Glyphosate Summary of Analytical Chemistry and Residue Data DP#: 394964

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Executive Summary

Background: Glyphosate is a nonselective Group 9 herbicide that disrupts the synthesis of the 

aromatic amino acids tyrosine, tryptophan, and phenylalanine through inhibition of 5-

enolpyruvylshikimate-3-phosphate synthase (EPSPS). Glyphosate is currently registered for preand post-emergence application to a variety of fruit, vegetable, and field crops. Tolerances are 

currently established for residues of glyphosate in/on various plant commodities at 0.2-400 ppm 

(40 CFR §180.364(a)(1); canola seed at 20 ppm) and for the combined residues of glyphosate 

and N-acetyl-glyphosate (expressed in glyphosate equivalents) in/on field corn, soybean, 

aspirated grain fractions (AGF), and livestock commodities at 0.10-310 ppm (40 CFR 

§180.364(a)(2)). The glyphosate Reregistration Eligibility Decision (RED) document was issued 

September 1993. A Registration Review Scoping document was completed in 2009 (D362745, 

J. Langsdale et al., 3-Jun-2009). 

The current proposal concerns application of glyphosate to the glyphosate-tolerance canola 

variety DuPontTM Optimum® GLY Canola (also called canola (Event DP-Ø73496-4)). The 

petitioner indicated that Optimum® GLY Canola has been genetically modified to express the 

gat4621 gene derived from the soil bacterium Bacillus licheniformis (e-mail from DuPont; Tim 

McPherson; 21-Oct-2011; 10:07 am). The protein encoded by the gat4621 gene confers 

tolerance to glyphosate via acetylation of the secondary amine group of glyphosate resulting in 

the formation of the non-herbicidal N-acetyl-glyphosate. 

Proposed Use: The petitioner submitted a proposed supplemental label for DuPontTM AbunditTM 

S Herbicide (EPA Reg. No. 352-832; water soluble liquid; isopropylamine salt of glyphosate; 3 

lb ae/gal) which includes directions for application to DuPontTM Optimum® GLY Canola. Table 

3 is a summary of the proposed end-use product and Table 4 is a summary of the proposed 

application scenario. Based on the currently available confined/field rotational crop data, a 30-

day plant-back interval (PBI) for all non-labeled crops is appropriate following application to 

Optimum® GLY canola. A revised Section B is requested. 

Nature of the Residue – Primary Crops: Metabolism studies conducted with non-transgenic 

corn, cotton, soybeans, and wheat were previously submitted and reviewed. Based on these data, 

HED concluded that the residue of concern in non-transgenic plants is glyphosate (Memo, R. 

Perfetti, 19-Oct-1992; RED, R. Perfetti, 27-Oct-1992; Memo, R. Perfetti, 17-Mar-1994). 

Metabolism studies have also been submitted on glyphosate-tolerant canola (RT73; D242628, T. 

Bloem, 30-Nov-1998) and glyphosate-tolerant field corn (Roundup Ready®field corn; D217539, 

G. Kramer, 14-Mar-1996). The glyphosate-tolerant canola and field corn varieties were 

genetically modified to express the EPSPS gene derived from Agrobacterium sp. (strain CP4) 

which codes for an EPSPS protein that is not inhibited by glyphosate. The glyphosate-tolerant 

canola and corn were also genetically engineered to express the oxireductase gene which 

converts glyphosate to the non-herbicidal AMPA (aminomethyl phosphonic acid). Metabolism 

in these varieties of transgenic canola and corn was essentially the same as the non-transgenic 

plants. Therefore, it was concluded that the terminal residue to be regulated, in non-transgenic 

plants and transgenic field corn and canola modified to express the Agrobacterium sp. EPSPS 

and oxireductase genes, is glyphosate.

Subsequent to these decisions, HED approved DuPont requests permitting the commercialization 
Glyphosate Summary of Analytical Chemistry and Residue Data DP#: 394964

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of Optimum GAT soybean and Optimum GAT field corn (D346713, T. Bloem, 12-Mar2008; D357880, T. Bloem, 29-Oct-2008; D361315, T. Bloem, 14-Jan-2010). Optimum

GAT soybean was engineered to express the gat4601 gene (derived from Bacillus 

licheniformis; soil bacterium) and Optimum GAT field corn was engineered to express the 

gat4621 gene (derived from Bacillus licheniformis) and the zm-hra gene (modified version of 

maize acetolactate synthase (ALS) gene). The protein encoded by the gat4601 and the gat4621 

genes confer tolerance to glyphosate via acetylation of the secondary amine group of glyphosate 

resulting in the formation of the non-herbicidal N-acetyl-glyphosate; the zm-hra gene in 

Optimum GAT field corn, encodes for an ALS protein which is not sensitive to the ALSinhibiting herbicides. As a result of the introduction of these seed lines, HED concluded that the 

residues of concern in plants for tolerance expression and risk assessment should change from 

glyphosate to the combined residues of glyphosate and N-acetyl-glyphosate (expressed in 

glyphosate equivalents; D346713, T. Bloem, 12-Mar-2008). Subsequent to this decision, it was 

determined that only the tolerance expression for field corn and soybean would change from 

glyphosate to the combined residues of glyphosate and N-acetyl-glyphosate (expressed in 

glyphosate equivalents); the tolerance expression for all other crops would remain as glyphosate. 

The current action is requesting application of glyphosate to Optimum® GLY Canola (also called 

canola (Event DP-Ø73496-4)). Optimum® GLY Canola is a new variety of canola genetically 

modified to express the gat4621 gene derived from the soil bacterium Bacillus licheniformis (email from DuPont; Tim McPherson; 21-Oct-2011; 10:07 am). The protein encoded by the 

gat4621 gene confers tolerance to glyphosate via acetylation of the secondary amine group of 

glyphosate resulting in the formation of the non-herbicidal N-acetyl-glyphosate. Based on the 

results of a metabolism study conducted with canola genetically modified to express the 

microbial gat gene (preemergence and foliar applications) and previous conclusions pertaining to 

the toxicity of AMPA (Memo, R. Perfetti, 17-Mar-1994) and N-acetyl-AMPA (D345926, T. 

Bloem et al., 18-Mar-2008), HED concludes that with the introduction of Optimum® GLY 

Canola the residues of concern in canola, for tolerance expression and risk assessment purposes,

are glyphosate and N-acetyl-glyphosate (expressed in glyphosate equivalents). 

Nature of the Residue - Livestock: The qualitative nature of the residue in livestock following 

dosing with glyphosate and AMPA is adequately understood. Studies with lactating goats and 

laying hens fed a mixture of glyphosate and AMPA indicate that the primary route of elimination 

was by excretion (urine and feces). HED determined that the terminal residue to be regulated in 

livestock is glyphosate (Memo, R. Perfetti, 19-Oct-1992; RED, R. Perfetti, 27-Oct-1992; Memo, 

R. Perfetti, 17-Mar-1994). 

DuPont previously submitted summaries of in vitro (rumen fluid, fertile hen egg, and rat liver S9 

supernatant) and in vivo (rat, goat, and hen metabolism studies) studies conducted with the Nacetyl-glyphosate metabolite. Based on these data and the glyphosate metabolism studies, HED 

concluded that the residues of concern in livestock following consumption of glyphosate and Nacetyl-glyphosate, for tolerance expression and risk assessment purposes, are glyphosate and Nacetyl-glyphosate (expressed in glyphosate equivalents; D346713, T. Bloem, 12-Mar-2008; 

D361315, T. Bloem, 14-Jan-2010). Based on these data and since Optimum® GLY Canola does

not result in any addition metabolites, HED concludes that the previous decisions concerning the 

residues of concern in livestock remain appropriate. 
Glyphosate Summary of Analytical Chemistry and Residue Data DP#: 394964

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Magnitude of the Residue - Canola: A tolerance for residues of glyphosate per se in/on canola 

seed at 20 ppm is currently established. The submitted field trials conducted with canola (Event 

DP-Ø73496-4) are considered adequate to support the proposed application scenario and resulted 

in combined glyphosate and N-acetyl-glyphosate (expressed in glyphosate equivalents) residues 

in/on seed of ≤2.0 ppm (no additional field trial data are required). In addition, an adequate 

canola seed processing study was submitted which indicates that combined residues of 

glyphosate and N-acetyl-glyphosate do not concentrate in canola meal (≤1.1x) or oil (<0.005x). 

Based on the available data, HED agrees with the petitioner’s proposal to move the currentlyestablished 20-ppm canola seed tolerance from 180.364(a)(1) where the tolerance expression is 

for residues of only glyphosate to 180.364(a)(2) where the tolerance expression is for combined 

residues of glyphosate and N-acetyl-glyphosate (expressed in glyphosate equivalents). 

Magnitude of the Residue - Livestock: The current petition results only in a change to the 

canola seed glyphosate tolerance expression (tolerance values are not changing). Therefore, the 

currently-established livestock tolerances remain appropriate. 

Nature/Magnitude of the Residue - Rotational Crops: Based on previously-reviewed confined 

rotational crop studies (3.71 lb ae/acre (2.2x); MRIDs 41543201 and 41543202, A. Abramovitch, 

14-Oct-1992) which resulted in glyphosate residues of >0.01 ppm in barley grain only (0.018 

ppm; 125-day PBI) and the limit of quantitation (LOQ) for the tolerance enforcement method of 

0.05 ppm, HED previously concluded that a 30-day PBI was appropriate for all non-labeled crops 

(D200041, G. Kramer, 12-May-1994; field rotational crop study has not been submitted). Based 

on the LOQ for the tolerance-enforcement method and since the confined rotational crop study

employed an exaggerated rate to bare soil, a 30-day PBI for all non-labeled crops is acceptable

following application to Optimum® GLY canola. A revised Section B is requested. 

Analytical Enforcement Method: DuPont liquid chromatography/mass spectrometry/mass 

spectrometry (LC/MS/MS) Methods 15444 (corn and soybean) and 20009 (livestock) have been 

determined to be acceptable for enforcement of the current tolerances for the combined residues 

of glyphosate and N-acetyl-glyphosate in/on field corn, soybean, and livestock and these methods 

were forwarded to the Food and Drug Administration (FDA; D357880, T. Bloem, 29-Oct-2008; 

D361315, T. Bloem, 14-Jan-2010). The canola field trial and processing studies submitted as 

part of the current petition were analyzed for residues of glyphosate, N-acetyl-glyphosate, 

AMPA, and N-acetyl AMPA using the DuPont LC/MS/MS Method 15444. Since adequate 

validation data were submitted as part of the canola field trail and processing data, HED 

concludes that DuPont LC/MS/MS Method 15444 is appropriate for data collection and 

enforcement purposes for canola. 

Recommendations: Provided the petitioner submits a revised Section B specifying a 30-day PBI 

for all non-labeled crops following application to Optimum® GLY canola, HED concludes that 

the residue chemistry database supports an unconditional registration for the proposed 

application scenario. A human-health risk assessment will be prepared as a separate document

(D387535). 
Glyphosate Summary of Analytical Chemistry and Residue Data DP#: 394964

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Background

Glyphosate is a nonselective Group 9 herbicide which disrupts the synthesis of the aromatic 

amino acids tyrosine, tryptophan, and phenylalanine through inhibition of EPSPS. Glyphosate is 

currently registered for pre- and post-emergence application to a variety of fruit, vegetable, and 

field crops. Tolerances are currently established for residues of glyphosate in/on various plant 

commodities at 0.2-400 ppm (40 CFR §180.364(a)(1)) and for the combined residues of 

glyphosate and N-acetyl-glyphosate (expressed in glyphosate equivalents) in/on field corn, 

soybean, AGF, and livestock commodities at 0.1-310 ppm (40 CFR §180.364(a)(2)). The 

glyphosate RED document was issued September 1993. A Registration Review Scoping 

document was completed in 2009 (D362745, J. Langsdale et al., 3-Jun-2009). The chemical 

structure/nomenclature and physicochemical properties of glyphosate are presented in Tables 1

and 2, respectively. 

Table 1: Glyphosate Nomenclature.

Compound

P

O

OH

OH

N

H

HO

O

Common name Glyphosate (PC code 471300)

Company experimental name DPX-B2856

IUPAC/CAS name N-(phosphonomethyl)glycine

CAS registry number 1071-83-6

Table 2: Physicochemical Properties of Technical Grade Glyphosate.

Melting point 189.5 ± 0.5 °C

The Pesticide Manual, 13th Edition

pH 1.9 at 20 °C

Density 1.705 g/cm3at 20 °C

Water solubility 10.5 g/L at 20 °C

Solvent solubility

acetone 0.078 g/L

methanol 0.231 g/L

hexane 0.026 g/L

ethyl acetate 0.012 g/L

dichloromethane 0.233 g/L

n-octanol 0.020 g/L

propan-2-ol 0.020 g/L

toluene 0.036 g/L

European Commission: Glyphosate 

6511/VI/99-final, 1/21/02

Vapor pressure 1.31 x 10-2 mPa at 25 °C The Pesticide Manual, 13th Edition

Dissociation constant, pKa

0.8 (1st phosphoric), 2.3 

(carboxylate), 6.0 (2nd phosphoric), 

and 11.0 (amine)

Knuuttila. 1979 Acta Chem. Scand. B 

33:623-626

Octanol/water partition coefficient, Log(KOW) -3.2 (pH 2-5, 25 °C) European Commission: Glyphosate 

UV/visible absorption spectrum ε = 0.086 (295nm) 6511/VI/99-final, 1/21/02
Glyphosate Summary of Analytical Chemistry and Residue Data DP#: 394964

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860.1200 Directions for Use

The petitioner submitted a proposed supplemental label for DuPontTM AbunditTM S Herbicide 

(EPA Reg. No. 352-832) which includes directions for application to DuPontTM Optimum® GLY 

Canola. Table 3 is a summary of the proposed end-use product and Table 4 is a summary of the 

proposed application scenario. Based on the currently available confined/field rotational crop 

data, a 30-day PBI for all non-labeled crops is appropriate following application to Optimum® 

GLY canola. A revised Section B is requested. 

Table 3: Summary of Proposed End-Use Products.

Trade Name EPA Reg. No. concentration Form. Label Date Target Crops Target Pests

DuPontTM AbunditTM 

S Herbicide 352-832

glyphosate as the 

isopropylamine salt 

4 lb ai/gal (3 lb ae/gal)

water 

soluble 

liquid

not

indicated

DuPontTM 

Optimum® GLY 

Canola

numerous annual and 

perennial grasses and 

broadleaf weeds

Table 4: Summary of Proposed Application Scenario for DuPontTM Optimum® GLY Canola.

1

Formulation

(EPA Reg. No.) App. Timing App. Rate(lb ae/acre)

Max. App.

Rate (lb ae/acre) Use Directions and Limitations

DuPontTM AbunditTM 

S Herbicide (EPA Reg.

No. 352-832)

broadcast preplant

or preemergence

not specified 1.125

-No more than two post-emergent over-the-top 

applications may be made from crop emergence to the 

6-leaf stage (minimum retreatment interval of 10 

days).

-Applications may be made with ground (3-40 gallons 

per acre (GPA)) or aerial (3-15 GPA) equipment.

-The last application must be made at least 60 days 

before harvest.

-The main label indicates that there are no rotational 

crop restrictions following application of this product. 

-The main label indicates that addition of ammonium 

sulfate may increase product performance (17 lbs/100 

gallon). 

emergence through 

6-leaf stage 0.234-0.398 0.562

1

 The proposed product is not currently registered for application to canola. HED recently recommended in favor of a petition 

concerning the application of Roundup WeatherMAX®(EPA Reg. No. 524-537) and Roundup PowerMAX®(EPA Reg. No. 

524-549) to Roundup Ready®Flex Canola (D394004, T. Bloem, 15-Sep-2011). 

860.1300 Nature of the Residue - Plants and Livestock (48398601.der)

In support of the current proposal, the petitioner submitted a metabolism study conducted with a 

canola variety genetically modified to express the microbial glyphosate N-acetyltransferase (gat) 

gene (0827 canola; 48398601.der). A summary of the study is presented below. See the 

Executive Summary for information detailing HED’s conclusions pertaining to the residues of 

concern in plants and livestock.

48398601.der: The canola variety used in the study (0827 canola) was genetically modified to 

express the microbial glyphosate N-acetyltransferase (gat) gene which confers tolerance to 

glyphosate via acetylation of the secondary amine group of glyphosate resulting in the formation 

of the non-herbicidal N-acetyl-glyphosate. Radiolabeled glyphosate was applied as a 

preemergence soil application at a rate of 4.02 lb ae/acre, followed by three foliar applications at 

0.84-0.91 lb ae/acre/application for a total application rate of 6.65 lb ae/acre (greenhouse grown). 

Foliar applications were started 21 days after the soil application, and were made at the BBCH 

12, 15, and 87 growth stages, with 14- and 90-day RTIs. Samples of immature foliage (BBCH 

69) were collected 38 days after the second foliar application (midway between the second and 
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third foliar application), preharvest foliage and pods with seed (BBCH 87) were collected 90 

days after the second foliar application (immediately prior to the third foliar application), and 

mature seed (BBCH 89) was collected 7 days after the last foliar application.

Table 5 is a summary of the total radioactive residues (TRRs) and a summary of the 

identified/characterized residues. Glyphosate was a major residue in mature seed (20.8% TRR, 

0.448 ppm), but was identified at a much lower level in immature foliage (3.0% TRR, 0.179 

ppm), and was not identified in preharvest foliage or pods (with seed). The major residue in all 

matrices was N-acetyl-glyphosate at 89.5% TRR (5.351 ppm) in immature foliage, 93.0% TRR 

(1.442 ppm) in preharvest foliage, 79.6% (1.013 ppm) in preharvest pods (with seed), and 51.1% 

TRR (1.101 ppm) in mature seed. N-acetyl AMPA was also a major residue in mature seed 

(14.7% TRR, 0.316 ppm) and was identified at a lower level in immature foliage (3.4% TRR, 

0.203 ppm). AMPA was a minor metabolite in immature foliage and mature seed at 1.4% and 

1.9% TRR, respectively. In mature seed, unidentified metabolites accounted for a total of 7.1% 

TRR (0.152 ppm), with no individual metabolite exceeding 1.0% TRR (0.022 ppm).

Based on the results of the metabolism study, the petitioner proposed that glyphosate is 

acetylated in 0827 canola plants to form N-acetyl-glyphosate. N-acetyl-glyphosate and 

glyphosate are further metabolized to N-acetyl AMPA. Low levels of AMPA detected in canola 

fractions (foliage and mature seed) could result, in part, from low levels of [14C]AMPA in the 

treatment solutions or from glyphosate or N-acetyl AMPA metabolism in the plant.

Table 5: Summary of Characterization and Identification of Radioactive Residues.

Metabolite Fraction

Immature Foliage Preharvest Foliage Preharvest Pods 

(with Seed) Mature Seed

TRR = 5.979 ppm TRR = 1.550 ppm TRR = 1.273 ppm TRR = 2.155 ppm

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

Glyphosate 3.0 0.179 -- -- -- -- 20.8 0.448

AMPA 1.4 0.084 -- -- -- -- 1.9 0.041

N-Acetyl AMPA 3.4 0.203 -- -- -- -- 14.7 0.316

N-Acetyl-glyphosate 89.5 5.351 93.0 1.442 79.6 1.013 51.1 1.101

Unidentified -- -- -- -- -- -- 7.1 0.152

Amylase hydrolysate -- -- 1.7 0.026 -- -- 11.7 0.252

Amyloglucosidase/cellulose hydrolysate -- -- 0.5 0.008 -- -- 2.3 0.050

Base hydrolysate -- -- 0.8 0.012 -- -- 3.2 0.069

Acid hydrolysate -- -- 0.4 0.006 -- -- 0.9 0.019

Total identified 97.3 5.817 93.0 1.442 79.6 1.013 88.5 1.906

Unidentified residues from HPLC analysis -- -- -- -- -- -- 7.140.1524

Total extractable197.3 5.818 93.0 1.442 79.6 1.013 78.4 1.690

Total hydrolyzed2-- -- 3.4 0.052 -- -- 18.1 0.390

Unextractable (PES)32.7 0.161 3.6 0.056 20.4 0.260 3.5 0.075

Accountability 100 100 100 100

1

 1% formic acid:methanol; 96:4; v:v.

2

 Combined amylase, amyloglucosidase/cellulase, base, and acid hydrolysates. Only the preharvest foliage and mature seed were 

hydrolyzed and only the mature seed hydrolysates were HPLC analyzed (glyphosate: 1.6% TRR; AMPA: 1.9% TRR; N-acetylAMPA: 0.4% TRR; N-acetyl-glyphosate: 6.3% TRR).

3

 Residues remaining after exhaustive extractions.

4

 No individual unidentified residue exceeded 1.0% TRR (0.022 ppm).
Glyphosate Summary of Analytical Chemistry and Residue Data DP#: 394964

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860.1340 Residue Analytical Methods

DuPont LC/MS/MS Methods 15444 (corn and soybean) and 20009 (livestock) have been 

determined to be acceptable for enforcement of the current tolerances for the combined residues of 

glyphosate and N-acetyl-glyphosate in/on field corn, soybean, and livestock and these methods were 

forwarded to FDA (D357880, T. Bloem, 29-Oct-2008; D361315, T. Bloem, 14-Jan-2010). The 

canola field trial and processing studies submitted as part of the current petition were analyzed for 

residues of glyphosate, N-acetyl-glyphosate, AMPA, and N-acetyl AMPA using the DuPont Method 

15444. Since adequate validation data were submitted as part of the canola field trail and 

processing data, HED concludes that DuPont Method 15444 is appropriate for data collection and 

enforcement purposes for canola. Standards and fortification solutions were prepared for the 

analytes in glyphosate equivalents, and all residues were quantitated as glyphosate equivalents. The 

method LOQ (determined as the lowest level of method validation (LLMV)) was 0.05 ppm for each 

analyte/matrix; the limit of detection (LOD) was defined as 0.02 ppm (1/3 LOQ). Residues in/on 

controls were generally <LOD and/or insignificant when compared to residues in/on the treated 

samples. The study indicated that concurrent recoveries were corrected for apparent residues ≥0.01 

ppm in control samples (treated samples were not corrected).

860.1360 FDA Multiresidue Methods (MRMs) Protocols

The FDA Pestrak database (1990) indicate that recoveries are not likely for glyphosate under the 

FDA MRMs. Data concerning the behavior of N-acetyl-glyphosate under the FDA MRMs has also 

been previously provided (47133201.der.doc). Using Protocol A, N-acetyl-glyphosate was 

determined not to be naturally fluorescent. Using Protocols B and C, derivatized (methylated) and 

underivatized N-acetyl-glyphosate were tested using the nitrogen-specific Modules DG5 (DB-1 

column) and DG17 (DB-17 column) which employ a nitrogen- phosphorus detector (NPD) and the 

more general Module DG18 (DB-225 column) which employs an electron-capture detector (ECD). 

N-acetyl-glyphosate was not chromatographable using DB-1, DB-17, or DB-225 columns with NPD 

or ECD; therefore, no testing was performed under Protocols D, E, or F. Because the test substance 

is not a substituted urea, no testing under Protocol G is required. The results indicate that the 

MRMs are not suitable for the determination of N-acetyl-glyphosate. These data were forwarded to 

FDA (D349698, T. Bloem, 5-Mar-2008).

860.1380 Storage Stability (48398603.der)

The canola commodities from the field trial and processing studies were stored frozen for the 

following durations: 302 days for forage, 293 days for seed, 53 days for meal, and 46 days for oil. 

Extracts were stored for up to 108 days prior to analysis. Storage stability data are available which 

indicate that residues of glyphosate, N-acetyl-glyphosate, and AMPA are stable for up to 12 months 

and residues of N-acetyl AMPA are stable for up to18 months in/on DP-356Ø43-5 soybean forage, 

seed, and hay ( ~-20 °C; 48398603.der). DP-356Ø43-5 soybean is soybean genetically modified to 

express the microbial glyphosate N-acetyltransferase (GAT) gene (gat4601) and the gm-hra gene 

which confers tolerance to ALS inhibiting herbicides. Based on these data and since the extracts for 

the concurrent recovery samples were stored for the same intervals as the extracts for the treated 

samples and acceptable recoveries were attained, the storage intervals for the extracts and RACs 

have been validated. Although storage stability data are not available for the processed commodities, 

HED will not require additional storage stability data for meal and oil in consideration of the 

physical similarities between canola meal and seed and the relatively short storage duration for oil.
Glyphosate Summary of Analytical Chemistry and Residue Data DP#: 394964

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860.1480 Meat, Milk, Poultry, and Eggs

The current petition results only in a change to the canola seed glyphosate tolerance expression 

(tolerance values are not changing). Therefore, the currently-established livestock tolerances remain 

appropriate. 

860.1500 Crop Field Trials (48398602.der1)

Canola field trials were conducted during the 2009 growing season in Canada and the U.S. 

reflecting different use patterns for the two countries. Ten trials were conducted using the 

Canadian application scenario in the North American Free Trade Agreement (NAFTA) Growing 

Zones 5 (n=1), 7 (n=2), and 14 (n=7) and six trials were conducted using the U.S. application 

scenario in Zones 2 (n=1), 5 (n=1), 7 (n=2), and 11 (n=2). HED notes that these figures

incorporate a reduction in the number of the trails conducted in Zone 14 (14 to 7; Canadian 

application scenario) and 11 (3 to 2; U.S. application scenario) due to classification of trials 

conducted at the same location as replicates and incorporates a reduction in the number of trials 

conducted in Zone 5 (2 to 1; U.S. application scenario) due elimination of trial as a result of 

overspray from a neighboring field. 

At each field trial location, canola (Event DP-Ø73496-4) was planted and treated with the 4.17 lb 

ae/gal soluble soluble-concentrate/liquid (SL/L) formulation of the potassium salt of glyphosate 

(Touchdown® Total herbicide; EPA Reg. No. 100-1169). Table 6 is a summary of the Canadian 

and U.S. application scenarios. For the Canadian field trials, white mesh tents were used to 

cover the field plots. The tents reflected some light, resulting in plants that were taller and 

thinner than normal. The tents also created a warmer, more humid environment, which 

prolonged flowering and delayed maturity beyond that for normal un-tented plants. The study 

indicated that consequently: the trials ran late into the season; it was harder to determine staging 

and timing; and in some provinces, the crops were affected by frost before sampling occurred. 

Canola forage samples were harvested from the Canadian and U.S. trials at the earliest forage 

stage but not before 60 days after planting. Forage preharvest intervals (PHIs) were 35-82 days 

for the Canadian trials and 27-55 days for the U.S. trials (interval after the second application for 

both the Canadian and U.S. trials). Mature canola seed was harvested 6-8 days following the 

third application in the Canadian trials, and 55-77 days following the second application in the 

U.S. trials. In some trials, seed was allowed to dry in the field for 3-13 days prior to collection. 

Residue decline samples were also collected at one of the Canadian U.S. trials. The harvested 

samples were analyzed for residues of glyphosate, N-acetyl-glyphosate, AMPA, and N-acetyl 

AMPA using the DuPontTM HPLC/MS/MS method 15444 (all residues were quantitated in 

glyphosate equivalents). The method was adequately validated and is appropriate for data 

collection (storage interval has also been validated). Based on the LLMV, the LOQ is 0.05 ppm 

for analyte/matrices. Table 7 is a summary of the residues. 

The residue decline trial conducted using the Canadian application scenario indicated the 

following: (1) forage - residues of N-acetyl-glyphosate showed a slight decrease as the PHI 

increased (residues of glyphosate, AMPA, and N-acetyl-AMPA were <LOQ) and (2) seed -

residues of glyphosate increased and residues of N-acetyl-glyphosate were consistent as the PHI 

increased (combined glyphosate and N-acetyl-glyphosate showed an increase as the PHI 

increased; residues of AMPA and N-acetyl-AMPA were <LOQ). 
Glyphosate Summary of Analytical Chemistry and Residue Data DP#: 394964

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The residue decline trial conducted using the U.S. application scenario indicated the following: 

(1) forage - residues of N-acetyl-glyphosate increased as the PHI increased (residues of 

glyphosate, AMPA, and N-acetyl-AMPA were <LOQ) and (2) seed - residues of N-acetylglyphosate were consistent as the PHI increased (residues of glyphosate, AMPA and N-acetylAMPA were <LOQ). 

Conclusions: A tolerance for residues of glyphosate per se in/on canola seed at 20 ppm is 

currently established (HED does not consider canola forage to be a food/feed commodity). The 

submitted field trial data resulted in combined glyphosate and N-acetyl-glyphosate residues in/on 

seed of ≤2.0 ppm following treatment at 1.3x the proposed maximum seasonal application rate 

(PHI = 55-77 days; glyphosate and N-acetyl-glyphosate are the residues of concern in DuPontTM 

Optimum® GLY Canola). To fulfill the geographical data requirements specified in OPPTS 

860.1500, additional canola field trials in NAFTA Growing Zones 5 (n=1) and 11 (n=1) are 

required. Since combined residues of glyphosate and N-acetyl-glyphosate residues were less than 

the currently established tolerance even under the higher rates associated with Canadian 

application scenario, HED concludes that no additional field trial data are required. Based on the 

available data and the revisions to the canola seed tolerance expression as a result of the 

introduction of DuPontTM Optimum® GLY Canola, HED concludes that the currentlyestablished 20 ppm canola seed tolerance should be deleted from 180.364(a)(1) and established 

in 180.364(a)(2). 

Table 6: Summary of Application Scenarios.

n

1

lb ae/acre (kg ae/ha)

preemergence 6-leaf stage (foliar) 7-days before 

harvest (foliar) total

Canadian Application 

Sceario1 10 0.59-0.62(0.66-0.70)

0.59-0.62

(0.66-0.70)

0.78-0.83

(0.88-0.93)3

forage - 1.18-1.24 (1.32-1.40)4

seed - 1.99-2.26 (2.23-2.53)4

U.S. Application 

Scenario2 6

1.55-1.60

(1.73-1.80)

0.56-0.57

(0.62-0.64) -- forage and seed - 2.11-2.17 (2.36-2.43)

1

 RTIs were 30-50 days between the first and second applications and 72-93 days between the second and third applications. All 

applications were made in spray volumes of 5.3-10.7 GPA (50-100 L/ha). An adjuvant was not added to the spray mixture for 

any application.

2

 RTIs were 20-34 days, and applications were made in spray volumes of 5-30 GPA (47-281 L/ha). Ammonium sulfate fertilizer 

was added to the tank mixtures at ~10-17 lb/100 gal. 

3

 One trial had a final application rate of 1.06 lb ae/acre (1.193 kg ae/ha); this trial did not result in the maximum residues.

4

 Forage harvested after the 6-leaf stage application and prior to the final application; seed harvested after the final application. 
Glyphosate Summary of Analytical Chemistry and Residue Data DP#: 394964

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Table 7: Summary of Residue Data from DP-Ø73496-4 Canola Crop Field Trials with Glyphosate.

Comm.

Total App. Rate,

lb ae/acre (kg ae/ha)

PHI 

(days)

Residue Levels, Glyphosate Equivalents (ppm)1

n

Sample

Min.

Sample

Max. LAFT HAFT Median Mean Std. Dev.

Canadian Trials2

Glyphosate

Forage 1.18-1.28 (1.32-1.44) 35-82 10 <0.05 0.26 0.05 0.11 0.05 0.06 0.02

Seed 1.99-2.26 (2.23-2.53) 6-8 10 0.29 9.9 0.44 9.0 1.7 2.7 2.7

N-acetyl-glyphosate

Forage 1.18-1.28 (1.32-1.44) 35-82 10 <0.05 1.1 0.08 0.81 0.26 0.31 0.23

Seed 1.99-2.26 (2.23-2.53) 6-8 10 0.17 14 0.18 12 0.35 1.7 3.5

AMPA

Forage 1.18-1.28 (1.32-1.44) 35-82 10 <0.05 0.07 0.05 0.06 0.05 0.05 <0.01

Seed 1.99-2.26 (2.23-2.53) 6-8 10 <0.05 0.09 0.05 0.07 0.05 0.05 0.01

N-acetyl AMPA

Forage 1.18-1.28 (1.32-1.44) 35-82 10 <0.05 <0.05 0.05 0.05 0.05 0.05 NA

Seed 1.99-2.26 (2.23-2.53) 6-8 10 <0.05 0.35 0.05 0.34 0.05 0.08 0.09

Combined glyphosate and N-acetyl-glyphosate

Forage 1.18-1.28 (1.32-1.44) 35-82 10 <0.10 <1.2 0.13 0.86 0.31 0.36 0.22

Seed 1.99-2.26 (2.23-2.53) 6-8 10 0.54 15 0.71 12 2.7 4.4 3.9

U.S. Trials3

Glyphosate

Forage 2.11-2.17 (2.36-2.43) 27-55 6 <0.05 <0.05 0.05 0.05 0.05 0.05 NA

Seed 55-77 6 <0.05 <0.05 0.05 0.05 0.05 0.05 NA

N-acetyl-glyphosate

Forage 2.11-2.17 (2.36-2.43) 27-55 6 <0.05 2.8 0.07 2.7 0.38 0.73 1.0

Seed 55-77 6 0.06 1.9 0.14 1.5 0.26 0.47 0.51

AMPA

Forage 2.11-2.17 (2.36-2.43) 27-55 6 <0.05 <0.05 0.05 0.05 0.05 0.05 NA

Seed 55-77 6 <0.05 <0.05 0.05 0.05 0.05 0.05 NA

N-acetyl AMPA

Forage 2.11-2.17 (2.36-2.43) 27-55 6 <0.05 0.14 0.05 0.13 0.05 0.06 0.03

Seed 55-77 6 <0.05 0.69 0.05 0.55 0.06 0.14 0.20

Combined glyphosate and N-acetyl-glyphosate

Forage 2.11-2.17 (2.36-2.43) 27-55 6 <0.10 <2.9 0.12 2.8 0.43 0.78 1.0

Seed 55-77 6 <0.11 2.0 0.19 1.6 0.30 0.53 0.53

1

 Except for sample min/max, values reflect per-trial averages; n = no. of field trials. For calculation of LAFT (lowest-averagefield-trial), HAFT (highest-average-field-trial), median, mean, and standard deviation, the LOQ (0.05 ppm) was used for any 

results reported as <LOQ.

2

 Includes the side-by-side trial reflecting the Canadian use pattern conducted in the U.S. (Trial 21).

3

 Trial 19 is not included as the study indicated that this trial may have received overspray due to drift from a neighboring field.
Glyphosate Summary of Analytical Chemistry and Residue Data DP#: 394964

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860.1520 Processed Food and Feed (48398602.der2)

The petitioner submitted an adequate processing study conducted with canola (Event DPØ73496-4). Three trials were established with each trial receiving application of a 4.17 lb ae/gal 

potassium glyphosate SL/L formulation as follows: (1) a preemergence soil application at 0.60-

0.64 lb ae/acre; (2) a foliar broadcast application made at the 6-leaf stage at 0.60-0.64 lb ae/acre; 

and (3) a foliar broadcast application made 6-7 days before harvest at 4.02-4.11 lb ae/acre. The 

total seasonal rate was 5.24-5.39 lb ae/acre (3.2x). Canola seed was harvested at a 6-7 days after 

the final application and were allowed to dry in the field for 4-10 days prior to collection. The 

seed from each trial was processed meal and oil using cold press and solvent extraction 

procedures. 

The seed and processed samples were analyzed for residues of glyphosate, N-acetyl-glyphosate, 

AMPA, and N-acetyl AMPA using the DuPontTM HPLC/MS/MS method 15444 (all residues 

were quantitated in glyphosate equivalents). The method was adequately validated and is 

appropriate for data collection (storage interval has also been validated). Based on the LLMV, 

the LOQ is 0.05 ppm for analyte/matrices. 

Table 8 is a summary of the canola oil and meal processing factors. Residues of glyphosate, Nacetyl-glyphosate, AMPA, and N-acetyl AMPA do not concentrate in refined oil. Residues of Nacetyl-glyphosate and N-acetyl-AMPA did concentrate in canola meal with processing factors of 

1.1-1.5x and 1.9-2.1x, respectively. Residues of glyphosate, AMPA, and combined residues of 

glyphosate and N-acetyl-glyphosate did not concentrate in canola meal. Based on these results, a

separate tolerance in/on canola meal and oil for the combined residues of glyphosate and Nacetyl-glyphosate are unnecessary. 

Table 8: Summary of Average Processing Factors.

Analyte canola meal canola oil

cold press solvent extraction cold press solvent extraction

glyphosate 0.8x 0.6x <0.004x

N-acetyl-glyphosate 1.5x 1.1x <0.05x

AMPA 0.4x <0.4x <0.4x

N-acetyl-AMPA 2.1x 1.9x <0.1x

combined glyphosate and N-acetyl-glyphosate 1.1x 0.9x <0.005x
Glyphosate Summary of Analytical Chemistry and Residue Data DP#: 394964

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860.1850 & 860.1900 Confined and Field Accumulation in Rotational Crops

Based on previously-reviewed confined rotational crop studies (3.71 lb ae/acre (2.2x); MRIDs 

41543201 and 41543202, A. Abramovitch, 14-Oct-1992) which resulted in glyphosate residues 

of >0.01 ppm in barley grain only (0.018 ppm; 125-day PBI) and the LOQ for the tolerance 

enforcement method of 0.05 ppm, HED previously concluded that a 30-day PBI was appropriate 

for all non-labeled crops (D200041, G. Kramer, 12-May-1994; field rotational crop study has not 

been submitted). Based on the LOQ for the tolerance-enforcement method and since the 

confined rotational crop study employed an exaggerated rate and employed application to bare 

soil, a 30-day PBI for all non-labeled crops is acceptable following application to Optimum® 

GLY canola. A revised Section B is requested. 

860.1550 Proposed Tolerances

Based on the available data, HED agrees with the petitioner’s proposal to move the currentlyestablished 20-ppm canola seed tolerance from 180.364(a)(1) where the tolerance expression is 

for residues of only glyphosate to 180.364(a)(2) where the tolerance expression is for combined 

residues of glyphosate and N-acetyl-glyphosate (expressed in glyphosate equivalents). Canadian 

and Codex maximum residue limits (MRLs) are currently established in/on canola seed at 10 

ppm (combined residues of glyphosate and AMPA) and 20 ppm (residues of glyphosate only), 

respectively (see attachment 2). HED notes that DuPont has also petitioned the Canadian 

Pesticide management Regulatory Agency (PMRA) concerning application of glyphosate to 

Optimum® GLY; PMRA indicated that they are willing to increase the canola seed glyphosate 

MRL to 20 ppm. Therefore, the U.S., Canadian, and Codex tolerance levels will be harmonized 

but the tolerance expressions will be different. 

Attachment 1 - Chemical Name and Structure Table

Attachment 2 - IRL Sheet

RDI: RAB1 Chemists (16-Nov-2011)

T. Bloem:S10945:Potomac.Yard 1:703-605-0217:7509P:RAB1
Glyphosate Summary of Analytical Chemistry and Residue Data DP#: 394964

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Attachment 1 - Chemical Structures

Compound Structure

Glyphosate

N-(phosphonomethyl)glycine

P

O

OH

OH

N

H

HO

O

N-acetyl-glyphosate

N-acetyl-N-(phosphonomethyl)glycine

AMPA

(aminomethyl)phosphonic acid

P

O

OH

OH

H2N

N-acetyl-AMPA

[(acetylamino)methyl]phosphonic acid

P

O

OH

OH

N

H

H3C

O

P

O

OH

OH

N

HO

O

H3C O
Glyphosate Summary of Analytical Chemistry and Residue Data DP#: 394964

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Attachment 2: IRL

International Residue Limits: Glyphosate (417300; 11/02/2011)

Summary of US and International Tolerances and Maximum Residue Limits 

Residue Definition

US Canada Mexico2 Codex3

40 CFR 180.364(a)(2): glyphosate (N-

(phosphonomethyl) glycine) and its metabolite N-acetylglyphosate (N- acetyl-N-(phosphonomethyl)glycine)

calculated as the stoichiometric equivalent of glyphosate

Plants: N-(phosphonomethyl)glycine, 

including the metabolite 

aminomethylphosphonic acid

glyphosate

Tolerance/Maximum Residue Limit (ppm)

Commodity1 US Canada Mexico2 Codex3

canola seed 20 10 20

Completed: M. Negussie; 11/03/11

1

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

proposed by the applicant.

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. 

MRLs indicated as proposed have not been finalized by the CCPR and the CAC.