**.#'f.f>"1" 4~** **~**

# ~~**_)** UNITED S TATE S EN V I RO NME N T A L PR O T EC TI O N A G ENC Y

WASHINGTO N

, D.C. 20460



### MEMORAN D UM

Date: March 29, 20 1 6 OFFICE OF

CHEMICAL SAFETY AND
POLLUTION PREVENTION



### SU BJECT: Dicamba. Sec tion 3 Registration fo r the Amended Use ofDicamba on Dicamb a-Tolerant

Cotton. Summary of Analytical Chemistry and Residue Data.

PC Code: 02980 1

. 029802. 029806, 128931

, 1 2894 4

, & 

### 129043

D ec i s ion No.: 46 7997

Petitio n No.: 2F8067

R isk Assess m en t Ty p e: NA

TX R No.: NA

MR IO Nos.: 48728701-48728704



### FROM: Pet e r Savoia

. C h e mi s t 

# OA- -le 

# L

Risk Assessment Branch V N II OP Barc o de: 0 408384

Rc2 i stra ti o n No.: 524-582

R eg ul ato r y Act i oi 1 : Amended Section 3 R eg i s trati o n

Case No.: 0065

CAS No.: 1918-00-9

40 CFR : 

### § 180. 227



### He alth Effects Di v i s ion ( 7509P ) )

. ,



### Mich ae l S. Met zge r
, Branch

# Chief**~** t-.ltf.<_ 

# I }

, T HRU:

TO: Donna S. Davi s
, Chemist

Risk Assessme nt Branch V / VII

Health Effects Di v i s ion (7509P)

William A. I rw in

, Ri s k Assessor

Risk Assessme nt Branch V N II

Health Effec t s Divi s ion (7509P)

And
Kathryn Montague / Michae l Walsh

, Ri s k Manager

Herbicid e Branch

R egistrat ion Division (7505P)



### E x ec u t i ve S um mary

Monsanto has s ubmitt e d p e tition PP #2F 8067 requ es tin g Sect ion 3 re g i s tration for the amended u se of

dicamba on dican1ba-tol e rant cotto n

. Mon s anto i s r e questing to amend th e co tt o n undelinted seed

t o l e r a nce from 0.2 ppm to 3.0 ppm and establish a n ew tolerance for co tton g in byproducts at 70.0

ppm. Dicamba i s a se l ec ti ve benzoic acid h erb i c id e u sed for controllin g w ee ds prior to their

e mergence

. It i s avai l ab l e for use in either acid o r sa lt forms with regist e r e d u ses being maintained o n

a wide variety of crop and li ves tock commoditie s. Permanent tolerance s are es tablished under 40

C FR § l 80.227 (a)( l ) for d i camba and it s 3

, 6-dichloro-5 -h yd rox y b enzo i c ac id (5

- 0H dicamba )

m e t abolite. Additional t o leranc es are established und e r 40CFR§180

. 227(a)(2) for dicamba and it s 3,6 -dichloro-2-h y dro xybe n zo ic acid ( als o known as 3.6 -diclorosalic ycl ic ac id or DCSA ) metab o lit e
,

as well as und e r 40 CF R § J 

### 80.227(a)(3) for dicamba, 5-0H dicamba

, and th e DCSA metabolit e.

P age I of26

### Dicamba Summary of Analytical Chemistry and Residue Data DP No. D408384



## Page 2 of 26 Dicamba is registered for pre

- plant application on cotton but not for post

- emergence treatment because crop injury could occur if it were to come in contact with cotton roots, stems, or foliage. To enable post

- emergence application, Monsanto has developed a dicam ba

- tolerant variety of cotton (MON 88701) capable of receiving treatment up to seven days before harvest. T he M1691 Herbicide (EPA Reg. No. 524

- 582) soluble liquid concentrate (SL) digl y colamine salt formulation of dicamba is proposed for use on dicamba

- t olerant cotton

. For pre

- emergent application, up to 32 fluid ounces of the M1691 herbicide (1.0 lb ae/A) may be used for pre

- plant or at planting prior to crop emergence. Using a maximum single in crop application of up to 16 fluid ounces per acre (0.5 l b ae/A)

, post

emergence treatment s are also allowed not to exceed the seasonal total in a given year

. Applications are to be made following a 7

- day re

- treatment interval (RTI) and a 7

- day pre

- harvest interval (PHI). A combined total of all applications i s not to exceed 64 fluid ounces per acre (2.0 lb ae/A) in a given

year.

Pr eviously submitted wheat, grape, asparagus, sugarcane, cotton, and soybean metabolism studie s demonstrate that dicamba is metabolized in plants by demethylation and hydroxylation. A new metabolism study on dicamba

- tolerant cotton shows a generally similar metabolic pathway to other plants; however, when compared to cotton which is not dicamba

- tolerant , signi ficantly more DCSA is

found with the dicamba

- tolerant cotton. Therefore, the newly submitted metabolism study supports including parent, 5

- OH dicamba, as well as the DCSA metabolite as the residues of concern for

dicamba

- resistant cotton for both toleranc e setting purposes and risk assessment

. Based on previously submitted metabolism studies on ruminants and poultry, the nature of the residue in livestock commodities for tolerance setting purposes and risk assessment is adequately understood to

be dicamba and DCSA.

There are adequate methods available for the enforcement of the newly proposed cotton seed and cotton gin byproducts tolerances

. PAM Vol. II Method I using Gas Chromatography/Electron Capture Detection (GC/ECD) analyses is adequate for enforcement

. New tolerances are not proposed for animal commodities; therefore a discussion of animal enforcement methods is not relevant to this petition. Dicamba is recoverable through the FDA multi

- residue method (MRM) testing protoco ls, but no data have been provided for any of its regulated metabolites. However, s ingle analyte methods

are available for enforce ment and additional MRM data are not needed.

The number and locations of the field trial studies submitted for dicamba

- resi stant cotton were in accordance with the OCSPP 860.1500 test guidelines. The data were generated using a validated data collection method and are supported by adequate storage stability data. Total residues of dicamba

and its metabolites 5

- OH dicamba, an d DCSA, expressed as parent equivalents, ranged from 0.11 ppm to 1.72 ppm in dicamba

- resistant cotton undelinted seed treated at a maximum seasonal application rate of 2.0 lbs ae/A with a single late

- season foliar application of 0.5 lbs ae/A and a PHI

of 7

- days. For gin byproducts in dicamba

- tolerant cotton , total residues of dicamba ranged from 5.06 ppm to 29.6 ppm following this same pattern of use.

An adequate processing study was submitted. Residue data was generated using an adequately

validated dat a collection method. Samples were analyzed within 30

- days; therefore, storage stability data is not required. Residues of dicamba, 5

- OH dicamba, and DCSA did not concentrate in any processed fraction.

The residue values obtained from these fiel d trial studies and the processing study were evaluated using the Organization for Economic Cooperation and Development (OECD) calculation procedures

### Dicamba Summary of Analytical Chemistry and Residue Data DP No. D408384



## Page 3 of 26 for estimating tolerances/M aximum Residue Limits (MRLs)

. Using the OECD calculation procedures, and input ting the total residue, which includes the parent compound, and its metabolites

5

- OH dicamba, and DCSA, expressed as parent, recommended tolerances of 3 .0 ppm for cotton

undelinted seed and 70 .0 ppm for cotton gin byproducts are appropriate

.

For this acti on, the establishment of a tolerance on cotton gin byproducts will not increase livestock dietary burden; therefore, no new revised tolerances on livestock commodities are require d to support this petition.

There are MRLs of 0.2 ppm in Mexico and 0.04 ppm established by Codex on cotton seed currently

established. There are currently no Mexican, Canadian or Codex MRLs established for cotton gin byproducts. Since the registrant has requested a late season use of dicamba on dicamba

- tolerant cotton, the currently established international tolerances are not adequate to cover residues likely from the newly proposed use in the U.S. In addition, the dicamba residues of concern for dicamba

tolerant cotton also include the DCSA metabolite which i s not found nor regulated in the other

common varieties of cotton

. Therefore, harmonization is not possible at this time for cotton seed. Since there are no international tolerances on cotton gin byproducts, there is no issue of international

harmonizati on relevant to that tolerance.

T he December 2005 Reregistration Eligibility Decision (RED) for dicamba requires additional rotational crop studies in order to satisfy data requirements . However, these data are not need ed if a 120

- day plantback interval is specified when dicamba is applied at the maximum seasonal rate of 0.75 lb ae/A. For greater seasonal application rates of 0.75

- 2.0 lb ae/A, only crops with established tolerances can be rotated for planting. No new rotational crop data have since bee n submitted ; therefore, the plantback restrictions noted in the 2005 RED are appropriately specified on the product

label for treating dicamba

- tolerant cotton ; and additional rotational crop data are not needed at this

time.

Regulatory Recommendations and Residue Chemistry Deficiencies

P rovided the label is amended as specified below , there are no residue chemistry deficiencies that preclude establishing permanent tolerances on cotton raw agricultural commodities (RACs) for

d icamba. A revised human health risk assessment to support the requested use on dicamba

- resistant

cotton is forthcoming.

The tolerance expressions specified in the CFR for dicamba are in compliance with HED’s Interim

Guidance on T olerance Expressions (0 5/27/2009, S.Knizner) . F or this action, the recommended

dicamba tolerances for cotton RACs are to be set under 40 CFR §180. 227(a)(3) as follows :

Tolerances are established for residues of the herbicide dicamba, 3,6

- dichloro

- o

- anisic acid, including

its metabolites and degradates, in or on the commodities in the table below. Compliance with the tolerance levels is to be determined by measuring only the residues of dicamba, 3,6

- dichloro

- o

- anisic acid, and its metabolites, 3,6

- dichloro

- 5

- hydroxy

- o

- anisic acid, and 3,6

- dichloro

- 2

- hydroxybenzoic acid, calculated as the stoichiometric equivalent of dicamba, in or on the following commodities:

Cotton, gin byproducts

................................ ....................... 70 .0 ppm Cotton, undelinted seed

................................ ........................ 3.0 ppm

### Dicamba Summary of Analytical Chemistry and Residue Data DP No. D408384



## Page 4 of 26 Note to RD

The 40 CFR §180. 227(a)(1) citation for dicamba currently lists a 0.2 ppm tolerance for cotton

, undelinted seed. This entry in the CFR should be removed, and replaced with the tolerances shown

above under 40 CFR §180. 227(a)(3).

860.1200 Directions for Use

An amended Section B is require d noting that no more than two (2) post

- emergence applications may be made past the first open boll stage when treating dicamba

- tolerant cotton is allowed.

Background

Dicamba

, 3,6

- dichloro

- 2

- methoxybenzoic acid

, is a selective benzoic acid herbicide registered for controlling a wide variety of broadleaf weeds and woody plants prior to their emergence. It has similar hormonal properties to natural auxins which induce abnormal and uncontrollable growth to disrupt normal plant functions at high concentrations

. D icamba end

- use products are available as

e ither acid or salt formulations with r egistered uses being maintained on a wide variety of crop and

livestock commodities. The chemical structure and nomenclature of dicamba and its metabolites 5

- OH dicamba, and DSCA are presented in Table 1 . The physicochemical properties of the technical grade of dicamba acid are presented in Table 2.

T able 1. Test Compound Nomenclature : Dicamba and its Residues of Concerm

. Compound

Common name Dicamba

Company experimental name MON 11900

IUPAC name 3,6

- dichloro

- o

- anisic acid or 3,6

- dichloro

- 2

- methoxybenzoic acid

CAS name 3,6

- dichloro

- 2

- methoxybenzoic acid

CAS registry number 1918

- 00

- 9 (dicamba acid), 104040

- 79

- 1 (diglycolamine salt), or 53404

- 28

- 7 (monoethanolamine salt)

End

- use product Clarity® Herbicide: SL formulation containing 4 lb ae/gal

Compound

Common name 5

- Hydroxy

- dicamba
Company experimental name 5

- OH dicamba

IUPAC/CAS name 2,5

- dichloro

- 3

- hydroxy

- 6

- methoxybenzoic acid

CAS registry number 7600

- 50

- 2

### Dicamba Summary of Analytical Chemistry and Residue Data DP No. D408384



## Page 5 of 26 T able 1. Test Compound Nomenclature : Dicamba and its Residues of Concerm

. Compound

Common name DCSA ; 3,6

- dichlorosalicylic acid

Company experimental name MON 52708

IUPAC/CAS name 3,6

- dichloro

- 2

- hydroxybenzoic acid

CAS registry number 3401

- 80

- 7



### T able 2. Physicochemical Properties of the Technical Grade Test Compound: Dicamba.

Parameter Value Reference

Melting point 114

- 116  C (PAI)

90

- 100  C (87% TGAI) Residue Chemistry Chapter of the

Dicamba RED ( D317699, C. L. Olinger

, 12/20/2005). pH 2.5

- 3.0 (87% TGAI)

Density 1.57 g/mL at 25  C (87% TGAI)

Water solubility 0.5 g/100 mL at 25  C (PAI)

Solvent solubility g/100 mL at 25  C (PAI)

dioxane 118.0

ethanol 92.2 isopropyl alcohol 76.0 methylene chloride 26.0

acetone 17.0

toluene 13.0 xylene 7.8 heavy aromatic naphthalene 5.2

Vapor pressure 3.4 x 10

- 5 mm Hg at 25  C (PAI)

Dissociation constant, pK a 1.97 (PAI)

Octanol/water partition coefficient, Log(K OW ) 0.1 (PAI)

UV/visible absorption spectrum neutral: 511 (275 nm) acidic (pH 0

- 1): 1053 (281 nm) basic (pH 13

- 14): 469 (274 nm)



## 860.1200 Directions for Use

The dicamba product used for treating dicamba

- tolerant cotton proposed for registration is the M1691 Herbicide (EPA Reg. No. 524

- 582) which is a soluble liquid concentrate (SL) formulation. This end

– use product contains 56.8% active ingredient in the form of the d iglycolamine salt of dicamba (equivalent to 4.0 lb ae/gal). A summary of the proposed directions for use taken directly from th e supplemental M1691 herbicide label provided by the registrant are presented below in Table.

### Dicamba Summary of Analytical Chemistry and Residue Data DP No. D408384



## Page 6 of 26 Table 3. Summary of Directions for Use of Dicamba

. Formulation [EPA Reg.
No.] Applic.

Timing, Type, and Equip. Max.

Applic.

Rate (lb ae 1 

### /A) Max.

No. Applic.

per Season Max.

Seasonal Applic. Rate (lb ae/A) Combined

Max. Seasonal Applic. Rate (lb ae/A) RTI 2



### (days) PHI 3 

### (days) Use Directions

and Limitations 4



### MON 88701 Cotton

M1691 4.0 lb ae/gal

SL [524

- 582] Pre

- plant,

at planting,

and P re

emergence Broadcast (20 gal/A) 1.0 NS 5 

### 1.0 2.0 7 7 Use of a COC or

MSO is not

recommended with Roundup

branded

herbicides. These adjuvants are only

used when other products require

them. For best results apply at min spray rate of

10 GPA

. Apply with ground equipment only; aerial application is prohibited. Post

emergence, Broadcast (20 gal/A) 0.5 NS 2.0

1 

### ae = Acid Equivalents

2 

### RTI = Re

- Treatment Interval

3 

### PHI = Pre

- Harvest Interval

4 

### COC = Crop Oil Concentrate; MSO = Methylated Seed Oil.

5 

### NS = Not Specified



## Conclusions

. The proposed directions for use were compared to the submitted field trial data. The field trial studies were conducted covering the expected range of applications allowed under the proposed label. This includes pre

- emergent applications of up to 1.0 lb ae/A, as well as multiple early to late post

- emergent treatments of up to 0.5 lb a e/A each for a combined annual total of 2.0 lb

ae/A. Treatments may include the use of adjuvants and can be re

- applied following a 7

- day RTI and

a 7

- day PHI. As such, the product label a llow s growers great flexibility in using dicamba to control

weeds wh en cultivating cotton. The residue data provided examining th is broad pattern of use shows

that later post

- emergence treatments give much higher residues then those made at earlier growth

stages. Following the pattern of late season use demonstrated by t he field trial data, no more than two (2) post

- emergence applications may be made after the first open boll stage when treating dicamba

tolerant cotton. Therefore, the registrant should amend the product label to include this restriction since the data pr ovided only support this pattern of use.

860.1300 Nature of the Residue

- Plants

Re registration Eligibility Decision Memo, D 317 699

, 12 / 20 / 20 05, C. L. Olinger

DER Reference: 4 8728701 .der.doc x (Nature of the Residue in Dicamba

- Tolerant Cotton )



## For non

- dicamba resistant plants, t he nature of the residue was previously determined to be

understood (D317699, C. L. Olinger, 12/20/2005). Prior plant metabolism studies were reviewed in part with the 1983 residue chemistry chapter of the dicamba registrati on standard. The studies

### Dicamba Summary of Analytical Chemistry and Residue Data DP No. D408384



## Page 7 of 26 demonstrate that dicamba is rapidly absorbed and translocated by grasses, grapes, black valentine beans, wheat and bluegrass, a s well as in soybeans. Dicamba is metabolized in plants mainly by demethylation and hydroxylation.

The registrant has developed a dicamba

- tolerant cotton (known as MON88701), which contains a gene that expresses an enzyme to catalyze the breakdown of dicamba. This enzyme

, O

- demethylase

, works by cleav ing a specific carbon

- oxygen bond converting the aro matic methoxyl group of dicamba into an aromatic hydroxyl group to form the non

- herbicidal DSCA metabolite. DSCA is a

known metabolite of dicamba found in soil, plants, and livestock

. The petitioner has submitted a new metabolism study for dicamba

- tolera nt (MON 88701) cotton (48728701.der, P . S avoia, 03/25/2013). A summary of the characterization and identification of residues is shown in Appendix I along with

the chemical names and structures of the identified metabolites illustrated in Appendix II. A summary of the data and findings follow.

For this study, m etabolism was found to occur at an appreciable rate with the f irst step in the process being demethylation of parent into the DCSA metabolite. Residue levels were considerably lower for the pre

- emergence samples in comparison to those obtained following post

- emergence treatment.
Greater residue concentrations were obtained for the cotton leaves/stems (surrogate gin byproducts) as opposed to seed . These data indicate that residues tend to be on the surface of the dicamba

- tolerant

cotton plant. Parent dicamba was identified in all the matrices tested which include both gin byproducts and in seed at lower levels. Its metabolites DCSA glucoside, DCSA, and DCGA glucoside were found to be present i n all matrices as well. The 5

- OH dicamba metabolite was not identified in any matrix. For dicamba

- tolerant cotton, DCSA glucoside was the major metabolite

obtained with the highest levels being found in the gin byproducts samples.

Based on these results, the petitioner proposes that the metabolism of dicamba in MON 88701 cotton proceeds by the demethylation of dicamba to form the DCSA metabolite. The formation of DCSA

occurs through the action of the dicamba O

- demethylase enzyme produced by the dicamba mono

oxygenase gene introduced into this variety of cotton for tolerating application of the herbicide. The petitioner notes that due to the predominance of the demethylation pathway, 5

- OH dicamba is not

observed as a metabolite fo und in either dicamba

- tolerant cotton or soybean.

In consultation with the HED Residues of Concern K nowledgebase Subcommittee (ROCKS) co

chairs on March 18, 2013, the residues of concern (ROC) in dicamba

- tolerant cotton were determined

for both toleranc e setting and risk assessment purposes

. The newly submitted data supports including parent and the DCSA metabolite, as well as the 5

- OH dicamba metabolite found in non

- resistant varieties as the residues of concern in cotton for tolerance expression and r isk assessment.

The rationale for the ROC decision is as follows. Residues present in both cotton and dicamba

tolerant cotton were comprised of dicamba, and its metabolites, 5

- OH dicamba, DCSA, and DCGA.

While the DCGA metabolite was detected in field trial samples, and can be considered of comparable toxicity to the parent compound, the available analytical enforcement methodology is not considered validated for the DCGA metabolite. However, dicamba, 5

- OH dicamba, and DCSA account for the majority of the residues in both tolerant and non

- tolerant cotton and will provide sufficient residues

with which to monitor for misuse for both tolerant and non

- tolerant cotton; therefore, the ROC for tolerance setting purposes is dicamba, 5

- OH dicamba, and DCSA.

### Dicamba Summary of Analytical Chemistry and Residue Data DP No. D408384



## Page 8 of 26 For the purpose of risk assessment, HED considered both the exposure and hazard profiles for dicamba, 5

- OH dicamba, DCSA, and DCGA and is including dicamba, 5

- OH dicamba, and DCSA as

the ROC for tolerant and non

- tolerant cotton. Based on available toxicit y studies and structural similarities, HED considers the parent and all three metabolites to be of comparable toxicity. While DCGA may be of comparable toxicity, it was present in the cotton metabolism studies at less than

10% of the total radioactive res idue (TRR), and is only detected in livestock feed items, not in human

food items. Furthe r , inclusion of this metabolite as a ROC in feed items would have no material impact on the livestock dietary burden since calculation of the reasonably balanced live stock diets are driven by other feed items with far higher residues. Thererfore, HED is not including the DCGA

metabolite as an ROC for risk assessment. The residues of concern that were concluded for tolerance expression and risk assessment in crop comm odities based on these data are presented below in Table

4.

Table 4

. Dicamba Residue s of Concern

. Matrix Tolerance Expression Residues for Risk Assessment
Barley, corn, grasses, oats, proso millet, sorghum, sugarcane, and

wheat Dicamba + 5

- OH dicamba Dicamba + 5

- OH dicamba

Asparagus Dicamba + DCSA 1 

### Dicamba + DCSA

Cotton, Soybeans and aspirated grain fractions (AGFs) Dicamba + 5

- OH dicamba + DCSA Dicamba + 5

- OH dicamba + DCSA

1 

### DCSA also referred to as 3,6

- dichloro

- 2

- hydroxybenzoic acid or as 3,6

- dichlorosalicylic acid.



## 860.1300 Nature of the Residue

- Livestock

Re registration Eligibility Decision Memo, D 317 699

, 12 / 20 / 20 05, C. L. Olinger



## The nature of the residue in livestock is adequately understood based upon acceptable metabolism

stud ies made on ruminants and poultry

. The residues of concern in meat, milk poultry, and eggs are

dicamba and its DCSA metabolite established under 40 CFR §180. 227(a)(2). A n ew livestoc k commodity storage stability study and ruminant feeding study made at an increased dicamba parent feeding level of 1000 ppm was provided in support of reregistration . Both studies were determined to be adequate and no further livestock storage stability or ruminant feeding studies are required (D320564 & D322482, C. L. Olinger, 12/08/2005).

860.1340 Residue Analytical Methods

Plant commodities

Re registration Eligibility Decision Memo, D 317 699

, 12 / 20 / 20 05, C. L. Olinger

DER Reference: 4 8728702 .der.doc x (Residue Analytical Method for Dicamba

- Tolerant Cotton )



## E nforcement method: Adequate methods available for the enforcement of the newly proposed cotton seed and cotton gin byproducts tolerances. The Pesticide Analytical Manual (PAM) Vol. II li sts Method I (AM 02658A) which uses GC/ECD analysis to carry out the determination of dicamba in/on crop commodities. The sensitivity of this method is listed to be 0.05 ppm and it has been validated by the Agency using corn and sorghum commodities. An i mproved plant enforcement method using GC/ECD analysis that includes an acid hydrolysis step for improved extraction efficiency has been developed as a modification to Method I. For this method

, the reported l imit of

### Dicamba Summary of Analytical Chemistry and Residue Data DP No. D408384



## Page 9 of 26 q uantitation (LOQ) is 0.02 ppm

, and pe nding revisions suggested by the Agency

, it will likewise be

included in PAM Vol. II.

Data collection methods: The cotton field trial samples analyzed in support of this petition were

made u sing a validated Liquid Chromatography/Mass Spectrometer/Mass Spectrometer (LC/MS/MS) analytical method. Developed by Monsanto as method AG

- ME

- 13801

- 01, it is entitled “Analytical Method for the Determination of Dicamba and its Major Metabolites in Cotton Matrices by

LC/MS/MS”. For this method

, samples are first ex tracted using a mixture of acetonitrile:water (40:60, v:v). An aliquot of the extract is then hydrolyzed with 1 N HCl at 95 °C in an oven followed by partitioning with a mixture of ethyl acetate:isooctane (20:80), v:v). Water is then added to the organic phase and the sample is concentrated by evaporation until only the aqueous solution remains. The resulting samples are acidified and analyzed by LC/MS/MS determination. A complete description of the method and validation data determining dicamba a long wi th its 5

- OH dicamba, DSCA, and DCGA metabolites were included in this submission (MRID No. 48728702 , P. Savoia, 03/25/2013)

. The method determined residues of parent, 5

- OH dicamba, and DCSA separately. The method limit of quantitation ( LOQ ) was 0.02 ppm determined for each analyte in cotton undelinted seed, meal, hulls, and alkali re f ined oil . The LOQ was 0.04 ppm for cotton gin byproducts. The

method was validated using samples of the various cotton matrices fortified with each analyte at 0.02, 0.20, a nd 10 ppm except for cotton gin byproducts which was fortified at 0.04, 0.40, and 10 ppm. Mean recoveries were within the acceptable range of 70% to 120% with the coefficient of variation

less than 20% for each matrix tested to demonstrate the method is a dequate for data collection. The tolerance is expressed as residues of dicamba and its metabolites expressed as dicamba; therefore, the

residues of 5

- OH dicamba, DCSA, and DCGA are converted to parent equivalents using molecular weight conversion factor s of 0.933, 1.068, and 0.991, respectively.

Livestock commodities

The analysis of animal commodities is n ot relevant to this petition because no new livestock tolerances are being proposed for this action.

860.1360 Multiresidue Methods

Re registration Eligibility Decision Memo, D 317 699

, 12 / 20 / 20 05, C. L. Olinger



## A ppendix II of PAM Volume I specifies that dicamba is completely recovered using Section 402 E2 of Protocol B but is only partially recovered using Section 402 E1 of Protocol B. There are no recovery data for MRM determining any of the dicamba metabolites of concern (5

- OH dicamba and DCSA), and these data have been required

. However, b ecause s ingle analyte methods for enforcement have thus been provided, the additional MRM data a re not needed.

860.1380 Storage Stability

DER Reference: 4 8728704 .der.doc x ( S t ability of Dicamba Residues in Dicamba

- Tolerant Cotton )



## In support of this petition, Monsanto has submitted storage stability data for dicamba along with its

DSCA and DCGA metabolites in/on MON 88701 cotton. Data on the stability of the 5

- OH dicamba

metabolite was not provided, and is not required since residues of that metabolite are not found in

### Dicamba Summary of Analytical Chemistry and Residue Data DP No. D408384



## Page 10 of 26 cotton undelinted seed. The storage durations and conditions o f the dicamba

- tolerant ( MON 88701 ) cotton samples studied are presented below in Table 5

.

Tab le 5

. Summary of Storage Conditions for Cotton Commodities

. Matrix (RAC or Extract) Storage

Temp. ( °C) Analyte Maximum Storage Duration (Days) Interval of Demonstrated Storage Stability

Cotton, undelinted

seed Held below

freezing at the

field sites and

at

- 2 0 °C at the laboratory. Dicamba 13

- 1 9 6 days (0.4

- 6.4 months) Residues of dicamba, DCGA, and DCSA are

stable during freezer storage for up to 277 days (9 months) in undelinted cotton seed (MRID No. 48728704)

.

The stability of 5

- OH dicamba was not determined concurrently because it was not

detected in cotton undelinted seed. 5

- OH Dicamba 13

- 122 days ( 0 .4

-

4.0 months)

DCGA 13

- 285 days (0.4

-

9.4 months)

DCSA 13

- 121 days ( 0.4

-

4.0 months)
Cotton, gin byproducts Held below

freezing at the

field sites and

at

- 2 0 °C at the laboratory. Dicamba 17

- 27 days (0.6

- 0.9 months) Storage stability data are not required since the se

samples were analyzed within 30 days of harvest. 5

- OH Dicamba

DCGA

DCSA

Cotton, hulls Ambient at

field sites,

- 12 °C at processor, and

- 2 0 °C at laboratory. Dicamba 33

- 40 days (1.1

- 1.3 months) S torage stability data are not required as the se

processed fractions were generally analyzed within 30 days of processing. 5

- OH Dicamba 33

- 34 days (1.1

months) DCGA

DCSA

Cotton, meal Ambient at

field sites,

- 12 °C at processor, and

- 2 0 °C at laboratory. Dicamba 26

- 27 days ( 0.9 months) 5

- OH Dicamba

DCGA

DCSA

Cotton, alkali

refined oil Ambient at

field sites,

- 12 °C at processor, and

- 2 0 °C at laboratory. Dicamba 24

- 25 days ( 0.8 months) 5

- OH Dicamba

DCGA

DCSA

Cotton, RBD 1 oil Ambient at

field sites,

- 12 °C at processor, and

- 2 0 °C at laboratory. Dicamba 19

- 20 days (0.6

- 0.7 months) 5

- OH Dicamba

DCGA

DCSA

1 RBD

– Refined Bleached Deodorized.



## Conclusions

. Adequate storage stability data for the residues of dicamba in/on dicamba

- tolerant

cotton commodities have b een provided to support the studies provided in this submission

.

860.1400 Water, Fish, and Irrigated Crops

This guideline requirement is not relevant to the current action

.

860.1460 Food Handling

This guideline requirement is not relevant to the current action

.

### Dicamba Summary of Analytical Chemistry and Residue Data DP No. D408384



## Page 11 of 26 860.1480 Meat, Milk, Poultry, and Eggs

Re registration Eligibility Decision Memo, D 317 699

, 12 / 20 / 20 05, C. L. Olinger



## For this action, a 70 .0 ppm tolerance will be established for cotton gin byproducts which is a

livestock feedstuff fed to beef cattle. The dicamba maximum theoretical dietary burdens to livestock were previously calculated in the residue chemistry considerations for the 2005 RED

. While HED is

recom mending the establishment of a 70 ppm tolerance for cotton gin byproducts, roughage, a tolerance of 250 ppm on grass hay, another livestock roughage feed item, was already included in the maximum theoretical dietary burden calculation; therefore, the addition of this new livestock tolerance will not materially impact the previously calculated dietary burden, as the calculation would default to the roughage with the highest impact, in theoretical dietary burden, but a maximum reasonably balanced dietary burden (MRBD) which considers reasonable diets for livestock, rather than assuming livestock consume any of the feed stuff with the highest residue in any quantity. If HED were to recalculate the MRBD burden for dicamba at this time, not only would the cotton gin byproducts be supers eded by the grass hay, so not be materially included in the calculation, but it is also likely that the overall MRBD burden would be substantially reduced from that calculated maximum theoretically dietary burden from 2005. Tolerances in livestock commodi ties are based on that 2005 RED dietary burden, which is likely an overestimate of livestock exposure, and not impacted by this new feedstuff tolerance; therefore, no new animal tolerances are needed to support

this new use on dicamba

- tolerant cotton.

86 0.1500 Crop Field Trials

DER Reference: 4 8728703 .der 1 .doc x ( Magnitude of Dicamba Residue s in Dicamba

- Tolerant Cotton)



## Monsanto has submitted crop field trial data for dicamba on dicamba

- tolerant (MON 88701) cotton.

T hirteen (13) field trials were con ducted during the 2010 growing season in the representative North American Free Trade Agreement (NAFTA) Growing Zones

. These studies were performed in

NAFTA Growing Zones 2 (GA and SC; 2 trials), 4 (AR, LA, and MO; 3 trials), 6 (TX; 1 trial), 8 (OK

and TX ; 5 trials), and 10 (CA; 2 trials). Each trial site consisted of one untreated and three to four

treated test plots.

Each treated test plot received the maximum total seasonal application rate of 2.0

- 2.1 lb ae/A applied following one (1) of four ( 4 ) distinct use patterns evaluated for study. The first use pattern (TRT 1)

was the cultivation of the control plot which received no application of the test substances.

The second use pattern (TRT 2) was performed as three foliar broadcast applications applied at pre

emergence, the 6

- leaf stage, and at 15

- days past the 1 st 

## white flower. The third use pattern (TRT 3) was also made as three foliar broadcast applications applied at pre

- emergence, 1 st 

## open boll, and 7

- days prior to harvest. The forth use p attern (TRTs 4 & 5) w as done using four post

- emergence foliar broadcast applications applied at the 6

- leaf stage, 15

- days past the 1 st 

## white flower, 1 st 

## open boll, and

7

- days prior to harvest. For this use pattern

, TRT 4 was carried out using a 4.0 lb ae / gal SL diglycolamine salt product proposed for registration

. TRT 5 was a bridging study made using a 5.0 lb ae/gal monoethanolamine salt formulation of dicamba for testing its equivalency to the diglycolamine salt product. A summary of the use patterns is provided below in Table 6

.

### Dicamba Summary of Analytical Chemistry and Residue Data DP No. D408384



## Page 12 of 26 T able 6

. Summary of Field Trial Use Patterns for Treating Cotton with Dicamba.

TRT Number

of Sites EP Growth Stages/ Target Application Rates lb ae/A (kg ae/ha)

Pre

- emergence 6 Leaf 1 st White Flower + 15 days First Open

Boll 7 Days Prior

to Harvest

1 13

-- -- -- -- -- -- 2 13 4 lb ae/gal SL 1.0 (1.1) 0.50 (0.56) 0.50 (0.56)

-- -- 3 13 4 lb ae/gal SL 1.0 (1.1)

-- -- 0.50 (0.56) 0.50 (0.56)

4 13 4 lb ae/gal SL

-- 0.50 (0.56) 0.50 (0.56) 0.50 (0.56) 0.50 (0.56)

5 4 5 lb ae/gal SL

-- 0.50 (0.56) 0.50 (0.56) 0.50 (0.56) 0.50 (0.56)



## All treatments were made using ground equipment (backpack, hand

- held or tractor

- mounted sprayers)

with a non

- ionic surfactant and a mmonium sulfate added to the spray mixture at spray volumes of

18.8

- 21.6 gal/A

. The use patterns evaluated for study followed the maximum labeled rate with pre

emergent application s all being made at 1.0 lb ae/A . The post

- emergence treatments likewise

followed the label using a maximum single in crop application rate of 0.5 lb ae/A made at a 7

- day

RTI and a 7

- day PHI. The c ombined total of all applications appropriately followed the maximum

labeled rate n ot exceed ing 2.0 lb ae/A or 64 fluid ounces per acre in a given ye ar. Additional s amples

were also collected from two trial site s for the TRT 2 and TRT 4 study use patterns to assess residue

decline.

To determine the magnitude of the residue, the cotton RACs were analyzed by LC/MS/MS analysis according to Monsanto Method AG

- ME

- 1381

- 01. The reported LOQ for this LC/MS/MS method is 0.02 ppm for each analyte in cotton undelinted seed and 0.04 ppm in gin byproducts. The method was successfully validated prior to and in conjunct ion with the analysis of the field trial samples. It is important to note that the analytical method does not specify protocol for conversion of metabolite residues to parent equivalents . The quantifiable residues of 5

- OH dicamba, DCSA, and DCGA were

con verted to parent equivalents using molecular weight conversion factor s of 0.933, 1.068, and 0.991, respectively.

The average residues for dicamba and the 5

- OH dicamba and DCSA metabolites of concern for cotton undelineted seed ranged from < 0. 06

- 1.72 ppm. Cotton gin byproducts were also harvested as stripper cotton from 3 field trials following the TRT 2 use pattern, as well as from 3 additional trials from the TRT 4 use pattern. The number of samples provided for this RAC is adequate because the

data fro m s tripper only cotton for gin byproducts generally contains much more trash than picker

cotton and is wors t case. For the cotton gin byproducts samples that were analyzed, average residues for dicamba, and the 5

- OH dicamba and DCSA metabolites of concern ranged from 0.53

- 29.6 ppm. These data examine the broad pattern of use offered by the label which shows that later post

emergence treatments give much higher residues then those made at earlier growth stages. No residues of dicamba were appropriately ob served above the reported LOQ of 0.02 ppm for cotton undelinted seed or 0.04 ppm in gin byproducts for any of the untreated control samples. The results of the residue decline studies performed for the TRT 2 use pattern yielded residues in cotton

undelint ed seed at or below the 0.02 ppm LOQ. Therefore, residue decline could not be assessed for

these studies. For the residue decline studies following the TRT 4 use pattern, residues of dicamba

were found to decrease in undelinted cotton seed with an increa sing PHI. A summary of the residue data acquired in these studies are presented in Table 7

.

### Dicamba Summary of Analytical Chemistry and Residue Data DP No. D408384



## Page 13 of 26 T able 7

. Summary of Residue Data from Cotton Crop Field Trials with Dicamba.

Commodity Analyte Total App.

Rate

lb ae/A (kg ae/ha) PHI

(days) Residue Levels (ppm) 1

n Sample

Min. Sample

Max. L AFT 2 HAFT 2 Median Mean Std. Dev.

TRT 2 (Applications at Preemergence, 6

- leaf stage, and first white flower + 15 days; EP: Clarity)

Undelinted

Cotton seed Dicamba 2.0 (2.2) 49

- 105 13 < 0.02 < 0.02 < 0.02 < 0.02 0.02 0.02 N/A

5

- OH Dicamba 13 < 0.02 < 0.02 < 0.02 < 0.02 0.02 0.02 N/A

DCSA 13 < 0.02 0.23 < 0.02 0.23 0.02 0.04 0.06

Combined Residues 13 < 0.06 < 0.28 < 0.06 < 0.28 0.06 0.09 0.06

Gin byproducts Dicamba 2.0 (2.2) 82

- 84 3 < 0.04 < 0.04 < 0.04 < 0.04 0.04 0.04 N/A

5

- OH Dicamba 3 < 0.04 < 0.04 < 0.04 < 0.04 0.04 0.04 N/A

DCSA 3 0.39 1.73 0.43 1.58 0.67 0.89 0.61

Combined Residues 3 < 0.47 < 1.82 < 0.53 < 1.66 0.75 0.97 0.61

TRT 3 (Applications at Preemergence, first open boll stage, and 7 days prior to harvest ; EP: Clarity)

Undelinted

Cotton seed Dicamba 2.0 (2.2) 6

- 8 13 0.06 1.97 0.06 1.38 0.65 0.64 0.43

5

- OH Dicamba 13 < 0.02 0.02 < 0.02 < 0.02 0.02 0.02 N/A

DCSA 13 < 0.02 0.25 < 0.02 0.16 0.03 0.05 0.05

Combined Residues 13 < 0.12 < 2.24 < 0.1 0 < 1. 56 0. 71 0. 71 0.4 8

TRT 4 (Applications at 6

- leaf

, first white flower + 15 days, first open boll , and 7 days prior to harvest ; EP: Clarity)

Undelinted

Cotton seed Dicamba 2.0

- 2.1 (2.2

- 2.4) 6

- 8 13 0.09 1.54 0.12 1.42 0.47 0.61 0.41

5

- OH Dicamba 13 < 0.02 < 0.02 < 0.02 < 0.02 < 0.02 < 0.02 N/A

DCSA 13 0.02 0.27 0.02 0.27 0.06 0.08 0.07

Combined Residues 13 < 0.1 3 < 1.83 < 0. 16 < 1. 72 0. 56 0. 71 0. 48

TRT 4 (Applications at 6

- leaf

, first white flower + 15 days, first open boll , and 7 days prior to harvest ; EP: Clarity)

Gin byproducts Dicamba 2.0 (2.2) 6

- 7 3 3.09 23.6 3.13 23.0 14.9 13.7 10.0

5

- OH Dicamba 3 < 0.04 0.04 < 0.04 < 0.04 0.04 0.04 N/A

DCSA 3 1.70 6.29 1.78 6.17 4.50 4.15 2.22

Combined Residues 3 < 4.83 29.9 < 5.06 < 29.6 19.7 18.1 12.2

TRT 5 (Applications at 6

- leaf

, first white flower + 15 days, first open boll , and 7 days prior to harvest ; EP: MON 11968 )

Undelinted

Cotton seed Dicamba 2.0 (2.2) 7

- 8 4 0.17 0.72 0.20 0.62 0.41 0.41 0.23

5

- OH Dicamba 4 < 0.02 < 0.02 < 0.02 < 0.02 0.02 0.02 N/A

DCSA 4 0.02 0.17 0.02 0.12 0.04 0.06 0.04

Combined Residues 4 < 0.2 1 < 0.91 < 0.2 4 < 0. 76 0. 47 0. 49 0.2 7

1 Except for sample min/max, values reflect per trial averages; n = no. of field trials. For calculation of median, mean, and standar d deviation, the LOQ (0.02 ppm each analyte in undelinted cotton seed and 0.04 ppm for each analyte in cotton gin byproducts) w as used for any results reported as <LOQ in Table C.3. Combined residues of dicamba, 5

- OH dicamba, DCGA, and DCSA are expressed in parent equivalents. Individual analyte results are reported as per se. N/A = Not applicable.

2 LAFT = lowest

- average

- field

- t rial; HAFT = highest

- a verage

- f ield

- t rial.



## Conclusions

. The submitted crop field trial results provided by Monsanto for MON 88701 cotton treated with dicamba are adequate to fulfill data requirements. The number and location of these field trials are sufficient in meeting the O CSPP 860.1500 Residue Chemistry Test Guidelines for determining the magnitude of the residue in/on cotton RACs. T here were no unusual meteorological conditions during the course of the field trials which would ad versely affect the quality and integrity

of the s tudy. An acceptable method was used for quantitation of residues in /on cotton RACs and adequate supporting storage stability data are available.

### Dicamba Summary of Analytical Chemistry and Residue Data DP No. D408384



## Page 14 of 26 860.1520 Processed Food and Feed

DER Reference: 4 8728703 .der 2 .doc x ( Magnitude of Dicamba Residue s in Dicamba

- Tolerant Cotton)



## Monsanto has submitted processing study data for dicamba

- tolerant MON 88701 cotton treated with

dicamba. Samples used for processing were generated from the two (2) trials held in Missouri and Texas during the 2010 growing season discussed above

. Each trial site maintained two treated test plots which received the maximum total seasonal application rate of 2 .0

- 2.1 lb ae/A applied following one (1) of two (2) distinct use patterns evaluated for study.

All applications were made at the maximum labeled rate with the 4 lb ae/gal SL diglycolamine salt

formulation of dicamba . Treatments were made using ground e quipment at spray volumes of 19.7

- 20.5 gal/A (184

- 192 l/ha) and included a nonionic surfactant ( NI S) and ammonium sulfate (AMS) adjuvants. Samples of cotton undelinted seed were harvested using a mechanical picker at maturity

which followed a PHI of 71

- 79 days for TRT 2 and 6

- 7 days for TRT 4. The resulting cotton

undelinted seed samples were then processed into meal, hulls

, alkali

- refined oi l, and RBD ( Refined

Bleached Deodorized ) oil by GLP Technologies located in Navasota Texas using procedures simulating commercial processing.

To analyze for residues of dicamba, the cotton RAC and its processed commodities were determined using the Monsanto LC / MS/MS method AG

- ME

- 1381

- 01. The reported LOQ for this LC/MS/MS method is 0.02 ppm for each analyte in cotton undelinted seed as well as for its processed

commodities. The method was successfully validated prior to and in conjunction with the analysis of the field trial samples. As previously noted, this analytical method does not specify protocol for

co nversion of metabolite residues to parent equivalents . The quantifiable residues of 5

- OH dicamba, DCSA, and DCGA were converted to parent equivalents using molecular weight conversion factor s of 0.933, 1.068, and 0.991, respectively.

For the two use patt erns tested in this study , quantifiable residues were only obtained for the four

post

- emergence applications comprising TRT 4. Using these data, the concentration factors for each dicamba residue of concern was calculated for the processed commodities of dicamba

- tolerant cotton

seed. These data indicate that the combined residues of dicamba do not concentrate in hulls, meal, alkali refined oil, or RBD oil by yielding processing factors of ≤ 1.0x for each dicamba analyte

, respectively

. A summary of the dat a acquired from these processing studies is presented below in

Table 8.

### Dicamba Summary of Analytical Chemistry and Residue Data DP No. D408384



## Page 15 of 26 T able 8

. Summary of Residue Data from the Cotton Processing Study made with Dicamba.

RAC: MRID No. Processed Commodity Total Rate

lb ae /A (Kg ae/A) TRT 1 Residues 2 (ppm) Avg. Processing Factor 3

Parent 5

- OH DCSA Parent 5

- OH DCSA

Cotton/ MRID

No. 48728703 Undelinted Seed (RAC) 2.0 (2.2) 4 0.65 <0.02 0.04

-- -- --
Hulls 0.43 <0.02 0.04 0.66x NC 1.0x

Meal <0.02 <0.02 <0.02 <0.03x NC <0.5x

Alkali Refined Oil <0.02 <0.02 <0.02 <0.03x NC <0.5x

RBD Oil <0.02 <0.02 <0.02 <0.03x NC <0.5x

1 TRT

– notes the use pattern that was followed for study.

2 Residues of dicamba include parent along with the 5

- OH dicamba and DCSA metabolites of concern expressed in parent equivalents obtained for the TRT 4 use pattern trials. The LOQ is 0.02 ppm for each analyte in every given matrix. Concentrations found at or
below the LOQ are denoted as < 0.02 ppm and included in the totals as 0.02 ppm.

3 NC

– “not calculated” since there are not quantifiable residues in the RAC.



## Conclusions: The cotton processing study is adequate to satisfy O CSPP 860.15 2 0 data requirements.

R esidues of dicamba do not concentrate in the processed commodities of dicamba

- tolerant ( MON

88701 ) cotton . Therefore, separate tolerance s are not required for the processed commodities of

dicamba

- tolerant ( MON 88701 ) cotton.

860.1650 Submittal of Analytical Reference Standards

An alytical standards for dicamba and its metabolites of concern are currently available in the EPA National Pesticide Standards Repository ( personal communication with Theresa Cole

, BEAD

, 0 1 / 30 /201 3 )

. The current stock of standards is set to expire on 11 / 30 /201 5

.

860.1850 / 860.1900 Confined and Field Accumulation in Rotational Crops

Re registration Eligibility Decision Memo, D 317 699

, 12 / 20 / 20 05, C. L. Olinger



## The December 2005 Reregistration Eligibility Decision (RED) for dicamba requires additional rotational crop studies in order to satisfy data requirements. However, these data are not needed if a

120

- day plantback interval is specified when dicamba is appl ied at the maximum seasonal rate of 0.75 lb ae/A. For greater seasonal application rates of 0.75

- 2.0 lb ae/A, only crops with established tolerances can be rotated for planting. Because no new rotational crop data have been submitted, the plantback restr ictions noted in the 2005 RED are required, and have been appropriately reflected on the pro posed pro duct label for dicamba

- tolerant (MON 88701) cotton.

860.1550 Proposed Tolerances

Permanent t olerances are e stablished under 40 CFR §180. 227(a)( 1) for dicamba and its 3,6

- dichloro

5

- hydroxybenzoic acid (5

- OH dicamba) metabolite. There are also a dditional tolerances l isted under 40 CFR §180. 227(a)(2) for dicamba and its 3,6

- dichloro

- 2

- hydroxybezoic acid metabolite, and under 40 CFR §180. 227(a)(3) for dicamba, and the 5

- OH dicamba and DSCA metabolite s

. The tolerance expression recommended for cotton commodities based on the new metabolism data provided in support of this action is parent

, 5

- OH dicamba

, and DSCA established under 40 CFR §180. 227(a)( 3). However, the 40 CFR §180. 227(a)(1) citation for dicamba currently lists a 0.2 ppm tolerance for cotton, undelinted seed. This registration action supersedes the 40 CFR §180. 227(a)(1) cotton

undelinted seed tolerance which requires it to be removed from the federal register. Therefore all

### Dicamba Summary of Analytical Chemistry and Residue Data DP No. D408384



## Page 16 of 26 tolerances for cotton RACs are now to be established under 40 CFR §180. 227(a)(3) as recommended

above.

The International Residue Limit (IRL) summary is shown in Attachment 1 of this memorandum. There are MRLs of 0.2 ppm in Mexico and 0.04 ppm established by Codex on cotton seed currently established. Mexico adopts existing U.S. crop tolerances for export purposes which in this instance is

the current cotton unde linted seed tolerance of 0.2 ppm. There are currently no Mexican, Canadian or Codex MRLs established for cotton gin byproducts. Because the registrant is now requesting a late

season use of dicamba on dicamba

- tolerant cotton, the currently established in ternational tolerances are not adequate to cover residues likely from the newly proposed use in the U.S. In addition, the

dicamba residues of concern for dicamba

- tolerant cotton also include the DCSA metabolite which is not found nor regulated in the othe r common varieties of cotton. Therefore, harmonization with respect to the tolerance expression is not possible at this time for cotton seed. Since there are no international tolerances on cotton gin byproducts, there is no issue of international harmoni zation

relevant to that tolerance.

T he cotton field trial residue data reflecting late season treatments at the labeled 7

- day PHI minimum (TRT 3 & TRT 4) w ere evaluated by the OECD calculation procedures to determine an appropriate tolerance level (see Appendix I I I ) to yield a recommended tolerance of 3 .0 ppm for cotton undelinted

seed . Using the OECD calculation procedures, the cotton gin by products data reflecting late season

treatments at the labeled 7

- day PHI minimum (TRT 4) w ere also evaluated an d a tolerance of 70 .0 ppm is recommended (see Appendix I I I ). As previously noted , no concentration in cotton seed processed commodities was seen; therefore, separate tolerances are not required for cotton processed

commodities. The final tolerance recom mendations are shown below in Table 9

.

Table 9

. Tolerance Summary for Dicamba

. Commodity Established

Tolerance (ppm) Proposed

Tolerance (ppm) Recommended

Tolerance (ppm) Comments; Correct Commodity Definition

Tolerances Proposed for 40 CFR §180. 227 (a) (3)

Cotton, gin byproducts

-- 70.0 70.0

Cotton, undelinted seed 0.2 3.0 3.0

### Dicamba Summary of Analytical Chemistry and Residue Data DP No. D408384



## Page 17 of 26 References

DP No.: D 317699 Subject: Dicamba

. Residue Chemistry Considerations for the Reregistration Eligibility

Decision (RED) Document. Summary of Analytical Chemistry and Residue Data.

From: C. L. Olinger

To: K. Tyler

Date: 12 / 20 / 200 5

MRIDs: None

DP No.: None Subject: Dicamba Registration St andard

From: C. L. Trichilo

To: A. Rispin and R. Taylor

Date: 08 /1 2 / 1983

MRIDs: None

DP No.: D320564 and D322842 Subject: Re re gistration of Dicamba. Livestock Storage Stability Study and Ruminant Feeding

Study

From: C. L. Olinger

To: K. Tyler

Date: 12 / 08 / 2005

MRIDs: 44891303 and 46668101

Attachments:

International Residue Limit Status S heet Appendix I

- Characterization and Identification of Radioactive Residues in Dicamba

- Tolerant Cotton Appendix I I

- Chemical Names and St ructures of Dicamba and its Metabolites Appendix I I I

- Tolerance Assessment Calculations

### Dicamba Summary of Analytical Chemistry and Residue Data DP No. D408384



## Page 18 of 26 

# International Residue Limits

Dicamba (029801, 029802, 029806, 128931, 128944 & 129043; 01/16/2013)

Summary of US and International Tolerances and Maximum Residue Limits Residue Definition 1 

### :
US Canada Mexico 2 

### Codex 3



### 40 CFR 180.227: Plant: Compliance with the tolerance levels is to be determined by measuring only the residues of dicamba, 3,6

- dichloro

- 0

- anisic acid, and its metabolites, 3,6

- dichloro

- 5

- hydroxy

- 0

- anisic acid, and 3,6

- dichloro

- 2

- hydroxybenzoic acid, calculated as the stoichiometric equivalent of dicamba benzoic acid, 3,6

- dichloro

- 2

- methoxy

- , including the

metabolite benzoic acid, 2,5

- dichloro

3

- hydroxy

- 6

- methoxy

Plants: Dicamba

Animals:

sum of dicamba and 3,6

- dichlorosalicylic acid (DCSA) expressed as

dicamba.

The residue is not

fat

- soluble.



## Commodity 4 

## Tolerance (ppm) /Maximum Residue Limit (mg/kg)

US Canada Mexico 2 

## Codex 3



## Cotton, undelinted seed 3 0.04 cotton seed (*) Cotton, gin byproducts 70

Completed: M. Negussie; 01/17/2013

1 

### The U.S. Residue definition for cotton commodities now includes the DCSA metabolite not found nor regulated in the

common varieties of cotton.

2 

### Mexico adopts US tolerances and/or Codex MRLs for its export purposes.

3 

### * = absent at the limit of quantitation; Po = postharvest treatment, such as treatment of stored grains. PoP = processed postharvest treated commodity, such as processing of treated stored wheat. (fat) = to be measured on the fat portion of the sample. MR Ls indicated as proposed have not been finalized by the CCPR and the CAC.

4 

### Includes only commodities of interest for this action. Tolerance values should be the HED recommendations and not those proposed by the applicant.

### Dicamba Summary of Analytical Chemistry and Residue Data DP No. D408384



## Page 19 of 26 Appendix I

– Ch aracterization a nd Identification of Radioactive Residues in Dicamba

- Tolerant

Cotton

Appendix I. Summary of Characterization and Identification of Radioactive Residues in Dicamba

Tolerant Cotton Matrices Following Application of [Phenyl

- U

14 

### C]dicamba at 2.094 lb ae/A (pr eemergence) or 2.065 lb ae/A (postemergence)

Preemergence Postemergence

Seed Gin byproducts Seed Gin byproducts

TRR = 0.1621 ppm TRR = 0.8493 ppm TRR = 0.9778 ppm TRR = 60.024 ppm

% TRR ppm %TRR ppm %TRR ppm %TRR ppm

Dicamba 0.09 0.0001 0.50 0.004 0.85 0.008 4.48 2.691

DCSA glucoside 0.73 0.0012 27.77 0.236 3.42 0.033 16.83 10.101

DCSA 0.06 0.0001 5.42 0.046 1.91 0.019 13.39 8.035

DCGA glucoside 0.09 0.0001 0.68 0.006 1.1 0.011 2.46 1.476

MCTHBA glucoside

-- -- -- -- -- -- 3.33 1.998

MCDHBA glucoside

sulfate -- -- -- -- -- -- 4.71 2.828

MCDHBA glucoside

-- -- -- -- -- -- 0.84 0.505

MCTHBA cyc. glucoside -- -- 2.94 0.025

-- -- 2.76 1.656

Dicamba amide

-- -- 3.22 0.027

-- -- -- -- Fatty acids 16.05 0.026

-- -- 10.07 0.098

-- -- Glycerol 2.86 0.005

-- -- 1.35 0.013

-- -- Sugars/natural products 4.59 0.0074 8.94 0.076 5.61 0.055 2.69 1.613

Phosphate rinse 3.87 0.006 3.25 0.028 1.96 0.019 6.20 3.72

Starch 2.79 0.005 2.73 0.023 2.19 0.021 7.05 4.234

Protein 17.71 0.029 1.71 0.015 10.10 0.099 2.25 1.350

Pectin 7.40 0.012 1.91 0.016 4.04 0.039 2.66 1.595

Lignin 6.18 0.010 5.28 0.045 14.00 0.136 6.60 3.962

Cellulose 4.37 0.007 0.90 0.008 3.13 0.031 0.62 0.374

Hemicellulose 14.43 0.023 5.35 0.045 13.49 0.132 2.80 1.679

Acid Hydrolysates 5.07 0.008

-- -- 3.21 0.031

-- -- Unidentified (oil) 1.94 0.003

-- -- 0.84 0.008

-- --
Discrete unknowns 3.03 0.0049 15.96 0.136 5.36 0.052 0.86 0.518

Total identified 0.97 0.0015 40.53 0.344 7.28 0.071 48.80 29.29

Total characterized 90.29 0.1463 46.03 0.392 75.35 0.734 31.73 19.045

Total extractable 92.65 0.150 97.38 0.827 90.34 0.883 99.39 59.659

Unextractable (PES) 1 7.35 0.012 2.62 0.022 9.66 0.094 0.61 0.365

Accountability 2 100 100 100 100

### Dicamba Summary of Analytical Chemistry and Residue Data DP No. D408384



## Page 20 of 26 Appendix I I

- Chemical Names and Structures of Dicamba and its Metabolites

Appendix I I . Summary of Chemical Names and Structures of Dicamba and its Metabolites.

Common name/code Chemical name Chemical structure

Dicamba 3,6

- dichloro

- 2

- methoxybenzoic acid

DCSA Glucoside 3,6

- dichloro

- 2

- (β

- D

glucopyranosyloxy)benzoic acid

DCSA 3,6

- dichloro

- 2

- hydroxy benzoic acid

DCGA Glucoside 2,5

- dichloro

- 3

- (β

- D

- glucopyranosyloxy)

- 6

- hydroxybenzoic acid

MCTHBA Glucoside 2

- Chloro

- 5,6

- dihydroxy

- 3

- (3,4,5

- trihydroxy

- 6

- hydroxymethyl

tetrahydropyran

- 2

- yloxy)benzoic acid

MCDHBA Glucoside Sulfate 6

- Chloro

- 3

- hydroxy

- 2

- (3,4,5

- trihydroxy

6

- sulfooxymethyltetrahydro

- pyran

- 2

- yloxy)benzoic acid

MCDHBA Glucoside 6

- Chloro

- 3

- hydroxy

- 2

- (3,4,5

- trihydroxy

6

- hydroxymethyltetrahydro

- pyran

- 2

- yloxy)benzoic acid

### Dicamba Summary of Analytical Chemistry and Residue Data DP No. D408384



## Page 21 of 26 Appendix I I . Summary of Chemical Names and Structures of Dicamba and its Metabolites.

Common name/code Chemical name Chemical structure

MCTHBA Cyclic Glucoside 7

- Chloro

- 3,4,6

- trihydroxy

- 2

- hydroxymethyl

- 3,4,4a,9a

- tetrahydro

- 2 H

1,9.10

- trioxa

- anthracene

5

- carboxylic acid

Dicamba Amide 3,6

- dichloro

- 2

- methoxybenzamide

### Dicamba Summary of Analytical Chemistry and Residue Data DP No. D408384



## Page 22 of 26 Appendix I I I . Tolerance Assessment Calculations.

Cotton, undelinted seed

The dataset used to establish a tolerance for dicamba on cotton undelinted seed consisted of field t rial

results held in the representative NAFTA Growing Zones during the 2010 growing season. It was acquired to evaluate the registration of dicamba for post

- emergence use on dicamba

- tolerant cotton
since th is variety can tolerate application of the herbicide in this manner. The data reflect th e maximum seasonal application rate of 2.0 lb ae/A or 64 fluid ounces per acre in a given year following a 7

- day RTI and a 7

- day PHI. Treatments were made with a 4.0 lb ae/gal SL diglycolamine salt product which included a non

- ionic surfactant and a mmoniu m sulfate added to the spray mixture. Th ese field trial data quantified the expected range of residues in cotton undelinted

seed following treatment according to the proposed use pattern

. All cotton undelinted seed samples

had combined residues of dicamb a above the LOQ of 0.0 2 ppm

.

To establish an appropriate tolerance level on this cotton RAC

, only the data acquired using late

season treatments at the labeled 7

- day PHI minimum (TRT 3 & TRT 4) were entered into the Organization for Economic Cooperation and Development (OECD) calculation procedures f or determination. For th o se field trials where sampling replicates were taken, the average or mean value

was used for calculat ion

. T he mean residue values used to calculate the resulting tolerance level i s provided in the table below

. When using the OECD calculation procedures, the resulting MRL value is always found to be greater than or equal to the highest residue within the dataset. MRLs are

di splayed without decimal zeros after the last significant figure and are rounded in the last step of calculation to produce a result in the region of the 95 th 

## percentile. The MRL estimate calculated at

the “ CF x 3 Mean ” level gives a value of 3 .0 ppm for t he combined residues of dicamba

.

### Dicamba Summary of Analytical Chemistry and Residue Data DP No. D408384



## Page 23 of 26 Dicamba

Cotton, undilented seed

U.S.

PHI= 6

- 8 days

Total number of data (n) 26

Percentage of censored data 0%

Number of non

- censored data 26

Lowest residue 0.110

Highest residue 1.720

Median residue 0.700

Mean 0.726

Standard deviation (SD) 0.458
Correction factor for censoring (CF) 1.000

Proposed MRL estimate

- Highest residue 1.720

- Mean + 4 SD 2.560

- CF x 3 Mean 2.178

Unrounded MRL 2.560

Rounded MRL 3

Residues (mg/kg) n

0.11 1

0.16 1

0.17 1

0.19 1

0.23 1

0.26 1

0.36 1

0.38 1

0.43 1

0.46 1

0.52 2

0.68 1

0.72 1

0.75 1

0.8 1

0.92 1

### Dicamba Summary of Analytical Chemistry and Residue Data DP No. D408384



## Page 24 of 26 1 1

1.05 1

1.07 1

1.09 1

1.11 1

1.26 1

1.35 1

1.57 1

1.72 1

### Dicamba Summary of Analytical Chemistry and Residue Data DP No. D408384



## Page 25 of 26 Cotton, gin byproducts

The dataset used to establish a tolerance for dicamba on cotton gin byproducts consisted of field trial results held in the representative NAFTA Growing Zones during the 2010 growing season. It was acquired to evaluate the registration of dicamba for post

- emergence use on dicamba

- tolerant cott on since th is variety can tolerate application of the herbicide in this manner. The data reflect the maximum seasonal application rate of 2.0 lb ae/A or 64 fluid ounces per acre in a given year following a 7

- day RTI and a 7

- day PHI. Treatments were made with a 4.0 lb ae/gal SL diglycolamine salt product which included a non

- ionic surfactant and a mmonium sulfate added to the spray mixture. These field trial data quantified the expected range of residues in cotton gin byproducts following treatment according to the proposed use pattern. All cotton gin byproducts samples had combined residues of dicamba above the LOQ of 0.0 4 ppm

.

To establish an appropriate tolerance level on this cotton RAC, only the data acquired u sing late

season treatments at the labeled 7

- day PHI minimum (TRT 4) were entered into the Organization for Economic Cooperation and Development (OECD) calculation procedures for determination. For those field trials where sampling replicates were taken, the average or mean value was used for

calculation. The mean residue values used to calculate the resulting tolerance level is provided in the

table below

. When using the OECD calculation procedures, the resulting MRL value is always found to be greater than or equal to the highest residue within the dataset. MRLs are displayed without decimal zeros after the last significant figure and are rounded in the last step of calculation to produce a result in the region of the 95 th 

## percentile. The MRL estimate calculated at the “CF x 3 Mean” level gives a value of 70 .0 ppm for the combined residues of dicamba

.

### Dicamba Summary of Analytical Chemistry and Residue Data DP No. D408384



## Page 26 of 26 Dicamba

Cotton, gin byproducts

U.S.

PHI= 6

- 8 days

Total number of data (n) 3

Percentage of censored data 0%

Number of non

- censored data 3

Lowest residue 5.060

Highest residue 29.600

Median residue 19.700

Mean 18.120

Standard deviation (SD) 12.346
Correction factor for censoring (CF) 1.000

Proposed MRL estimate

- Highest residue 29.600

- Mean + 4 SD 67.504

- CF x 3 Mean 54.360

Unrounded MRL 67.504

Rounded MRL 70

High uncertainty of MRL estimate.

[Small dataset]

Residues (mg/kg) n

5.06 1

19.7 1

29.6 1

### Dicamba/ MON 11900 / PC Code 128931 / Mon sa nto Company

DACO 6.3 / 0PPTS 860.1300 / 0ECD 11 6.2.2, 6.2.3 & 

### II I A 8.2

, 8.4.1

, 8.4.2

Nat ur e of the Re s idu es in Plant s

- Dicamba-Tolerant Cotton (Event MON 8870 I)



### Prim ary Eva luator Date: 1 1/ 15 / 2012

Versar, Inc.

- 

### ' 

### Approved by 

# rJ-1-rK**.La__,,~** Date : 04 / 22 / 2013

Peter Savo i a
, Chemist

, OCSPP-HED-RAB 

### YNII

Note: This Data Evaluaiion Record ( DER) was originall y prepared under contract by Vcrsar. Inc. (6 850 Vasar Center. Springfield. VA 2215 1: s ubmitted I I / 15 /20 12). The D E R h as been re v iewe d b) H E D and re v i sed t o rencct current Office of

Pe s ticide Pr og r ams ( OPP ) policie s.



### STUDY R EP ORT :

48728701 Whitehead, T.; Mierkowsk i

, M.: Chott, R. (2011) Amended Report for MSL0021858:

Metaboli s m of (C arbon 14)-Dicarnba in Dicamba-Tolerant Corton. Project Number:

MSL0023760 / 0CR

, REG / 09 / 046, 1901 W. U npublished study prepared by Monsanto Company.

378 p.



### EXECUTIVE SUMMARY:

Monsanto has subm itt ed a s tudy investigating the metaboli sm of uniformly ring-labeled (phen yl

U-14C]dicamba (specific activity of9.85-9.9 1 mCi / mmol ) in dicamba

- and glufosinate-tolerant

cotton, identified as Event MON 88701 (fo rm erly GH_S26695). Event MON 88701 cotton

expresses a modifi ed dicamba mono-o xygenase (OM O) gene derived from 

### Stenotrophomonas maltophilia and the bialaphos resistance (BAR) gene i solated from 

### Streptomyces hygroscopicus.

The expression of DMO confers dicamba tolerance, and the expression of BAR confers glufosinate tolerance. The radiolabeled te s t s ubstance was formulated as an aqueous solution of

the diglycolamine ( D GA) sa lt and applied to cotton grown in outdoor box plots. One plot

received a s in gle pre-emergence soil application (PRE

- T) at 2.094 lb ae / A on the day of planting

after seed was planted, and t h e other plot received a s i ngle post-emergence folia r appl ication
(POE-T) at 2.065 lb ae / A at the first white flower stage (76 days after planting). Samples of seed

cotton and leaves / stems (su r TOgate gin byproducts) were collected at maturity: 180 days afte r

treatment ( DAT ) for the PRE-T plot and I 04 DAT for the POE-T plot. The in-lif e phase of the study was conducted by PTRL West

, Inc. ( J l ercules, CA) and Excel Research Services (Fresno

, CA), and the analyt i cal pha s e of the study was cond uct ed by Monsanto (St. Louis

, MO).

Total radioactive residues (TRR) were determined by combustion / LSC. Following a s in g l e preemergence so il application of [ 14 

### C]dicamba at 2.094 lb ae / A. TRR were 0.1621 ppm in seed and

0.8493 ppm in gin byproduct s harvested at a 180-da y pre-harvest interval ( PHI ). Following a single po s t-emergence foliar application at 2.065 lb ae / A

, TRR were 0

. 9778 ppm in seed and

60

. 0235 ppm in gin byproducts harvested at a I 04-day PHI.

Extraction with hexane released 18.91 % and 11.47% TRR from PRE-T and POE-T seed, respectively ; extraction with acetonitrile (ACN) and ACN / water together relea sed an additional

11.91 % and 26.78% TRR

, respectively. I n gin byproducts

, ext r ac ti on with ACN / water relea sed

the majority of the radioactivity: 76.25% and 71.21% TRR from PRE-T and PO E -T gin byproducts

, respectively. R ema ining non-extractable residues for all matrices were subjected to

0408384 / MRID No. 48728701 Page I of24

### Dicamba/ MON 11900/ PC Code 128931 / Monsanto Company

DACO 6.3/OPPTS 860.1300/OECD II 6.2.2, 6.2.3 & IIIA 8.2, 8.4.1, 8.4.2

Nature of the Residues in Plants

– Dicamba

- Tolerant Cotton (Event MON 88701)

D40 838 4/MRID No. 48728701 Page 2 of 24 

## sequential enzymatic and chemical hydrolysis and oxidation procedures , including a phosphate rinse followed by h ydrolysis with α

- a mylase and protease ; EDTA extraction ; o xidation with

chlorite ; hydrolysis with cellul a se
, 24% aqueous KOH

, and sulfuric acid ( seed only); a second

h ydrolysis with protease (POE

- T seed only ) ; and hot DMSO extraction (POE

- T seed only). These procedures released 52.12

- 61.82% TRR in seed and 21.13

- 28.18% TRR in gin byproducts

. Non

- extractable res idues remaining following hydrolysis procedures were 7.35% TRR (0.012 ppm) in PRE

- T seed, 9.66% TRR (0.094 ppm) in POE

- T seed, 2. 62% TRR (0.022 ppm) in PR E

- T gin byproducts , and 0.61% TRR (0.365 ppm) in POE

- T gin byproducts

. These procedures adequat e l y ex tracted the majority of residues (90.3

- 99.4%) from all cotton matrices. Extraction results were normalized; therefore, accountabilities were ~100%.

Up to 38 peaks were observed in the initial metabolic profiles of seed and gin byproducts. All metabolite peaks accounting for ≥2% TRR were isolated and purified by preparative high performance liquid chromatography ( HPLC ) for further investigation. After isolation, residues

were identifi ed by HPLC/UV and by mass spectral analysis including LC/MS and LC/MS/MS; Gas chromatography/electron ionization mass spectrometry ( GC/EI/MS ) was u sed for volatile

derivatives of the metabolit es or their hydrolysis products

. Isolated metabolites were subjected to partition ing with ethyl acetate, acid, base or enzymatic hydrolysis, and/or chemical derivatizations ( methylation and acetylation ) to provide additional structural information

. Comparison of the chromatographic and mass spectral properties of the metabolite deriva tives or hydrolysates to those of synthetic reference standards was used to confirm the identities of the

metabolites. The study is supported by adequate storage stability data.

P arent dicamba was identified in all matrices at low levels: 0.09

- 0.85% TRR ( 0.0001

- 0.008

ppm ) in seed and 0.5 0

- 4.48% TRR (0.004

- 2.691 ppm) in gin byproducts

. Metabolites DCSA glucoside, DCSA, and DCGA glucoside were also present in all matrices. DCSA g lucoside was the major identified metabolite in gin byproducts, at 16.83

- 27. 77% TRR (0.236

- 10.101 ppm) and

was present in seed at low levels (0.73

- 3.42% TRR)

. Free DCSA was the next most abundant

metabolite in POE

- T gin byproducts (13.39 % TRR, 8.035 ppm) and was present in PRE

- T gin byproducts at 5.42% TRR and in seed at 0.06

- 1.9 1%TRR

. DCGA g lucoside was identified at low levels in all matrices ( 0.09

- 2.46% TRR). The remaining identified metabolites were found in gin byproducts only and accounted for ≤4.71% TRR each, including MCTHBA g lucoside,

MCDHBA g lucoside s ulfate

, and MCDHB A g lucoside in POE

- T gin byproducts , MCTHBA

c yc lic g lucoside in both PRE

- T and POE

- T gin byproducts , and d icamba a mide in PRE

- T gin byproducts only

. Because dicamba amide was only observed in PRE

- T plants, it is believed to be a soil metabolite that was taken up by the plants.

Remaining extractable radioactivity in cotton seed and gin byproducts was characterized as triglycerides and sugars / natural products. In the hexane extracts of seed, triglycerides inclu ding fatty acids ( palmitic, stearic, linoleic, and oleic ) accounted for 10.07

- 16.05% TRR (0.026

- 0.098 ppm), while glycerol accounted for 1.35

- 2.86% TRR. In the ACN/water extracts of all matrices, sugars/natural products (Peak 1) accounted for 2.69

- 8.94% T RR (0.0074

- 1.613 ppm). Th is peak was characterized extensively, indicating that it was a mixture of polar materials, including sugars such as glucose, sucrose, and raffinose, and amino acids such as phenylalanine and glutamic acid. U nidentified component s in oil accounted for 0.84

- 1.94% TRR in seed ; d iscrete

### Dicamba/ MON 11900/ PC Code 128931 / Monsanto Company

DACO 6.3/OPPTS 860.1300/OECD II 6.2.2, 6.2.3 & IIIA 8.2, 8.4.1, 8.4.2

Nature of the Residues in Plants

– Dicamba

- Tolerant Cotton (Event MON 88701)

D40 838 4/MRID No. 48728701 Page 3 of 24 

## u nknown s in the ACN/water extracts together accounted for 0.86

- 15.96% TRR (0.00 49

- 0.518 ppm) in seed and gin byproducts , with none present at > 2.91% TRR

.

Radioactivity in non

- extractable residues was characterized by s equential e nzymatic and

chemical hydrolysis/oxidation . In seed

, these procedures indicated that the majority of radioactivity was incorporat ed into hemicellulose (13. 49

- 14.4 3 % TRR, 0.023

- 0.132 ppm ), protein (10.1 0

- 17.7 1 % TRR, 0.029

- 0.099 ppm ), and lignin (6. 18

- 14.0% TRR, 0.01

- 0.136 ppm ).

Other fractions (inclu ding starch, pectin, and cellulo se) individually accounted for ≤ 7.4% TRR.

In gin byproducts

, radioactivity was incorporated primarily into starch (2.7 3

- 7. 05 % TRR, 0.023

- 4.234 ppm ), lignin (5. 28

- 6.6 0 % TRR, 0.045

- 3.962 ppm ), and hemicellulose (2.8 0

- 5. 35 % TRR,

0.045

- 1.679 ppm )

. Other fractions (including protein, pectin, and cellulose) individually

accounted for ≤ 6.2 % TRR.

Based on the results of the metabolism study on dicamba

- tolerant cotton , the petitioner proposes

that the metabolism of dicamba proceeds by initial demethylation of dicamba to form DCSA through the action of the dicamba O

- demethylase enzyme , which is the product of the DMO

gene introduced to confer dicamba tolerance. The petitioner noted that, due to the predominance

of the O

- demethylation pathway, 5

- OH

- dicamba is not observed as a metabolite in dicamba

tolerant cotton or soybean. While free DCSA was observed in cotton matrices (POE

- T and PRE

T gin byproducts in particular), the majority of DCSA is found as its conjugate DCSA glucoside. Some DCSA glucoside is further modified by hydroxyl replacement of chlorine to form MCDHBA glucoside which is then further conjugated by sulfation to form MCDHBA gl ucoside sulfate. In a minor pathway, DCSA is hydroxylated at the 5

- position, presumably by a P

- 450 enzyme or other oxygenase, to form DCGA. DCGA is not observed as a free metabolite in cotton matrices but is converted to DCGA glucoside in which the gluco se moiety is attached to the hydroxyl group at the 5

- position of DCGA (based upon comparison to the same metabolite

identified in dicamba

- tolerant soybean). DCGA glucoside is further converted to MCTHBA glucoside by hydroxyl replacement of chlorine at the 3

- position. Alternatively, MCTHBA cyclic glucoside may form from DCGA glucoside through displacement of chlorine at the 6

- position of the DCGA portion of the molecule by a hydroxyl group of the glucose moiety. D icamba amide

was a minor metabolite that w as only observed in PRE

- T matrices. Th e petitioner concluded that

this metabolite is most likely the result of the metabolic conversion of the carboxylic acid moiety of dicamba to an amide functional moiety via soil microbial metabolism.

STUDY/WAIVER AC CEPTABILITY/DEFICIENCIES/CLARIFICATIONS :

Under the conditions and parameters used in the study, the plant metabolism data are classified as scientifically acceptable. The acceptability of this study for regulatory purposes is addressed in the forthcoming U.S. EPA Residue Chemistry Summary Document D 40 8384

.

COMPLIANCE :

S igned and dated Good Laboratory Practi ce (GLP), Quality Assurance and Data Confidentiality statements were provided. No deviations from regulatory requirements were reported which would have an impact on the validity of the study

.

### Dicamba/ MON 11900/ PC Code 128931 / Monsanto Company

DACO 6.3/OPPTS 860.1300/OECD II 6.2.2, 6.2.3 & IIIA 8.2, 8.4.1, 8.4.2

Nature of the Residues in Plants

– Dicamba

- Tolerant Cotton (Event MON 88701)

D40 838 4/MRID No. 48728701 Page 4 of 24 

## A. BACKGROUND INFORMATION

Dicamba (3,6

- dichloro

- 2

- methoxybenz oic acid ) is a selective benzoic acid herbicide registered for the control of weeds prior to their emergence. The d icamba Reregistration Eligibility Decision (RED) was issued December 2005. The chemical structure and nomenclature of

dicamba and its metab olites 5

- OH dicamba, DSCA, and DCGA are presented in Table A.1. The physicochemical properties of the technical grade of dicamba acid are presented in Table A.2.

TABLE A.1. Test Compound Nomenclature.

Compound

Common name Dicamba

Company experimental name MON 11900

IUPAC name 3,6

- dichloro

- o

- anisic acid or 3,6

- dichloro

- 2

- methoxybenzoic acid

CAS name 3,6

- dichloro

- 2

- methoxybenzoic acid

CAS registry number 1918

- 00

- 9 (dicamba acid), 104040

- 79

- 1 (diglycolamine salt), or 53404

- 28

- 7 ( monoethanolamine salt)

End

- use product Not applicable.

Compound

Common name 5

- Hydroxy

- dicamba
Company experimental name 5

- OH dicamba

IUPAC/CAS name 2,5

- dichloro

- 3

- hydroxy

- 6

- methoxybenzoic acid

CAS registry number 7600

- 50

- 2

Compound

Common name DCSA ; 3,6

- dichlorosalicylic acid

Company experimental name MON 52708

IUPAC/CAS name 3,6

- dichloro

- 2

- hydroxybenzoic acid

CAS registry number 3401

- 80

- 7

### Dicamba/ MON 11900/ PC Code 128931 / Monsanto Company

DACO 6.3/OPPTS 860.1300/OECD II 6.2.2, 6.2.3 & IIIA 8.2, 8.4.1, 8.4.2

Nature of the Residues in Plants

– Dicamba

- Tolerant Cotton (Event MON 88701)

D40 838 4/MRID No. 48728701 Page 5 of 24 TABLE A.1. Test Compound Nomenclature.

Compound

Common name DCGA ; 3,6

- dichlorogentistic acid

Company experimental name MON 52724

IUPAC/CAS name 2,5

- dichloro

- 3,6

- dihydroxybenzoic acid

CAS registry number 18688

- 01

- 2



### TABLE A.2. Physicochemical Properties of the Technical Grade Test Compound: Dicamba.

Parameter Value Reference

Melting point 114

- 116  C (PAI)

90

- 100  C (87% TGAI) Residue Chemistry Chapter of the Dicamba RED, DP# 317699, 12/20/05, C. Olinger pH 2.5

- 3.0 (87% TGAI)

Density 1.57 g/mL at 25  C (87% TGAI)

Water solubility 0.5 g/100 mL at 25  C (PAI)

Solvent solubility g/100 mL at 25  C (PAI)

dioxane 118.0

ethanol 92.2 isopropyl alcohol 76.0 methylene chloride 26.0

acetone 17.0

toluene 13.0 xylene 7.8 heavy aromatic naphthalene 5.2

Vapor pressure 3.4 x 10

- 5 mm Hg at 25  C (PAI)

Dissociation constant, pK a 1.97 (PAI)

Octanol/water partition coefficient, Log(K OW ) 0.1 (PAI)

UV/visible absorption spectrum neutral: 511 (275 nm) acidic (pH 0

- 1): 1053 (281 nm) basic (pH 13

- 14): 469 (274 nm)



## B. EXPERIMENTAL DESIGN

B.1. Test Site and Crop Information

The in

- life and analytical phases of the study were conducted by PTRL West, Inc. (Hercules, CA) and Excel Research Services (Fresno, CA)

. Cotton was grown from seed planted in loam y sand s oil in outdoor box plots . Separate boxes were used for pre

- and po st

- emergence applications. The plots were located in Madera, CA

.

Crops were maintained under normal agricultural conditions

. Fertilizer was applied during the study period ; however, maintenance pesticides were not applied

. Weather data for the site wa s provided, including minimum/maximum temperatures and daily rainfall. Weather conditions were reported to be normal for the study duration. When rain threatened, the plots were covered

### Dicamba/ MON 11900/ PC Code 128931 / Monsanto Company

DACO 6.3/OPPTS 860.1300/OECD II 6.2.2, 6.2.3 & IIIA 8.2, 8.4.1, 8.4.2

Nature of the Residues in Plants

– Dicamba

- Tolerant Cotton (Event MON 88701)

D40 838 4/MRID No. 48728701 Page 6 of 24 

## so that no rain fell on the plots. Plots were watered by hand as ne eded. Soil characteristics are presented in Table B.1.1, and crop information is presented in Table B.1.2.

TABLE B.1.1. Test Site Information.

Type Method Soil characteristics

Type %OM pH CEC meq/100g

Preemergence soil treatment (PRE

- T) Cotton was grown from seed in a 4 ft. x 4

ft. (16 ft 2 ; PRE

- T) or 3 ft. x 3 ft. (9 ft 2 ; POE

- T) wooden box filled with ~18 in of soil and lined with heavy

- gauge plastic. Loamy sand 0.2 6.8 6.3

Postemergence foliar treatment (POE

- T)



### TABLE B.1.2. Crop Information.

Crop/crop group Variety 1 Growth stage at application Growth stage at

harvest Harvested

Matrix

Cotton; Oilseed group

20 Event MON

88701 PRE

- T: day of planting after planting

seed Maturity Seed; leaves and stems (gin byproducts)

POE

- T: first white flower stage ( 50% of the plants had first white flower)

1 Dicamba

- and glufosinate

- tolerant cotton was used in the study, identified as Event MON 88701 (formerly GH_S26695)

. Event

MON 88701 cotton expresses a modified dicamba mono

- oxygenase (DMO) gene derived from Stenotrophomonas maltophilia and the bialaphos resistance (BAR) gene isolated from Streptomyces hygroscopicus

. The expression of DMO confers dicamba

tolerance

, and the expression of BAR confers glufosinate tolerance to MON 88701



## B.2. Test Materials

The radiolabeled test substance

, [ p henyl

- U

14 

## C]dicamba ( specific activity 45.0 mCi/mmol, 7.53 mBq/mg), was dissolved in ACN and isotopically diluted with unlabeled dicamba

. The characteristics of the test substance are presented in Table B.2.1.

TABLE B.2.1. Test Material Characteristics.

Preemergence Treatment Postemergence Treatment

Chemical structure

Radiolabel position [Phenyl

- U

14 C]dicamba

Lot No. 2344

- 2 2344

- 9

Purity 98.4% prior to and after application 98.0 % prior to application; 98.1% after application

Specific activity 9.91 mCi/mmol (99548 dpm/μg or 1.66 MBq/mg) 9.85 mCi/mmol (98946 dpm/μg, 1.65 MBq/mg)



## B.3. Study Use Pattern

The isotopically diluted test substance was formulated as an aqueous solution of the DGA salt

and diluted with water. Applications were made using a hand

- held sprayer

. Applications were

made as a single pre

- emergence soil broadcast application (PRE

- T) at 2.094 lb ae/A on the day

### Dicamba/ MON 11900/ PC Code 128931 / Monsanto Company

DACO 6.3/OPPTS 860.1300/OECD II 6.2.2, 6.2.3 & IIIA 8.2, 8.4.1, 8.4.2

Nature of the Residues in Plants

– Dicamba

- Tolerant Cotton (Event MON 88701)

D40 838 4/MRID No. 48728701 Page 7 of 24 

## of planting afte r seed was planted or as a single post

- emergence foliar application (POE

- T) at
2.065 lb ae/A 76 days after planting, when >50% of the plants had reached the first white flower growth stage. Samples of seed and leaves/stems were harvested at PHI s of 180 days for the

PRE

- T plot and 1 04 days for the POE

- T plot. The study use pattern is summarized in Table

B.3.1.

TABLE B.3.1. Use Pattern Information.

Chemical name [Phenyl

- U

14 C]dicamba

Application method The test material was formulated as an aqueous solution of the DGA salt, diluted with water, and applied as a single preemergence soil application (PRE

- T plot) or a single p ostemergence foliar application (POE

- T plot)

. Application rate PRE

- T : 2.094 lb ae / A ( 2,347 g ae /ha)

POE

- T : 2.065 lb ae /A ( 2,315 g ae /ha)

Number of applications PRE

- T : 1

POE

- T : 1

Timing of application PRE

- T : Single preemergence soil application made on the day of planting after seed was planted

POE

- T : Single p ostemergence application made at the first white flower stage (76 days after planting).

PHI PRE

- T : 180 days

POE

- T : 104 days



## B.4. Identification/ Characterization of Residues

B.4.1. Sample Handling and Preparation

Samples of seed and leaves/stems (gin byproducts) were harvested 180 days after application for

the PRE

- T plot and 104 days after application for the POE

- T plot. Th e seed cotton was ginned and delinted using a small

- scale gin at Excel Research Services, Inc. prior to shipment to the analytical laboratory (Monsanto Environmental Services Technology Center, St . Louis, MO).

The leaves and stems were shipped to PTRL West (Hercules, CA), where they were processed by grinding in a food proce sser in the presence of dry ice prior to shipment to the analytical laboratory. At the laboratory, the undelinted seed was further processed by cryogenic milling using a SPEX CertiPrep Freezer/Mill ; no further processing was required for the gin byproducts samples

. All samples were stored frozen at Monsanto (<

- 2 0 °C) prior to and after processing

when not in use.

Cotton seed samples were extracted three times with hexane followed by a single ACN

extract ion to remove oils

, after which they were extracted four times with ACN :water ( 40:60 ; v/v)

. Cotton gin byproducts were extracted four times with ACN :water ( 40:60 ; v /v)

. Following centrifugation, t he ACN/water extracts of each matrix were combined and rotary evaporated to

remove the ACN and part of the water. Evaporation was stopped a t near dryness for gin byproducts

, while seed extract s were not evaporated to dryness b ecause of issu es caused by insoluble materials. All samples were re

- dissolved in water containing 0.5% formic acid and

ACN (92 :8 , v:v ) and reserved for HPLC analysis.

Separate subsamples of PRE

- T and POE

- T seed were al so subjected to exhaustive hexane extraction for analysis of fatty acids. The samples were extracted with hexane (3x), and the

### Dicamba/ MON 11900/ PC Code 128931 / Monsanto Company

DACO 6.3/OPPTS 860.1300/OECD II 6.2.2, 6.2.3 & IIIA 8.2, 8.4.1, 8.4.2

Nature of the Residues in Plants

– Dicamba

- Tolerant Cotton (Event MON 88701)

D40 838 4/MRID No. 48728701 Page 8 of 24 

## hexane extracts were combined and evaporated to near dryness. Methanol and concentrated sulfuric acid were added, and the sampl e s w ere refluxed for 2.5 hours, then cooled and partitioned with hexane (3x). The combined extracts were evaporated to near dryness and

reconstituted in hexane for analysis by HPLC.

The non

- extractable residues remaining following ACN / water extraction of seed and gin byproducts were subjected to the following sequential enzymatic and chemical hydrolys i s /oxidation procedures ; mixtures were centrifuged or filtered after each step. Both seed and gin trash: 1) Phosphate rinse (in 0.05 M phosp hate buffer, pH 7.0, at RT , for 20 min.) 2) α

- Amylase hydrolysis for starch ( in 0.05 M phosphate buffer, pH 7.0, at 30 °C for 20 hr. ) 3) P rotease hydrolysis for protein ( in TRIS buffer, pH 7.0, at 30 °C for 19 hr. ) 4) EDTA extraction for pectin ( with 0.05 M EDTA:0.05 M sodium acetate buffer, pH 5.0, at

80 °C for 5 hr. , then at RT overnight ) 5) Chlorite o xidation for lignin ( in glacial acetic acid containing sodium chlorite at 70 °C

for 1 hr.

, then repeated with water, acetic acid, and sodium chlorite ) 6) Cellulase hydrolysis for cellulose ( in 0.05 M sodium acetate buffer, pH 5.0, at 37 °C for

20 hr. ) 7) 24% KOH h ydrolysis for hemicellulose (from 37 to 24 °C for several hours, then at 24 °C overnight) Seed only: 8) Concentrated sulfuric acid (at RT , overnight) 9) Protease hydrolysis (POE

- T seed; in 0.05 M TRIS buffer , pH 7.0, at 30 °C for 2 3 hr. ) 10) Hot DMSO extraction ( POE

- T seed ; heated to 78 °C over 1.5 hr.

, at RT for 17 hr. , heated

to 70 °C over several hr.

, at RT for 2 hr. , heated to 77 °C over 2.5 hr. , cooled t o RT )

Partitioning with ethyl acetate was used to investigate organic/aqueous partitioning behavior of dicamba residues and to facilitate quantitation of the metabolites or to facilitate isolation and purification of metabolites

. T he combined ACN / water extracts of seed and gin byproducts were rotary evaporated to remove ACN

, and the remaining aqueous solution was acidified to pH 1

- 2 with concentrated HCl and partitioned three times with ethyl acetate. The ethyl acetate phase was evaporated to dryn ess
, and the residue was reconstituted in 0.5% aqueous formic acid and acetonitrile for HPLC analysis

. The hydrolysates resulting from protease hydrolysis of PRE

- T seed and 24% KOH hydrolysis of PRE

- T and POE

- T seed were acidified to pH <2 with

concentrat ed HCl, then partitioned three times with ethyl acetate. For each digest, the combined ethyl acetate extracts and the remaining aqueous layer were aliquoted for LSC analysis

. Quantitative results for the partitioning procedures are not included herein.

B.4.2. Analytical Methodology

TRR were determined by combustion/LSC

. Radioactivity in extracts was determined by direct LSC, and radioactivity in non

- extractable fractions was determined by combustion/LSC. The limit of quantitation for TRR determination s was not reported.

### Dicamba/ MON 11900/ PC Code 128931 / Monsanto Company

DACO 6.3/OPPTS 860.1300/OECD II 6.2.2, 6.2.3 & IIIA 8.2, 8.4.1, 8.4.2

Nature of the Residues in Plants

– Dicamba

- Tolerant Cotton (Event MON 88701)

D40 838 4/MRID No. 48728701 Page 9 of 24 

## Seed and gin byproducts extracts were subjected to HPLC /UV analysis for quantitation and generation of the metabolite profile. The HPLC systems were equipped with various columns, a UV detector (280 , 254, or 210 nm), and a radioactive flow detector

. Two methods were employed for detection and quantitation of radioactivity: HPLC with radioactive flow detection (HPLC/RAD ) in which the effluent from the UV detector was pumped through the flow detector

to a detection cell for an analyti cal scale or semi

- prepa rative scale HPLC column, and HPLC

with fraction collection and LSC detection (HPLC/LSC) , in which fractions were collected at

0.3

- min. intervals. HPLC Method B (C

- 18 column, gradient mobile phase of 0.5% formic acid and ACN) using HPLC/RAD and HPLC/LSC was the primary method used for residue quantitation and for generation of the metabolite profile.

Up to 38 metabolite peaks were observed in cotton seed and gin byproducts . All metabolite peaks accounting for ≥2% TRR were isolated and purified by preparative HPLC for further investigation. Metabolites were isolated by preparative HPLC fractionation using HPLC method (Method B). The isolated metabolite fractions were typically purified by preparative fractionation using HPLC Metho d D (phenyl column, gradient mobile phase of 0.5% formic acid and ACN). HPLC Method C (C18 column, gradient mobile phase of 0.5% formic acid and

methanol ) was used to further separate isolated metabolites from matrix contaminants.

The primary technique for identification of metabolites was mass spectral analysis which provided molecular weights and some structural information. LC/ MS or LC/MS/MS with

e lectrospray ionization (ESI) was conducted using an ion trap mass spectrometer int erfaced with an Agilent 1200 RRLC system and HPLC methods M (C18 column, gradient mobile phase of 0.1% formic acid and methanol), N and O (phenyl column, gradient mobile phase of 0.1% formic acid and ACN), P (amino column, isocratic mobile phase of 0.03% N H 4 

## OH and ACN), and Q (amino column, isocratic mobile phase of water and ACN). Typically, negative ion MS was utilized for the metabolites and their hydrolysis products

, and positive ion MS was utilized

for neutral derivatives of the metabolites and for t he two amides (dicamba amide and DCSA amide). GC/EI/MS conducted in the positive ion mode was utilized for volatile derivatives of

the metabolit es or their derivatives or hydrolysis products.

HPLC co

- chromatography and/or retention time comparisons , as well as comparison of the chromatographic and mass spectral properties of the metabolite derivatives or hydrolysates to those of synthetic reference standards was used to confirm the identities of the metabolites. T he

chemical names and structures of the reference standards used in the study are presented in Appendix I.

For the purpose of identification, metabolite peaks were isolated primarily from the POE

- T gin byproducts ; m etabolites were not specifically isolated from the seed due to low residue levels ( with the exceptions of Peaks 19, 34, and 36 from POE

- T seed ). I t was assumed that metabolite peaks in the seed extracts represent the same metabolite peaks in the gin byproducts with the

same or similar retention times. T o confirm the ide ntity of the peaks in seed, the PRE

- T seed

extract was co

- injected with a mixture of standards corresponding to metabolites identified in

POE

- T and PRE

- T gin byproducts

, and the retention times were compared and analyzed by HPLC/RAD using Method B.

### Dicamba/ MON 11900/ PC Code 128931 / Monsanto Company

DACO 6.3/OPPTS 860.1300/OECD II 6.2.2, 6.2.3 & IIIA 8.2, 8.4.1, 8.4.2

Nature of the Residues in Plants

– Dicamba

- Tolerant Cotton (Event MON 88701)

D40 838 4/MRID No. 48728701 Page 10 of 24 

## Prior t o analysis by HPLC or mass spectrometry methods, isolated metabolites were characterized using a variety of methods alone or in combination to provide additional structural information, including:

 

## Partitioning with ethyl acetate (see above)

 

## Acid hydrolysis (to effect removal of sugar moieties) with 2 N HCl (at 95

- 100 °C for 1

- 2

hr. ) or 1 N HCl at (60 °C for 4 hr. )

 

## Base hydrolysis with 2 N NaOH (at 60 °C for 4 hr. )

 

## Methylation (for detection of carboxylic acid and/or phenolic groups) with

trim ethylsilyldiazomethane at RT for 35 min.

 

## Acetylation (for detection of hydroxyl or phenolic groups) with acetic anhydride (at RT

for 1

- 2 hr. or overnight)

Identification of the metabolites DCGA glucoside (Peak 6) and DCSA glucoside (Peak 19) were

achieved by LC/MS and/or HPLC/RAD comparisons with the same metabolites isolated from soy forage from a metabolism study on dicamba

- tolerant soybean (Report No. MSL0022659; no MRID found)

.

Metabolite Peak 1 (found in all mat r ices) was isolated from ACN/water extr acts HPLC Method

B. Peak 1 was also isolated from POE

- T seed extracts using C18 solid phase extraction and sequential elution with water, ACN, and ethyl acetate. Peak 1 was also characterized by a number of procedures including : partitioning with ethyl acetate at acidic and neutral pH (<1.5 and 6) ; acid hydrolysis followed by HPLC; HPLC using Method J (amino column and isocratic mobile phase of 0.03% NH 4 

## OH in water and ACN) to isolate possible saccharides and amino

acids ; and HPLC (PRE

- T seed) using Method R (amino column, gradient mobile phase of water and ACN) to resolve component peaks in the organic phase following ethyl acetate partitioning

. The Peak 1 radioactive components were also analyzed by negative ion electrospray LC/MS using HPLC Method Q and were compared to representative mono

- , di

- and tri

- saccharide

reference standards and amino acids

. Confirmation of the identities of the mono and disaccharides observed in the LC/MS analyses was achieved by derivatization and GC/EI/MS analys es.

For identification of triglycerides, including fatty acids and glycerol in the hexane extracts of

seed

, the hexane phase following acidic methanolysis was analyzed by HPLC Method I ( silver

impregnated strong cation exchange column, isocratic mobile p hase of 1% ACN in hexane and

hexanes )

. The peaks observed were consistent with those of the fatty acid methyl ester reference standards of stearic, oleic, and linoleic acid. Radioactivity remaining in the methanol fraction was designated glycerol.

C. RESULTS AND DISCUSSION

T he sample storage conditions and durations for dicamba

- tolerant cotton matrices are presented

in Table C.1

. 1. TRR determinations were completed within 10 days of harvest. The petitioner

stated that i nitial HPLC a nalyses were conducted for POE

- T seed and gin trash extracts within 30 days of sample receipt to establish baseline profiles. Based on the dates of sample collection ( 10/28/09 ) and the experimental completion date ( 2/18/11, storage stability study ), final analysis

### Dicamba/ MON 11900/ PC Code 128931 / Monsanto Company

DACO 6.3/OPPTS 860.1300/OECD II 6.2.2, 6.2.3 & IIIA 8.2, 8.4.1, 8.4.2

Nature of the Residues in Plants

– Dicamba

- Tolerant Cotton (Event MON 88701)

D40 838 4/MRID No. 48728701 Page 11 of 24 

## of all samples w as conducted within 47 8 days (15.7 months) of sample collection. To demonstrate storage stability, extracts of PRE

- T and POE

- T seed and gin byproduct s were analyzed at the beginning of the study (47

- 99 days after collection) , at a midway poin t (296

- 308 days after collection) and again at 47 5

- 478 days after collection (POE

- T samples only). The extractability of radioactivity was similar at all time intervals (Table C.1.2) , and comparison of the metabolite profiles for the POE

- T seed and gin byproduct samples following initial and final analysis indicated only negligible change s in distribution of residues

. These data are sufficient to support the sample storage conditions and intervals from the submitted study.

TRR in dicamba

- resistant matrices are reported in Table C.2.1. TRR were determined by combustion/LSC. Following a single pre

- emergence soil application of [ 14 

## C]dicamba at 2.094 lb ae/A, TRR were 0.1621 ppm in seed and 0.8493 ppm in gin byproducts harvested at a 180

- day

PHI. Fol lowing a single post

- emergence foliar application at 2.065 lb ae/A, TRR were 0.9778 ppm in seed and 60.0235 ppm in gin byproducts harvested at a 104

- day PHI.

The distribution of radioactivity in dicamba

- tolerant cotton matrices is presented in Table C. 2.2.

Extraction with hexane released 18.91% and 11.47% TRR from PRE

- T and POE

- T seed, respectively; extraction with acetonitrile (ACN) and ACN/water together released an additional 11.91% and 26.78% TRR, respectively. In gin byproducts, extraction with A CN/water released the majority of the radioactivity: 76.25% and 71.21% TRR from PRE

- T and POE

- T gin byproducts, respectively. Remaining non

- extractable residues for all matrices were subjected to sequential enzymatic and chemical hydrolysis and oxidation procedures, including a phosphate rinse followed by hydrolysis with α

- amylase and protease; EDTA extraction; oxidation with chlorite; hydrolysis with cellulase, 24% aqueous KOH, and sulfuric acid (seed only); a second hydrolysis with protease (POE

- T seed o nly); and hot DMSO extraction (POE

- T seed only). These procedures released 52.12

- 61.82% TRR in seed and 21.13

- 28.18% TRR in gin byproducts. Non

- extractable residues remaining following hydrolysis procedures were 7.35% TRR (0.012 ppm) in PRE

- T seed, 9.66% TRR (0.094 ppm) in POE

- T seed, 2.62% TRR (0.022 ppm) in PRE

- T gin byproducts, and 0.61% TRR (0.365 ppm) in POE

- T gin byproducts. These procedures adequately extracted the majority of residues (90.3

- 99.4%) from all cotton matrices.

Extraction results wer e normalized; therefore, accountabilities were ~100%.

Up to 38 peaks were observed in the initial HPLC/UV profiles of seed and gin byproducts. All metabolite peaks accounting for ≥2% TRR were isolated and purified by preparative HPLC for further investig ation. After isolation, r esid ues were identifi ed by HPLC /UV and by mass spectral analysis including LC/MS and LC/MS/MS; GC/EI/MS was u sed for volatile derivatives

of the metabolit es or their hydrolysis products

. Isolated metabolites were subjected to partitioning with ethyl aceta t e, acid, base or enzymatic hydrolysis, and/or chemical derivatizations ( methylation and acetylation ) to provide additional structural information

. Comparison of the chromatographic and mass spectral properties of the metaboli te derivatives or hydrolysates to those of synthetic reference standards was used to confirm the identities of the

metabolites.

The characterization and identification of residues in dicam b a

- tolerant cotton matrices are

summarized in Table C.2.3. The parent, dicamba, was identified in all matrices at low levels:

### Dicamba/ MON 11900/ PC Code 128931 / Monsanto Company

DACO 6.3/OPPTS 860.1300/OECD II 6.2.2, 6.2.3 & IIIA 8.2, 8.4.1, 8.4.2

Nature of the Residues in Plants

– Dicamba

- Tolerant Cotton (Event MON 88701)

D40 838 4/MRID No. 48728701 Page 12 of 24 

## 0.09

- 0.85% TRR (0.0001

- 0.008 ppm) in seed and 0.50

- 4.48% TRR (0.004

- 2.691 ppm) in gin byproducts. Metabolites DCSA glucoside, DCSA, and DCGA glucoside were also present in all

matrices. DCSA glucoside was the major identified metabolite in gin byproducts, at 16.83

- 27.77% TRR (0.236

- 10.101 ppm) and was present in seed at low levels (0.73

- 3.42% TRR). Free

DCSA was the next most abundant metabolite in POE

- T gin byproducts (13.39% TRR, 8.035 ppm) and was present in PRE

- T gin byproducts at 5.42% TRR and in seed at 0.06

- 1.91%TRR. DCGA glucoside was identified at low levels in all matrices (0.09

- 2.46% TRR). The remaining identified metabolites were found in gin byproducts only and accounted fo r ≤4.71% TRR each, including MCTHBA glucoside, MCDHBA glucoside sulfate, and MCDHBA glucoside in POE

T gin byproducts, MCTHBA cyclic glucoside in both PRE

- T and POE

- T gin byproducts, and

dicamba amide in PRE

- T gin byproducts only. Because dicamba amide wa s only observed in

PRE

- T plants, it is believed to be a soil metabolite that was taken up by the plants.

Remaining extractable radioactivity in cotton seed and gin byproducts was characterized as triglycerides and sugars / natural products. In the hexane extracts of seed, triglycerides including fatty acids (palmitic, stearic, linoleic, and oleic) accounted for 10.07

- 16.05% TRR (0.026

- 0.098 ppm), while glycerol accounted for 1.35

- 2.86% TRR. In the ACN/water extracts of all matrices, sugars/natural product s (Peak 1) accounted for 2.69

- 8.94% TRR (0.0074

- 1.613 ppm). Th is peak was characterized extensively, indicating that it was a mixture of polar materials, including sugars such as glucose, sucrose, and raffinose, and amino acids such as phenylalanine and

g lutamic acid. These sugars and amino acids are likely produced from extensive metabolism of dicamba to small molecules which are incorporated into natural plant constituents (such as carbohydrate, protein and oil) through normal plant metabolic processes. Unidentified components in oil accounted for 0.84

- 1.94% TRR in cotton seed; discrete unknowns in the ACN/water extracts together accounted for 0.86

- 15.96% TRR (0.00 49

- 0.518 ppm) in seed and gin byproducts, with none present at >2.91% TRR.

Radioactivit y in non

- extractable residues was characterized by sequential enzymatic and chemical hydrolysis/oxidation. In seed, these procedures indicated that the majority of radioactivity was incorporated into hemicellulose (13.49

- 14.43% TRR, 0.023

- 0.132 ppm),

prot ein (10.10

- 17.71% TRR, 0.029

- 0.099 ppm ), and lignin (6.18

- 14.0% TRR, 0.01

- 0.136 ppm). Partitioning of the hemicellulose and protein fractions with ethyl acetate following acidification resulted in low levels of radioactivity in the organic phase, indicati ng that the residues are polar in nature and unlikely to be related to dicamba metabolites. Other fractions (including starch, pectin, and cellulose) individually accounted for ≤7.4% TRR. In gin byproducts, radioactivity was incorporated primarily into s tarch (2.73

- 7.05% TRR, 0.023

- 4.234 ppm), lignin (5.28

- 6.60% TRR, 0.045

- 3.962 ppm), and hemicellulose (2.80

- 5.35% TRR, 0.045

- 1.679 ppm). Other fractions (including protein, pectin, and cellulose) individually accounted for ≤6.2% TRR. The

conclusion from t hese analyses is that the metabolism of dicamba to small organic molecules results in the incorporation of the label into endogenous plant components (e.g., proteins, sugars, lignin, etc.) through normal metabolic processes.

The following peaks were desig nated unidentified following characterization/identification attempts; however, the petitioner provided the following tentative identifications:

 Peak 5: a highly conjugated dicamba

- related compound

### Dicamba/ MON 11900/ PC Code 128931 / Monsanto Company

DACO 6.3/OPPTS 860.1300/OECD II 6.2.2, 6.2.3 & IIIA 8.2, 8.4.1, 8.4.2

Nature of the Residues in Plants

– Dicamba

- Tolerant Cotton (Event MON 88701)

D40 838 4/MRID No. 48728701 Page 13 of 24 

##  Peak 9: conjugate of DCSA amide (Peak 27)

 Peak 14 : conjugate of DCSA amide ( Peak 27 ), or 3

- chloro

- 2,6

- dihydroxybenzoic acid

 Peak 27: primary amide of DCSA (DCSA amide)

 Peak 32 : conjugate of DCSA

C.1. Storage Stability

TRR determinations were completed within 10 days of harvest. The petitioner stated that i nitial

HPLC a nalyses were conducted for POE

- T seed and gin trash extracts within 30 days of sample receipt to establish baseline profiles. Based on the dates of sample collection ( 10/28/09 ) and the experimental completion date ( 2/18/11, storage stability study ), final analysis of all samples w as conducted within 478 days (15.7 months) of sample collection. To demonstrate storage stability,

extracts of PRE

- T and POE

- T seed and gin byproducts were analyzed at the beginning of the study (47

- 99 days after collection), at a midway point (296

- 308 days after collection) and again

at 475

- 478 days after collection (POE

- T samples only). The extractability of radioactivity was similar at all time intervals (Table C.1.2), and comparison of the metabolite pr ofiles for the POE

T seed and gin byproduct samples following initial and final analysis indicated only negligible changes in distribution of residues. These data are sufficient to support the sample storage

conditions and intervals from the submitted stu dy.

TABLE C.1. 1. Summary of Storage Conditions

Matrix Storage Temperature (°C) Actual Study Duration 1 Interval of Demonstrated Storage Stability

Seed <

- 20 47 days 47 8 days (15.7 months)

Gin byproducts 79

- 99 days 47 5 days (15.6 months)

1 In terval between harvest and ini tial analysis of extracts by HPLC

.



### TABLE C.1.2. Comparison of Extraction Results Obtained at the Beginning, Middle, and End of the

Analytica l phase.

Matrix Normalized Extractability (%)

Initial (47

- 79 days) Mid (296

- 308 days) Final (475

- 478 days)

PRE

- T Seed 27.42 30.84 Not analyzed

PRE

- T Gin byproducts 73.57 76.24

POE

- T Seed 35.07 38.26 40.37

POE

- T Gin byproducts 67.76 71.28 69.10

1 In terval between harvest and ini tial analysis of extracts by HPLC.

### Dicamba/ MON 11900/ PC Code 128931 / Monsanto Company

DACO 6.3/OPPTS 860.1300/OECD II 6.2.2, 6.2.3 & IIIA 8.2, 8.4.1, 8.4.2

Nature of the Residues in Plants

– Dicamba

- Tolerant Cotton (Event MON 88701)

D40 838 4/MRID No. 48728701 Page 14 of 24 

## C.2. Identification, Characterization, and Distribution of Residues

TABLE C.2.1. Total Radioactive Residues (TRR) in Dicamba

- Tolerant Cotton Matrices

. Matrix Timing and Applic. No. PHI (days) Residues (ppm [ 14 C]dicamba)

Seed Gin byproducts

Dicamba

- tolerant

cotton Single pre

- emergence:

2.094 lb ae / A (2,347 g ae /ha) 180 0.1621 0.8493

Single post

- emergence: 2.065 lb ae /A (2,315 g ae /ha) 104 0.9778 60.0235



### TABLE C.2.2. Distribution of the Parent and the Metabolites in Dicamba

- Tolerant Cotton Matrices

Following Application of [Phenyl

- U

14 

### C]dicamba at 2.094 lb ae /A (pre

- e mergence) or 2.065

lb ae /A (post

- emergence)

. 1

Metabolite Fraction Preemergence Postemergence

Seed Gin byproducts Seed Gin byproducts

TRR = 0.1621 ppm TRR = 0.8493 ppm TRR = 0.9778 ppm TRR = 60.02 4 ppm

% TRR ppm %TRR ppm %TRR ppm %TRR ppm

Hexane [initial extracts] 2 20.25 0.03 28 11. 5 1 0.11 3

Triglycerides 14.41 0.0230 8.80 0. 0 86

Unidentified 5.84 0.00 98 2.71 0.027

H exane [ acidic methanolysis ] 3 18.91 0.031 11.47 0.112

Hexane: F atty acid s 4 16. 05 0. 026 10.07 0.098

Acidi c MeOH: Glycerol 2.86 0.005 1.35 0.013

Unidentified (oil) 1.94 0.003 0.84 0.008

ACN 0.44 0.001 0.48 0.005

ACN/ water 11. 47 0.019 76.2 5 0.648 26. 3 0 0.2 57 71.2 1 42.7 44

Combined ACN, ACN/water 5 11.91 0.020 26.78 0.262

D icamba 0.09 0.0001 0.50 0.004 0.85 0.008 4.48 2.691

DCSA g lucoside 0.73 0.0012 27.77 0.236 3.42 0.033 16.83 10.101

DCSA 0.06 0.0001 5.42 0.046 1.91 0.019 13.39 8.035

DCGA g lucoside 0.09 0.0001 0.68 0.006 1.10 0.011 2.46 1.476

MCTHBA g lucoside

-- -- -- -- -- -- 3.33 1.998

MCDHBA g lucoside

s ulfate -- -- -- -- -- -- 4.71 2.828

MCDHBA g lucoside

-- -- -- -- -- -- 0.84 0.505

MCTHBA c yc. g lucoside

-- -- 2.94 0.025

-- -- 2.76 1.656

Dicamba a mide

-- -- 3.22 0.027

-- -- -- -- Sugars /natural products 4.59 0.0074 8.94 0.076 5.61 0.055 2.69 1.613

Discrete u nknowns 6 3.03 0.0049 15.96 0.136 5.36 0.052 0.86 0.518

Non extractable 69.16 0.112 23.76 0.202 61.74 0.604 28.72 17.239

Phosphate rinse 3.87 0.006 3.25 0.028 1.96 0.019 6.20 3.72

α

- Amylase (s tarch ) 2.79 0.005 2.73 0.023 2.19 0.021 7.05 4.234

Protease (p rotein ) 17.71 0.029 1.71 0.015 7.43 0.073 2.25 1.350

EDTA extract (p ectin ) 7.40 0.012 1.91 0.016 4.04 0.039 2.66 1.595

Chlorite (l ignin ) 6.18 0.010 5.28 0.045 8.43 0.082 6.60 3.962

Cellulase (cellulose) 4.37 0.007 0.90 0.008 3.13 0.031 0.62 0.374

24% KOH (hemicellulose) 14.43 0.023 5.35 0.045 13.49 0.132 2.80 1.679

### Dicamba/ MON 11900/ PC Code 128931 / Monsanto Company

DACO 6.3/OPPTS 860.1300/OECD II 6.2.2, 6.2.3 & IIIA 8.2, 8.4.1, 8.4.2

Nature of the Residues in Plants

– Dicamba

- Tolerant Cotton (Event MON 88701)

D40 838 4/MRID No. 48728701 Page 15 of 24 TABLE C.2.2. Distribution of the Parent and the Metabolites in Dicamba

- Tolerant Cotton Matrices

Following Application of [Phenyl

- U

14 

### C]dicamba at 2.094 lb ae /A (pre

- e mergence) or 2.065

lb ae /A (post

- emergence)

. 1

Metabolite Fraction Preemergence Postemergence

Seed Gin byproducts Seed Gin byproducts

TRR = 0.1621 ppm TRR = 0.8493 ppm TRR = 0.9778 ppm TRR = 60.02 4 ppm

% TRR ppm %TRR ppm %TRR ppm %TRR ppm

Sulfuric acid 5.07 0.008 3.21 0.031

Protease (second p rotein ) 2.67 0.026

Hot DMSO (second lignin) 5.57 0.054

Non extractable 7.35 0.012 2.62 0.022 9.66 0.094 0.61 0.365

1 Extraction results were normalized for the original extraction procedures and for characterization of nonextractable residues (See Tables 14 and 15). Shading indicates that the extraction step and/or characterization analysis was not conducted for the matrix in question.

2 Results as reported in Tables 9 and 11.

3 Calculated by the study reviewer to show distribution of triglycerides as fatty acids and glycerol, based on normalized values reported in Table 13.

4 Including palmitic, stearic, oleic, and linoleic acids

5 Calculated by the study reviewer by summing.

6 Unknowns (discrete quantitated peaks) are comprised of 19 components in PRE

- T seed (each ≤0.45% TRR, ≤0.0007 ppm); 12 components in POE

- T seed (each ≤0.81% TRR, ≤0.008 ppm); 11 components in PRE

- T gin byproducts (each ≤2.91% TRR, ≤0.025 ppm); and 1 component in PO E

- T gin byproducts (0.86% TRR, 0.518 ppm)

.



### TABLE C.2.3. Summary of Characterization and Identification of Radioactive Residues in Dicamba

Tolerant Cotton Matrices Following Application of [Phenyl

- U

14 

### C]dicamba at 2.094 lb ae /A (pre

- e mergence) or 2.065 lb ae /A (post

- emergence)

Preemergence Postemergence

Seed Gin byproducts Seed Gin byproducts

TRR = 0.1621 ppm TRR = 0.8493 ppm TRR = 0.9778 ppm TRR = 60.02 4 ppm

% TRR ppm %TRR ppm %TRR ppm %TRR ppm

Dicamba 0.09 0.0001 0.5 0 0.004 0.85 0.008 4.48 2.691

DCSA g lucoside 0.73 0.0012 27.77 0.236 3.42 0.033 16.83 10.101

DCSA 0.06 0.0001 5.42 0.046 1.91 0.019 13.39 8.035

DCGA g lucoside 0.09 0.0001 0.68 0.006 1.1 0.011 2.46 1.476

MCTHBA g lucoside

-- -- -- -- -- -- 3.33 1.998

MCDHBA g lucoside

s ulfate -- -- -- -- -- -- 4.71 2.828

MCDHBA g lucoside

-- -- -- -- -- -- 0.84 0.505

MCTHBA c yc.

g lucoside -- -- 2.94 0.025

-- -- 2.76 1.656

Dicamba a mide

-- -- 3.22 0.027

-- -- -- -- Fatty acids 16.05 0.026

-- -- 10.07 0.098

-- -- Glycerol 2.86 0.005

-- -- 1.35 0.013

-- -- Sugars /natural products 4.59 0.0074 8.94 0.076 5.61 0.055 2.69 1.613

Phosphate rinse 3.87 0.006 3.25 0.028 1.96 0.019 6.2 0 3.72

Starch 2.79 0.005 2.73 0.023 2.19 0.021 7.05 4.234

Protein 17.71 0.029 1.71 0.015 10.1 0 0.099 2.25 1.35 0

Pectin 7.4 0 0.012 1.91 0.016 4.04 0.039 2.66 1.595

### Dicamba/ MON 11900/ PC Code 128931 / Monsanto Company

DACO 6.3/OPPTS 860.1300/OECD II 6.2.2, 6.2.3 & IIIA 8.2, 8.4.1, 8.4.2

Nature of the Residues in Plants

– Dicamba

- Tolerant Cotton (Event MON 88701)

D40 838 4/MRID No. 48728701 Page 1 6 of 24 TABLE C.2.3. Summary of Characterization and Identification of Radioactive Residues in Dicamba

Tolerant Cotton Matrices Following Application of [Phenyl

- U

14 

### C]dicamba at 2.094 lb ae /A (pre

- e mergence) or 2.065 lb ae /A (post

- emergence)

Preemergence Postemergence

Seed Gin byproducts Seed Gin byproducts

TRR = 0.1621 ppm TRR = 0.8493 ppm TRR = 0.9778 ppm TRR = 60.02 4 ppm

% TRR ppm %TRR ppm %TRR ppm %TRR ppm Lignin 6.18 0.01 0 5.28 0.045 14 .0 0 0.136 6.6 0 3.962

Cellulose 4.37 0.007 0.9 0 0.008 3.13 0.031 0.62 0.374

Hemicellulose 14.43 0.023 5.35 0.045 13.49 0.132 2.8 0 1.679

Acid Hydrolysates 5.07 0.008

-- -- 3.21 0.031

-- -- Unidentified (oil) 1.94 0.003

-- -- 0.84 0.008

-- --

Discret e unknowns 3.03 0.0049 15.96 0.136 5.36 0.052 0.86 0.518

Total identified 0.97 0.0015 40.53 0.344 7.28 0.071 48.8 0 29.29

Total characterized 90.29 0.1463 46.03 0.392 75.35 0.734 31.73 19.045

Total extractable 92.6 5 0.15 0 97.38 0.827 90.34 0.883 99.39 59.659

Unextractable (PES) 1 7.35 0.012 2.62 0.022 9.66 0.094 0.61 0.365

Accountability 2 100 100 100 100

1 Residues remaining after exhaustive extractions.

2 Accountability = (Total extractable + Total unextractable)/(TRR from combustion analysis; see TABLE C.2.1) * 100.



## C.3. Proposed Metabolic Profile

Based on the results of the metabolism study on dicamba

- tolerant cotton, the petitioner proposes

that the metabolism of dicamba proceeds by initial demethylation of dicamba to form DCSA through the action of the dicamba O

- demethylase enzyme, which is the product of the DMO gene introduced to confer dicamba tolerance. The petitioner noted that, due to the predominance

of the O

- demethylation pathway, 5

- OH

- dicamba is not observed as a metabolite in dicamba

tolerant cotton or soybean. While free DCSA was observed in cotton matrices (POE

- T and PRE

T gin byprod ucts in particular), the majority of DCSA is found as its conjugate DCSA glucoside. Some DCSA glucoside is further modified by hydroxyl replacement of chlorine to form MCDHBA glucoside which is then further conjugated by sulfation to form MCDHBA glucoside sulfate. In a minor pathway, DCSA is hydroxylated at the 5

- position, presumably by a P

- 450 enzyme or other oxygenase, to form DCGA. DCGA is not observed as a free metabolite in cotton matrices but is converted to DCGA glucoside in which the glucose moie ty is attached to the hydroxyl group at the 5

- position of DCGA (based upon comparison to the same metabolite

identified in dicamba

- tolerant soybean). DCGA glucoside is further converted to MCTHBA glucoside by hydroxyl replacement of chlorine at the 3

- posi tion. Alternatively, MCTHBA cyclic glucoside may form from DCGA glucoside through displacement of chlorine at the 6

- position of the DCGA portion of the molecule by a hydroxyl group of the glucose moiety. Dicamba amide was a minor metabolite that was only observed in PRE

- T matrices. The petitioner concluded that this metabolite is most likely the result of the metabolic conversion of the carboxylic acid moiety of dicamba to an amide functional moiety via soil microbial metabolism.

### Dicamba/ MON 11900/ PC Code 128931 / Monsanto Company

DACO 6.3/OPPTS 860.1300/OECD II 6.2.2, 6.2.3 & IIIA 8.2, 8.4.1, 8.4.2

Nature of the Residues in Plants

– Dicamba

- Tolerant Cotton (Event MON 88701)

D40 838 4/MRID No. 48728701 Page 17 of 24 

## The proposed metabolic pathway is depicted in Figure C.3.1, which was copied without

alteration from MRID 48728701

.

FIGURE C.3.1. Proposed Metabolic Profile of Dicamba in Dicamba

- Tolerant Cotton

TABLE C.3.1. Identification of Compounds from Metabolism Study

Common name/code Chemical name Chemical structure

Dicamba

(Peak 36) 3,6

- dichloro

- 2

- methoxybenzoic acid

DCSA Glucoside

( Peak 19 ) 3,6

- dichloro

- 2

- (β

- D

glu copyranosyloxy)benzoic acid

### Dicamba/ MON 11900/ PC Code 128931 / Monsanto Company

DACO 6.3/OPPTS 860.1300/OECD II 6.2.2, 6.2.3 & IIIA 8.2, 8.4.1, 8.4.2

Nature of the Residues in Plants

– Dicamba

- Tolerant Cotton (Event MON 88701)

D40 838 4/MRID No. 48728701 Page 18 of 24 TABLE C.3.1. Identification of Compounds from Metabolism Study

Common name/code Chemical name Chemical structure

DCSA

( Peak 34 ) 3,6

- dichloro

- 2

- hydroxy benzoic acid

DCGA Glucoside

( Peak 6 ) 2,5

- dichloro

- 3

- (β

- D

- glucopyranosyloxy)

- 6

- hydroxybenzoic acid

MCTHBA Glucoside

( Peak 12 ) 2

- Chloro

- 5,6

- dihydroxy

- 3

- (3,4,5

- trihydroxy

- 6

- hydroxymethyl

tetrahydropyran

- 2

- yloxy)benzoic acid

MCDHBA Glucoside Sulfate

( Peak 16 ) 6

- Chloro

- 3

- hydroxy

- 2

- (3,4,5

- trihydroxy

6

- sulfooxymethyltetrahydro

- pyran

- 2

- yloxy)benzoic acid

MCDHBA Glucoside

( Peak 18 ) 6

- Chloro

- 3

- hydroxy

- 2

- (3,4,5

- trihydroxy

6

- hydroxymethyltetrahydro

- pyran

- 2

- yloxy)benzoic acid

MCTHBA Cyclic Glucoside

( Peak 21 ) 7

- Chloro

- 3,4,6

- trihydroxy

- 2

- hydroxymethyl

- 3,4,4a,9a

- tetrahydro

- 2 H

1,9.10

- trioxa

- anthracene

5

- carboxylic acid

### Dicamba/ MON 11900/ PC Code 128931 / Monsanto Company

DACO 6.3/OPPTS 860.1300/OECD II 6.2.2, 6.2.3 & IIIA 8.2, 8.4.1, 8.4.2

Nature of the Residues in Plants

– Dicamba

- Tolerant Cotton (Event MON 88701)

D40 838 4/MRID No. 48728701 Page 19 of 24 TABLE C.3.1. Identification of Compounds from Metabolism Study

Common name/code Chemical name Chemical structure

Dicamba Amide

( Peak 29 ) 3,6

- dichloro

- 2

- methoxybenzamide



## D. CONCLUSION

The results of the metabolism study adequately delineate the nature of the residue in dicamba

tolerant cotton tr eated with dicamba

. Following a single pre

- emergence soil application of
[ 14 

## C]dicamba at 2.094 lb ae/A, TRR were 0.1621 ppm in seed and 0.8493 ppm in gin byproducts

harvested at a 180

- day PHI. Following a single post

- emergence foliar application at 2.065 lb ae/A, TRR were 0.9778 ppm in seed and 60.0235 ppm in gin byproducts harvested at a 104

- day

PHI.

The parent, dicamba , was identified in all matrices at low levels: 0.09

- 0.85% TRR in seed and

0.50

- 4.48% TRR in gin byproducts. Metabolites DCSA glucoside, DCSA, and DCGA glucoside

were also present in all matrices. DCSA glucoside was the major identified metabolite in gi n byproducts, at 16.83

- 27.77% TRR and was present in seed at low levels (0.73

- 3.42% TRR).

Free DCSA was the next most abundant metabolite in PO E

- T gin byproducts (13.39% TRR ) and was present in PRE

- T gin byproducts at 5.42% TRR and in seed at 0.06

- 1.91%TR R. DCGA glucoside was identified at low levels in all matrices (0.09

- 2.46% TRR). The remaining identified metabolites were found in gin byproducts only and accounted for ≤4.71% TRR each, including MCTHBA glucoside, MCDHBA glucoside sulfate, and MCDHBA gl ucoside in POE

T gin byproducts, MCTHBA cyclic glucoside in both PRE

- T and POE

- T gin byproducts, and

dicamba amide in PRE

- T gin byproducts only. Because dicamba amide was only observed in

PRE

- T plants, it is believed to be a soil metabolite that was taken up by the plants.

Remaining extractable radioactivity in seed and gin byproducts was characterized as triglycerides and sugars / natural products. In the hexane extracts of seed, triglycerides including fatty acids (palmitic, stearic, linoleic, and oleic) accounted for 10.07

- 16.05% TRR, while glycerol accounted for 1.35

- 2.86% TRR. In the ACN/water extracts of all matrices, sugars/natural products (Peak 1) accounted for 2.69

- 8.94% TRR. Unidentified components in oil

accounted for 0.84

- 1.94% TRR in seed ; d iscrete unknowns in the ACN/water extracts together

accounted for 0.86

- 15.96% TRR in seed and gin byproducts, with none present at >2.91% TRR. Radioactivity in non

- extractable residues was characterized by sequential enzymatic and chemical hydrolysis/ox idation. In seed, these procedures indicated that the majority of radioactivity was incorporated into hemicellulose (13.49

- 14.43% TRR), protein (10.10

- 17.71%

TRR ), and lignin (6.18

- 14.0% TRR ). Other fractions (including starch, pectin, and cellulose)

ind ividually accounted for ≤7.4% TRR. In gin byproducts, radioactivity was incorporated primarily into starch (2.73

- 7.05% TRR), lignin (5.28

- 6.60% TRR), and hemicellulose (2.80

-

### Dicamba/ MON 11900/ PC Code 128931 / Monsanto Company

DACO 6.3/OPPTS 860.1300/OECD II 6.2.2, 6.2.3 & IIIA 8.2, 8.4.1, 8.4.2

Nature of the Residues in Plants

– Dicamba

- Tolerant Cotton (Event MON 88701)

D40 838 4/MRID No. 48728701 Page 20 of 24 

## 5.35% TRR). Other fractions (including protein, pectin, and cellulose) individua lly accounted
for ≤6.2% TRR.

Based on the results of the metabolism study on dicamba

- tolerant cotton, the petitioner proposes

that the metabolism of dicamba proceeds by initial demethylation of dicamba to form DCSA through the action of the dicamba O

- dem ethylase enzyme, which is the product of the DMO gene introduced to confer dicamba tolerance. The petitioner noted that, due to the predominance

of the O

- demethylation pathway, 5

- OH

- dicamba is not observed as a metabolite in dicamba

tolerant cotton or soy bean. While free DCSA was observed in cotton matrices (POE

- T and PRE

T gin byproducts in particular), the majority of DCSA is found as its conjugate DCSA glucoside. Some DCSA glucoside is further modified by hydroxyl replacement of chlorine to form

MCDHB A glucoside which is then further conjugated by sulfation to form MCDHBA glucoside sulfate. In a minor pathway, DCSA is hydroxylated at the 5

- position, presumably by a P

- 450 enzyme or other oxygenase, to form DCGA. DCGA is not observed as a free metaboli te in cotton matrices but is converted to DCGA glucoside in which the glucose moiety is attached to the hydroxyl group at the 5

- position of DCGA (based upon comparison to the same metabolite

identified in dicamba

- tolerant soybean). DCGA glucoside is furth er converted to MCTHBA glucoside by hydroxyl replacement of chlorine at the 3

- position. Alternatively, MCTHBA cyclic glucoside may form from DCGA glucoside through displacement of chlorine at the 6

- position of the DCGA portion of the molecule by a hydroxy l group of the glucose moiety. Dicamba amide was a minor metabolite that was only observed in PRE

- T matrices. The petitioner concluded that this metabolite is most likely the result of the metabolic conversion of the carboxylic acid moiety

of dicamba to an amide functional moiety via soil microbial metabolism.

E. REFERENCES

None.

F. DOCUMENT TRACKING

Petition Number(s): 2F8067 DP Barcode(s): 408384

PC Code: 029801, 029802, 029806, 128931, 128944, & 129043

Template Version June 2005.

### Dicamba/ MON 11900/ PC Code 128931 / Monsanto Company

DACO 6.3/OPPTS 860.1300/OECD II 6.2.2, 6.2.3 & IIIA 8.2, 8.4.1, 8.4.2

Nature of the Residues in Plants

– Dicamba

- Tolerant Cotton (Event MON 88701)

D40 838 4/MRID No. 48728701 Page 21 of 24 APPENDIX I. Chemical Names and Structures of Reference Standards Used in Cotton Metabolism Study.

### Dicamba/ MON 11900/ PC Code 128931 / Monsanto Company

DACO 6.3/OPPTS 860.1300/OECD II 6.2.2, 6.2.3 & IIIA 8.2, 8.4.1, 8.4.2

Nature of the Residues in Plants

– Dicamba

- Tolerant Cotton (Event MON 88701)

D40 838 4/MRID No. 48728701 Page 22 of 24 APPENDIX I. Chemical Names and Structures of Reference Standards Used in Cotton Metabolism Study (cont.).

### Dicamba/ MON 11900/ PC Code 128931 / Monsanto Company

DACO 6.3/OPPTS 860.1300/OECD II 6.2.2, 6.2.3 & IIIA 8.2, 8.4.1, 8.4.2

Nature of the Residues in Plants

– Dicamba

- Tolerant Cotton (Event MON 88701)

D40 838 4/MRID No. 48728701 Page 23 of 24 APPENDIX I. Chemical Names and Structures of Reference Standards Used in Cotton Metabolism Study (cont.).

### Dicamba/ MON 11900/ PC Code 128931 / Monsanto Company

DACO 6.3/OPPTS 860.1300/OECD II 6.2.2, 6.2.3 & IIIA 8.2, 8.4.1, 8.4.2

Nature of the Residues in Plants

– Dicamba

- Tolerant Cotton (Event MON 88701)

D40 838 4/MRID No. 48728701 Page 24 of 24 APPENDIX I. Chemical Names and Structures of Reference Standards Used in Cotton Metabolism Study (cont.).**:-:..d I**  

### Dicacmoba/MON 1 1900 / PC 

### Co
1 de I 

### 28S93 

### 8 1

6 

## r
0 Mons
0 

### a 1



### n 

### 0 to Ccompany
2 5 

### 4 2 6 

### d
4 ,, "·"·

... . DA 7.2.1

, 7.2.2

, and 7.2.3 OPPT .134 E D llA 4

.. , .. an

. .J 

### Residue Analytical M ethod

- Cotton



### Primary Evaluato r

Versar. Inc.

,? 

### Approved by

## ("**~~1 ~** 

## A'~~~ Pet e 1 :-

Savoia, Chemist, OCSPP-HED-RAB 

### VNII Date: 11/15 / 2012

Date: 04 / 22 / 20 13

Note: Thi s 

### Dat a Eva luation Re co rd 

### ( D E R ) was originall y prep a red under contract b y Ver sar

. Inc. ( 685 0 Vcr sa r Ce nter. Sprin g fi e ld. VA 22151: 

### submiucd 11 11 5 11 2 )

. 

### The DER h as 

### been re v iewed b y 

### ll E D a nd re v i s ed 

### to n:nc c t c urr e nt Offic e of

Pe s ticide Pr og ram s ( OPP ) p o licie s.



### S T U DY R E POR TS:

48728702 Foster, 

### J. ; Mierkowski

, M. ; Miller. M. (2011) Amended Report for MSL0023267: Analytical Method for the D e c erm inati on o f Dicamba and it s Major Metabolites in Cotton

Matrices by LC/MS/MS. Project Numbe r : AG/ME / 1 381101/0CR

, MSL0023759. Unpublished

s tudy prepared by Monsanto Co mpany. 93p.



### EXECUT I VE SUM M A R Y:

Monsanto has su bmitt ed an analytical method description and validation data for a data

collection method, AG-ME-138 1-01

, for the determination of residues of dicamba and its

metabolites

, 5-hydroxydicamba (5 -0H dicamba), 3,6

- dichloro sa lic yl ic acid (OCSA) and 3,6- dichlorog e nti s ic ac i d (DCGA) in cotton matrice s using hi g h performance liquid chromatography

with trip l e-quadro ple mass spec trometric (LC-MS / MS) analysis. The method was u sed for data

collection in the cotton crop field trial, processing, and storage stability studies reviewed under

DP # 404821.

Briefly, samp l es were extracted wit h acetonitrile:water (40:60, v:v)

. An aliquot of the extract

was hydrolyz e d in 1 N H C l at 95 °C in an oven. The hydroly sa te was partition ed with ethyl

acetate:isooctane (20 :80

, v:v). Water was added to the organic phase and the sam ple was
concentrated by evaporation until only the aqueous solut i on remained. The samples were
acidified and analyzed using LC/MS/MS. Stable -label ed internal standards for each analyte

(' 3 

### C 6 -Di carnba

, 13 

### C 6 -5-Hydroxydicamba

, 13 

### C 6 -DCSA, and 13 

### C 6 -DCGA) were used to normalize

for matrix effects and procedural recovery. The method monitors a single ion transition for eac h analyte.

The limit of quantitation (LOQ)

, determined as the lowest limit of method validation (L LMV ).

for dicamba. 5 -0H dicamba

, DCSA and DCGA was 0.02 ppm fo r each analyte in undelinted

cotton seed, meal, hulls

, and alkali refin ed oi l ; and 0.04 ppm for gin byproducts. The limit of

detection (LOO) in cotton seed, statistically determined from the LLMV, was 0.0089, 0.0057

, 0.0035

, and 0.0041 ppm fo r dicamba, 5-0H dicamba, DCSA and DCGA

, re spec ti ve l y. LODs

were not determined for co tt on gi n byproduct o r processed commodities.

The method was successfully validated using samples of undelinted cotton seed

, meal, hulls

, and

oil fortified with each ana l yte at 0.02, 0.20

, and I 0 ppm; and cotton g in byproducts fort ified with

0408384 / MRID No. 48728702 P age I oflO

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.2.1, 7.2.2, and 7.2.3/OPPTS 860.1340/OECD IIA 4.2.5, 4.2.6 and 4.3 Residue Analytical Method

- Cotton

D408 384 /MRID No. 487 2 8702 Page 2 of 10 

## each analyte at 0.04, 0.40, and 10 ppm

. M ean recoveries were within the acceptable range of

70% to 120% with the coefficient of variation less than 20% for each matrix tested

.

No confirmatory analysis procedures were included for the method, and no interfere nce study

was submitted ; however, because the method has not been proposed for enforcement, confirmatory analysis procedures are not required.

An independent laboratory validation (ILV) was not submitted and is not required in support of a

data collection method.

Monsanto submitted radiovalidation data to evaluate the efficiency of the extraction methodology presented in Method AG

- ME

- 1381

- 01. Average extractabilities from gin byproduct and cotton seed using Method AG

- ME

- 1381

- 01 extraction procedures (6 7.07% and 24.08%), were similar to normalized extractabilities obtained from gin byproduct and cotton seed samples in the metabolism study (MRID 48728701) using multiple acetonitrile:water extractions (67.76% and 24.97%). Additionally, residue levels of d icamba, DCGA, and DCSA in the acidic

post

- evaporation solutions obtained using LC/MS/MS method AG

- ME

- 1381

- 01, were similar to the residue levels in the metabolism samples as determined by 14 

## C

- HPLC profiles of hydrolyzed

extracts. The extraction efficienci es for dicamba, DCGA, and DCSA, respectively, as determined by the reviewer, were 106%, 101%, and 121% in the undelinted seed samples, and 83%, 79%, and 87% in the gin byproduct samples. Monsanto also investigated the recovery of radioactivity through the post

- extraction steps of method AG

- ME

- 1381

- 01. Overall post

extraction recovery was 31.31% in undelinted cotton seed and 50.20% in cotton gin byproducts. The majority of the loss was in the partitioning step.

STUDY/WAIVER ACCEPTABILITY/DEFICIENCIES/CLARIFICATIONS :

Under the conditions and parameters used in the study, analytical method test data are classified

as scientif ically acceptable

. The acceptability of this study for regulatory purposes is addressed

in the forthco ming U.S. EPA Residue Chemistry Summary Document D408384

.

COMPLIANCE :

Signed and dated Good Laboratory Practice (GLP), Quality Assurance and Data Confidentiality

statements were provided. No deviations from regulatory requirements were reported which

wo uld have an impact on the validity of the study.

A. BACKGROUND INFORMATION

Dicamba (3,6

- dichloro

- 2

- methoxybenzoic acid ) is a selective benzoic acid herbicide registered for the control of weeds prior to their emergence. The d icamba Reregistration Eligibility Decision (RED) was issued December 2005. The chemical structure and nomenclature of

dicamba and its metabolites 5

- OH dicamba, DSCA, and DCGA are presented in Table A.1. The physicochemical properties of the technical grade of dicamba acid ar e presented in Table A.2.

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.2.1, 7.2.2, and 7.2.3/OPPTS 860.1340/OECD IIA 4.2.5, 4.2.6 and 4.3 Residue Analytical Method

- Cotton

D408 384 /MRID No. 487 2 8702 Page 3 of 10 TABLE A.1. Test Compound Nomenclature.

Compound

Common name Dicamba

Company experimental name MON 11900

IUPAC name 3,6

- dichloro

- o

- anisic acid or 3,6

- dichloro

- 2

- methoxybenzoic acid

CAS name 3,6

- dichloro

- 2

- methoxybenzoic acid

CAS registry number 1918

- 00

- 9 (dicamba acid), 104040

- 79

- 1 (diglycolamine salt), or 53404

- 28

- 7 (monoethanolamine salt)

End

- use product Not applicable

Compound

Common name 5

- Hydroxy

- dicamba
Company experimental name 5

- OH dicamba

IUPAC/CAS name 2,5

- dichloro

- 3

- hydroxy

- 6

- methoxybenzoic acid

CAS registry number 7600

- 50

- 2

Compound

Common name DCSA ; 3,6

- dichlorosalicylic acid

Company experimental name MON 52708

IUPAC/CAS name 3,6

- dichloro

- 2

- hydroxybenzoic acid

CAS registry number 3401

- 80

- 7

Compound

Common name DCGA ; 3,6

- dichlorogentistic acid

Company experimental name MON 52724

IUPAC/CAS name 2,5

- dichloro

- 3,6

- dihydroxybenzoic acid

CAS registry number 18688

- 01

- 2

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.2.1, 7.2.2, and 7.2.3/OPPTS 860.1340/OECD IIA 4.2.5, 4.2.6 and 4.3 Residue Analytical Method

- Cotton

D408 384 /MRID No. 487 2 8702 Page 4 of 10 TABLE A.2. Physicochemical Properties of the Technical Grade Test Compound: Dicamba.

Parameter Value Reference

Melting point 114

- 116  C (PAI)

90

- 100  C (87% TGAI) Residue Chemistry Chapter of the Dicamba RED, DP# 317699, 12/20/05, C. Olinger pH 2.5

- 3.0 (87% TGAI)

Density 1.57 g/mL at 25  C (87% TGAI)

Water solubility 0.5 g/100 mL at 25  C (PAI)

Solvent solubility g/100 mL at 25  C (PAI)

dioxane 118.0

ethanol 92.2 isopropyl alcohol 76.0 methylene chloride 26.0

acetone 17.0

toluene 13.0 xylene 7.8 heavy aromatic naphthalene 5.2

Vapor pressure 3.4 x 10

- 5 mm Hg at 25  C (PAI)

Dissociation constant, pK a 1.97 (PAI)

Octanol/water partition coefficient, Log(K OW ) 0.1 (PAI)

UV/visible absorption spectrum neutral: 511 (275 nm) acidic (pH 0

- 1): 1053 (281 nm) basic (pH 13

- 14): 469 (274 nm)



## B. MATERIALS AND METHODS

B.1. Data

- Gathering Method

Monsanto has submitted an analytical method description and validation data for a data collection method, AG

- ME

- 1381

- 01, for the determination of residues of dicamba and its metabolites, 5

- OH dicamba , DCSA and DCGA in cotton matrices using LC / MS/MS analysis.

B.1.1. Principle of the Method:

Briefly, samples were extracted with acetonitrile : water ( 4 0: 6 0, v:v). An aliquot of the extract

was hydrolyzed in 1 N HCl at 95 °C in an oven

. The hydrolysate was partitioned with ethyl acetate:isooctane (20:80, v:v). Water was added to the organic phase and the sample was concentrated by evaporation until only the aqueous solution r emained. The samples were acidified and analyzed using LC/MS/MS. Stable

- labeled internal standards for each analyte ( 13 

## C 6



## - Dicamba, 13 

## C 6



## - 5

- Hydroxydicamba, 13 

## C 6



## - DCSA, and 13 

## C 6



## - DCGA) were used to normalize for matrix effects and procedural recovery. The method monitors a single ion transition for each analyte.

TABLE B.1.1. Summary Parameters for the Analytical Meth od Used for the Quantitation of the

Residues of Dicamba and Dicamba Metabolites in Cotton Matrices.

Method ID AG

- ME

- 1381

- 01 Analyte(s) Dicamba, 5

- OH dicamba , DCSA and DC GA Extraction solvent/technique C otton matrices were extracted with acetonitrile (ACN)/water (40:60, v:v).

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.2.1, 7.2.2, and 7.2.3/OPPTS 860.1340/OECD IIA 4.2.5, 4.2.6 and 4.3 Residue Analytical Method

- Cotton

D408 384 /MRID No. 487 2 8702 Page 5 of 10 TABLE B.1.1. Summary Parameters for the Analytical Meth od Used for the Quantitation of the

Residues of Dicamba and Dicamba Metabolites in Cotton Matrices.
Cleanup strategies An aliquot of the extract was hydrolyzed in 1 N HCl at 95 °C in an oven. The hydrolysate was partitioned with ethyl acetate:isooctane (20:80, v:v). Water was added to the organic phase and the sample was concentrated by evaporation until only the aqueous solution

remained. The sample was acidified and analyzed by LC/MS/MS.

Instrument/Det ector LC/MS/MS using a Betasil Phenyl column. Quantitation of residues is performed using

MS/MS t urbo ion spray i on ization in the negative ion MRM mode

. The following ions are monitored:

Dicamba: 219 →175 m/z

5

- OH dicamba: 237 →193 m/z

DCSA: 205 →161 m/z

DCGA: 221 →177 m/z

Internal Standards

13 C 6

- Dicamba: 225 →181 m/z

13 C 6

- 5

- Hydroxydicamba: 243 →199 m/z

13 C 6

- DCSA: 211 →167 m/z

13 C 6

- DCGA: 227 →183 m/z

Standardization method Internal ( 13 C 6

- Dicamba, 13 C 6

- 5

- Hydroxydicamba, 13 C 6

- DCSA, and 13 C 6

- DCGA) and external

standardization using mixed standards was used . A calibration curve is constructed where the

p eak area is plotted versus the analyte concentration using linear regression and a correlation coefficient of 0.990 or greater is obtained. Stability of std sol utions Stock solutions (100 μg/ml) were reported to be stable for at least 201 days. Calibration standards (0.005 to 0.500 μg/mL) were reported to be stable for dicamba, 5

- OH

dicamba and DCSA at the 0.005 μg/mL concentration and above for at least 175 days and for DCGA at the 0.010 μg/mL concentration and above for at least 32 days

. Retention times (minutes) Dicamba: 2.7

5

- OH dicamba : 1.3

DCSA: 5.6

DCGA: 1.7



## B.2. Enforcement Method

A gas chromatography method with electron capture detection (GC/ECD) is the current enforcement method for residues of dicamba in/on plant commodities (Method AM

- 0691B

0297

- 4).

C. RESULTS AND DISCUSSION

C.1. Data

- Gathering Method

Method validation data are presented in Table C.1.1. Acceptable method validation recoveries

for method AG

- ME

- 1381

- 01 were obtained for all analytes from samples undelinted cotton seed

at 0.0 2, 0. 20, and 10 ppm; cotton meal, hulls, and oil at 0.0 2 and 0. 20 ppm ; and cotton gin

b yproducts at 0.04, 0.40, and 10 ppm

. M ean recoveries were within the acceptable range of 70%

to 120% with the coefficient of variation less than 20% for each matrix tested . Apparent residues of dicamba and its metabolites, 5

- OH dicamba, DCSA and DCGA wer e each < LLMV in/on two samples each of seed (<0.02 ppm), gin byproducts (<0.04 ppm), meal, hulls, and alkali refined oil (<0.02 ppm). The fortification levels used in method validation are adequate to bracket expected residue levels in/on cotton matrices

.

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.2.1, 7.2.2, and 7.2.3/OPPTS 860.1340/OECD IIA 4.2.5, 4.2.6 and 4.3 Residue Analytical Method

- Cotton

D408 384 /MRID No. 487 2 8702 Page 6 of 10 

## The method characteristics for AG

- ME

- 1381

- 01 are summarized in Table C.1.2. The LOQ,

determined as the LLMV, was 0.02 ppm for each analyte in undelinted cotton seed, meal, hulls, and alkali refined oil; and 0.04 ppm for each analyte in gin byproducts. The LOD in cotton seed, statistically determine d from the LLMV, was 0.0089, 0.0057, 0.0035, and 0.0041 ppm for dicamba, 5

- OH dicamba, DCSA and DCGA, respectively. LODs were not determined for cotton gin byproduct or processed commodit ies.

No confirmatory analysis procedures were included for the method, and no interference study was submitted; however, because the method has not been proposed for enforcement, confirmatory analysis procedures are not required.

Monsanto also submitted radiovalidation data. In the first part of the study, Monsanto compared the amount of radioactivity extracted from cotton seed and gin byproduct samples obtained from the metabolism study (MRID 48728701) using multiple acetonitrile: water extractio ns , to t he amount of radioactivity extracted using the analytical method AG

- ME

- 1381

- 01 extraction procedures. The extractabilities were similar : for gin byproducts and undelinted cotton seed

, respectively, normalized extractabilities from the metabolism study wer e 67.76% and 24.97% and average extractabilities from method AG

- ME

- 1381

- 01 were 67.07% and 24.08%.

In the second part of the radiovalidation study, Monsanto investigated the recovery of radioactivity through the post

- extraction steps of method AG

- ME

- 1381

- 01. Overall post

extraction recovery was 31.31% in undelinted cotton seed and 50.20% in cotton gin byproducts. The majority of the loss was in the partitioning step. Average percent recoveries for the hydrolysis, organic partition, aqueous partition, an d acidic post

- evaporation steps, respectively, were 87.06%, 57.21%, 62.98%, and 62.87% in cotton seed and 82.65%, 67.98%, 39.10%, and

89 .34% in gin byproducts. The study notes that losses in the partitioning step may be due to 1) water solubility of a per centage of the residues (e.g., polar residues formed by degradation of dicamba to small molecules and incorporation into natural products such as sugars), especially for seed, and, 2) incomplete partitioning of the analytes into the organic phase. The stu dy also

notes that the use of internal standards in this method compensates for any procedural losses incurred in the hydrolysis, partitioning, and evaporation steps.

In the third part of the radiovalidation study Monsanto compared the residue quantitation results

of the acidic post

- evaporation solutions obtained using LC/MS/MS method AG

- ME

- 1381

- 01

, with the actual residue levels in the metabolism samples as determined by 14 

## C

- HPLC profiles of hydrolyzed extracts (MRID 48728701)

. These results a re presented in Table C.1.3. The

extraction efficiencies for dicamba, DCGA, and DCSA, respectively, were determined by the reviewer to be 106%, 101%, and 121% in the undelinted seed samples

, and 83%, 79%, and 87% in the gin byproduct samples.

TABLE C. 1. 1 Recovery Res ults from Method Validation of Cotton Matrices using the Data

- Gathering

Analytical Method. Standards were prepared in acetonitrile.

Matrix Spiking Level (ppm) Recoveries Obtained 1
(%) Mean Recovery ± SD (CV) 1
(%)

Dicamba

Cotton seed 0.005 187, 73, 172, 105, 102 128 ± 49 ( 38)

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.2.1, 7.2.2, and 7.2.3/OPPTS 860.1340/OECD IIA 4.2.5, 4.2.6 and 4.3 Residue Analytical Method

- Cotton

D408 384 /MRID No. 487 2 8702 Page 7 of 10 TABLE C. 1. 1 Recovery Res ults from Method Validation of Cotton Matrices using the Data

- Gathering

Analytical Method. Standards were prepared in acetonitrile.

Matrix Spiking Level (ppm) Recoveries Obtained 1
(%) Mean Recovery ± SD (CV) 1
(%)

0.01 129, 149, 93, 78, 125 115 ± 29 (25)

0.02 119, 100, 123, 101 , 124 113 ± 12 (10 )

0.20 94.0, 96.0, 94.5, 96.5, 99.5 96.1 ± 2.2 (2.2)

10 68.3, 90.5, 90.6, 100, 98.4 89.6 ± 13 (14 ) Cotton gin byproducts 0.04 99.8, 92.5, 88.0, 122, 119 104 ± 15 ( 15 )

0.40 97.0, 98.5, 101, 97.5, 104 99.5 ± 2.7 (2.7)

10 87.8, 90.5, 82.4, 85.1, 87.8 86.7 ± 3.1 (3.5)

Cotton meal 0.02 102, 101, 98.0, 111, 108 104 ± 5.3 (5.1)

0.20 96.5, 98.5, 95.5, 100, 94.0 96.9 ± 2.4 (2.5)

Cotton hulls 0.02 96.5, 100, 106, 99.0, 98.5 100 ± 3.4 (3.4)

0.20 96.5, 101, 102, 100, 99.0 100 ± 2.1 (2.1)

Cotton alkali refined oil 0.02 99.0, 96.0, 97.0, 98.5, 97.5 97.6 ± 1.2 (1.2)

0.20 99.0, 100, 105, 98.5, 97.0 100 ± 2.8 (2.9)

5

- OH dicamba

Cotton seed 0.005 68, 56, 101, 84, 105 82.9 ± 21 (25)

0.01 106, 97, 76, 110, 82 94.1 ± 15 (16)

0.02 96.0, 97.5, 115, 117, 114 108 ± 10 (9.3)

0.20 97.5, 104, 105, 104, 101 102 ± 3 .0 (2.9)

10 69.1, 89.6, 79.4, 93.2, 104 87.1 ± 13 (15 ) Cotton gin byproducts 0.04 124, 106, 98.0, 96.5, 108 106 ± 11 (10 )

0.40 111, 113, 106, 116, 109 111 ± 3.6 (3.3)

10 113, 98.7, 99.4, 101, 84.7 99.4 ± 10 (10 )

Cotton meal 0.02 72.0, 108, 92.0, 77.0, 91.5 88.1 ± 14 (16 )

0.20 98.0, 93.5, 87.5, 98.5, 89.5 93.4 ± 4.9 (5.3)

Cotton hulls 0.02 86.5, 78.0, 106, 89.0, 97.0 91.2 ± 10 (11 )

0.20 114, 107, 117, 98.5, 105 108 ± 7.4 (6.8)

Cotton alkali refined oil 0.02 94.5, 101, 79.0, 92.0, 88.0 90.8 ± 8.0 (8.8)

0.20 110, 103, 88.0, 110, 100 102 ± 9.0 (8.8)

DCGA

Cotton seed 0.005 140, 115, 88, 98, 129 114 ± 22 (19)

0.01 90, 108, 96, 79, 104 95.6 ± 11 (12)

0.02 81.0, 98.0, 92.5, 77.0, 86.0 86.9 ± 8.5 (9.8)

0.20 90.0, 87.0, 88.0, 84.0, 95.5 88.9 ± 4.3 (4.8)

10 73.4, 95.7, 89.2, 94.4, 90.4 88.6 ± 8.9 (10 ) Cotton gin byproducts 0.04 66.8, 87.5, 88.3, 89.8, 69.5 80.4 ± 11 (14 )

0.40 99.3, 95.5, 99.0, 94.8, 96.5 97.0 ± 2.0 (2.1)

10 101, 94.1, 105, 98.9, 104 101 ± 4.4 (4.3)

Cotton meal 0.02 98.5, 108, 102, 103, 102 102 ± 3.3 (3.2)

0.20 87.5, 86.0, 84.5, 87.5, 84.5 86.0 ± 1.5 (1.7)

Cotton hulls 0.02 116, 117, 115, 121, 112 116 ± 3.4 (2.9)

0.20 88.5, 86.0, 85.5, 84.0, 82.0 85.2 ± 2.4 (2.8)

Cotton alkali refined oil 0.02 100, 102, 98.5, 103, 113 103 ± 5.7 (5.5)

0.20 96.5, 89.0, 91.5, 91.5, 90.5 91.8 ± 2.8 (3.1)

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.2.1, 7.2.2, and 7.2.3/OPPTS 860.1340/OECD IIA 4.2.5, 4.2.6 and 4.3 Residue Analytical Method

- Cotton

D408 384 /MRID No. 487 2 8702 Page 8 of 10 TABLE C. 1. 1 Recovery Res ults from Method Validation of Cotton Matrices using the Data

- Gathering

Analytical Method. Standards were prepared in acetonitrile.

Matrix Spiking Level (ppm) Recoveries Obtained 1
(%) Mean Recovery ± SD (CV) 1
(%)

DCSA

Cotton seed 0.005 88, 95, 90, 134, 98 101 ± 19 (19)

0.01 70, 84, 102, 96, 98 89.8 ± 13 (15)

0.02 99.5, 82.0, 97.0, 97.0, 94.5 94.0 ± 6.9 (7.4)

0.20 94.5, 99.5, 105, 95.5, 101 98.9 ± 4.0 (4.1)

10 80.7, 92.1, 95.1, 93.1, 96.9 91.6 ± 6.4 (6.9) Cotton gin byproducts 0.04 79.0, 87.3, 75.5, 75.3, 80.5 79.5 ± 4.9 (6.1 )

0.40 90.5, 83.5, 92.8, 87.0, 91.0 89.0 ± 3.7 (4.2)

10 110, 101, 91.8, 109, 98.8 102 ± 7.6 (7.4)

Cotton meal 0.02 101, 95.5, 100, 89.5, 97.5 96.5 ± 4.4 (4.5)

0.20 82.0, 83.5, 85.5, 91.0, 89.0 86.2 ± 3.8 (4.4)

Cotton hulls 0.02 109, 102, 93.5, 97.0, 96.0 100 ± 6.1 (6.2)

0.20 95.5, 102, 91.5, 100, 99.0 97.4 ± 3.9 (4.0)

Cotton alkali refined oil 0.02 102, 93.0, 89.5, 92.5, 100 95.2 ± 5.1 (5.3)

0.20 103, 106, 100, 104, 105 103 ± 2.1 (2.0)

1 As calculated by the reviewer.



### TABLE C.1. 2 Characteristics for the Data

- Gathering Analytical Method Used for the Quantitation of

Dicamba and Dicamba Metabolites in Cotton Matrices

. Analyte(s) Dicamba, 5

- OH dicamba, DCSA and DCGA Equipment ID Shimadzu Prominence HPLC equipped with a Betasil Phenyl HPLC column (50 mm x 3

mm x 3 μm). Quantitation of residues is performed using a n Applied Biosystems API 5500

MS/MS using turbo ion spray ionizatio n operated in the negati ve

- ion multiple reaction monitoring (MRM) mode. Limit of quantitation (LOQ) The LOQs, determined as the LLMV, were 0.0 2 ppm for each analyte in cotton seed, meal, hulls, and oil; and 0.04 ppm in gin by products.

C alculated LOQ s were determined for cotton seed based on replicate determinations at the LLMV. The LOQ s were 0.0266 ppm for dicamba, 0.0171 ppm for 5

- OH dicamba, 0.0105 ppm for DCSA, and 0.0121 ppm DCGA.
Limit of detection (LOD) The LOD s, defined as the t 0.99 confidence interval times the standard deviation of the fortified replicates at the LLMV, were determined for cotton seed. The LODs were 0.0089

ppm for dicamba, 0.0057 ppm for 5

- OH dicamba, 0.0035 ppm for DCSA and 0.0041 ppm

for DCGA. Accuracy/Precision Percent recoveries and coefficients of variance (CVs) indicate acceptable accuracy/precision at 0.02, 0.20, and 10 ppm for d icamb a and its metabolites in seed; at 0.02 and 0.20 ppm in meal, hulls, and oil; and at 0.04, 0.40, and 10 ppm for gin byproducts

. Average recovery ranges (and CVs) were 86.7

- 113% (1.22

- 14.6) for dicamba, 87.1

- 111% ( 2.90

- 16.1 ) for 5

- OH dicamba , 79.5

- 103% (2.01

- 7.40) for DCSA, and 80.4

- 116 (1.74

- 14.0) for DC G A. See

Table C.1.1 above. Reliability of the Method/ [ILV] An independent laboratory method validation [ILV] was not conducted. Linearity The method/detector response was linear (coefficient of determination, r 2 = ≥0.99 ) within the range of 0.005

- 0.50 μg/mL.

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.2.1, 7.2.2, and 7.2.3/OPPTS 860.1340/OECD IIA 4.2.5, 4.2.6 and 4.3 Residue Analytical Method

- Cotton

D408 384 /MRID No. 487 2 8702 Page 9 of 10 TABLE C.1. 2 Characteristics for the Data

- Gathering Analytical Method Used for the Quantitation of

Dicamba and Dicamba Metabolites in Cotton Matrices

. Specificity The control chromatograms generally have no peaks above the chromatographic background and the spiked sample chromatograms contain only the analyte peak of interest. In cotton gin byproducts, however, DCGA and 5

- OH dicamba each had matri x interference for at

least one lot that exceeded 30% of the LLMV. The study notes that chromatographic interference may be a problem in some matrice s . Gradient conditions and use of MS with MRM analysis greatly reduces the interferences for these samples . The study also notes that there are no interferences from 2,4

- dichlorophenoxyacetic acid or 2,4

- dichlorophenylacetic acid, which have the sam e mass transitions as dicamba an d DCSA, respectively. Interferences from other pesticides are unknown, however, none are expected due to the high level of specificity of the LC/MS/MS analysis.

Peaks were well defined and symmetrical. There appeared to be no carryover to the following chromatograms.

Table C.1.3. Extraction Efficiency of Method AG

- ME

- 1381

- 01 As Compared to the Metabolism Study Extraction

Method

Matrix Analyte Extraction Method Radioactive Residues (ppm) [Average] Extraction Efficiency 1
(%)

Seed Dicamba Metabolism study 0.0102 106

Method AG

- ME

- 1381

- 01 0.0098, 0.0119 [0.0108]

DCGA Metabolism study 0.0224 101

Method AG

- ME

- 1381

- 01 0.0227, 0.0227 [0.0227]

DCSA Metabolism study 0.0532 121

Method AG

- ME

- 1381

- 01 0.0634, 0.0658 [0.0646]

Gin Byproduct Dicamba Metabolism study 2.83 83

Method AG

- ME

- 1381

- 01 2.31, 2.37 [2.34]

DCGA Metabolism study 4.63 79

Method AG

- ME

- 1381

- 01 3.82, 3.50 [3.66]

DCSA Metabolism study 16.1 87

Method AG

- ME

- 1381

- 01 13.99, 14.09 [14.04]

1 Extraction efficiency = (residues determined by residue method ÷ residues determined in metabolism study)*100.



## C.2. Enforcement Method

A gas chromatography method with electron capture detection (GC/ECD) is the current

enforcement method for residues of dicamba in/on plant commodities (Method AM

- 0691B

0297

- 4).

C.3. Independent Laboratory Validation

An independent laboratory validation (ILV) study was not included as a part of this submission; however, no additional information is required because ILV is not g enerally required for data

collection methods.

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.2.1, 7.2.2, and 7.2.3/OPPTS 860.1340/OECD IIA 4.2.5, 4.2.6 and 4.3 Residue Analytical Method

- Cotton

D408 384 /MRID No. 487 2 8702 Page 10 of 10 

## D. CONCLUSION

Adequate method validation data have been submitted for the a nalytical m ethod AG

- M E

- 1381

- 01 for the determination of residues of dicamba, 5

- OH dicamba , DCSA and DCGA in cotton

commodities. The method has been adequately validated with accuracy and precision over the

range of 0.0 2 to 10 ppm for cotton seed ; 0.0 2 to 0.20 ppm for meal, hulls, and oil , and 0.04 to 10 ppm for cotton gin byproducts

. M ean recoveries were within acceptable range of 70% to 120%

with the coefficient of variation less than 20% for each matrix tested

. The data are sufficiently representative of the expected residue levels in cotton seed, gin byproducts , meal, hulls, and oil

. Acceptable radiovalidation data were submitted.

No confirmatory analysis procedures were included for the method, and no interference study was submitted; however, because the method has not been proposed for enforcement, confirmatory analysis procedur es are not required.

An ILV was not submitted and is not required in support of a data collection method.

E. REFERENCES

DP No.: D317699 Subject: Dicamba. Residue Chemistry Considerations for the Reregistration Eligibility

Decision (RED) Document. S ummary of Analytical Chemistry and Residue Data.

From: C. L. Olinger

To: K. Tyler

Date: 12 / 20 / 2005

MRIDs: None

F. DOCUMENT TRACKING

Petition Number(s): 2F8067 DP Barcode(s): 408384

PC Code: 029801, 029802, 029806, 128931, 128944, & 129043

Template Version September 2003

### Dicarnb a/ MON 1 1 900 / PC Code 1 2 8931 / M onsanto Co mpany

# r¥~**··** _::::. I

- 

# · - OA CO 7.4. 1 /7.4.2 / 0PPTS 860.1500 / 0ECD ll A 6.3.1

, 6.3.2, 6.3.3 a nd JllA 8.3

. 1

, 8. 3.2

, 8 .3 .3 Crop F i e ld Trial / R e s idue Decline

- Dicamba-Tolerant Conon (Event MO 8870 I)



### Primary Evaluator D ate: 11115 / 2012

Versar

, I nc.
,7 

### Approved b y 

# /**.rr- ~** **A,....,.-,,~ ;......___./**  

### Date: 04 / 22 / 20 13

_.-" 

### Pe te r Savoia, Chemi s t, OCSPP- H ED-RAB VN i f

Thi s DER was o ri ginall y prepared under contrac t by Vcr s ar. In c. (6850 Vcr s ar Center. Sprin g fidd. VA 22 1 5 1 : s ubmiued ( 11115 / 20 12 ). The D E R h as been reviewed b y I-JED and revi s ed to renec1 current Office o f Pe s ticide Pro g ram s ( OPP ) policie s.



### S T U DY REPORT:

48728703 Mah e r
, D. ; Fo s ter

, 

### J. (2011) Ma g nitude of Dicamba Re s idue s in Cotton Raw Agricultural and Processed Commodities following Applications o f Dicamba-Based

Formulat i ons to MON 88701. Project N umber: REG / I 0 / 062 / 0CR

, MSL002407 2. U npubli s h e d study prepared by Mo n santo Company, Ca rr inge r s, I nc

. a n d GLP Tec h no l ogie s

. 603 p

.



### E X ECU TIV E SU MMAR Y:

Mon s anto ha s s ubmitted fie l d t r ial data for dicamba on d i camba-tolerant cotton

, identified a s
Eve n t MON 8870 1 (forme rl y GH

_ S26695). Event MON 88701 cotton expresses a modified

dic a mba mono- oxyge nase ( DMO ) gene derived from 

### S t enotrophomonas malt ophi lia and the bialaphos re s i s t ance ( B AR) ge ne iso l ated from 

### Streptomyces hygroscopicus. The exp res s i o n of

DMO confers dicamba tolerance and th e ex pression o f BAR confers g lufosinat e tolerance to

Ev e nt MON 8870 I cotton

. Th i rtee n field tr i als were cond u cted in the Un i ted Sta t es during t h e
20 1 0 growi n g seaso n in the North American Free Trad e Agreement (NA FTA) Growing Zon es 2
(GA a nd SC; 2 trial s), 4 ( AR. LA

, and MO: 3 trials). 6 (TX; l trial )

, 8 (O K and TX : 5 trials). a nd

1 0 (CA; 2 trial s).

Eac h tria l con s i s t e d of one u ntreated and three to four t r ea ted plots. Eac h treated p l ot fo ll owed

one of three treatment types: three broadca s t applications applied at pre-emergence. 6-leaf, and

1 5 1 

### white flo we r + 15 da ys [TRT 2]; three broadcast applications applied at pre-emergence
, fir s t

open boll, and 7-days prior t o harvest [T R T 3]; and four foliar broadca s t applications applied at

6-l eaf

, l s t 

### white flower + 15 days, fir s t open bo ll

, and 7 days prior to harvest [ T R Ts 4 and 5]. Applications to p l ots 2, 3

, and 4 were mad e u s i ng a 4 lb ae / gal wat e r so lub l e concentrate (SL)

formu l at ion of dicamba di g l yco lamine sa lt (C larit y Herb i cide; MO N 54 140 ); plot 5 applications

were made u si n g a 5 lb a e / ga l S L formulation of dicamb a monoethan o lamine sa lt (M ON 11 96 8 )

to provide bridg i ng data. Th e tota l application rate fo r a ll treatmen t types was 2.0-2.l l b ae / A
(2.2-2.4 kg ae / ha). For T RT 2, app li cation I was app l i e d at 0

. 98-1.0 l b ae / A (1. 1 kg ae/ha) and appl i cations 2 and 3 were applied at 0.49- 0.5 l lb a e / Napp lication (0.55 -0.57 kg

ae / h a/app licati on). Retreatm e nt interval s ( RTis) were 24 -4 9 da ys between the fi r s t and seco nd applications and 36 -49 day s between th e seco nd and thi r d applications. For TRT 3

. app li cation 1

was app li ed at 0.99-1

. 0 lb ae / A ( 1 .1 kg ae/ha) and app li cations 2 and 3 were app l ied at 0.49-0

. 52

lb ae / N applic at i o n ( 0.55-0.58 kg ae / h a/ap plication ), for a total se a so nal rate of 2.0 l b ae / A (2.2 kg ae / ha ). RT l s we re 97-178 d ays between the fir s t and seco nd applications and 5 -63 days

between the second and third a pplication s

. For TRT s 4 a nd 5

, applic a tions were applied at 0.49-

0408 384 f MRID No. 4872 870 3 P age I of28

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.4.1/7.4.2/OPPTS 860.1500/OECD IIA 6.3.1, 6.3.2, 6.3.3 and IIIA 8.3.1, 8.3.2, 8.3.3 Crop Field T rial/ Residue Decline

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D40 838 4 /MRID No. 48 728703 Page 2 of 28 

## 0.52 lb ae/A/application ( 0.55

- 0.58 kg ae/ha/application)

. RTIs were 36

- 49 days between th e first and second applications, 32

- 88 days between the second and third applications , and 5

- 63 days between the third and fourth applications

. A summary of the use patterns is provided in the

table below.

TRT Number

of Sites EP Growth Stages/ Target Application Rates lb ae/A (kg ae/ha)

Pre

- emergence 6 Leaf 1 st Wh ite Flower + 15 days First Open

Boll 7 Days

Prior to

Harvest

1 13

-- -- -- -- -- -- 2 13 4 lb ae/gal SL 1.0 (1.1) 0.50 (0.56) 0.50 (0.56)

-- -- 3 13 4 lb ae/gal SL 1.0 (1.1)

-- -- 0.50 (0.56) 0.50 (0.56)

4 13 4 lb ae/gal SL

-- 0.50 (0.56) 0.50 (0.56) 0 .50 (0.56) 0.50 (0.56)

5 4 5 lb ae/gal SL

-- 0.50 (0.56) 0.50 (0.56) 0.50 (0.56) 0.50 (0.56)



## Applications for all treatments were made using ground equipment (backpack, hand

- held and

tractor

- mounted sprayers) in spray volumes of 18.8

- 21.6 gal/A (176

- 202 l/ha) and included an nonionic surfactant ( NIS ) and ammonium sulfate ( AMS ) at all trial sites.

Undelinted cotton seed was collected mechanically with a cotton picker (n=3) or a cotton stripper (n=3) or by hand (n=7) at normal harvest : 49

- 105

- day prehar vest interval ( PHI ) for TRT 2 and 6

- 8

- day PHI for TRTs 3

- 5

. Additional samples of cotton seed were collected from two trials at

PHIs of 43/99, 49/105, 56/112, and 63/119 days for TRT 2 and 1, 7, 14, and 21 days for TRT 4

to assess residue decline. Cotton gin byproduct samples were collected from three sites using a cotton stripper at normal harvest (82

- 84

- day PHI for TRT 2; 6

- 7

- day PHI for TRT 4)

.

Samples were analyzed for residues of dicamba and metabolite s 5

- hydroxydicamba (5

- OH dicamba), 3,6

- dichlor osalicylic acid (DCSA), and 3,6

- dichlorogentisic acid (DCGA) using a high performance liquid chromatography method with tandem mass spectrometry detection (LC /MS/MS). The limit of quantitation (LOQ, equivalent to the lowest level of method validation; LLM V) was 0.0 2 ppm for all analytes in undelinted cotton seed and 0.04 ppm for all analytes in cotton gin byproducts . The method was adequate for data validation based on acceptable method and concurrent recovery data. The fortification levels used in concu rrent method recovery were adequate to bracket expected residue levels. Concurrent recoveries were corrected for apparent residues in controls; residues in treated samples were not corrected for

residues in controls. We note that the analytical method do es not specify conversion of metabolite residues to parent equivalents; therefore, quantifiable residues of 5

- OH dicamba, DCSA, and DCGA were converted to parent equivalents by the study reviewer using a molecular weight conversion factor of 0.933, 1.068, and 0.991, respectively.

Dicamba, 5

- OH dicamba, and DCSA undelinted cotton seed s amples were stored for 13

- 146 days (0.4

- 4.8 months) from collection to analysis; DCGA undelinted cotton seed samples were

stored for 13

- 285 days (0.4

- 9.4 months). Gin byprod uct samples for all analytes were stored

from 17

- 27 days (0.6

- 0.9 months) from collection to analysis

. All samples were analyzed within

0 to 2 day s of extraction. Samples were stored be low freezing at the field sites and at

- 20 °C at the analytical labor atory prior to extraction. A storage stability study for dicamba, DCGA , and

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.4.1/7.4.2/OPPTS 860.1500/OECD IIA 6.3.1, 6.3.2, 6.3.3 and IIIA 8.3.1, 8.3.2, 8.3.3 Crop Field T rial/ Residue Decline

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D40 838 4 /MRID No. 48 728703 Page 3 of 28 

## DCSA on cotton was submitted in conjunction with the field trial study (refer to the 860.1380 der for MRID 48728704). The stability of 5

- OH dicamba was not determined concurrentl y because it was not found in undelinted cotton seed. These data are acceptable to support the storage conditions and durations of the undelinted cotton seed samples from the submitted field

trials

. Storage stability data for cotton gin byproduct was not provided; however, storage stability data is not required as the samples were analyzed within 30 days of harvest.

Following three broadcast applications of the 4 lb ae/gal SL formulation of dicamba (formulated

as the diglycolamine salt), which were appli ed at pre

- emergence, the 6

- leaf stage, and first white flower + 15 days for a total rate of 2.0 lb ae/A (2.2 kg ae/ha) ( TRT 2 ), combined residues (and per trial averages) of dicamba, 5

- OH dicamba, DCGA, and DCSA (expressed in parent equivalents ) were <0.08

- <0.39 (<0.08

- < 0 .39) ppm in/on undelinted cotton seed collected at PHIs

of 49

- 105 days and <0.67

- <2.34 (<0.73

- <2.33) ppm in/on gin byproducts collected at PHIs of

82

- 84 days

.

Following three broadcast applications of the 4 lb ae/gal SL formulation of dic amba (formulated

as the diglycolamine salt), which were applied at pre

- emergence, the first open boll stage, and 7 days prior to harvest for a total rate of 2.0 lb ae/A (2.2 kg ae/ha) ( TRT 3 ), combined residues (and per trial averages) of dicamba, 5

- OH dic amba, DCGA, and DCSA (expressed in parent equivalents) were <0.12

- <2.24 (<0.13

- <1.66) ppm i n/on undelinted cotton seed collected at a 6

- 8

- day PHI.

Following four foliar broadcast applications of the 4 lb ae/gal SL formulation of dicamba (formulated as the diglycolamine salt)

, which were applied at the 6

- leaf stage, the first white flower + 15 days, the first open boll, and 7 days prior to harvest for a total rate of 2.0

- 2.1 lb ae/A (2.2

- 2.4 kg ae/ha) ( TRT 4 ), combined residues (and per trial averages) of d icamba, 5

- OH dicamba, DCGA, and DCSA (expressed in parent equivalents) were <0.15

- <2.02 (<0.21

- <1.86) ppm in/on undelinted cotton seed collected at a 6

- 8

- day PHI and <5.50

- 34.6 (<5.51

- <33.7) ppm in/on gin byproducts collected at a 6

- 7

- day PHI

. Correspondin g combined residues (and per trial averages) in side

- by

- side trials treated with the 5 lb ae/gal SL formulation of dicamba formulated

as the monoethanolamine salt ( TRT 5 ) were <0.24

- <1.00 (<0.29

- <0.82) ppm in/on undelinted cotton seed. Average combined re sidues in the diglycolamine salt formulation trials were slightly higher (approximately 1.5x) than the corresponding residues in the monoethanolamine

salt formulation trials.

Treatment plots 3 and 4 showed the highest average combined residues (0.76 ppm ), followed closely by treatment plot 5 (average of 0.52 ppm). Treatment plot 2 had the lowest average combined residues (average of 0.12 ppm). Lower residue level s obtained from treatment plot 2

are consistent with the earlier application timings at the se plots

.

For the TRT 2 residue decline samples, average residues of dicamba, 5

- OH dicamba, DCGA,

and DCSA in/on undelinted cotton seed were at or <LOQ in both studies; therefore, residue

decline could not be assessed. For the TRT 4 residue decline sam ples, average residues of

dicamba decreased in/on undelinted cotton seed with increasing PHI. Average residues of

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.4.1/7.4.2/OPPTS 860.1500/OECD IIA 6.3.1, 6.3.2, 6.3.3 and IIIA 8.3.1, 8.3.2, 8.3.3 Crop Field T rial/ Residue Decline

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D40 838 4 /MRID No. 48 728703 Page 4 of 28 

## DCGA and DCSA increased with increasing PHI in the SC1 trial, and remained relatively

constant in the TX1 trial. Residues of 5

- OH dicamba we re <LOQ in both studies; therefore,

residue decline could not be assessed.

STUDY/WAIVER ACCEPTABILITY/DEFICIENCIES/CLARIFICATIONS :

Under the conditions and parameters used in the study, the field trial residue data are classified

as scie ntifically acce ptable

. T he acceptability of this study for regulatory purposes is addressed in the forthcoming U.S. EPA Residue Chemistry Summary Document D 408 384

.

COMPLIANCE :

Signed and dated Good Laboratory Practice (GLP), Quality Assurance

, and D ata Confidentiality

statements were provided. No deviations from regulatory requirements were reported which would have an impact on the validity of the study.

A. BACKGROUND INFORMATION

Dicamba (3,6

- dichloro

- 2

- methoxybenzoic acid ) is a selective benz oic acid herbicide registered for the control of weeds prior to their emergence. The Dicamba Reregistration Eligibility Decision (RED) was issued December 2005. The chemical structure and nomenclature of

dicamba and its metabolites 5

- OH dicamba, DSCA, an d DCGA are presented in Table A.1. The physicochemical properties of the technical grade of dicamba acid are presented in Table A.2.

TABLE A.1. Test Compound Nomenclature.

Compound

Common name Dicamba

Company experimental name MON 11900

IUPAC nam e 3,6

- dichloro

- o

- anisic acid or 3,6

- dichloro

- 2

- methoxybenzoic acid

CAS name 3,6

- dichloro

- 2

- methoxybenzoic acid

CAS registry number 1918

- 00

- 9 (dicamba acid), 104040

- 79

- 1 (diglycolamine salt), or 53404

- 28

- 7 (monoethanolamine salt)

End

- use product Clarity ® Herbicide: SL formulation containing 4 lb ae/gal

MON 11968 : SL formulation containing 5 lb ae/gal

Compound

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.4.1/7.4.2/OPPTS 860.1500/OECD IIA 6.3.1, 6.3.2, 6.3.3 and IIIA 8.3.1, 8.3.2, 8.3.3 Crop Field T rial/ Residue Decline

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D40 838 4 /MRID No. 48 728703 Page 5 of 28 TABLE A.1. Test Compound Nomenclature.

Common name 5

- Hydroxy

- dicamba
Company experimental name 5

- OH dicamba

IUPAC/CAS name 2,5

- dichloro

- 3

- hydroxy

- 6

- methoxybenzoic acid

CAS registr y number 7600

- 50

- 2

Compound

Common name DCSA ; 3,6

- dichlorosalicylic acid

Company experimental name MON 52708

IUPAC/CAS name 3,6

- dichloro

- 2

- hydroxybenzoic acid

CAS registry number 3401

- 80

- 7

Compound

Common name DCGA ; 3,6

- dichlorogentistic a cid

Company experimental name MON 52724

IUPAC/CAS name 2,5

- dichloro

- 3,6

- dihydroxybenzoic acid

CAS registry number 18688

- 01

- 2



### TABLE A.2. Physicochemical Properties of the Technical Grade Test Compound: Dicamba.

Parameter Value Reference

Melting p oint 114

- 116  C (PAI)

90

- 100  C (87% TGAI) Residue Chemistry Chapter of the Dicamba RED, DP# 317699, 12/20/05, C. Olinger pH 2.5

- 3.0 (87% TGAI)

Density 1.57 g/mL at 25  C (87% TGAI)

Water solubility 0.5 g/100 mL at 25  C (PAI)

Solvent solubility g/ 100 mL at 25  C (PAI)

dioxane 118.0

ethanol 92.2 isopropyl alcohol 76.0 methylene chloride 26.0

acetone 17.0

toluene 13.0 xylene 7.8 heavy aromatic naphthalene 5.2

Vapor pressure 3.4 x 10

- 5 mm Hg at 25  C (PAI)

Dissociation constant, pK a 1. 97 (PAI)

Octanol/water partition coefficient, Log(K OW ) 0.1 (PAI)

UV/visible absorption spectrum neutral: 511 (275 nm) acidic (pH 0

- 1): 1053 (281 nm) basic (pH 13

- 14): 469 (274 nm)

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.4.1/7.4.2/OPPTS 860.1500/OECD IIA 6.3.1, 6.3.2, 6.3.3 and IIIA 8.3.1, 8.3.2, 8.3.3 Crop Field T rial/ Residue Decline

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D40 838 4 /MRID No. 48 728703 Page 6 of 28 

## B. EXPERIMENTAL DESIGN

B.1. Study Site Information

Thirteen fie ld trials were conducted on cotton in the United States during the 20 10 growing

season in NAFTA Zones 2 (GA and SC; 2 trials), 4 (AR, LA, and MO; 3 trials), 6 (TX; 1 trial), 8 (OK and TX; 5 trials), and 10 (CA; 2 trials).

Each trial consisted of one unt reated and three to four treated plots. The plots were planted with

dicamba

- tolerant cotton, identified as Event MON 88701 (formerly GH_S26695). Event MON

88701 cotton expresses a modified dicamba mono

- oxygenase (DMO) gene derived from Stenotrophomonas maltophilia and the bialaphos resistance (BAR) gene isolated from Streptomyces hygroscopicus . The expression of DMO confers dicamba tolerance and the expression of BAR confers glufosinate tolerance to Event MON 88701 cotton. Each treated plot

followed one of three treatment types: three broadcast applications applied at pre

- emergence, 6

- leaf, and 1 st 

## white flower + 15 days [TRT 2]; three broadcast applications applied at pre

emergence, first open boll, and 7

- days prior to harvest [TRT 3]; and four foliar b roadcast applications applied at 6

- leaf, 1 st 

## white flower + 15 days, first open boll, and 7 days prior to harvest [TRTs 4 and 5]. Applications to plots 2, 3, and 4 were made using a 4 lb ae/gal SL

formulation of dicamba diglycolamine salt (Clarity Herbici de; MON 54140); plot 5 applications were made using a 5 lb ae/gal SL formulation of dicamba monoethanolamine salt (MON 11968) to provide bridging data. The total application rate for all treatment types was 2.0

- 2.1 lb ae/A (2.2

- 2.4 kg ae/ha). F or TRT 2

, application 1 was applied at 0.98

- 1.0 lb ae/A (1.1 kg ae/ha) and applications 2 and 3 were applied at 0.49

- 0.51 lb ae/A/application (0.55

- 0.57 kg ae/ha/application). RTIs were 24

- 49 days between the first and second applications and 36

- 49 days between the second and third applications. For TRT 3

, application 1 was applied at 0.99

- 1.0 lb ae/A (1.1 kg ae/ha) and applications 2 and 3 were applied at 0.49

- 0.52 lb ae/A/application ( 0.55

- 0.58 kg ae/ha/application). RTIs were 97

- 178 days between the first and se cond applications and 5

- 63 days between the second and third applications. For TRTs 4 and 5

, applications were applied at 0.49

- 0.52 lb ae/A/application ( 0.55

- 0.58 kg ae/ha/application)

. RTIs

were 36

- 49 days between th e first and second applications, 32

- 8 8 days between the second and third applications , and 5

- 63 days between the third and fourth applications

. A summary of the use patterns is provided in the table below.

TRT Number

of Sites EP Growth Stages/ Target Application Rates lb ae/A (kg ae/ha)

Pre

- emergence 6 Leaf 1 st White Flower + 15 days First Open

Boll 7 Days

Prior to

Harvest

1 13

-- -- -- -- -- -- 2 13 4 lb ae/gal SL 1.0 (1.1) 0.50 (0.56) 0.50 (0.56)

-- -- 3 13 4 lb ae/gal SL 1.0 (1.1)

-- -- 0.50 (0.56) 0.50 (0.56)

4 13 4 lb ae/gal SL

-- 0.50 (0.56) 0.50 (0.56) 0.50 (0.56) 0.50 (0.56)

5 4 5 lb ae/gal SL

-- 0.50 (0.56) 0.50 (0.56) 0.50 (0.56) 0.50 (0.56)

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.4.1/7.4.2/OPPTS 860.1500/OECD IIA 6.3.1, 6.3.2, 6.3.3 and IIIA 8.3.1, 8.3.2, 8.3.3 Crop Field T rial/ Residue Decline

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D40 838 4 /MRID No. 48 728703 Page 7 of 28 

## Applications for all treatments were made using ground equipment (backpack, hand

- held and

tractor

- mounted sprayers) in spray volu mes of 18.8

- 21.6 gal/A (176

- 202 l/ha) and included an

NIS and an AMS at all trial sites.

Undelinted cotton seed was collected mechanically with a cotton picker (n=3) or a cotton stripper (n=3) or by hand (n=7) at normal harvest : 49

- 105

- day preharvest in terval ( PHI ) for TRT 2 and 6

- 8

- day PHI for TRTs 3

- 5

. Additional samples of cotton seed were collected from two trials

conducted in SC and TX at PHIs of 43/99, 49/105, 56/112, and 63/119 days for TRT 2 and 1, 7, 14, and 21 days for TRT 4 to assess residue decline; decline sampling times corresponded to 1 day following the 7

- day PHI application, normal harvest, 7±1 days following normal harvest, and 14±1 days following normal harvest. Cotton gin byproduct samples were collected from three sites (in TX and O K) using a cotton stripper at normal harvest (82

- 84

- day PHI for TRT 2;

6

- 7

- day PHI for TRT 4)

.

For each field trial, the petitioner presented the average minimum and maximum monthly temperatures and the total monthly rainfall for the study period, as we ll as the historical averages. Irrigation was used to supplement rainfall as needed. The petitioner reported that n o unusual weather events occurred that negatively affected the residue trials; however,

p recipitation was greater than normal at the CA2, T X1, and TX4 sites and less than normal at the LA1, OK1, and OK2 sites . These conditions did not cause adverse crop growth and development, except at CA2 where cool temperatures and excess moisture in October and November kept the cotton from maturing at a normal rate.

Trial conditions are presented in Table B.1.1. The study use pattern is presented in Table B.1.2, and the crop varieties grown are identified in Table C.3.

TABLE B.1.1 Trial Site Conditions

. Trial Identification: City, C oun ty, State; Y ear (Trial No.) Soil characteristics
Type %OM 1 pH 1 CEC 1 meq/ 100 g Proctor, Crittenden, AR; 2010 (AR1) Clay 1.1 6.3 17.4
Porterville, Tulare, CA; 2010 (CA1) Sandy loam 0.5 8.2 Not reported
Visalia, Tulare, CA; 2010 (CA2) Sandy loam 1.11 7.2 5.6

Chula , Tift, GA; 2010 (GA1) Loamy sand Not reported Cheneyville, Rapides, LA; 2010 (LA1) Silt loam 0.88 8.07 13.77
Fisk, Butler, MO; 2010 (MO1) Silt loam 1.4 5.2 9.0
Hinton, Caddo, OK; 2010 (OK1) Fine sandy loam 1.6 6.6 12.5 Dill City, Washita, OK; 2010 (OK 2) Loamy sand 0.6 7.4 6.0
Elko, Barnwell, SC; 2010 (SC1) Loamy sand 1.0 6.1 3.6 Raymondville, Willacy, TX; 2010 (TX1) Clay loam 1.1 8.2 26.7 Levelland, Hockley, TX; 2010 (TX2) Sandy loam 0.59 7.9 10.35
Wolfforth, Lubbock, TX; 2010 (TX3) Sandy clay loam 0.6 7.6 10.1
Uvalde, Uvalde, TX; 2010 (TX4) Clay loam 2.2 7.9 25.4

1 These parameters are optional except in cases where their value affects the use pattern for the chemical.

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.4.1/7.4.2/OPPTS 860.1500/OECD IIA 6.3.1, 6.3.2, 6.3.3 and IIIA 8.3.1, 8.3.2, 8.3.3 Crop Field T rial/ Residue Decline

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D40 838 4 /MRID No. 48 728703 Page 8 of 28 TABLE B.1.2. Study Use Pattern.

Location City, County, State;Year (Trial I D) EP 1 TRT Application Tank Mix/ Adjuvants 3 Harvest

Procedures Method; Timing Volume gal/A (l/ha) Rate

lb

a e /A (kg

ae/ ha ) RTI 2 (days) Total

Rate

lb a e /A (kg

ae/ ha )

Proctor, Crittenden, AR; 2010 (AR1) 4 lb

ae/gal

SL 2 1. Soil b roadcast ; Preemergen ce 20.1 (188) 1.0

(1.1) -- 2.0

(2.2) NIS + AMS Mechanical picker
2. Foliar broadcast; 6

- leaf 20.0 (187) 0.50 (0.56) 29

3. Foliar broadcast; 1 st WF + 15 days 20.1 (188) 0.50 (0.56) 38

4 lb ae/gal

SL 3 1. Soil b roadcast ; Preemergence 20.1 (18 8) 1.0

(1.1) -- 2.0

(2.2) NIS + AMS Mechanical picker
2. Foliar broadcast; 1 st open boll 20.1 (188) 0.50 (0.56) 99

3. Foliar broadcast; 7 days prior

to harvest 20.1 (188) 0.50 (0.56) 31

4 lb ae/gal

SL 4 1. Foliar broadcast; 6

- leaf 20.1 (188 ) 0.50 (0.56) -- 2.0

(2.2) NIS + AMS Mechanical picker
2. Foliar broadcast; 1 st WF + 15 days 20.2 (189) 0.50 (0.56) 38

3. Foliar broadcast; 1 st open boll 20.1 (188) 0.50 (0.56) 32

4. Foliar broadcast; 7 days prior

to harvest 20.1 (188) 0. 50 (0.56) 31

5 lb ae/gal

SL 5 1. Foliar broadcast; 6

- leaf 20.0 (187) 0.50 (0.56) -- 2.0

(2.2) NIS + AMS Mechanical picker
2. Foliar broadcast; 1 st WF + 15 days 20.2 (189) 0.50 (0.56) 38

3. Foliar broadcast; 1 st open boll 20.2 (189) 0.50 (0. 56) 32

4. Foliar broadcast; 7 days prior

to harvest 20.1 (188) 0.50 (0.56) 31

Porterville, Tulare, CA; 2010 (CA1) 4 lb

ae/gal

SL 2 1. Soil b roadcast ; Preemergence 20.5 (192) 1.0

(1.1) -- 2.0

(2.2) NIS + AMS Harvested

by hand

2. Foliar broadc ast; 6

- leaf 20.3 (190) 0.50 (0.56) 48

3. Foliar broadcast; 1 st WF + 15 days 20.1 (188) 0.51 (0.57) 43

4 lb ae/gal

SL 3 1. Soil b roadcast ; Preemergence 20.3 (190) 1.0

(1.1) -- 2.0

(2.2) NIS + AMS Harvested

by hand
2. Foliar broadcast; 1 st op en boll 20.2 (189) 0.51 (0.57) 136

3. Foliar broadcast; 7 days prior

to harvest 20.4 (191) 0.51 (0.57) 28

4 lb ae/gal

SL 4 1. Foliar broadcast; 6

- leaf 20.2 (189) 0.50 (0.56) -- 2.0

(2.2) NIS + AMS Harvested

by hand
2. Foliar broadcast; 1 st WF + 15 days 20.3 (190) 0.52 (0.58) 43

3. Foliar broadcast; 1 st open boll 20.2 (189) 0.51 (0.57) 45

4. Foliar broadcast; 7 days prior

to harvest 20.2 (189) 0.51 (0.57) 28

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.4.1/7.4.2/OPPTS 860.1500/OECD IIA 6.3.1, 6.3.2, 6.3.3 and IIIA 8.3.1, 8.3.2, 8.3.3 Crop Field T rial/ Residue Decline

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D40 838 4 /MRID No. 48 728703 Page 9 of 28 TABLE B.1.2. Study Use Pattern.

Location City, County, State;Year (Trial I D) EP 1 TRT Application Tank Mix/ Adjuvants 3 Harvest

Procedures Method; Timing Volume gal/A (l/ha) Rate

lb

a e /A (kg

ae/ ha ) RTI 2 (days) Total

Rate

lb a e /A (kg

ae/ ha )

Porterville, Tulare, CA; 2010 (CA1)

(continued) 5 lb

ae/gal

SL 5 1 . Foliar broadcast; 6

- leaf 20.2 (189) 0.50 (0.56) -- 2.0

(2.2) NIS + AMS Harvested

by hand
2. Foliar broadcast; 1 st WF + 15 days 20.2 (189) 0.52 (0.58) 43

3. Foliar broadcast; 1 st open boll 20.1 (188) 0.50 (0.56) 45

4. Foliar broadcast; 7 days prior

to harvest 20.0 (187) 0.50 (0.56) 28

Visalia, Tulare, CA; 2010 (CA2) 4 lb

ae/gal

SL 2 1. Soil b roadcast ; Preemergence 20.5 (192) 1.0

(1.1) -- 2.0

(2.2) NIS + AMS Harvested

by hand
2. Foliar broadcast; 6

- leaf 20.1 (188) 0.50 (0.56) 41

3. Foliar broadcast; 1 st WF + 15 days 20.7 (194) 0.51 (0.57) 49

4 lb ae/gal

SL 3 1. Soil b roadcast ; Preemergence 20.4 (191) 1.0

(1.1) -- 2.0

(2.2) NIS + AMS Harvested

by hand
2. Foliar broadcast; 1 st open boll 20.3 (190) 0.51 (0.57) 178

3. Foliar broadcast; 7 days prior

to harvest 20.5 (192) 0.50 (0.56) 7

4 lb ae/gal

SL 4 1. Foliar broadcast; 6

- leaf 20.6 (193) 0.51 (0.57) -- 2.0

(2.2) NIS + AMS Harvested

by hand
2. Foliar broadcast; 1 st WF + 15 days 20.3 (190) 0.50 (0.56) 49

3. Foliar broadcast; 1 st open boll 20.4 (191) 0.51 (0.57) 88

4. Foliar broadcast; 7 days prior

to harvest 20.8 (195) 0.51 (0.57) 7

Chula, Tift,
GA; 2010 (GA1) 4 lb

ae/gal

SL 2 1. Soil b roadcast ; Preemergence 20.5 (192) 0.99

(1.1) -- 2.0

(2. 2) NIS + AMS Harvested

by hand
2. Foliar broadcast; 6

- leaf 20.5 (192) 0.50 (0.56) 26

3. Foliar broadcast; 1 st WF + 15 days 20.2 (189) 0.51 (0.57) 43

4 lb ae/gal

SL 3 1. Soil b roadcast ; Preemergence 20.6 (193) 1.0

(1.1) -- 2.0

(2.2) NIS + AM S Harvested

by hand
2. Foliar broadcast; 1 st open boll 20.4 (191) 0.52 (0.58) 104

3. Foliar broadcast; 7 days prior

to harvest 20.4 (191) 0.50 (0.56) 42

4 lb ae/gal

SL 4 1. Foliar broadcast; 6

- leaf 20.9 (195) 0.51 (0.57) -- 2.0

(2.2) NIS + AMS Harvested

by hand
2. Foliar broadcast; 1 st WF + 15 days 20.0 (187) 0.50 (0.56) 43

3. Foliar broadcast; 1 st open boll 19.8 (185) 0.51 (0.57) 35

4. Foliar broadcast; 7 days prior

to harvest 20.4 (191) 0.50 (0.56) 42

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.4.1/7.4.2/OPPTS 860.1500/OECD IIA 6.3.1, 6.3.2, 6.3.3 and IIIA 8.3.1, 8.3.2, 8.3.3 Crop Field T rial/ Residue Decline

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D40 838 4 /MRID No. 48 728703 Page 10 of 28 TABLE B.1.2. Study Use Pattern.

Location City, County, State;Year (Trial I D) EP 1 TRT Application Tank Mix/ Adjuvants 3 Harvest

Procedures Method; Timing Volume gal/A (l/ha) Rate

lb

a e /A (kg

ae/ ha ) RTI 2 (days) Total

Rate

lb a e /A (kg

ae/ ha )

Chula, Tift, GA; 2010 (GA1)

(continued) 5 lb

ae/gal

SL 5 1. Foliar broadcast; 6

- leaf 20.9 (195) 0.51 (0.57) -- 2.0

(2.2) NIS + AMS Harvested

by hand
2. Foliar broadcast; 1 st WF + 15 days 20.0 (187) 0.50 (0.56) 43

3. Foliar broadcast; 1 st open boll 19.5 (182) 0 .50 (0.56) 35

4. Foliar broadcast; 7 days prior

to harvest 20.2 (189) 0.50 (0.56) 42

Cheneyville, Rapides, LA; 2010 (LA1) 4 lb

ae/gal

SL 2 1. Soil b roadcast ; Preemergence 20.5 (192) 1.0

(1.1) -- 2.0

(2.2) NIS + AMS Harvested

by hand

2. Folia r broadcast; 6

- leaf 19.0 (178) 0.51 (0.57) 27

3. Foliar broadcast; 1 st WF + 15 days 19.2 (180) 0.51 (0.57) 37

4 lb ae/gal

SL 3 1. Soil b roadcast ; Preemergence 20.8 (195) 1.0

(1.1) -- 2.0

(2.2) NIS + AMS Harvested

by hand

2. Foliar broadcast ; 1 st open boll 20.1 (188) 0.50 (0.56) 97

3. Foliar broadcast; 7 days prior

to harvest 21.6 (202) 0.52 (0.58) 33

4 lb ae/gal

SL 4 1. Foliar broadcast; 6

- leaf 19.2 (180) 0.52 (0.58) -- 2.0

(2.2) NIS + AMS Harvested

by hand

2. Foliar broadcas t; 1 st WF + 15 days 19.4 (181) 0.51 (0.57) 37

3. Foliar broadcast; 1 st open boll 20.5 (192) 0.51 (0.57) 33

4. Foliar broadcast; 7 days prior

to harvest 21.2 (198) 0.51 (0.57) 33

Fisk, Butler,
MO; 2010 (MO1) 4 lb

ae/gal

SL 2 1. Soil b roadc ast ; Preemergence 20.0 (187) 1.0

(1.1) -- 2.0

(2.2) NIS + AMS Mechanical picker
2. Foliar broadcast; 6

- leaf 20.0 (187) 0.50 (0.56) 29

3. Foliar broadcast; 1 st WF + 15 days 20.0 (187) 0.50 (0.56) 42

4 lb ae/gal

SL 3 1. Soil b roadcast ; Preeme rgence 20.0 (187) 1.0

(1.1) -- 2.0

(2.2) NIS + AMS Mechanical picker
2. Foliar broadcast; 1 st open boll 20.0 (187) 0.50 (0.56) 113

3. Foliar broadcast; 7 days prior

to harvest 20.0 (187) 0.50 (0.56) 30

4 lb ae/gal

SL 4 1. Foliar broadcast; 6

- leaf 20.3 (190) 0.51 (0.57) -- 2.0

(2.2) NIS + AMS Mechanical picker
2. Foliar broadcast; 1 st WF + 15 days 20.1 (188) 0.50 (0.56) 42

3. Foliar broadcast; 1 st open boll 20.0 (187) 0.50 (0.56) 42

4. Foliar broadcast; 7 days prior

to harve st 20.0 (187) 0.50 (0.56) 30

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.4.1/7.4.2/OPPTS 860.1500/OECD IIA 6.3.1, 6.3.2, 6.3.3 and IIIA 8.3.1, 8.3.2, 8.3.3 Crop Field T rial/ Residue Decline

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D40 838 4 /MRID No. 48 728703 Page 11 of 28 TABLE B.1.2. Study Use Pattern.

Location City, County, State;Year (Trial I D) EP 1 TRT Application Tank Mix/ Adjuvants 3 Harvest

Procedures Method; Timing Volume gal/A (l/ha) Rate

lb

a e /A (kg

ae/ ha ) RTI 2 (days) Total

Rate

lb a e /A (kg

ae/ ha )

Hinton, Caddo, OK; 2010 (OK1) 4 lb

ae/gal

SL 2 1. Soil b roadcast ; Preemergence 18.8 (176) 0.99

(1.1) -- 2.0

(2.2) NIS + AMS Harvested

by hand
2. Foliar broadcast; 6

- leaf 20.3 (190) 0.51 (0.57) 24

3. Foliar broadc ast; 1 st WF + 15 days 20.1 (188) 0.50 (0.56) 49

4 lb ae/gal

SL 3 1. Soil b roadcast ; Preemergence 19.0 (178) 1.0

(1.1) -- 2.0

(2.2) NIS + AMS Harvested

by hand
2. Foliar broadcast; 1 st open boll 19.8 (185) 0.50 (0.56) 107

3. Foliar broadcast ; 7 days prior

to harvest 20.6 (193) 0.51 (0.57) 27

4 lb ae/gal

SL 4 1. Foliar broadcast; 6

- leaf 20.3 (190) 0.51 (0.57) -- 2.0

(2.2) NIS + AMS Harvested

by hand
2. Foliar broadcast; 1 st WF + 15 days 19.9 (186) 0.49 (0.55) 49

3. Foliar broad cast; 1 st open boll 20.2 (189) 0.51 (0.57) 34

4. Foliar broadcast; 7 days prior

to harvest 20.6 (193) 0.51 (0.57) 27

5 lb ae/gal

SL 5 1. Foliar broadcast; 6

- leaf 19.8 (185) 0.50 (0.56) -- 2.0

(2.2) NIS + AMS Harvested

by hand

2. Foliar broa dcast; 1 st WF + 15 days 19.8 (185) 0.49 (0.55) 49

3. Foliar broadcast; 1 st open boll 19.7 (184) 0.50 (0.56) 34

4. Foliar broadcast; 7 days prior

to harvest 20.3 (190) 0.50 (0.56) 27

Dill City, Washita, OK; 2010 (OK2) 4 lb

ae/gal

SL 2 1. S oil b roadcast ; Preemergence 19.0 (178) 0.98

(1.1) -- 2.0

(2.2) NIS + AMS Mechanical

stripper
2. Foliar broadcast; 6

- leaf 20.1 (188) 0.50 (0.56) 28

3. Foliar broadcast; 1 st WF + 15 days 19.7 (184) 0.49 (0.55) 49

4 lb ae/gal

SL 3 1. Soil b roa dcast ; Preemergence 19.2 (180) 0.99

(1.1) -- 2.0

(2.2) NIS + AMS Harvested

by hand
2. Foliar broadcast; 1 st open boll 20.3 (190) 0.50 (0.56) 109

3. Foliar broadcast; 7 days prior

to harvest 20.1 (188) 0.50 (0.56) 43

4 lb ae/gal

SL 4 1. Foli ar broadcast; 6

- leaf 20.2 (189) 0.51 (0.57) -- 2.0

(2.2) NIS + AMS Mechanical

stripper
2. Foliar broadcast; 1 st WF + 15 days 20.1 (188) 0.50 (0.56) 49

3. Foliar broadcast; 1 st open boll 20.3 (190) 0.50 (0.56) 32

4. Foliar broadcast; 7 day s prior

to harvest 19.7 (184) 0.49 (0.55) 43

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.4.1/7.4.2/OPPTS 860.1500/OECD IIA 6.3.1, 6.3.2, 6.3.3 and IIIA 8.3.1, 8.3.2, 8.3.3 Crop Field T rial/ Residue Decline

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D40 838 4 /MRID No. 48 728703 Page 12 of 28 TABLE B.1.2. Study Use Pattern.

Location City, County, State;Year (Trial I D) EP 1 TRT Application Tank Mix/ Adjuvants 3 Harvest

Procedures Method; Timing Volume gal/A (l/ha) Rate

lb

a e /A (kg

ae/ ha ) RTI 2 (days) Total

Rate

lb a e /A (kg

ae/ ha )

Elko, Barnwell, SC; 2010 (SC1) 4 lb

ae/gal

SL 2 1. Soil b roadcast ; Preemergence 20.3 (190) 1.0

(1.1) -- 2.0

(2.2) NIS + AMS Harvested

by hand
2. Foliar broadcast; 6

- leaf 20.8 (195) 0.50 (0.56) 29

3. Foliar broadcast; 1 st WF + 15 days 20.6 (193) 0.50 (0.56) 38

4 lb ae/gal

SL 3 1. Soil b roadcast ; Preemergence 20.2 (189) 1.0

(1.1) -- 2.0

(2.2) NIS + AMS Harvested

by hand
2. Foliar broadcast; 1 st open boll 20.8 (195) 0.50 (0.56) 102

3. Foliar broadcast; 7 days prior

to harvest 20.0 (187) 0.50 (0.56) 63

4 lb ae/gal

SL 4 1. Foliar broadcast; 6

- leaf 20.7 (194) 0.50 (0.56) -- 2.0

(2.2) NIS + AMS Harvested

by hand
2. Foliar broadcast; 1 st WF + 15 days 20.6 (193) 0.50 (0.56) 38

3. Foliar broadcast; 1 st open boll 20.6 (193) 0.50 (0.56) 35

4. Foliar broadcast; 7 days prior

to harvest 19.9 (186) 0.50 (0.56) 63

Raymondville, Willacy, TX; 2010 (TX1) 4 lb

ae/gal

SL 2 1. Soil b roadcast ; Preemergence 20.3 (190) 1.0

(1.1) -- 2 .0

(2.2) NIS + AMS Harvested

by hand
2. Foliar broadcast; 6

- leaf 20.7 (194) 0.52 (0.58) 36

3. Foliar broadcast; 1 st WF + 15 days 20.9 (195) 0.52 (0.58) 46

4 lb ae/gal

SL 3 1. Soil b roadcast ; Preemergence 20.4 (191) 1.0

(1.1) -- 2.0

(2.2) NI S + AMS Harvested

by hand
2. Foliar broadcast; 1 st open boll 20.7 (194) 0.52 (0.58) 119

3. Foliar broadcast; 7 days prior

to harvest 20.6 (193) 0.52 (0.58) 5

4 lb ae/gal

SL 4 1. Foliar broadcast; 6

- leaf 20.7 (194) 0.52 (0.58) -- 2

. 1

(2.4 ) N IS + AMS Harvested

by hand
2. Foliar broadcast; 1 st WF + 15 days 20.8 (195) 0.52 (0.58) 46

3. Foliar broadcast; 1 st open boll 20.7 (194) 0.52 (0.58) 37

4. Foliar broadcast; 7 days prior

to harvest 20.9 (195) 0.52 (0.58) 5

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.4.1/7.4.2/OPPTS 860.1500/OECD IIA 6.3.1, 6.3.2, 6.3.3 and IIIA 8.3.1, 8.3.2, 8.3.3 Crop Field T rial/ Residue Decline

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D40 838 4 /MRID No. 48 728703 Page 13 of 28 TABLE B.1.2. Study Use Pattern.

Location City, County, State;Year (Trial I D) EP 1 TRT Application Tank Mix/ Adjuvants 3 Harvest

Procedures Method; Timing Volume gal/A (l/ha) Rate

lb

a e /A (kg

ae/ ha ) RTI 2 (days) Total

Rate

lb a e /A (kg

ae/ ha )

Levelland,

H ockley, TX; 2010 (TX2) 4 lb

ae/gal

SL 2 1. Soil b roadcast ; Preemergence 20.4 (191) 1.0

(1.1) -- 2.0

(2.2) NIS + AMS Mechanical

stripper
2. Foliar broadcast; 6

- leaf 20.6 (193) 0.51 (0.57) 36

3. Foliar broadcast; 1 st WF + 15 days 20.1 (188) 0.50 ( 0.56) 41

4 lb ae/gal

SL 3 1. Soil b roadcast ; Preemergence 20.3 (190) 1.0

(1.1) -- 2.0

(2.2) NIS + AMS Mechanical

stripper
2. Foliar broadcast; 1 st open boll 19.7 (184) 0.49 (0.55) 118

3. Foliar broadcast; 7 days prior

to harvest 19.8 (185) 0.49 (0.55) 37

4 lb ae/gal

SL 4 1. Foliar broadcast; 6

- leaf 20.0 (187) 0.49 (0.55) -- 2.0

(2.2) NIS + AMS Mechanical

stripper
2. Foliar broadcast; 1 st WF + 15 days 20.0 (187) 0.49 (0.55) 41

3. Foliar broadcast; 1 st open boll 20.4 (191) 0.50 (0.56) 41

4. Foliar broadcast; 7 days prior

to harvest 20.0 (187) 0.49 (0.55) 37

Wolfforth, Lubbock, TX; 2010 (TX3) 4 lb

ae/gal

SL 2 1. Soil b roadcast ; Preemergence 20.1 (188) 1.0

(1.1) -- 2.0

(2.2) NIS + AMS Mechanical

stripper

2. Foliar b roadcast; 6

- leaf 20.7 (194) 0.51 (0.57) 41

3. Foliar broadcast; 1 st WF + 15 days 19.8 (185) 0.49 (0.55) 36

4 lb ae/gal

SL 3 1. Soil b roadcast ; Preemergence 20.2 (189) 1.0

(1.1) -- 2.0

(2.2) NIS + AMS Mechanical

stripper
2. Foliar broadcast; 1 st open boll 20.0 (187) 0.49 (0.55) 125

3. Foliar broadcast; 7 days prior

to harvest 19.7 (184) 0.49 (0.55) 30

4 lb ae/gal

SL 4 1. Foliar broadcast; 6

- leaf 20.2 (189) 0.50 (0.56) -- 2.0

(2.2) NIS + AMS Mechanical

stripper

2. Foliar broadc ast; 1 st WF + 15 days 20.1 (188) 0.50 (0.56) 36

3. Foliar broadcast; 1 st open boll 20.2 (189) 0.50 (0.56) 48

4. Foliar broadcast; 7 days prior

to harvest 20.1 (188) 0.50 (0.56) 30

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.4.1/7.4.2/OPPTS 860.1500/OECD IIA 6.3.1, 6.3.2, 6.3.3 and IIIA 8.3.1, 8.3.2, 8.3.3 Crop Field T rial/ Residue Decline

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D40 838 4 /MRID No. 48 728703 Page 14 of 28 TABLE B.1.2. Study Use Pattern.

Location City, County, State;Year (Trial I D) EP 1 TRT Application Tank Mix/ Adjuvants 3 Harvest

Procedures Method; Timing Volume gal/A (l/ha) Rate

lb

a e /A (kg

ae/ ha ) RTI 2 (days) Total

Rate

lb a e /A (kg

ae/ ha )

Uvalde, Uvalde, TX; 2010 (TX4) 4 lb

ae/gal

SL 2 1. Soil b r oadcast ; Preemergence 20.2 (189) 1.0

(1.1) -- 2.0

(2.2) NIS + AMS Mechanical picker
2. Foliar broadcast; 6

- leaf 20.1 (188) 0.50 (0.56) 49

3. Foliar broadcast; 1 st WF + 15 days 19.9 (186) 0.50 (0.56) 47

4 lb ae/gal

SL 3 1. Soil b roadcast ; Pr eemergence 19.9 (186) 1.0

(1.1) -- 2.0

(2.2) NIS + AMS Mechanical picker
2. Foliar broadcast; 1 st open boll 21.5 (201) 0.52 (0.58) 132

3. Foliar broadcast; 7 days prior

to harvest 19.7 (184) 0.49 (0.55) 29

4 lb ae/gal

SL 4 1. Foliar broadca st; 6

- leaf 20.0 (187) 0.50 (0.56) -- 2.0

(2.2) NIS + AMS Mechanical picker
2. Foliar broadcast; 1 st WF + 15 days 19.7 (184) 0.49 (0.55) 47

3. Foliar broadcast; 1 st open boll 20.5 (192) 0.49 (0.55) 36

4. Foliar broadcast; 7 days prior

to h arvest 20.2 (189) 0.51 (0.57) 29

1 EP = E nd

- use Product ; Clarity [ SL formulation containing 480 g /L dicamba formulated as the diglycolamine salt ( 4 lb ae /gal) ] and MON 11968 [ SL formulation containing 600 g/L dicamba formulated as the monoethanolamine salt (5 lb ae/gal)

. 2 Retreatment Interval.

3 NIS = nonionic surfactant

. AMS = ammonium sulfate.

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.4.1/7.4.2/OPPTS 860.1500/OECD IIA 6.3.1, 6.3.2, 6.3.3 and IIIA 8.3.1, 8.3.2, 8.3.3 Crop Field T rial/ Residue Decline

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D40 838 4 /MRID No. 48 728703 Page 15 of 28 TABLE B.1.3. Trial Numbers and Geographical Locations

.

NAFTA Growing Zones Cotton

Submitted Requested 1

Canada U.S.

1

-- -- --

1A

-- -- --
2 2

-- 1/1

3

-- -- --
4 3

-- 3/2

5

-- -- --
5A

-- -- --
5B

-- -- --

6 1

-- 1/1

7

-- -- --

7A

-- -- --
8 5

-- 4/3

9

-- -- --
10 2

-- 3/2

11

-- -- --

12

-- -- --
13

-- -- --
14

-- -- --
15

-- -- --
16

-- -- --
17

-- -- --
18

-- -- --
19

-- -- --
20

-- -- --
21

-- -- --

Total 13 12 / 9

1 A s per OCSPP 860.1500, Tables 1 and 5 for cotton ; the second number reflects a 25% reduction in the number of trials allowed for the crop as a representative commodity in support of a crop group/subgroup to lerance or when application results in no quantifiable residues.

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.4.1/7.4.2/OPPTS 860.1500/OECD IIA 6.3.1, 6.3.2, 6.3.3 and IIIA 8.3.1, 8.3.2, 8.3.3 Crop Field T rial/ Residue Decline

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D40 838 4 /MRID No. 48 728703 Page 16 of 28 

## B.2. Sample Handling and Preparation

At all test sites, one untreated and duplicate treated cotton seed samples were harvested at normal

harvest at a PHI of 49

- 105 days for TRT 2 and 6

- 8 days for TRTs 3

- 5

. Two site s ( SC1 and TX1 ) collected additional treated decline samples. The cotton seed samples were harvested by mechanical picker, stripper or by hand and ginned to yield ~1.0 kg of undelinted seed samples (all sites) and ~0.5

- 1.0 kg of gin byproduct samples ( 3 sites only). Samples were ginned at the

field sites within 3 hours after harvest , except for sites OK2, TX2, and TX3 where the cotton was ginned 1 day after harvest . The treated samples were placed in frozen storage at the fie ld trials

within 3. 1 hours after sampling . All samples were shipped 0 to 41 days after collection by

freezer truck or shipped by FedEx overnight to Monsanto ( St. Louis, MO ) for residue analysis. Samples were maintained frozen (

- 20 °C) at the analytical l aboratory prior to homogenization and analysis. In preparation for analysis, the cotton gin by product samples were initially ground with dry ice in a vertical cutter/mixer

. Undelinted c otton seed and gin byproduct samples were further processed by millin g in a cryomill, which pulverizes samples into a fine powder at cryogenic temperatures.

B.3. Analytical Methodology

S amples were analyzed for residues of dicamba and the metabolite s DCGA, DCSA, and 5

- hydoxy dicamba using a LC /MS/MS method from Monsant o Company : Analytical Method AG

ME

- 1381

- 01

. 

## For a complete d escription of the LC/MS/MS method, ref er to the 860.1340 DER

for MRID 48728702

. We note that the analytical method does not specify conversion of metabolite residues to parent equivalents; there fore, quantifiable res idues of 5

- OH dicamba,

DCSA , and DCGA were converted to parent equivalents by the study reviewer using a molecular weight conversion factor of 0.933, 1.068, and 0.991, respectively.

Briefly, samples were extracted with acetonitrile : w ater ( 4 0: 6 0, v:v). An aliquot of the extract

was hydrolyzed in 1 N HCl at 95 °C in an oven

. The hydrolysate was partitioned with ethyl acetate:isooctane (20:80, v:v). Water was added to the organic phase and the sample was concentrated by evaporation un til only the aqueous solution remained. The samples were acidified and analyzed using LC/MS/MS.

The LOQ , determined as the LLMV, was 0. 02 ppm for each analyte in undelinted cotton seed and 0.04 ppm for each analyte in cotton gin by

- products

. The 0.020 p pm fortification level was used to statistically determine the LODs and LOQs for the four analytes of the method in

undelinted cotton seed. The LODs for the analytes in undelinte d cotton seed were 0.0035, 0.0041, 0.0057 and 0.0089 ppm for DCSA, DCGA, 5

- hyd roxydicamba and dicamba, respectively. The LOQs for the analytes in undelinte d cotton seed were 0.0105, 0.0121, 0.0171

and 0.0266 ppm for DCSA, DCGA, 5

- hydroxydicamba and dicamba, respectively (reported as analyte found per se). The LODs and LOQs were no t statistically determined in cotton gin byproducts

.

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.4.1/7.4.2/OPPTS 860.1500/OECD IIA 6.3.1, 6.3.2, 6.3.3 and IIIA 8.3.1, 8.3.2, 8.3.3 Crop Field T rial/ Residue Decline

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D40 838 4 /MRID No. 48 728703 Page 17 of 28 

## The analytical method was validated prior to and in conjunction with the analysis of field trial samples

.

C. RESULTS AND DISCUSSION

Sample storage conditions and durati ons are summarized in Table C. 2. Dicamba, 5

- OH dicamba, and DCSA undelinted cotton seed s amples were stored for 13

- 146 days ( 0.4

- 4.8 months) from collection to analysis; DCGA undelinted cotton seed samples were stored f or 13

- 285 days (0.4

- 9.4 months). Gin byproduct samples for all an alytes were stored from 17

- 27 days ( 0.6

- 0.9 months) from collection to analysis

. All samples were analyzed within 0 to 2 day s of

extraction. Samples were stored be low freezing at the field sites and at

- 20 °C at the analytical laboratory prior to extract ion. A storage stability study for dicamba

, DCGA , and DCSA on cotton was submitted in conjunction with the field trial study (refer to the 860.1380 der for

MRID 4872870 4 ). Residues of dicamba, DCGA, and DCSA were stable during freezer storage for up to 2 77 days (9 months) in undelinted cotton seed. The stability of 5

- OH dicamba was not determined concurrently because it was not found in undelinted cotton seed. These data are acceptable to support the stora ge conditions and durations of the undelinted co tton seed s amples

from the submitted field trials

. Storage stability data is not required for cotton gin byproduct samples because the samples were analyzed within 30 days of harvest.

Method validation and c oncurrent method recovery data are presented in Table C.1. The LC/MS/MS method used to analyze cotton samples for residues of dicamba, 5

- OH dicamba, DCGA, and DCSA was adequate for data collection based on acceptable method and concurrent method recovery data. Concurrent recoveries were generally wit hin the acceptable range of 70

- 120% in cotton fortified with dicamba, 5

- OH dicamba, DCGA, and DCSA each at 0.0 2

- 5.00 ppm in undelinted cotton seed and 0.0 4

- 100.0 ppm in gin byproducts . The fortification levels were adequate to bracket residues found in tr eated samples. Apparent residues of dicamba, 5

- OH dicamba, DCGA, DSGA were below the LOQ (<0.0 2 ppm or <0.04 ppm ) in/on all samples

of untreated undelinted cotton seed and cotton gin by

- products

. Concurrent recoveries were corrected for apparent for resi dues in controls; residues in treated samples were not corrected for

residues in controls.

Residue data from the cotton field trials are reported in Table C.3. A summary of residue data is presented in Table C.4.

Following three broadcast applications of the 4 lb ae/gal SL formulation of dicamba (formulated

as the diglycolamine salt), which were applied at pre

- emergence, the 6

- leaf stage, and first white flower + 15 days for a total rate of 2.0 lb ae/A (2.2 kg ae/ha) ( TRT 2 ), combined residues (and

per trial averages) of dicamba, 5

- OH dicamba, DCGA, and DCSA (expressed in parent equivalents) were <0.08

- <0.39 (<0.08

- < 0 .39) ppm in/on undelinted cotton seed collected at PHIs

of 49

- 105 days and <0.67

- <2.34 (<0.73

- <2.33) ppm in/on gin byproducts collected at PHIs of

82

- 84 days.

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.4.1/7.4.2/OPPTS 860.1500/OECD IIA 6.3.1, 6.3.2, 6.3.3 and IIIA 8.3.1, 8.3.2, 8.3.3 Crop Field T rial/ Residue Decline

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D40 838 4 /MRID No. 48 728703 Page 18 of 28 

## Following three broadcast applications of the 4 lb ae/gal SL formulation of dicamba (formulated

as the diglycolamine salt), which were applied at pre

- emergence, the first open boll stage, and 7 days prior to harvest for a total rate of 2.0 lb ae/A (2.2 kg ae/ha) ( TRT 3 ), combined residues (and per trial averages) of dicamba, 5

- OH dicamba, DCGA, and DCSA (expressed in parent equivalents) were <0.12

- <2.24 (<0.13

- <1.66) ppm i n/on undelinted cotton seed collected at a 6

- 8

- day PHI.

Follow ing four foliar broadcast applications of the 4 lb ae/gal SL formulation of dicamba (formulated as the diglycolamine salt) , which were applied at the 6

- leaf stage, the first white flower + 15 days, the first open boll, and 7 days prior to harvest for a tot al rate of 2.0

- 2.1 lb ae/A (2.2

- 2.4 kg ae/ha) ( TRT 4 ), combined residues (and per trial averages) of dicamba, 5

- OH dicamba, DCGA, and DCSA (expressed in parent equivalents) were <0.15

- <2.02 (<0.21

- <1.86) ppm in/on undelinted cotton seed collected at a 6

- 8

- day PHI and <5.50

- 34.6 (<5.51

- <33.7) ppm in/on gin byproducts collected at a 6

- 7

- day PHI. Corresponding combined residues (and per trial averages) in side

- by

- side trials treated with the 5 lb ae/gal SL formulation of dicamba formulated

as the monoethanolam ine salt ( TRT 5 ) were <0.24

- <1.00 (<0.29

- <0.82) ppm in/on undelinted cotton seed. Average combined residues in the diglycolamine salt formulation trials were slightly higher (approximately 1.5x) than the corresponding residues in the monoethanolamine

salt formulation trials.

Treatment plots 3 and 4 showed the highest average combined residues (0.76 ppm), followed closely by treatment plot 5 (average of 0.52 ppm). Treatment plot 2 had the lowest average combined residues (average of 0.12 ppm). Lower re sidues from treatment plot 2 is consistent with the earlier application timings at these plots.

For the TRT 2 residue decline samples, average residues of dicamba, 5

- OH dicamba, DCGA,

and DCSA in/on undelinted cotton seed were at or <LOQ in both studies ; therefore, residue decline could not be assessed. For the TRT 4 residue decline samples, average residues of

dicamba decreased in/on undelinted cotton seed with increasing PHI. Average residues of DCGA and DCSA increased with increasing PHI in the SC1 trial, and remained relatively

constant in the TX1 trial. Residues of 5

- OH dicamba were <LOQ in both studies; therefore,

residue decline could not be assessed.

T ABLE C.1. Summary of Method and Concurrent Recoveries of Dicamba and its Metabolites from

Cotton

Matrix Analyte Spike

Level (ppm) Sample Size (n) Recoveries (%) 1 Mean ±

Std. Dev. (%) 2

Method Validation Recoveries

Undelinted

Cotton seed Dicamba 0.005 5 187, 73, 172, 105, 102 128 ± 49

0.010 5 129, 149, 93, 78, 125 115 ± 29

0.020 5 119, 1 00, 123, 101, 124 113 ± 12

0.200 5 94, 96, 95, 97, 100 96.1 ± 2.2

10.0 5 68, 91, 91, 100, 98 89.6 ± 13

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.4.1/7.4.2/OPPTS 860.1500/OECD IIA 6.3.1, 6.3.2, 6.3.3 and IIIA 8.3.1, 8.3.2, 8.3.3 Crop Field T rial/ Residue Decline

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D40 838 4 /MRID No. 48 728703 Page 19 of 28 T ABLE C.1. Summary of Method and Concurrent Recoveries of Dicamba and its Metabolites from

Cotton

Matrix Analyte Spike

Level (ppm) Sample Size (n) Recoveries (%) 1 Mean ±

Std. Dev. (%) 2

Undelinted

Cotton seed (continued) 5

- OH

Dicamba 0.005 5 68, 56, 101, 84, 105 82.9 ± 21

0.010 5 106, 97, 76, 110, 82 94.1 ± 15

0.020 5 96, 98 , 115, 117, 114 108 ± 10

0.200 5 98, 104, 105, 104, 101 102 ± 3.0

10.0 5 69, 90, 79, 93, 104 87.1 ± 13

DCGA 0.005 5 140, 115, 88, 98, 129 114 ± 22

0.010 5 90, 108, 96, 79, 104 95.6 ± 11

0.020 5 81, 98, 93, 77, 86 86.9 ± 8.5

0.200 5 90, 87, 88, 84, 96 88.9 ± 4.3

10.0 5 73, 96, 89, 94, 90 88.6 ± 8.9

DCSA 0.005 5 88, 95, 90, 134, 98 101 ± 19

0.010 5 70, 84, 102, 96, 98 89.8 ± 13

0.020 5 100, 82, 97, 97, 95 94.0 ± 6.9

0.200 5 95, 100, 105, 96, 101 98.9 ± 4.0

10.0 5 81, 92, 95, 93, 97 91.6 ± 6.4

Gin Byproducts Dicamba 0.040 5 100, 93, 88, 122, 119 104 ± 15

0.400 5 97, 99, 101, 98, 104 100 ± 2.7

10.0 5 88, 91, 82, 85, 88 86.7 ± 3.1

5

- OH

Dicamba 0.040 5 124, 106, 98, 97, 108 106 ± 11

0.400 5 111, 113, 106, 116, 109 111 ± 3.6

10.0 5 113, 99, 99, 101, 85 99.4 ± 10

DCGA 0.040 5 67, 88, 88, 90, 70 80.4 ± 11

0.400 5 99, 96, 99, 95, 97 97.0 ± 2.0

10.0 5 101, 94, 105, 99, 104 101 ± 4.4

DCSA 0.040 5 79, 87, 76, 75, 81 79.5 ± 4.9

0.400 5 91, 84, 93, 87, 91 89.0 ± 3.7

10.0 5 110, 101, 92, 109, 99 102 ± 7.6

Concurrent Recoveries

Undelinted

Cotton seed Dicamba 0.02 19 103, 97.7, 93.5, 104, 104, 98.0, 91.6, 75.5, 96.5, 77.5, 116, 109, 110,
99.9, 99.7, 103, 108, 119, 110 101 ± 11

0.04 13 105, 106, 84.3, 105, 101 , 80.8, 97.6, 119, 114, 110, 100, 108, 109 103 ± 11

0.20 20 94.5, 105, 97.3, 103, 99.9, 107, 103, 98.5, 102, 95.0, 98.0, 108, 93.9, 97.8, 99.5, 105, 96.5, 101, 105, 94.5 100 ± 4.3

5.00 1 108 108

5

- OH

Dicamba 0.02 13 94.9, 100, 91.0, 79.5, 95.0, 110 , 69.5, 68.5, 82.0, 83.0, 88.0, 92.0,

94.0 88.2 ± 12

0.04 13 93.5, 94.3, 76.5, 86.3, 125, 121, 80.0, 90.3, 105, 110, 109, 107, 92.8 99.3 ± 15

0.20 15 93.2, 97.5, 97.0, 110, 93.9, 83.7, 115, 86.0, 104, 97.0, 93.5, 94.5, 77.0, 87.5, 108 95.8 ± 10

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.4.1/7.4.2/OPPTS 860.1500/OECD IIA 6.3.1, 6.3.2, 6.3.3 and IIIA 8.3.1, 8.3.2, 8.3.3 Crop Field T rial/ Residue Decline

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D40 838 4 /MRID No. 48 728703 Page 20 of 28 T ABLE C.1. Summary of Method and Concurrent Recoveries of Dicamba and its Metabolites from

Cotton

Matrix Analyte Spike

Level (ppm) Sample Size (n) Recoveries (%) 1 Mean ±

Std. Dev. (%) 2

Unde linted

Cotton seed (continued) DCGA 0.02 16 83.5, 71.4, 93.0, 86.0, 105, 83.5, 107, 101, 114, 120, 116, 87.5, 79.5, 95.5, 93.0, 66.9 93.8 ± 16

0.04 12 74.0, 83.9, 90.5, 81.3, 77.5, 101, 88.0, 88.8, 93.0, 79.8, 80.5, 80.3 84.8 ± 7.6

0.20 15 88.0, 92.2 , 94.5, 105, 82.5, 86.0, 98.1, 88.0, 90.5, 87.5, 92.5, 77.1, 96.0, 81.5, 100 90.6 ± 7.5

5.00 1 106 106

DCSA 0.02 18 105, 101, 100, 94.0, 122, 101, 86.0, 101, 91.0, 98.5, 89.5, 98.0, 103, 106, 101, 92.3, 87.0, 118 99.6 ± 9.4

0.04 13 84.0, 114, 89.3, 96.3, 108, 95.0, 93.8, 103, 106, 102, 103, 108, 106 101 ± 8.5

0.20 17 90.5, 100, 95.0, 107, 102, 97.0, 103, 99.5, 103, 101, 92.3, 100, 105, 97.2, 98.2, 101, 103 99.7 ± 4.3

5.00 1 99.8 99.8

Gin Byproducts Dicamba 0.04 4 120 , 90.5, 88.5, 94.0 9 8.2 ± 15

0.40 1 97. 2 97.2

5.00 3 108, 102 , 98.6 103 ± 4.8

100.00 3 96.7, 95.9, 104 98.9 ± 4.5

5

- OH

Dicamba 0.04 4 89.7 , 96.7, 107 , 98. 9 98.0 ± 6.9

0.40 1 103 103

5.00 3 102 , 93. 3 , 87. 3 94.1 ± 7.3

100.00 3 111 , 94. 5 , 99. 2 102 ± 8.5

DCGA 0.04 4 88.5, 70.0 , 93. 8 , 108 90.1 ± 16

0.40 1 98.0 98.0

5.00 3 110, 98.4, 105 105 ± 5.7

100.00 3 118, 104, 107 110 ± 7.4

DCSA 0.04 4 88.0, 101

, 103

, 98.0 97.4 ± 6.6

0.40 1 91.0 91.0

5.00 3 125, 114, 123 121 ± 5.9

100.00 3 99.2, 101, 106 102 ± 3.5

1 The concurrent recoveries were corrected for apparent residues in the unfortified control samples.

2 Standard deviation is not applicable for sample size s n < 3 samples.

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.4.1/7.4.2/OPPTS 860.1500/OECD IIA 6.3.1, 6.3.2, 6.3.3 and IIIA 8.3.1, 8.3.2, 8.3.3 Crop Field T rial/ Residue Decline

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D40 838 4 /MRID No. 48 728703 Page 21 of 28 TABLE C.2. Summary of Storage Co nd itions

. Matrix Analyte Storage Temp erature Actual Storage Duration 1 Interval of Demonstrated Storage Stability 2

Undelinted

Cotton seed Dicamba Below freezing at

field sites

- 20 oC at analytical

lab 13

- 146 days

(0.4

- 4.8 months) Residues of dicamba, DCGA, and DCSA

are stable during freezer storage for up to 277 days (9 months) in undelinted cotton

seed.

The stability of 5

- OH dicamba was not determined concurrently because it was

not detected in undelinted cotton seed



## . 5

- OH Dicamba 13

- 122 days (0.4

- 4.0 months)

DCGA 13

- 285 days (0.4

- 9.4 months)

DCSA 13

- 121 days (0.4

- 4.0 months)

Gin Byproducts Dicamba Below freezing at

field sites

- 20 oC at analytical

lab 17

- 27 days

(0.6

- 0.9 months) Not provided. Storage stability data is not

required as the samples were analyzed within 30 days of harvest. 5

- OH Dicamba

DCGA

DCSA

1 Interval from harvest to analysis ; all samples were analyzed within 0 to 2 days of extraction.

2 R efer to the 860.13 8 0 der for MRID 48728704.



### TABLE C. 3. Residue Data for Dicamba and its Metabolites from Cotton Field Trials

Trial ID (City, County, State; Year) Zone Com

modity 1 Total

Rate

lb a e /A (kg

ae/ ha ) TRT 2 PHI

(days) 3 Residues (ppm) [Average] 4, 5

Dicamba 5

- OH

Dicamba DCGA DCS A Combined

Residues

Proctor, Crittenden, AR; 2010 (AR1) 4 Und elinted

Cotton

seed 2.0

(2.2) 2 70 (0.01)

, ND [<0.0 2 ] ND

, ND

[<0.02] 0.0 3, 0.0 3

[ 0.0 3 ] 0.0 5, 0.0 4

[ 0.0 5 ] <0.12,

<0.11 [<0.12]

3 7 1.20 6 , 0.59 6

[0.89] ND, ND

[<0.02] 0.04, 0.03

[0.03] 0.10, 0.05

[0.08] <1.37,

<0.69 [<1.03]

4 7 0.33 6 , 0. 58 6

[ 0.46] ND, ND

[<0.02] 0.04, 0.05

[0.04] 0.05, 0.07

[0.06] <0.45,

<0.72 [<0.58]

5 7 0.72, 0.51

[0.62] ND, ND

[<0.02] 0.08, 0.04

[0.06] 0.17, 0.06

[0.12] <1.00,

<0.64 [<0.82]

Porterville, Tulare, CA; 2010 (CA1) 10 Undelinted

Cotton

seed 2.0

(2.2) 2 81 ND, ND [<0.02] ND, ND

[<0.02] (0.01),

(0.01) [<0.02] (0.01),

(0.01) [<0.02] <0.08,

<0.08 [<0.08]

3 8 0.18, 0.19

[0.18] ND, ND

[<0.02] ND, ND

[<0.02] (0.01), 0.02

[<0.02] <0.24,

<0.25 [<0.25]

4 8 0.47, 0.39

[0.43] ND, ND

[<0.02] 0.04,

(0.018)
[ < 0.03] 0.09, 0.04

[0.06] <0.63,

<0.47 [<0.55]

5 8 0.23, 0.17

[0.20] ND, ND

[<0.02] 0.02, 0.02

[0.02] 0.06, 0.02

[0.04] <0.34,

<0.24 [<0.29]

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.4.1/7.4.2/OPPTS 860.1500/OECD IIA 6.3.1, 6.3.2, 6.3.3 and IIIA 8.3.1, 8.3.2, 8.3.3 Crop Field T rial/ Residue Decline

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D40 838 4 /MRID No. 48 728703 Page 22 of 28 TABLE C. 3. Residue Data for Dicamba and its Metabolites from Cotton Field Trials

Trial ID (City, County, State; Year) Zone Com

modity 1 Total

Rate

lb a e /A (kg

ae/ ha ) TRT 2 PHI

(days) 3 Residues (ppm) [Average] 4, 5

Dicamba 5

- OH

Dicamba DCGA DCS A Combined

Residues

Visalia, Tulare, CA; 2010 (CA2) 10 Undelinted

Cotton

seed 2.0

(2.2) 2 103 ND, ND [<0.0 2] ND, ND

[<0.02] (0.01),

(0.01) [<0.02] (0.01),

(0.01) [<0.02] <0.08,

<0.08 [<0.08]

3 8 0.66, 0.64

[0.65] ND, ND

[<0.02] ND, (0.01)

[<0.02] 0.02, 0.05

[0.04] <0.72,

<0.73 [<0.73]

4 8 1.05, 0.82

[0.94] ND, ND

[<0.02] 0.03, 0.03

[0.03] 0.14, 0.16

[0.15] <1.25,

<1.03 [<1.14]

Chula, Tift, GA; 2010 (GA1) 2 Undelinted

Cotton

seed 2.0

(2.2) 2 84 ND, ND [<0.02] ND, ND

[<0.02] (0.01),

(0.01) [<0.02] (0.01),

(0.01) [<0.02] <0.08,

<0.08 [<0.08]

3 7 0.76 6

, 1.30 6

[0.90] ND, ND

[<0.0 2] (0.01), ND

[<0.02] (0.016)

,

( 0.0 16)
[ < 0.02] <0.82,

<1.36 [<1.09]

4 7 0.80, 0.71

[0.76] ND, ND

[<0.02] 0.03, 0.02

[0.03] 0.03, 0.02

[0.02] <0.88,

<0.78 [<0.83]

5 7 0.68, 0.50

[0.59] ND, ND

[<0.02] 0.02,

( 0.0 17)
[ < 0.02] (0.016),

(0.01 6) [<0.02] <0.74,

<0.5 6 [<0.65]

Cheneyville, Rapides, LA; 2010 (LA1) 4 Undelinted

Cotton

seed 2.0

(2.2) 2 73 ND, ND [<0.02] ND, ND

[<0.02] (0.01),

(0.01) [<0.02] (0.01),

(0.01) [<0.02] <0.08,

<0.08 [<0.08]

3 7 0.33, 0.45

[0.39] ND, ND

[<0.02 ] 0.02,

( 0.0 19)
[ < 0.02] (0.01),

(0.01) [<0.02] <0.39,

<0.51 [<0.45]

4 7 0.82, 0.65

[0.74] ND, ND

[<0.02] 0.03, 0.03

[0.03] 0.03, 0.06

[0.04] <0.90,

<0.76 [<0.83]

Fisk, Butler, MO; 2010 (MO1) 4 Undelinted

Cotton

seed 2.0

(2.2) 2 79 ND, ND [<0 .02] ND, ND

[<0.02] ( 0.0 15)

,

(0.015)
[ < 0.02] 0.02,

( 0.0 17)
[ < 0.02] <0.08,

<0.08 [<0.08 ]

3 7 0.97, 1.00

[0.98] ND, ND

[<0.02] (0.019)

,

(0.01) [<0.02] (0.01), 0.06

[<0.03] <1.03,

<1.10 [<1.07]

4 7 0.56 6

, 1.17 6

[0.87] ND, ND

[<0.02] 0.02, 0.02

[0.02] 0.02, 0.03

[0.03] <0.62,

<1.25 [<0.94]

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.4.1/7.4.2/OPPTS 860.1500/OECD IIA 6.3.1, 6.3.2, 6.3.3 and IIIA 8.3.1, 8.3.2, 8.3.3 Crop Field T rial/ Residue Decline

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D40 838 4 /MRID No. 48 728703 Page 23 of 28 TABLE C. 3. Residue Data for Dicamba and its Metabolites from Cotton Field Trials

Trial ID (City, County, State; Year) Zone Com

modity 1 Total

Rate

lb a e /A (kg

ae/ ha ) TRT 2 PHI

(days) 3 Residues (ppm) [Average] 4, 5

Dicamba 5

- OH

Dicamba DCGA DCS A Combined

Residues

Hinton, Caddo, OK; 2010 (OK1) 8 Undelinted

Cotton

seed 2.0

(2.2) 2 68 ND, ND [<0.02] ND, ND

[<0.02] 0.03, 0.03

[0.03] 0.03, 0.03

[0.03] <0.10,

<0.10 [<0.10]

3 8 0.33, 0.23

[0.28] ND, ND

[< 0.02] 0.05, 0.03

[0.04] 0.08, 0.03

[0.06] <0.49,

<0.31 [<0.40]

4 8 0.35 6 , 0.18 6

[0.27] ND, ND

[<0.02] 0.09, 0.05

[0.07] 0.11, 0.07

[0.09] <0.57,

<0.33 [<0.45]

5 8 0.23, 0.23

[0.23] ND, ND

[<0.02] 0.03, 0.04

[0.04] 0.02, 0.07

[0.04] <0 .30,

<0.37 [<0.33]

Dill City, Washita, OK; 2010 (OK2) 8 Undelinted

Cotton

seed 2.0

(2.2) 2 82 ND, ND [<0.02] ND, ND

[<0.02] 0.03, 0.02

[0.03] 0.03, (0.01)

[<0.02] <0.10,

<0.08 [<0.09]

3 7 0.07, 0.06

[0.06] ND, ND

[<0.02] ND, ND

[<0.02] (0.01 ),

(0.01) [<0.02] <0.13,

<0.12 [<0.13]

4 7 0.12, 0.16

[0.14] ND, ND

[<0.02] 0.02, 0.03

[0.03] 0.03, 0.03

[0.03] <0.19,

<0.24 [<0.22]

Gin byproducts 2 82 ND, ND [<0.0 4 ] (0.01),

(0.01) [<0.0 4 ] 0.17, 0.23

[0.20] 0.39, 0.46

[0.43] <0.67,

<0 .80 [<0.73]

4 7 3.17, 3.09

[3.13] (0.01),

(0.01) [<0.0 4 ] 0.49, 0.40

[0.45] 1.70, 1.85

[1.78] <5.52,

<5.50 [<5.51]

Elko, Barnwell, SC; 2010 (SC1) 2 Undelinted

Cotton

seed 2.0

(2.2) 2 99 ND, ND [<0.02] ND, ND

[<0.02] (0.01),

(0.01) [<0.02] (0. 01),

(0.01) [<0.02] <0.08,

<0.08 [<0.08]

105 ND, ND [<0.02] ND, ND

[<0.02] (0.01),

(0.01) [<0.02] (0.01),

(0.01) [<0.02] <0.08,

<0.08 [<0.08]

112 ND, ND [<0.02] ND, ND

[<0.02] (0.01),

(0.01) [<0.02] (0.01),

(0.01) [<0.02] <0.08,

<0.0 8 [<0.08]

119 ND, ND [<0.02] ND, ND

[<0.02] (0.01),

(0.01) [<0.02] (0.01),

(0.01) [<0.02] <0.08,

<0.08 [<0.08]

3 7 0.18, 0.06

[0.12] ND, ND

[<0.02] ND, 0.02

[<0.02] (0.01), 0.02

[<0.02] <0.24,

<0.12 [<0.18]

4 1 0.21, 0.31

[0.26] ND, ND

[<0.02] 0.02, 0.03

[0.03] 0.02, 0.03

[0.03] <0.28,

<0.39 [<0.34]

7 0.14, 0.09 [0.12] ND, ND

[<0.02] 0.06,

( 0.0 18)
[ < 0.04] 0.05, 0.02

[0.04] <0.27,

<0.15 [<0.21]

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.4.1/7.4.2/OPPTS 860.1500/OECD IIA 6.3.1, 6.3.2, 6.3.3 and IIIA 8.3.1, 8.3.2, 8.3.3 Crop Field T rial/ Residue Decline

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D40 838 4 /MRID No. 48 728703 Page 24 of 28 TABLE C. 3. Residue Data for Dicamba and its Metabolites from Cotton Field Trials

Trial ID (City, County, State; Year) Zone Com

modity 1 Total

Rate

lb a e /A (kg

ae/ ha ) TRT 2 PHI

(days) 3 Residues (ppm) [Average] 4, 5

Dicamba 5

- OH

Dicamba DCGA DCS A Combined

Residues

14 0.09, 0.07 [0.08] ND, ND

[<0.02] 0. 10 7 , 0.0 5 7

[0.05] 0.06, 0 .03

[0.05] <0.28,

<0.17 [<0.22]

21 0.04, 0.08 [0.06] ND, ND

[<0.02] 0. 11 7 , 0.0 4 7

[0.06] 0.09, 0.03

[0.06] <0.26,

<0.17 [<0.22]

Raymondvill e, Willacy, TX; 2010 (TX1) 6 Undelinted

Cotton

seed 2.0

(2.2) 2 43 ND, ND [<0.02] ND, ND

[<0.02] 0.02,

(0.019) [<0.02] 0.03, (0.01)

[<0.02] <0.09,

<0.08 [<0.09]

49 ND, ND [<0.02] ND, ND

[<0.02] (0.01), 0.02

[<0.02] 0.02, (0.01)

[<0.02] <0.08,

<0.08 [<0.08]

56 ND, ND [<0.02] ND, ND

[<0.02] (0.01),

(0.019) [<0.02] (0.01), 0.03

[<0.02] <0.08,

<0.09 [<0.08]

63 ND, ND [<0.02] ND, ND

[<0.02] (0.018),

(0.01) [<0.02] (0.016),

(0.01) [<0.02] <0.08,

<0.08 [<0.08]

3 7 0.58 6 , 0.19 6

[0.38] ND, ND

[<0.02] ND, ND

[<0.02] 0.07, (0.01)

[<0.04] <0.70,

<0.25 [<0.48]

6 Undelinted

Cotton

seed 2.1

(2.4) 4 1 0.28, 0.23 [0.25] ND, ND

[<0.02] 0.03, 0.03

[0.03] 0.03, 0.03

[0.03] <0.36,

<0.31 [<0.33]

7 0.19, 0.17 [0.18] ND, ND

[<0.02] 0.03, 0.02

[0.03] 0.09, 0.04

[0.06] <0.34,

<0.25 [<0.29]

14 0.07, 0.04 [0.05] ND, ND

[<0.02] 0.03, 0.02

[0.03] 0.04, 0.03

[0.04] <0.17,

<0.11 [<0.14]

21 0.03, 0.04 [0.03] ND, ND

[<0.02] 0.03, 0.03

[0.03] 0.03, 0.03

[0.03] <0.12,

<0.13 [<0.12]

Levelland, Hockley, TX; 2010 (TX2) 8 Undelinted

Cotton

seed 2.0

(2.2) 2 84 ND, ND [<0.02] ND, ND

[<0.02] 0.11, 0.11

[0.11] 0.23, 0.23

[0.23] <0.39,

<0.39 [<0.39]

3 6 0.84 6 , 1.31 6

[1.07] (0.01), ND

[<0.02] 0.07, 0.05

[0.06] 0.07 6 , 0.25 6

[0.16] <1.00,

<1.64 [<1.32]

4 6 1.30, 1.54

[1.42] ND, (0.01)

[<0.02] 0.11, 0. 17

[0.14] 0.26, 0.27

[0.27] <1.71,

<2.02 [<1.86]

Gin byproducts 2 84 ND, (0.01) [<0.04] ND, (0.01)

[<0.04] 0.42, 0.71

[0.57] 1.73, 1.43

[1.58] <2.34,

<2.31 [<2.33]

4 6 16.4, 13.4

[14.9] 0.04, (0.03)

[<0.04] 3.25, 1.56

[2.41] 6.02, 2.97

[4.50] <26.1,

<18.2 [<22.1]

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.4.1/7.4.2/OPPTS 860.1500/OECD IIA 6.3.1, 6.3.2, 6.3.3 and IIIA 8.3.1, 8.3.2, 8.3.3 Crop Field T rial/ Residue Decline

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D40 838 4 /MRID No. 48 728703 Page 25 of 28 TABLE C. 3. Residue Data for Dicamba and its Metabolites from Cotton Field Trials

Trial ID (City, County, State; Year) Zone Com

modity 1 Total

Rate

lb a e /A (kg

ae/ ha ) TRT 2 PHI

(days) 3 Residues (ppm) [Average] 4, 5

Dicamba 5

- OH

Dicamba DCGA DCS A Combined

Residues

Wolfforth, Lubbock,
TX; 2010 (TX3) 8 Undelinted

Cotton

seed 2.0

(2.2) 2 84 ND, ND [<0.02] ND, ND

[<0.02] 0.07, 0.04

[0.05] 0.10, 0.04

[0.07] <0.22,

<0.12 [<0.17]

3 6 1.97 6 , 0.80 6

[1.38] (0.01), ND

[<0.02] 0.07, 0.1 0

[0.09] 0.16, 0.16

[0.16] <2.24,

<1.09 [<1.66]

4 6 1.29, 1.08

[1.19] ND, ND

[<0.02] 0.08, 0.12

[0.10] 0.11, 0.16

[0.14] <1.51,

<1.39 [<1.45]

Gin byproducts 2 84 ND, ND [<0.04] (0.01),

(0.01) [<0.04] 0.32, 0.31

[0.32] 0.85, 0.49

[0.67] <1.30,

<0.91 [<1.11]

4 6 23.6, 22.4

[23.0] (0.03),

(0.03) [<0.04] 4.31, 3.97

[4.14] 6.29, 6.05

[6.17] <34.6,

<32.8 [<33.7]

Uvalde, Uvalde, TX; 2010 (TX4) 8 Undelinted

Cotton

seed 2.0

(2.2) 2 71 ND, ND [<0.02] ND, ND

[<0.02] (0.01),

(0.01)
[ <0.02] (0.01),

(0.01) [<0.02] <0.08,

<0.08 [<0.08]

3 6 0.86, 1.22

[1.04] ND, ( 0.0 16)

[<0.02] 0.02, 0.03

[0.03] 0.03, 0.04

[0.03] <0.93,

<1.31 [<1.12]

4 6 0.49, 0.44

[0.47] ND, ND

[<0.02] 0.03, 0.03

[0.03] 0.03, 0.04

[0.04] <0.58,

<0.53 [<0.55]

1 Each trial site was planted with Event MON 88701 cotton that is tolerant to dicamba herbicide and glufosinate

- ammonium

herbicide.

2 TRT 2: Applications with 4 lb ae/gal SL formulation made at preemergence, the 6

- leaf stage, and at the fir st w hite flower + 15 days stage; TRT 3: Applications with 4 lb ae/gal SL formulation made at preemergence, the first open boll stage, and at 7 days prior to harvest; TRT 4: Applications with 4 lb ae/gal SL formulation made at the 6

- leaf stage, the first whi te flower + 15

- days stage, the first open boll stage, and at 7 days prior to harvest; TRT 5: Applications with 5 lb ae/gal SL formulation made at the 6

- leaf stage, the first white flower + 15

- days stage, the first open boll stage, and at 7 days prior to h arvest

. 3 For the residue decline trials, the target PHI is bolded

. 4 ND = <LOD. The LODs for the analytes in undelinte d cotton seed were 0.0035, 0.0041, 0.0057 and 0.0089 ppm for DCSA, DCGA, 5

- hydroxydicamba and dicamba, respectively. The LOQ was 0.0 2 ppm for each analyte in undelinted cotton seed and 0.04 ppm in cotton gin byproducts

. Values between the LOQ and LOD are in parenthesis

. 5 Residues for individual analytes are presented as per se. Combined residues of dicamba, 5

- OH dicamba, DCGA, and DC SA are expressed in parent equivalents . R esidues of 5

- OH dicamba, DCSA, and DCGA were converted to parent equivalen ts by the study reviewer using molecular weight conversion factor s of 0.933, 1.068, and 0.991, respectively

. W hen calculating per trial

avera ges, the LOQ was used for values reported as ND or between the LOD and LOQ. NOTE TO EPA REVIEWER: DCGA is included in the combined residues as it is a major metabolite of dicamba.

6 Average of multiple analyses.

7 The data in Table 13 (p. 53) does not ma tch the raw data in the Master Summary Table (p. 361). It appears that the DCGA data in Table 13 was copied from the DCSA data. The study reviewer used the raw data for DCGA.

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.4.1/7.4.2/OPPTS 860.1500/OECD IIA 6.3.1, 6.3.2, 6.3.3 and IIIA 8.3.1, 8.3.2, 8.3.3 Crop Field T rial/ Residue Decline

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D40 838 4 /MRID No. 48 728703 Page 26 of 28 TABLE C.4. Summary of Residue Data from Cotton Crop Field Trials with Dicam ba.

Commodity Analyte Total App.

Rate

lb ae/A (kg ae/ ha ) PHI

(days) Residue Levels (ppm) 1

n Sample

Min. Sample

Max. L AFT 2 HAFT 2 Median Mean Std.

Dev.

TRT 2 (Applications at Preemergence, 6

- leaf stage, and first white flower + 15 days; EP: Clarity)

Undelinted

Cotton seed Dicamba 2.0 (2.2) 49

- 105 13 < 0.02 < 0.02 < 0.02 < 0.02 0.02 0.02 N/A

5

- OH Dicamba 13 < 0.02 < 0.02 < 0.02 < 0.02 0.02 0.02 N/A

DCGA 13 < 0.02 0.11 < 0.02 0.11 0.02 0.03 0.03

DCSA 13 < 0.02 0.23 < 0.02 0.23 0.02 0.04 0.06

Combined

Residues 13 < 0.0 8 < 0.39 < 0.08 < 0.39 0.08 0.12 0.09

Gin byproducts Dicamba 2.0 (2.2) 82

- 84 3 < 0.04 < 0.04 < 0.04 < 0.04 0.04 0.04 N/A

5

- OH Dicamba 3 < 0.04 < 0.04 < 0.04 < 0.04 0.04 0.04 N/A

DCGA 3 0.17 0.71 0.20 0.57 0.32 0.36 0.19

DCSA 3 0.39 1.73 0.43 1.58 0.67 0.89 0.61

Combined

Residues 3 < 0.67 < 2.34 < 0.73 < 2.33 1.11 1.39 0.84

TRT 3 (Applications at Preemergence, first open boll stage, and 7 days prior to harvest ; EP: Clarity)

Undelinted

Cotton seed Dicamba 2.0 (2.2) 6

- 8 13 0.06 1.97 0.06 1.38 0.65 0.64 0.43

5

- OH Dicamba 13 < 0.02 0.02 < 0.02 < 0.02 0.02 0.02 N/A

DCGA 13 < 0.02 0.10 < 0.02 0.09 0.02 0.03 0.02

DCSA 13 < 0.02 0.25 < 0.02 0.16 0.03 0.05 0.05

Combined

Residues 13 < 0.12 < 2.24 < 0.13 < 1.66 0.73 0.76 0.49

TRT 4 (Applicati ons at 6

- leaf

, first white flower + 15 days, first open boll , and 7 days prior to harvest ; EP: Clarity)

Undelinted

Cotton seed Dicamba 2.0

- 2.1 (2.2

- 2.4 ) 6

- 8 13 0.09 1.54 0.12 1.42 0.47 0.61 0.41

5

- OH Dicamba 13 < 0.02 < 0.02 < 0.02 < 0.02 < 0.02 < 0.02 N/A

DCGA 13 0.02 0.17 0.02 0.14 0.03 0.05 0.04

DCSA 13 0.02 0.27 0.02 0.27 0.06 0.08 0.07

Combined

Residues 13 < 0.15 < 2.02 < 0.21 < 1.86 0.58 0.76 0.49

Gin byproducts Dicamba 2.0 (2.2) 6

- 7 3 3.09 23.6 3.13 23.0 14.9 13.7 10.0

5

- OH Dicamba 3 < 0.0 4 0.04 < 0.04 < 0.04 0.04 0.04 N/A

DCGA 3 0.40 4.31 0.45 4.14 2.41 2.33 1.85

DCSA 3 1.70 6.29 1.78 6.17 4.50 4.15 2.22

Combined

Residues 3 < 5.50 34.6 < 5.51 < 33.7 22.1 20.4 14.2

TRT 5 (Applications at 6

- leaf

, first white flower + 15 days, first o pen boll , and 7 days prior to harvest ; EP: MON 11968 )

Undelinted

Cotton seed Dicamba 2.0 (2.2) 7

- 8 4 0.17 0.72 0.20 0.62 0.41 0.41 0.23

5

- OH Dicamba 4 < 0.02 < 0.02 < 0.02 < 0.02 0.02 0.02 N/A

DCGA 4 0.02 0.08 0.02 0.06 0.03 0.04 0.02

DCSA 4 0.02 0.17 0.02 0.12 0.04 0.06 0.04

Combined

Residues 4 < 0.24 < 1.00 < 0.29 < 0.82 0.49 0.52 0.26

1 Except for sample min/max, values reflect per trial averages; n = no. of field trials. For calculation of median, mean, and standard deviation, the LOQ (0.0 2 ppm each analyte in undelinted cotton seed and 0.04 ppm for each analyte in cotton gin

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.4.1/7.4.2/OPPTS 860.1500/OECD IIA 6.3.1, 6.3.2, 6.3.3 and IIIA 8.3.1, 8.3.2, 8.3.3 Crop Field T rial/ Residue Decline

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D40 838 4 /MRID No. 48 728703 Page 27 of 28 byproducts ) was used for any results reported as <LOQ in Table C.3. Combined residues of dicamba, 5

- OH dicamba, DCGA, and DCSA are expressed in parent equivalents. Ind ividual analyte results are reported as per se. NOTE TO EPA REVIEWER: DCGA is included in the combined residues as it is a major metabolite of dicamba. N/A = Not applicable.

2 LAFT = lowest

- average

- field

- trial; HAFT = highest

- a verage

- f ield

- t rial.



## D. CO NCLUSION

The submitted cotton field trial data are adequate and reflect the use of three to four broadcast applications of the 4 lb ae/gal SL formulation of dicamba (formulated as the diglycolamine salt),

at a total rate of 2.0

- 2.1 lb ae/A (2.2

- 2.4 kg ae/ ha). Following three applications at pre

emergence, the 6

- leaf stage, and first white flower + 15 days (TRT2)

, combined residues (and per trial averages) of dicamba, 5

- OH dicamba, DCGA, and DCSA (expressed in parent equivalents) were <0.08

- <0.39 (<0.08

- < 0 .39) ppm in/on undelinted cotton seed collected at PHIs

of 49

- 105 days and <0.67

- <2.34 (<0.73

- <2.33) ppm in/on gin byproducts collected at PHIs of

82

- 84 days. Following three applications at pre

- emergence, the first open boll stage, and 7 days prior to ha rvest (TRT 3) combined residues (and per trial averages) of dicamba, 5

- OH dicamba, DCGA, and DCSA (expressed in parent equivalents) were <0.12

- <2.24 (<0.13

- <1.66) ppm i n/on

undelinted cotton seed collected at a 6

- 8

- day PHI. Following four applications at the 6

- leaf stage, the first white flower + 15 days, the first open boll, and 7 days prior to harvest (TRT 4)

, combined residues (and per trial averages) of dicamba, 5

- OH dicamba, DCGA, and DCSA (expressed in parent equivalents) were <0.15

- <2.02 (<0.21

- <1.8 6) ppm in/on undelinted cotton

seed collected at a 6

- 8

- day PHI and <5.50

- 34.6 (<5.51

- <33.7) ppm in/on gin byproducts

collected at a 6

- 7

- day PHI. Corresponding combined residues (and per trial averages) in side

- by

side trials treated with the 5 lb ae/gal S L formulation of dicamba formulated as the monoethanolamine salt ( TRT 5 ) were <0.24

- <1.00 (<0.29

- <0.82) ppm in/on undelinted cotton seed. Average combined residues in the diglycolamine salt formulation trials were slightly higher (approximately 1.5x) than the corresponding residues in the monoethanolamine salt

formulation trials.

Treatment plots 3 and 4 showed the highest average combined residues (0.76 ppm), followed closely by treatment plot 5 (average of 0.52 ppm). Treatment plot 2 had the lowest av erage combined residues (average of 0.12 ppm). Lower residues from treatment plot 2 is consistent with the earlier application timings at these plots.

For the TRT 2 residue decline samples, average residues of dicamba, 5

- OH dicamba, DCGA,

and DCSA in/o n undelinted cotton seed were at or <LOQ in both studies; therefore, residue decline could not be assessed. For the TRT 4 residue decline samples, average residues of

dicamba decreased in/on undelinted cotton seed with increasing PHI. Average residues of DCGA and DCSA increased with increasing PHI in the SC1 trial, and remained relatively

constant in the TX1 trial. Residues of 5

- OH dicamba were <LOQ in both studies; therefore,

residue decline could not be assessed.

An acceptable method was used for re sidue quantitation, and a dequate data were submitted to support sample storage intervals and conditions.

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company DACO 7.4.1/7.4.2/OPPTS 860.1500/OECD IIA 6.3.1, 6.3.2, 6.3.3 and IIIA 8.3.1, 8.3.2, 8.3.3 Crop Field T rial/ Residue Decline

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D40 838 4 /MRID No. 48 728703 Page 28 of 28 

## E. REFERENCES

None.

F. DOCUMENT TRACKING

Petition Number(s): 2F8067 DP Barcode(s): 408384

PC Code: 029801, 029802, 029806, 128931, 128944, & 129043

Template Version June 2005.

### Dicamba/MON 11900 / P

C Code 1 2893 1 / Monsanto Compa ny
DA CO 7.4.5 / 0PPT S 860.1520 / 0ECD llA 6.5.4 and lllA 8.5

Processed Food and Feed

- Dicamba-Tolerant Cotton (Eve nt MON 88701)



### Primar y Eva l uator

Versar, In c.



## - ;;f 

### Approved by 

# ( J,-r;-v-**4a.~<-~** Peter Savoia, Chemist, OCSPP-HED-RAB VN H Date: 11/15 / 2012

Date: 04 / 22 / 2013

Note: This DER wa s origina ll y prepared under contract by Versar. In c. (6850 Versar Center. Springfield. V /\ 22151: s ubmitted

I 1/15112 )

. The DER ha s been reviewed b y the Health Elkcts Division (I JED ) and revi s ed t o renect current Office of Pe s ticide

Pr og ram s ( OPP ) p o li c i es.



### STU DY REPORTS:

48728703 Maher, D. ; Foster. J. (201 1 ) Magnitude of Oicamba Residues in Cotton Raw Agricultural and Processed Commodities following Applications of Dicamba-Ba se d

Fo rmulation s to MON 8870 l. Proj ect N umb e r: RE G/I 0/062/0CR

, MSL0024072. Unpubli s hed study prepared by Monsanto Company, Carringers, Inc. and GL P Technolog i es. 603 p.



### EXECUT I VE SUMMARY:

Monsanto has s ubmitted a processi n g study for dicamb a on dicamba-tolerant cotton, identified as

Event MON 88701 (formerly GH_S26695). Event MON 88701 cotton expresses a modified

dicamba mono-o xyge nase (DM O ) gene derived from 

### S t enotrophomonas maltophilia and the bialaphos resistance (BAR) gene isolated from 

### Streptomyces hygro scopicus. Th e ex pression of

D MO confers dicamba tolerance and the express i on of BAR confer s g lu fos in ate tolerance to

Event MON 8870 l cotton. Two trials were conducted in MO and TX during the 20 I 0 growing

season.

Eac h trial conta in ed two treat ed p l ots that fo ll owed one of two treatm e nt types: thr ee broadcast

app li cations applied at pre-em e rgence
, 6-leaf

, and 

## pt white flower + 1 5 days [TRT 2] or four

foliar broadcast applications applied at 6-leaf, l 5 1 

### white flower + 15 days

, first open boll, a n d 7 days prior to har vest (TRT 4]. Applications were made at the field rate using a 4 lb ae / gal wacer

so luble concentrate (SL) fo rmul atio n of dicamba diglycolamine sa l t (C l arity H erbicide; MON
54140)

. T h e total application rate for both tr ea tm ent types was 2.0 lb ae / A (2.2 kg ae/ha)

. For TRT 2

, application l was applied at 1.0 lb ae / A ( 1.1 kg ae/ha) and applications 2 and 3 were applied at 0

. 50 lb ae / A/app li cation (0.56 kg ae/ha/applicacion). Re t reatment interva l s ( RTi s)

were 24

- 49 days between the first and second applications and 42-47 days between the second

and third applications. For TRT 4

, all four app licati ons we r e applied at 0.49-0.51 l b

ae / A/ap plication (0.55

- 0.57 kg ae / ha/applic a tion). RTi s were 42-47 days between the first and

second applications, 36

- 42 days between the seco nd and third applications. and 29-30 days

between the third and fourth app li cations.

All app li cations we r e m ade u s ing ground equipment in sp ray volumes of 1 9. 7-20.5 gal/ A (184-

192 I / ha) and included a noni o nic surfactant (N IS ) and a mmonium su lfate (AMS) at a ll trial si t es. Samples of undelinted cotton seed were harvested using a mechanical picker at maturity (pre har vest inter va l [PHI] of 7 I -79 day s for TRT 2 and 6- 7 days for TRT 4 ). Samp l es of cotton

0408384 / MRJD No. 48728703 Pag e I of 1 5

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company

DACO 7.4.5/OPPTS 860.1520/OECD IIA 6.5.4 and IIIA 8.5

Processed Food and Feed

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D408 384 /MRID No. 48 728703 Page 2 of 15 

## seed were processed into meal, hulls

, alkali

- refined oi l, and RBD oil by GLP Technologies (Navasota, TX) using procedures simulating commercial processing.

Samp les were analyzed for residues of dicamba and metabolite s 5

- hydroxydicamba (5

- OH dicamba), 3,6

- dichlorosalicylic acid (DCSA), and 3,6

- dichlorogentisic acid (DCGA) using a high performance liquid chromatography method with tandem mass spectrometry detection (LC /MS/MS). The limit of quantitation (LOQ, equivalent to the lowest level of method validation; LLMV) was 0.0 2 ppm for all analytes in undelinted cotton seed (RAC) and the processed commodities . The method was adequate for data validation based on acce ptable

method and concurrent recovery data. The fortification levels used in concurrent method recovery were adequate to bracket expected residue levels. Concurrent recoveries were corrected for apparent residues in controls; residues in treated samples were not corrected for

residues in controls. We note that the analytical method does not specify conversion of metabolite residues to parent equivalents; therefore, quantifiable residues of 5

- OH dicamba, DCSA, and DCGA were converted to parent equivalents by the study reviewer using a molecular weight conversion factor of 0.933, 1.068, and 0.991, respectively

.

Samples were stored frozen (~

- 20 °C) from harvest /processing to analysis for 152

- 196 days (5.0

- 6.4 months) for cotton (RAC) and 19

- 40 days (0.6

- 1.3 months) for the processed commodities. All samples were analyzed on the day of extraction. Processing took place within 118

- 169 day s of harvest. A storage stability study for dicamba, DCGA , and DCSA on undelinted cotton seed was submitted in conjunctio n with the field trial study (refer to the 860.1380 der for MRID 48728704). The stability of 5

- OH dicamba was not determined concurrently because it was not

found in undelinted cotton seed. These data are acceptable to support the stora ge conditions and

durations of the undelinted cotton seed s amples from the submitted field trials . Storage stability data for the processed commodities was not provided; however, storage stability data is not required as the processed commodities were generally analyzed wi thin 30 days of processing.

Following three or four applications of the 4 lb ae/gal SC formulation at a total rate of 2 lb ae/A (2.2 kg ae/ha)

, using PHIs of 71

- 79 days and 6

- 7 days, respectively, the combined residues of dicamba, 5

- OH dicamba, DCGA, and DCSA (in parent equivalents) were below the LOQ ( < 0.08)

- <0.78 ppm in/on undelinted cotton seed (RAC), <0.081

- <0.79 ppm in hulls, below the LOQ ( < 0.08)

- <0.28 ppm in meal, and below the LOQ (<0.08 ppm) in alkali refined oil and RBD

oil.

The processing dat a indicate that the combined residues do not concentrate in hulls, meal, alkali refined oil, or RBD oil (average processing factors of 0. 86 x, 0. 49 , 0. 1 1x, and 0.11x, respectively ).

T he processing factors calculated in this study for combined residues of dicamba, 5

- OH dicamba, DCGA, and DCSA in cotton were less than the theoretical concentration factor of 3.8x for hulls, 2.2x for meal, and 6.3x for oil (based separation of components; OPPTS 860.1520, Table 3).

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company

DACO 7.4.5/OPPTS 860.1520/OECD IIA 6.5.4 and IIIA 8.5

Processed Food and Feed

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D408 384 /MRID No. 48 728703 Page 3 of 15 

## STUDY/WAIVER ACCEPTABILITY/DEFICIENCIES/CLA RIFICATIONS :

Under the conditions and parameters used in the study, the field trial residue data are classified as scientifically acceptable. The acceptability of this study for regulatory purposes is addressed in the forthcoming U.S. EPA Residue Chemist ry Summary Document D408 384

.

COMPLIANCE :

Signed and dated Good Laboratory Practice (GLP), Quality Assurance

, and D ata Confidentiality

statements were provided. No deviations from regulatory requirements were reported which

would ha ve an impact on the validity of the study.

A. BACKGROUND INFORMATION

Dicamba (3,6

- dichloro

- 2

- methoxybenzoic acid is a selective benzoic acid herbicide registered for the control of weeds prior to their emergence. The Dicamba Reregistration Eligibility

D ecision (RED) was issued December 2005. The chemical structure and nomenclature of

dicamba and its metabolite s 5

- OH dicamba , DSCA, and DCGA are presented in Table A.1. T he physicochemical properties of the technical grade of dicamba acid are presented in Table A.2.

TABLE A.1. Test Compound Nomenclature.

Compound

Common name Dicamba

Company experimental name MON 11900

IUPAC name 3,6

- dichloro

- o

- anisic acid or 3,6

- dichloro

- 2

- methoxybenzoic acid

CAS name 3,6

- dichloro

- 2

- methoxybenzoic acid

CAS regi stry number 1918

- 00

- 9 (dicamba acid), 104040

- 79

- 1 (diglycolamine salt), or 53404

- 28

- 7 (monoethanolamine salt)

End

- use product Clarity® Herbicide: SL formulation containing 4 lb ae/gal

Compound

Common name 5

- Hydroxy

- dicamba
Company experimental name 5

- OH dicamba

IUPAC/CAS name 2,5

- dichloro

- 3

- hydroxy

- 6

- methoxybenzoic acid

CAS registry number 7600

- 50

- 2

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company

DACO 7.4.5/OPPTS 860.1520/OECD IIA 6.5.4 and IIIA 8.5

Processed Food and Feed

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D408 384 /MRID No. 48 728703 Page 4 of 15 TABLE A.1. Test Compound Nomenclature.

Compound

Common name DCSA ; 3,6

- dichlorosalicylic acid

Company experimental name MON 52708

IUPAC/CAS name 3,6

- dichloro

- 2

- hydroxybenzoic acid

CAS registry number 3401

- 80

- 7

Compound

Common name DCGA ; 3,6

- dichlorogentistic acid

Company experimental name MON 52724

IUPAC/CAS name 2,5

- dichloro

- 3,6

- dihydroxybenzoic acid

CAS registry number 18688

- 01

- 2



### TABLE A.2. Physicochemical Properties of the Technical Grade Test Compound: Dicamba.

Parameter Value Reference

Melting point 114

- 116  C (PAI)

90

- 100  C (87% TGAI) Residue Chemistry Chapter of the Dicamba RED, DP# 317699, 12/20/05, C. Olinger pH 2.5

- 3.0 (87% TGAI)

Density 1.57 g/mL at 25  C (87% TGAI)

Water solubility 0.5 g/100 mL at 25  C (PAI)

Solvent solubility g/100 mL at 25  C (PAI)

dioxane 118.0

ethanol 92.2 isopropyl alcohol 76.0 methylene chloride 26.0

acetone 17.0

toluene 13.0 xylene 7.8 heavy aromatic naphthalene 5 .2

Vapor pressure 3.4 x 10

- 5 mm Hg at 25  C (PAI)

Dissociation constant, pK a 1.97 (PAI)

Octanol/water partition coefficient, Log(K OW ) 0.1 (PAI)

UV/visible absorption spectrum neutral: 511 (275 nm) acidic (pH 0

- 1): 1053 (281 nm) basic (pH 13

- 14): 469 (274 nm)

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company

DACO 7.4.5/OPPTS 860.1520/OECD IIA 6.5.4 and IIIA 8.5

Processed Food and Feed

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D408 384 /MRID No. 48 728703 Page 5 of 15 

## B. EXPERIMENTAL DESIGN

B.1. Application and Crop Information

In two crop field trial s conducted in MO and TX

, one untreated and two treated plots were planted with dicamba

- tolerant cotton ( Event MON 88701) containing a dicamba mono

oxygenase (DMO) gene to confer tolerance to dicamba and a bialophos resistance ( BAR ) gene to

conf er tolerance to the glufosinate

. Each treated plot followed one of two treatment types: thre e broadcast applications applied at pre

- emergence, 6

- leaf, and 1 s t 

## white flower + 15 days [TRT 2] or four foliar broadcast applications applied at 6

- leaf, 1 st 

## white flower + 15 days, first open boll, and 7 days prior to harvest [TRT 4]. Applications were made at the field rate using a 4 lb ae/gal

SL formulation of dica mba diglycolamine salt (Clarity Herbici de; MON 54140). The total application rate for both treatment types was 2.0 lb ae/A (2.2 kg ae/ha). For TRT 2 , application 1 was applied at 1.0 lb ae/A (1.1 kg ae/ha) and applications 2 and 3 were applied at 0.50 lb

a e/A/application (0.56 kg ae/ha/application). RTIs were 24

- 49 days between the first and second applications and 42

- 47 days between the second and third applications. For TRT 4

, all four applications were applied at 0.49

- 0.51 lb ae/A/application (0.55

- 0.5 7 kg ae/ha/application)

. RTIs

were 42

- 47 days between th e first and second applications, 36

- 42 days between the second and third applications , and 29

- 30 days between the third and fourth applications.

All a pplications were made using ground equipment i n spray volumes of 1 9

. 7

- 20.5 gal/A (184

- 192 l/ha) and included an NI S and an AMS at all trial sites. Samples of undelinted cotton seed

were harvested using a mechanical picker at maturity (PHI of 71

- 79 days for TRT2 and 6

- 7 days

for TRT4 ). Study use patt ern data are reported in Table B.1.

TABLE B.1

. Study Use Pattern.

Location City, County, State;Year (Trial ID) EP 1 TRT Application Tank Mix/ Adjuvants 3 Harvest

Procedures Method; Timing Volume gal/A (l/ha) Rate

lb a e /A (kg ae/A) RTI 2

(days) Tot al

Rate

lb a e /A (kg ae/A)

Fisk, Butler, MO; 2010 (MO1) 4 lb

ae/gal

SL 2 1. Broadcast ; Preemergence 20.0 (187) 1.0

(1.1) -- 2.0

(2.2) NIS + AMS Mechanical picker
2. Foliar broadcast; 6

- leaf 20.0 (187) 0.50

(0.56) 29

3. Foliar broadcast; 1 st WF + 15 days 20.0 (187) 0.50

(0.56) 42

4 lb ae/gal

SL 4 1. Foliar broadcast; 6

- leaf 20.3 (190) 0.51

(0.57) -- 2.0

(2.2) NIS + AMS Mechanical picker
2. Foliar broadcast; 1 st WF + 15 days 20.1 (188) 0.50

(0.56) 42

3. Foliar broadcast; 1 st open boll 20.0 (187) 0.50

(0.56) 42

4. Foliar broadcast; 7 days prior to harvest 20.0 (187) 0.50

(0.56) 30

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company

DACO 7.4.5/OPPTS 860.1520/OECD IIA 6.5.4 and IIIA 8.5

Processed Food and Feed

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D408 384 /MRID No. 48 728703 Page 6 of 15 TABLE B.1

. Study Use Pattern.

Location City, County, State;Year (Trial ID) EP 1 TRT Application Tank Mix/ Adjuvants 3 Harvest

Procedures Method; Timing Volume gal/A (l/ha) Rate

lb a e /A (kg ae/A) RTI 2

(days) Tot al

Rate

lb a e /A (kg ae/A)

Uvalde, Uvalde, TX; 2010 (TX4) 4 lb

ae/gal

SL 2 1. Broadcast ; Preemergence 20.2 (189) 1.0

(1.1) -- 2.0

(2.2) NIS + AMS Mechanical picker

2. Foliar broadcast; 6

- leaf 20.1 (188) 0.50

(0.56) 49

3. Foliar broadcast; 1 st WF + 15 days 19.9 (186) 0.50

(0.56) 47

4 lb ae/gal

SL 4 1. Foliar broadcast; 6

- leaf 20.0 (187) 0.50

(0.56) -- 2.0

(2.2) NIS + AMS Mechanical picker

2. Foliar broad cast; 1 st WF + 15 days 19.7 (184) 0.49

(0.55) 47

3. Foliar broadcast; 1 st open boll 20.5 (192) 0.49

(0.55) 36

4. Foliar broadcast; 7 days prior to harvest 20.2 (189) 0.51

(0.57) 29

1 EP = E nd

- use Product ; Clarity [water soluble concentrat e (SL) formulation containing 480 g/L dicamba formulated as the diglycolamine salt (4 lb ae/gal) ]

. 2 Retreatment Interval.

3 NIS = nonionic surfactant. AMS = ammonium sulfate.



## B.2. Sample Handling and Processing Procedures

Single bulk samples of undelin ted cotton seed (~19

- 32 kg) from the control and treated plots were collected and ginned at the field sites within 3 hours of harvest . Samples were placed in frozen storage within 1.0 hour of collection. The samples were stored frozen for 5

- 25 days after collection then shipped frozen via freezer truck to the processing facility, GLP Technologies (Navasota, TX). Cotton seed samples were stored frozen (~<

- 12 °C) at GLP Technologies until processing and were processed within 118

- 1 69 days of harvest. Sampl es from both trials were processed into meal, hulls, alkali

- refined oil , and RBD oil using procedures simulating commercial processing. Samples were stored frozen after processing and were shipped frozen

within 6

- 20 days after processing via FedEx to Mons anto Company ( St. Louis, MO ) for residue analysis. Samples were maintained frozen (~

- 20 °C) at the analytical laboratory. In preparation for analysis, samples (except oil) were milled in a cryomill, which pulverizes samples into a fine powder at cryogeni c temperatures.

The petitioner submitted adequate descriptions of the processing procedures including material

balance summaries. The cotton processing procedure is summari zed below in Figure 1, which

w as copied without alteration from MRID 48728703

.

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company

DACO 7.4.5/OPPTS 860.1520/OECD IIA 6.5.4 and IIIA 8.5

Processed Food and Feed

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D408 384 /MRID No. 48 728703 Page 7 of 15 

## FIGURE 1. Processing Flowchart for Cotton

.

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company

DACO 7.4.5/OPPTS 860.1520/OECD IIA 6.5.4 and IIIA 8.5

Processed Food and Feed

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D408 384 /MRID No. 48 728703 Page 8 of 15 

## B.3. Analytical Methodology

Samples were analyzed for residues of dicamba and the metabolites DCGA, DCSA, and 5

- hydoxy dicamba using a LC/MS/MS method from Monsanto Company: Analytical Method AG

ME

- 13 81

- 01. For a complete description of the LC/MS/MS method, refer to the 860.1340 DER

for MRID 48728702

. We note that the analytical method does not specify conversion of metabolite residues to parent equivalents; therefore, quantifiable residues of 5

- OH d icamba, DCSA, and DCGA were converted to parent equivalents by the study reviewer using molecular weight conversion factor s of 0.933, 1.068, and 0.991, respectively.

Briefly, samples were extracted with acetonitrile : water ( 4 0: 6 0, v:v). An aliquot of the extract

was hydrolyzed in 1 N HCl at 95 °C in an oven

. The hydrolysate was partitioned with ethyl acetate:isooctane (20:80, v:v). Water was added to the organic phase and the sample was concentrated by evaporation until only the aqueous solution remained . The samples were acidified and analyzed using LC/MS/MS.

The LOQ, determined as the LLMV, was 0.02 ppm for each analyte in each cotton matrix

. The

0.020 ppm fortification level was used to statistically determine the LODs and LOQs for the

four analytes of the method in undelinted cotton seed (RAC)

. The LODs for the analytes in

undelinte d cotton seed were 0.0035, 0.0041, 0.0057 and 0.0089 ppm for DCSA, DCGA, 5

- hydroxydicamba and dicamba, respectively. The LOQs for the analytes in undelinte d cotton
seed were 0.0105, 0.0121, 0.0171 and 0.0266 ppm for DCSA, DCGA, 5

- hydroxydicamba and dicamba, respectively (reported as analyte found per se). The LODs and LOQs were no t statistically determined in the processed fractions

.

The analytical method was validated prior to and in conjunction with the analysis of the

submitted samples

.

C. RESULTS AND DISCUSSION

Sample storage conditions and dura tions for samples of cotton and processed commodities are reported in Table C.2. Samples were stored under frozen (

- 20 °C) at the analytical laboratory

from harvest

/processing to analysis . Processing took place within 118

- 169 day s of harvest. Sample analyses were completed within 152

- 196 days (5.0

- 6.4 months) for cotton (RAC) and

19

- 40 days (0.6

- 1.3 months) for processe d commodities. All samples were analyzed on the day

of extraction. A storage stability study for dicamba, DCGA , and DCSA on cotton was submitted in conjunction with the field trial study (refer to the 860.1380 der for MRID 48728704 ).

Residues of dicamba , DCGA, and DCSA were stable during freezer storage for up to 277 days (9 months) in undelinted cotton seed. The stability of 5

- OH dicamba was not determined concurrently because it was not found in undelinted cotton seed. These data are acceptable to

su pport the stora ge conditions and durations of the undelinted cotton seed s amples from the

submitted field trials . Storage stability data for the processed commodities was not provided; however, storage stability data is not required as the processed commo dities were generally analyzed within 30 days of processing.

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company

DACO 7.4.5/OPPTS 860.1520/OECD IIA 6.5.4 and IIIA 8.5

Processed Food and Feed

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D408 384 /MRID No. 48 728703 Page 9 of 15 

## Method validation and concurrent method recovery data for the LC/MS/MS method are presented in Table C.1. The LC/MS/MS method used to analyze cotton samples for residues of dicamba, 5

- OH dicamb a, DCGA, and DCSA was adequate for data collection based on acceptable method and concurrent method recovery data. Concurrent recoveries were generally within the acceptable range of 70

- 120% in cotton and processed commodity samples fortified

with dicamba , 5

- OH dicamba, DCGA, and DCSA each at 0.0 2

- 0.200 ppm. The fortification levels were adequate to bracket residues found in treated samples. Apparent residues of dicamba, 5

- OH dicamba, DCGA, DSGA were below the LOQ (<0.02 ppm) in/on all samples of

untreat ed undelinted cotton seed (RAC), hulls, meal, alkali refined oil, and RBD oil

. Concurrent recoveries were corrected for apparent for residues in controls; residues in treated samples were not corrected for residues in controls.

Residue data from the cott on processing study are reported in Table C.3

. Following three or four applications of the 4 lb ae/gal SC formulation at a total rate of 2 lb ae/A, using PHIs of 71

- 79 days and 6

- 7 days, respectively, the combined residues of dicamba, 5

- OH dicamba, DCGA, and DCSA (in parent equivalents) were below the LOQ ( < 0.08)

- <0.78 ppm in/on undelinted cotton seed (RAC), <0.081

- <0.79 ppm in hulls, below the LOQ ( < 0.08)

- <0.28 ppm in meal, and below the LOQ (<0.08 ppm) in alkali refined oil and RBD oil.

The processing data indicate that the combined residues do not concentrate in hulls, meal, alkali refined oil, or RBD oil (average processing factors of 0.86x, 0.49, 0.11x, and 0.11x, respectively ).

T he processing factors calculated in this study for combined residue s of dicamba, 5

- OH dicamba, DCGA, and DCSA in cotton were less than the theoretical concentration factor of 3.8x for hulls, 2.2x for meal, and 6.3x for oil (based separation of components; OPPTS 860.1520, Table 3).

T ABLE C.1. Summary of Method and Concu rrent Recoveries of Dicamba and its Metabolites from

Cotton Matrices

Matrix Analyte Spike

Level (ppm) Sample Size (n) Recoveries (%) 1 Mean ±

Std. Dev. (%) 2

Method Validation Recoveries

Undelinted

Cotton seed (RAC) 3 Dicamba 0.005 5 187, 73, 172, 105, 102 128 ± 49

0.010 5 129, 149, 93, 78, 125 115 ± 29

0.020 5 119, 100, 123, 101, 124 113 ± 12

0.200 5 94, 96, 95, 97, 100 96.1 ± 2.2

10.0 5 68, 91, 91, 100, 98 89.6 ± 13

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company

DACO 7.4.5/OPPTS 860.1520/OECD IIA 6.5.4 and IIIA 8.5

Processed Food and Feed

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D408 384 /MRID No. 48 728703 Page 10 of 15 T ABLE C.1. Summary of Method and Concu rrent Recoveries of Dicamba and its Metabolites from

Cotton Matrices

Matrix Analyte Spike

Level (ppm) Sample Size (n) Recoveries (%) 1 Mean ±

Std. Dev. (%) 2

Undelinted

Cotton seed (continued) 5

- OH

Dicamba 0.005 5 68, 56, 101, 84, 105 82.9 ± 21

0.010 5 106, 97, 76, 110, 82 94.1 ± 15

0.020 5 96, 98, 115, 117, 114 108 ± 10

0.200 5 98, 104, 105, 104, 101 102 ± 3.0

10.0 5 69, 90, 79, 93, 104 87.1 ± 13

DCGA 0.005 5 140, 115, 88, 98, 129 114 ± 22

0.010 5 90, 108, 96, 79, 104 95.6 ± 11

0.020 5 81, 98, 93, 77, 86 86.9 ± 8.5

0.200 5 90, 87, 88, 84, 96 88.9 ± 4.3

10.0 5 73, 96, 89, 94, 90 88.6 ± 8.9

DCSA 0.005 5 88, 95, 90, 134, 98 101 ± 19

0.010 5 70, 84, 102, 96, 98 89.8 ± 13

0.020 5 100, 82, 97, 97, 95 94.0 ± 6 .9

0.200 5 95, 100, 105, 96, 101 98.9 ± 4.0

10.0 5 81, 92, 95, 93, 97 91.6 ± 6.4

Meal Dicamba 0.0 20 5 102, 101, 98, 111, 108 104 ± 5.3

0. 2 00 5 97, 99, 96, 100, 94 96.9 ± 2. 4

5

- OH

Dicamba 0.0 2 0 5 72, 108, 92, 77, 92 88.1 ± 1 4

0. 2 00 5 98, 94, 88, 99, 90 93.4 ± 4.9

DCGA 0.0 2 0 5 99, 108, 102, 103, 102 102 ± 3.3

0. 2 00 5 88, 86, 85, 88, 85 86 .0 ± 1.5

DCSA 0.0 2 0 5 101, 96, 100, 90, 98 96 .5 ± 4. 4

0. 2 00 5 82, 84, 86, 91, 89 86.2 ± 3. 8

Hulls Dicamba 0.020 5 97, 100, 106, 99, 99 100 ± 3.4

0.200 5 97, 101, 102, 100, 99 100 ± 2.1

5

- OH

Dicamba 0.020 5 87, 78, 106, 89, 97 91.2 ± 10

0.200 5 114, 107, 117, 99, 105 108 ± 7.4

DCGA 0.020 5 116, 117, 115, 121, 112 116 ± 3.4

0.200 5 89, 86, 86, 84, 82 85.2 ± 2.4

DCSA 0.020 5 109, 102, 94, 97, 96 99.5 ± 6.1

0.200 5 96, 102, 92, 100, 99 97.4 ± 3.9

Alkali

Refined Oil Dicamba 0.020 5 99, 96, 97, 99, 98 97.6 ± 1.2

0.200 5 99, 100, 105, 99, 97 100 ± 2.8

5

- OH

Dicamba 0.020 5 95, 101, 79, 92, 88 90.8 ± 8.0

0.200 5 110, 103, 88, 110 , 100 102 ± 9.0

DCGA 0.020 5 100, 102, 99, 103, 113 103 ± 5.7

0.200 5 97, 89, 92, 92, 91 91.8 ± 2.8

DCSA 0.020 5 102, 93, 90, 93, 100 95.2 ± 5.1

0.200 5 103, 106, 100, 104, 105 103 ± 2.1

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company

DACO 7.4.5/OPPTS 860.1520/OECD IIA 6.5.4 and IIIA 8.5

Processed Food and Feed

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D408 384 /MRID No. 48 728703 Page 11 of 15 T ABLE C.1. Summary of Method and Concu rrent Recoveries of Dicamba and its Metabolites from

Cotton Matrices

Matrix Analyte Spike

Level (ppm) Sample Size (n) Recoveries (%) 1 Mean ±

Std. Dev. (%) 2

Concurrent Recoveries

Undelinted

Cotton seed Dicamba 0.02 3 78. 1 , 118, 105 10 0 ± 20

0. 20 3 91. 9 , 103, 10 8 10 1 ± 8.3

5

- OH

Dicamba 0.02 2 85.5, 96.5 91.0

0. 20 2 90.0, 89.5 89.8

DCGA 0.02 2 108

, 119 113

0. 20 2 109 , 114 112

DCSA 0.02 2 84.9, 100 92.4

0. 20 2 96.4 , 98.7 97.6

RBD Oil Dicamba 0.0 2 2 1 15

, 110 112

0. 20 2 106

, 105 105

5

- OH

Dicamba 0.0 2 2 119

, 101 110

0. 20 2 98.5, 108 103

DCGA 0.0 2 2 112

, 106 109

0. 2 0 2 97.0, 89.5 93.3

DCSA 0.0 2 2 108

, 123 115

0. 2 0 2 104, 101 103

Alkali

Refined Oil Dicamba 0.02 2 103

, 109 106

0.20 2 1 01, 104 102

5

- OH

Dicamba 0.02 2 94.5, 75.5 85.0

0.20 2 92.0, 114 103

DCGA 0.02 2 120, 119 120

0.20 2 99.5, 99.0 99.3

DCSA 0.02 2 113 , 97.5 105

0.20 2 109, 105 107

Hulls Dicamba 0.02 3 109, 120 , 101 110 ± 9.3

0.20 3 79.0, 107, 103 96.4 ± 15

5

- OH

Dicamba 0.02 2 87.5, 99.5 93.5

0.20 2 84.5, 91.5 88.0

DCGA 0.02 2 112, 106 109

0.20 2 106, 104 105

DCSA 0.02 2 95. 9 , 96.4 96.1

0.20 2 98. 1 , 97. 2 97.6

Meal Dicamba 0.02 2 98.8, 114 106

0.20 2 101, 103 102

5

- OH

Dicamba 0.02 2 100 , 79.0 89.5

0.20 2 91.5, 92.5 92.0

DCGA 0.02 2 104

, 104 104

0.20 2 111

, 109 110

DCSA 0.02 2 79. 8 , 87. 4 83.6

0.20 2 94.1, 105 99.8

1 The concurrent recoveries were corrected for apparent residues in the unfortified control samples.

2 Standard deviation is not applicable for sample size s n < 3 samples.

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company

DACO 7.4.5/OPPTS 860.1520/OECD IIA 6.5.4 and IIIA 8.5

Processed Food and Feed

– Dicamba

- Tolerant Cotton ( Event MON 88701)

D408 384 /MRID No. 48 728703 Page 12 of 15 3 Recoveries for method validation undelinted cotton seed are also presented in the crop field trial DER (refer to 48728703.DER1).



### TABLE C.2. Summary of Storage Conditions

. Matrix Analyte Stor age Temperature Actual Storage

Duration 1 Limit of Demonstrated Storage Stability 2

Dicamba

Undelinted Cotton

Seed Dicamba Ambient at field sites;

- 12 °C at processor;

- 20 °C at analytical laboratory 152

- 196 days

(5.0

- 6.4 months) Residues of dicamba are st able during freezer

storage for up to 277 days (9 months) in

undelinted cotton seed.

Hulls 33

- 40 days (1.1

- 1.3 months) Storage stability data for the processed

fractions was not provided. However, storage stability data is not required as the process ed fractions were generally analyzed within 30 days of processing. Meal 26

- 27 days (0.9 months)

Alkali Refined Oil 24

- 25 days (0.8 months)

RBD Oil 19

- 20 days (0.6

- 0.7 months)

Dicamba M etabolites

Undelinted Cotton

Seed 5

- OH

Dicamba, DCGA,

D CSA Ambient at field sites;

- 12 °C at processor;

- 20 °C at analytical laboratory 152

- 189 days

(5.0

- 6.2 months) Residues of dicamba, DCGA, and DCSA are

stable during freezer storage for up to 277 days (9 months) in undelinted cotton seed. The stability of 5

- OH dicamba was not determined concurrently because it was not

found in undelinted cotton seed.

Hulls 33

- 34 days (1.1 months) Storage stability data for the processed

fractions was not provided. However, storage stability data is not required as the p rocessed fractions were generally analyzed within 30 days of processing. Meal 26

- 27 days (0.9 months)

Alkali Refined Oil 24

- 25 days (0.8 months)

RBD Oil 19

- 20 days (0.6

- 0.7 months)

1 Interval from harvest

/processing to analysis; all samples were analyzed on the day of extraction. The samples were processed

within 118

- 169 days of harvest.

2 R efer to the 860.1380 der for MRID 48728704.

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company

DACO 7.4.5/OPPTS 860.1520/OECD IIA 6.5.4 and IIIA 8.5

Processed Food and Feed

– Dicamba

- Tolerant Cotton (Event MON 88701)

DP# 404821 /MRID No. 48 728703 Page 13 of 15 TABLE C.3. Residue Data for Dicamba and Metabolites from Cotton Processing Studies 1

RAC Processed

C ommodity Total

Rate

lb ae/A (kg ae/A) TRT 2 PHI

(days) Residues (ppm) 3 Processing Factor 4,5

Dicamba 5

- OH

Dicamba DCGA DCSA Combined

Residues Dicamba 5

- OH

Dicamba DCGA DCSA Combined

Residues

Fisk, Butler, MO; 2010 (MO1)

Cotton Undelinted Seed 2.0 (2 .2) 2 79 <0.02 <0.02 <0.02 < 0.02 <0.08

-- -- -- -- -- Hulls <0.02 <0.02 0.02 0.02 <0.08 NC 5 NC >1.0x 1.0x 1.1x

Meal <0.02 <0.02 <0.02 <0.02 <0.08 NC NC NC NC NC

Alkali Refined

Oil <0.02 <0.02 <0.02 <0.02 <0.08 NC NC NC NC NC

RBD Oil <0. 02 <0.02 <0.02 <0.02 <0.08 NC NC NC NC NC

Cotton Undelinted Seed 2.0 (2.2) 4 7 0.70 <0.02 0.04 0.03 <0.78

-- -- -- -- -- Hulls 0.70 <0.02 0.05 0.03 <0.79 1.0x NC 1.3x 1.0x 1.0x

Meal 0.22 <0.02 <0.02 <0.02 <0.28 0.31x NC 0.50x <0.67x 0.35x

Alka li Refined

Oil <0.02 <0.02 <0.02 <0.02 <0.08 <0.03x NC <0.50x <0.67x 0.10x

RBD Oil <0.02 <0.02 <0.02 <0.02 <0.08 <0.03x NC <0.50x <0.67x 0.10x Uvalde, Uvalde, TX; 2010 (TX4)

Cotton Undelinted Seed 2.0 (2.2) 2 71 <0.02 <0.02 <0.02 <0.02 <0.08

-- - - -- -- -- Hulls <0.02 <0.02 <0.02 0.02 <0.08 NC NC NC >1.0x 1.0x

Meal <0.02 <0.02 <0.02 <0.02 <0.08 NC NC NC >1.0x 1.0x

Alkali Refined

Oil <0.02 <0.02 <0.02 <0.02 <0.08 NC NC NC NC NC

RBD Oil <0.02 <0.02 <0.02 <0.02 <0.08 NC NC NC NC N C

Cotton Undelinted Seed 2.0 (2.2) 4 6 0.60 <0.02 0.04 0.05 <0.69

-- -- -- -- -- Hulls 0.16 <0.02 0.08 0.06 <0.26 0.27x NC 2.0x 1.2x 0.3 7 x Meal <0.02 <0.02 <0.02 <0.02 <0.08 <0.03x NC <0.50x <0.40x 0.12x

Alkali Refined

Oil <0.02 <0.02 <0.02 <0.02 <0.08 <0.03x NC <0.50x <0.40x 0.12x

RBD Oil <0.02 <0.02 <0.02 <0.02 <0.08 <0.03x NC <0.50x <0.40x 0.12x

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company

DACO 7.4.5/OPPTS 860.1520/OECD IIA 6.5.4 and IIIA 8.5

Processed Food and Feed

– Dicamba

- Tolerant Cotton (Event MON 88701)

DP# 404821 /MRID No. 48 728703 Page 14 of 15 TABLE C.3. Residue Data for Dicamba and Metabolites from Cotton Processing Studies 1

RAC Processed

C ommodity Total

Rate

lb ae/A (kg ae/A) TRT 2 PHI

(days) Residues (ppm) 3 Processing Factor 4,5

Dicamba 5

- OH

Dicamba DCGA DCSA Combined

Residues Dicamba 5

- OH

Dicamba DCGA DCSA Combined

Residues

Average

Cotton Undelinted Seed 2.0 (2.2) 2 71

- 79

-- -- -- -- -- Hulls NC NC 1.0x 1.0x 1.0x

Meal NC NC NC 1.0x 1.0 x Alkali R efined

Oil NC NC NC NC NC

RBD Oil NC NC NC NC NC

Cotton Undelinted Seed 2.0 (2.2) 4 6

- 7

-- -- -- -- -- Hulls 0.63x NC 1.6x 1.1x 0.69x

Meal 0.17x NC 0.50x 0.53x 0.2 3 x Alkali Refined

Oil 0.03x NC 0.50x 0.53x 0.11x

RBD Oil 0.03x NC 0.50x 0.53x 0.11x

Cotton Undelinted Seed 2.0 (2.2) 2

- 4 6

- 79

-- -- -- -- -- Hulls 0.63 x NC 1.4 x 1.1 x 0.86 x Meal 0.17 x NC 0.50 x 0.69 x 0.49 x Alkali Refined

Oil 0.03 x NC

0.50 x 0.53 x 0.11 x RBD Oil 0.03 x NC 0.50 x 0.53 x 0.11 x

1 Each trial site was planted with Event MON 88701 cotton that is tolerant to dicamba herbicide and glufosinate

- ammonium herbicide.

2 TRT 2: Applications made at preemergence, the 6

- leaf stage, and at the first white flower + 15 days stage; EP: Clarity. TRT 4: applications made at the 6

- leaf stage, the

first white flower + 15

- days stage, the first open boll stage, and at 7 days prior to harvest; EP: Clarity.

3 The LOQs were 0.02 ppm per analyte per matrix. Residues for individual analytes are presente d per se and reflect the raw data. Combined residues were calculated by the study reviewer and are expressed in terms of parent equivalents. Q uantifiable residues of 5

- OH dicamba, DCSA, and DCGA were converted to parent equivalents by the study reviewer using molecular weight conversion factor s of 0.933, 1.068, and 0.991, respectively

. W hen calculating c ombined r esidues , the LOQ was used for values reported as ND or between the LOD and LOQ.

4 Processing Factor = [Measured residue for analyte in the proc essed fraction] / [Measured residue for analyte in the RAC].

5 NC = Not calculated; residues were below the LOQ in the RAC and processed fraction.

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company

DACO 7.4.5/OPPTS 860.1520/OECD IIA 6.5.4 and IIIA 8.5

Processed Food and Feed

– Dicamba

- Tolerant Cotton (Event MON 88701)

DP# 404821 /MRID No. 48 728703 Page 15 of 15 

## D. CONCLUSION

The submitted cotton processing study is acceptable. Following three or four applicatio ns of the 4 lb ae/gal SC formulation at a total rate of 2 lb ae/A, using PHIs of 71

- 79 days and 6

- 7 days, respectively, the combined residues of dicamba, 5

- OH dicamba, DCGA, and DCSA (in parent equivalents) were below the LOQ ( < 0.08)

- <0.78 ppm in/on undeli nted cotton seed (RAC),

<0.081

- <0.79 ppm in hulls, below the LOQ ( < 0.08)

- <0.28 ppm in meal, and below the LOQ (<0.08 ppm) in alkali refined oil and RBD oil. The processing data indicate that the combined

residues do not concentrate in hulls, meal, alkali refined oil, or RBD oil (average processing factors of 0.86x, 0.49, 0.11x, and 0.11x, respectively ). The observed concentration factors were

less than the theoretical concentration factor s.

Samples were analyzed using acceptable methods, and the study is supported by a dequate storage stability dat a

.

E. REFERENCES

None.

F. DOCUMENT TRACKING

Petition Number(s): 2F8067 DP Barcode(s): 408384

PC Code: 029801, 029802, 029806, 128931, 128944, & 129043

Template Version June 2005

### Dicamba/MON 11900 / PC Code 1 2 8 93 1 /M on sa nto Co mpany
DA CO 7.3 / 0PPTS 860.1 3 80 / 0ECD ll A 6. 1.1 and lll A 8.1.1

Stora g e Stability

- Dicamba-Toleranl Canon (Event MO 8870 I)



### Prim a r y Evaluat o r

Versar. [n c.



## - 

# ,., A ppro ve d b y (

### _./**~** ---f-c-,i.**~**

### A:~,.__._;._ __/



### P eter Savo ia

, Ch e mi st

, OCS PP- HED-RAB V / Vll Date: 11115 / 2012

Date: 04 / 22 / 2013

Note: Thi s Data Eva lu a ti o n Record ( DER ) was or i g inall y pr epa r ed under contra c t by Versar, In c. (6850 V e r sa r Center

, Sp rin gfield

. VA 22151; submitted 11 /1 5 /1 2 )

. The D E R ha s been re viewed by HED and revi s ed to reflect

current Office of P e s ti cide P rogram s (O PP ) polici es

.



### STUDY REPORT:

48728704

. Maher. D. ; F os t e r
, 

### J. (2 01 2) D e t e rmination o f the Stability of Dicamb a Re s idu es in

Di ca mba-Tol era nt Cotton und e r Frozen Storage Conditi o n s. P roject N umber:

R EG/I 0 / 504/ 0CR

, MSL00 2305 8

. Unpub li s h ed st ud y prepared by Monsanto Co mpan y. l 44p.



### EX EC UTIVE SUMMA RY :

Monsanto ha s s ubmitt e d th e r es ults of a s t o ra ge stabili t y s tud y with fi e ld-in curred residues of

dicamba and it s metabolites 3,6 -dichloro sa lic y li c acid ( DCSA) and 3

, 6 -dichl o r oge nti sic acid
( D C GA ) in/on dicamba-tolerant co tton identified as E ve nt MON 88701 ( form e rl y GH_S26695).

Eve nt M O N 88701 cotton expresses a modified dicamb a mono-oxygenase ( OMO) ge ne deri ved

from 

### Ste notroph o mona s malt oph i/ia and the b i alaphos r es istance (BA R ) gene i so lat e d from 

### Streptomyces h yg roscopicu s

. T he exp re ss i on of OMO co nf e r s dicamba toleranc e and th e

expre ss i o n of BAR confers glufos inate toler a nce to Event MON 88701 cotton. Sa mples of

und e linted cotton see ds were ob tained from a crop fi e ld tria l conducted b y Monsan t o (M RID
48728703). Samp l es were collec t ed 6 days following the l ast of four post-emergence broad cas t applicatio n s of a 4 lb ae / gal water solub l e concentrate (S L ) formulati o n of dicamba diglycolan1ine sa lt (C larit y H erbic ide ; M ON 54140) at 0.5 lb ae / A (0.56 kg ae / ha)

, for a total application rat e of2 .0 lb ae / A (2.2 k g ae / h a)

. The applicat i ons were made at th e 6- l ea f stage.

fir s t w hite flo wer + 15 da ys s t age, first open bo ll s ta ge a nd 7 -da ys preharvest. Samp le s we r e plac ed in fro ze n s torage ( :5 -1 0 ° C) from th e time of initi a l sa mplin g / milling until a nalysis at

- 0

, 1 , 2

, 4

, 6 and 9 months from th e initiation o f the s tora ge s t ab ility s tud y

.

Unde lint ed cott o n see d s ampl es we re anal yzed fo r residues of dicamb a. DCSA and DCG A u s in g
l iquid c hrom atography with t andem ma ss spec trometr y d e tection ( L C / MS/MS), Monsanto

m e th o d AG-ME-1381. Th e va lidated limit o f quantitation (L OQ ; determined as the lo west l eve l

of m e thod va lid a ti on, LLMV) was 0

. 02 ppm for dicamba. DCSA a nd D CGA. The method was acceptable for data collection based o n adequate meth o d va lidati on a nd co n current r ecove r y data.

Th e r es ults of th e s tud y indic a t e that field-incurred re s idu es of dicamba and met abo lites D CSA

and D C GA are r e lati ve l y s t ab l e in / on samp l es of und e lin t e d cotton see d sto r e d fro ze n at :5 -10 ° C

for up to 9 month s

. Corrected mean recoverie s at 9 m o nth s ( calculat e d a s percentage of th e 0-

month r es ult s) were 76%

, 117 % and 95%, r es pecti ve l y. for d ic amba. DCGA a nd D CSA.

04 08384 / MRlD No. 48728704 Page I 

### of9

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company

DACO 7.3/OPPTS 860.1380/OECD IIA 6.1.1 and IIIA 8.1.1 Storage Stability

– Dicamba

- Tolerant Cotton (Event MON 88701)

D 408 384 /MRID No. 48728704 Page 2 of 9 

## STUDY/WAIVER ACCEPTABILITY/DEFICIENCIES/CLARIFICATIONS :

Under the conditions an d parameters used in the study , the storage stability data are classified as scientifically acceptable. The acceptability of th is stud y for regulatory purposes is addressed in the forthcoming U.S. EPA Residue C hemistry Summary Document, D 408 384

.

COMPLIANCE :

Signed and dated Good Laboratory Practice (GLP), Quality Assurance and No Data Confidentiality statements were provided. No deviations from regulatory requirements were reported which would have an impact on the validity of the study.

A. BACKGROUND INFORMATION

Dicamba (3,6

- dichloro

- 2

- methoxybenzoic acid ) is a selective benzoic acid herbicide registered

for the control of weeds prior to their emergence. The Dicamba Reregistration Eligibility Decision (RED) was issued December 2005. The chemical structure and nomenclature of

dicamba and its metabolites DSCA and DCGA are presented in Table A.1. The physicochemi cal properties of the technical grade of dicamba acid are presented in Table A.2.

TABLE A.1. Test Compound Nomenclature.

Compound

Common name Dicamba

Company experimental name MON 11900

IUPAC name 3,6

- dichloro

- o

- anisic acid or 3,6

- dichloro

- 2

- methoxybenzoic acid

CAS name 3,6

- dichloro

- 2

- methoxybenzoic acid

CAS registry number 1918

- 00

- 9 (dicamba acid), 104040

- 79

- 1 (diglycolamine salt), or 53404

- 28

- 7 (monoethanolamine salt)

End

- use product Clarity® Herbicide: SL formulation cont aining 4 lb ae/g al

Compound

Common name DCSA

Company experimental name 3,6

- dichlorosalicylic acid ; MON 52708

IUPAC/CAS name 3,6

- dichloro

- 2

- hydroxybenzoic acid

CAS registry number 3401

- 80

- 7

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company

DACO 7.3/OPPTS 860.1380/OECD IIA 6.1.1 and IIIA 8.1.1 Storage Stability

– Dicamba

- Tolerant Cotton (Event MON 88701)

D 408 384 /MRID No. 48728704 Page 3 of 9 TABLE A.1. Test Compound Nomenclature.

Compound

Common name DCGA

Company experimental name 3,6

- dichlorogentisic acid (DCGA); MON 52724

IUPAC/CAS name 2,5

- dichloro

- 3,6

- dihydroxybenzoic acid

CAS registry number 18688

- 01

- 2



### TABLE A.2. Physicochemical Properties of the Technical Grade Test Compound: Dicamba.

Parameter Value Reference

Melting point 114

- 116  C (PAI)

90

- 100  C (87% TGAI) Residue Chemistry Chapter of the Dicamba RED, DP# 317699, 12/20/05, C. Olinger pH 2.5

- 3.0 (87% TGAI)

Density 1.57 g/mL at 25  C (87% TGAI)

Water solubility 0.5 g/100 mL at 25  C (PAI)

Solvent solubility g/100 mL at 25  C (PAI)

dioxane 118.0

ethanol 92.2 isopropyl alcohol 76.0 methylene chloride 26.0

acetone 17.0

toluene 13.0 xylene 7.8 heavy aromatic naphthalene 5.2

Vapor pressure 3.4 x 10

- 5 mm Hg at 25  C (PAI)

Dissociation constant, pK a 1.97 (PAI)

Octanol/water partition coefficient, Log(K OW ) 0.1 (PAI)

UV/visible absorption spectrum neutral: 511 (275 nm) acidic (pH 0

- 1): 1053 (281 nm) basic (pH 13

- 14): 469 (274 nm)



## B. EXPERIMENTAL DESIGN

B.1. Sample Handling and Preparation

The storage stability study was conducted by Monsant o Company at its Environmental Sciences Technology Center in St. Louis, Missouri

.

Undelinted cotton seed samples were obtained from a crop field trial study conducted by

Monsanto (MRID 48728703) using dicamba

- tolerant cotton identified as Event MON 88701 (formerly GH_S26695). Event MON 88701 cotton expresses a modified dicamba mono

oxygenase (DMO) gene derived from Stenotrophomonas malt ophilia and the bialaphos resistance (BAR) gene isolated from Streptomyces hygroscopicus

. The expression of DMO confers dicamba tolerance and the expression of BAR confers glufosinate tolerance to Event

MON 88701 cotton.

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company

DACO 7.3/OPPTS 860.1380/OECD IIA 6.1.1 and IIIA 8.1.1 Storage Stability

– Dicamba

- Tolerant Cotton (Event MON 88701)

D 408 384 /MRID No. 48728704 Page 4 of 9 

## In one trial conducted in Hockley, TX (Site TX2), samples of undelinted cotton seeds were collected from treatment plot 4 (TRT 4) following the last of four post

- emergence broadcast applications of a 4 lb ae/gal SL formulation of dicamba diglycolamine salt (Clarity Herbicide; MON 54140) at 0.5 lb ae/A (0.56 kg ae /ha ) , for a total application rate of 2.0 lb ae/A (2.2 kg

ae/ha ). The applications were made at the 6

- leaf stage, f irst white flower + 15 days stage, first open bo ll stage and 7

- days preharvest usin g a backpack sprayer in spray volumes of ~20 gal/A (187 L/ha) . A non

- ionic surfactant (NIS) and ammonium sulfate (AMS) were added to the spray

mixture.

Single composite cotton seed samples (~1 kg) were collected from the control and treated plot 6 days after the last application and ginned at the field site one day after harvest

. Samples were stored at ambient temperature until ginning. The undelinted cotton seed samples were placed in

fr ozen storage at the field trial site within 1 hour of sampl ing and shipped via ACDS freezer

truck to Monsanto the next day (2 days after harvest)

. The s amples were placed in frozen storage at Monsanto (

- 10 °C) and were removed from frozen storage and analyzed at intervals of

~ 0, 1, 2, 4, 6 and 9 months from the initiation of the storage stability study, which occurred 121 days

after harvest. Actual storage durations were 0, 31, 60, 125, 194, and 277 days. T he undelinted

cotton seed subsample used for the storage stability study was prepared at Monsant o two days before the Day 0 analysis (119 days after harvest) using a cryomill, which pulverizes samples into a fine powder at cryogenic temperatures. Concurrent recovery samples were fortified at

each interval at 0.04, 0.3 and 2.0 ppm, using a mixed stan dard containing dicamba and DCSA and a separate solution containing only DCGA.

A subsample of the treated undelinted cotton seed sample was originally milled 20 days after harvest and analyzed 38 days after harvest, as part of the crop field trial study (see above) . Due to an inadequate amount of milled material, new subsamples were milled 119 days after harvest for use in the storage stability study. Th re e replicates of the original milled subsample w ere re

analyzed shortly after the init iation of the storage stability study to assess stability of residues during this period. The results of the original and repeat analyses were similar. The Study

Author attributed differences between residues in the original analysis for the crop field trial study and the Day 0 analysis for the storage stability study to variability between subsamples

.

B.2. Analytical Methodology

S amples were analyzed for residues of dicamba and the metabolites DCGA and DCSA using a LC/MS/MS method from Monsanto: Analytical Method AG

- ME

- 1381

- 01. For a complete description of the LC/MS/MS method, refer to the 860.1340 DER for MRID 48728702.

Briefly, samples were extracted with acetonitrile : water ( 4 0: 6 0, v:v). An aliquot of the extract

was hydrolyzed in 1 N HCl at 95 °C in an oven

. The hydrolysate was partitioned with ethyl acetate:isooctane (20:80, v:v). Water was added to the organic phase and the sample was concentrated by evaporation until only the aqueous solution remained. The samples were acidified and analyzed us ing LC/MS/MS.

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company

DACO 7.3/OPPTS 860.1380/OECD IIA 6.1.1 and IIIA 8.1.1 Storage Stability

– Dicamba

- Tolerant Cotton (Event MON 88701)

D 408 384 /MRID No. 48728704 Page 5 of 9 

## The LOQ (determined as the LLMV) was 0.02 ppm for dicamba, DCSA and DCGA in

undelinted cotton seed. We note that t he petitioner also presented calculated values statistically

determined for the LOD and LOQ based on recoveries obtained at the 0.020 ppm fortification

level from the method validation study

. The LODs were 0.0089, 0.0035 and 0.0041 ppm for dicamba, DCSA and DCG A, respectively. The LOQs were 0.0266, 0.0105 and 0.0121 ppm for dicamba, DCSA and DCGA, respectively

.

The analytical method was validated prior to the storage stability study and in conjunction with analysis of frozen storage stability samples.

C. RESULTS AND DISCUSSION

C oncurrent recovery data are presented in Tab le C.1

. The method validation data are presented

in the DER for MRID 48728703. The data indicate that the LC/MS/MS was adequate for determination of residues of dicamba, DCSA and DCGA in undelinted cotton seed. C oncurrent recoveries were within the acceptable range of 70

- 120% for all analytes in samples fortified at 0.04, 0.30, and 2.0 ppm each. Apparent residues in control samples were less than the method LOQ (<0.02 ppm), and adequate example chromatograms were provided. Concurrent recoveries were corrected for residues found in the control samples.

The results of the storage stability stud y are presented in Table C.2. The study resu lts indic a te that residues of dicamba, DCSA and DCG A a re stable in undelint ed cotton seed stored fro zen ( ≤

- 10 ° C ) for up to 9 months

. Corrected mean recoveries were 76 %

, 117 % and 95 %

, respectively,

for dicamba, DC G A and DC S A in/on undelinted cotton seed at 9 months (calculated as percentage of the 0

- month resul ts).

A g raph depicting the storage stability of field

- incurred residues of dicamba and its metabolites

DCS A and DCG A in /on undelinted cotton seed is presented in Figure C .1

.

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company

DACO 7.3/OPPTS 860.1380/OECD IIA 6.1.1 and IIIA 8.1.1 Storage Stability

– Dicamba

- Tolerant Cotton (Event MON 88701)

D 408 384 /MRID No. 48728704 Page 6 of 9 TABLE C.1 Summary of Concurrent Recoveries of Dicamba and its Metabolites from Undeli nted Cotton

Seed.

Matrix Analyte Storage

Interval (Months) Fortification

Level (ppm) Sample size (n) Recoveries 1
(%) Mean ± Std

Dev. 2
(%)

Undelinted

cotton seed Dicamba 0 0.04 1 79.8

92.9 ± 11.5 0.30 1 98.0

2.00 1 101

1 0.04 1 90.3

97.0 ± 7.39

0.30 1 96.3

2.00 1 105

2 0.04 1 97.3

97.6 ± 1.77

0.30 1 96.0

2.00 1 99.5

4 0.04 1 107

111 ± 4.00

0.30 1 111

2.00 1 115

6 0.04 1 117

91.1 ± 22.3

0.30 1 80.2

2.00 1 76.7

9 0.04 1 112

105 ± 6.26

0.30 1 105

2.00 1 99.5

DCGA 0 0.04 1 103

105 ± 2.08 0.30 1 107

2.00 1 104

1 0.04 1 83.5

84.7 ± 1.43

0.30 1 84.4

2.00 1 86.3

2 0.04 1 96.7

100 ± 3.22

0.30 1 101

2.00 1 103

4 0.04 1 104

103 ± 3.21

0.30 1 99.0

2.00 1 105

6 0.04 1 94.0

93.4 ± 2.32

0.30 1 95.3

2.00 1 90.8

9 0.04 1 95.0

105 ± 9.54

0.30 1 114

2.00 1 106

DCSA 0 0.04 1 99.9

112 ± 11.3 0.30 1 115

2.00 1 122

1 0.04 1 115

105 ± 12.3

0.30 1 108

2.00 1 91.0

2 0.04 1 99.4

94.5 ± 4.25

0.30 1 92.2

2.00 1 91.9

4 0.04 1 98.7

95.0 ± 6.29

0.30 1 98.5

2.00 1 87.7

6 0.04 1 79.0

76.0 ± 4.71

0.30 1 70.6

2.00 1 78.5

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company

DACO 7.3/OPPTS 860.1380/OECD IIA 6.1.1 and IIIA 8.1.1 Storage Stability

– Dicamba

- Tolerant Cotton (Event MON 88701)

D 408 384 /MRID No. 48728704 Page 7 of 9 TABLE C.1 Summary of Concurrent Recoveries of Dicamba and its Metabolites from Undeli nted Cotton

Seed.

Matrix Analyte Storage

Interval (Months) Fortification

Level (ppm) Sample size (n) Recoveries 1
(%) Mean ± Std

Dev. 2
(%)

9 0.04 1 98.5

97.7 ± 3.01

0.30 1 94.2

2.00 1 100

1 Recovery values were corrected for background levels in controls.

2 Standard deviations were determined by the study reviewer.



### TABLE C .2 Stability of Field

- Incurred Residues of Dicamba, DCGA and DCSA in Dicamba

- Resistant

Undelinted Cotton Seed Following Storage at

- 1 0°C

Commodity Analyte Spike

Level (ppm) Storage

interval (months ) Recovered

Residues (ppm) Mean

Recovered

Residues (ppm) Mean

Recovery 1
(%) Corrected

Recovered

Residues 2 (ppm) Corrected

Mean

R ecovery 3
(%)

Undelinted

cotton seed Dicamba Incurred 4

0 1.01, 0.91, 0.78, 0.83, 0.78, 0.82 0.85 100 0.91

--

1 0.82, 0.76,

0.79, 0.75 0.78 92 0.80 88

2 0.70, 0.89,

0.90, 0.88 0.84 99 0.86 94

4 0.94, 0.82,

0.88, 1.00 0.91 107 0.82 90

6 0.54, 0.62,

0.70, 0.72, 0.64, 0.72 0.66 78 0.72 79

9 0.72, 0.70,

0.80, 0.75, 0.72, 0.71 0.73 86 0.70 76

DCGA Incurred 4

0 0.13, 0.12, 0.12, 0.13,
0.11, 0.12 0.12 100 0.11

--

1 0.14, 0.14,

0.16, 0.15 0.14 117 0.17 145

2 0.12, 0.13,

0.13, 0.14 0.13 108 0.13 114

4 0.14, 0.13,

0.12, 0.12 0.13 108 0.13 110

6 0.13, 0.14,

0.15, 0.13, 0.12, 0.14 0.13 108 0.14 1 22

9 0.14, 0.13,

0.14, 0.15, 0.14, 0.13 0.14 117 0.13 11 7

DCSA Incurred 4

0 0.24, 0.24,
0.24, 0.22, 0.24, 0.23 0.23 100 0.21

--

1 0.22, 0.21,

0.23, 0.24 0.22 96 0.21 102

2 0.20, 0.24,

0.22, 0.25 0.23 100 0.24 1 19

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company

DACO 7.3/OPPTS 860.1380/OECD IIA 6.1.1 and IIIA 8.1.1 Storage Stability

– Dicamba

- Tolerant Cotton (Event MON 88701)

D 408 384 /MRID No. 48728704 Page 8 of 9 TABLE C .2 Stability of Field

- Incurred Residues of Dicamba, DCGA and DCSA in Dicamba

- Resistant

Undelinted Cotton Seed Following Storage at

- 1 0°C

Commodity Analyte Spike

Level (ppm) Storage

interval (months ) Recovered

Residues (ppm) Mean

Recovered

Residues (ppm) Mean

Recovery 1
(%) Corrected

Recovered

Residues 2 (ppm) Corrected

Mean

R ecovery 3
(%)

4 0.21, 0.20,

0.19, 0.21 0.20 87 0.21 103

6 0.12, 0.18,

0.18, 0.17, 0.18, 0.18 0.17 7 4 0.22 109

9 0.20, 0.17,

0.18, 0.21,
0.18, 0.18 0.19 83 0.19 95

1 Recoveries for stored samples calculated by Versar reviewer as percentage of 0

- month results.

2 Corrected by the study reviewer for mean concurrent recovery (Table C.1.)

3 Calculated by the study reviewer as (ppm corrected recovered residues in sample ÷ ppm corrected recovered residues in 0

- day sample).

4 Residues incurred following four application s of MON 54140, a SL formulation of dicamba diglycolamine salt containing 4 lb ae/gal (480 g/L)

, to cotton a t application rates of ~ 0.5 lb ae /A /application (0.5 6 kg ae /ha)

.



## F igure C.1 Graph of Dicamba Storage Stability in Undelinted Cotton Seed

### Dicamba/ MON 11900 /PC Code 128931 / Monsanto Company

DACO 7.3/OPPTS 860.1380/OECD IIA 6.1.1 and IIIA 8.1.1 Storage Stability

– Dicamba

- Tolerant Cotton (Event MON 88701)

D 408 384 /MRID No. 48728704 Page 9 of 9 

## D. CONCLUSION

The storage stability data are adequate and demonstrate the stability of field

- incurred residue s of

dicamba and metabolites DCSA and DCG A in/on samples of undelinted cotton seed stored frozen for up to 9 months at ≤

- 10 °C. An acceptable method was used for quantitation of

residues.

E. REFERENCES

Residue Chemistry Chapter of the Dicamba RED, DP# 317699, 12/20/05, C. Olinger

F. DOCUMENT TRACKING

Petition Number(s): 2F8067 DP Barcode(s): 408384

PC Code: 029801, 029802, 029806, 128931, 128944, & 129043