Document ID: EPA-HQ-OPP-2006-0239-0034
Agency: epa
Document Type: Supporting & Related Material
Title: 
Posted Date: 2009-04-22T04:00Z

Appendix B – Ecological Effects Data

This appendix contains toxicity data from three sources:  Ecotoxicology
guideline studies in Agency files, data located by ECOTOX in open
literature, and data as presented in the registration review ERA. 
Reviews for open literature data included in the table, or reviewed and
not found valid for quantitative or qualitative use, are also included.

Table   SEQ Table \* ARABIC  1   Ecological Guideline StudiesM for
Fomesafen

Guideline #	Requirement Description	Test Product	Data Valid for Risk
Assessment?	MRID	Study ClassificationC

71-1	Avian Acute Oral	Technical 	Yes	163168E	Core

71-2	Avian Subacute Dietary	Technical 	Yes

Yes	103022

163384E	Core

Core

71-4	Avian Reproduction	Technical

Technical	Yes	135639E

144862	Core

Core

72-1	FW Fish Acute	Formulation	Yes	103023E	CoreC

72-2	FW Aquatic Invertebrate Acute	Formulation	Yes	163169E	CoreC

72-3 (a)	SW Fish Acute	Formulation	Yes	135651E	CoreC

72-4	SW Aquatic Invertebrate	Formulation	Yes	135647E	CoreC

72-4	FW Fish Early Life Stage	No study located

72-4	SW Fish Early Life Stage	Formulation	Yes	135644E	CoreC

72-4(b)	FW Aquatic Invertebrate Reproduction	Formulation	Yes	135642E
CoreC

123-1(a)	Seedling EmergenceA	Formulation

Reflex 2LC	Yes	46673801 E	Acceptable

123-1 (b)	Vegetative VigorA	Formulation

Reflex 2LC	Yes	46673802E	Acceptable

123-2	Aquatic PlantA	Technical	Yes

Yes

Yes

Yes

Yes	46673803E

46673804E

46673805

46673806E

46673807	Acceptable

Supplemental

Acceptable

Acceptable

Acceptable

141-1	Honey Acute Contact	Formulation

Flex 2 LC	YesR	135651	CoreC

	Honey Bee Acute Oral

Non	Earthworm	Technical	Yes	135652	Core

Non	Soil Invertebrates	Technical	Yes	135656	Supplemental

M Not including mammalian studies, which are reviewed by HED

C Studies submitted and classified in 1980’s, when tests on formulated
product were permitted

A Not reported in OPPIN

E Used as endpoint in assessment  

R Currently, there is no standard method for using this type of data in
the risk assessment.Table   SEQ Table \* ARABIC  2   Summary of
Registrant Submitted Acute AquaticToxicity Studies

Species	LC50

(mg/L)	Slope	95% C.I.

(mg/L)	NOAEC

(mg/L)	MRID	Toxicity

Category	Study Classification	Notes

Freshwater

Green Alga

(Selenastrum capricornutum)	0.092	2.4	0.078-0.110	0.0095	46673804	NA
Supplemental

(72-hr study)	Technical

Freshwater diatom

(Navicula pelliculosa)	3.5	2.4	2.3-5.4	ND

EC05 =0.74	46673805	NA	Acceptable	Technical

Blue-green alga

(Anabaena flos-aquae)	71	5.6	60-84	27.3	46673807	NA	Acceptable	Technical

Duckweed

(Lemna gibba)	0.210	1.9	0.150-0.290	0.064	46673803	NA	Acceptable
Technical

Water flea

(Daphnia magna)	376	5.6	323-437	ND	1631169	Practically nontoxic	Core
Formulation1,2

Rainbow trout

(Onchorhyncus mykiss)	126	14.6	117-135	ND	103023	Practically nontoxic
Core	Formulation1,2

Bluegill sunfish

(Lepomis macrochirus)	1507	ND	ND	ND	163169	Practically nontoxic
Supplemental	Formulation1,3

Saltwater

Saltwater diatom

(Skeletonema costata)	0.490	1.6	0.270-0.900	0.360	46673806	NA	Acceptable
Technical

Mysid shrimp

(Mysidopsis bahia)	25	ND	19-38	ND	135647	Slightly toxic	Core	Technical

Sheepshead minnow

(Cyrpinodon variegatus)	>163	ND	ND	NR	135644	Practically nontoxic	Core
Formulation1,2

1 Data are from studies originally reviewed and classified in 1984, some
of which used formulated product. 2  For risk assessment purposes, test
concentration were adjusted for percent a.i., if necessary, and
endpoints were re-calculated using TOXANAL software, prior to 2006
Section 3 Ecological Risk Assessment.  2  For risk assessment purposes,
test concentration were adjusted for percent a.i..  ND-Not determined 
NA-Not applicable, no toxicity category scale for aquatic plants.

Table   SEQ Table \* ARABIC  3   Summary of Registrant Submitted
Chronic AquaticToxicity Studies

Species	NOAEC

(mg/L)	LOAEC

(mg/L)	Endpoints Affected	MRID	Study Classification	Notes

Freshwater

Water flea

(Daphnia magna)	50	100	Reduced growth, total # of offspring	MRID 1354642
Core	Formulation1,2

Saltwater

Mysid shrimp

(Mysidopsis bahia)	0.7	1.7	Parental mortality	MRID 1354648	Core
Technical

Sheepshead minnow

(Cyrpinodon variegatus)	12.2	20.1	Reduced larval survival	MRID 1354644
Core	Formulation1,2

1 Data are from studies originally reviewed and classified in 1984, some
of which used formulated product. 2  For risk assessment purposes, test
concentration were adjusted for percent a.i., if necessary, and
endpoints were re-calculated using TOXANAL software, prior to 2006
Section 3 Ecological Risk Assessment.  

Table   SEQ Table \* ARABIC  4   Summary of Registrant Submitted Acute
Terrestrial Toxicity Studies

Species	LC50/LD50

(mg/kg)	Slope	95% C.I.	MRID	Toxicity

Category	Study Classification1	Notes

Acute oral dose studies on technical (sodium salt of fomesafen)

Mallard duck

(Anas playrhynchos)	>5,000	ND	ND	163168	Practically non-toxic	Core	none

Guinea pig	(F) 607	ND	ND	164901	Slightly toxic	Minimum	none

Mouse	(F) 745

(M) 766	1.6(0.6-2.6)

1.6(0.6-2.5)	512-1286

525-1341

Rat	(F) 1499

(M) 1858	5.4(2.0-8.9)

2.4(0.8-4.1)	1302-1749

1420-2546

Avian sub-acute dietary on technical (sodium salt of fomesafen)

Mallard duck

(Anas playrhynchos)	>20,000	ND	ND	163384	Practically non-toxic	Core	none

Bobwhite quail (Colinus virginianus))	>20,000	ND	ND	103022	Practically
non-toxic	Core	none

Honey bee acute 

Honey bee

(Apis mellifera)	>50 mg/bee	ND	ND	135651	Practically non-toxic	Core	Oral
dose

	>100 mg/bee	ND	ND

Contact

1Studies originally reviewed and classified in 1980s.

ND Not determined  NR Not reported

NA Not applicable, non-definitive endpoint



Table   SEQ Table \* ARABIC  5   Summary of Registrant Submitted Acute
Mammal Toxicity Studies on Formulations

Species	LC50/LD50

(mg/kg)	Slope	95% C.I.	MRID	Toxicity

Category	Study Classification	Notes

Mammal acute oral dose on formulations

Rat 	(F) 3863

(M) >5000	NR	NR	44030101	Practically non-toxic	Acceptable	22.1%
fomesafen/

Trade: Flexstar HL

Rat	5,000	ND	2,865-8390	46952103	Practically non-toxic	Acceptable	10.1%
fomesafen/

46.5% 

s-metolachlor

Trade: Prefix

Rat	>1210 and<1780	NR	NR	40031302	Slightly toxic	Core	7.2% fomesafen/

29% Bentazon 

Trade: 

Faster

Rat	(F) 1210

(M) 1470	NR	NR

1000-2160	158241

also

40340502	Slightly toxic	Core	12.9% fomesafen/

29% Bentazon 

Trade: 

BASF 530 04

ND Not determined  NR Not reported

Table   SEQ Table \* ARABIC  6   Summary of Registrant Submitted Chronic
Terrestrial Toxicity Studies

Species	NOAEC

(mg/kg)	LOAEC

(mg/kg)	Endpoints Affected	MRID	Study Classification1	Notes

Mallard duck

(Anas playrhynchos)	46	ND	No endpoints affected	135639	Core	Technical

Rat	1,000	ND	No reproductive endpoints affected.

Parents and offspring showed modifications in liver histopathology at
1000 ppm	144862	Acceptable	Technical

1Studies originally reviewed and classified in 1980s.

ND Not determined  



Table   SEQ Table \* ARABIC  7   Summary of Registrant Submitted Acute
Toxicity Studies for Terrestrial Plants

Test	Species	Most Sensitive Endpoint	EC25

(lb ai/A)	95% C.I.

(lb ai/A)	EC05

(lb ai/A)	NOAEC

(lb ai/A)	MRID	Classification

Seedling emergence	Onion	Biomass	0.084	0.024-0.29	0.024	0.030	46673801
Acceptable

	Barnyard grass	Biomass	0.28	Not reported	0.076	0.12

	Corn	None	>0.48	N/A	>0.48	0.48

	Oat	None	>0.48	N/A	>0.48	0.48

	Tomato	% survival	0.0046	0.00048-0.044	0.00021	<0.0038

	Oilseed Rape	% survival	0.073	0.012-0.462	0.028	0.060

	Radish	% survival	0.10	0.004-3.14	0.023	0.060

	Lettuce	Biomass	0.22	0.123-0.444	0.15	0.030

	Sugarbeet	% survival	0.35	Not reported 	0.11	0.12

	Soybean	None	>0.48	N/A	>0.48	0.48

Vegetative vigor	Barnyard Grass	Biomass	0.29	0.14-0.59	0.097	0.24
46673802	Acceptable

	Corn	None	>0.47	N/A	>0.47	0.47

	Oat	Biomass	1.9	0.12-0.30	0.10	0.17

	Onion	Biomass	3.0	0.0007-14,000	0.045	0.47

	Lettuce	Biomass	0.0020	0.0011-0.0035	0.00042	<0.0019

	Radish	Plant height	0.0031	0.003-0.018	0.00037	0.00090

	Oilseed Rape	Biomass	0.0080	0.0050-0.013	0.0036	0.0019

	Sugarbeet	Biomass	0.15	0.12-0.19	0.064	0.0073

	Tomato	Biomass	0.24	0.16-0.36	0.045	0.060

	Soybean	None	>0.47	N/A	>0.47	0.47

Table   SEQ Table \* ARABIC  8   Summary of Valid Aquatic ECOTOX
Toxicity Studies on Fomesafen

Species	Measurement	Type of Effect	Endpoint	Concentration 	ECOTOX Ref#

Aquatic

Algae

(Kirchneriella lunaris)

(Scenedesmus obliquus)

(Cryptomonas sp.)	Reduced growth, extent not specified	40 g/L

72 hours	87334

Caquet 2005

Aquatic mesocosm

Plankton

Caldoceran

Common backswimmer

(Notonecta glauca)	Modifications of trophic structure, as evaluated by
stable isotope analysis	Community	“perturbation”

~LOAEC	30-60 g/L

15 weeks	87334

Caquet 2006

Aquatic mesocosm

Planktonic community	Decreases in dissolved oxygen	Community	NOAEC	~60
g/L

48 hours	87334

Pershberger 1997

Freshwater snail

(Lymnaea stagnalis)	Reactive oxygen species (ROS)

Phagocytic activity

Lysosomal fragility	Biochemical

Histological	LOAEC	10 g/L

24 hours	E95100

Russo 2007a

	Mortality	Mortality	NOAEC	270 g/L

21 days

Growth	Growth	NOAEC

	Increase in hemocyte density	Biochemical

Histological	LOAEC	10 g/L

24 hours	E95590

Russo 2007b

Table   SEQ Table \* ARABIC  9   Summary of Valid Terrestrial ECOTOX
Toxicity Studies on Fomesafen

Species	Measurement	Type of Effect	Endpoint	Concentration 	ECOTOX Ref#

Fungus

(Bipolaris euphobiae)	Reduction in mycelium growth 	Growth	NOAEC	1L/ha

Product

10 days	E95538

Nemoto

	Speed of conidial germination

	1L/ha

Product

12 hours

	Mice

(Mus domesticus)

Various lab strains	Body weight gain	Growth	LOAEC	2,500 mg/kg diet
E95026

Krijit 1993

	Increased relative liver weight	Growth	LOAEC	100 mg/kg diet	E95026

Krijit 1993

	Elevated liver and fecal porphyrins	Biochemical	LOAEC	100 mg/kg diet
E95026

Krijit 1993

Open Literature Review Summaries for Fomesafen

Studies Classified Valid for Qualitative Use

Open Literature Review Summary

Chemical Name:  Fomesafen 

PC Code:  123802

ECOTOX Record Number and Citation:

This review evaluates information from two related aquatic mesoscosm
experiments evaluating population- and community-level effect of
fomesafen and a fomesafen-surfactant mixture.  The mesocosoms used in
this experiment appear to be the same mesocosms descirbed in Jumel et
al. 2002 (ECOTOX 64953), although authors focus on different organisms
in the mesocosoms.  Other related papers are Russo et. al, 2007 (ECOTOX
95100), and Russo & Medec 2007 (ECOTOX 95590).  All papers have been
reviewed.

87334  Caquet, T, Deydier-Stephan, L, Lacroix, G, Le Rouzic, B, and
Lescher-Moutoué, F.  (2005)  Effects of fomesafen, alone and in
combination with an adjuvant, on plankton communities in freshwater
outdoor pond mesocosm.  Environmental Toxicology and Chemistry
24:1116-1124. 

95935  Caquet, T.  2006.  Use of carbon and nitrogen stable isotope
ratios to assess the effects of environmental contaminants on aquatic
food webs.  Environmental Pollution 141:54-59.  

Purpose of Review (DP Barcode or Litigation):  Registration Review

Date of Review:  4/24/08

Summary of Study Findings:

Caquet, et. al 2005.  Authors evaluated the effects of fomesafen and a
mixture of fomesafen and Agral-90 (a polyethoxylated nonylphenol
surfactant) on planktonic communities in outdoor mesocosms.  Mesoscosms
were treated with fomesafen (nominal concentration 40 g/L) or
fomesafen/Agral-90 mixture (nominal concentrations 40 g/L and 90
g/L respectively) every three weeks, with a total of five treatments.
 Water concentrations of fomesafen measured at the end of the treatment
period were 62.5 g/L in the fomesafen only treatments, and 19.4
g/L in the mixture treatments.  Phytoplankton and zooplankton were
collected, enumerated and identified at regular intervals throughout the
experiment, which included a pre-treatment period, a treatment period,
and a post-treatment period..  For algae, abundance data were used to
calculate  Margalef’s diversity index and Simpson’s dominance index.
 Concurrent laboratory toxicity tests were conducted on six algal
species: Chlorella vulgaris, Kirchneriella lunaris, Scenedesmus
obliquus,  and Staurastrum polymorphum (Chlorophyceae), Cryptomonas sp.
(Cryptophyceae) and Cyclotella pseudoselligera (Bacillariophyceae).

g/L fomesafen (nominal) had a negative effect on the growth of 3 of
the 6 species tested (K. lunaris, S. obliquus, and Cryptomonas sp.). 
Authors do not specify the extent of the effect, merely noting it was
statistically significant.

Authors conclude that Chlororphyceae were slightly inhibited in the
fomsafen only treatments, and this may be due to the fact that a greater
number of species in this class are more sensitive to fomesafen. 
However, they were apparently replaced in the ecosystem by another less
sensitive species.  They also note the between pond heterogeneity, which
was high, may have decreased the statistical sensitivity of the study. 
Fomesafen alone had no significant effect on zooplankton.

Caquet (2006).  In this study, author used stable isotopes of carbon and
nitrogen to evaluate potential trophic effects associated with
application of fomesafen to outdoor mesocosms.  The organism evaluated
was 4th and 5th instars of the common backswimmer, Notonecta glauca L.,
which is an aquatic predator that feeds on zooplankton.  Upper level
predators can integrate changes occurring in lower trophic levels, and
the stable isotope ratios can be used to track these changes.  Authors
measured stable isotopes of carbon and nitrogen in the backswimmer, but
not lower trophic levels such as their cladoceran and copepod prey, or
the primary producers.  The level of enrichment in 13C can be used to
trace the origin of carbon in the food chain, and enrichments in 15N
indicate trophic level.

Experimental design of the mesoscosms is as described above.  N. glauca
nymphs were collected at two different times: once four weeks after
initiating treatments (following two applications of fomesafen) and once
seven weeks after initiating treatments (following three applications). 
All nymphs collected the first time were 4th instar, and those collected
the second time were 5th instar.  Instars were processed separately for
stable isotopes.  Treatments did not have any effect on size or
abundance of the organisms.

Based on information from the plankton study, summarized above, there
were some changes in the composition of the plankton communities in the
treated ponds, although no change in abundance.  Data on zooplankton
communities presented in this paper (Table 2, pg 57) show an increase in
rotifers and a decrease in calanoid copepods in both the fomesafen only
and the mixture ponds, with the shift being more apparent in the mixture
ponds.  According to author, copepods are a prey of the nymphs, along
with cladocerans (no significant differences were found in cladoceran
populations).

Comparisons of the stable isotope ratios (Fig 1, pg 56) shows two
distinct groupings, comprised of the 4th instar and the 5th instar. 
There was a significant effect of larval instar on 15N, indicating a
shift in trophic level.  Post-hoc tests comparing the controls and the
treatments showed fomesafen had an effect on 13C (indicating a potential
shift in primary production) and the mixture had effects on both 13C and
15N ( indicating potential shifts in both primary production and trophic
level).  [note:  author does not specify if post hoc test described were
for both 4th and 5th instar, or just for 5th instar where differences
were more discernable or if there was any interaction effect between
treatment and instar.]

Information Applicable to Fomesafen Ecological Risk Assessment 

Based on the Work of Caquet et. al.

g/L(“maximum mean”) in the fomesafen only ponds and as 19.4 ±7.6
g/L (“maximum mean”) in the fomesafen-Agral 90 ponds.  Although
concentrations are reported only in Caquet et al., 2005, reviwer has
assumed they are valid for both papers.  Authors make no mention of
measuring concentrations in other compartments in the mesocosm or
performing any mass balance calculations to determine fate of fomesafen
in the mixture mesocosms, although they speculate (Caquet et al., 2005)
it may have accumulated at the interfaces.

g/L) and 60 -day (6-13 g/L) EECs predicted by PRZM-EXAMS when
fomesafen is applied in accordance with current labels (0.375-0.5 lb
ai/A) (Fomesafen ERA, DP 302766, 314014, 314112) is lower than the
concentrations evaluated in the mesocosm tests.  The most sensitive
freshwater plant based on guideline tests was a green alga (Selenastrum
capricornutum), with an LC50 of 92 g/L and an NOAEC of 9.5 g/L
(MRID 46673806).

Description of Use in Document (QUAL, QUAN, INV):  QUAL

Rationale for Use:  Provides insight into potential effects on
phytoplankton and zooplankton communities exposed to fomesafen, although
concentrations used in the study are above modeled EECs.

Limitations of Study:  Measured concentrations of fomesafen are only
explicitly stated for the end of the treatment period.  None of the
papers associated with this mesocosm study have presented the water
concentrations in a form that allows determination of a mean-measured or
time-averaged concentration.   

Primary Reviewer:

Paige D. Doelling, Ph.D., Fisheries Biologist, ERB1

Secondary Reviewer 

Christine Hartless, Ph. D., Wildlife Biologist, ERB1

Open Literature Review Summary

Chemical Name:  Fomesafen 

PC Code:  123802

ECOTOX Record Number and Citation:

This review includes a summary of data from five papers written by the
same first author (Jan Krijit) describing and evaluating effects of
fomesafen on mice liver hepatocytes.  Papers were published in various
journals over a period extending from 1993-2003.  Evaluations of the
papers are presented in chronological order.

E95026  Krijit, J, van Hosteijn, I, Vokurka, M, and Blaauboer, B. (1993)
 Effect of diphnyl ether herbicides and oxadiazon on porphyrin
biosynthesis in mouse liver, rat primary hepatocyte culture and HepG2
cells.  Archives of Toxicology 67:255-261.

E95399  Krijit, J, Vokurka M, Sanitrák, J, Janousek, V, van Hosteijn,
I, and Blaauboer, B. (1994a)  Effect of the protophyrinogen
oxidase-inhibiting herbicide fomesafen on liver uroporphyrin and
heptacarboxylic prophyrin in two mouse strains.  Food Chemical
Toxicology 32(7): 641-649.

E95589  Krijit, J, Vokurka M, Sanitrák, J, and Janousek, V. (1994b) 
Effect of Protophyrinogen Oxidase Inhibitors on Mammalian Porphyrin
Metabolism.  American Chemical Society Symposium Series 559:247-254.

E95400  Krijit, J, Stránska, P, Sanitrák, J, Chlumská, and Fakan, F.
(1999)  Liver preneoplastic changes in mice treated with the herbicide
fomesafen. Human and Experimental Toxicology 18:338-344.

E95588  Krijit, J, Pšenák, O, Vokurka, M, Chlumská, A, and Fakan, F.
(2003)  Experimental hepatic uroprophhyria induced by the diphnyl-ether
herbicide fomesafen in male DBA/2 mice. Toxicology and Applied
Pharmacology 189:28-38.

Purpose of Review (DP Barcode or Litigation):  Registration Review

Date of Review:  4/14/08

Summary of Study Findings:

Fomesafen is a peroxidizing enzyme, inhibiting the enzyme
protoporphyrin-III oxidase, which catalyzes the formation of
protoporphyrin IX.  Protoporphyrin IX is a precursor molecule common to
both plants and animals.  In plants, it is a step in chlorophyll
biosynthesis, and in animals, it is a step in hemoglobin biosynthesis. 
Some chemicals of this class (diphenyl-ether herbicides, which also
includes aciflourfen, oxyflourfen, lactofen, and nitrofen) have been
associated with peroxisome proliferation, which can induce
hepatocellualr carcinomas in rodents.  (Smith and Elcombe 1989, Ashby et
al., as cited in Krijit et al., 1999).

Krijit, et. al, 1993:  Authors investigated the effects of fomesafen (
and other chemicals) in vivo (mouse livers), and in vitro (rat primary
hepatocyte culture and Hep G2 cells).  Based on a range-finding test,
authors tested “0.25% (w/w) concentration of the herbicides in the
diet” for a 10-day period.  Conversion of 0.25% to ppm in diet yields
2,500 ppm.

For the in vivo tests each herbicide and the control were tested with 4
male C57B1/6J mice per treatment.  Authors report statistically
significant effects (p<0.05, student t-test) in the fomesafen-treated
group for the endpoints of body weight (decrease) , relative liver
weight (increase), liver  porphyrins (increase), fecal porphyrins
(increase), liver microsomal ethoxyresorufin and pentoxyresorufin
O-dealkylation (EROD and PROD) activity (increase), and peroxisomal
β-oxidation activity( increase).  Effects on cytochrome P-450 level
were not significant.  Raw data were not available to confirm
statistical analysis.  

Based on this study, there appears to be in decrease in body weight and
a modification in liver activity for mouse fed a diet containing 2,500
ppm fomesafen for 10 days.

(Note:  The ECOTOX database lists an LOAEL of 100 ppm on mouse
growth-weight for this paper.  This appears to be based on a change in
relative liver weight (Table 1, pg 257) and has not been used as an
endpoint.)

Krijit et. al.,1994a.  Authors investigated effects of long-term (5
months) dietary exposure to fomesafen on the enzymes protoporphyrin-II
oxidase (enzyme 7 in heme biosynthesis) and uroporphyrinogen-III
decarboxylase (enzyme 5 in heme biosynthesis).  They also evaluated the
effects of administering iron separately and in conjunction with
fomesafen on the biochemical endpoints.  

Male ICR mice (n=4) were fed 0.25% w/w (2,500 ppm) fomesafen in diet for
5 months.  Treatment groups were a control, a group treated with just
iron (subcutaneaous injection, 1 time, prior to feeding trial), a group
treated with fomesafen (in diet), and a goup pre-treated with iron and
fed fomesafen.  Authors note iron pretreatment can induce experimental
prophyria.  Fomesafen decreased body weight, increased relative liver
weight, and bile prophyrin content by a statistically significant amount
(p<0.05, test not specified, Table 1).Some of the mice exhibited
dark-colored livers.  Serum alanine aminotranferase (ALT), and liver
uroporphyrinogen decarboxylase (UROD) activity and liver porphyrins were
also affected.  Elevated ALT and UROD activities indicated
hepatotoxicity.

A similiar experiment was conducted with male C57Bl/6J mice, although in
this case they were fed 0.20% w/w (2,000 ppm) in diet, and oxyfluorfen
and an oxyfluorfen-iron combination were tested in addition to
fomesafen.  Results for body weight, relative liver weight, liver
porphyrin and bile porphyrin were similar.  In this group of mice,
authors also measured acitivity of liver cytochrome P-450 and associated
enzymes, as well as peroxisomal β-oxidation activity.  Peroxisomal
β-oxidation activity was affected in all treatment groups, and enzyme
activity levels were elevated in most treatment groups, including those
treated with fomesafen..

Authors concluded that it “appears fomesafen . . . can block two
enzymes in the porphyrin biosynthetic pathway-protoporphyrinogen oxidase
and uroporphyrinogen decarboxylase.  Whereas the inhibition of
proyoporphyrinogen oxidase requires relatively low doses of the
herbicide (Krijit et al., 1993) [100 to 2,500 ppm in diet for 10 days]
the inhibition of URO-D can be observed only after feeding of high doses
of fomesafen for prolonged periods [2,000 to 2,500 ppm for 3 to 5
months, depending on strain of mouse].” (pg 648)

Krijit et. al, 1994b.  Authors investigated the effects of fomesafen
administered in the diet on biochemical endpoints (primarily prophyrin
concentrations) in the liver, kidney, and bile of male ICR mice.  In all
cases, treatment groups appeared to consist of 3 or 4 mice.  Results are
presented in what appears to be (but is not specifically noted as) mean
± SD.  Table III (pg 249) presents statistical comparisons (p<0.05,
test not specified), but the others do not.  A diet including 2,500 ppm
of fomesafen fed to male ICR strain mice increases liver, bile, and
fecal porphyrin content compared to controls (Table I, pg 248).  Smaller
increases in liver and bile porphyrin content were noted in the same
strain of mice fed 100 ppm fomesafen for 10 days.  In an experiment
where the male ICR mice were fed 5000 ppm fomesafen in the diet for 10
days, followed by a 5-day period of control diet, porphyrin
concentrations in the liver, bile and feces returned to levels not
different than the controls (authors do not specify if these results
were compared statistically.  Fomesafen fed to male C57Bl/6J for 3
months at a concentration of 2,500 ppm did not cause a change in total
microsomal cytochrome P-450 or EROD, but did cause an increase in PROD
as compared to controls.  Raw data were not available to confirm
statistical analysis.  

Authors conclude “rather high doses are needed to produce relatively
slight changes in porphyrin concentrations in vivo.  Also, the
accumulation of liver porphyrins induced by fomesafen treatment is
clearly reversible.”  (pg 253)

Krijit, et al. 1999.  Authors investigated effects of long-term (50
weeks to 14 months), high-dose (2,300-3,000 ppm in diet) consumption of
fomesafen on biochemical and histological endpoints in two strains of
mice (C57Bl/6J mice and ICR mice).  Biochemical endpoints measured were
associated with porphyrin biosynthesis and histological endpoints
evaluated were precursors to hepatocellular carcinomas.  Fomesafen
caused preneoplastic changes in both strains of mice, as well as
modifications in enzyme activity.  Only the ICR mice developed
uroporphyria.  Iron injections increased response in both strains of
mice.  In a portion of the study which included a recovery period after
long-term treatment with a high dose(3,200 ppm for 9 months, followed by
a 3 month recovery), authors noted that liver histology returned to
normal, and liver porphyrin content decreased, although still remaining
above control levels.

Based on information contained in this paper, authors conclude the liver
porphyrin biosynthesis pathway is the most sensitive biomarker for
fomesafen exposure.  They also note the toxicological significance of
liver porphyrin accumulation is uncertain.  The extent of the response
differs depending on the strain of mouse.

Krijit et al., 2003  Authors evaluated the effects of dietary fomesafen
on two strains of male mice (C57Bl/6N and DBA/2N).  These strains of
mice were selected because “PAH-induced uroporphyria readily develops
in animals of the C57Bl6 strain, whereas DBA/2N mice are almost
completely resistant” (pg 35).  Fomesafen content in the diet was
0.25% w/w (equivalent to 2,500 ppm, author converted to a daily dose of
350 mg/kg for DBA/2N group) for 18 weeks.

In the DBA/2N mice, administration of induced hepatic uroporphyria,
indicating there was some inhibition of the uroporphyrinogen III
decarboxylase enzyme (5th enzyme in the heme biosysthesis pathway) in
addition to inhibition of protophyrinogen oxidase (7th enzyme in the
heme biosysthesis pathway).  The C57Bl/6N mice showed a slight (although
statistically significant) elevation in liver porphyrins.  In this
study, authors also measured cytochrome P-450 activities, performed
histological examinations, and looked at messenger-RNA in an attempt to
elucidate biochemical processing differences between the two strains.

Based on comparisions between biochemical endpoints modified by exposure
to fomesafen, and the known uroporphyria inducers TCDD and PCBs, authors
concluded that there may be a “yet unknown mechanism of uroporphyria
induction (p28).”  Extent and nature of the response to fomesafen
differs depending on the strain of mouse.

Information Applicable to Fomesafen Ecological Risk Assessment 

Based on the Work of Krijit et. al.

The work of Krijit et. al., demonstrates that fomesafen ingested by mice
at concentrations ranging from 100-2,500 ppm diet for 10 days causes
elevated liver and fecal porphyrins, primarily via inhibition of the 7th
enzyme of the heme biosynthesis protoprophyrin III oxidase (Krijit et.
al,1993).  Body weight gain is statistically significantly affected when
fomesafen is administered at concentrations of 2,500 ppm in diet for 10
days (Krijit et. al,1993).  The treatment groups in all investigations
are small (n=3 or 4).  Mice used in the experiments were generally in
the size range of 30-35 grams.  In some cases, fomesfen may also inhibit
uroporphyrinogen III decarboxylase, but specific biochemical responses
may vary dependent on the strain of mouse (Krijit et. al,1994b, Krijit
et. al, 2003).  .  Consumption of high doses (2,300-3,000 ppm) for
extended periods of time (50 weeks to 14 months) caused development of
precancerous cells in the liver (Krijit et. al,1999).  Even with
extended exposure , mice appeared to recover following a period of clean
diet (3months) with liver porphyrins decreasing, and liver histology not
noticeably different from controls (Krijit et. al,1999).  In both
studies that included a recovery period, liver porphyrin concentrations
decreased rapidly following withdrawal from fomesafen, but did not
return to the same levels as controls during the observation period
(Krijit et. al,1999).  

Description of Use in Document (QUAL, QUAN, INV):  QUAL

Rationale for Use:  

Provides insight as to how chronic effects on mammals (and by extension,
other animals) are mediated.  Subacute (10 day) growth endpoint derived
from Krijit et. al 1993 (LOAEC decreased weight at 2,500 ppm diet) is
higher than current guideline-derived NOAEC of 250 ppm and LOAEC of 1000
ppm and higher than acute dose LD50 (607 mg/kg).

(Note:  The ECOTOX database lists an LOAEL of 100 ppm on mouse
growth-weight for this paper.  This appears to be based on a change in
relative liver weight (Table 1, pg 257) and has not been used as an
endpoint.)

Limitations of Study:

It is uncertain how modifications in liver porphyrin levels might affect
wild mammals at the individual or population level.

Treatment groups were small (n=3or 4).

Strains of laboratory mice are inbred, and lack genetic variability
found in wild populations that may affect response to the toxicant. 
Different strains of mice were shown to react to a different extent, but
all exhibited some type of negative effects.

Doses used in laboratory testing were generally an order of magnitude
above what animals are anticipated to encounter in a treated field based
on TREX calculations.  Kenaga upper bound EEC for short grass at 0.5 lb
ai/A (highest application rate as of May 2008) is 118 mg/kg.  Kenaga
upper bound dose for short grass-consuming 35 g mammal is 78 mg/kg bwt.

Primary Reviewer:

Paige D. Doelling, Ph.D., Fisheries Biologist, ERB1

Secondary Reviewer 

Christine Hartless, Ph. D., Wildlife Biologist, ERB1

Open Literature Review Summary

Chemical Name:  Fomesafen 

PC Code:  123802

ECOTOX Record Number and Citation:

95538 Nemoto,MCM, Nahas, E, Pitelli, RA, and Coelho, L.  (2002) 
Germination and mycelial growth of Bipolaris euphorbiae Muchovej &
Carvalho as influenced by herbicides and surfactants.  Brazilian Journal
of Microbiology 33:352-357.

Purpose of Review (DP Barcode or Litigation):  Registration Review

Date of Review:  4/25/08

Summary of Study Findings:

Authors investigated the effects of herbicides and surfactants on the
mycelium growth and conidial germination of Bipolaris euphorbiae, a
fungus which serves as a biological control for milkweed in soybean
fields.  Authors describe test concentrations of 1L/ha for fomesafen,
but do not specify the commercial product used in the test, thus a
determination of dose in terms of active ingredient (ai) is not
possible.  Fomesafen tested at this concentration had no effect on
either mycelium biomass or conidial germination of the test organism. 
Authors conclude tank mixes of Bipolaris euphorbiae and fomesafen are
not contraindicated.

Description of Use in Document (QUAL, QUAN, INV):  QUAL

Rationale for Use:  Provides information of effects on underrepresented
taxa.

Limitations of Study:  Determination of dose in terms of active
ingredient (ai) is not possible.  Test quantity should be presented in
terms of L/ha (end-use product).

Primary Reviewer:

Paige D. Doelling, Ph.D., Fisheries Biologist, ERB1

Secondary Reviewer 

Christine Hartless, Ph.D., Wildlife Biologist, ERB1

Open Literature Review Summary

Chemical Name:  Fomesafen 

PC Code:  123802

ECOTOX Record Number and Citation:

E53095  Perschbacher, PW, Stone, N, Ludwig, GM, Guy, Jr. CB. (1997)
Evaluation of effects of common aerially-applied soybean herbicides and
propanil on the plankton communities of aquaculture ponds.  Aquaculture
157:117-122.

Purpose of Review (DP Barcode or Litigation):  Registration Review

Date of Review:  4/25/08

Summary of Study Findings:

Authors investigated the effects of spray drift deposition on aquatic
mesocosms for herbicides associated with soybean production.  Herbicides
were sprayed across the surface of outdoor 500-L fiberglass pools at
concentrations of full label rate, 1/10 label rate (described as “high
drift”) and 1/100 label rate (described as “low drift”).  Pools
were filled with water drawn from nearby catfish aquaculture ponds, but
authors do not specify total water volume or note whether water was
tested for any other toxicants.  Following treatment, authors evaluated
phytoplankton productivity, morning dissolved oxygen (DO) levels
chlorophyll a, pheophytin a, total ammonia nitrogen, nitrite nitrogen,
and performed a light bottle/dark bottle estimation of phytoplankton
productivity and respiration.  Zooplankton and phytoplankton were
apparently identified and enumerated for concurrent investigations, but
results are not reported in this paper.  Authors do note cyanobacteria
dominated the phytoplankton population.

Fomesafen (as the product Reflex) was applied at the rate of 0.43 kg
ai/ha (equivalent to 0.38 lb ai/A; probably based on the aerial
application rate for soybeans of 0.375 lb ai/A).  Authors note “the
highest rate of application of 0.57 kg/ha was equivalent to 0.06 mg/L
(pg 121)”, but do not specify if this value was calculated or
measured.

Dissolved oxygen was found to be the most sensitive parameter.  For
fomesafen, there was no significant difference in dissolved oxygen
content in the treatments as compared to the control for any
concentration.

Information Applicable to Fomesafen Ecological Risk Assessment 

Based on the Work of Pershbacher et. al.

work, one could interpret that short-term effects on the phytoplankton
abundance and production from concentrations of fomesafen ≤0.06 mg/L
are not anticipated.  This should be caveated with the fact that the
mesocosm community was dominated by cyanobacteria (which appear to be
less sensitive to fomesafen than green algae based on
registrant-submitted studies: MRID 46673806, green alga EC50 92 g/L;
MRIDs 46673807, blue-green alga EC50 710 g/L).  Also, measurement
endpoints reported by authors evaluate overall primary productivity of
the system, rather than community structure, which may be more
responsive.

Description of Use in Document (QUAL, QUAN, INV):  QUAL

Rationale for Use:  Confirmatory data for other non-vascular aquatic
plant information.  Exposure scenario closely mimics current EFED spray
drift evaluation.

Limitations of Study:  Aquatic concentrations were not measured, and can
only be estimated.  Effects should be associated with technical end-use
product Reflex, rather than technical fomesafen.

Primary Reviewer:

Paige D. Doelling, Ph.D., Fisheries Biologist, ERB1

Secondary Reviewer 

Christine Hartless, Ph. D., Wildlife Biologist

Open Literature Review Summary

Chemical Name:  Fomesafen 

PC Code:  123802

ECOTOX Record Number and Citation:

This review evaluates two papers regarding fomesafen effects on the
hemocytes of the freshwater snail Lymnaea stagnalis.

E95100  Russo, J, Lefeuvre-Orfila, l, and Lagadic, L.  (2007)
Hemocyte-specific responses to the peroxidizing herbicide fomesafen in
the pond snail Lymnaea stagnalis (Gastropoda, Pulmonata). Environmental
Pollution 146:420-427.

E 95590  Russo, J, Madec, L.  (2007)  Hemocyte apoptosis as a general
cellular immune response of the snail, Lymnaea stagnalis, to a toxicant.
 Cell Tissue Research 328:431-441.

Purpose of Review (DP Barcode or Litigation):  Registration Review

Date of Review:  4/23/08

Summary of Study Findings:

Fomesafen acts by inhibiting protoporphyrinogen oxidase (Grimm 1999, as
cited in Russo et. al, 2007a) an enzyme in the porphyrin biosynthesis
pathway for chlorophyll (plants) and hemoglobin (animals).  This results
in production of reactive oxygen species (ROS), which can disrupt
cellular membranes in plants (Wakabayashi and Böger 1999, as cited in
Russo et. al, 2007b) and in animals Elcombe et. al, 1996, Krijit et. al,
1999 as cited in Russo et. al, 2007b).  The papers evaluated in this
review are concerned with effects on the circulating hemocytes of
freshwater snails exposed to fomesafen in aqueous solution.

Russo et. al, 2007  Snails were exposed to concentrations of 10, 30, 90,
and 270 g/L fomesafen (nominal)  for 21 days in a laboratory setting.
 Fomesafen was solubilized in acetone, with the concentration of solvent
in exposure media maintained below 1 L/mL.  Authors maintained both
solvent and neutral controls.  Endpoints measured included intracellular
production of reactive oxygen species (ROS), phagocytic activity, and
membrane integrity.  Measurements were made at 24 hours and 504 hours of
exposure.  Results were evaluated with two-way ANOVA for time and
treatment, and one-way ANOVA to compare different exposure regimens for
each exposure duration.

●  Even at the highest concentration (270 g/L), there were no
observable effects on snail mortality or hemocyte cell viability.

	●  Intracellular production of ROS was enhanced in all concentrations
at both time durations except for the 270 g/L-21 day group.  Based on
results from the two-way ANOVA, exposure time was significant
(p=0.0001).  Results were not presented for the interaction term.

	●  Phagocytic activity followed approximately the same pattern as
proportion of ROS-producing hemocytes, with reductions in activity even
at the 10 g/L concentration, and greater inhibition of phagocytic
activity in the hemocytes from snails exposed for 504 hours.  Based on
two-way ANOVA analysis, the concentration of fomesafen was significant
(p=0.049), but the effects of time, and the interaction of time and
concentration were not significant.

	●  Authors evaluated the “oxidative burst,” a cellular immune
defense, in snail hemocytes stimulated by E. coli (low stimulation) and
phorbol 12-myristate 13-acetate (PMA, high stimulation).  In the E. coli
stimulated cells, the oxidative burst was statistically significantly
reduced in all treatment groups when measured at 504 hours, but not when
measured at 24 hours.  In the PMA stimulated cells, the reverse
situation occurred, where the burst was statistically significantly
reduced in all treatment groups when measured at 24 hours, but not when
measured at 504 hours.

●    Lysosomal fragility (as indicated by neutral-red dye retention
time) was statistically significantly decreased in all treatment groups,
and length of exposure also had a significant effect, with great
fragility noted in the 504 hour exposure.  Authors estimate the LOAEC
for lysosomal fragility is 10 g/L for both exposure periods (Williams
test, p=0.05, pg 423).  In their discussion, they note tha “lysosome
stability represents a key component of cell responses to chemical
stress (page 426) but caution “the significance of physiological
changes in the snails needs to be carefully defined (p 426.)”

Russo & Medec 2007

(Reveiwer’s note:  Snails described in this paper may be the same
snails described in Russo et al., 2006, with the second paper focused on
different endpoints.)

Authors investigated the effect of exposure to fomesafen on apoptosis
(programmed cellular death) in freshwater snail circulating hemocytes. 
Exposure to fomesafen can generate reactive oxygen species (ROS) in the
cells, which may initiate the apoptosis sequence as cellular membranes
are compromised.  Exposure periods were 24 hrs, 96 hrs, and 504 hrs (21
days).  The experiment evaluated two events which occur in apoptosis: 
the disruption of the mitochondrial transmembrane potential, and
membrane asymmetry with phophatidylserine (PS) release.  

g/L).  Cell viability was consistently 94%.  Authors state exposure
to fomesafen at 30 g/L and higher concentrations, had a significant
effect on hemocyte density compared to the control (Dunnett’s test,
p=0.01 to 0.0003).  ,Authors note time of exposure did not affect
hemocyte density (p=0.08), and for this endpoint, comparisons appear to
have been averaged over time.  Based on Figure 1 (pg 434, tables with
specific values not included), exposure to acetone increased the
hemocyte density, although authors do not note if the increase is
significantly different from the neutral control.

Oxidative stress (ROS production) was measured by exposing the hemocytes
to a fluorescing dye, and then measuring the response using flow
cytometry.  Cells from the treatment groups were compared to the neutral
control and to a positive control.  Flow cytometry showed “two
functional hemocyte populations,” distinguished by the the wavelength
at which they fluoresced.  Authors state that “according to the four
criteria used for the multivariate test (criteria not specified) the
effect of fomesafen exposure on ROS production was highly
significant.”  Both populations of hemocytes from exposed snails
displayed an increased magnitude of flouresence compared with control
and PMA-induced samples.  At 504 hours there was a marked decrease in
ROS production.

g/L treatment groups, but not in the 90 and 270 g/L groups.  

Membrane asymmetry, which the authors describe as an indicator of
intermediate stage apoptosis, was evaluated by using two flourescing
dyes, one of which binds to a cellular surface marker, and one of which
intercalates DNA.  Hemocytes were placed into groups based on the
proportions of these dyes present in the cells.  The groups were viable
cells, apoptic cells, necrotic cells, and dead cells.  At 96 hours, the
samples from fomesafen-treated snails included a higher percentage of
cells in the apoptic and necrotic stage than the control.  Authors note
“the subsequent increase of necrotic cells did not imply that these
cells passed into the population of dead cells.”  At 504 hours, there
were no significant differences between any of the treatment groups.

Authors conclude fomesafen is “responsible for increasing plasma
membrane permeability and affects cyltoplamic manifestations but does
not provoke an advanced stage of the [cellular] death process.”

Information Applicable to Fomesafen Ecological Risk Assessment 

Based on the Work of Russo et. al.

≤270 g/L for durations ≤504 hours (21 days) these was no
observable abnormal behavior, mortality, of reduction of hemocyte cell
viability in the snails (Russo et. al 2007, Russo and Madec 2007). 
Based on this work, it appears that fomesafen induces an immune response
in freshwater snails at concentrations that may occur in the
environment, but how these responses translate to measurable effects at
an organism level (e.g. reduced growth, tumor activation, increase
susceptibility to disease) are unknown.

Description of Use in Document (QUAL, QUAN, INV):  QUAL

Rationale for Use:  Provides insight into mechanisms whereby fomesafen
affects aquatic wildlife.

Limitations of Study:  Concentrations were nominal, but fomesafen is
relatively persistent in aquatic systems.  Effects at biochemical level
are not readily translated to the growth, survival, and reproduction
endpoints used in the EFED risk assessment process.

Primary Reviewer:

Paige D. Doelling, Ph.D., Fisheries Biologist, ERB1

Secondary Reviewer 

Christine Hartless, Ph.D., Wildlife Biologist, ERB1

Open Literature Review Summaries for Fomesafen

Studies Classified Invalid

Open Literature Review Summary

Chemical Name:  Fomesafen 

PC Code:  123802

ECOTOX Record Number and Citation:

64953  Jumel, A. Coutellec, M-A, Cravedic J-P, and Lagadic, L.  (2002) 
Nonylphenol polyethoxylate adjuvant mitigates the reproductive toxicity
of fomesafen on the freshwater snail Lymnaea stagnalis in outdoor
experimental ponds.  Environmental Toxicology and Chemistry
21:1876-1888.

Purpose of Review (DP Barcode or Litigation):  Registration Review

Date of Review:  4/22/08

Summary of Study Findings:

Background:  Nonylphenol, a common surfactant has been associated with
endocrine-disrupting effects.  Fomesafen is a diphenyl ethers (DPE)
herbicide, which are protopophyrinogen oxidase inhibitors.  DPEs induce
production of reactive oxygen species (ROS), resulting in cellular
membrane disruption.  This investigation was “designed to study the
influence of polyethoxylated nonylphenols formulated as Agral 90 on the
toxicity of fomesafen in the pond snail L. stagnalis, with particular
attention to the reproductive performance and underlying energetic and
hormonal processes (pg 1877).

Tests were preformed in both the laboratory and in mesocosms.

Snails tested in the laboratory, were housed in stainless steel tanks
with a density of  1.75-2.5 snails/L.  For the laboratory experiment,
test concentrations were 22 g/l technical (99% purity) fomesafen and
a combination of 22 g/l technical (99% purity) fomesafen and 50
g/l Agral 90.  No solvent was used.

Outdoor mesocosms contained phytoplantkton, zooplankton, aquatic
invertebrates and plants.  Snails were caged within the mesocosms.
Mesocosms were treated five times with either 40 g/l formulated
fomesafen or 40 g/l formulated fomesafen and 90 g/l Agral 90 at
three week intervals beginning in April.  Three replicates were used for
each treatment.  Fomesafen concentrations in both the water and the
snails was analytically confirmed.  Limit of detection was 1g/L
for water and 2 ng/g for snails.  Limit of quantitation was 5g/L
for water and 10 ng/g for snails.  

Endpoints evaluated were:

Organism level: survival, fecundity, egg development and hatching rate,
and F1 growth rate.

	

 

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4s; glycogen phosphorylase, UDP-galactose 4-epimerase, polysaccharide
content (glycogen and galactogen), protein, and gonad steroids
(testosterone and estradiol-17

Results:

Authors state intermesocosm variability for several endpoints was
significant, and was taken into account in the data analysis

g/L in water (estimated from Fig 1) to a peak of 60 g/L at
the conclusion of the exposure period (reported in text). 
Concentrations of fomesafen in the mixture treatment were consistently
lower than in the pesticide only treatment throughout the exposure
period.  Concentrations ranged from a low ~5 g/L in water
(estimated from Fig 1) to a peak of 30 g/L at the conclusion of
the exposure period (reported in text).  Authors did not measure
concentration of nonylphenol.

Reproductive performance:  Trends in reproductive effects appeared
similar between the laboratory and mesocosm experiments, although
specific values are difficult to compare given both the exposure
concentrations and exposure times were different.  There was no
significant effect on either number of egg masses or number of eggs per
mass in the laboratory experiment.  In the mesocosm experiment, there
was a significant effect on number of egg masses in the fomesafen-only
treatment after the first application, (Fig 4, pg 1881), but data
presented in Table 1 (pg 1882) indicate the snails in the fomesafen-only
group were depositing less egg masses even before treatment.  The
decrease in egg masses may not be treatment related.  There are no
significant differences between either of the treatments and the
controls at any other time period.  There was no significant differences
in development of egg masses, hatching rate, or growth of the F1
generation in the treatment groups.

Energetic biomarkers:  Laboratory exposures to fomesafen and the
fomesafen-Agral 90 mixture did not affect glycogen or glycogen
phosphorylase activity.  In the mesocosms there were significant
differences in glycogen content (at 12 days, Fig 5, pg 1883) and
glycogen phosphorylase (at 8 days and 35 days, Fig 5, pg 1883) in the
fomesafen treated group, but these reduced responses did not correlate
with one another in a time sequence, and trends were not consistent over
the exposure period.  There was no significant response in galactogen
content or UDP-galactose 4-epimerase activity in the treatment groups as
compared to the controls.

Hormonal biomarkers:  There were no significant differences in
testosterone-like or estradiol-like steroids in the  treatment groups as
compared to the control groups in the laboratory experiment.  Mean
steroid concentrations in the gonadodigestive complex were slightly
lower in the laboratory treatment groups than in the controls.  In the
mesocosms, authors state “steroid-like level values were significantly
lower in fomesafen-exposed snails as compared to the controls and the
snails exposed to the fomesafen-Agral 90 mixture (treatment effect
F[2,24]=3.00 p=0.069, and F[2,24]=5.04, p=0.015 for testosterone-like
and estradiol-17-like, respectively, Tukey =0.05). (pg 1884).”

g/L at the end of the contamination period) and the snails strongly
suggest that the surfactant properties of the adjuvant were responsible
for such an overwhelming effect (pg 1886).”

Reviewer’s Note:  In both laboratory and mesocosm experiments, control
mortality was high (20% and 29% respectively), indicating potential
problems with either viability of the organisms and/or husbandry issues.
 Given that both exposure durations and concentrations varied between
the laboratory experiment and the mesocosm experiment it is difficult to
correlate specific effects.  However, the glycogen content effects that
were significantly different in the mesocosms were not observed in the
laboratory as well as not occurring consistently over time or (based on
data presented in Fig 5, pg 1883) or occurring in any obvious time
sequence associated with the applications.  Although no variability
information was provided, based on the values in Table 1 (pg 1882), it
appears that the snails in the fomesafen treatment group evidenced
depressed egg production compared to the other groups even prior to
fomesafen exposure.  Authors do not provide any explanation as to why
this may have occurred.  Thus, any conclusions regarding fomesafen
effects on the snail reproduction based on these endpoints or related
endpoints, such as the lower steroid levels in the gonadodigestive
complex are questionable.   It is the reviewer’s opinion that no firm
conclusions regarding the effect of fomesafen on L. stagnalis can be
drawn based on the data presented in the paper.  Reviewer also believes
that application of the surfactant Agral 90 reduces exposure in the
dissolved phase (as evidence by the much lower measured water column
concentrations of fomesafen in the mixture mesocosm) rather than
necessarily reducing effects.

Description of Use in Document (QUAL, QUAN, INV):  INV  (Due to high
control mortality and depressed reproductive performance in fomesafen
treatment group prior to exposure.)

Rationale for Use:  Not used

Limitations of Study:  High control mortality and depressed reproductive
performance in fomesafen treatment group prior to exposure. 
Mean-measured value for mesocosm exposures can only be estimated from
figure, and analytical verifications do not appear to have been spaced
equally over time.  Fomesafen concentrations in laboratory experiment do
not appear to have been analytically verified.  Thus it cannot be
determined if fomesafen concentrations in the mixture treatment were the
same as the fomesafen only treatment or if the surfactant also reduced
dissolved phase fomesfen in the lab experiment.

Primary Reviewer:

Paige D. Doelling, Ph.D., Fisheries Biologist, ERB1

Secondary Reviewer 

Christine Hartless, Ph.D., Wildlife Biologist, ERB1

Open Literature Review Summary

Chemical Name:  Fomesafen 

PC Code:  123802

ECOTOX Record Number and Citation:

95404  Singh, KJ, Thakur RC, and Singh, OP.  (1988)  Toxicity of
insecticides, on eggs, nymphs, and adults of tur-pod bug (Calvigralla
gibbosa) and its ovipositional behavior on pigeonpea (Cajanus cajan). 
Indian Journal of Agricultural Sciences 58(8): 621-623.

Purpose of Review (DP Barcode or Litigation):  Registration Review

Date of Review:  4/25/08

Summary of Study Findings:

전・ý尀

resumably of the 1,200L/ha applied).  They report significant mortality
of nymphs (70-100%) and adults (67-90%) in the 72 hour observation
period following treatment.

Reviewer’s Note:  Fomesafen is not generally classed as an
insecticide.  Authors do not report specific source of pesticides used
in experiment.  An internet search for the product the authors describe
using, PP 21 5 EC, returned no hits, although references to PP 021 were
located.  PP 021 is not currently registered for use in the United
States.  Registrant-submitted data regarding fomesfen effects on
terrestrial insects (MRID135656), honeybees (ACC 135651), and earthworms
(ACC 135652) indicate relatively low or no toxicity for these classes of
organisms.  It is the reviewer’s opinion that either 1) the product
used was a multiple active ingredient product containing an insecticidal
active ingredient, or 2) the plants in the fomesafen plot were
contaminated by one of the other active ingredients evaluated, some of
which are insecticides toxic at low concentrations.

Given these uncertainties, and information contradictory to the bulk of
the available data, this study is classified invalid.

Description of Use in Document (QUAL, QUAN, INV):  INV

Rationale for Use:  Not used.

Limitations of Study:  Actual treatment concentration cannot be
calculated.  Cross-contamination by other test materials may have
occurred.

Primary Reviewer:

Paige D. Doelling, Ph.D., Fisheries Biologist, ERB1

Secondary Reviewer 

Christine Hartless, Ph.D., Wildlife Biologist, ERB1

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