Docket Number EPA-HQ-OPP-2013-0086 www.regulations.gov



## Trichloromelamine Preliminary Work Plan

Registration Review: Initial Docket

Case Number 3144

Approved by:

Date: 

## March 2013

Susan Lewts Acting Director

Antimicrobials Division

Page 1 of37Page 2 of 37 Trichloromelamine Registration Review Team

Human Health & Environmental Effects
William Hazel
Tim Dole Jenny Tao Siroos Mostaghimi
Srinivas Gowda James Breithaupt David Bays

Risk Management
Seiichi Murasaki
Lance Wormell

Office of General Counsel Philip RossPage 3 of 37 

### Table of Contents

1 Introduction .................................................................................................................. ........... 5

1.1 Statutory and Regulatory Authority ................................................................................. 5

1.2 Case Overview ................................................................................................................. 7

1.3 Chemical Identification and Properties ............................................................................ 7

1.4 Use/Usage Description ..................................................................................................... 8

1.4.1 Registrations ............................................................................................................. 8

1.4.2 Summary of Regist ered Uses .................................................................................... 8

1.4.3 Usage Information .................................................................................................... 8

1.5 Regulatory History ........................................................................................................... 8

1.5.1 Recent/Pending Regulatory Actions ......................................................................... 8

1.5.2 Tolerance Information .............................................................................................. 9

1.6 Incidents ..................................................................................................................... ...... 9

1.6.1 Human Health ........................................................................................................... 9

1.6.2 Ecological ................................................................................................................. 9

2 Anticipated Data Needs ........................................................................................................ .. 9

3 Human Health Risk Assessment ........................................................................................... 11

3.1 Existing Toxicological Endpoints .................................................................................. 11

3.2 Dietary Exposure ............................................................................................................ 1 2

3.2.1 Food ........................................................................................................................ 1 2

3.2.2 Drinking Water ....................................................................................................... 12

3.3 Occupational and Residential Exposures ....................................................................... 12

3.3.1 Occupational Handler Exposure ............................................................................. 12

3.3.2 Residential Exposures ............................................................................................. 14

3.4 Aggregate and Cumulative Exposure ............................................................................. 14

3.4.1 Aggregate Exposures .............................................................................................. 14

3.4.2 Cumulative Exposures ............................................................................................ 14

4 Environmental Risk Assessment........................................................................................... 14

4.1 Environmental Conceptual M odel Exposure Pathways ................................................. 15

4.2 Ecological Effects Assessment ....................................................................................... 16

4.2.1 Mechanism of Action .............................................................................................. 16

4.2.2 Measures of Effect (Ecotoxicology Endpoints) ...................................................... 16

4.3 Exposure and Effects Analysis Plan ............................................................................... 17

5 Endocrine Disruptor Screen ing Program (EDSP) ................................................................. 17

6 Guidance for Commenters .................................................................................................... 18

6.1 Preliminary Work Plan ................................................................................................... 18

6.1.1 Trade Irritants .......................................................................................................... 18

6.1.2 Water Quality .......................................................................................................... 18

6.1.3 Environmental Justice ............................................................................................. 18

6.1.4 Structure Activity Relationships ............................................................................. 19

6.1.5 Additional Information ........................................................................................... 20

7 Next Steps .................................................................................................................... ......... 20

8 References .................................................................................................................... ......... 21

Appendix A Toxicology Profile ............................................................................................... 23 Appendix B Environmental Fate ............................................................................................. 29Page 4 of 37 List of Tables Table 1 – Summary of Anticipated Risk Assessments and Data Needs: TCM .............................. 6 Table 2 – Anticipated Regist ration Review Schedule .................................................................... 6
Table 3 – Chemical Identification of TCM ..................................................................................... 7 Table 4 – Physical-Chemical Properties for TCM .......................................................................... 7 Table 5 – Summary of TCM Registered Uses ................................................................................ 8 Table 6 – Studies Anticipated to be Needed for the Registration Review of TCM ...................... 10 Table 7 – Toxicological Endpoints from the 2005 RED .............................................................. 11 Table 8 – TCM Occupational MOEs included in the RED .......................................................... 13 Table 9 – Occupational Handler Exposure Scenarios for TCM ................................................... 13 Table 10 – Residential Expos ure Scenarios for TCM .................................................................. 14 Table 11 – Existing Effects Endpoints for Risk Assessment of TCM .......................................... 16 Table 12 – Acute Toxicity Studies for TCM ................................................................................ 23 Table 13 – Summary of Subchronic and Developm ental Toxicity and Mutagenicity Data for

TCM ........................................................................................................................... ................... 24
Table 14 – Environmental Fate Data for TCM and Related Compounds C ontaining a Symmetric Triazine Ring ................................................................................................................. ............... 32 Table 15 – Changes in Protolysis (Hydrolysis) of Solutions C ontaining Hypochlorite Ion. ........ 36

List of Figures Figure 1 – Conceptual Model for Ecological E xposure and Effects of Melamine, Primary Degradate of TCM, and Its Major Reaction Produ cts/Degradates to Aquatic and Terrestrial Organisms ..................................................................................................................... ................ 15 Figure 2 – Formation and Decline of Cyanuric Ac id from Compounds with a Symmetric Triazine Ring. ......................................................................................................................... ..................... 30Page 5 of 37 

## 1 Introduction

This document is the Environmen tal Protection Agency’s (EPA or “the agency”) Preliminary Work Plan (PWP) for trichloromelamine (TCM ). The PWP document explains what EPA's Office of Pesticide Programs knows about TCM, highlighting anticipated data and assessment needs, identifying the types of information that would be especially useful to the agency in conducting the review, and providing an antici pated timeline for completing TCM’s review.

The registration review process was designed to include a publi c participation component to solicit input from interested stak eholders. The agency intends, by sh aring this information in the docket, to inform the public of what it knows ab out TCM and what types of new data or other information would be helpful for the agency to receive as it moves toward a decision on TCM. The agency encourages all interested stakeholde rs to review the PWP and to provide comments and additional information that will help the agen cy’s decision-making process for this chemical.



## 1.1 Statutory and Regu latory Authority

The Food Quality Protection Act (FQPA) of 1996 mandated a regi stration review program. All pesticides distributed or sold in the United Stat es generally must be registered by EPA based on scientific data showing that th ey will not cause unreasonable risks to human health or the environment when used as directed on produc t labeling. The registrati on review program is intended to make sure that, as the ability to assess risk evolves and as policies and practices change, all registered pesticides continue to meet the statutory standard of no unreasonable
adverse effects to human health or the environm ent. Changes in scienc e, public policy, and pesticide use practices will occur over time. Th rough the registration review program, the agency periodically reevaluates pesticides to make sure that as change occurs, products in the marketplace can be used safely. Inform ation on this program is provided at http://www.epa.gov/oppsrrd1/ registration_review/.

The agency is implementing the registration revi ew program pursuant to Section 3(g) of the Federal Insecticide, Fungicide, and Rodenticide Act (FIFRA) and will review each registered pesticide every 15 years to determine whether it continues to meet th e FIFRA standard for registration. The regulations governing registra tion review begin at 40 CFR 155.40. The agency will consider benefits information and data as required by FIFRA. The public phase of registration review begins when the initial docket is opened for each case. The docket is the agency’s opportunity to state what it knows about the pesticide and what additional risk analyses
and data or information it believes are needed to make a registrati on review decision. After reviewing and responding to comments and data received in the docke t during this initial comment period, the agency will develop and commit to a Final Work Plan (FWP) and anticipated schedule for the Trichloromelamine Case.

Documents associated with this registration re view can be viewed at http://www.regulations.gov in docket EPA-HQ-OPP-2013-0086. Below is a summ ary of the issues relevant to this registration review case.Page 6 of 37 Table 1 – Summary of Anticipated Risk Assessments and Data Needs: TCM

Risk Assessment Assessment Necessary to Support Registration
Review Date of

Most

Recent
Assessment Type of
Assessment Required (New/Updated) Data Anticipated as Needed (See Table 6 for details)

Dietary (food) Yes 5/26/2005 Updated Food Contact Surface Residue

in vivo Mutagenicity Data

Dietary (drinking water) No (see 3.2.2) N/A N/A None

Occupational Handler Yes 9/15/2005 Updated Indoor Exposure Data

in vivo Mutagenicity Data

Residential Handler Yes N/A New Indoor Exposure Data

in vivo Mutagenicity Data

Residental Post Application - Incidental
Oral and Dermal Yes N/A New Surface Residue Data

in vivo Mutagenicity Data

Aggregate Yes N/A New Indoor Exposure Data
Food Contact Surface Residue
Surface Residue Data

in vivo Mutagenicity Data

Cumulative No (see 3.4.2) N/A N/A None

Tolerance Review Yes N/A New None

Ecological No (see 4.2) N/A N/A None

N/A = Not applicable

Table 2 – Anticipated Regi stration Review Schedule

Anticipated Activity Target
Date* Completion Date

Phase 1: Openin g the Docket Open Docket and 60-Day Comment Period for Preliminary Work Plan 2013-03 2013-03-27

Close Public Comment Period 2013-05 Phase 2: Case Development

Issue Final Work Plan 2013-09 Issue Data Call-In (DCI) 2014-09

Receive Data to be Considered in Risk Assessment 2016-09 Open 30-Day Public Comment Period for Preliminary Risk Assessment(s) 2019-03

Close Public Comment Period 2019-04 Phase 3: Registration Review Decision and Implementation Open 60-Day Public Comment Period for Proposed Decision 2019-09

Close Public Comment Period 2019-11

Issue Final Decision 2020-03 Begin Post-Decision Follow-up 2020 Total (years) 7

*The anticipated schedule will be revised as necessary (e.g., need arising under the Endocrine Disruptor Screening Program with respect to the active ingredients in this case).Page 7 of 37 

## 1.2 Case Overview

The docket for the Trichloromelamine Case (Case 3144) has been established at http://www.regulations.gov in doc ket number EPA-HQ-OPP-2013-0086.



## 1.3 Chemical Identification and Properties

Table 3 presents the active ingredient to be asse ssed in Case 3144: Trichloromelamine (TCM) (PC Code 077101).

Table 3 – Chemical Identification of TCM 1

Chemical Name Trichloromelamine (TCM) Classification Triazine; Amine; Organochlorine

PC Code 077101

CAS No. 7673-09-8

Molecular Formula C 3 H 3 Cl 3 N 6 Molecular Weight 229.46 g/mol

% Available chlorine 91.5%

Molecular Structure

The TCM product chemistry information relevant to risk assessment is summarized in Table 4.

Table 4 – Physical-Chemical Properties for TCM 1

OPPTS
Guideline No. Physical and Chemical Properties TCM

830.1550 Product identity and composition Refer to Table 3.
830.1650 Description of Formulation Proces s Confidential Business Information (CBI)
830.6302 Color Cream or white powder
830.6303 Physical State Solid

830.6304 Odor Chlori ne- or bleach-like odor

830.6313 Stability to sunlight, normal and elevated temperature, metals/metal ions Stable under normal conditions.

830.7000 pH 4.7 (5% w/w slurry in water)

830.7050 UV/Visible Absorption The chemical structure suggests that it will not absorb in the UV spectral region (stable to sunlight for 40 minutes).
830.7200 Melting point >300 °C
830.7220 Boiling point Not applicable (substance is solid).
830.7300 Density 0.34 g/cm 3 at 20oC 830.7370 Dissociation Constant Not applicable
830.7550 Partition coefficient ( n -octanol/water) 1.43

830.7840 Solubility in water 0.064 g/100 mL (20 °C). Soluble in aqueous caustic solution, glacial ace tic acid and acetone.
830.7950 Vapor pressure 7.08 x 10 -5 mmHg

1 Source: MRIDs 42131001, 42131101, 42283401, 42385101, 43350301, 43350302, 43350303, and 43395401, Trichloromelamine RED (2005) and EPI Suite v4.1Page 8 of 37 

## 1.4 Use/Usage Description

1.4.1 Registrations

There are six EPA-registered products that cont ain TCM as an active i ngredient (a.i.). The products include two technical or manufacturing use products and four end-use products formulated as powders and packaged in canisters or small packets.



### 1.4.2 Summary of Registered Uses

Table 5 presents a summary of the registered uses of TCM that will be assessed in this registration review. TCM can be applied by the following application methods: mop, sponge, cloth, spray, and immersion.

Table 5 – Summary of TCM Registered Uses

Use Application

Method Application Rate (In Terms of Available Chlorine)

Food service areas (e.g., restaurants, schools, nursing homes, hospitals, food processing plants, hotels, day care centers, and homes)

Dishware, Utensils, Glassware Immersion 2 100 to 200 ppm

Nonporous hard surfaces such as tabl e tops, countertops, sinks, splash backs, trays, sealed floors and food processing / preparation equipment. Mop, sponge, cloth

or spray 100 to 200 ppm

Soft serve, yogurt, shake, slush and food equipment Immersion 3 100 to 200 ppm



### 1.4.3 Usage Information

There is no mention of TCM in the Kline Biocides Report fo r 2004/2005 (Kline, 2005), however, production volume data in the EPA Section Se ven Tracking System (SSTS) for the years 2008 through 2010 indicate that no more than 50,000 pounds of TCM were sold per year in the United

States 4

.



## 1.5 Regulatory History

The first product containing TCM as an active ingr edient was registered in the United States in 1959. The Agency completed a Reregistration El igibility Decision (RED) for TCM in 2005.



### 1.5.1 Recent/Pending Regulatory Actions

The post-RED Data Call-In (DCI) was issu ed in 2008. A follow-up DCI requiring skin
sensitization data was issued in 2010.

2 Applied in last sink prior to air drying

3 Air dried

4 Data for the years 2011and 2012 were not used in this estimate since data collection is still in progress.Page 9 of 37 

### 1.5.2 Tolerance Information

A tolerance exemption has been established fo r TCM used as a sani tizer on food processing equipment and utensils [40 CFR 180.940(c)] 5 . TCM must be used such that the end-use concentration of all di- or trichloroisocyanuric ac id chemicals in the solution is not to exceed 200 ppm determined as total available chlorine. Th e wording of this exemption will be revisited during the risk assessment stage of this regist ration review. In addition, although uses in public eating places have been approved, a tolerance exemption for TCM us ed as a sanitizer in public eating places has not been established at 40 CFR 180.940(a). The Agency will assess the need for a tolerance during this re gistration review. The U.S. F DA has cleared TCM sanitizing solution to be used on food-processing equipmen t and utensils and other food-contact articles except milk containers or equipment at 100-200 ppm [(21 CFR 178.1010(b)( 10)]. TCM has also been cleared by FDA for use in adhesives for food-packaging mate rial (21 CFR §175.105) although no products are currently registered for that use.



## 1.6 Incidents

1.6.1 Human Health

A total of 48 human health inci dents are listed in the OPP Inci dent Data System (IDS) for TCM for the time period spanning 2005 to 2012. Ten of th ese incidents are rated as moderate (Code HC) and 38 are rated as minor (Code HD).

Five of the moderate incidents involved skin effect s including: rash, b listering, peeling skin, open sores, and in one case, a second-degree ch emical burn. The skin in cidents occurred after exposure to the diluted product as it was being used to treat gl assware. In most cases, gloves
were not worn. Three incidents occurred when du st from the undiluted product got into the eye causing irritation, and in one cas e, a corneal burn which required antibiotic treatment. One
incident occurred when a worker accidentally inhaled some und iluted product and experienced
shortness of breath. Another inci dent occurred when a restaura nt patron experienced severe
throat irritation and vocal cord effects after drinking water allegedly contaminated with the product.

Symptoms reported for the 38 minor incident s included dermal, ocular, respiratory, and gastrointestinal irritation as we ll as vomiting. Because the incident s were classified as minor, the causes of exposure were not incl uded in the incident reports.



### 1.6.2 Ecological

No TCM ecological incidents ha ve been reported in the Ag ency’s Ecological Incident Information System (EISS) for the time period spanning 2005 to 2012.



## 2 Anticipated Data Needs

Table 6 presents a summary of the data anticipate d as being needed to su pport this registration

review.

5 Expressed as 1,3,5-Triazine, N,N ′ ,N ′′ -trichloro-2,4,6-triamino-.Docket Number EPA-HQ-OPP-2013-0086www.regulations.govPage10of37Table 6 – Studies Anticipated to be Neededfor the Registration Review of TCMGLNStudy NameTestSubstanceTime Frame(Measured inmonths from DCIReceipt)RiskAssessment(s)Data Will SupportUse Site(s)TriggeringAnticipated DataRequirementApplicable Exposure ScenarioStudies Anticipated to be Required through the Registration Review DCI870.5385 or870.5395In VivoMammalian Bone MarrowChromosomal Aberration Test orIn VivoMammalian ErythrocyteMicronucleus TestTGAI 12 AllAll (see Mutagenicitydiscussion inAppendix A fordetails)All scenarios875.1700 Product Use Information TGAI12 All AllAll scenarios.875.23006Indoor Surface ResidueDissipationTGAI 24 ResidentialSurface disinfection ofhousehold floorsDermal and incidental oral exposuresto TCM and/or melamine areexpected for toddlers who sit or playon floors disinfected with TCMproducts.SpecialStudyResidue Migration Studies FromFood Contact SurfacesTEP 24 Dietary Food Contact SurfacesDietary exposure from sanitized foodcontact surfacesStudies Required through the Post-RED DCI(GDCI-077101-27850) Issued October 8, 2008875.12007Indoor Exposure, DermalTGAI 12Occupational andResidentialHard SurfaceDisinfection1. Mix “last sink” solutions2. Soak food service utensils3. Mop Application4: Trigger Spray and WipeApplication875.14007Indoor Exposure, Inhalation875.28007Description of Human ActivitiesStudies No Longer Requiredthrough the Post-RED DCI870.3465890-day inhalation toxicity study TGAI 24Occupational andResidentialHard SurfaceDisinfectionMilitary mess kits6In the absence of surface residue data,the standard assumption of 100 percent transferable surface residue will be used.7It may be possible to use proprietary surrogate exposure data if data compensation issues are appropriately and adequately addressed.8The agency has determined that this study can be waived since the only product registered for use on mess kits was voluntarilycancelled effective July 19, 2009.Docket Number EPA-HQ-OPP- 2013-0086 www.regulations.gov

Page 11 of 37 

## 3 Human Health Risk Assessment

The agency anticipates the need to conduct a human health risk assessment for TCM. The agency expects to require additional data for use in conducting the registration review.



## 3.1 Existing Toxicological Endpoints

EPA anticipates the need to revise the existing t oxicological endpoints as pa rt of this registration review. Table 7 presents the endpoints used in EPA’s most recent human health risk assessment in support of the 2005 RED. Available information including existing toxicology studies, valid scientific literature, and the studies that are expe cted to be required for registration review will be considered in establishing the endpoints to be us ed in the registration re view risk assessment. A detailed description of the toxicity studies is provided in Appendix A .

Table 7 – Toxicological Endpoints from the 2005 RED

Exposure Scenario Dose Used in Risk Assessment (mg/kg/day) Target Margin of Exposure (MOE), Uncertainty Factor (UF), FQPA Safety Factor (SF) 9 Study and Toxicological Effects (See Appendix A for details)

Acute Dietary (General Population including infants and children) Acute Reference Dose (RfD) = 0.3 mg/kg/day UF = 100 FQPA SF = 1 acute Population Adjusted Dose (aPAD) = acute RfD FQPA SF= 0.3 mg/kg/day 90-Day Oral Rodent Study (MRID 43064301)
Lowest Observed Adverse Effect Level (LOAEL) = 150 mg/kg/day
No Observed Adverse Effect Level (NOAEL) = 30 mg/kg/day
Chronic Dietary (All populations) Chronic RfD = 0.1 mg/kg/day UF = 300 10
FQPA SF = 1 chronic Population Adjusted Dose (cPAD) = chronic RfD FQPA SF= 0.1 mg/kg/day 90-Day Oral Rodent Study (MRID 43064301) LOAEL = 150 mg/kg/day NOAEL = 30 mg/kg/day

Short-Term Incidental Oral (1-30 days) Oral Study Target MOE = 100

UF = 100 See study for Acute/Chronic Dietary Endpoint

Intermediate-Term Incidental Oral (1- 6 months) Oral Study Target MOE = 100

UF = 100 See study for Acute/Chronic Dietary Endpoint

Short/Intermediate-Term Dermal Exposure Oral Study / Dermal Absorption=100% Target MOE =100

UF = 100 See study for Acute/Chronic Dietary Endpoint

Short/Intermediate-Term Inhalation Exposure No appropriate route-specific study was available. The oral endpoint of 30 mg/kg with a MOE of 100 (10x interspecies extrapolation, 10x intraspecies variation) is used. An additional 10x route-to-route extrapolation is used to determine if a confirmatory inhalation toxicity study is warranted.

9 Developmental toxicity studies showed a lack of effects in offspring up to and including the highest doses tested, and the LOAELs were based on general systemic effect s that were not considered severe. Studies were welldesigned and provided an adequate dose-response for TCM. Therefore, the FQPA Safety Factor was removed.

10 UF of 300 includes a 10x interspecies extrapolation, 10x intraspecies variation, and 3x extrapolation from a subchronic study to a chronic duration.Page 12 of 37 

## 3.2 Dietary Exposure

3.2.1 Food

In the 2005 RED, acute and chronic dietary risk s to military personnel using treated mess kits and consuming treated fruits and vegetables were ≤ 7.1% and ≤ 21% of the aPAD and cPAD, respectively. Because the only product registered for use on mess kits and for fruit and vegetable washes was voluntarily cancelled effective Ju ly 19, 2009, these risks do not need to be reassessed. Acute dietary risks to civili ans eating in public eating places were ≤ 32% of the aPAD and chronic dietary risks were ≤ 54% of the cPAD with childr en being the most exposed population subgroup.

EPA anticipates the need to revise the acute and chronic dietary (food) risk assessment conducted in support of the 2005 RED. TCM is expect ed to be rapidly hydr olyzed to melamine and hypochlorous acid. The melamine is further degraded to ammeline and ammelide before degrading to cyanurate/isocyanur ate. Residue chemistry data ar e expected to be needed to determine dietary exposure to the modera tely stable degradates melamine and cyanurate/isocyanurate following sa nitization of food contact surf aces of food processing plants and food-handling facilities (inc luding public eating places). Acute and chronic dietary risk assessments are anticipated to be needed to estimate dietary exposure of all population subgroups to melamine and cyanurate/isocyanurate.



### 3.2.2 Drinking Water

EPA does not anticipate the need to conduct a drinking water assessment since dietary exposure from drinking water is not expected to occur. TCM is registered only for indoor use as a food or

nonfood contact surface sanitizer. Ther e are no approved uses expected to result in direct aquatic exposure or discharges of TCM a nd its degradates into a water system. As a result, the agency maintains its determination in the RED that a drinking water assessment is not necessary for TCM. No residue chemistry data are expected to be needed to char acterize drinking water

exposures.



## 3.3 Occupational and Residential Exposures

The agency anticipates the need to revise the occupational and resident ial assessments conducted in support of the 2005 RED since Margins of E xposure (MOE) were calculated using exposure data sets in which the agency has limited confidence; these MOEs will need to be reassessed using updated values. Uses of TCM that may re sult in occupational ha ndler and residential handler/postapplication exposure are pr esented in Table 9 and Table 10.



### 3.3.1 Occupational Handler Exposure

EPA anticipates the need to revise the occupa tional handler assessment conducted in support of the 2005 RED. Occupational handler dermal and i nhalation exposures to TCM were assessed in the RED for the Army uses involving the dip trea tment of mess kits, fruit and vegetables, and for food handling establishment uses involving the treatment of uten sils and wiping of countertops.Page 13 of 37 The specific scenarios included opening and pouri ng packets into treatment solutions; dipping mess kits, utensils, and fruit/vegetables into treatment solutions; and wiping countertops with

treatment solutions. The inhalation exposures were calculated using unit exposure data from the Chemical Manufacturers Association (CMA) anti microbial exposure study and the Pesticide Handlers Exposure Database (PHED). Dermal exposures for the opening/pouring and wiping scenarios were calculated using unit exposure da ta and dermal exposures for the dip treatment scenarios were calculated using the film thic kness approach. Both the inhalation and dermal MOEs were calculated using the NOAEL of 30 mg/ kg/day from the 90-day or al rodent study that had been selected at the time of the RED (MRID 43064301).

A summary of the MOEs from the 2005 RED is in cluded in Table 8. All of the MOEs exceeded the target MOE of 100 which meant that the risks were not of concern.

Table 8 – TCM Occupational MOEs included in the RED

Exposure Scenario Daily Amount Used (lb ai) Dermal Dose (mg/kg/day) Dermal
MOE Inhalation Dose (mg/kg/day) Inhalation
MOE Mess Kit and Fruit and Vegetable Treatments Open and Pour Packets 0.115 11 0.125 240 0.0000677 440,000 Dip Treat Mess Kits 0.0058 5200 0.0455 660 Dip Treat Fruits and Vegetables 0.0072 2200 0.0455 660

Restaurant and Bar Treatments Open and Pour Packets 0.00292 12 0.0032 1,7x10 7 0.0000017 9,400 Wipe Countertops 0.12 250 0.00116 26,000 Dip Treat Utensils 0.00488 6100 0.0016 26,000

Because the only product registered for use on mess kits and for fruit and vegetable washes was voluntarily cancelled effective July 19, 2009, thes e scenarios will not need to be reassessed during registration review. However, all of the other handler scenarios assessed in the RED will need to be revised during regist ration review upon rece ipt of the unit exposure data that are anticipated to be required to supplement or repl ace the unit exposure data that were used in the RED. In addition, a new scenario for spray application is expected to be assessed to account for products that were registered af ter the RED. Occupational handler scenarios to be assessed are presented in Table 9.

Table 9 – Occupational Handler Exposure Scenarios for TCM

Occupational Exposure Scenario Exposure Route(s) Duration

Mix “last sink” solutions for fo od service utensils Dermal &
Inhalation Short- and
Intermediate-term

Soak food service utensils in TC M “last sink” solutions Dermal &
Inhalation Short- and
Intermediate-term
Mop and wipe TCM disinfectant solutions onto hard surfaces in food service areas and food processing plants Dermal &
Inhalation Short- and
Intermediate-term
Trigger spray apply TCM solutions to hard surfaces in food service areas and food processing plants Dermal &
Inhalation Short- and
Intermediate-term

11 Two packets each containing 4.77 oz of product containing 19.3% a.i.

12 One packet containing 0.25 oz of product containing 18.7% a.i.Page 14 of 37 

### 3.3.2 Residential Exposures

EPA anticipates the need to conduct a residential handler assessment. The risk assessment for the RED indicated that a separate risk assessment for residential uses had not previously been performed because the assessment for occupational handlers in restaurants was considered to be a conservative and protective surrogate for reside ntial uses. In addition, th e products appeared to be intended primarily for food service and pr eparation areas in commercial and institutional facilities. EPA Reg. No. 65169-1, however, includes day care centers and homes as use sites and, therefore, it will be necessary to assess postappli cation dermal and incidental oral exposure of toddlers playing on floors treated with this produ ct. This scenario is included in Table 10.

Table 10 – Residential Exposure Scenarios for TCM

Residential Exposure Scenario Exposure Route(s) Duration
Toddlers playing on floor treated with TCM products Der mal & Incidental Oral Short- and Intermediate-term



## 3.4 Aggregate and Cumulative Exposure

3.4.1 Aggregate Exposures

EPA anticipates the need to conduct an aggreg ate exposures assessment. Upon a reevaluation and selection of toxicological endpoints, combined with the hu man health exposure assessments expected to be needed as a pa rt of this registration review case, aggregate exposures including dietary (food) exposures and residential exposures, will need to be assessed.



### 3.4.2 Cumulative Exposures

With respect to cumulative exposure, unlike other pesticides for which EPA has followed a cumulative risk approach based on a common me chanism of toxicity, EPA has not made a common mechanism of toxicity finding as to TC M and any other substances and TCM does not appear to produce a toxic metabolite produced by other substances. For the purposes of this registration review, th erefore, EPA has not assumed that TCM has a common mechanism of toxicity with other s ubstances. For information regarding EPA’s efforts to determine which chemicals have a common mechanism of toxicity a nd to evaluate the cumulative effects of such chemicals, see http://www.ep a.gov/pesticides/cumulative/.



## 4 Environmental Risk Assessment

The agency does not anticipate the need to c onduct a quantitative ecologi cal risk assessment for TCM or its degradates. The regi stered use pattern as a food and non-food contact sanitizer is expected to result in minor or insignificant am ounts reaching surface waters because of the rapid dechlorination to the degradates melamine, hypo chlorous acid, and cyanuric/isocyanuric acid (see Appendix B ).

Based on the low hazard and exposure, the agency inte nds to conclude that the registered uses of TCM will have 'no effect' (NE) on endangered or thre atened terrestrial or aquatic species, or their designated critical habitats, as listed by the U.S. Fish and Wildlife Service (USFWS) and thePage 15 of 37 National Oceanic and Atmospheric Administration' s (NOAA) National Mari ne Fisheries Service (NMFS). EPA anticipates conducting no further an alysis of potential ri sks to endangered or threatened species unless public comment or othe r submissions provide data or information that would otherwise inform the agency's determination.



## 4.1 Environmental Conceptual Model Exposure Pathways

Based on the chemical/physical properties summarized in Table 4, environmental fate information (Appendix B ), and the registered use as a food surface sani tizer in Table 5, the agency has developed a conceptual model for the environmental exposure to TCM and its metabolites/degradates. For the registered us e as a food and non-food contact sanitizer, the agency assumes that environmental exposures will be minimal. Bioconcentration of any residues
that reach surface water will not be si gnificant because of their low log K
ow (log P) and high water solubility values. Any environmental exposure is expected to be of short duration in the presence of acclimated microorganisms or an aerobic conditions (Zeyer et al., 1980, Ernst and Rehm, 1990, Jessee et al., 1983, Cook et al., 1985, Saldick et al., 1974). Figure 1 illustrates the conceptual model for the use of TCM products in cluding as a hard-surface, food contact sanitizer which can go down the drain after use. Table 14 in Appendix B cont ains the details of
environmental fate studies reviewed.

Figure 1 – Conceptual Model for Ecological Exposure and Effects of Melamine, Primary Degradate of TCM, and Its Major Reaction Products/Degrada tes to Aquatic and Terrestrial OrganismsPage 16 of 37 

## 4.2 Ecological Effects Assessment

Ecological effects data are used as measures of di rect and indirect effect s to biological receptors. Acute and chronic toxicity data from studies submitted by pesticide registrants along with the available open literature are used to evaluate th e potential direct and indirect effects of TCM and its degradates on aquatic a nd terrestrial receptors.



### 4.2.1 Mechanism of Action

TCM is used as a water solution which form s melamine and hypochlorous acid (HClO), an oxidizing agent and the biocidal agent. HClO is a weak acid and, depending upon pH, can further disassociate into a hydrogen ion and a hypochlorite ion (ClO - ), in a reversible reaction. The antimicrobial activity of chlorine compounds depends primarily on HClO which can rapidly

cross cell membranes. While ClO - has some disinfection potential it requires longer contact time.

It is HClO and ClO - that are the active di sinfecting moieties of TC M (see Hypochlorous Acid section in Appendix B ).



### 4.2.2 Measures of Effect (Ecotoxicology Endpoints)

Current selected ecotoxicologi cal endpoints are summarized in Table 11. The results of the

submitted studies indicate that TCM is practically nontoxic to B obwhite Quail on an acute oral basis. It is slightly toxic to Bobwhite quail and practically nont oxic to Mallard duck on a dietary basis. It is moderately toxic to Bluegill sunfish and Rainbow trout. It is highly toxic to Daphnia.

An agency memorandum (EPA, 1983), indicates th at there is some evidence that melamine, a degradate of TCM, may be chroni cally toxic to fish and invertebrates by causing adverse effects on reproduction and embryonic development based on an FDA review of acute toxicity studies

for melamine. The memorandum noted the FDA c onclusion that, although there is insufficient evidence to determine the potential environmental toxicity of melamine, there is some basis for concern based on the structure-activity-re lationship (SAR) of melamine to metaphenylenediamine, and the readily available st udies on melamine’s reproductive effects on embryonic development of fish and invertebra tes. Based on SARs of melamine to metaphenylenediamine, the 96-hour EC 50 for melamine for green algae is expected to be 2.4 mg/L (moderately toxic). The LC 50 for daphnids is expected to be 5 .9 mg/L (moderately toxic), and the MATC is expected to range between 0.05 and 0.09 mg/L.

Due to the limited use pattern of this chemical, TC M and its degradates are not expected to enter

the environment and come into contact with non-target terrestrial a nd aquatic organisms. Therefore, based on lack of expos ure, risk to terrestrial and aq uatic organisms is not expected.

Table 11 – Existing Effects Endpoints for Risk Assessment of TCM

Receptor Group Surrogate Species Risk
Scenario Toxicity Endpoint as

a.e. Reference

Birds Bobwhite quail ( Colinus virginianus ) Acute LD 50 > 5,000 mg/kg NOEC = 5,000 mg/kg 42250801Page 17 of 37 Receptor Group Surrogate Species Risk
Scenario Toxicity Endpoint as

a.e. Reference

Bobwhite quail ( Colinus virginianus ) Acute LD 50 > 2,150 mg/kg 42280801

Mallard duck ( Anas platyrhynchos ) Acute LC 50 > 5,000 mg/kg NOEC = 2,500 mg/kg 42247401

Freshwater fish Bluegill sunfish ( Lepomis
macrochirus ) Acute LC 50 = 4.5 mg/L NOEC = 1.0 mg/L 41934901

Rainbow Trout Acute LC 50 = 4.0 mg a.i./L NOEC = 0.56 mg a.i./L 42010601

Freshwater
invertebrates Daphnia Acute EC 50 = 0.80 mg ai/L NOEC = 0.32 mg ai/L 42020801



## 4.3 Exposure and Effects Analysis Plan

Based on low likelihood of exposure, a quantitativ e risk assessment is not anticipated to be conducted. No additional ecological effects or envir onmental fate data are expected to be needed.



## 5 Endocrine Disruptor Screening Program (EDSP)

As required by FIFRA and FFDCA, EPA reviews num erous studies to asse ss potential adverse outcomes from exposure to chemicals. Collectiv ely, these studies includ e acute, subchronic and chronic toxicity, including asse ssments of carcinogenicity, ne urotoxicity, developmental, reproductive, and general or systemic toxicity. These studies include e ndpoints which may be susceptible to endocrine influe nce, including effects on endocri ne target organ histopathology, organ weights, estrus cyclicity, sexual maturatio n, fertility, pregnancy rates, reproductive loss, and sex ratios in offspring. For ecological hazard assessments, EPA evaluates acute tests and chronic studies that assess gr owth, developmental and reprod uctive effects in different taxonomic groups. As part of its 2005 reregistrati on decision for TCM, EPA reviewed these data and selected the most sensitive endpoints for relevant risk assessment scenarios from the existing hazard database. However, as required by FFD CA section 408(p), TCM is subject to the endocrine screening part of the Endocrine Disruptor Screenin g Program (EDSP).

EPA has developed the EDSP to determine whet her certain substances (including pesticide active and other ingredients) may have an effect in humans or wildlife similar to an effect produced by a “naturally occurring estrogen, or ot her such endocrine effect s as the Administrator may designate.” The EDSP employs a two-tiered approach to making th e statutorily required
determinations. Tier 1 consists of a battery of 11 screening assays to identify the potential of a chemical substance to interact with the estrog en, androgen, or thyroid (E, A, or T) hormonal systems. Chemicals that go th rough Tier 1 screening and are found to have the potential to interact with E, A, or T hormonal systems will proceed to the next stag e of the EDSP where EPA will determine which, if any, of th e Tier 2 tests are necessary base d on the available data. Tier 2 testing is designed to identify a ny adverse endocrine-related eff ects caused by the substance, and establish a dose-response relationship between the dose and the E, A, or T effect.

Under FFDCA section 408(p), the agency must scr een all pesticide chemicals. Between October 2009 and February 2010, EPA issued test orders/dat a call-ins for the firs t group of 67 chemicals, which contains 58 pesticide activ e ingredients and 9 inert ingr edients. TCM is not among thePage 18 of 37 group of 58 pesticide active ingredients on the in itial list to be screened under the EDSP. Accordingly, as part of registration review, EPA will issue future EDSP orders/data call-ins, requiring the submission of EDSP screening assays for TCM.

For further information on the status of the ED SP, the policies and procedures, the list of 67 chemicals, future lists, the test guidelines and the Tier 1 screening ba ttery, please visit our website: http://www.epa.gov/endo/.



## 6 Guidance for Commenters

6.1 Preliminary Work Plan

The public is invited to comment on EPA’s Preliminary Work Plan and rationale. The agency will carefully consider all comments as well as any additional information or data provided in a timely manner prior to issuing a final work plan for the Trichloromelamine registration review

case.



### 6.1.1 Trade Irritants

Through the registration review process, the ag ency intends to solic it information on trade irritants and, to the extent feas ible, take steps toward facilitating irritant resolution. The agency will work to harmonize tolerances and international maximum residue limits (MRLs) and may modify tolerance levels to do so, when possible. Stakeholders are asked to comment on any trade irritant issues resulting fr om lack of MRLs or disparities between U.S. tolerances and MRLs in key export markets, providing as much specificity as possible re garding the nature of
the concern.



### 6.1.2 Water Quality

TCM is not identified as a cause of impairmen t for any water bodies listed as impaired under section 303(d) of the Clean Water Act 13 . In addition, no Total Maximum Daily Loads (TMDL) have been developed for TCM 14 . More information on impaired water bodies and TMDLs can be
found at EPA’s website 15

. The agency invites submission of water quality data for this pesticide. To the extent possible, data should confor m to the quality standards in Appendix A of
the OPP Standard Operating Procedure: Inclus ion of Impaired Water Body and Other Water Quality Data in OPP’s Registration Revi ew Risk Assessment and Management Process 16 in order to ensure they can be used quantitatively or qualitatively in pesticide risk assessments.



### 6.1.3 Environmental Justice

EPA seeks to achieve environmenta l justice, the fair tr eatment and meaningful involvement of all people, regardless of race, color, national origi n, or income, in the deve lopment, implementation, and enforcement of environmental laws, regulati ons, and policies. To help address potential

13 http://iaspub.epa.gov/tmdl_waters10/attains_nation_cy.cause_detail_303d?p_cause_group_id=885

14 http://iaspub.epa.gov/tmdl_waters10/attains_nation.tmdl _pollutant_detail?p_pollutant_group_id=885&p_pollutant _group_name=PESTICIDES

15 http://www.epa.gov/owow/tmdl/

16 http://www.epa.gov/oppsrrd1/registration_review/water_quality_sop.htmPage 19 of 37 environmental justice issues, the agency seeks information on any groups or segments of the population who, as a result of thei r location, cultural practices, or other factors, may have atypical, unusually high exposure to TCM compared to the general population. Please comment if you are aware of any sub-populations that may have atypical, unusually high exposure compared to the general population.



### 6.1.4 Structure Activity Relationships

EPA must rely upon information of appropriate qu ality and reliability for each decision made by the agency. In the Office of Pesticide Program s (OPP), the evaluation process for a pesticide chemical traditionally begins with the applicant’ s submission of a set of studies conducted with the specific pesticide chemical of interest. The us e of the results of such testing (measured data) is a logical, scientifically rigorous proce ss that identifies the physical, chemical, and environmental fate properties of the pesticide, as well as the dose and endpoints at which an adverse effect can occur in various animal species.

Today, there is significant interest in alternat ive techniques, i.e., tech niques other than data generation that could significantly inform the agency’s decision-making process. Recently, OPP has made increasing use of stru cture activity relationship (SAR ) as part of its regulatory decision-making process. In the SAR process, a ch emical's molecular structure is compared to
that of other chemicals for which data are availabl e. These structural similarities are then used to make predictive judgments about a chemical’s physical, chemical, and biological properties. Thus, the chemical’s physical, chemical, and biol ogical properties are a f unction of (or directly related to) the chemical’s molecular structure. Quantitative SAR is referred to as QSAR. To develop a QSAR, a selected set of measured da ta on a single physical, chemical, or biological property is used to derive a model (an equatio n) to predict the valu e of that property.

Since SAR assessments and QSAR modeling are anothe r set of tools that are available to agency scientists, OPP has begun a proce ss shift that envisions shifting from the current study-by-study approach to an approach in which the use of pr edicted data, generated us ing validated models, is considered along with information from open litera ture and studies specifically generated under Part 161 requirements. All relevant information w ould be considered as pa rt of a weight-of-theevidence evaluation.

At this time, EPA believes that for certain e ndpoints, especially physi cal/chemical and fate properties, that SAR and QSAR might be effect ively utilized to fulfill these data requirements for many antimicrobial pesticide chemicals. When considering biological prope rties, at this time, EPA believes that SAR and QSAR can be most effectively utilized in the evaluation of chemicals that exhibit lower toxicity for human health and/or ecotoxicity parameters. This is appropriate because the risk assessment for lower toxicity chemicals can be stream-lined, i.e., a screening-level assessment procedure rather than multiple tiers of assessments with progressively more data requirements.

If stakeholders believe that submission of predic ted data can fulfill one of the data needs for the
Trichloromelamine Case, then the agency invites submission of this information. The submitter would be expected to supply a ra tionale describing the utility of the information and provide
documentation on the scientific va lidity of the information. The de termination that the predictedPage 20 of 37 data fulfills the data requirement would be at the sole discretion of the agency. Pre-submission consultation with the agency is encouraged.



### 6.1.5 Additional Information

Stakeholders are also specifically asked to provid e available information a nd data that will assist the agency in refining its risk assessments, including any species-spec ific ecological effects determinations. The agency is interested in receiving the following information:

1. Confirmation on the following label information:

A. Sites of application

B. Formulations

C. Application methods and equipment

D. Maximum application rates

E. Frequency of application, applicatio n intervals and maximum number of applications

F. Geographic limitations on use

2. Use or potential use distribution

3. Use history

4. Usage/use information for non-agricultura l uses (e.g., material s preservation)

5. Typical application interval

6. State or local use restrictions

7. Ecological incidents (non-target plant damage and avian, fish, reptilian, amphibian and mammalian mortalities) not alr eady reported to the agency

8. Monitoring data



## 7 Next Steps

After the 60-day comment period closes in May 2013, the agency will review and respond to any comments received in a timely manner, and then issue a Final Work Plan for the

Trichloromelamine Case.Page 21 of 37 

## 8 References

An L, Li Z, and Yang Z, et al. (2011) Cognitive defic its induced by melamine in rats. Toxicol Lett. 2011 Oct 30;206(3):276-80. Epub 2011 Aug 22. Bentivegna, S. (2001) Trichloromelamine: A 3-Month Oral (Capsule) Toxicity Study in Dogs with a One-Month Recovery Period: Lab Project Number: 003445:85MA0970: DAAD05-00-P8190. Unpublished study prepared by Huntingdon Life Sciences. 523 p. (MRID# 45603301) Bowman, J. (1986) Acute Toxicity of Trichloromelamine to Bluegill Sunfish (Lepomis macrochirus): Lab Project Number: 34811. Unpublished study prepared by ABC Labs, Inc. 17 p. (MRID# 41934901) Bowman, J. (1986) Acute Oral Toxicity Trichloromelam ine to Rainbow Trout (Salmo gairdneri): Lab Project Number: 34812. Unpublished study prepared by Analytical Bio-chemistry Laboratories, Inc. 17 p. (MRID# 42010601) Brown CA, Jeong KS, Poppenga RH, et al. (2007) Outbreaks of renal failure associated with melamine and cyanuric acid in dogs and cats in 2004 and 2007. J Vet Diagn Invest. 2007 Sep;19(5):525-31. Burgess, D.; Frazier, S.; Schoen, L. (1991) Acute Toxicity Trichloromelamine to Daphnia magna: Lab Project Number: 34813. Unpublished study prepared by ABC Laboratories, Inc. 15 p. (MRID# 42020801) Cifone, M. (1986) Evaluation of Trichloromelamine in the Rat Hepatocyte scheduled DNA Synthesis Assay: Lab Project Number: 20991 Unpublished study prepared by Ha zleton Laboratories America, Inc. 25 p. (MRID# 42021701) Dobson, RL, Motlagh, S, and Quijano, M, et al. (2008) Identification and characterization of toxicity of contaminants in pet food leading to an outbreak of renal toxicity in cats and dogs. Toxicol Sci. 2008 Nov;106(1):251-62. Epub 2008 Aug 9. Early RJ, Yu H, and Mu XP, et al. (2012) Repeat oral dose toxicity studies of melamine in rats and monkeys. Arch Toxicol. 2012 Sep 28. Fletcher, D.; Pedersen, C. (1988) Trichloromelamine: 8-Day Acute Dietary LC 50 Study in Mallard Ducklings: Lab Project Number: 88 DC 109. Unpublished study prepared by Bio-Life Associates, Ltd. 26 p. (MRID# 42247401) Fletcher, D.; Pedersen, C. (1988) Trichloromelamine: 21-Day Acute Dietary LD 50 Study in Bobwhite Quail: Lab Project Number: BLAL 88 QD 109. Unpublished study prepared by Bio-Life Assocs. 31 p. (MRID# 42250801) Fletcher, D.; Pedersen, C. (1988) Trichloromelamine: 8-Day Acute Dietary LC 50 Study in Bobwhite Quail: Lab Project Number: BLAL 88 QC 110. Unpublished study prepared by Bio-Life Associates, Ltd. 25 p. (MRID# 42280801) Gamboa da Costa, G, Jacob, CC, and Von Tungeln, LS, et al. (2012) Dose-response assessment of nephrotoxicity from a twenty-eight-day combined-exposure to melamine and cyanuric acid in F344 rats. Toxicol Appl Pharmacol. 2012 Jul 15;262(2):99-106. Epub 2012 May 2. Glaza, S. (1994) Acute Dermal Toxicity Study in Trichloromelamine (TCM), #14468W46 in Rabb its: Final Report: Lab Project Number: HWI 31102386: TP3016. Unpublished study prepared by Hazleton Wisconsin, Inc. 31 p. (MRID# 43159901) Glaza, S. (1994) Primary Eye Irritation Study of Trichlorom elamine (TCM), #14468W46 in Rabbits: Final Report: Lab Project Number: HWI 31102388: TP3015. Unpublished study prepared by Hazleton Wisconsin, Inc. 26 p. (MRID# 43159902) Glaza, S. (1994) Primary Dermal Irritation Study of Tr ichloromelamine (TCM), #14468W46 in Rabbits: Final Report: Lab Project Number: HWI 31102387: TP3014. Unpublished study prepared by Hazleton Wisconsin, Inc. 24 p. (MRID# 43159903) Glaza, S. (1994) Dermal Sensitization Study of Trichloromelamine (TCM), #14468W46 in Guin ea Pigs--Closed Patch Technique: Final Report: Lab Project Number: HWI 31102389: TP2008. Unpublished study prepared by Hazleton Wisconsin, Inc. 38 p. (MRID# 43159904) Glaza, S. (1994) Acute Oral Toxicity Study of Trichloromelamine (TCM), #14468W46 in Rats: Final Report: Lab Project Number: HWI/31102385. Unpublished study prepared by Hazleton Wisconsin, Inc. 44 p. (MRID# 43165701) Ivett, J. (1986) Clastogenic Evaluation of Trichloromelamine in an In Vitro Cytogenic Assay Measuring Chromosomal Aberration Freq- uencies in Chinese Hamster Ovary (CHO) Cells: Lab Project Number : 20990. Unpublished study prepared by Hazelton Laboratories America, Inc. 33 p. (MRID# 42021801)Page 22 of 37 Jacob, CC, Reimschuessel, R, and Von Tungeln, LS, et al . (2010) Dose-response assessment of nephrotoxicity from a 7-day combined exposure to melamine and cyanuric acid in F344 rats. Toxicol Sci. 2011 Feb;119(2):391- 7. Epub 2010 Oct 28. Jacob CC, Von Tungeln LS, and Vanlandingham M, et al. (2012) Pharmacokinetics of me lamine and cyanuric acid and their combinations in F344 rats. Toxicol Sci. 2012 Apr;126(2):317-24. Epub 2012 Jan 6. Jagannath, D. (1987) Evaluation of Trichloromelamine, (...) in Ames Salmonella/ Microsome Reverse Mutation Assay: Lab Project Number: 20988. Unpublished study prepared by Hazleton Labs., America, Inc. 26 p. (MRID# 42148801) Kim SH, Lee IC, and Lim JH, et al. (2011) Effects of melamine on pregnant dams and embryo-fetal development in rats. J Appl Toxicol. 2011 Jun 27. doi: 10.1002/jat.1703. Michie, M. (1989) Trichloromelamine--90-Day Subchronic Study in Rats: LabProject Number: 75-51-0743-88: 40- 0743-88. Unpublished study prepared by U.S. Army Environmental Hygiene Agency. 1177 p. (MRID# 43064301) OECD SIDS Dossier for Melamine, UNEP Publications, June, 2002. http://www.inchem.org/documents/sids/sids/1 08781.pdf accessed in November, 2012. Pang J, Li GQ, and Li CR, et al. (2011) Toxicokinetic st udy of melamine in the presence and absence of cyanuric acid in rats. J Appl Toxicol. 2011 Nov 17. doi: 10.1002/jat.1764. Park D, Kim TK, and Choi YJ, et al. (2011) Increased nephrotoxicity after combined administration of melamine and cyanuric Acid in rats. Lab Anim Res. 2011 Mar;27(1):25-8. Epub 2011 Mar 26. Puschner B, Poppenga RH, Lowenstine LJ, et al. (2007) As sessment of melamine and cyanuric acid toxicity in cats. J Vet Diagn Invest. 2007 Nov;19(6):616-24. Tyl, R.; Marr, M.; Myers, C. (1995) Developmental Toxicity Evaluation of Trichloromelamine Administered by Gavage to New Zealand White Rabbits: Final Re port: Lab Project Number: 65C-5742-300/400. Unpublished study prepared by Research Tr iangle Institute. 215 p. (MRID# 43614301) Tyl, R.; Marr, M.; Myers, C. (1995) Developmental Toxicity Evaluation of Trichloromelamine Administered by Gavage to CD (Sprague-Dawley) Rats: Final Report: Lab Project Number: 65C-5742-100/200. Unpublished study prepared by Research Tr iangle Institute. 246 p. (MRID# 43614302) U. S. EPA, 1983. Meeting with FDA Toxicology Department concerning their evaluation of pituitary tumor
incidence in male Fischer 344 rats and bone marrow leukemia incidence in female F 344 rats in the 30- month melamine study submitted by American Cyanamin (1983). Memorandum from Stephanie April, Toxicology Branch, to Ti m Gardner, Registration Division. December 14, 1983. U.S. EPA (1984) Memorandum: Melamine Studies. From Stephanie P. April, Toxicology Branch, to James Davis, Pesticide and Toxic Substances. May 2, 1984. U.S. EPA (1993) Memorandum: Carcinogenicity Peer Review of Melamine. From Stephen C. Dapson, Senior Pharmcologist, HED, and Esther Rinde, Manager of Carcinogenicity Peer Review Committee, HED, to Phil Hutton, Insecticide-Rodenticide Branch , Registration Division. July 21, 1993. U.S. EPA (1994) Memorandum: Review of Acute Toxicity Studies Submitted in Response to DCI. From Jess Rowland, Toxicologist, HED to K. Davis and B. Alder, Product Managers, Reregistration Division. July 21, 1994. U. S. EPA, 2004. Overview of the Ecological Risk Assessment Process in the Office of Pesticide Programs U.S. Environmental Protection Agency - Endangered and Threatened Species Effects Determinations, Appendix A, Section IIB, pg.81. US Environmental Protection Agency. January 24, 2004. http://www.epa.gov/oppfead1/endanger/c onsultation/ecorisk-overview.pdf. U.S. EPA (2005a) Reregistration Eligibility Decision Document (RED) for Trichloromelamine (Case 3144), September 2005. http://www.regulations.gov/#!documentDetail;D=EPA-HQ-OPP-2005-0262-0002 U.S. EPA (2005b) Trichloromelamine Risk Assessment fo r the Reregistration Eligibility Decision. September 15, 2005. http://www.regulations.gov/#!documentDetail;D=EPA-HQ-OPP-2005-0262-0003 Viau, A. (1994) An Evaluation of the Acute Toxicity of Inhaled Trichloromelamine Dry Powder Formulation in the Albino Rat: Lab Project Number: 90906. Unpublished study prepared by Bio- Research Lab., Ltd. 190 p. (MRID# 43368501)Docket Number EPA-HQ-OPP-2012-0682 www.regulations.gov

Page 23 of 37 

# Appendix A Toxicology Profile

Acute Toxicity for Product Labeling

As listed in Table 12, TCM is Toxicity Catego ry I for primary eye irritation, Category II for acute oral, inhalation toxicity, and primary skin irritation, and Category III for acute dermal toxicity. It is not a skin sensitizer. All acute t oxicity data are adequate for currently registered uses. The agency does not anticipate that any ad ditional technical acute t oxicity information will be needed for the registration review of TCM.

Table 12 – Acute Toxicity Studies for TCM

Guideline No./ Study Type MRID Results Toxicity Category
870.1100 / Acute Oral (93% a.i.) 43165701 LD 50 = 398 mg/kg II

870.1200 / Acute Dermal 43159901 LD 50 > 2000 mg/kg III

870.1300 / Acute Inhalation (100% a.i.) 43368501 LC 50 = 0.4 mg/L, males and 0.780 mg/L, females II

870.2400 / Primary Eye Irritation (95% a.i.) 43159902 Severe conjunctival irritation I

870.2500 / Primary Dermal Irritation (95.1% a.i.) 43159903 Severe irritant II

870.2600 / Dermal Sensitization (95.1% a.i.) 43159904 No sensitization reactions for group treated with 0.1% [w/v] solution N/A

N/A=Not available



## Subchronic Toxicity

TCM caused mortality in mid- a nd high-dose groups in a 90-day oral toxicity study in rats (MRID 43064301). Gross necropsy revealed hemorrhag ic erosions of the stomach mucosa as well as stomach and intestinal distention, and congestion of the lung w ith white spotting at varying degrees. Significantly (p <0.05) lower body weight gain was seen in the high-dose group. Histopathology examination revealed engorgement of the small blood ve ssels of the adrenal gland, brain, kidneys, liver, l ung, and pituitary gland as well as pulmonary edema. TCM also
induced mild to moderate chronic inflammati on and moderate hyperplasia of the non-glandular stomach at the high-dose level as well as a moderate hyperplasia in one mid-dose male. A NOAEL of 30 mg/kg/day (low-dose) and a LO AEL of 150 mg/kg/day (mid-dose) were established, based on hi stopathological lesions.



## Developmental Toxicity

TCM did not show developmental effects in a rat developmental study (MRID 43614302). Clinical signs, including al opecia, gasping, altered respir ation, salivation, lethargy, and chromodacryorrhea were observed at one or mo re dose levels. Reduction in body weight gains and decreased food consumption at all dose leve ls were also observed. A maternal LOAEL of 62.5 mg/kg/day and a developmental NOAEL of 500 mg/kg/day (highest dose tested [HDT])Page 24 of 37 were established. A maternal NOA EL could not be determined at the lowest dose tested (LDT) of 62.5 mg/kg/day. TCM did not show developmenta l effects in a rabbit developmental study but produced statistically significan t dose-related depressions in mean maternal body weights and decreased mean feed consumption (MRID 43614301). A maternal systemic toxicity LOAEL of 30 mg/kg/day and a developmental NOAEL of 12 0 mg/kg/day (HDT) were established. No maternal NOAEL could be determined even at the LDT of 30 mg/kg/day.



## Reproductive Toxicity

Based on the review of available data (e.g., subc hronic and developmental studies), the database for reproduction is adequate to support TCM’s direct and indirect food uses.



## Chronic Toxicity

Chronic toxicity studies are not expected to be needed si nce long-term duration exposure scenarios are not expected based on current registered uses of TCM.



## Carcinogenicity

The need for carcinogenicity studies will be re-a ssessed following the agency’s review of the in

vivo mutagenicity study expected to be required.



## Mutagenicity

The agency reviewed the submitted gene mutation studies and found they are acceptable. TCM is positive for mutagenicity in Salmonella strains TA98 and TA100, with and without metabolic activation, in an Ames assay (MRID 42148801). It is also positive in an in vitro mammalian chromosomal aberration assay in Chinese hams ter ovary (CHO) cells, with and without S9 activation (MRID 42021801). In an unscheduled DNA synthesis (UDS ) in mammalian cells test, no mutagenic response was observed in primary rat hepatocytes (MRID 42021701).

The available in vitro data have shown positive results, and neither an in vivo mammalian bone
marrow chromosomal aberration test nor an in vivo mammalian erythrocyte micronucleus test is
available to examine the effect of TCM on an intact mammalian system. A dditionally, there is indirect food-contact exposure, when TCM is us ed on food-contact surfaces and hand-to-mouth
behavior on nonfood contact treate d surfaces. To further evaluate mutagenicity and potential carcinogenicity of TCM via dietary exposure, th e agency expects to require one of these in vivo mutagenicity studies.

Table 13 – Summary of Subchronic and Developm ental Toxicity and Mutagenicity Data for
TCM

Guideline

No. Study Type/Purity (%) MRID Results

Subchronic ToxicityPage 25 of 37 Guideline

No. Study Type/Purity (%) MRID Results

870.3100 90-day Oral Study (Rodent) (technical grade TCM) 43064301 NOAEL = 30 mg/kg/day LOAEL = 150 mg/kg/day, based on the observation of histological lesions (engorgement of small blood vessels of the adrenal gland, brain, kidneys, liver, lung, and pituitary gland).
Developmental Toxicity

870.3700 Developmental Toxicity (TCM, 94.4%) 43614301(Rabbit) Systemic Toxicity NOAEL < 30 mg/kg/day (LDT) and LOAEL

≤ 30 mg/kg/day, based on depressions in mean maternal body weights and decreased mean feed consumption values.
Developmental toxicity NOAEL = 120 mg/kg/day (HDT)

43614302 (Rat) Maternal NOAEL < 62.5 mg/kg/day (LDT) and LOAEL ≤ 62.5 mg/kg/day, based on clinical signs at several or all dose levels (alopecia, gasping, altered respiration, salivation, lethargy, chromodacryorrhea), reduction in body weight gains, and statistically significant trends fo r decreased food consumption at
all dose levels.
Developmental toxicity NOAEL = 500 mg/kg/day (HDT)
Mutagenicity Studies

870.5300 Ames Assay (TCM, 95.9%) 42148801 Positive result at 25 μg/plate for TA98 and TA100, both with
and without metabolic activation.

870.5375 In vitro Mammalian
Chromosomal Aberration Assay (TCM, 95.9%) 42021801 Positive result at 50 μg/mL in Chinese hamster ovary (CHO)
cells both with and without metabolic activation.

870.5550 In vitro Unscheduled DNA Synthesis (UDS) in Mammalian
Cells (TCM, 95.9%) 42021701 Negative result in primary rat hepatocytes.

Notes: NOAEL = No Observed Adverse Effects Level; LOAEL = Lowest Observed Adverse Effects Level; NA = Not Applicable; LDT=Lowest Dose Tested; HDT=Highest Dose Tested



## Other Toxicological Effects

As indicated in the environmental fate section (Appendix B ), TCM breaks down into hypochlorous acid and melamine in water; mela mine is further degraded to ammeline and ammelide before yielding cyanuric /isocyanuric acid. Melamine is relatively stable and would be expected to remain as a re sidue on food contact surfaces.



## Melamine

Melamine was administered to Spragu e–Dawley rats (Crl:CD [SD]VAF/Plus ® ; 6-7 weeks old) (8/sex/group) by gavage for 14 consecutive days, followed by an 8-day recovery period for the high-dose group, and monkeys were treated with me lamine by gavage for 3 months, followed by a 4-week recovery period for the high-dose group (Early RJ, et al. , 2012). Mortality was reported in the high-dose rats, as well as compound-rela ted clinical signs (re d urine), decreased body weights, and changes in clini cal pathology (increased serum ur ea nitrogen and creatinine) and anatomical pathology (renal tubu lar cell debris, crystal depositi on, and hyperactiv e regeneration of renal tubular epitheliu m) in the mid-dose rats. For the monkey study, the kidneys and/or liverPage 26 of 37 weights were increased in high- dose monkeys with increased serum alanine aminotransferase level, decreased red blood cell, hemoglobin, a nd hematocrit and increased reticulocyte counts with microscopic findings, and demonstrated in creased extramedullary hematopoiesis in bone marrow, liver, or adrenals. Microscopic examin ation revealed nephrotoxicity, pericarditis, and increased hematopoiesis. Nephrotoxicity was also noted in mid-dose group. The NOAEL was determined to be 140 mg/kg/day (low dose) for rats following a 14-day oral administration and 60 mg/kg/day (low dose) for m onkeys in the 3-month study.

Pregnant Sprague-Dawley rats administered melamine by gavage demonstrated maternal toxicity, including increased inci dences of clinical signs and death, lower body weight gain and food intake, and increases in heart, adrenal gland and kidne y weights, at the high-dose. Histopathological examinations revealed an increase in inci dences of congestion, tubular necrosis/degeneration, crystals, cas ts, inflammatory cells in tubul es, tubular dilation and tubular hyaline droplets in the maternal kidneys. Developmental toxic effects included a decrease in fetal weight, an increase in the incidence of skeletal variations, and a delay in fetal ossification; however, no histopathological change s were observed in fetal kidne ys (one fetus/litter). The NOAEL was determined to be 400 mg/kg/day for both maternal and developmental toxicity (Kim SH, et al. , 2011).

An et al. (2010) investigated the hypothesis that melamine could impair cognitive function through impairment of hippocampal function. Male Wi star rats were administered melamine for four weeks at a dose of 300 mg/kg by oral ga vage. The result of Morris water maze (MWM) testing showed a significant defi cit in learning and memory in the melamine group compared to controls, and the field excitatory postsynaptic potentials (fEPSPs ) slopes were significantly lower in the melamine group as well, as demonstrat ed by the long term potentiation (LTP) from Schaffer collaterals to CA1 region in the hippocampus test.



## Melamine and Cy anuric Acid

Although melamine alone presents low toxicity in a range of species, availa ble data showed that melamine and cyanuric acid together increased renal toxicity when co-ingested by animals. Renal failure and the presence of renal crystals were reported in studies in which cats were fed increasing amounts of melamine and cyanuric acid (Brown et al. , 2007; Puschner et al. , 2007). Such effects were confirmed in a study (Dobson et al ., 2008) in which rats were administered ammeline or ammelide alone (both analogs of mela mine), a mixture of melamine and cyanuric acid, and a mixture of all four compounds. The te st results showed that neither ammeline nor ammelide alone produced any renal effects, but th e mixtures produced significant renal damage and crystals in nephrons. Recent studies conclude d the same findings at the high-dose level in rats, even after only 3 days e xposure to melamine in combination with cyanuric acid (Park D, et al. , 2011).

Recently, a NOAEL of 2.1mg/kg bw/day for males and <2.6 mg/kg bw/day for females, and benchmark dose lower confidence limit (BMDL) values of 1.6 mg/kg bw/day for both sexes were established in a 28-day dietary study of co -exposure of melamine and cyanuric in F344 rats (Gamboa da Costa, G, et al., 2012).Page 27 of 37 Pharmacokinetics studies of melamine and cyanuric acid and th eir combinations in rats revealed that the pharmacokinetic profile was significantly altered when animals were administered equal amounts of melamine and cyanuric acid as a preformed melamine cyanurate complex, with reduced bioavailability of both compounds, lo wer observed maximum serum concentrations, delayed peak concentrations, and prol onged elimination half-lives (Jacob CC, et al. , 2012). Additional studies (Pang J, et al. , 2011) showed that animals co -administered equal amounts of melamine and cyanuric acid resulted in signifi cant changes in toxicokinetic parameters of melamine, including lower maximum concentra tion and area under curve, higher plasma
elimination half-life and volume of distributi on, and significantly high er concentration of melamine in the kidneys, when compared with melamine alone group (P < 0.05).



## National Toxicology Program (NTP) Studies on Melamine and Cyanuric Acid

When analyzing the outbreak of kidney failure and death of hundreds of cats and dogs in the USA caused by intentionally adu lterating pet food with melami ne and several degradates (including cyanuric acid), labor atory animals were exposed simultaneously to melamine and cyanuric acid. It was discovered that melamine cya nurate crystals formed and that these crystals occluded the nephron’s distal t ubules which resulted in the ne phrotoxicity observed in these animal species.

An investigation of the high incidence of hos pitalization of many infants and toddlers with nephrolithiasis (kidney stone), some with renal failure, and some confirmed deaths in China revealed that the cause was powdered infant form ula intentionally adulterated with melamine.

Human exposure to melamine and/or cyanuric acid from sources other than food are outlined in the NTP's document “NTP Research concept: melamine/cyanuric acid” (http://ntp.niehs.nih.gov/ntp/Noms/Final_ResC oncept/Melamine_Cyanuric.pdf) as follows.

(1) "Melamine resin, a hard thermosetting polym er made from melamine and formaldehyde, is widely used in the U.S. in the form of k itchenware, including plates, bowls, mugs and
utensils.......leach considerable am ounts of melamine monomer. A migration of up to 2.5 mg
melamine/100 cm 2 was observed under conditions that simulate an exposure to ho t acidic foods";

(2) "Cyanuric acid is an FDA-accepted component of feed-grade biuret, a ruminant feed additive. The additive can legally contain up to 30% of cyanuric acid (and triuret)";

(3) "Cyanuric acid is used as a stabilizer and disinfectant in sw imming pool water......the maximum allowable limits, dictated by state or local codes, are typi cally 100 ppm (National Swimming Pool Foundation (2007) Instructor Manual Certified Pool-SPA Operator Handbook

, Appendices A-17. Available at http://www.nspf.com/Docum ents/CPOI_Manual_2007.pdf ) A study on the water ingestion during swimming activi ties in a pool indicates that the amount of water swallowed by non-adults (under 18 years ol d) was on average 37 mL and reached 154 mL. Considering this scenario and a cyanuric ac id concentration of 100 ppm in a swimming pool, these volumes translate into an ingestion of up to 15.4 mg of cyanuric acid by non-adults. Furthermore, the possibility that non-prof essionally managed pools and hot tubs mayPage 28 of 37 accidentally be treated with much higher con centrations of cyanuric acid should not be disregarded."

The information outlined above indicated that th ere are a number of possi ble sources of exposure to melamine and cyanuric acid in the normal daily life of an average indi vidual. In addition, the human food chain could be possibly indirectly or directly contamin ated with melamine and/or its derivatives, includi ng cyanuric acid.

The National Toxicology Program (NTP) proposed to further investigate and determine: (1) the combined toxicological implications of a concurrent exposure of different species to melamine and derivatives, (2) the possible occurrence of biomarkers predictive of renal failure upon exposure to melamine and derivatives, and (3) th e pharmacokinetics of melamine and cyanuric
acid to better understand the full toxicokinetic profile of the individua l triazines and their combinations. The studies, conducted with co-admin istration of melamine and cyanuric acid “in a phased manner”, are outlined in the final research proposal as follows:

(1) “A pharmacokinetic study in rats with mela mine and cyanuric aci d when administered individually, in a time-staggered manner as a separate base and acid, simultaneously as a separate base and acid and simultaneously as a pre-formed compound (melamine cyanurate)” This study was completed and publis hed (Jacob, CC, et al., 2012).

(2) “28-day and 90-day studies in rats to dete rmine the NOAEL of a combined treatment with melamine and cyanuric acid. Additional studies in cluding an evaluation of systemic toxicity and/or specialized organ systems toxicity after long term (chronic) exposure will be considered following an evaluation of the subchronic study data.” The 28-d ay gavage study in rats was completed and published (Gamboa da Costa, G, et al., 2012).

(3) “Metabolomic and proteomic early biomar kers of melamine + cyanuric acid-induced nephrotoxicity......by non-i nvasive methods.”

(4) “Investigate the pharmacokinetics and dete rmine the NOAEL of a combined exposure to melamine and cyanuric acid in a miniature pig model deemed to be representative of the human kidney anatomy and physiology.”

The agency will be in contact with the NTP Proj ect Leader to follow up with testing progress and
obtain test results once available.Page 29 of 37 

# Appendix B Environmental Fate

Environmental Fate Properties

The agency has reviewed environmental fate da ta submitted in support of registration review from submitted OCSPP 835 laboratory studies, the EPI-WEB 4.1 program (http://www.epa.gov/oppt/expos ure/pubs/episuitedl.htm ), and the open literature. Many of these
literature sources were included in the High Production Volume (HPV) submissions (http://www.epa.gov/hpv/pubs/summaries/tricltrz/c 14659rs.pdf), while others were obtained directly from the Science Dire ct website (http://www.sciencedir ect.com/), and the European Food Safety Authority (EFSA, http://dar.ef sa.europa.eu/dar-web/provision, search for cyromazine).

These data sources indicate th at the parent compound TCM readily dechlorinates in water to form hypochlorous acid and the degradate mela mine (CAS # 108-78-1, dechlorinated parent compound). As a result, environmental exposure to TCM from the registered uses will be minimal. In addition, environmental exposure to th e TCM degradates is expected to be minimal
based on the limited indoor use as a food or nonf ood contact surface sanitizer and the further microbial degradation of TCM de gradates. Melamine degrades by microbial action to sequential metabolites ammeline (CAS # 645-92-1, once-deaminated and hydroxylated melamine), ammelide (CAS # 645-93-2, twice-deaminated and hydroxylated melamine), and cyanuric/isocyanuric acid (CAS # 108-80-5, deam inated melamine with three hydroxyl groups), biuret (CAS # 108-19-0, two urea molecules co mbined), and urea (CAS # 57-13-6) which ultimately degrades to form ammonia (NH 3 ) and carbon dioxide (CO 2 ). Other than biuret and urea, all of the above compounds contain a symm etric triazine ring. Cyanuric/isocyanuric acids are tautomers with readily-interchangeable ke to and enol groups, depending on the pH.

TCM and its degradates/metabolites are highly wate r soluble, weakly sorbed, and will be in the water phase in both soil and wa stewater treatment plants (WWTPs). Melamine and its metabolites do not degrade under abiotic conditions (presence of water and/or light without microorganisms). However, the metabolites cont aining the triazine ring are more degradable under anaerobic conditions in the presence of mi croorganisms (acclimated or unacclimated) or when acclimated microorganisms are present under aerobic conditions. Th e degradation pathway
of TCM leads to the formation of CO 2 and NH 3 ; which are released into the air or incorporated
into microbial biomass. The environmental fate properties of TCM, its metabolites, and related compounds are summarized in Table 14.

Figure 2 illustrates the degrad ation pathway for a number of compounds with a symmetric triazine ring (including melamine) to form cyanur ic/isocyanuric acid. As a result, environmental fate of these compounds depends on the environmental fate of cyanuric acid. Cyanuric/isocyanuric acid is not prone to abiotic degradation (hydr olysis or phototolysis) and is more resistant to degradation under aerobic cond itions or in the presence of unacclimated microbes. However, anaerobic conditions or acclimated microbes faci litate its eventual degradation to CO 2 and NH 3

.Page 30 of 37 Figure 2 – Formation and Decline of Cyanuric Acid from Compounds with a Symmetric Triazine Ring.

TCM and Metabolites

TCM and its metabolites melamine, ammeline, amme lide, and cyanuric/isocyanuric acid share a symmetric triazine ring with s-tr iazine herbicides (e.g. atrazine, simazine) and an insect growth regulator (cyromazine). However, TCM and its metabolites are not to be confused with agronomic herbicides that are cl assified as s-triazine compounds which have alkyl side chains, and therefore, it is not considered as part of the triazine cumula tive assessment (See Section 3.4.2 above). Therefore, a total toxic residue appro ach, as would be used for s-triazine compounds, will not be applied to assess environmental exposure.

TCM (parent compound) is registered as a food- contact sanitizer and di sinfectant with indoor uses and produces hypochlorous acid (an oxidi zing agent) when dissolved in water. Hypochlorous acid is a weak acid (pKa (dissociation constant) of 3.7 x 10 -8 at 25 °C). Based on this pKa, 50% will be in hypochlorous acid form (HOCl) and 50% will be in hypochlorite ion (- OCl) form at pH 7.4 (pH = -log10 (3.7 x 10 -8 ). As the pH increases above 7.4, the amount of hypochlorite ion present will incr ease. Conversely, as the pH decreases below pH 7.4, the amount of hypochlorous acid present will increa se. Hypochlorous acid reacts readily with ammonia, amines, sulfides, sulfites, and nitrites present in the water. Antimicrobial activity
results from the oxidation reacti ons with microbial enzyme syst ems. These oxidation reactions consume residual free chlorine, with the concu rrent reduction of hypochlor ous acid/hypochlorite to chloride anions. However, the release of hypochl orous acid is reversible and it can reattach to the melamine, ammeline, ammelide, and cyanuric acid molecules if additional free hypochlorous
acid is added. Cyromazine Atrazine

Simazine Formation and Decline of Cyanuric Acid from Compounds with Symmetric Triazine Ring

Chlorinated
Isocyanurates

Melamine Trichloromelamine

Ammeline

Ammelide

Cyanuric/Isocyanuric Acid

Biuret

Urea

CO 2 NH 3Page 31 of 37 The TCM degradates, cyanuric acid and isocyanuri c acid (dechlorinated parent compound) exist in a pH-dependent equilibrium. These two aci ds are keto/enol tautom ers which interconvert readily depending on the pH of the environment. The cyanuric acid (enol form with three hydroxyl groups (-OH) attached to tr iazine ring) is the primary fo rm found at lower pH values, while the isocyanuric acid (keto form, with three oxygens double-bonded w ith triazine ring) would be found at higher pH values. The pKa values for cyanuric acid are 6.88, 11.4, and 13.5 (http://www.drugfuture.com/chemdata /cyanuric-acid.html ). For more details, please see Table

14.Docket Number EPA-HQ-OPP-2012-0682www.regulations.govPage32of37Table 14 – Environmental Fate Data for TCM and Related Compounds Containing a Symmetric Triazine RingTest Guideline(Number)Test SubstanceResultReferenceCommentHydrolysis(835.2120)Isocyanuric acidand sodiumisocyanurateRapid degradation to cyanuric acid Hu, 19822MRID00056479Cyanuric acid was stablepp. 10-12 of HPV document1Photodegradationin Water(835.2240)MonosodiumcyanurateRapid degradation to cyanuric acid from hydrolysis (nophotodegradation)_Hu, 19822MRID00056479Cyanuric acid was stablepp. 10-12 of HPV document1Batch Equilibrium(835.1230, OECD106)Cyanuric acid Average Kocof 51 ml/g Michael andCummings,19823Similar to OECD 106Unknown Cyanuric acidAerobic Conditions:Degrades slowly under aerobicconditions. Some microorganisms can degrade cyanuricacid if acclimated and organic nutrients are presentAnaerobic conditions:Readily biodegradableZeyer et al.,19804Cyanuric acid formed CO2and NH3p. 15 of HPV documentUnknown Cyanuric acidAerobic conditions:30 and 15% degradation ofcyanuric acid in 48 h forPseudomonas sp. AndAchromobacter sp. (mesophilic bacteria)Ernst andRehm. 19905Cyanuric acid formed CO2and NH3pp. 17-18 of HPV documentBacteria isolatedfrom corn fieldUnknown Cyanuric acidAerobic conditions:58% degradation of cyanuric acidin 8 days in FeS medium under aerobic conditionsAnaerobic conditions:100% degradation of cyanuricacid in 4 days under anaerobic conditions in cysteinemediumJessee et al.,19836Unknown bacterium from acclimated bottomsediment downstream of WWTP in Illinois thattreats wastewater from trichloro isocyanuric acidmanufacturing facilityCyanuric acid formed CO2and NH3pp. 19-20 of HPV documentUnknown Cyanuric acidAnaerobic conditions:100% degradation of triazinering when additional source of nitrogen was presentNo cell growth in absence of additional nitrogen sourceCook et al.,19857PseudomonasNRRL B-12227 and B-12228 andKlebsiellapneumoniae(mesophilic bacteria)Cyanuric acid formed CO2and NH3pp.21-22Unknown Cyanuric acidAerobic conditions:100% degradation after 8 h (aeratedactivated sludge, 2.5 mg/l O2); 42% degradation after 9 h(aerated activated sludge, 8.7 mg/l O2); 0% degradation(highly aerobic system, O2concentration not specified)Anaerobic conditions:25-50% degradation in 48 h and100% in 72 h (sewage); 95-98% degradation in 72 h(nutrient broth); 52-100% degradation in 9-23 days (6soils and muds)Saldick, 19748Cyanuric acid formed CO2and NH3pp. 22-23 of HPV documentPage33of37Test Guideline(Number)Test SubstanceResultReferenceCommentUnknown Cyanuric acid 100% degradation in unknown time Sisodia et al.,19969Degradation of cyanuricacid occurs in absenceof nitrogen when triazine-acclimated bacteria arepresentCyanuric acid formed CO2and NH3pp. 24-25 of HPV documentUnknown Cyanuric acid BOD5and COD of 6.5 and 11 mg/l Unknownreference,possibly Zeyeret al., 19804p. 15 of HPV documentAerobic AquaticMetabolism(835.4300)Atrazine 70-80% of atrazine mineralization in constructed wetlandfed by river draining treated corn fields; Wetland waterdemonstrated 60-80% of atrazine mineralization in onemonth; 75-81% mineralization in river water samplescontaining acclimated microorganisms; Cedar bog didnot demonstrate mineralization of atrazine becausemicroorganisms were not adaptedAnderson etal., 200210Atrazine totally mineralized in the river andconstructed wetland that containedmicroorganisms thatare acclimated todegradation of triazine rings from herbicides.Atrazine did not mineralize in wetland withoutacclimated microorganisms. Atrazine formedCO2and NH3None NumeroustriazineherbicidesCyanuric acid was end product (degradate) of S-triazinesthat reached WWTPs and was not degraded significantlyKontchou andGschwind,199811Mixed culture of microorganisms in laboratorytest.None Melamine resin 85 and 54% of melamine in sludge degraded by 84 daysin waste paper and plant residue compost, respectively.Initial melamine concentration of 200 mg/kg dry matter;Clumps of melamine resin slowed degradation process;Half-lives of melamine in waste paper and plant residuecomposts were 45 and 84 days, respectively. Sequentialdegradates ammeline, ammelide, and cyanuric acidreached up to 150, 2,400,and 4,000 mg/kg dry matter(ammelide and cyanuric acid degraded slower than thepile shrank)Tian et al.,201212Composting study using melamine resin fromwaste paint sludge (WPS)Melamine resin (melamine formaldehyde) is apolymer that can exist as an adhesive, a foam, orrubber13and can be used to create moldedproducts, adhesives, and surface coatings14Temperatures reached up to 55oC (131oF) instudy (thermophilic phase.) Most temperaturesbetween 25 and 40oC (78-104oF) in study(mesophilic phase).Rates were 0, 20 and 100 g/kg WPS-waste papercompost and 20 and 100 g/kg WPS-plant residuecompostBoth composts increased plant growth incucumber (Cucumis sativus), radish (Raphanussativus), and lettuce (Lactuca sativa)Page34of37Test Guideline(Number)Test SubstanceResultReferenceCommentNone MelamineformaldehyderesinMelamine degraded 73-95% and 55-74% in inoculatedvs uninoculated compost, respectively. Melamineconcentrations increased in the thermophilic phasebecause degradation was slower than pile shrinkage.Degradate ammeline formedand degraded quickly todegradates ammelide and cyanuric acid in thermophilicphase. Ammelide and cyanuric acid degradation slowerthan pile shrinkage (leadingto higher concentrations).Melamine degraded more slowly in uninoculatedcompost than in inoculated compost (Mesophilic phase).Ammeline degraded quickly in mesophilic phase toammelide and cyanuric acid which degraded faster ininoculated conditionsTian et al.,201215Temperatures reached up to 55oC (131oF) instudy for 91 days (thermophilic phase) andstayed at 30oC (86oF) for 56 days (mesophilicphase). Study had mostly thermophilic phasewhich concentrated melamine. Pile shrank 50%by 91 days of thermophilic conditionsNone Atrazine Atrazine degradation (removal of side chains) occurredprimarily by fungi. Atrazine degradation did not occur inabsence of functional bacteria or fungiLevanon,199316Delanco silt loam soil from monoculture cornfield with acclimated organismsNone Melamine,Cyanuric acid,DeethylsimazineComplete (100%) mineralization of melamine, cyanuricacid, and deethyl simazine (triazine herbicide) as solesource of nitrogen in fluidized bed for waste-specifictreatmentHogrefe et al.,198517Pseudomonas sp.(mesophilic bacteria) wasprimary removal organism835.4100 Melamine fromappliedcyromazineHalf-lives of 88-217 days in laboratory aerobic soil(topsoil) at 20oCEuropean FoodSafetyAuthority(EFSA), vol.118Aerobic soil metabolism (top layer of soil underoxygenated conditions); Cyromazine is asymmetric triazine used as an insect growthregulator; CO2and non-extractable residueswere primary degradates of melamine indicatingthat mineralization of intermediary compoundssuch as ammeline, ammelide, and cyanuric acidwas complete835.4200 Melamine fromappliedcyromazineSlower mineralization of cyromazine in laboratoryanaerobic soil metabolism studies with no detectablemelamineEuropean FoodSafetyAuthority(EFSA), vol.118Anaerobic soil metabolism (top layer of soilfollowing rain); Cyromazine is a symmetrictriazine used as an insect growth regulator; Evenunder sterile conditions, non-extractable residueswere produced indicatingthat some degradationoccurredPage35of37Test Guideline(Number)Test SubstanceResultReferenceComment835.6100 Melamine fromappliedcyromazineHalf-lives of 24-54 days in terrestrial field dissipationstudies in Canada and Europe (colder climate); Medianand 90thpercentile half-lives of 31 and 52 days,respectivelyEuropean FoodSafetyAuthority(EFSA), vol.118Cyromazine is a symmetric triazine registeredfor use as an insectgrowth regulator1.http://www.epa.gov/hpv/pubs/summaries/tricltrz/c14659rs.pdf2.H. Hu (1981) Hydrolysis study of aqueous sodium salt solutionsof 2,4,6-trihydroxy-1,3,5-triazine. Princeton Chemical Researchand Development Centre. MRID 00056478.3.Michael, P., and Cummings, I., “Cyanuric Acid AbsorptionOn Soils”, Monsanto Report ES-82-SS-94, December, 1982.Method:OECD 106.4 An Index of Refractory Organics, T. Helfgott, F. Hart andR. Bedard, EPA 600/2-77-174, 1977, pp. 1-7, 24-25. Process for thebiological degradation of cyanuric acid, J. Zeyer, R. Hütter, and P.Mayer, UK Patent App. 2,025,919 (Jan. 30, 1980).5 Degradation of Cyanuric Acid by Immobilized Bacteria, C. Ernst and H. Rehm, DECHEMA Biotechnol. Conf., 4(Pt. A), 1990, 577-580.Method:J. Biol. Chem. 195, 265-275 (1951).6 Anaerobic Degradation of Cyanuric Acid, Cysteine, and Atrazineby a Facultative Anaerobic Bacterium, J. Jessee, R. Benoit, A.Hendricks, G. Allen, and J. Neal, Appl. and Environ. Microbiol.,1983, 45(1) 97-102.Method: J. Water Pollution Control Fed., 39, 181-187 (1967);J. Water Pollution Control Fed., 40, 306-319 (1968)7 Ring cleavage and degradative pathway of cyanuric acid in bacteria, A. Cook, P. Beilstein, J. Grossenbacher, and R. Hütter, Biochem. J., 1985, 231, 25-30.Method: J. Water Pollution Control Fed.,39, 181-187 (1967); J. Water Pollution Control Fed., 40, 306-319 (1968)8 Biodegradation of Cyanuric Acid, J. Saldick, Applied Microbiology, 1974, 28(6) 1004-1008.Method:(1967) J. Water Pollut. Contr.Fed. 39:181-187; Biodegradabilityof organic chemicals isolatedfrom rivers. (1963) Proc. 18th Ind.Waste Conf. Purdue Univ. 115:278-282.9 Continuous Culture Biodegradation of Simazine’s Chemical Oxidation Products, S. Sisodia, A. Weber, and J. Jensen, Water Research, 1996, 30(9) 2055-2064.Method:S-triazine biodegradationwas investigated as a function of continuous culture process growth rate.10 Anderson, K.L., K.A. Wheeler, J.B. Robinson, and O.H. Tuovinen. 2002. Atrazine Mineralization Potential in Two Wetlands. Water Research 36(2002): 4785-4794.11 Kontchou, C.Y and N. Gschwind. 1999. Biodegradation of S-Triazine Compounds by a Stable Mixed Bacterial Community. Ecotoxicology and Environmental Safety 43 (1999): 47-56.12 Tian, Y., L. Chen, L. Gao, F.C. Michel, Jr., H.M. Keener,M. Klingman, and W.A. Dick. 2012. Composting of waster paint sludge containing melamine resin and the compost’s effect on vegetablegrowth and soil water quality. J. Hazard. Mater. http://dx.doi.org/10.1016/j.jhazmat.2012.09.013. (Article in press)13 http://en.wikipedia.org/wiki/Thermosetting_plastic14 http://www.thefreedictionary.com/melamine+resin15 Tian, Y., L. Chen, L. Gao, F.C. Michel, Jr., C. Wan, Y.Li, W.A. Dick. 2012. Composting of waster paint sludge containing melamine resin as affected by nutrients and gy7psum addition andmicrobial inoculation. Environmental Pollution 162:129-137.16 Levanon, D. 1993. Roles of fungi and bacteria in the mineralization of the pesticides atrazine, alachlor, malathion, and carbofuran in soil. Soil Biol. Biochem. (25):1097-1105.17 Hogrefe, W., H. Grossenbacher, Y. Kido, A.M. Cook., and R.Hutter. 1985. Biodegradation in a fluidized bed of real wastes containing s-triazines. Conservation and Recycling (8):85-90.18 http://dar.efsa.europa.eu/dar-web/provision (search for cyromazine)Page 36 of 37 Hypochlorous Acid 17

TCM is used as a water solution wh ich forms melamine and hypochlorous acid (HClO), an oxidizing agent and the biocidal agent. HClO is a weak acid and, depending upon pH, can further disassociate into a hydr ogen ion and a hypochlorite ion (ClO - ), in a reversible reaction. The antimicrobial activity of chlorine compounds depends primarily on HClO which can rapidly
cross cell membranes. While ClO - has some disinfection potential it requires longer contact time.
It is HClO and ClO - that are the active disi nfecting moieties of TCM.

HClO

ு௬௣௢௖௛௟௢௥௢௨௦ ௔௖௜ௗ ՞ H +

ு௬ௗ௥௢௚௘௡ ௜௢௡ ൅ ClO -

ு௬௣௢௖௛௟௢௥௜௧௘ ௜௢௡ Eqn. 1

The acid dissociation constant (K diss or K
a ) of HClO is 3.4 x 10 -8 (or pK

a 18 = 7.7). Hence at any given time, the concentration of HClO is not ve ry high; however, as it is consumed, more HClO is formed. It also means that at pH 7.7, the ratio of HClO to ClO - is 50:50. A literature report (Connell, 1996) estimated the changes due to protolysis (hydrolysis) of a solution containing
ClO - . The graphical data is converted into the num erical data on the pe rcent of HClO and ClO from pH 4 to 11 as shown in Table 15. In dilute aqueous solutions these re actions occur rapidly.

Table 15 – Changes in Protolysis (Hydrolysis) of Solutions Containing Hypochlorite Ion.

pH % HClO % ClO 4.0 100 0

6.3 95 5

7.2 70 30

7.7 50 50

8.0 21 79

11 0 100 Disinfection is more efficient at lower pH than at higher pH because HClO is a more efficient
disinfectant than ClO - , and as indicated above there is a higher proportion of HC lO at lower pH than at higher pH values.

Available chlorine (AC) is a measure of the equiva lent weights of oxidizing agents, with chlorine gas as a basis— i.e ., normalizes chlorine-containing compoun ds to allow comparison of oxidizing potential. The concentration of hypochlorite (o r any other oxidizing disinfectant) may be expressed as available chlorine by determini ng the electrochemical equivalent amount of elemental chlorine (Cl 2 ) to that compound (USEPA 1986).

A mole of chlorine gas reacts with water to fo rm a mole of HClO and a mole of protons (or hydrogen ions) and a mole of chloride ions. The oxida tion state of chlorine is 0 in chlorine gas,
+1 in HClO and -1 in Cl - , therefore chlorine is both oxidi zed and reduced in this reaction.

Cl 2

஼௛௟௢௥௜௡௘,௚௔௦ ൅ H 2 O

ௐ௔௧௘௥ ՜ HClO

ு௬௣௢௖௛௟௢௥௢௨௦ ௔௖௜ௗ ൅ H +

ு௬ௗ௥௢௚௘௡ ௜௢௡ ൅ Cl -

஼௛௟௢௥௜ௗ௘ ௜௢௡ Eqn. 2

17 Details regarding Hypochlorous Acid can be viewed in the Na & Ca Hypochlorite Registration Review Docket at http://www.regulations.gov in docket EPA-HQ-OPP-2012-0004.

18 pK

a = -log 10 K

aPage 37 of 37 Elemental Cl 2 oxidizing potential : one mole of elemental chlorine is capable of reacting with two electrons to form inert chloride, as shown in the reaction equation below (USEPA, 1986). The AC is 70.91 grams per mole (identical to the molecular weight of Cl 2 ).

Cl 2

஼௛௟௢௥௜௡௘ ൅ 2݁
ି

ா௟௘௖௧௥௢௡௦ ൌ 2Cl -

஼௛௟௢௥௜ௗ௘ ௜௢௡ Eqn. 3

ClO - oxidizing potential : one mole of ClO - may react with two electrons to form chloride, as shown in the reaction equation below. Hence, one mole of hypochl orite is equivalent (electrochemically) to one mole of elemental chlo rine, and may be said to contain 70.91 grams of AC (identical to the molecular weight of Cl 2 ) (USEPA, 1986).

ClO ି

ு௬௣௢௖௛௟௢௥௜ௗ௘ ൅ 2݁
ି

ா௟௘௖௧௥௢௡௦ ൅ 2H +

ு௬ௗ௥௢௚௘௡ ௜௢௡ ൌ Cl -

஼௛௟௢௥௜ௗ௘ ௜௢௡ ൅ 2H 2 O

ௐ௔௧௘௥ Eqn. 4

Chlorine, in addition to providi ng disinfection or oxidation of microorganisms, is consumed through oxidation or substitution reactions with i norganic substances such as ammonia, iron, manganese, sulfides and some othe r organic substances (e.g., phenols).

In the surface water and wastew ater streams, possible transfor mation products like chlorine, HOCl, and ClO 3 - may be present, but th e determination of exact amounts of these moieties cannot be calculated as these will depend on envi ronmental conditions (e.g., pH of the medium, temperature, ionic strength).