UNITED STATES ENVIRONMENTAL PROTECTION AGENCY

WASHINGTON, D.C. 20460

PREVENTION, 

PESTICIDES AND

TOXIC SUBSTANCES

July 13, 1999

MEMORANDUM

SUBJECT: Trifloxystrobin. Results of HED Metabolism Assessment 

Review Committee Meeting held 6/15/99. DP Barcode D257835

TO: George Kramer, Executive Secretary

HED Metabolism Assessment Review Committee

FROM: Fred Ives, Chemist

Health Effects Division (7509C)

and

Bill Greear, Toxicologist

Registration Action Branch 3

Health Effects Division (7509C)

THRU: Richard Loranger, Chair

HED Metabolism Assessment Review Committee

Health Effects Division (7509C)

A. Summary of Deliberations

The Committee reviewed and discussed material in the HED briefing 

memorandum for the fungicide trifloxystrobin 

[(E,E)-methoxyimino-[2-[1-(3-trifluoromethy-phenyl)-ethylideneam

inooxymethyl]-phenyl]acetic acid methyl ester, (code 

CGA-279202)(Fred Ives, Maxie Nelson, William Greear, 6/9/99) and in 

an EFED memo for the same chemical on water (Ron Bloom, 6/9/99). 

Included in the HED memo was a summary of plant (apple, cucumber, 

peanut, wheat) and animal (rat, goat, hen) metabolism, rotational 

crop, analytical methodology and magnitude of the residue studies. 

Also included in the HED memo were two directed questions:

1. Does the Committee concur with the petitioner’s recommendation 

that trifloxystrobin be the only residue regulated in plant and 

animal commodities?

2. What residue does the committee recommend for risk assessment 

purposes.

The Committee considered a number of factors in its deliberations: 
2

that both acute (2.5 mg/kg bw)and chronic (0.05 mg/kg bw) reference 

doses had been estimated, that EFED had some concern for the 

environmental persistence of the primary environmental degradant and 

acid metabolite CGA-321113, the lack of any substantial toxicology 

information for CGA-321113, that metabolism indicated a substantial 

qualitative similarity among animals and to a large extent between 

animals and plants, that generally the acid metabolite was the 

primary residue in animal metabolism, and its presence (or that of 

its conjugates) was also observed in plants. The Committee further 

noted substantial differences between metabolism in fruiting 

vegetables and fruits compared to peanuts (especially the nutmeat) 

and wheat, the low amount of TRR identified in wheat, the relatively 

high proportional levels of metabolite L7a (taurine conjugate of 

trifloxystrobin) especially in liver (to a lesser extent in milk and 

kidney) and the presence of trifluoroacetic acid as the major residue 

(on the order of 0.1 ppm or slightly higher) in rotational crops. 

There was some discussion on the significance of cyano compounds 

among the metabolites. The meeting also was unable to discern 

whether the configurational isomers of trifloxystrobin and its acid 

metabolite are determined by the analytical method. It did note that 

the analytical method will determine both parent and acid metabolite 

(as separate GC peaks) and that the analytical method does not contain 

a hydrolysis step.

B. MARC Decisions

1. Regarding the two questions posed to the Committee, the MARC 

concluded that trifloxystrobin and the free form of its acid 

metabolite CGA-321113 were residues of concern in both plants and 

animals for both regulatory and risk assessment purposes. 

2. Additionally, for risk assessment purposes the MARC concluded 

that the contribution of metabolite L7a from liver should be taken 

into account and assumed to have equal toxicity to 

trifloxystrobin.

3. Concerning the cyano compounds, it was concluded that these were 

not of concern as they are aromatic nitriles (as opposed to 

aliphatics). 

4. Concerning the trifluoroacetic acid metabolite found in the 

rotational crop studies, MARC concluded that it was not of 

toxicological concern at the 0.2 ppm levels observed. 
3

5. The meeting concluded that the fate of residues in animals is 

adequately understood. In the case of plants the MARC concluded 

that the fate of residues is understood for fruits, fruiting 

vegetables, cucurbit vegetables and peanuts. 
4

6. The MARC concluded that any future petitions for uses on leafy 

vegetables, cereals or other crops not specified in 5 above would 

require additional metabolism studies.

7. The MARC recommended that clarification be sought from the 

registrant as to whether the configurational isomers of 

trifloxystrobin and its acid metabolite CGA-321113 are determined 

by the analytical procedure.

8. The residues of concern in drinking water are the parent and acid 

metabolite. The latter is more persistent and more mobile than 

the parent.

C. Attendance

1. MARC Members attending

Sanjivani Diwan Richard Loranger (chair)

Leung Cheng Chris Olinger

George Kramer Alberto Protzel

Nancy Dodd William Wassell

2. MARC Members absent

Kit Farwell, Nelson Thurman, John Doherty

3. Other responsible scientists participating

Mary Rust Maxie Nelson

Fred Ives Stephen Dapson

William Greear Ghazi Dannan

Ron Bloom Stephen Carey

cc: HED Metabolism Committee file (G.Kramer), Cynthia 

Giles-Parker/Janet Whitehurst (PM 22), Francis (Dick) 

Griffith (BEAD-7503C), Mary Rust, PP#8F04955

T:\HED\SARC\METABOLI\129112MT.MEM

c:\fives\wpdoc\HED99\trflxyMARCdec3LAN.wpd
UNITED STATES ENVIRONMENTAL PROTECTION AGENCY

WASHINGTON, D.C. 20460

OFFICE OF

 PREVENTION, PESTICIDES AND

TOXIC SUBSTANCES

MEMORANDUM

DATE: June 9, 1999

SUBJECT: PP#8F04955. Reduced Risk New Chemical: Trifloxystrobin (ISO-proposed common name 

CGA-279202) in or on imported bananas, cucurbit vegetables, grapes, peanuts, peanut hay, pome fruit 

and apple pomace. Briefing memorandum for June 15, 1999 Meeting of the MARC.

FROM: Fred Ives, Chemist

HED (7509C)

and

Maxie Jo Nelson, Chemist

RAB2/HED (7509C)

and

William Greear, Toxicologist

Registration Action Branch 3/HED (7509C)

THRU: Richard Loranger, Branch Senior Scientist

RAB2/HED (7509C)

TO: George Kramer, MARC Executive Secretary

RAB1/HED (7509C)

This memo provides a briefing for the June 15, 1999 MARC consideration of trifloxystrobin. The chemistry 

portion is based on the contractor review and is intended to be in accordance with the 6/23/98 MARC SOP. 

Attached to this memo for reference you will find:

1. Table 1 - Trifloxystrobin metabolism in plants - a summary. This 

summary information was extracted as representative information 

from tables presented in the contractor review.

2. Tables 18a and 18b. These are summaries (TFMP and GP labels 

respectively) of the goat metabolism study taken from the 

contractor review. 

3. Table 21a taken from the contractor review. It summarizes 

metabolites found in poultry with the TFMP label and is intended 

to be representative for the committee, although information on 

the GP label is also available.
2

4. Plant metabolism schematics - No metabolism schematics are 

included in the contractor review. Fig. 35 apple metabolism 

schematic is a good representative for plant metabolism. It is 

taken from MRID 444968-22. MRID schematics from original studies 

are also attached for cucumber (Fig. 18), wheat (Fig. 14) and 

peanuts (Fig. 204).

5. Animal metabolism schematics - Fig. 42, a goat metabolism study 

from MRID 444968-18 is a good representative for animal 

metabolism. Also attached are hen metabolism schematics (Fig. 

45 and 32).

6. Fig. 34 Rat metabolism as part of the toxicology contribution.

7. Attachment II, Fig. 1 taken from the contractor review. It 

summarizes chemical names and structures of trifloxystrobin in 

plants and animals and identifies the matrix in which each was 

identified.
3

TRIFLOXYSTROBIN

RESIDUE CHEMISTRY SECTION

Use information/Identification of Chemical

Trifloxystrobin 

[(E,E)-methoxyimino-[2-[1-(3-trifluoromethyl-phenyl)-ethylideneam

inooxymethyl]-phenyl]acetic acid methyl ester], [code CGA-279202] is 

one of a new class of β-methoxyacryl ester (MAE) fungicides which are 

synthetic analogs of strobilurin A, an antifungal metabolite of the 

fungus Strobilurus tenacellus. Via PP#8F04955 Novartis Crop 

Protection, Inc. proposes permanent tolerances for CGA-279202 (parent 

compound only) in conjunction with Section 3 uses of a 50% water 

dispersible granular formulation (CGA-279202 50WG) on imported 

bananas (0.1 ppm), cucurbit vegetables (0.25 ppm), grapes (1.5 ppm), 

peanuts (0.02 ppm), peanut hay (4.0 ppm), pome fruits (0.4 ppm) and 
4

apple pomace 1.5 ppm). While three conformational isomers of 

trifloxystrobin (CGA-331409, CGA-357261 and CGA-357262) may occur at 

low levels, they are not proposed to be included in the tolerance nor 

is whether they would be determined by the proposed analytical method 

addressed. It is likely that they would be determined if present.
5

Uses in summary:

No. Interval lb ai/

PHI

Formulation Appl. Days lb. ai/appl 

seaso

n

Days

bananas* 75 g/L EC 10 — 0.1 1

—

cucurbits 50% WDG 8 7-14 0.063-0.125 1 0

grapes 50% WDG 6 7-21 0.063-0.094 0.563

14

peanuts 50%WDG 8 10-14 0.063-0.094 1

14

pome fruits 50%WDG 5 7-14 0.063-0.094 0.3125

14

* Good Agricultural Practices (GAP = use information) for 

bananas is lacking. That indicated was the use in field 

trials.

Summary of Plant and Livestock Metabolism Studies

Plants

The nature of residue studies for trifloxystrobin in plants have been 

conducted on apples, cucumbers, peanuts and wheat. In separate 

experiments for each crop trifloxystrobin is labeled in the 

glyoxylphenyl (GP) or trifluoromethylphenyl (TFMP) rings. Initial 

extractions generally were with acetonitrile (ACN):water, except in 

the case of peanut nutmeats it was with hexane. A wide variety of 

separative techniques were utilized, including liquid/liquid 

partitioning, column chromatography and analyses by TLC (1-D, 2-D, 

normal phase, reversed phase), LSC or combustion/LSC, HPLC, MS 

(different types), NMR and confirmations via derivatization. 

Various fractions were subjected to acid, base or enzymatic hydrolysis 

prior to further analyses. Table 1, extracted from the contractor’s 

review, briefly summarizes representative results for apples, 

cucumbers, peanuts and wheat.

Metabolism indicated for plants includes hydrolysis of the methyl 

ester to the CGA-321113 acid metabolite, hydroxylation on the TFMP 

ring, oxidation of the aminooxymethyl group on the bridge, 
6

conjugations with sugars and breakdown of the methoxyiminoacetic acid 

side chain and minor cis/trans isomerization.

Apples - 67-95% of TRR was characterized/identified in foliage, 

surface rinse, peel and flesh. From the GP label, radioactivity was 

86.9, 11.2 and 1.9% in surface rinse, peel and flesh one hour after 

the fourth GAP application (similar with the TFMP label). At the 14 

day proposed GAP PHI with both labels trifloxystrobin was the major 

residue (ca. 81-83%) in whole apple with three cis-trans isomers of 

trifloxystrobin (CGA-331409, 357261, and 357262) at < 5.2% each or 

<11% total of the TRR. Approximately 1% was the acid metabolite 

CGA-321113 or its two isomers CGA-373466 or -373465. Lesser 

metabolites were also reported. Results were similar with both 

labels.
7

Cucumbers - Residues were characterized from the GP and TFMP labels 

in foliage and in both small and large greenhouse grown cucumbers which 

were sampled after 3 applications at approximate GAP, both at the 0 

day PHI (1 hr.) and after one day and 7 days. Although the TRR in 

absolute terms was higher (3-8 times) after 7 days in the small 

cucumbers (2.3 ppm GP label) compared to large cucumbers (0.3 ppm GP 

label) and somewhat less with the TFMP label, similar metabolite 

profile results were obtained at all PHIs, with both labels and with 

either cucumber size. Metabolite profile results were similar for 

fruit and foliage and markedly similar to those obtained with apples. 

As an example, after 7 days residues as %TRR in fruit of small cukes 

was 87% trifloxystrobin, and its cis/trans isomers 2% CGA-331409, 0.9% 

-357261, and <0.1% -357262. Trifloxystrobin’s acid metabolite 

CGA-321113 was 2.3%. Low residues of other metabolites were also 

reported.

Peanuts - Starting at bloom stage peanut plants were treated 4 times 

(the latter at nut maturity) at ~0.5 lb. ai/A or ~2X the GAP seasonal 

rate with either the GP or TFMP labels. Nutmeat, hay and immature 

vine samples were taken. Immature vines were sampled on the day of 

1

st application and 14 days after the 2nd application. Mature plants 

were sampled 14 days (i.e. at GAP) after final application and 

separated into nutmeats, hulls and hay. 

The residue profile in nutmeat was markedly different than in apple 

and cucumbers and to a lesser extent even in hay and immature vines. 

This resulted in subjection of peanuts samples to more elaborate 

separative and analytical approaches than required for apples and 

cucumbers. In nutmeat with the GP label only about 45% of the TRR 

was identified. Only 2% of the TRR was trifloxystrobin, ~24% was 

shown to be in the form of triglycerides, 3.4% “metabolite B”, 8.8% 

“metabolite A” (glucoside conjugate of metabolite B hydrolyzed to the 

acid at the methyl ester) and 6% phthalic acid. Unidentified TRR was 

characterized with via different solvent partitions and after 

hydrolysis with acid, base and enzymes, but were unidentified. A 

somewhat similar result was found with the TFMP label, especially the 

low relative portion of trifloxystrobin and relatively similar 

triglyceride levels.

In the case of hay and immature vines the residue profiles were 

similar, and closer to that found in apple and cucumber. 

Trifloxystrobin was on the order of 30% TRR, plus about 6% of TRR in 

the form of cis/trans isomers of trifloxystrobin and about 10-15% as 

the acid metabolite CGA-321113, its isomers, or conjugates.
8

Wheat - Although uses are not proposed for wheat, metabolism studies 

were conducted on spring wheat with both the GP and TFMP labels. 

Included were penetration investigations. Plants were treated at late 

boot (41 days) and early ear emergence (58 days) at ~0.22 lb ai/A (~0.44 

lb ai/season). Wheat shoots were collected 1 hr. after the 1

st and 

2

nd applications, at 50% maturity (24 hrs after 2nd application) and 

at maturity (52 days after 2nd application). 

Contrasted to the apple, cucumber and peanut, only low levels (<15% 

of TRR) were identified in the metabolism studies. In stalk for the 

GP label only about 4% was trifloxystrobin, about 2% its 

conformational isomers. Similar results with somewhat lower 

trifloxystrobin was found for grain, hulls and straw. Different from 

studies with apple, cucumber and peanuts were significant levels 

(5-10%TRR) of metabolites NOA-413161 (methyl group of 

ethylideneaminooxymethyl bridge of trifloxystrobin metabolite 

CGA-321113 oxidized to the acid) and its conformational isomer 

NOA-413163 and in hulls and straw 4 and 5.4% respectively of metabolite 

NOA-414412 (CGA-321113 hydroxylated on the CF ring). Also noteworthy 

was the 10-15%TRR in stalk and grain as sugar conjugates. Results 

for the TFMP label were similar.

In the penetration part of the investigation one hour after a single 

application trifloxystrobin accounted for 89% of surface rinse, 

decreasing to 76.5% after 14 days. Of the penetrated radioactivity 

trifloxystrobin accounted for 73.5% after 1 hour and only 9.1% after 

14 days. Most of the remaining radioactivity in the surface rinse or 

penetrated consisted of trifloxystrobin conformational isomers or its 

acid metabolite CGA-321113.

Livestock

Metabolism studies were conducted in lactating goats and poultry. 

Indicated metabolism for livestock includes hydrolysis of the methyl 

ester to form the CGA-321113 acid metabolite, demethylations, ring 

and chain hydroxylations and conjugation. 

Goats 

Two lactating goats were administered either GP- or TFMP-labeled 

trifloxystrobin for 4 consecutive days via gelatin capsule at 

approximately 100 ppm in the diet (ca. 23X maximum theoretical dietary 

burden). Milk was collected twice daily and the animals were 

sacrificed 6 hours after the last dose for collection of liver, kidney, 

bile, fat (perirenal and omental) and muscle (leg and tenderloin). 
9

Using procedures similar to those for plant metabolism studies, 

samples were analyzed for TRR and for 

identification/characterization. Results are summarized in Tables 

18a (TFMP label) and 18b (GP label) taken from the contractor’s review.

Except for 54.7% and 60.3% in liver with the TFMP and GP labels 

respectively, > 68.9% of TRR was identified in all matrices. 

Trifloxystrobin was a significant or major portion of the residue in 

all matrices except liver and kidney (1.5-2.1% TRR) and somewhat for 

muscle where it was < 23% TRR. Where triflxoystrobin was not a 

significant or major portion of the TRR in the matrices, its acid 

metabolite CGA-321113 was. 

In milk 48.5-60% of the TRR was trifloxystrobin and other metabolites 

each approximately 4% or less, except for 10.3% metabolite L7a 

(aminoethylsulfonic acid conjugate of the acid metabolite (TFMP 

label). In muscle trifloxystrobin was 16.8 - 22.8% of TRR and its 

acid metabolite CGA-321113 43.9-49.3% of TRR. Other identified 

metabolites were <2.4% of TRR each. In fat trifloxystrobin 

accounted for 68-71.8% of TRR and its acid metabolite CGA-321113 

8.8-9.3%. Other identified metabolites were each <0.5% TRR.

In liver slight quantitative differences are observed between the GP 

and TFMP labels, although as indicated, in both cases unchanged 

trifloxystrobin accounted for only a small portion of the TRR. The 

acid metabolite CGA-321113 was 10.1-31.6% of TRR, metabolite L7a 

4.2-20.3% TRR, metabolite L7b (glycine conjugate of CGA-321113) ca. 

9% TRR. Evidence suggests that CGA-357276 at 7% TRR with the TFMP 

label resulted from CGA-321113 due to a microwave extraction step. 

CGA-166988 was reported at 8% of TRR with the GP label. Other 

identified metabolites were less than 2.2% TRR.

In kidney a somewhat similar profile to that in liver was observed, 

although the acid metabolite CGA-321113 was higher at 47.2-64.4% TRR 

and other identified metabolites <4.4% of TRR, except for MET L7a at 

11% of TRR.

Poultry

Two separate groups of five hens were administered via gelatin capsule 

either GP- or TFMP-labeled trifloxystrobin for 4 consecutive days at 

approximately 100 ppm dietary equivalent, estimated to be ~20,000X 

the maximum theoretical dietary burden. Eggs were collected twice 

daily and hens sacrificed 6 hours after that last dose for collection 

of liver, kidney, thigh and breast muscle, peritoneal fat and skin 
10

with attached fat. Approximately 74-87% of administered dose was 

eliminated via excreta. After 78 hours most of the radioactivity in 

eggs was in the yolk (2-3.6 ppm trifloxystrobin equivalent) compared 

to 0.07-0.3 ppm in the white. As determined by scintillation counting 

in individual tissues trifloxystrobin equivalent residues were 

similar between the two labels TFMP/GP, e.g. liver 3.9-8.6/4.6-8.1 

ppm, muscle 0.13-0.35/0.13-0.20 ppm, skin (with fat) 0.8-1.8/0.7-0.8 

ppm and peritoneal fat 1.9-2.8/0.8-1.3 ppm..

Separative and analytical procedures employed were somewhat similar 

to those used for ruminants and results for pooled tissues are 

summarized in tables 21a (TFMP label) taken from the contractor 

review. Trifloxystrobin was found in all matrices, except egg white, 

albeit at low levels (0.3-1.1% TRR) in liver. Compared to the 

ruminant studies, somewhat less of the radioactivity was identified 

in poultry matrices, especially in egg yolk ( 4.3-17.3 in egg yolk; 

27-61% in muscle; 34-50% in liver and higher at in 57-74% TRR in egg 

white; and 60-85% in fat + skin). Except in egg white, the acid 

metabolite CGA-321113 accounted for less of the TRR than in the 

ruminant studies with fewer individual metabolites contributing a 

major portion of identified residue.

In contrast to other poultry matrices, in egg white acid metabolite 

CGA-321113 was a significant portion of the TRR (10.3-19.5%). While 

CGA-357276 was reported at 28.9% of egg white TRR (TFMP label), it 

was thought to have resulted from sample work up degradation of MET 

1U or a conjugate thereof. Therefore, MET 1U (demethylated acid 

metabolite CGA-321113) is likely a major component of egg white TRR 

as confirmed at 21.7% with the GP label (not of other poultry 

matrices). The next highest was MET L13b (trifloxystrobin 

hydroxylated in the TFMP ring) at 3.6-8.5% TRR. As noted, in egg 

yolk <17.3% of TRR was identified. Up to 7% of that was 

trifloxystrobin with remaining metabolites common to other matrices, 

but each <4% of TRR. About 26-35% of TRR was designated as 

unidentified metabolite EGR10.

Although only 27% of TRR was identified in muscle with the GP label, 

with the TFMP label ca. 24% of TRR was trifloxystrobin. Significant 

residues with this label were also reported for MET 12U (8.8% TRR), 

MET 2F (demethylated trifloxystrobin) (8.1% TRR) and MET L14 (7.3% 

TRR), with other metabolites at <6% TRR each. In fat + skin 26-53% 

of TRR was reported as trifloxystrobin. Other major identified 

metabolites were MET 2F (12-14% TRR) and MET L13b (trifloxystrobin 

hydroxylated in the TFMP ring) (2.6-10.7% TRR). Other identified 

metabolites were <4.1% TRR, except for ME 3U (trifloxystrobin 
11

hydroxylated on ethylideneamino bridge and nitrated on the acetic 

acid methyl ester group) at 3.5-5.9% TRR

In liver only 0.3-1.1 % of TRR was trifloxystrobin. The major 

identified metabolites were MET 13b (6.4-11.2% TRR) and Met L14 (10.5% 

TRR, TFMP label). Except for CGA-357276 (5.6% TRR, GP label), other 

identified metabolites were each less than 5% TRR. 

Rotational Crop Studies

Rotational crop studies were conducted with both the GP- and TFMP 

labels in silt loam soil with applications at 2 lb ai/A (2X rate 

proposed for cucurbits and peanuts) to bare soil plots. Planted were 

turnips, spinach and wheat, components of each analyzed for TRR and 

for identification/characterization of residues at 30 and 120 days 

after application. Trifloxystrobin-equivalent residues were higher 

for the TFMP label than with the GP label. For the TFMP label 

trifloxystrobin equivalent after 30 /120 days were: turnip leaves 

0.06/0.04 ppm, turnip roots 0.02 ppm, spinach 0.25/0.26 ppm, 25% 

mature wheat forage 0.28/0.19 ppm, mature wheat forage 0.14/0.1 ppm, 

wheat straw 0.17/0.2 ppm and wheat grain 0.07/0.06 ppm. Generally 

with the GP label only a relatively small portion of the TRR in aqueous 

and organic soluble fractions was identified/characterized (6 - 46% 

TRR of each) with trifloxystrobin being <2% of TRR in each case, 

although one of its three conformational isomers also at about the 

same levels. The acid metabolite CGA-321113 was higher in some 

matrices (e.g. up to 8.5% of the DCM fraction of turnip leaves and 

17.6% for turnip root (0.003 ppm). 

With the TFMP label the identified/characterized % TRR was much 

higher, especially in aqueous fractions where 37- >100% TRR was 

characterized/identified.. Again, only low levels of 

trifloxystrobin, its conformational isomers, the acid metabolite 

and its isomers and phthalic acid were reported, all generally <0.01 

ppm. Trifluoroacetic acid was by far the major residue identified 

with the TFMP label, ranging generally 20 - 94% of a given matrix TRR 

with the levels even for this generally being less than 0.2 ppm. 

Residue Analytical Methods

Analytical method AG-659A is proposed for enforcement of tolerances 

for trifloxystrobin on plant and animal commodities. It also 

determines residues of the acid metabolite of trifloxystrobin 

CGA-321113, although that is not proposed for inclusion in the residue 

for enforcement. It is based on extraction of samples (homogenized 
12

solid samples) with ACN:water (80:20 v:v, filtration, liquid:liquid 

partitioning with a three solvent system (sodium chloride saturated 

water, toluene and hexane), clean up on a C18 solid extraction column, 

partitioned into methyl-tert-butyl ether:hexane, concentrated to 

dry, taken up in 0.1% polyethylene glycol in acetone for GC analysis 

with a nitrogen phosphorus detection detector using a DBWAX capillary 

column. The reported limit of quantitation is 0.02 ppm in all 

matrices except peanut hay at 0.05 ppm LOQ and in milk at 0.01 ppm 

LOQ. 

The petitioner conducted method validation of both trifloxystrobin 

and CGA-321113 on fortified plant and animal commodities for the most 

part at the reported LOQ’s and at higher levels with average recoveries 

for CGA-321113 ranging from 70-120% and for trifloxystrobin at 

74-107%. An independent laboratory validation was conducted on 

method AG-659, which AG-659A supercedes. AG-659 (or REM 177.03 which 

is the same except with EC detection) was used in the field trials, 

rotational crop, storage stability and field trial studies. An 

independent validation was also conducted on AG-659A, but only for 

soil residues. As previously noted, while three conformational 

isomers of trifloxystrobin (CGA-331409, CGA-357261 and CGA-357262) 

may occur at low levels, they are not proposed to be included in the 

tolerance nor is whether they would be determined by the proposed 

analytical method addressed. It is likely that they would be 

determined if present. These same comments apply to CGA-3321113 and 

its conformational isomers CGA-3734466 and CGA-373465.

Multi-residue Methods

Study 44496829 (1997) reports on multiresidue testing for recovery 

of trifloxystrobin and CGA-321113. Trifloxystrobin was tested in 

accordance with PAM I Appendix II, through Protocols C (GLC systems), 

D (complete method without cleanup) and E (standards and fortified 

samples through cleanup columns with different solvent systems) and 

its acid metabolite CGA-32113 through Protocols B (methylation) and 

C. 

Trifloxystrobin gave adequate responses through protocol C; it was 

completely recovered from fortified apple sampes when analzyed 

through the Section 302 multiresidue method (Protocol D); standard 

solutions thereof were completely recovered through Protocol E 

Section 303 C2 (MeCL Florisil cleanup) and recovery was complete from 

fortified apple samples through Protocol E Section 303 E4/C2 complete 

method. It was not recovered though Section 303 C1 Florisil columns 

(ethyl ether/petroleum ether) or Section 303 E4/C1. 
13

CGA-321113 - Methylation through protocol B was effective. 

Unmethylated CGA-321113 gave adequate responses only on one GC 

system, but the methylated derivative (trifloxystrobin) on several. 

CGA-321113 was completely recovered through Section 402 C1a gel 

permeation cleanup (Protocol B - GPC) and residues from CGA-321113 

fortified apples was completely recovered through Section 402 E2/C1 

multiresidue method (extraction with methylene chloride).

Crop Field Trials and Livestock Feeding Studies

Crop Field Trials

Supervised trials have been conducted on apple, pear, peanuts, 

cucumber, cantaloupe, summer squash, grapes, processed products and 

bananas.

Bananas - The proposed tolerance is 0.1 ppm. At 0 or 7 days after 

that last of 10-11 applications of a 75 g/L EC formulation at 0.1 lb 

ai/A (presumed 1X GAP) maximum residues were 0.06 ppm in bagged bananas 

and 0.03 ppm in unbagged bananas on day 0, but 0.05 ppm on day two. 

2X data were also available. CGA-321113 was <0.02 ppm. Data are 

considered adequate if GAP is confirmed to be the same as that at which 

the trials were conducted. 

Cucurbits - The proposed tolerance is 0.25 ppm. At the proposed 1X 

per application and seasonal rate and proposed 0 day PHI residues of 

trifloxystrobin were 0.02-0.22 ppm in cucumber, 0.03-0.024 ppm in 

cantaloupe and <0.02-0.23 ppm in summer squash. In all cases 

CGA-321113 was <0.02 ppm. In other experiments at 1X seasonal rate, 

but 2X the per application rate maximum residues for trifloxystrobin 

were 0.03-0.28 ppm in cucumber, 0.03-0.58 ppm in cantaloupe and 

0.04-0.33 ppm in summer squash. CGA-321113 was 0.02 ppm except two 

summer squash samples at 0.02 -0.03 ppm. Decline data were also 

available. Geographic representation is judged to be inadequate, but 

an additional trial was to be conducted in Region 5, which will be 

submitted as an amendment. Adequacy of the proposed tolerance will 

be reconsidered at that time. HED likely will not support a 

fractional tolerance as proposed.

Grapes - The proposed tolerance is 1.5 ppm. At the 14 day proposed 

PHI and at 1.3X the proposed per application and per season rate 

triflolxystrobin residues were <0.02-1.1 ppm. CGA-321113 was 

<0.02-0.068 ppm. Data are considered adequate. HED may suggest 2 

ppm. While residues do not concentrate in juice, conflicting 
14

studies preclude a conclusion on the need for MRLs for raisins. In 

one study trifloxystrobin concentrated 2-2.5X and in another with 

measurable residues in grapes residues did not concentrate. A 3 or 

4 ppm limit for raisins may be needed.

Peanuts - The proposed tolerance is 0.02 ppm in peanut and 4.0 ppm 

in peanut hay. Triflxoystrobin or CGA-321113 residues were <0.02 

ppm from 2/3 to 4X proposed maximum GAP rates (0.5-3X seasonal rates). 

Trifloxystrobin residues in peanut hay from maximum proposed GAP were 

0.15-3.7 ppm and CGA-321113 0.089-0.67 ppm, at 14-17 days (14 days 

GAP PHI). Data are considered adequate. HED may suggest 5 ppm for 

peanut hay. Residues are not expected to result in meal or refined 

oil.

Pome fruits - The proposed tolerance is 0.4 ppm in pome fruit. At 

the approximate proposed per application rates and 1.1X seasonal rate 

and the proposed 14 day PHI trifloxystrobin residues were 0.09-0.37 

ppm in apple and 0.05-0.23 ppm in pears. CGA-321113 was <0.02-<0.03 

ppm in apple and <0.02-0.04 ppm in pears. Data are considered 

adequate. HED may suggest 0.5 ppm for pome fruit. A 1.5 ppm 

tolerance is proposed for apple pomace. That is judged too low. A 

4 or 5 ppm tolerance may be recommended for apple pomace based on 

the highest average field trial residue of 0.3 ppm and the average 

concentration factor (from 6 processing studies) of 13.3X.
15

Livestock Feeding Studies

The petitioner does not propose tolerances for animal commodities, 

but HED will require such since the maximum feeding levels were only 

4.6X compared to the agency policy to base the decision to exclude 

the need for tolerances on a 10X feeding rate.

Poultry

A poultry feeding study was not submitted. Since peanut meal is the 

only poultry feeding item and residues are not expected to exceed the 

0.02 ppm limit of determination, the expected dietary burden has been 

estimated at 0.005 ppm. Poultry metabolism studies were conducted 

at ca. 100 ppm (20,000X). Maximum trifloxystrobin-equivalent 

residues were 8.6 ppm (liver). On this basis no residues of 

trifloxystrobin are expected in poultry meat or eggs, thus no feeding 

study is required for the proposed uses.

Cattle

Feeding studies were conducted at 2, 6 and 20 ppm (0.5, 1.4 and 4.6 

times the 4.4 ppm dietary burdens estimated for dairy cattle 

respectively (5.2 ppm for beef cattle). Trifloxystrobin residues 

were less than the LOQ for milk, muscle, kidney and liver, but in 

omental and perirenal fat maximum residues were 0.05 and 0.06 ppm 

respectively. Acid metabolite was below the LOQ’s except for kidney 

(0.02 ppm) and liver (up to 0.09 ppm). If the metabolism committee 

concludes that trifloxystrobin is the only residue in need of 

regulation, the contractor recommends LOQ tolerances for milk, 

muscle, kidney and liver and 0.06 ppm for fat. 0.1 ppm may be 

preferable for fat.

International Considerations

No Codex, Canadian or Mexican limits are established for 

trifloxystrobin.

TOXICOLOGY SECTION

On May 11, 1999, the Health Effects Division's Hazard Identification Assessment Review Committee (HIARC) 

evaluated the toxicology data base of trifloxystrobin and selected doses and endpoints for acute and chronic dietary, 

as well as occupational and residential exposure risk assessments, and addressed the sensitivity of infants and 

children from exposure to trifloxystrobin as required by the Food Quality Protection Act (FQPA) of 1996. The HIARC's 

conclusions are presented below.
16

Acute Reference Dose (Acute RfD) Females (13 +years)

Study Selected: Developmental Toxicity - Rabbit §83-3b

MRID No.: 44496709

Executive Summary: In a developmental study (MRID# 44496709), 5 groups of 19 THOMAE RUSSIAN, 

Chbb:HM Rabbits from Dr. K. Thomae GmbH (Chemisch-Pharmazeutische Fabrik, 7950 Biberach, 

Germany) received CGA 279202 Technical (Trifloxystrobin; Purity: 96.4%; Batch No.: P.405009) in a 0.5% 

w/w aqueous solution of sodium carboxymethylcellulose at either 0, 10, 50, 250 or 500 mg/kg/day by oral 

gavage from gestation days 7 through 19, inclusive. Maternal observations and measurements included daily 

mortality, cage-side observations, body weights and food consumption determined on days 4, 7, 12, 16, 20, 24 

and 29 post coitum. The maternal animals were sacrificed on gestation day 29 and subject to a gross 

necropsy, the uterine contents were examined and the fetuses were examined for external, visceral or skeletal 

anomalies by standard techniques. 

Maternal systemic toxicity was seen as reduced body weights in the 250 and 500 mg/kg/day dose groups 

during dosing and reduced body weight gains in the 50 mg/k/g day dose group and above during the dosing 

period (gestation days 7-19, statistically significant at the 2 higher doses, p<0.01), for the dosing period plus 

post dosing period (gestation days 7-29) and for the entire gestation period (gestation days 0-29), also for the 

net body weight gain (gestation days 7-29) and corrected body weight gain (gestation days 0-29) and there was 

a rebound in body weight gain in the 250 and 500 mg/kg/day dose groups for the post dosing period (gestation 

days 19-29). There was reduced food consumption in the 50 mg/kg/day dose groups and above during the 

dosing period, for the dosing plus post dosing period and for the entire gestation period with a rebound in food 

consumption noted during the post dosing period. There was reduced food efficiency noted during the same 

time periods for the 50 mg/kg/day dose groups and above. The Maternal Toxicity NOAEL was 10 

mg/kg/day and the Maternal Toxicity LOAEL was 50 mg/kg/day based on reduced body weights, body 

weight gains, food consumption and food efficiency.

Developmental toxicity was observed as a very marginal increase in fetal and litter incidence of fused 

sternebrae #3 and 4 at the 500 mg/kg/day dose group, no other observations were noted. The 

Developmental Toxicity NOAEL was 250 mg/kg/day and the Developmental Toxicity LOAEL was 500 

mg/kg/day based on a marginal increased incidence of skeletal anomalies.

This study is classified as Acceptable-Guideline and satisfies the requirement (OPPTS 870.3700, OPP 

§83-3b) for a developmental toxicity study in rabbits. 

Dose and Endpoint for Risk Assessment: Developmental NOAEL based on increase in fetal incidence of 

fused sternebrae #3 and 4 at 500 mg/kg/day (LOAEL).

Comments about Study/Endpoint: The developmental effects are presumed to occur after a single expsoure 

(dose). Since this in an in utero effect, it is applicable only to the population subgroup Females 13+.

Uncertainty Factor (UF): 100 (includes 10x for intra-species extrapolation and 10x for inter-species 

variation).

Acute RfD = 250 mg/kg/day (NOAEL) = 2.5 mg/kg/day

100 (UF)

Chronic RfD

Study Selected: Chronic Toxicity - Dog §83-1b

MRID No.: 44496704
17

Executive Summary: In a chronic toxicity study (MRID # 44496704), 4 beagle dogs/sex/group were treated 

orally with 0, 2, 5, 50 or 200 mg/kg/day of CGA-279202 Technical (purity: 96.4%) in gelatin capsules for 12 

months. No mortality resulted from the treatment. Animals in the 200 mg/kg/day group suffered from an 

increased frequency and severity of diarrhea and vomiting. Dark discoloration of the hair, skin of the paws, 

thorax and abdomen was noted in animals in the 50 and 200 mg/kg/day treatment groups. Food consumption 

was significantly reduced (p<0.05) in the high dose female group for weeks 2 and 3. However, treatment did 

not affect the mean body weight values. Although some hematological parameters for the treated animals were 

statistically different from those of the control animals, none of the parameters demonstrated a significant 

treatment effect. Mean serum albumin levels were reduced for the males in the 50 (p<0.05) and 200 mg/kg 

groups (p<0.01) for the three sampling times (weeks 13, 26, and 52). The albumin level for the females was 

not affected. Total bilirubin was reduced at weeks 13 and 26 for the 200 mg/kg males (p<0.05) and all three 

sampling times for the 200 mg/kg females (13 and 52 weeks, p<0.05, 26 weeks, p<0.01). Triglycerides were 

increased for the 200 mg/kg males at 13 and 52 weeks (p<0.01) and for the 200 mg/kg females at 13 (p<0.05) 

and 26 weeks (p<0.01). Mean serum alkaline phosphatase activities were increased for the 200 mg/kg males at 

all sampling times (p<0.01) and for the 50 mg/kg males at 26 (p<0.05) and 52 weeks (p<0.01). Serum alkaline 

phosphatase activities for the 200 mg/kg females were increased at week 52 (p<0.05). There were no 

treatment-related effects upon the urinalysis parameters.

At the necropsy, the mean absolute liver and testes weights were increased for the 50 (p<0.05) and 200 mg/kg 

males (p<0.01). The mean relative liver weight was increased for the 200 mg/kg male group (p<0.01). The 

mean relative testes weights were increased for the 50 (p<0.05) and 200 mg/kg groups (p<0.01). The mean 

relative liver weight was increased for the 200 mg/kg females (p<0.01). An increased number of animals 

exhibited hepatocellular hypertrophy in the 50 mg/kg females (3/4) and the 200 mg/kg males (3/4) and females 

(4/4), compared to the control group (1/4 males, 0/4 females). Likewise, bone marrow hypocellularity was 

noted for a greater number of animals in the 200 mg/kg group (males: 3 vs. 1 for the controls, females: 4 vs. 2 

for the controls). No potential adverse effect indicated.

The LOAEL is 50 mg/kg/day, based on The NOAEL is 5 mg/kg/day. 

This chronic toxicity study in the dog is Acceptable-Guideline, and satisfies the requirement for a chronic oral 

study (83-1b) in dogs.

Dose and Endpoint for Establishing RfD: NOAEL = 5 mg/kg/day based on increased incidence of clinical 

signs, increased mean liver weight, and hepatocellular hypertrophy. The LOAEL is 50 mg/kg/day.

Uncertainty Factor(s): 100 (includes 10x for intra-species extrapolation and 10x for inter-species variation).

Chronic RfD = 5 mg/kg/day (NOAEL) = 0.05 mg/kg/day

100 (UF)

Comments about Study/Endpoint/Uncertainty Factor(s): This study was selected because it is a chronic study 

and the NOAEL (systemic) of 5 mg/kg/day is lower that the NOAEL of 9.8 mg/kg/day derived in the chronic 

rat study. Also, the toxic effects observed (liver effects and/or body weight decreases) are also seen in the 

chronic rat and the multigeneration study in rats.

Classification of Carcinogenic Potential

The classification of trifloxystrobin was made by an ad hoc subcommittee of the Cancer Assessment Review 

Committee on May 27, 1999, that trifloxystrobin would be classified as a “Not Likely Human Carcinogen”.

HAZARD CHARACTERIZATION
18

The database is incomplete in that an acute neurotoxicity study in the rat is a data gap. Trifloxystrobin is Toxicity 

III-IV for acute toxicity. However, trifloxystrobin is a strong dermal sensitizer. Trifloxystrobin is positive for 

mutagenicity in Chinese Hamster V79 cells, albeit at cytotoxic dose levels. However, trifloxystrobin is negative in 

the remaining 7 mutagenicity studies. Developmental NOAELs and LOAELs for both rats and rabbits occurred at 

either the same dose levels or were above the NOAELs and LOAELs for maternal toxicity. The NOAEL for offspring 

effects in the 2-generation rats reproduction study occurred at a dose level equivalent to the NOAEL for parental 

findings. Based on these data, there was no increased susceptibility demonstrated under FQPA for infants and 

children. Trifloxystrobin was not carcinogenic in mice following dietary administration to Tif: MAGf (SPF) mice for 

18 months. Trifloxystrobin was negative for carcinogenicity in rats in a 2-year study at dose levels up to 1500 ppm 

(62.2 mg/kg/day). . There is a high degree of confidence in the studies upon which the acute and chronic RfDs are 

based. The study from which an acute RfD (females 13+) was derived was a developmental toxicity study in rabbits. 

It is considered to be appropriate for an acute risk assessment by the HIARC because the skeletal anomalies seen at 

the LOAEL are presumed to occur after a single dose. An appropriate endpoint could not be determined for the risk 

assessment for the general population acute RfD. The study selected for the chronic RfD was a chronic toxicity study 

in dogs. This study exposed a common strain of laboratory dog (beagle) to a wide range of doses of trifloxystrobin. 

The dose selected upon which to base the RfD was 5 mg/kg/day (NOAEL).

Rat Excreta Metabolites

In a rat metabolism study (MRID # 44496822), urine, feces, and bile that were recovered from a previous 

CGA-279202 pharmacokinetics study (MRID 44496821) were pooled by group and sex, subjected to methods 

involving solvent extractions, chemical purification (LC and HPLC), quantification (HPLC), and metabolites 

characterization (MS and NMR), and the information was used to elucidate and propose a metabolic pathway. 

Based on the identified structures of more than 30 metabolites, schemes of metabolic pathways were proposed 

(copies attached) with the following major reaction types: 1) Hydrolysis of the methyl ester to the 

corresponding acid (e.g., CGA 321113). 2) O-Demethylation of the methoxyimino group to the 

hydroxyimino derivative(s) (e.g., NOA 405637). 3) Oxidation of the methyl side chain to the primary 

alcohol (e.g., MET 2U) with further partial oxidation to the corresponding carboxylic acid (e.g., MET 13U). 

The metabolism of the methyl side chain to a primary alcohol was more pronounced in female than in male 

rats resulting in various sex specific major urinary metabolites. There were other minor reaction types 

including chain shortening by oxidative decarboxylation of the glyoxyl moiety to a benzoic acid amide (MET 

13U), hydrolysis of the hydroxyimino group to a ketone (MET 5U) followed by oxidative decarboxylation to 

a benzoic acid derivative (MET 4U), oxidation of the hydroxyimino group to a nitro group (MET 3U), and 

hydroxylations of the phenyl rings. In addition, nearly 10% of the administered dose was accounted for by 

cleavage between the glyoxyl-phenyl and the trifluoromethyl-phenyl moieties with further metabolism by one 

or more of the above described reactions to yield several one phenyl ring metabolites including ortho-phthalic 

acid and meta-trifluoromethyl benzoic acid, among others. 

SUMMARY OF TOXICOLOGY ENDPOINT SELECTION

The doses and toxicological endpoints selected for various exposure scenarios are summarized below. 

EXPOSURE

SCENARIO

DOSE

(mg/kg/day)

ENDPOINT STUDY

Acute Dietary

Females 13+

NOAEL= 250

UF = 100

Increased fetal skeletal anomalies Developmental 

Toxicity-Rabbit

Acute RfD = 2.5 mg/kg 

Chronic Dietary NOAEL =5

UF = 100

Increased incidence of clinical signs, increased mean 

liver weight, and hepatocellular hypertrophy

Chronic Toxicity - Dog
19

Chronic RfD = 0.05 mg/kg/day 

Short-Term

(Dermal)

Dermal

NOAEL= 100

Increases in liver and kidney weights 28-Day Dermal Toxicity 

Study in Rats

Intermediate-Term

(Dermal)

Dermal NOAEL = 

100 

Increases in liver and kidney weights 28-Day Dermal Toxicity 

Study in Rats

Long-Term

(Dermal)a

NOAEL = 5 Increased incidence of clinical signs, increased mean 

liver weight, and hepatocellular hypertrophy

Chronic Toxicity - Dog

Inhalation

Short

----------------------

Inter/Long

NOAEL = 250

------------------

NOAEL = 5

Increased fetal skeletal anomalies

------------------------------------------------------

Increased incidence of clinical signs, increased mean 

liver weight, and hepatocellular hypertrophy

Developmental 

Toxicity-Rabbit

------------------------

Chronic Toxicity - Dog

a= Since an oral NOAEL was selected, a dermal absorption rate of 33 % should be used for route-to-route extrapolation
2020

RESIDUES IN WATER SECTION

See separate submisison.

QUESTIONS TO THE COMMITTEE

H. Does the committee concur with the petitioner’s recommendation 

that trifloxystrobin be the only residue regulated in plant and 

animal commodities?

I. What residue does the committtee recommend for risk assessment 

purposes?

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