymerase-Silver 

Binding and RNA Transcription Inhibition in Erythroid Progenitor Cells. ACS Nano 7: 4171-

4186. 

Wang, Y., Westerhoff, P., Hristovski, K.D. 2012a. Fate and biological effects of silver, titanium 

dioxide, and C60 (fullerene) nanomaterials during simulated wastewater treatment processes. 

Journal of Hazardous Materials 201-202:16-22 
77 

Wang, Z, Chen, J., Li, J. et al. 2012b. Aquatic Toxicity of Nanosilver Colliods to Different 

Trophic Organisms: Contributions of Particles and Free Silver Ion. Environmental Toxicology 

and Chemistry 31: 2408-2413. 

Wigginton, N.S., De Titta, A., Piccapietra, F., et al. 2010. Binding of Silver Nanoparticles to 

Bacterial Proteins Depends on Surface Modifications and Inhibits Enzymatic Activity. 

Environmental Science Technology 44:2163. 

Wu, Y., Zhou, Q. 2012. Silver Nanoparticles Cause Oxidative Damage and Histological Changes 

in Medaka (Oryzias latipes) after 14 days of Exposure. Environmental Toxicology and 

Chemistry, 32:165-173. 

Yang, Y., Chen, Q., Wall, J.D., Hu, Z. 2012. Potential nanosilver impact on anaerobic digestion 

at moderate silver concentrations. Water Research 46: 1176-1187. 
Appendix A - Page 1

Appendix A –

Evaluation of Nanosilva LLC’s Data Waiver Requests

August 27, 2013
Appendix A - Page 2

I. Introduction

On August 10, 2009, Nanosilva LLC submitted an application to register Nanosilva as a new 

active ingredient (PRIA A420). In 2009, as an applicant for registration of a new active 

ingredient, Nanosilva LLC was required to submit all the applicable information and studies 

under 40 CFR part 161. EPA published a Notice of Proposed Rulemaking on October 8, 2008

proposing to revise and update the existing data requirements under 40 CFR part 161 for 

antimicrobial pesticides. These data requirements were made final as 40 CFR part 185W on May 

8, 2013 (78 FR 26936). In 2009, the Nanosilva LLC application included a request to waive the 

requirement to generate data for Nanosilva using the following test guidelines:

1. Hydrolysis (OPPTS 835.2120)

2. Aquatic Invertebrate Acute Toxicity Test, Freshwater Daphnids (OPPTS 850.1010)

3. Fish Acute Toxicity Test, Freshwater and Marine (OPPTS 850.1075)

4. Avian Acute Oral Toxicity Test (OPPTS 850.2100)

5. 90-Day Dermal Toxicity (OPPTS 870.3250)

6. Prenatal Developmental Toxicity Study ( OPPTS 870.3700)

7. Bacterial Reverse Mutation Test (OPPTS 870.5100)

8. Detection of Gene Mutations in Somatic Cells in Culture (OPPTS 870.5300)

9. In Vitro Mammalian Cytogenetics (OPPTS 870.5375)

10. Mammalian Bone Marrow Chromosome Aberration Test (OPPTS 870.5385)

11. Immunotoxicity (OPPTS 870.7800)

The test guidelines in items 1 through 4 are cited in 40 CFR part 161 and were required at the 

time Nanosilva applied to registered Nanosilva. Guidelines in items 5 through 11 were proposed 

in 2008 to be included in 40 CFR part 158W, which was made final in 2013 and therefore, the 

test guidelines in items 5 through 11 were not required at the time Nanosilva LLC applied to 

register Nanosilva. Even though these guidelines were not required at the time of application, 

EPA evaluated the request to waive each of these data requirements.

Waivers for the above studies were requested because of the lack of acute toxicity noted in the 

six acute toxicity studies (MRID 47828918 through 47828923), because Nanosilva has a specific 

gravity greater than water, because Nanosilva has low solubility in water as determined by the 

water column generator method (MRID 47828917), and due to the non-leaching characteristic of 

Nanosilva because silver at concentrations greater than the analytical detection limit were not 

detected in leaching studies conducted with a plastic incorporating Nanosilva (MRID 47828925).

The data requirements listed above in items 1 through 11 are either waived or satisfied. In 

general, the data requirement is met or satisfied because:

• certain guideline studies are not appropriate for nanosilver and where appropriate are 

being modified and required as terms and conditions on the registration (i.e., 

Reproduction/Developmental Toxicity Screening Test)
Appendix A - Page 3

• certain environmental data are satisfied because the environmental hazard labeling 

statement was determined using literature studies completed with nanosilver

• Dermal guideline studies are not appropriate given there was no risk concern

• in vitro mutagenicity study requirements are satisfied and have shown that nanosilver is 

possibly mutagenic

• the immunotoxicity study is initially waived and only triggered after the potential 

immune toxicity of nanosilver is determined during the 90-day inhalation toxicity study

required as a term and condition on the registration

Each of the above studies is addressed in the Environmental Fate, Wildlife and Aquatic 

Organism, and Toxicology data requirements listed in 40 CFR part 161 for antimicrobial 

pesticides. The following sections contain an evaluation of each waiver request.

A. Environmental Fate Data Requirements

The following test for environmental fate is required for antimicrobials under 40 CFR 161.202 to 

assess the exposure to pesticides after application:

1. Hydrolysis, OPPTS 835.2120

This test is waived because nanosilver undergoes dissolution with the generation of ionic silver 

rather than undergoing hydrolytic transformation.

B. Wildlife and Aquatic Organism Data Requirements

The following tests are required for solid-formulation manufacturing use antimicrobials under 40 

CFR 161.440:

2. Aquatic Invertebrate Acute Toxicity Test, Freshwater Daphnids (OPPTS 850.1010)

3. Fish Acute Toxicity Test, Freshwater and Marine (OPPTS 850.1075)

4. Avian Acute Oral Toxicity Test (OPPTS 850.2100)

The acute ecotoxicity testing waiver request was based on the non-leaching characteristic of 

Nanosilva. However, the results of these tests are required for use in determining the appropriate 

precautionary labeling on packages containing the Nanosilva liquid suspension prior to 

incorporating into a product. Therefore, the non-leaching characteristic is not an appropriate 

basis for waiving these tests. As summarized in Table 19 of the Decision Document, EPA has 

information on the acute toxicity of nanosilver for Daphnids and Freshwater Fish. Because the 

EC50 values were less than 1 mg/L, as indicated in Chapter 8 of the Pesticide Label Review 

Manual, the following environmental hazard statement is required on the Nanosilva label:
Appendix A - Page 4

This pesticide is toxic to fish and aquatic invertebrates.

Given that there are no test results for Avian species available for nanosilver, EPA requires that 

the environmental hazard statement for the Nanosilva label be expanded to include:

This pesticide is toxic to fish and aquatic invertebrates, and birds.

EPA is requiring that Nanosilva LLC place the above environmental hazard statement on the 

pesticide label for Nanosilva. However, if Nanosilva LLC wishes to remove birds from this 

statement then they can submit results from an Avian Acute Oral Toxicity Test (OPPTS 

850.2100) showing that the nanosilver in Nanosilva is non-toxic to birds with an LD50 of greater 

than 100 mg/kg.

In addition, Nanosilva LLC is required to place the following statement on the label for 

Nanosilver liquid suspension:

“Do not discharge effluent containing this product into lakes, streams, ponds, estuaries, 

oceans, or other waters unless in accordance with the requirements of a National Pollutant 

Discharge Elimination System (NPDES) permit and the permitting authority has been 

notified in writing prior to discharge. Do not discharge effluent containing this product to 

sewer systems without previously notifying the local sewage treatment plant authority. For 

guidance contact your State Water Board or Regional Office of the EPA”.

Thus, the data requirements in items 2 through 4 are satisfied by data in the public literature for 

nanosilver and required labeling.

C. Toxicology Data Requirements

The following tests are required for antimicrobials under 40 CFR part 158.2230 (i.e., part 

158W):

5. 90-Day Dermal Toxicity (OPPTS 870.3250

6. Prenatal Developmental Toxicity Study (OPPTS 870.3700)

The requirement for the 90-Day Dermal Toxicity study is waived because the calculated Margin 

of Exposures (MOEs) for dermal and incidental exposure to products incorporating Nanosilva 

where greater than 1,000,000 as compared to the highest target MOE of 3,000. However, if 

nanosilver is determined to be released during the leaching test of plastics incorporating 

Nanosilva in amounts well above those found in the plastics leaching study already submitted 

such that the MOE calculated using the newly determined nanosilver release rate results in a risk 
Appendix A - Page 5

concern, then the following route-specific subchronic tests is proposed to be required to confirm

the risk assessment for the nanosilver in Nanosilva.

• Dermal penetration study in rats (OSCPP 870.7600)

The requirement for the Prenatal Developmental Toxicity Study is waived because the following 

study has been determined to be more appropriate for Nanosilva:

• Modified 870.3550/ OECD TG 421: Reproduction/Developmental Toxicity Screening 

Test

The following tests are required for pesticides under 40 CFR part 158.2230 (e.g., 158W):

7. Bacterial Reverse Mutation Test (OPPTS 870.5100)

8. Detection of Gene Mutations in Somatic Cells in Culture (OPPTS 870.5300)

9. In Vitro Mammalian Cytogenetics (OPPTS 870.5375)

10. Mammalian Bone Marrow Chromosome Aberration Test (OPPTS 870.5385)

11. Immunotoxicity (OPPTS 870.7800)

The mutagenicity studies [Bacterial Reverse Mutation Test (OPPTS 870.5100), Detection of 

Gene Mutations in Somatic Cells in Culture (OPPTS 870.5300), In Vitro Mammalian 

Cytogenetics (OPPTS 870.5375), Mammalian Bone Marrow Chromosome Aberration Test

(OPPTS 870.5385)] are satisfied because there are in vitro studies available for nanosilver in the 

scientific literature which address/satisfy these data needs and show that nanosilver is possibly 

mutagenic. Thus, the Agency is requiring as a term and condition on the registration an in-vivo

bone marrow assay to clarify the mutagenic properties of the nanosilver in Nanosilva.

The immunotoxicity (OPPTS 870.7800) is waived at this time because OPP has published a 

document (U.S. EPA, 2013) recommending that immune related endpoints be evaluated during 

the inhalation toxicity test, required as a term and condition on the registration, before 

determining if the immunotoxicity test is required. The currently available oral toxicity studies 

indicate that nanosilver causes liver and kidney toxicity in laboratory animals where silver is 

distributed to all organs and tissues with accumulation of silver in the brain and male animal 

testes. Inhalation toxicity studies also identified liver toxicity as well as lung effects including 

chronic alveolar inflammation. The toxicology database for nanosilver does not reveal any 

evidence of treatment-related effects on the immune system, which suggests that nanosilver does 

not directly target the immune system.

U.S. EPA, 2013. Part 158 Toxicology Data Requirements: Guidance for Neurotoxicity Battery, 

Subchronic Inhalation, Subchronic Dermal and Immunotoxicity Studies. May 1st. Available at: 

http://www.epa.gov/pesticides/regulating/part158-tox-data-requirement.pdf
Appendix B - Page 1

Appendix B –

Data Requirements, Enforceable Schedule, and 

Conditions of the Proposed Conditional Registration 

for Nanosilva

August 27, 2013
Appendix B - Page 2

I. Introduction

Nanosilva is a liquid suspension containing 1% nanosilver by weight where the nanosilver

active-ingredient is covalently bonded to crystalline silica via a thiolate bond. The diameter of 

the spherical silica core is 320 nm on average (Lee et al., 2006) and the nanosilver particles that 

are attached to the silica core have mean diameters between 6.9 and 10.6 nm (Lee et al., 2007). 

The complex is formed by reacting spherical silica particles modified with thiol groups with 

silver nitrate and polyvinylpyrrolidone (PVP) in ethanol (Lee et al. 2007). Thus, the resulting 

liquid suspension contains 1% nanosilver by weight where the nanosilver surface is coated with 

sulfur and PVP, and is attached to silica. Reference to the nanosilver in Nanosilva (whether when 

part of the composite or when broken away from the complex) is to the “new active ingredient” 

and reference to “Nanosilva” or the Nanosilva “complex” is to the end-use product. 

II. Exposure Pathways

The human and environmental exposures resulting from the use of Nanosilva, and use and 

disposal of plastics and textiles incorporating Nanosilva will largely be a function of what 

materials are available for inhalation or dermal exposure during treatment of plastics and textiles 

or what materials leach or break away from the plastics and textiles incorporating Nanosilva 

during use and disposal. EPA anticipates that humans and the environmental will potentially be 

exposed to the following materials:

1. Silver ions released from Nanosilva; 

2. Nanosilva complex; and/or

3. Nanosilver that might break away from the Nanosilva complex.

EPA expects that occupational inhalation and dermal exposures to Nanosilva and the 

nanoparticles that might break away from Nanosilva are likely to occur during the following use 

scenarios:

1. Mixing and loading of Nanosilver during preparation of a master batch

2. Mixing, loading, and applying the Nanosilva containing master batch during treatment of 

plastics and textiles

3. Handling plastics and textiles incorporating Nanosilva 

EPA expects consumer exposures to Nanosilva, the silver ions derived from the Nanosilva 

complex, and the nanoparticles that break away from the Nanosilva complex could potentially 

occur during the following use scenarios:

1. Incidental oral exposure to plastics and textiles incorporating Nanosilva

2. Dermal exposure to plastics and textiles incorporating Nanosilva

Leaching of plastics and textiles incorporating Nanosilva is anticipated to be the primary route 

by which Nanosilva, silver ions, and nanosilver reach the environment. The silver released 
Appendix B - Page 3

during leaching of plastics and textiles incorporating Nanosilva could be discharged to the 

sanitary sewer system leading to publically owned wastewater treatment and privately owned 

septic systems, also known as the down-the-drain discharge scenario. Leaching of building 

material such as plastic siding and decks and outdoor furniture incorporating Nanosilva will be 

discharged directly into the environment. Once Nanosilva, silver ions, or nanosilver reach 

wastewater treatment and septic systems they will most likely complex with sulfide and partition 

to biosolids. However, some fraction of the silver compounds will reach surface water and may 

potentially impact aquatic organisms. 

III. Data Needed to Confirm the Estimates for Risks of Exposure to Nanosilva

As a condition of the registration, EPA is requiring the Nanosilva LLC to conduct a number of 

studies, based on a tiered approach, which will allow the Agency to confirm the findings of the 

risk assessment completed for the proposed conditional registration and discussed in the decision 

document. These tests include Tier I studies to determine the nature and quantity of silver 

released from plastics incorporating Nanosilva under conditions of use, product characterization 

testing, and silver release characteristics from the Nanosilva complex. If nanosilver is determined 

to be released from plastics during the leaching tests as part of Tier I testing and the weight of 

evidence based on physical and chemical properties, toxicity, and exposure result in a risk 

concern then route-specific toxicity and environmental fate tests as part of Tier II studies to 

confirm the nanosilver risk assessment will be triggered. Data must be submitted within four 

years and according to the schedule provided in Table 3B to avoid cancellation of the conditional 

registration. 

The duration of four years was chosen to allow time for protocol reviews prior to initiation of the 

studies, completion 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 confirm the Agency’s determination that the product is not expected to cause 

unreasonable adverse effects to human health and the environment. If EPA determines that 

Nanosilva LLC has failed to take appropriate steps to initiate the required studies, or failed to 

submit the protocols or studies, as required pursuant to this Appendix , EPA will issue a notice of 

intent to cancel Nanosilva’s registration under FIFRA section 6(e). 

The following factors were considered in developing the data requirements for Nanosilva:

• Submitted as a new active ingredient application, the Nanosilva nanosilver was 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, nontarget 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 use pattern. Information provided by Nanosilva LLC and information from the 

literature was used to tailor the data requirements to the proposed use pattern.

• Additional studies in the area of physical and chemical characterization that are not 
Appendix B - Page 4

specifically included in 40 CFR Part 161 are needed because Nanosilva contains 

nanosilver. These studies are needed because nanosized materials may 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 (FIFRA SAP, 2009) and 

by the MINChar Initiative (2008).

In addition, the following recommendations from the FIFRA SAP report were considered in the 

development of the data requirements as terms and conditions on the proposed conditional 

registration of Nanosilva:

• Both nano-sized particles of silver (Ag) as well as ionic silver (Ag+

) can contribute to 

toxic effects of nanosilver. The rate of silver ion production, as well as the distribution of 

nanosilver in tissues and the environment, may differ substantially between nanosilver

and other forms of silver, as nanosilver can potentially deliver silver ions directly to 

specific tissues, cell membranes or inside cells – places where other forms of silver 

compounds cannot reach. Therefore, the hazard profile of nanosilver may differ from 

other forms of silver.

• Particle size can substantially impact particle 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. 

• 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 registration of Nanosilva is included in Tables 1B

and 2B. The studies included in Table 1B are considered to be Tier I, meaning that their need is 

not based on the results of other studies. The studies listed in Table 2B are considered to be Tier 

II, because they may or may not be required depending upon the results of the Tier I studies. The 

Nanosilva complex and plastics/textiles incorporating Nanosilva will be the test material during 

Tier I studies and the test material for Tier II studies will depend on the results of the Tier I 

leaching and dissolution studies. Figure 1B contains a conceptual diagram outlining the Tier I 

and Tier II testing approach.
Appendix B - Page 5

Figure 1B - Test material and tiered approach for Nanosilva and plastic incorporating 

Nanosilva. See Tables 1B and 2B for the specific tests required for each material.

Tier II - Testing

Nanosilva Complex

Environmental Fate

Tier II – Testing

Silver Nanoparticle

Product Chemistry

Health Effects

Ecological Effects

Environmental Fate

Tier I - Testing

Nanosilva Complex

Product Chemistry

• Ultraviolent-Visible Absorption

• Particle Size Distribution

• Particle Surface Area

• Particle Stability

Occupational Inhalation Exposure

• 90-Day Inhalation Toxicity with Neuro and Muta

• Repro/Developmental Screening by Inhalation Route

Tier I - Testing

Nanosilva Treated Plastic

Leaching Study

Weight of evidence is 

based on physical and 

chemical properties, 

toxicity, and exposure.

Low risk potential, high 

confidence in available data, 

potential for no further testing.

Additional characterization of 

toxicity profile needed.
Appendix B - Page 6

IV. Non Data-Related Terms and Conditions of Registration

Based on Nanosilva LLC’s original application, amendments to the proposed label language are 

required. The pesticide label must include the following:

1. The application rate must be stated as being less than 30 ppm (mg/kg) or 0.003% silver 

on a weight basis for plastics incorporating Nanosilva.

2. Nanosilva may only be incorporated into linear low-density polyethylene (LLDPE) 

plastic and polyethylene terephthalate (PET) based textiles consistent with the materials 

used in the leaching studies.

3. Nanosilva may only be used to preserve plastics and textiles and may not be used for 

drinking water or food contact uses.

4. It is required that the following mitigation measures be employed when mixing and 

loading Nanosilva during treatment of articles: 

• Closed system loading of Nanosilva containing suspension.

• NIOSH certified full-face respirators with P100 or equivalent filter cartridges

immediately available for use in an emergency.

• Gloves which are chemically resistant to all of the components of the Nanosilva 

liquid suspension.

• A long-sleeve shirt, long pants, shoes plus socks.

5. The label must have an environmental hazard statement consisting of the following:

“This pesticide is toxic to fish and aquatic invertebrates, and birds, and can’t be used 

for irrigation purposes.”

“Do not discharge effluent containing this product into lakes, streams, ponds, estuaries, 

oceans, or other waters unless in accordance with the requirements of a National 

Pollutant Discharge Elimination System (NPDES) permit and the permitting authority 

has been notified in writing prior to discharge. Do not discharge effluent containing this 

product to sewer systems without previously notifying the local sewage treatment plant 

authority. For guidance contact your State Water Board or Regional Office of the 

EPA”.

V. Enforceable Schedule

Before EPA will issue a final determination on this application, Nanosilva LLC must provide 

information to satisfy the outstanding guideline items listed in Table 1B under Product 

Chemistry. EPA will not issue a registration until all Product Chemistry information is submitted 

by Nanosilva LLC and accepted by the EPA.
Appendix B - Page 7

EPA has prepared an enforceable schedule that is presented in Table 3B. This schedule is an 

estimation of the time required for developing and submitting protocols for review, conducting 

the studies, and submitting the resulting data, as well as EPA’s review of the submitted data. 

However, unforeseen technical issues may arise due to the unique nature of Nanosilva (a 

difficult-to-test substance), which may cause a delay in testing. If such a case arises, Nanosilva 

LLC shall submit a written request justifying the nature of the delay. In addition to technical 

delays, there may be delays by EPA in reviewing protocols and data submitted by Nanosilva 

LLC. In this case, the EPA shall submit a written statement to Nanosilva LLC outlining the 

nature of the delay. In either case, if EPA determines a delay in the enforceable schedule is 

appropriate, it will amend the terms and conditions on the conditional registration. 

Notwithstanding technical delays or delays in reviewing data, if Nanosilva LLC fails to take 

appropriate steps to initiate the required studies, or fails to submit the protocols or data, as 

required pursuant to this Appendix, EPA will issue a notice of intent to cancel Nanosilva LLC’s 

registration under FIFRA section 6(e). Specifically, Nanosilva LLC shall:

1. Submit protocols modified for Nanosilva LLC to EPA’s satisfaction for each of the 

data requirements listed in Table 1B. 

2. Perform each test and submit the results from each test as described in Table 1B and 

2B. 

These items shall be submitted according to the schedule provided in Table 3B. If EPA 

determines that Nanosilva LLC has failed to initiate or to submit the required studies by the dates 

indicated in Table 3B, then EPA will issue notice of intent to cancel Nanosilva LLC’s 

registration under FIFRA section 6(e). In addition, Nanosilva LLC’s conditional registration for 

Nanosilva will automatically expire four years after being issued. Nanosilva LLC will have to 

submit an application for an unconditional registration if they wish to continue to sell and 

distribute Nanosilva in the United States.

EPA will use this data to confirm EPA’s determination that the conditional registration of 

Nanosilva will not cause unreasonable adverse effects on the environment, taking into account 

the terms and conditions on the registration.

All tests shall be conducted using laboratories that comply with the Good Laboratory Practice 

(GLP) standards of 40 CFR Part 160. In accordance with those regulations, the pesticide testing 

facility and the sponsor must meet GLPs in the following areas:

1. Organization and personnel

2. Facilities

3. Equipment

4. Testing facilities operation

5. Test, control, and reference substances

6. Protocol for and conduct of a study

7. Records and reports
Appendix B - Page 8

Thus, the EPA expects that all studies performed to support, among other things, the registration 

or continued registration of a pesticide will address those requirements. However, studies 

deviating from those requirements may be considered if, as set forth in 40 CFR 160, a statement 

describing in detail all differences between the practices used in the study and those required by 

40 CFR Part 160 is provided [see 40 CFR Part 160.12(b)]. As part of its review of the subject 

study, EPA will evaluate any such differences in determining if the study is acceptable for use in 

supporting, among other things, product registration or continued registration.

VI. Time-Limited Conditional Registration

The conditional registration for Nanosilva will automatically expire four years after being issued. 

Nanosilva LLC should request an amendment to remove the expiration date once they have 

submitted the required data if they wish to continue to sell and distribute Nanosilva in the United 

States.

References:

FIFRA SAP, 2009. FIFRA Scientific Advisory Panel Meeting held November 3 - 5, 2009 on the 

Evaluation of Hazard and Exposure Associated with Nanosilver and Other Nanometal Pesticide 

Products Available at:

http://www.epa.gov/scipoly/sap/meetings/2009/november/110309ameetingminutes.pdf

Lee, J.M., Kim, D.W., Jun, Y.D., Oh, S.G. 2006. Preparation of silica-silver heterogeneous 

nanocomposite particles by one-pot preparation strategy using polyol process: Size-controlled 

immobilization of silver nanoparticles. Materials Research Bulletin 41:1407-1416.(MRID 

48379901 and 48379903)

Lee, J.M., Kim, D.W., Kim, T.H., Oh, S.G. 2007. Facile route for preparation of silica-silver 

heterogeneous nanocomposite particles using alcohol reduction method. Materials Letters 

61:558-1562. (MRID 48379902 and 48379904)

The Minimum Information for Nanomaterial Characterization (MINChar) Initiative, 2008. 

Available at: http://characterizationmatters.files.wordpress.com/2008/11/mincharparameters-list.pdf
Appendix B - Page 9

Table 1B - Summary of Tier I Required Data for Nanosilva

OSCPP Data Requirement (Note 1)

Guideline Number: Study Title Reason for Study Test Material Comments

Product Chemistry

830.1620: Production Process

Required for antimicrobial 

pesticides per 40 CFR part 

158. Nanosilva

Completely describe quantities of reactants, reaction conditions, and reaction 

time.

830.1650: Formulation Process Completely describe details regarding the formulation of the master batch.

830.1670: Formation of Impurities Describe all possible reaction by-product impurities.

830.1700: Preliminary Analysis Distinguish between the amount of silver and silver ions present.

830.1800: Analytical Method ICP-AES only provides amount of silver present, also need method to quantify 

the amount of nanosilver present.

830.1900: Submittal of Samples Nanosilva LLC must provide samples of Nanosilva for EPA use in verifying 

analysis and identity of Nanosilva.

830.6314: Oxidation/Reduction; Chemical 

Incompatibility

MSDS states that halogen salts and chlorides should be avoided; however, 

Nanosilva LLC states that the Nanosilva complex is stable. Need to fully 

describe stability of Nanosilva to other chemicals.

830.6317: Storage Stability Only the concentration of silver was determined. Need to describe changes in 

particle size and UV/Visible spectroscopy with storage time.

830.7050: UV-Visible Light Adsorption UV/Visible spectroscopy is required to evaluate the assessment of the structural 

integrity of the active ingredient.

Nanosilva Characterization

Non-Guideline: Particle Size and Diameter 

(size) Distribution

Required to characterize 

product. Determine both the size of the silica core particle and the nanosilver particles.
Appendix B - Page 10

Table 1B - Summary of Tier I Required Data for Nanosilva

OSCPP Data Requirement (Note 1)

Guideline Number: Study Title Reason for Study Test Material Comments

Non-Guideline: Surface Area Determination Required to characterize 

product.

Determine both the surface area of the silica core particle and the nanosilver 

particles.

Non-Guideline: Particulate Stability Required to assess stability of 

Nanosilva. Nanosilva

Nanosilva LLC claims that Nanosilva is a stable complex that does not release 

nanosilver. This test should be designed to demonstrate Nanosilva stability to 

mechanical stress (normal and shear) and common reactants and solvents. EPA 

will use this information to evaluate Nanosilva LLC claims of Nanosilva 

stability. 

Human Exposure

Non-Guideline: Plastic Leaching Study

(Confirm Nanosilva LLC study results)

Required to confirm 

assessment on human and 

environmental exposure.

This test will determine what 

test substance will be used 

for Tier II studies.

Nanosilva

Treated 

Plastics

A leaching study is needed to determine the quantity and form of silver that is 

released from Nanosilva treated plastics under conditions of use. Nanosilva 

LLC submitted a leaching study using food simulants and determined the total 

silver concentration. EPA requires an additional, follow-up study using 

physiological fluids and methods to determine the form of silver released from 

Nanosilva treated plastic. The exposure data will be used in to confirm the 

Agency’s evaluation of the risk of exposure to Nanosilva.

Health Effects

870.3465: 90-Day Inhalation Toxicity (Rat)

(Replace Song et al., 2012 study results)

Modified to include in vivo bone marrow 

assay and functional observational battery, 

motor activity and detailed neuropathology

Required to confirm the 

adequacy of the 10 fold 

database uncertainty factor, to 

reduce the uncertainties 

related to differences in the 

physical properties of the 

nanosilver.

Nanosilva The inhalation study is a route-specific study required to evaluate the effects of 

inhaling nanosilver when mixing and loading Nanosilva. 

Modified 870.3550/ OECD TG 421: 

Reproduction/Developmental Toxicity 

Screening Test1

Required because there are 

currently no acceptable studies 

on the reproductive and 

developmental toxicity for 

nanosilver.

Occupational exposure to 

workers of reproductive age. 

Nanosilva 

The combined repeated-dose toxicity study with the 

reproduction/developmental toxicity screening test will provide initial 

information on possible effects on reproduction and/or development. In 

addition, the study may also provide a toxicity endpoint applicable to a risk 

assessment for oral incidental exposure.

Ecological Effects – No Tier I Studies Required
Appendix B - Page 11

1

The use of a combined study that utilizes the 2-generation reproduction study in rodents as a basic protocol for the addition of other 

endpoints or functional assessments in the immature animal is encouraged. 

Table 2B - Summary of Tier II Required Data for Nanosilva

OSCPP Data Requirement (Note 1)

Guideline Number: Study Title Reason for Study Test Material Comments

Product Chemistry

830.7050: UV-Visible Light Adsorption

These studies are required to 

characterize the nanosilver if released 

during Tier I stability, dissolution or 

leaching studies.

Nanosilver

Non-Guideline: Particle Size and Diameter 

(size) Distribution

Non-Guideline: Surface Area Determination

830.7840: Solubility

Non-Guideline: Zeta Potential and Surface 

Charge Determination

Human Exposure – No Tier II Studies are Required

Health Effects

Non-Guideline: peri- and post-natal 

exposure to Nanosilva

Children’s exposure to nanosilver while 

contacting Nanosilva treated plastics

and textiles. Occupational exposure to 

nanosilver for workers of child bearing 

age.

See Note 2

Based on the results of the plastics leaching study, the Agency may 

require additional toxicity data to assess potential adverse health 

outcomes in young children resulting from incidental oral exposure to 

Nanosilva-treated plastic toys (i.e., children chewing on toys).

Ecological Effects

850.1850: Modified Aquatic Food Chain 

Transfer

Required to determine bioavailability 

and biomagnifications. See Note 2

Traditionally use Fish and Oyster BCF to estimate bioaccumulation for 

chemicals, however, mesocosm tests more likely to yield useful 

information.

850.4100: Terrestrial Plant Toxicity

Required to determine effects to plants 

during early critical stages in their 

development.

See Note 2

Nanosilver is likely to partition to biosolids during wastewater 

treatment. If those biosolids are then used in land farming, nanosilver

may impact growth of plants in farm fields.

850.4400: Aquatic Plant Toxicity, Tier 2 Required to determine the toxicity to 

freshwater and aquatic plants. See Note 2

Aquatic plants are the primary source of cellular carbon and chemical 

energy for aquatic environments. Impacts to these primary producers 

would have broad implications for the aquatic food chain.

850.5400: Algal Toxicity, Tier 2 Required to determine the phytotoxicity 

to freshwater and marine algae. See Note 2

Algae are the primary source of cellular carbon and chemical energy for 

aquatic environments. Impacts to these primary producers would have

broad implications for the aquatic food chain.

Non-Guideline: Measuring the Chronic 

Effects of Freshwater Sediment-Associated 

Contaminants on Chironomus dilutes 

Required to determine chronic impact 

to freshwater sediment dwelling 

organisms.

See Note 2

Silver nanoparticles if released into aquatic environments are likely to 

partition to sediment. Chronic tests on a freshwater benthic emergent 

insect (Chironomus dilutes, formerly Chironomus tentans) with 
Appendix B - Page 12

Table 2B - Summary of Tier II Required Data for Nanosilva

OSCPP Data Requirement (Note 1)

Guideline Number: Study Title Reason for Study Test Material Comments

epibenthic ecological niche will be used to estimate potential risks to 

freshwater benthic organisms.

Non-Guideline: Measuring the Chronic 

Effects of Freshwater Sediment-Associated 

Contaminants on Hyalella azteca

Required to determine chronic impact 

to freshwater sediment dwelling 

organisms.

See Note 2

Silver nanoparticles if released into aquatic environments are likely to 

partition to sediment. Chronic tests on a freshwater benthic amphipod 

(Hyalella azteca ) with infaunal ecological niche will be used to 

estimate potential risks to freshwater benthic organisms.

Non-Guideline: Measuring the Chronic 

Effects of Marine and Estuarine SedimentAssociated Contaminants on Leptocheirus 

plumulosus

Required to determine chronic impact 

to marine sediment dwelling 

organisms.

See Note 2

Silver nanoparticles if released into aquatic environments are likely to 

partition to sediment. Chronic tests on an esuarine/marine benthic 

amphipod (Leptocheirus plumulosus) will be used to estimate potential 

risks to marine benthic organisms.

850.1075: Acute Toxicity Test with 

Freshwater and Marine Fish

Required to confirm the chronic effects

of nanosilver. See Note 2

Acute toxicity test with fish species is to help in the assessment of 

possible risk to similar species in natural environments, as an aid in 

determination of possible water quality criteria for regulatory purposes, 

and for use in correlation with acute testing of other species for 

comparative purposes.

850.1300: Daphnid Chronic Toxicity Test Required to determine the chronic 

effects of nanosilver. See Note 2

This guideline prescribes a chronic toxicity test in which daphnids are 

exposed to a chemical either in a static-renewal or a flow through 

system.

850.1400: Early-Life Stage Toxicity Test 

for Freshwater, Estuarine, and Marine Fish

Required to determine the chronic 

effects of nanosilver. See Note 2

Tests with the early-life stages of fish are intended to define the lethal 

and sublethal effects of chemicals on the stages and species tested. They 

yield information of value for the estimation of the chronic lethal and 

sublethal effects of the substance on other fish species.

Environmental Fate

Non-Guideline: Rate of Deposition

The rates of aggregation and 

sedimentation are required for 

confirming estimates on the 

environmental fate and potential 

ecological impacts of these materials. 

See Note 2

Knowing the rate of aggregation and sedimentation of the products 

released from textiles would give primary information on the behavior 

of these compounds in the aquatic environment. 

835.1100: Activated Sludge Sorption 

Isotherm

Required to determine wastewater 

treatment removal efficiency. See Note 2

The EPA uses Guideline 835.1110 test results to estimate the removal 

efficiency of a chemical as it passes through a wastewater treatment 

plant.
Appendix B - Page 13

Table 2B - Summary of Tier II Required Data for Nanosilva

OSCPP Data Requirement (Note 1)

Guideline Number: Study Title Reason for Study Test Material Comments

835.1230: Adsorption/Desorption 

(Batch Equilibrium)

Required to determine partitioning to 

solids. See Note 2

If the material is removed during wastewater treatment, it may be 

deposited on land through the deposition of sludge (i.e. land farming). 

If the material is not removed during wastewater treatment it may be 

released into aquatic environments and may bind to sediment. 

Adsorption/desorption equilibrium studies required to determine the 

mobility of Nanosilva or nanosilver in the environment.

835.1240: Leaching Studies (Soil Column 

Tests)

Required to determine the mobility in 

subsurface environment. See Note 2

The distance that nanoparticles move in soil and groundwater is thought 

to depend on interaction with soil grains. Attachment of nanoparticles to 

soil grains depends on the physical processes of sedimentation, 

interception, and diffusion rather than partitioning to natural organic 

matter. Nanoparticle to soil grain interaction depends on nanoparticle 

diameter, aqueous chemistry, and the arrangement of soil grains and 

must be measured using soil column tests.

850.6800: Modified Activated Sludge, 

Respiration Inhibition Test for Sparingly 

Soluble Chemicals

Required to determine impact to 

wastewater treatment systems.

See 

Note 2

Silver from industrial processes (e.g., film processing) has been shown 

to reduce microbial activity in wastewater treatment systems. The 

purpose of the study is to assess the impact of Nanosilva or nanosilver

on microbial activity during wastewater treatment. 

Note 1: These guidelines only provide general guidance. Protocols shall be submitted prior to conducting these studies.

Note 2: The test material shall include materials that are released during the stability and plastics leaching studies. These materials include nanosilver released from the 

Nanosilva complex or plastics treated with Nanosilva and or the Nanosilva complex released from plastics treated with Nanosilva.
Table 3B – Enforceable Schedule

The Nanosilva complex and plastics treated with Nanosilva will be the test material during Tier I 

studies. EPA anticipates that data will be developed in a phased approach. Thus, the schedule is 

separated into phases where Phase 1 – Product Characterization and Phase 2 – Product Testing 

will occur prior to developing protocols for Phase 3 – Release Characteristics/Exposure, and 

Phase 4 – Health Effects. 

Tier I: Nanosilva and Nanosilva Treated Plastics Testing

Prepare and Submit

Protocols*

Perform Study and 

Submit Results*

Submittal of samples

Guideline Phase 1 – Product Characterization 2 6

Non-Guideline Particle Size Distribution

Non-Guideline Surface Area

Phase 2 – Product Testing 5 9

Non-Guideline Stability

Phase 3 – Release 

Characteristics/Exposure

21 27

Non-Guideline Dissolution Kinetics

Non-Guideline Leaching Test of Plastic

Phase 4 – Health Effects 27 37

870.3250 90-Day Inhalation

870.3550 Modified Reproduction/Developmental 

Toxicity Screening Test

*Number of months after conditional registration is issued.
Appendix B - Page 15

Table 3B – Enforceable Schedule

The test material for Tier II studies will depend on the results of the Tier I leaching and dissolution 

studies. EPA anticipates that data will be developed in a phased approach. Thus, the schedule is 

separated into phases where Phase 5 – Characterization will occur prior to developing protocols for 

Phase 7 – Health Effects, Phase 7 – Ecological Effects, and Phase 8 – Environmental Fate.

Tier II: Testing for Nanosilver and /or Nanosilva Released during Tier I Tests

Prepare and Submit

Protocols*

Perform Study and 

Submit Results*

Guideline Phase 5 – Characterization 19† 24

830.7050 UV-Vis

Non-Guideline Particle Size Distribution

Non-Guideline Surface Area

830.7840/7860 Solubility

Non-Guideline Zeta-potential

Phase 6 - Health Effects 27 37

Non-Guideline peri- and post-natal exposure to Nanosilva

Phase 7 – Ecological Effects 25 36

850.1850 Modified Aquatic Food Chain Transfer

850.4100 Terrestrial Plant Toxicity, Seedling 

Emergence

850.4400 Aquatic Plant Toxicity, Tier II

850.5400 Algal Toxicity, Tier II

Non-Guideline Measuring the Chronic Effects of 

Freshwater Sediment-Associated 

Contaminants on Chironomus dilutes 

Non-Guideline Measuring the Chronic Effects of 

Freshwater Sediment-Associated 

Contaminants on Hyalella azteca

Non-Guideline Measuring the Chronic Effects of Marine 

and Estuarine Sediment-Associated 

Contaminants on Leptocheirus plumulosus

850.1075 Acute Toxicity Test with Freshwater and 

Marine Fish

850.1300 Daphnid Chronic Toxicity Test

850.1400 Early-Life Stage Toxicity Test for 

Freshwater, Estuarine, and Marine Fish

Phase 8 – Environmental Fate 25 36

Non-Guideline Rate of Deposition

850.1100 Activated Sludge Sorption Isotherm

835.1230 Adsorption/Desorption 

835.1240 Leaching Studies (Soil Column Tests)

850.6800

Modified Activated Sludge, Respiration 

Inhibition Test for Sparingly Soluble 

Chemicals

* Number of months after conditional registration is issued.