UNITED STATES ENVIRONMENTAL PROTECTION AGENCY

WASHINGTON D.C., 20460

	PC Code:  000325

DP Barcodes:  357063

MEMORANDUM	Date:  January 28, 2010

SUBJECT:	Metrafenone:  Ecological Risk Assessment For Proposed Use on
Grapes

FROM:	Tanja Crk, M.A., Biologist

		James A. Hetrick, Ph.D., Senior Scientist

		Dana Spatz, Branch Chief

Environmental Risk Branch III

		Environmental Fate and Effects Division (7507P)

		Office of Pesticide Programs

TO:		Mary Waller, Product Manager

Fungicide Branch

		Registration Division (7505P)

		Office of Pesticide Programs

This memorandum transmits the Environmental Fate and Effects
Division’s (EFED) environmental risk assessment for the active
ingredient, metrafenone, as a fungicide for uses on grapes to treat
powdery mildew; it is used for preventive but not curative measures.
The proposed end-use product Metrafenone 300 is a suspension concentrate
containing 25.2% metrafenone.  Ground (liquid) application is the
proposed method of application. The maximum single application rate is
0.300 lbs a.i./A; at 6 applications per season, the seasonal maximum
rate is 1.80 lbs a.i./A. The minimum application interval is 14 days.

While metrafenone is expected to be slightly mobile, its major routes of
degradation are aqueous photolysis and aquatic metabolism, in both
aerobic and anaerobic conditions.  Soil metabolism is slow, with
laboratory half-lives of 6 months to a year and accumulation over two
years observed in field studies.  In aquatic environments, the measured
photolysis half-life was 6.4 days and metabolism in both aerobic and
anaerobic occurred with half-lives of one to four weeks. Exposure is
expected to be dominated by runoff and spray drift. Metrafenone exposure
may also result from off-site movement in runoff water and on
metrafenone -bearing soil particulates to adjacent fields (soil
erosion). The moderate Koc’s (Koc 1073 to 22517 L/kg-oc) of
metrafenone suggest that leaching to ground water/recharge to surface
water would not be a route of exposure. Long-range transport of
metrafenone in the gas phase is not considered a significant route of
exposure. While the high log Kow (4.3 at 25oC, pH 4) would suggest the
potential for bioaccumulation, the lipid normalized BCF (between 140 and
530) indicates that metrafenone is not expected to accumulate in tissues
of aquatic organisms.

The results of this screening-level assessment indicate a potential for
direct adverse effects to non-target mammals (dose-based RQs 1.14-2.91)
following chronic exposure; these RQs also exceed the federally listed
species LOC. Due to the potential for direct adverse effects to mammals
associated with the application of metrafenone on grapes, indirect
effects may consequently affect other aquatic and terrestrial species.
To reduce chronic risk to mammals, several components of the application
protocol would have to change. For example, in order to have no chronic
LOC exceedances (i.e., all RQs < 1) for mammals, the minimum single
application rate (0.2 lbs a.i./A) would have to be cut by 25% (i.e., to
0.15 lbs a.i./A) yet considered the maximum instead, the application
interval would have to nearly double (from 14 days up to 26 days), and
the maximum allowed number of applications would have to be cut from 6
to 5. Alternatively, in order to have no chronic LOC exceedances for
mammals, the minimum single application rate (0.2 lbs a.i./A) can still
be applied yet considered the maximum instead, but the application
interval would have to nearly double (from 14 days up to 26 days), and
the maximum number of applications would have to be cut by 50% (i.e.,
from 6 to 3). Furthermore, in order to have no chronic LOC exceedances
in nearly all cases for mammals –the one exception being an exceedance
for the 15g size glass consuming short grass where the calculated RQ is
1.02 – the minimum single application rate (0.2 lbs a.i./A) would be
changed to the maximum single application rate and the application
interval (14 days) could remain as currently prescribed by the label,
but the maximum number of applications would have to be reduced from 6
to 2.

Data were either not submitted or were deemed invalid for freshwater
invertebrates and marine/estuarine fish via chronic exposure. Without
data, risk cannot be ruled out for these taxa (either non-listed or
federally listed species).

Non-definitive endpoints – in this case, where total concentrations
were estimated instead of the preferred dissolved concentrations in the
majority of aquatic studies on metrafenone – suggest potential chronic
risk to federally listed freshwater fish and acute risk to federally
listed marine/estuarine invertebrates. More detailed risk conclusions
are provided in the environmental risk assessment document.

Data Needs

The ecotoxicity and environmental fate data needs are summarized in
Tables 1 and 2 below.  Further explanation and characterization of these
data needs can be found in the executive summary of the ecological risk
assessment.

Table 1. Ecological Toxicity Data Gaps

Study	Reason

850.1075 Acute toxicity freshwater fish	Non-definitive endpoint
w/effects observed

850.1400 Freshwater fish early-life stage (chronic)	Non-definitive
endpoint w/effects observed

850.1300 Aquatic freshwater invertebrate life-cycle (chronic)	Invalid,
CFR data gap

850.1025 Acute marine/estuarine invertebrate (oyster)

     Or

850.1035 Acute marine/estuarine invertebrate (mysid)	Non-definitive
endpoint w/effects observed

OECD 218 Freshwater invertebrate whole sediment study (chronic)	Invalid,
CFR data gap

850.4400 Aquatic vascular plant growth, Tier II	Non-definitive endpoint
w/effects observed

850.5400 Aquatic non-vascular plant growth, Tier II	Non-definitive
endpoint w/effects observed

850.2100 Avian acute oral toxicity test (passerine species)	Not
submitted, CFR data gap

850.3020 Honeybee acute contact toxicity (on TEP)	Address formulation
uncertainty

850.4100 Seedling Emergence, Tier II (on TEP)	Address formulation
uncertainty

850.4150 Vegetative Vigor, Tier II (on TEP)	Address formulation
uncertainty

Table 2.  Environmental Fate Data Gaps

835.6100 Independent Laboratory Validations

NEW CHEMICAL REGISTRATION

(Section 3)

      ECOLOGICAL RISK ASSESSMENT

Metrafenone: Fungicide

USEPA PC # 000325

Chemical Name(s):
(3-bromo-6-methoxy-2-methylphenyl)(2,3,4-trimethoxy-6-methylphenyl)metha
none (CAS)

	3’-bromo-2,3,4,6’-tetramethoxy-2’,6-dimethylbenzophenone (IUPAC)

Chemical Abstracts Service (CAS) Number: 220899-03-6

Chemical Family:   Benzophenone fungicide

Pesticidal Mode of Action:  Inhibits growth of mycelium on the leaf
surface, leaf penetration, formation of haustoria, and sporulation.
Likely affects actin proteins which play a role in cell function and
cell division.

Proposed End-use Product:  Metrafenone 300 SC (EPA Reg.No.7969-xxx);

                                              Suspension Concentrate,
25.2% (2.5 lbs a.i./gallon)

Target Pest(s):   Powdery mildew produced by Uncinula necator

Proposed Target Crop(s):   Grapes

Risk Assessors:	Tanja Crk, MA, Biologist

			James A. Hetrick, PhD, Senior Scientist

Environmental Risk Branch III

			Environmental Fate and Effects Division

Secondary Reviewers:	Pamela Hurley, PhD, Toxicologist

Environmental Risk Branch III

			Environmental Fate and Effects Division

Through:		Dana Spatz, Branch Chief

Environmental Risk Branch III

			Environmental Fate and Effects Division

Table of Contents

  TOC \o "1-5" \h \z \u    HYPERLINK \l "_Toc238026248"  I.	Executive
Summary	5

  HYPERLINK \l "_Toc238026249"  A.	Nature of the Chemical Stressor	5

  HYPERLINK \l "_Toc238026250"  B.	Potential Risks to Non-target
Organisms	5

  HYPERLINK \l "_Toc238026251"  C.	Environmental Fate Summary	6

  HYPERLINK \l "_Toc238026252"  D.	Ecological Effects Summary	7

  HYPERLINK \l "_Toc238026253"  E.	Uncertainties and Data Gaps	8

  HYPERLINK \l "_Toc238026254"  1.	Environmental Fate and Exposure	8

  HYPERLINK \l "_Toc238026255"  2.	Ecological Effects Data	9

  HYPERLINK \l "_Toc238026256"  F.	 Endangered Species Considerations	12

  HYPERLINK \l "_Toc238026257"  II.	Problem Formulation	13

  HYPERLINK \l "_Toc238026258"  A.	Nature of Regulatory Action	13

  HYPERLINK \l "_Toc238026259"  B.	Stressor Source and Distribution	13

  HYPERLINK \l "_Toc238026260"  1. 	Nature of the Chemical Stressor	13

  HYPERLINK \l "_Toc238026261"  a.	Mode of Action  (MoA) of Metrafenone
15

  HYPERLINK \l "_Toc238026262"  b.	Reactions of  Metrafenone in the
Environment	15   PAGEREF _Toc238026262 \h  Error! Bookmark not defined.

  HYPERLINK \l "_Toc238026263"  2.	Overview of Pesticide Usage	 15

  HYPERLINK \l "_Toc238026264"  C.	Receptors	15

  HYPERLINK \l "_Toc238026265"  1.	Aquatic and Terrestrial Effects	15

  HYPERLINK \l "_Toc238026266"  2.	Ecosystems Potentially at Risk	17

  HYPERLINK \l "_Toc238026267"  D.	Assessment Endpoints	17

  HYPERLINK \l "_Toc238026268"  E.	Conceptual Model	17

  HYPERLINK \l "_Toc238026269"  1.	Risk Hypothesis	17

  HYPERLINK \l "_Toc238026270"  2.	Conceptual Diagram	18

  HYPERLINK \l "_Toc238026271"  F.	Analysis Plan	 19

  HYPERLINK \l "_Toc238026272"  1.	Conclusions from Previous Risk
Assessments	19

  HYPERLINK \l "_Toc238026273"  2.	Preliminary Identification of Data
Gaps	19

  HYPERLINK \l "_Toc238026274"  3.	Measures of Exposure and Effects	23

  HYPERLINK \l "_Toc238026275"  a.	Aquatic Exposure Models	24

  HYPERLINK \l "_Toc238026276"  b.	Terrestrial Exposure Models	24

  HYPERLINK \l "_Toc238026277"  III.	Analysis	28

  HYPERLINK \l "_Toc238026278"  A.	Use Characterization	28

  HYPERLINK \l "_Toc238026279"  B.	Exposure Characterization	28

  HYPERLINK \l "_Toc238026280"  1.	Environmental Fate and Transport	28

  HYPERLINK \l "_Toc238026281"  2.	Measures of Aquatic Exposure	31

  HYPERLINK \l "_Toc238026282"  a.	Aquatic Exposure Modeling	31
HYPERLINK \l "_Toc238026283"

  HYPERLINK \l "_Toc238026287"  b.	Aquatic Exposure Monitoring and Field
Data	  PAGEREF _Toc238026287 \h  34

  HYPERLINK \l "_Toc238026288"  c.	Aquatic Bioaccumulation Assessment
PAGEREF _Toc238026288 \h  34

  HYPERLINK \l "_Toc238026289"  3.	Measures of Terrestrial Exposure	36

  HYPERLINK \l "_Toc238026290"  a.	Terrestrial Exposure Modeling	36

  HYPERLINK \l "_Toc238026292"  C.	Ecological Effects Characterization
38

  HYPERLINK \l "_Toc238026293"  1.	Aquatic Effects Characterization	38

  HYPERLINK \l "_Toc238026294"  a.	Aquatic Animals	38

  HYPERLINK \l "_Toc238026295"  (1)	 Acute Effects	38

  HYPERLINK \l "_Toc238026296"  (2)	Chronic Effects	47

  HYPERLINK \l "_Toc238026298"  b.	Aquatic Plants	49

  HYPERLINK \l "_Toc238026299"  2.	Terrestrial Effects Characterization
55

  HYPERLINK \l "_Toc238026300"  a.	Terrestrial Animals	55

  HYPERLINK \l "_Toc238026301"  (1)	Acute Effects	55

  HYPERLINK \l "_Toc238026302"  (2)	Chronic Effects	62

  HYPERLINK \l "_Toc238026304"  b.	Terrestrial Plants	67

  HYPERLINK \l "_Toc238026305"  IV.	Risk Characterization	68

  HYPERLINK \l "_Toc238026306"  A.	Risk Estimation - Integration of
Exposure and Effects Data	 68

  HYPERLINK \l "_Toc238026307"  1.	Risk to Aquatic Animals and Plants	68

  HYPERLINK \l "_Toc238026308"  a.	Aquatic Animals	68

  HYPERLINK \l "_Toc238026309"  (1)	Risk Following Acute Exposure	68

  HYPERLINK \l "_Toc238026310"  (2)	Risk Following Chronic Exposure	69

  HYPERLINK \l "_Toc238026311"  b.	Aquatic Plants	70

  HYPERLINK \l "_Toc238026312"  2.	Risk to Terrestrial Animals and
Plants	70

  HYPERLINK \l "_Toc238026313"  a.	Terrestrial Animals	70

  HYPERLINK \l "_Toc238026314"  (1)	Risk Following Acute Exposure	71

  HYPERLINK \l "_Toc238026315"  (2)	Risk Following Chronic Exposure	72

  HYPERLINK \l "_Toc238026316"  b.	Terrestrial Plants	75

  HYPERLINK \l "_Toc238026317"  B.	Risk Description	75

  HYPERLINK \l "_Toc238026318"  1.	Risk to Aquatic Animals and Plants	75

  HYPERLINK \l "_Toc238026319"  a.	Aquatic Animals	75

  HYPERLINK \l "_Toc238026320"  (1)	Risk Following Acute Exposure	75

  HYPERLINK \l "_Toc238026321"  (2)	Risk Following Chronic Exposure	78

  HYPERLINK \l "_Toc238026322"  b.	Aquatic Plants	79

  HYPERLINK \l "_Toc238026323"  2.	Risk to Terrestrial Animals and
Plants	80

  HYPERLINK \l "_Toc238026324"  a.	Terrestrial Animals	80

  HYPERLINK \l "_Toc238026325"  (1)	Risk Following Acute Exposure	80

  HYPERLINK \l "_Toc238026326"  (2)	Risk Following Chronic Exposure	82

  HYPERLINK \l "_Toc238026327"  b.	Terrestrial Plants	83

  HYPERLINK \l "_Toc238026328"  3.	Review of Incident Data	84

  HYPERLINK \l "_Toc238026329"  4.	Endocrine Effects	84

  HYPERLINK \l "_Toc238026330"  5.	Federally Threatened and Endangered
(Listed) Species..	85

  HYPERLINK \l "_Toc238026331"  a.	Action Area	86

  HYPERLINK \l "_Toc238026332"  b.	Taxonomic Groups Potentially at Risk
86

  HYPERLINK \l "_Toc238026333"  (1)	Discussion of Risk Quotients	89

  HYPERLINK \l "_Toc238026334"  (2)	Probit Dose Response Relationship	89

  HYPERLINK \l "_Toc238026335"  (3)	Data Related to Under-represented
Taxa	90

  HYPERLINK \l "_Toc238026336"  (4)	Implications of Sublethal Effects	90

  HYPERLINK \l "_Toc238026337"  c.	Indirect Effects Analysis	92

  HYPERLINK \l "_Toc238026338"  d.	Critical Habitat	93

  HYPERLINK \l "_Toc238026339"  e.	Co-occurrence Analysis	94

  HYPERLINK \l "_Toc238026340"  C.	Description of Assumptions,
Limitations, Uncertainties, Strengths and Data Gaps
………………………………………………………………
…………………....95

  HYPERLINK \l "_Toc238026341"  1.	Assumptions, Limitations, and
Uncertainties Related to Exposure For All
Taxa…………………………………………………………
………………………95  	a. Maximum Use
Scenario……………………………………………………95

  HYPERLINK \l "_Toc238026342"  2.	Assumptions, Limitations and
Uncertainties  Related to Exposure For Aquatic Species	95

  HYPERLINK \l "_Toc238026343"  Environmental Fate
Studies……………………………………………...…95

Aquatic Exposure Model	95

  HYPERLINK \l "_Toc238026344"  b.	Bioaccumulation Modeling	96

  HYPERLINK \l "_Toc238026345"  3.	Assumptions, Limitations and
Uncertainties Related to Exposure for Terrestrial Species	96

  HYPERLINK \l "_Toc238026346"  a.	Location of Wildlife Species	96

  HYPERLINK \l "_Toc238026347"  b.	Routes of Exposure	96

  HYPERLINK \l "_Toc238026348"  c.	Dietary Intake and Other Limitations
of Oral Studies in Terrestrial Species	96

  HYPERLINK \l "_Toc238026349"  d.	Incidental Releases Associated With
Use	98

  HYPERLINK \l "_Toc238026350"  e.	Residue Levels Selection	99

  HYPERLINK \l "_Toc238026351"  f.	TerrPlant Model	99

  HYPERLINK \l "_Toc238026352"  4.	Assumptions, Limitations and
Uncertainties Related to Effects Assessment	99

  HYPERLINK \l "_Toc238026354"  a.	Sublethal Effects	99

  HYPERLINK \l "_Toc238026355"  b.	Age Class and Sensitivity of Effects
Thresholds	99

  HYPERLINK \l "_Toc238026356"  c.	Use of Most Sensitive Species Tested
100

  HYPERLINK \l "_Toc238026357"  5.	Assumptions, Limitations,
Uncertainties, Strengths and Data Gaps Related to the Acute and Chronic
LOC’s	100

  HYPERLINK \l "_Toc238026358"  V.	Literature Cited	101

  HYPERLINK \l "_Toc238026359"  VI.	Appendices	 109

VII.   LIST OF
FIGURES………………………………………………………
…….109

VIII. LIST OF
TABLES…………………………………………………………
…...109

Executive Summary

Nature of the Chemical Stressor

Metrafenone
(3-bromo-6-methoxy-2-methylphenyl)(2,3,4-trimethoxy-6-methylphenyl)metha
none; CAS Reg. No. 220899-03-6) is a new pesticide active ingredient
submitted for registration in the United States. Metrafenone is a
benzophenone fungicide with resistance code U8 (unknown mode of action).
What is known is that the chemical inhibits growth of mycelium on the
leaf surface, leaf penetration, formation of haustoria, and sporulation
of the target fungus, Uncinula necator (EPA Pesticide Fact Sheet 2006).
Metrafenone likely affects actin proteins which play a role in cell
function and cell division (Opalski et al. 2006). It has a local
inhibition effect; in other words, it is not systemic and not a contact
killer.

As a new pesticide active ingredient, the actual usage of metrafenone is
not known. The end-use product Metrafenone 300 SC is proposed for use on
grapes to treat powdery mildew; it is used for preventive but not
curative measures. Ground (liquid) application is the proposed method of
application. The maximum single application rate is 0.300 lbs a.i./A; at
6 applications per season, the seasonal maximum rate is 1.80 lbs a.i./A.
The minimum application interval is 14 days.

Potential Risks to Non-target Organisms

The results of this screening-level assessment indicate a potential for
direct adverse effects to non-target mammals (dose-based RQs 1.14-2.91)
following chronic exposure; these RQs also exceed the federally listed
species LOC. Due to the potential for direct adverse effects to mammals
associated with the application of metrafenone on grapes, indirect
effects may consequently affect other aquatic and terrestrial species.
Data were either not submitted or were deemed invalid for freshwater
invertebrates and marine/estuarine fish via chronic exposure. Without
data, risk cannot be ruled out for these taxa (either non-listed or
federally listed species). Non-definitive endpoints – in this case,
where total concentrations were estimated instead of the preferred
dissolved concentrations in the majority of aquatic studies on
metrafenone – suggest potential chronic risk to federally listed
freshwater fish and acute risk to federally listed marine/estuarine
invertebrates.  Comparison of the non-definitive endpoints with aquatic
EECs indicates potential risk to the given federally listed taxa (see
Section I.F below, Table 3) but not the non-listed taxa summarized in
Table 1 and Table 2.

Table   SEQ Table \* ARABIC  1 .  Summary of Environmental Risk
Conclusions for Aquatic Animals and Plants from Metrafenone Use on
Grapes at the Maximum Proposed Application Rate (0.3 lb a.i./A, Assuming
6 Applications/Year)*

Taxonomic Group	Assessment Endpoint

Summarized Risk Characterization and Important Uncertainties

Freshwater Fish and Aquatic Phase Amphibians	Mortality	Acute risk is not
expected (from technical grade active ingredient, metabolites, and EU/UK
formulation).

	Reproduction, growth etc.	Chronic risk is not expected.

Freshwater Invertebrates	Mortality	Acute risk is not expected (from
technical grade active ingredient, metabolites, and EU/UK formulation).

	Reproduction, growth etc.	No acceptable studies available. Chronic risk
cannot be precluded.

Marine/

Estuarine Fish	Mortality	Acute risk is not expected.

	Reproduction, growth etc.	No studies submitted. Chronic risk cannot be
precluded.

Marine/

Estuarine Invertebrates	Mortality	Acute risk is not expected.

	Reproduction, growth etc.	Chronic risk is not expected.

Aquatic Plants	Acute Risk	Risk to vascular species is not expected from
technical grade active ingredient. In addition, risk to non-vascular
species is not expected (from the technical grade active ingredient,
metabolites, and EU/UK formulation).

* Consult ‘Risk Description’ section for further details. Also, risk
in this table implies risk to technical grade active ingredient unless
otherwise specified that metabolites and formulations were assessed as
well.

Table   SEQ Table \* ARABIC  2 .  Summary of Environmental Risk
Conclusions for Terrestrial Animals and Plants from Metrafenone Use on
Grapes at the Maximum Proposed Application Rate (0.3 lbs a.i./A,
Assuming 6 Applications/Year)*

Taxonomic Group	Risk Endpoint

Summarized Risk Characterization and Important Uncertainties

Birds, Reptiles and Terrestrial Phase Amphibians	Mortality	Acute risk is
not expected.

	Reproduction, growth etc.	Chronic risk is not expected.

Mammals	Mortality	Acute risk is not expected.

	Reproduction, growth etc.	Chronic risk is expected.

Non-target Invertebrates

Acute Risk	Acute risk to honeybees is not expected. Acute risk to
earthworms (from the technical grade active ingredient, a metabolite,
and EU/UK formulation) is not expected.

Terrestrial Plants	Acute Risk	Risk to terrestrial plants is not expected
(from EU/UK formulation).

* Consult ‘Risk Description’ section for further details. Also, risk
in this table implies risk to technical grade active ingredient unless
otherwise specified that metabolites and formulations were assessed as
well.

Environmental Fate Summary

While metrafenone is expected to be slightly mobile, its major routes of
degradation are aqueous photolysis and aquatic metabolism, in both
aerobic and anaerobic conditions.  Soil metabolism is slow, with
laboratory half-lives of 6 months to a year and accumulation over two
years observed in field studies.  In aquatic environments, the measured
photolysis half-life was 6.4 days and metabolism in both aerobic and
anaerobic occurred with half-lives of one to four weeks.

Soil photolysis and anaerobic aquatic metabolism of metrafenone led to
one major degradate, CL 377160 [Methanone,
(3-bromo-6-methoxy-2-methylphenyl)(3-hydroxy-2,4-dimethoxy-6-methylpheny
l)-], and anaerobic aquatic metabolism also led to six other major
degradates, but all were either unidentified or only tentatively
identified.  In addition, there were many minor degradates, which were
formed at low levels individually but reached substantial amounts as
groups.

μg/L for the daily peak, 8.82 μg/L for the 21 day average, and 6.21
μg/L for the 60 day average. The highest aquatic EECs for total
metrafenone residues were 20.22 μg/L for the daily peak, 17.67 μg/L
for the 21 day average, and 16.98 μg/L for the 60 day average.

No monitoring data are available to compare with model estimates.
Bioaccumulation modeling was conducted because metrafenone has a log
octanol:water coefficient > 4.  Maximum residue concentrations in fish
are expected to range from 17,636 to 19,586 µg/kg-ww.

Ecological Effects Summary

Aquatic Organisms

The greatest amount of uncertainty in the assessment stems from aquatic
studies which were largely based on total concentrations (both dissolved
and undissolved) of the test compound. In all cases (except for the
chronic study on the saltwater mysid, but including aquatic plants), the
risk quotient values were not calculated. However, given comparisons of
the total concentrations for most available studies and soluble
concentrations for the saltwater mysid (chronic) and sheesphead minnow
(acute) to respective EEC values the implication is that acute and
chronic risk (including sublethal effects) to aquatic organisms
(including aquatic plants) is not expected as a result of metrafenone
use on grapes. However, the taxa for which studies were either submitted
and deemed unacceptable (chronic freshwater invertebrate studies) or not
submitted at all (chronic marine/estuarine fish), the risk as a result
of metrafenone use cannot be precluded.

Terrestrial Organisms

The acute oral avian studies indicated no effects; the acute dietary
avian studies indicated a significant change in body weight that was not
associated with a dose-response pattern. In the acute mouse study on the
technical grade active ingredient, only one mortality was reported.
Therefore, no implications with lethal or sublethal acute effects can be
made for either avian or mammalian taxa. Metrafenone is classified as
‘practically non-toxic’ to honeybees and non-lethal to earthworms up
to the limit concentration. Therefore, the implication is that acute
risk to terrestrial invertebrates is not expected as a result of
metrafenone use on grapes. In addition, no terrestrial plant risks are
expected as a result of use of the EU/UK formulation; however, no data
on terrestrial plants are available on the U.S. formulation.

The avian chronic toxicity endpoint (NOAEC 848 mg a.i./kg diet) based on
egg production and hatchability did not yield chronic LOC exceedances.
The most sensitive chronic mammalian endpoint (NOAEL 35.9 mg/kg bw/day)
based on decreased body weights and body weight gain in F1 males as well
as body weights in F1 and F2 females, however, exceeded the chronic LOC,
which implies that sublethal chronic effects on mammals are expected
under field conditions. Therefore, given the LOC exceedance chronic risk
to mammals is expected as a result of metrafenone use on grapes.

Uncertainties and Data Gaps

Environmental Fate and Exposure

All but one of the environmental fate studies required to support the
registration of metrafenone for use on grapes were submitted.
Independent Laboratory Validations for the methods submitted for
analysis of metrafenone in soil and water remain a data gap. These data
gaps fall under the terrestrial field dissipation study guideline
(835.6100). For a list of submitted environmental fate studies for
metrafenone see Appendix F.

All of the environmental fate studies for the parent compound were
determined to be scientifically valid and therefore results from all of
the studies can be used to characterize the mobility and rates of
transformation of metrafenone.  However, many of the metabolism studies
have major uncertainties in the identification and pattern of formation
and decline of transformation products.  In all of the aquatic
metabolism studies, between 57% and 65% of the applied radioactivity
remains unidentified with incomplete characterization, and in two
aerobic soil metabolism studies, 15% and 44% of the applied
radioactivity is unidentified.   This includes at least four major
degradates that individually reach levels of 11% to 35% of the applied.
Other transformation products appear as groups of up to 15 components,
in some cases characterized as each being <5% of the applied
radioactivity, but in other cases, some individual components make up 9%
to 10% of the applied.  Even when individual components can all be
classified as minor degradates, these groups represent such a large
portion of the applied radioactivity overall that the possibility that
they may have some impact as a group cannot be precluded despite their
lower individual levels.  This is especially true given that the
degradation pathways suggest that groups of degradates may have a high
degree of structural similarity and so may have similar fate and effects
behavior.  Without information to adequately characterize the
degradates, it was necessary to assume that they are of equal toxicity
to the parent in order to quantify risks for metrafenone extractable
residues.

In addition to the studies submitted by the registrant, EFED was
provided with a Draft Assessment Report (DAR) on Metrafenone written by
the United Kingdom.  That assessment reports that the long dissipation
time observed in field studies triggered a requirement for longer term
field accumulation studies. Two years into those five year studies,
results show that metrafenone appears to be accumulating, and that
metrafenone residues have been detected as deep as 20-30 cm below the
surface. These studies would provide useful information regarding the
environmental fate of metrafenone and would assist in reducing
uncertainties.

Ecological Effects Data

The submitted ecotoxicity database is incomplete. The greatest amount of
uncertainty in the assessment stems from aquatic studies which were
largely based on total (both dissolved and undissolved) concentrations
of test compound; that is, the concentrations were measured without
centrifugation even though precipitate was observed or test was
conducted at the solubility limit of the metrafenone technical grade
active ingredient; the solubility limits of the metabolites are unknown.
To reduce uncertainty the majority of the toxicity studies, which are
considered supplemental, and cannot be used in a quantitative risk
estimation would need to be redone; especially those where mortality or
sublethal effects were observed. These aquatic studies are specified
below. For a list of submitted ecological effects studies for
metrafenone see Appendix G.

The following studies are considered data needs:

	Aquatic studies

Given the freshwater fish studies on metrafenone technical grade active
ingredient and assuming that the concentrations in the environment reach
the solubility limit, the effect of the compound is likely to be low.
However, according to model estimated EECs (which include metrafenone
and metrafenone residue scenarios: 0.00153 - 0.02 mg/L), levels of
metrafenone (TGAI, TEP) at the solubility limit (0.2-0.5 mg/L at 12oC)
and metrafenone metabolites at the tested concentrations are not
expected to occur in the environment given the proposed grape use.
Therefore, acute risk to freshwater fish and aquatic-phase amphibians is
not expected as a result of metrafenone use on grapes. In addition,
although the EU/UK formulation – with which the submitted studies were
conducted – closely matches the U.S. formulation (i.e., BAS 560 03F)
it is still not its equivalent. Therefore, the effect of the U.S.
formulated product on freshwater fish and aquatic-phase amphibians is
not known.

To reduce uncertainty in characterizing risk, which is currently based
on total concentrations, the freshwater fish studies on bluegill sunfish
(Lepomis macrochirus) are requested to make same species comparisons for
acute and chronic values. Bluegill was chosen over the rainbow trout due
to greater sensitivity (on an acute basis) of the bluegill to the TGAI
relative to the trout. Since the acute marine/estuarine fish study is
acceptable and given the above two studies, the acute to chronic ratio
can be utilized to estimate marine/estuarine chronic endpoints, negating
the need to fill this data gap. Special emphasis must be placed on
centrifuging the test samples prior to analytical determination of the
test compound.

 Acute: Freshwater fish toxicity (96-hour LC50 for Lepomis macrochirus)
(850.1075; 72-1), TGAI

 Chronic: Fish early-life stage (freshwater: Lepomis macrochirus)
(850.1400; 72-4), TGAI

Although acute freshwater invertebrate studies were based on total
concentrations, no effects were observed with the TGAI, negating the
need to request additional data. However, a chronic freshwater
invertebrate data gap still exists since the two reviewed studies were
deemed invalid.

Chronic: Aquatic invertebrate life cycle (freshwater: Daphnia magna)
(850.1300; 72-4), TGAI

Effects were observed in both acute marine/estuarine invertebrate
studies with the TGAI on the eastern oyster (Crassostrea viginica) and
saltwater mysid (Americamysis bahia) but the endpoints are based on
total concentrations. Therefore, should concentrations in the
environment reach the solubility limit, the acute risk to
marine/estuarine invertebrates may be expected. However, according to
the model estimated EECs (0.00153 - 0.02 mg/L, which includes
metrafenone and total metrafenone residue scenarios), levels of
metrafenone at the solubility limit are not expected to occur in the
environment given the proposed grape use. Given these studies and
assuming that metrafenone concentrations in the environment are not
likely to reach the solubility limit, the acute risk to marine/estuarine
invertebrates is not expected. However, to reduce uncertainty in
characterizing risk, either study is requested to be redone with special
emphasis on centrifuging the test samples prior to analytical
determination of the test compound.

Acute: Marine/estuarine invertebrate either Crassostrea virginica
(850.1025; 72-3) or Americamysis bahia (850.1035; 72-3), TGAI

A non-guideline chronic midge study (Chironomus riparius) with
metrafenone technical (97.1%) was deemed invalid on the basis of low
negative control emergence. Therefore, a chronic freshwater invertebrate
whole sediment study is requested. A protocol should be submitted for
approval prior to study initiation. The protocol should include spiked
sediment. The whole sediment study is requested because metrafenone has
a high logKow value (4.3 at 25oC, pH 4) and a high aerobic soil half
life (> 4 months), which indicates that the compound may partition to
sediment and persist.

Chironomus dilutus (freshwater) using TGAI. Consult EPA Test Method
100.5 Life-cycle Test for Measuring the Effects of Sediment Associated
Contaminants on Chironomus dilutus (formerly Chironomus tentans) and
OECD Guideline 218 Sediment-Water Chironomid Toxicity Test Using Spiked
Sediment

Non-target aquatic plant studies were also based on total concentrations
leading to uncertainty in exposure concentrations and thus endpoint
values. Had valid endpoints (i.e., those based on dissolved
concentrations) been determined using these studies, they would likely
be greater than the highest model predicted concentration in the
environment, which implies that risk to vascular and non-vascular
species is not expected as a result of metrafenone use on grapes.
However, without centrifugation, the amount of chemical that is freely
dissolved and bioavailable cannot accurately be determined. Therefore,
vascular and non-vascular aquatic plant studies are requested with
special emphasis placed on centrifuging the test samples prior to
analytical determination of the test compound. In addition,
cyanobacteria yielded no effects given total concentrations and does not
need to be redone.

Aquatic vascular plant growth (Lemna spp.), Tier II (850.4400), TGAI

Aquatic non-vascular plant growth, Tier II (850.5400), TGAI

     Terrestrial Studies

An avian acute oral toxicity test in passerine species is required.
Passerines are the most common birds (in terms of numbers and number of
species) in the United States.  Many utilize agricultural fields,
forests, residential areas and surrounding areas, and, therefore, have
the potential to be exposed to pesticides used in agricultural, forest,
and residential settings.  It is likely that, for the requested use
patterns, passerines are more likely to be exposed to metrafenone than
upland game species and waterfowl.  Passerines are smaller and have a
higher energy requirement than larger-sized birds.  As such, passerines
may be more sensitive than other birds.

Passerine bird toxicity study (EPA approved protocol is required prior
to study initiation): Avian acute oral toxicity test (850.2100, 71-1),
TGAI

Formulated Product Testing

The available formulation product studies were conducted using three
different UK/EU formulated products (i.e., BAS 560 00F, BAS 560 01F, and
BAS 560 02F), one of which (BAS 560 00F) closely matches the U.S.
formulation (i.e., BAS 560 03F). Formulation studies in this assessment
are based on BAS 560 00F because data on the U.S. formulation were not
submitted. In order to eliminate uncertainty in effects characterization
of formulated products used within the U.S. on given taxa, future
registrant submitted studies should be based on the U.S. formulation. At
this time the effect of the U.S. formulation on given taxa is not known,
however, greater toxicity of the EU/UK formulation on the honeybee
relative to the TGAI was observed. Similarly, data submitted for
terrestrial plants is based on the EU/UK formulation. As a result, the
following studies are requested:

Honeybee acute contact toxicity (850.3020, 141-1), TEP

Seedling emergence, Tier II (850.4225, 123-1), TEP

Vegetative vigor, Tier II (850.4250, 123-1), TEP

Endangered Species Considerations

Table 3 summarizes the listed species at risk associated with either
direct or indirect effects following application of metrafenone for the
proposed uses.

Concerns For Federally Listed as Endangered and/or Threatened Species

Table 3.  Listed Species Risks Associated With Direct or Indirect
Effects from Metrafenone use on Grapes at the Maximum Proposed
Application Rate (0.3 lbs a.i./A, Assuming 6 Applications/Year)  TC
"Table IB-1.  Listed species risks associated with direct or indirect
effects due to applications of propazine on sorghum" \f C \l "1"

Listed Taxon	Direct Effects	Indirect Effects

Terrestrial and semi-aquatic plants - monocots	No	Yes from effects to
mammals

Terrestrial and semi-aquatic plants – dicots	No	Yes from effects to
mammals

Terrestrial invertebrates	No	Yes from effects to mammals

Birds	No	Yes from effects to mammals, FW fish, FW inverts, M/E fish,
M/E inverts (mollusks)

Terrestrial-phase amphibians	No	Yes from effects to mammals

Reptiles	No	Yes from effects to mammals, FW fish, FW inverts, M/E fish,
M/E inverts (mollusks)

Mammals	Yes for chronic1	Yes from effects to mammals, FW fish, FW
inverts, M/E fish,  M/E inverts (mollusks)

  Aquatic non-vascular plants	No	Yes from effects to mammals, FW fish,
FW inverts, M/E fish,  M/E inverts (mollusks)

Aquatic vascular plants	No	Yes from effects to mammals, FW fish, FW
inverts, M/E fish,  M/E inverts (mollusks)

Freshwater (FW) fish	Yes for chronic2	Yes from effects to mammals, FW
fish, FW inverts, M/E fish,  M/E inverts (mollusks)

Aquatic-phase amphibians	Yes for chronic3	Yes from effects to mammals,
FW fish, FW inverts, M/E fish,  M/E inverts (mollusks)

Freshwater (FW) invertebrates	Yes for chronic4	Yes from effects to
mammals, FW fish, FW inverts, M/E fish,  M/E inverts (mollusks)

Marine/estuarine (M/E) fish	Yes for chronic4	Yes from effects to
mammals, FW fish, FW inverts, M/E fish,  M/E inverts (mollusks)

Marine/estuarine (M/E) invertebrates (mollusk)	Yes for acute5,

No for chronic6 	Yes from effects to mammals, FW fish, FW inverts, M/E
fish,  M/E inverts (mollusks)

1 The chronic LOC is exceeded on a dose basis for mammals in all size
classes eating short grass, for the 15 and 35 gram size classes eating
tall grass and broadleaf plants / small insects.  The chronic LOC on a
dietary basis is not exceeded for any of the food categories.

2 The total concentration based endpoint (NOAEC: 0.118 mg total a.i./L)
for the chronic freshwater fish (fathead minnow) study (MRID 47267449)
with an effect on post-hatch survival is approximately 7x greater than
the highest estimated EEC (0.016 mg/L), hence risk to federally listed
freshwater fish cannot be precluded.

3 Results from freshwater fish used as surrogate for assessing risk to
aquatic-phase amphibians

4 Studies not submitted or invalid for which risk cannot be precluded.

5 Mollusk (Eastern oyster); 6 Saltwater mysid

Problem Formulation

The purpose of this problem formulation is to provide the foundation for
the ecological risk assessment being conducted for the fungicide
metrafenone. As such, it articulates the purpose and objectives of the
risk assessment, evaluates the nature of the problem, and provides a
plan for analyzing the data and characterizing the risk (EPA, 1998).

Nature of Regulatory Action

Metrafenone is a new pesticide active ingredient being proposed as a
fungicide to control powdery mildew on grapes.  As a new active
ingredient submitted for registration, there are no previously prepared
ecological risk assessments by the Agency for metrafenone uses. However,
the European Commission has issued a Dossier for Metrafenone (DAR 2005).

Stressor Source and Distribution

The stressor is the metrafenone and its unidentified, extractable
degradation products when applied to grapes to control powdery mildew.
Therefore, metrafenone could potentially be used anywhere in the United
States where grapes are grown.

Nature of the Chemical Stressor

Figure 1 provides the chemical structure of metrafenone. Table 4
identifies the physical and chemical properties of metrafenone from
experimental data.

Figure 1. Chemical Structure of Metrafenone

Table 4. Environmental Fate Properties of Metrafenone

PARAMETER	VALUE(S) (units)	SOURCE	COMMENT

  Chemical Name
(3-Bromo-6-methoxy-2-methylphenyl)(2,3,4-trimethoxy-6-methylphenyl)-meth
anone	–	–

  Molecular Formula	C19H21BrO5.

  Molecular Weight	409	_	–

  Solubility (20 oC)	0.474 mg/L or ppm	MRID: 46415711	“Slightly
soluble” according to FAO Classification

  Vapor Pressure (20 oC)	1.15 x 10-6 mmHg	MRID: 46415713	–

  Henry’s Law constant	1.3 x 10-6 atm-m3/mole

Estimated from vapor pressure and water solubility.

  pKa (20 oC)	None	MRID: 46415714	–

Octanol-Water Partition Coefficient

 (Log KOW,  at 25 oC, pH 4)	4.3	MRID: 46415715	–

  Hydrolysis Half-life

  (pH 5, 7, 9; (50 oC))	Stable  	MRID: 47267422	Stable at all pHs.

  Aqueous Photolysis Half-life

  (pH 5)	t1/2  = 6.4 days	MRID: 47267423	Value corrected to represent
natural sunlight at 40°N latitude; uncorrected lab half-life of 3.2
days (continuous irradiation; xenon lamp).

  Soil Photolysis Half-life	t1/2  = 31 days	MRID: 47267424	–

Aerobic Soil Metabolism

Half-life	Silt loam:  	t1/2  = 178-277 d

Sandy loam:  	t1/2  = 277, 365 d

Clay loam: 	t1/2  = 299, 330 d

Loamy sand: 	t1/2  = 182 d

DT50 =  160 – 270 d	MRIDs:

47267425

47267426

47267427	8 systems, 4 conducted with foreign soils.

In 2 studies metrafenone residues exceeded 50% at study termination (120
& 210 d)

  Anaerobic Aquatic Metabolism

  Half-life	t1/2  = 21.6 d, 18.0 d

DT50 =  15 d, 3 d	MRIDs:

47267429

47267431	–

  Aerobic Aquatic Metabolism

  Half-life	t1/2  = 27.3 d, 24.1 d

DT50 =  10 d, 11 d	MRID: 47267430	–

  Organic Carbon Partition

  Coefficient (KOC)	1073, 2230, 2331,

22517, 2792 mL/gOC

	MRID: 47267420	“Slightly mobile” according to FAO Classification

  Soil Partition Coefficient (Kd)	15.1, 36.5, 40.1,

42.0, 86.5 mL/g	MRID: 47267420	--

Terrestrial Field Dissipation	t1/2  =144 d, 161 d, 210 d, 161 d

	MRID 47267432	Field studies were conducted in WA, ON,CA, FL

Bioaccumulation in Fish	BCF: 140 to 530	MRID 47267450	Lipid Normalized
BCF

Mode of Action (MoA) of Metrafenone

Metrafenone is a benzophenone fungicide. What is known is that the
chemical inhibits growth of mycelium on the leaf surface, leaf
penetration, formation of haustoria, and sporulation of the target
fungus, Uncinula necator (EPA Pesticide Fact Sheet 2006). Metrafenone
likely affects actin proteins which play a role in cell function and
cell division (Opalski et al. 2006).

Reactions of Metrafenone in the Environment

Metrafenone is expected to be slightly mobile.  Its major routes of
degradation are aqueous photolysis and aquatic metabolism, in both
aerobic and anaerobic conditions.  Soil metabolism is slow, with
laboratory half-lives of 6 months to a year and accumulation over two
years observed in field studies.  In aquatic environments, the measured
photolysis half-life was 6.4 days and metabolism in both aerobic and
anaerobic occurred with half-lives of one to four weeks.

Soil photolysis and anaerobic aquatic metabolism of metrafenone led to
one major degradate, CL 377160, and anaerobic aquatic metabolism also
led to six other major degradates, but all were either unidentified or
only tentatively identified.  In addition, there were many minor
degradates, which were formed at low levels individually but reached
substantial amounts as groups.

Overview of Pesticide Usage

As a new pesticide active ingredient, the actual usage of metrafenone is
not known. The end-use product Metrafenone 300 SC is proposed for use on
grapes.

Ground application is the proposed method of application.  The label
states, “do not apply directly to water, to areas where surface water
is present, or to intertidal areas below the mean high water mark.”
The chemical is to be applied at most 6 times per season at a maximum
application rate of 0.3 lbs a.i./A (Table 5). The label also states,
“do not make more than 2 sequential applications of Metrafenone 300 SC
fungicide before alternating to a labeled fungicide with a different
mode of action.”

Table 5.  Metrafenone use and application information based on the
proposed label for Metrafenone  300 SC

Use	Max. Single App. Rate

(lbs a.i./A)	# of App. /

Season	Seasonal Max. Rate (lbs a.i./A)	Minimum App. Interval (days)

Grapes	0.300	6	1.80	14 days

Receptors

Aquatic and Terrestrial Effects

The receptor is the biological entity that is exposed to the stressor
(EPA, 1998). Based on the proposed uses for metrafenone, it is expected
that the aquatic and terrestrial receptors will include freshwater fish
and invertebrates, marine/estuarine fish and invertebrates, aquatic
plants, terrestrial plants, birds, mammals, and terrestrial
invertebrates.

Consistent with the process described in the Overview Document (EPA,
2004), this risk assessment uses a surrogate species approach in its
evaluation of metrafenone.  Toxicological data generated from surrogate
test species, which are intended to be representative of broad taxonomic
groups, are used to extrapolate to potential effects on a variety of
species (receptors) included under these taxonomic groupings.

Acute and chronic toxicity data from studies submitted by pesticide
registrants are used to evaluate the potential direct effects of
metrafenone to the aquatic and terrestrial receptors identified in this
section.  This includes toxicity data on the technical grade active
ingredient, any major transformation products, and when available,
formulated products (e.g. “Six-Pack” studies).

Table 6 provides a summary of the taxonomic groups and the surrogate
species tested to help understand potential acute ecological effects of
pesticides to these non-target taxonomic groups.  In addition, the table
provides a preliminary overview of the potential acute toxicity of
metrafenone by providing the acute toxicity classifications.

Table 6.  Test Species Evaluated for Assessing Potential Ecological
Effects of Metrafenone and the Associated Acute Toxicity Classification

Taxonomic Group	Surrogate Species	Acute Toxicity Classification

Birds1	  SEQ CHAPTER \h \r 1 Mallard Duck (Anas platyrhynchos)

Bobwhite Quail (Colinus virginianus)

Passerine species	Practically nontoxic

Practically nontoxic

No available study

  SEQ CHAPTER \h \r 1 Mammals

	  SEQ CHAPTER \h \r 1 Laboratory rat (Rattus norvegicus)

Laboratory mouse	Practically non-toxic

Practically non-toxic

  SEQ CHAPTER \h \r 1 Insects	  SEQ CHAPTER \h \r 1 Honey bee (Apis
mellifera L.)	Practically non-toxic

  SEQ CHAPTER \h \r 1 Freshwater fish2		  SEQ CHAPTER \h \r 1 Bluegill
sunfish (Lepomis macrochirus)

Rainbow trout (Oncorhynchus mykiss)	Highly toxic

Highly toxic

  SEQ CHAPTER \h \r 1 Freshwater invertebrates	  SEQ CHAPTER \h \r 1
Water flea (Daphnia magna)	At most, highly toxic

  SEQ CHAPTER \h \r 1 Marine/estuarine fish	  SEQ CHAPTER \h \r 1
Sheepshead minnow (Cyprinodon variegatus)	Highly toxic

Marine/estuarine invertebrates	Mysid shrimp (Americamysis bahia)

Eastern oyster (Crassostrea virginica)	Highly toxic

Highly toxic

  SEQ CHAPTER \h \r 1 Terrestrial plants3	  SEQ CHAPTER \h \r 1 Monocots
– most sensitive species

Dicots – most sensitive species	No Classification

  SEQ CHAPTER \h \r 1 Aquatic plants and algae	  SEQ CHAPTER \h \r 1
Duckweed (Lemna gibba)

Cyanobacteria/blue-green algae (Anabaena flos-aquae)

Marine diatom (Skeletonema costatum)

Freshwater diatom (Navicula pelliculosa)

Algae (Pseudokirchneriella subcapitata; previously known as Selenastrum
capricornutum)	No classification

  SEQ CHAPTER \h \r 1 1 Birds represent surrogates for terrestrial-phase
amphibians and reptiles.

2 Freshwater fish may be surrogates for aquatic-phase amphibians.

3 Normally four species of two families of monocots, of which one is
corn; six species of at least four dicot families, of which one is
soybeans.

Ecosystems Potentially at Risk

The ecosystems at risk are often extensive in scope, and as a result it
may not be possible to identify specific ecosystems during the
development of a baseline risk assessment.  However, in general terms,
terrestrial ecosystems potentially at risk could include the treated
field and areas immediately adjacent to the treated field that may
receive drift or runoff.  Areas adjacent to the treated field could
include cultivated fields, fencerows and hedgerows, meadows, fallow
fields or grasslands, woodlands, riparian habitats, and other
uncultivated areas.

Aquatic ecosystems potentially at risk might include but are not
necessarily limited to water bodies adjacent to, or down stream from,
the treated field and might include impounded bodies such as ponds,
lakes and reservoirs, or flowing waterways such as streams or rivers.
For uses in coastal areas, aquatic habitat also includes marine
ecosystems, including estuaries.

Assessment Endpoints

Assessment endpoints represent the actual environmental value that is to
be protected, defined by an ecological entity (species, community, or
other entity) and its attribute or characteristics (EPA, 1998).
Generally, the ecological entities may include the following: freshwater
as well as marine/estuarine fish and invertebrates, aquatic and
terrestrial plants, birds, reptiles, amphibians, mammals, and non-target
insects. For metrafenone: chronic risk to mammals is expected. The
attributes for each of these entities may include growth, reproduction,
and survival.

Conceptual Model

A conceptual model provides a written description and visual
representation of the predicted relationships between metrafenone
residues, potential routes of exposure, and the predicted effects for
the assessment endpoint. A conceptual model consists of two major
components: risk hypothesis and a conceptual diagram (EPA, 1998).

Risk Hypothesis

For a pesticide to pose an ecological risk, it must reach ecological
non-target organisms (receptors) at biologically significant
concentrations. An exposure pathway is the means by which a pesticide
moves in the environment from the application site to non-target
organisms. The evaluation of the ecological exposure pathways in this
assessment includes an examination of the source and potential transport
pathways for metrafenone and the determination of exposure routes of
non-target species.

Metrafenone, when used in accordance with the label, results in
potential direct adverse effects upon the growth and reproduction of
mammals.  As a result, given the persistence of metrafenone, there is
potential for indirect effects to terrestrial and/or aquatic organisms.

Conceptual Diagram

The conceptual model diagram is a generic graphic depiction of the risk
hypotheses identified in the previous section.  It is assumed that
metrafenone is capable of affecting exposed terrestrial and aquatic
organisms if environmental concentrations are sufficiently elevated as a
result of proposed label uses.  Through a preliminary process of
examining fate and effects data, the risk hypotheses and conceptual
model have been refined to reflect possible exposure pathways and the
organisms that are most relevant and applicable to this assessment
(Figure 2).  If exposed at sufficient levels, mortality may occur, as
well as sublethal effects.  Direct effects on a taxonomic group may
result in indirect effects (i.e., loss of habitat, food resources) to
other taxonomic groups.  This assessment will examine the potential for
these effects to occur within the surrogate taxa with the intent to
extrapolate to actual effects within the environment.

In order for a chemical to pose an ecological risk, it must reach
ecological receptors in biologically significant concentrations.  An
exposure pathway is the means by which a contaminant moves in the
environment from a source to an ecological receptor.  For this pathway
to be complete, it must have a source, a release mechanism, an
environmental transport medium for metrafenone and/or its transformation
products, a point of exposure for ecological receptors, and a feasible
route of exposure.  The assessment of these pathways thus includes an
examination of the sources and potential migration pathways for
constituents, and the determination of potential exposure routes.

The conceptual model for both potential aquatic and terrestrial risk is
shown in Figure 2.  Exposure routes shown in dashed lines are not
quantitatively considered because the resulting exposures are expected
to be very low when compared to the major routes of exposure.

Exposure is expected to be dominated by runoff and spray drift.
Metrafenone exposure may also result from off-site movement in runoff
water and on metrafenone -bearing soil particulates to adjacent fields
(soil erosion). The moderate Koc’s (Koc 1073 to 22517 L/kg-oc) of
metrafenone suggest that leaching to ground water/recharge to surface
water would not be a route of exposure. Long-range transport of
metrafenone in the gas phase is not considered a significant route of
exposure.

This screening-level assessment for ground spray applications of
metrafenone only considered dietary exposure. Other routes of exposure
that were not considered in the assessment are incidental soil ingestion
exposure, inhalation exposure, dermal exposure, and drinking water
exposure.  These routes are not represented in the diagram (Figure 2).

Figure 2.  Conceptual Model Depicting Sources of Exposure from
Metrafenone as well as Metrafenone Residues, Potential Receptors, and
Adverse Effects from the Proposed Uses of Metrafenone on Grapes.

Analysis Plan

The structure of this risk assessment is based on guidance contained in
U.S. EPA’s Guidance for Ecological Risk Assessment (U.S. EPA, 1998)
and is consistent with procedures and methodology outlined in the
Overview Document (U.S. EPA, 2004).

Conclusions from Previous Risk Assessments

There are no previous ecological risk assessments because this is the
first registration petition for metrafenone for use on grapes in the
United States.

Preliminary Identification of Data Gaps

Review of the submitted studies indicated the following points:

Environmental Fate

All but one of the environmental fate studies required to support the
registration of metrafenone for use on grapes were submitted.
Independent Laboratory Validations for the methods submitted for
analysis of metrafenone in soil and water remain a data gap. For a list
of submitted environmental fate studies for metrafenone see Appendix F.

All of the environmental fate studies for the parent compound were
determined to be scientifically valid and therefore results from all of
the studies can be used to characterize the mobility and rates of
transformation of metrafenone.  However, many of the metabolism studies
have major uncertainties in the identification and pattern of formation
and decline of transformation products.  In all of the aquatic
metabolism studies, between 57% and 65% of the applied radioactivity
remains unidentified with incomplete characterization, and in two
aerobic soil metabolism studies, 15% and 44% of the applied
radioactivity is unidentified.  This includes at least four major
degradates that individually reach levels of 11% to 35% of the applied.
Other transformation products appear as groups of up to 15 components,
in some cases characterized as each being <5% of the applied
radioactivity, but in other cases, some individual components make up 9%
to 10% of the applied.  Even when individual components can all be
classified as minor degradates, these groups represent such a large
portion of the applied radioactivity overall that the possibility that
they may have some impact as a group cannot be precluded despite their
lower individual levels.  This is especially true given that the
degradation pathways suggest that groups of degradates may have a high
degree of structural similarity and so may have similar fate and effects
behavior.  Without information to adequately characterize the
degradates, it may be necessary to assume that they are of equal
toxicity to the parents in order to quantify risks.

In addition to the studies submitted by the registrant, EFED was
provided with a Draft Assessment Report (DAR) on Metrafenone written by
the United Kingdom.  That assessment reports that the long dissipation
time observed in field studies triggered a requirement for longer term
field accumulation studies. Two years into those five year studies,
results show that metrafenone appears to be accumulating, and that
metrafenone residues have been detected as deep as 20-30 cm below the
surface. These studies would provide useful information regarding the
environmental fate of metrafenone and would assist in reducing
uncertainties.

Ecotoxicity

The submitted ecotoxicity database is incomplete. The greatest amount of
uncertainty in the assessment stems from aquatic studies which were
largely based on total (both dissolved and undissolved) concentrations
of test compound; that is, the concentrations were measured without
centrifugation even though precipitate was observed or test was
conducted at the solubility limit of the metrafenone technical grade
active ingredient; the solubility limits of the metabolites are unknown.
To reduce uncertainty the majority of the toxicity studies, which are
considered supplemental, and cannot be used in a quantitative risk
estimation would need to be redone; especially those where mortality or
sublethal effects were observed. These aquatic studies are specified
below. For a list of submitted ecological effects studies for
metrafenone see Appendix G.

The following studies are considered data needs:

	Aquatic studies

Given the freshwater fish studies on metrafenone technical grade active
ingredient and assuming that the concentrations in the environment reach
the solubility limit, the effect of the compound is likely to be low.
However, according to model estimated EECs (which include metrafenone
and metrafenone residue scenarios: 0.00153 - 0.02 mg/L), levels of
metrafenone (TGAI, TEP) at the solubility limit (0.2-0.5 mg/L at 12oC)
and metrafenone metabolites at the tested concentrations are not
expected to occur in the environment given the proposed grape use.
Therefore, acute risk to freshwater fish and aquatic-phase amphibians is
not expected as a result of metrafenone use on grapes. In addition,
although the EU/UK formulation – with which the submitted studies were
conducted – closely matches the U.S. formulation (i.e., BAS 560 03F)
it is still not its equivalent. Therefore, the effect of the U.S.
formulated product on freshwater fish and aquatic-phase amphibians is
not known.

To reduce uncertainty in characterizing risk, which is currently based
on total concentrations, the freshwater fish studies on bluegill sunfish
(Lepomis macrochirus) are requested to make same species comparisons for
acute and chronic values. Bluegill was chosen over the rainbow trout due
to greater sensitivity (on an acute basis) of the bluegill to the TGAI
relative to the trout. Since the acute marine/estuarine fish study is
acceptable and given the above two studies, the acute to chronic ratio
can be utilized to estimate marine/estuarine chronic endpoints, negating
the need to fill this data gap. Special emphasis must be placed on
centrifuging the test samples prior to analytical determination of the
test compound.

 Acute: Freshwater fish toxicity (96-hour LC50 for Lepomis macrochirus)
(850.1075; 72-1), TGAI

 Chronic: Fish early-life stage (freshwater: Lepomis macrochirus)
(850.1400; 72-4), TGAI

Although acute freshwater invertebrate studies were based on total
concentrations, no effects were observed with the TGAI, negating the
need to request additional data. However, a chronic freshwater
invertebrate data gap still exists since the two reviewed studies were
deemed invalid.

Chronic: Aquatic invertebrate life cycle (freshwater: Daphnia magna)
(850.1300; 72-4), TGAI

Effects were observed in both acute marine/estuarine invertebrate
studies with the TGAI on the eastern oyster (Crassostrea viginica) and
saltwater mysid (Americamysis bahia) but the endpoints are based on
total concentrations. Therefore, should concentrations in the
environment reach the solubility limit, the acute risk to
marine/estuarine invertebrates may be expected. However, according to
the model estimated EECs (0.00153 - 0.02 mg/L, which includes
metrafenone and total metrafenone residue scenarios), levels of
metrafenone at the solubility limit are not expected to occur in the
environment given the proposed grape use. Given these studies and
assuming that metrafenone concentrations in the environment are not
likely to reach the solubility limit, the acute risk to marine/estuarine
invertebrates is not expected. However, to reduce uncertainty in
characterizing risk, either study is requested to be redone with special
emphasis on centrifuging the test samples prior to analytical
determination of the test compound.

Acute: Marine/estuarine invertebrate either Crassostrea virginica
(850.1025; 72-3) or Americamysis bahia (850.1035; 72-3), TGAI

A non-guideline chronic midge study (Chironomus riparius) with
metrafenone technical (97.1%) was deemed invalid on the basis of low
negative control emergence. Therefore, a chronic freshwater invertebrate
whole sediment study is requested. A protocol should be submitted for
approval prior to study initiation. The protocol should include spiked
sediment. The whole sediment study is requested because metrafenone has
a high logKow value (4.3 at 25oC, pH 4) and a high aerobic soil half
life (> 4 months), which indicates that the compound may partition to
sediment and persist.

Chironomus dilutus (freshwater) using TGAI. Consult EPA Test Method
100.5 Life-cycle Test for Measuring the Effects of Sediment Associated
Contaminants on Chironomus dilutus (formerly Chironomus tentans) and
OECD Guideline 218 Sediment-Water Chironomid Toxicity Test Using Spiked
Sediment

Non-target aquatic plant studies were also based on total concentrations
leading to uncertainty in exposure concentrations and thus endpoint
values. Had valid endpoints (i.e., those based on dissolved
concentrations) been determined using these studies, they would likely
be greater than the highest model predicted concentration in the
environment, which implies that risk to vascular and non-vascular
species is not expected as a result of metrafenone use on grapes.
However, without centrifugation, the amount of chemical that is freely
dissolved and bioavailable cannot accurately be determined. Therefore,
vascular and non-vascular aquatic plant studies are requested with
special emphasis placed on centrifuging the test samples prior to
analytical determination of the test compound. In addition,
cyanobacteria yielded no effects given total concentrations and does not
need to be redone.

Aquatic vascular plant growth (Lemna spp.), Tier II (850.4400), TGAI

Aquatic non-vascular plant growth, Tier II (850.5400), TGAI

     Terrestrial Studies

An avian acute oral toxicity test in passerine species is required.
Passerines are the most common birds (in terms of numbers and number of
species) in the United States.  Many utilize agricultural fields,
forests, residential areas and surrounding areas, and, therefore, have
the potential to be exposed to pesticides used in agricultural, forest,
and residential settings.  It is likely that, for the requested use
patterns, passerines are more likely to be exposed to metrafenone than
upland game species and waterfowl.  Passerines are smaller and have a
higher energy requirement than larger-sized birds.  As such, passerines
may be more sensitive than other birds.

Passerine bird toxicity study (EPA approved protocol is required prior
to study initiation): Avian acute oral toxicity test (850.2100, 71-1),
TGAI

Formulated Product Testing

The available formulation product studies were conducted using three
different UK/EU formulated products (i.e., BAS 560 00F, BAS 560 01F, and
BAS 560 02F), one of which (BAS 560 00F) closely matches the U.S.
formulation (i.e., BAS 560 03F). Formulation studies in this assessment
are based on BAS 560 00F because data on the U.S. formulation were not
submitted. In order to eliminate uncertainty in effects characterization
of formulated products used within the U.S. on given taxa, future
registrant submitted studies should be based on the U.S. formulation. At
this time the effect of the U.S. formulation on given taxa is not known,
however, greater toxicity of the EU/UK formulation on the honeybee
relative to the TGAI was observed. Similarly, data submitted for
terrestrial plants is based on the EU/UK formulation. As a result, the
following studies are requested:

Honeybee acute contact toxicity (850.3020, 141-1), TEP

Seedling emergence, Tier II (850.4225, 123-1), TEP

Vegetative vigor, Tier II (850.4250, 123-1), TEP

Measures of Exposure and Effects

EFED uses a tiered system of pesticide exposure modeling to assess
ecological risk following a registered application of that pesticide.
This tiered system is designed to minimize the amount of analysis which
is required to register any given chemical.  Each of the tiers is
designed to screen out pesticides by requiring higher, more complex
levels of investigation only for those that have not passed the next
lower tier.  Each tier screens out a percentage of pesticides from
having to undergo a more rigorous review prior to registration or
re-evaluation.

Aquatic Exposure Models

Tier II PRZM and EXAMS simulation models were used to estimate the
exposure concentrations of metrafenone in surface water for the proposed
use on grapes. The results are presented in Appendix B.

The data used as input parameters come solely from the environmental
fate studies and proposed product label submitted by the petitioner to
support in the United States the registration of metrafenone as a new
pesticide-active ingredient for grapes.

KABAM (KOW (based) Aquatic BioAccumulation Model) v.1.0 is used to
estimate potential bioaccumulation of hydrophobic organic pesticides
such as metrafenone (log Kow > 4) in freshwater aquatic food webs and
subsequent risks to mammals and birds via consumption of contaminated
aquatic prey. The bioaccumulation portion of KABAM is based upon work by
Arnot and Gobas (2004) who parameterized a bioaccumulation model based
on PCBs and some pesticides (e.g., lindane, DDT) in freshwater aquatic
ecosystems. KABAM relies on a chemical's octanol-water partition
coefficient (KOW) to estimate uptake and elimination constants through
respiration and diet of organisms in different trophic levels. Pesticide
tissue residues are calculated for different levels of an aquatic food
web. The model then uses pesticide tissue concentrations in aquatic
animals to estimate dose- and dietary-based exposures and associated
risks to mammals and birds consuming aquatic organisms, using an
approach that is similar to the T-REX model (USEPA 2008).

KABAM incorporates 7 trophic levels to describe bioaccumulation of a
pesticide in a model aquatic food web: phytoplankton, zooplankton (e.g.,
Daphnia sp.), benthic invertebrates (e.g., Chironomus sp., crayfish),
filter feeders (e.g., mussels, clams), small fish (e.g., young of the
year), medium sized fish (e.g., adult bluegill), and larger
upper-trophic level fish (e.g., largemouth bass).

Metrafenone concentrations in organisms of the aquatic trophic levels
listed above were used to estimate acute and chronic exposures of
piscivorous mammals and birds consuming fish. Applicable and available
acute and chronic toxicity data on metrafenone (mammals and birds) were
used to calculate risk quotients for estimated exposures due to
bioaccumulation of metrafenone in an aquatic ecosystem.

Terrestrial Exposure Models

T-REX Model

The focus of terrestrial wildlife exposure estimates is for birds (also
acting as surrogate for reptiles and terrestrial-phase amphibians) and
mammals with an exposure route emphasis on uptake through the diet.  The
residues in or on potential dietary sources for mammals and birds (e.g.,
vegetation, insects, and seeds) were estimated using the Tier I model
T-REX (Version 1.4.1, 2008).  In this Tier I assessment, it was assumed
that organisms are exposed to one active ingredient in a given exposure
scenario.  In all screening-level assessments, the organisms are assumed
to consume 100% of their diet as one food type and one food source.  The
T-REX output is presented in the Risk Characterization section of this
document as well as an example in Appendix C.

The approach used to estimate exposure of terrestrial animals to
metrafenone was based on potential foliar applications of metrafenone.
Upper-bound exposure levels were calculated for spray applications of
metrafenone using maximum proposed application rates for one application
for the proposed uses. The exposure estimates are based on a database of
pesticide residues on wildlife food sources associated with specified
application rates (Kenaga, 1972; Fletcher et al., 1994). Essentially,
for a single application, there is a linear relationship between the
amount of pesticide applied and the amount of pesticide residue present
on a given food item.  Food item residue levels are then linearly
adjusted based on application rate.  The upper-bound estimates are used
to estimate risks since these values represent the high-end exposure
that may be encountered for terrestrial species that consume food items
that have received label-specified pesticide application. Although these
represent higher-end estimates, they do not represent the highest
possible exposure estimates.

T-REX is a simulation model that, in addition to incorporating the
relationship between application rate and food item residue
concentrations, accounts for pesticide degradation in the estimation of
terrestrial EECs. T-REX calculates pesticide residues on each type of
food item on a daily interval for one year. A first-order decay function
is used to calculate the residue concentration at each day based on the
concentrations present from both initial and all subsequent
applications.  The decay rate is dependent on the foliar dissipation
half-life. The food item concentration on any given day is the sum of
all residues up to that day, taking into account the first-order
degradation. The initial application occurs on day 0 (t=0) and the model
runs for 365 days.  Over the 365-day run, the highest residue
concentration is the measure of exposure (EEC) used to calculate risk
quotients (RQs).

The foliar dissipation half-life and residue decline studies can be
important in estimating exposure because they essentially determine how
long the pesticide remains in or on food items after application. In
many cases, neither empirically determined foliar dissipation nor
residue decline half-life (with a day 0 residue) values are available,
in which case the default value of 35 days is used (Willis and McDowell,
1987). That was the case for this assessment.

TERRPLANT Model

The TerrPlant (Ver.1.2.2) model is used to predict EECs from terrestrial
uses for terrestrial plants located in dry and semi-aquatic areas
adjacent to the treated field or treated water body.  TerrPlant assumes
100% efficiency in ground and aerial applications. A semi-aquatic area
(wetland) is defined as a low-lying area of terrestrial habitat that is
wet but may dry up at times throughout the year.  TerrPlant incorporates
two similar conceptual models for depicting dry and semi-aquatic areas
of terrestrial habitats.  For both models, a non-target area is adjacent
to the target area. Pesticide exposures to plants in the non-target area
are estimated to receive runoff and spray drift from the target area.

For a dry area adjacent to the treatment area, runoff exposure is
estimated as sheet runoff. Sheet runoff is the amount of pesticide in
water that runs off of the soil surface of a target area of land which
is equal in size to the non-target area (1:1 ratio of areas).  In the
sheet runoff scenario, the treated area generating runoff is assumed to
drain into an area with equal size containing seedlings, resulting in 1,
2, or 5% of the application rate being deposited.  For semi-aquatic
areas, runoff exposure is estimated as channel runoff.  Channel runoff
is the amount of pesticide that runs off of a target area 10 times the
size of the non-target area (10:1 ratio of areas).  In the channel
runoff scenario, a ten-to-one ratio of watershed area to receiving area
results in 10, 20, or 50% of the application rate being deposited on
soil with emerging or emerged seedlings.  The magnitude of runoff is
assumed to be dependent on the water solubility of the pesticide active
ingredient.  For pesticides with a solubility of <10, 10 to 100, or >100
ppm, runoff fractions of 0.01, 0.02 or 0.05 respectively are selected by
the model user.

Exposures through runoff and spray drift are then compared to measures
of survival and growth (e.g. effects to seedling emergence and
vegetative vigor) to develop RQ values.

The model compares the combined deposition estimates from runoff and
spray drift to adverse effect levels measured in seedling emergence
studies.  In addition, RQs are derived for plants with consideration for
spray drift exposures.  For monocots and for dicots, TerrPlant compares
estimated spray drift deposition, without a runoff exposure component,
to the more sensitive measure of effect, either seedling emergence or
vegetative vigor (USEPA 2005).

Table 7 summarizes the measures of ecological effects and exposure used
to assess ecological risk following exposure to metrafenone with the
proposed uses.

Table 7.  Measures of Ecological Effects and Exposure for Metrafenone

Assessment Endpoint	Surrogate Species and Measures of Ecological
Effect1,2	Measures of Exposure

Birds3	Survival	Bobwhite Quail acute oral LD50, subacute dietary LC50

Mallard Duck acute oral LD50,  subacute dietary LC50

Upper bound residues on food items (foliar)

	Reproduction and growth	Bobwhite Quail reproduction NOAEC/LOAEC

	Mammals	Survival	Laboratory rat acute oral LD50

Laboratory mouse acute oral LD50

Reproduction and growth	Laboratory rat reproduction NOAEL/LOAEL

	Freshwater fish4	Survival	Rainbow trout 96-hr LC50

Bluegill sunfish 96-hr LC50	Peak EEC5

	Reproduction and growth	Fathead minnow NOAEC/LOAEC	60-day average EEC5

Freshwater invertebrates	Survival	Water flea 48-hr EC50	Peak EEC5

	Reproduction and growth	No acceptable study available	21-day average
EEC5

Marine/estuarine fish	Survival	Sheepshead minnow 96-hr LC50	Peak EEC5

	Reproduction and growth	No study available	60-day average EEC5

Marine/estuarine invertebrates	Survival	Eastern oyster 96-hr EC50

Saltwater mysid 96-hr LC50	Peak EEC5

	Reproduction and growth	Saltwater mysid NOAEC/LOAEC	21-day average EEC5

Terrestrial plants6	Survival and growth	Monocot Seedling emergence

EC25, NOAEC or EC05

Monocot Vegetative Vigor

EC25, NOAEC or EC05

Dicot Seedling emergence

EC25, NOAEC or EC05

Dicot Vegetative Vigor

EC25, NOAEC or EC05	Estimates of runoff and spray drift to non-target
areas

Insects	Survival 	Honey bee acute contact 48-hr LD50	Maximum application
rate

Aquatic plants and algae	Survival and growth	Duckweed 7-day EC50, NOAEC

Cyanobacteria 96-hr EC50, NOAEC

Marine diatom 96-hr EC50, NOAEC

Freshwater diatom 96-hr EC50, NOAEC

Green algae 96-hr EC50,  NOAEC

Algae 72-hr EC50, NOAEC

Green algae 72-hr EC50, NOAEC	Peak EEC5

1LD50 = Lethal dose to 50% of the test population; NOAEC = No observed
adverse effect concentration; LOAEC = Lowest observed adverse effect
concentration; LC50 = Lethal concentration to 50% of the test
population; EC50/EC25 = Effect concentration to 50%/25% of the test
population.

2 If species listed in this table represent most commonly encountered
species from registrant-submitted studies, risk assessment guidance
indicates most sensitive species tested within taxonomic group are to be
used for baseline risk assessments.

3 Birds represent surrogates for amphibians (terrestrial phase) and
reptiles.

4 Freshwater fish may be surrogates for amphibians (aquatic phase).

5 One in 10-year return frequency.

6 Four species of two families of monocots - one is corn, six species of
at least four dicot families, of which one is soybeans.

Analysis

Use Characterization

The proposed end-use product is “Metrafenone 300 SC” (EPA Reg. No.
7969-XXX), a suspension concentrate containing 25.2%
(3-bromo-6-methoxy-2-methylphenyl)(2,3,4-trimethoxy-6-methylphenyl)metha
none (metrafenone) at a concentration of 2.5 lbs metrafenone per gallon.
 The product is claimed to provide “optimum disease control when
applied in a regularly scheduled protective fungicide program and when
used in a spray program that rotates fungicides with different modes of
action.” The product is intended to control powdery mildew of grapes.

Ground application is the proposed method of application.  The label
states, “do not apply directly to water, to areas where surface water
is present, or to intertidal areas below the mean high water mark.”
The chemical is to be applied at most 6 times per season at a maximum
application rate of 0.3 lbs a.i./A (Table 5). The label also states,
“do not make more than 2 sequential applications of Metrafenone 300 SC
fungicide before alternating to a labeled fungicide with a different
mode of action.”

Exposure Characterization

Environmental Fate and Transport

Summary

Metrafenone is a new chemical of the benzophenone class that is expected
to be slightly mobile.  Its major routes of degradation are aqueous
photolysis and aquatic metabolism, in both aerobic and anaerobic
conditions.  Soil metabolism is slow, with laboratory half-lives of 6
months to a year and accumulation over two years observed in field
studies.  In aquatic environments, the measured photolysis half-life was
6.4 days and metabolism in both aerobic and anaerobic occurred with
half-lives of one to four weeks.

Soil photolysis and anaerobic aquatic metabolism of metrafenone led to
one major degradate, CL 377160 [Methanone,
(3-bromo-6-methoxy-2-methylphenyl)(3-hydroxy-2,4-dimethoxy-6-methylpheny
l)-]., and anaerobic aquatic metabolism also led to six other major
degradates, but all were either unidentified or only tentatively
identified.  In addition, there were many minor degradates, which were
formed at low levels individually but reached substantial amounts as
groups.

μg/L for the daily peak, 8.82 μg/L for the 21 day average, and 6.21
μg/L for the 60 day average. The highest aquatic EECs for metrafenone
were 20.22 μg/L for the daily peak, 17.67 μg/L for the 21 day average,
and 16.98 μg/L for the 60 day average.

No monitoring data are available to compare with model estimates.
Bioaccumulation modeling was conducted because metrafenone has a log
octanol:water coefficient > 4.  Maximum residue concentrations in fish
are expected to range from 17,636 to 19,586 µg/kg-ww.

Persistence

Metrafenone biodegrades slowly in terrestrial environments.  In
laboratory studies in a variety of soils, aerobic soil metabolism linear
half-lives ranged from 178 to 365 days.  In four of the eight soils
tested, data collection extended long enough to determine an empirical
DT50, of 124 to 188 days in two silt loam soils and 272 to 362 days in a
clay loam and sandy loam soil.  In these two soils, metabolism appeared
to slow after 272 days.  No data are available for anaerobic soil
metabolism rates.  Photolysis on soil was observed in laboratory studies
to occur with a linear half-life of 31 days.

In aquatic environments, metrafenone biodegrades in weeks to months in
both aerobic and anaerobic conditions (linear half-lives of 24-27 days
and 18-22 days, respectively).  Considering abiotic degradation,
metrafenone is photolyzed in water (half-life of 6.4 days), but is
stable to hydrolysis at pH values from 4 to 9.

Degradation Products

The only positively identified major degradate of metrafenone was CL
377160 [Methanone,
(3-bromo-6-methoxy-2-methylphenyl)(3-hydroxy-2,4-dimethoxy-6-methylpheny
l)-].  CL 377160 was formed through soil photolysis at up to 18.9% AR
and through anaerobic aquatic metabolism at up to 10.8% AR in the total
system. It was also formed as a minor degradate through aquatic
photolysis and aerobic aquatic metabolism.   A compilation of identified
degradation products are shown in Appendix A.

In one anaerobic aquatic metabolism study, another six degradates were
formed at >9%, but these were either unidentified or only tentatively
identified.  These included TN (tentatively identified as a lactone
compound; maximum 3.3%, 26.9% and 28.0% in the water layer, sediment and
total system, respectively), MB (tentatively identified as a
hydroxylation product of TN; maximum 1.1%, 11.6% and 11.7% in the water
layer, sediment and total system, respectively), and four unidentified
degradates formed in the total system at 9.3% to 13% AR.

Carbon dioxide was also a major degradate, formed through aquatic
photolysis at up to 25% AR and through all other measured processes as a
minor degradate.

Most processes also resulted in formation of a substantial amount of
minor degradates, some individually measured and identified, but most
not.  Although all of these degradates were present at less than 10%,
and some at very low levels, they still may be of exposure concern
because of the large overall amounts.  In some cases minor degradates,
mostly unidentified, make up more than 50% of the applied radioactivity
in a single sampling event.  In addition, based on the structure of the
parent compound and the proposed degradation pathways, many of the minor
degradates may have a high degree of structural similarity to each other
and/or the parent and so may have cumulative effects.

Mobility

Batch equilibrium data on metrafenone suggest that the compound will
sorb to organic surfaces and would be considered “slightly mobile”
according to the FAO classification scheme (organic carbon partition
coefficients range from 1,073 to 2,792 ml/g-oc).  Also according to FAO
classification, metrafenone’s solubility in water of 0.447 mg/L is
considered “slightly soluble.”  It showed low volatility from soil
and water with <0.5% volatilization in any laboratory study, consistent
with the vapor pressure of 1.15 x 10-6 mmHg.  Therefore, dissipation in
the environment is expected to occur predominantly via runoff of
suspended soil and sediments to which metrafenone is adsorbed.  Based on
terrestrial field dissipation studies in the UK , metrafenone was
detected at a soil depth of 20-30 cm after 2 years.

In aquatic systems, partitioning of metrafenone to sediment occurred
rapidly.  In one anaerobic system, half of the applied metrafenone was
detected in sediment on the first day, and in three other systems (one
anaerobic, two aerobic), at least 50% of the parent compound had
partitioned to sediment within three to seven days.  Including
degradates, partitioning was rapid in anaerobic systems but slower in
aerobic systems.  In two anaerobic aquatic metabolism studies, more than
half of the applied radioactivity (AR) was found in sediment within one
week and radioactivity in the water layer was down to <5% AR in two to
six weeks.  However, in aerobic aquatic metabolism studies in two
systems, from 8% to 28% AR remained in the water layer at study
termination (100 days).

Field Dissipation Studies

Field dissipation studies were conducted on bare plots in California
(CA), Washington (WA), Ontario (ON), and Florida (FL).  The first-order
dissipation half-life for metrafenone in surface soils (0-7.5 cm) was
144 days (DT 50 14-30 days; DT 90 272-360 days) in CA, 161.2 days (DT 50
181-269 days; DT 90 540-900 days) in WA, 210 days (DT 50 59-272 days; DT
90 598-710 days) in ON, and 161 days (DT 50 14-30 days; DT 90 360-451
days) in FL.  No degradation products were detected above the limit of
quantification (LOQ) in the field dissipation studies.

The reference degradation product in the field study was CL377160.
Degradation was not a clear route of metrafenone dissipation in field
studies because of the lack of detectable degradation products in soil.
Leaching is possible route of dissipation. There were sporadic
detections of metrafenone with soil depth in the Florida study with
maximum detection depth of 15-30 cm, Ontario study with maximum
detection depth of 60-75 cm, Washington study with a maximum depth depth
of  7.5-15 cm, and  California study with a maximum detection depth of
15-30 cm. The field studies were not designed to assess runoff and
volatilization of metrafenone residues.

Bioaccumulation in Fish

Bluegill sunfish exposed to metrafenone, at 5 and 50 µg/L, accumulated
metrafenone residues during a 28 day accumulation period. After the
accumulation phase, the fish were transferred to metrafenone-free water
for a 14 day depuration phase.  For the bromophenyl labeled metrafenone,
lipid normalized bioconcentration factors (BCFs) ranged 140 to 460 in
whole fish.  For trimethoxybenzene labeled metrafenone, lipid normalized
bioconcentration factors (BCFs) ranged 490 to 530 in whole fish. The
lipid normalized BCF range indicates that metrafenone is not expected to
accumulate in tissues of aquatic organisms. Major bioaccumulated
residues (>10% of TRR) were identified as
[(3-Bromo-6-methoxy-2-methylphenyl)(4-hydroxy-2,3-dimethoxy-6-ethylpheny
l)methanone (CL 434223),  CL1500699 (3-
bromo-6-methoxy-2-methylphenyl)[4-(beta-D-gluco pyran
uronosyloxy)-2,3-dimethoxy-6-methyphenyl]-methanone), and CL377160.
These degradation products were detected in whole fish, viscera, fillet,
and in the exposure water.  The bioaccumulation rate at 90% steady-state
of total radioactive residue and metrafenone ranged from 1.3 to 1.8 days
and 1.8 to 2.1 days, respectively.  The 95% depuration rate of total
radioactive residue and metrafenone ranged from 1.8 to 2.3 days and 1.8
to 2.1 days, respectively.

Measures of Aquatic Exposure

Aquatic Exposure Modeling

The estimated environmental concentrations (EECs) reported in the
assessment were calculated using the Tier II model for surface water
(PRZM/EXAMS). Sample inputs and outputs of the model are presented in
Appendix B.

PRZM (v3.12.2, May 2005) and EXAMS (v<IP_ADDRESS>, April 2005) are screening
simulation models coupled with the input shell pe5.pl (Aug 2007) to
generate daily exposures and 1-in-10 year EECs of metrafenone and total
metrafenone residues that may occur in surface water bodies adjacent to
application sites receiving metrafenone residues through runoff and
spray drift for specific scenarios.  PRZM simulates pesticide
application, movement and transformation on an agricultural field and
the resultant pesticide loadings to a receiving water body via runoff,
erosion and spray drift.  EXAMS simulates the fate of the pesticide and
resulting concentrations in the water body.  The standard scenario used
for ecological pesticide assessments assumes application to a 10-hectare
agricultural field that drains into an adjacent 1-hectare water body,
2-meters deep (20,000 m3 volume) with no outlet.  PRZM/EXAMS was used to
estimate screening-level exposure of aquatic organisms to metrafenone
residues. The measure of exposure for aquatic species is the 1-in-10
year return peak or rolling mean concentration.  The 1-in-10 year peak
is used for estimating acute exposures of direct effects to aquatic
organisms as well as indirect effects. The 1-in-10-year 60-day mean
concentration is used for assessing chronic exposure. The 1-in-10 year
21-day mean concentration is used for assessing chronic exposures to
aquatic invertebrates.

Input Parameters

The appropriate input parameters were selected from the
physical/chemical properties and environmental fate data submitted by
the petitioner to support registration of metrafenone.  Input parameters
were selected in accordance with US EPA-OPP EFED water model parameter
selection guidelines, Guidance for Selecting Input Parameters in
Modeling the Environmental Fate and Transport of Pesticides, Version
2.1, November 10, 2009.   Expanded information about the models,
selection of input parameters and scenarios can be obtained from
http://www.epa.gov /oppefed1/models/water/index.htm.

The use input parameters were selected from label for the end-use
product “Metrafenone 300 SC.”  The physical-chemical properties and
environmental fate input parameters were obtained from studies submitted
and reviewed by the Agency.

Total Residues Modeling Input Parameters

Calculation of total residue half-lives for metrafenone are based on the
concentration (expressed as percent of applied radioactivity) of
extractable residues (Table 8).  Extractable residues include
metrafenone, CL 377160, tentatively identified compounds, and
unidentified compounds (Appendix A).

Table 8. Total Extractable Metrafenone Residue Half-lives from
Laboratory Degradation Studies

Study Type	MRID1	Half-life (days)	Environmental Matrix	Extractable
Residues

Aqueous Photolysis	-23	27

Total Mass Balance - VOC – CO2

Soil Photolysis	-24	47.8

Acetonitrile, acetonitrile:0.2NHCl, 0.5 N NaOH

Aerobic Soil	-27	169.1	sandy loam soil	Acetonitrile,
acetonitrile:acetone, acetonitrile:water

Aerobic Soil	-26	210.0	clay loam soil	Acetone, methanol:water

Aerobic Soil	-25	277.2	silty loam soil	Acetone, methanol :water,
acetonitrile:0.5NHCl

Aerobic Soil	-27	277.3	clay loam soil	Acetonitrile,
acetonitrile:acetone, acetonitrile:water

Aerobic Soil	-27	301.4	silt loam soil	Acetonitrile,
acetonitrile:acetone, acetonitrile:water

Aerobic Soil	-26	315.1	sandy loam soil	Acetone, methanol:water

Aerobic Soil	-27	330.1	silt loam soil	Acetonitrile,
acetonitrile:acetone, acetonitrile:water

Aerobic Soil	-26	364.8	loamy sand soil	Acetone, methanol:water

Anaerobic Aquatic	-29	182.0	DI water- silty clay loam	Water + Acetone,
methanol acetic acid, acetonitrile, 0.5N NaOH:acetonitrile, ammonium
hydroxide:acetonitrile

Anaerobic Aquatic	-31	577.6	pond-sandy loam	Water +
acetone:acetonitrile, acetonitrile:formic acid, acetonitrile;water
formic acid, acetonitrile:water:triethylamine, acetonitrile:0.05 M
sodium phosphate dibasic, 0.5N ammonium hydroxide

Aerobic Aquatic	-30	91.2	river-loam	Water + acetone, methanol:acetic
acid, methylene chloride, acetonitrile:0.5N NaOH

Aerobic Aquatic	-30	123.8	pond-sand	Total Mass Balance - VOC – CO2

1- Prefix MRID number is 472674-

Surface water

Scenarios used to run PRZM and EXAMS (Tier II) for the proposed use are
shown in Table 9.  Input parameters for modeling in Tier II are shown in
Table 10.

Table 9.  Scenarios used to estimate Metrafenone concentrations in
surface water.

Agricultural Commodity	Crop Scenario	Met File	Scenario Characterization

Grapes	NY

CA	W14860.dvf

W93193.dvf	Standard Scenarios for assessing pesticides use on grapes

Table 10.  PRZM/EXAMS Input Parameters for Metrafenone and Total
Extractable Metrafenone Residues

PARAMETER	Metrafenone	Total Extractable Metrafenone  Residues	COMMENT
SOURCE

Application Rate per Event

Lb a.i./A (kg a.i./ha)	0.3 lb a.i./A	0.3 lb a.i./A

Metrafenone 300 SC Label

No. of Applications

 (Interval)	6 (14 day)	6 (14 day)

Metrafenone 300 SC Label

CAM

(Chemical App. Method)	2	2	Broadcast spray	Metrafenone 300 SC Label

Depth of Incorporation	0	0	Default

	Spray Drift Fraction / Application Efficiency	0.01 / 0.99	0.01 / 0.99
Assume ground spray.	EFED Input Guidance

Aerobic Soil Metabolism t1/(	303 days	313 days	Upper 90th percentile of
the mean half-life (n=8)	MRID 47267425

MRID 47267426

MRID 47267427

Aerobic Aquatic Degradation  t1/(  	28.2 days	158 days	Upper 90th
percentile of the mean half-life (n=2) 	MRID 47267430

Anaerobic Aquatic Degradation  t1/(	25.3 days	989 days	Upper 90th
percentile of the mean half-life (n=2)	MRID 47267429

MRID 47267431

Aqueous Photolysis t1/(	6.4 days	27.0 days	Corrected for dark	MRID
47267423

Hydrolysis t1/(	0 days	0 days	Stable	MRID 47267422

Soil Partition Coefficient (Koc)	2188 mL/goc	2188 mL/goc	Average Koc
MRID 47267420

Molecular Weight	409 g/mole	409 g/mole

MRID 47267423

Water Solubility @ 25°C	0.474 mg/L	0.474 mg/L

MRID 47267423

Vapor Pressure	1.15 x 10-6 mmHg	1.15 x 10-6 mmHg

EPA Fact Sheet (9/06)

Tier II- PRZM/EXAMS

Tier II estimated environmental concentrations for metrafenone and total
metrafenone residues are shown in Table 11.

Table 11.  Estimated Exposure Concentrations for Metrafenone and Total
Metrafenone Residues from Surface Water.  Concentrations are in μg/L
(ppb)

Agricultural Commodity/Scenario

	1-in-10 yr Peak	1-in-10 yr 21-Day Average	1-in-10 yr 60-Day Average

Metrafenone

NY Grapes	11.70	8.82	6.21

CA Grapes	1.53	1.10	0.75

Total Metrafenone  Residues*

NY Grapes	20.22	17.67	16.98

CA Grapes	2.25	1.89	1.58

*Parent metrafenone + identified and unidentified extractable residues

b.	Aquatic Exposure Monitoring and Field Data

Monitoring data for metrafenone in surface water and ground water are
not available in the United States or Europe.

c.	Aquatic Bioaccumulation Assessment

Available data on the octanol-water partition coefficient (Kow) for
metrafenone indicates that this pesticide may accumulate in aquatic food
webs.  Because the Log Kow is > 4.0, KABAM v.1.0 was used to estimate
concentrations of metrafenone in tissues of aquatic organisms resulting
from bioaccumulation. Input parameters are provided in Table 12 and
estimated concentrations of metrafenone in fish are provided in Table
13.  Sample KABAM output is provided in Appendix D.

Table 12.  Input Parameters and Chemical Characteristics of Metrafenone
Used in KABAM

Characteristic	Value	Comments

Log KOW	4.3

	KOW	19953

	KOC   (L/kg OC)	2188	Input value used in PRZM/EXAMS to derive EECs.

Time to steady state (TS; days)	8	This value is calculated automatically
from the Log KOW value entered above.

Pore water EEC (µg/L)	16.8	Value generated by PRZM/EXAMS benthic file
for New York Grapes scenario for metrafenone total residue and high
runoff scenario.  PRZM/EXAMS EEC represents the freely dissolved
concentration of the pesticide in the pore water of the sediment.

The appropriate averaging period of the EEC is dependent on the specific
pesticide being modeled and is based on the time it takes for the
chemical to reach steady state. 21-day average concentration (EEC) was
used as averaging period closest to the time to steady state calculated
above.

Water Column EEC (µg/L)	17.7	Value generated by PRZM/EXAMS water column
file for New York Grapes scenario for metrafenone total residue and high
runoff scenario.  PRZM/EXAMS EEC represents the freely dissolved
concentration of the pesticide in the water column. 21-day average
concentration (EEC) was used as averaging period closest to the time to
steady state calculated above (as discussed above for pore water).

Table 13. Estimated Concentrations of Metrafenone in Fish (based on
metrafenone total residue scenario)

Ecosystem Component	Total concentration (µg/kg-ww)	Lipid normalized
concentration (µg/kg-lipid)	Contribution due to diet (µg/kg-ww)
Contribution due to respiration (µg/kg-ww)

Small Fish	17,636	440909	862.94	16,773.41

Medium Fish	18,305	457626	1,651.07	16,653.98

Large Fish	19,586	489661	3,063.04	16,523.40

Measures of Terrestrial Exposure

Terrestrial Exposure Modeling

T-REX

Exposure of free-ranging terrestrial animals is a function of the timing
and extent of pesticide application with respect to the location and
behavior of those species. OPP’s terrestrial exposure model generates
exposure estimates assuming that the animal is present on the use site
at the time that pesticide levels are highest. The upper-bound pesticide
residue concentration on food items is calculated from both initial
applications and any additional applications, taking into account
pesticide degradation between applications. Although this approach is
conservative, it is reasonable, particularly when considering acute
risks.  For acute risks, the assumption is that the duration of exposure
is a single day and, again, occurs when residue levels are highest.  In
evaluating chronic risks, longer-term exposure estimates are also based
on the assumption that the animal is present on the use site when
residue levels are highest and furthermore that it repeatedly forages on
the use site.

The current screening-level approach does not directly relate timing of
exposure to critical or sensitive population, community, or ecosystem
processes. Given that the application timing and location is
crop-dependent, it is difficult to address the temporal and spatial
co-occurrence of metrafenone use and sensitive ecological processes.
However, pesticides are frequently used from spring through fall; crop
cultivation frequently starts in the spring, hence uses of metrafenone
are likely to occur in spring and perhaps summer depending on the
region. Spring and early summer are typically seasons of active
migrating, feeding, and reproduction for many wildlife species.  The
increased energy demands associated with these activities (as opposed to
hibernation, for example) can increase the potential for exposure to
pesticide-contaminated food items since agricultural areas can represent
a concentrated source of relatively easily obtained, high-energy food
items. In this assessment, the spatial extent of exposure for
terrestrial animal species is limited to the use area only and the area
immediately surrounding the use area.

Currently, the Agency does not require toxicity studies on reptiles and
amphibians in support of pesticide registrations. To accommodate this
data gap, birds are used as surrogates for terrestrial-phase amphibians
and reptiles. It is assumed that, given the usually lower metabolic
demands of reptiles and amphibians compared to birds, exposure to birds
would be greater due to higher relative food consumption. While this
assumption is likely true, there are no supported relationships
regarding the relative toxicity of a compound to birds and herpetofauna.
The lack of toxicity data on reptiles and amphibians represents a source
of uncertainty in this assessment.

Tables 14 and 15 list selected predicted EECs for birds, reptiles,
terrestrial amphibians, and mammals obtained from T-REX simulations for
the proposed use of metrafenone at the maximum label rates.

Table 14.  Terrestrial Food-Item Residue Estimates for Birds with
Metrafenone Proposed Use on Grapes at 0.3 lbs a.i./A (6 apps./year; 14
day app. interval) with a Foliar Dissipation Half-life default value of
35 Days.

Crop	Food Item	Maximum Dose-Based EECs (mg/kg)1	Maximum Dose-Based EECs
(mg/kg)2	Maximum Dose-Based EECs (mg/kg)3	Dietary-Based EECs (ppm)4

Grapes	Short grass	274.49	156.52	70.08	241.01

	Tall grass	125.81	71.74	32.12	110.46

	Broadleaf plants/ small insects	154.40	88.04	39.42	135.57

	Fruits, pods, seeds, lg. insects	17.16	9.78	4.38	15.06

1Based on 20 gram birds (acute)

2 Based on 100 gram birds (acute)

3 Based on 1000 gram birds (acute)

4 Dietary-based EECs apply to both acute and chronic exposure

Table 15. Terrestrial Food-Item Residue Estimates for Mammals with
Metrafenone Proposed Use on Grapes at 0.3 lbs a.i./A (6 apps./year; 14
day app. interval) with a Foliar Dissipation Half-life default value of
35 Days.

Size Class

(grams)	Adjusted

LD501	EECs

Short Grass	Tall Grass	Broadleaf Plants/

Small Insects	Fruits/Pods/

Seeds/

Large Insects	Granivore

Dose-Based	Dose-Based	Dose-Based	Dose-Based	Dose-Based

15	2225.56	229.78	105.32	129.25	14.36	3.19

35	1800.71	158.81	72.79	89.33	9.93	2.21

1000	778.86	36.82	16.88	20.71	2.30	0.51

Dietary-Based EECs2	241.01	110.46	135.57	15.06	Not applicable

1 Herbivores/ insectivores; Granivores

2  Dietary-based EECs apply to both acute and chronic exposure

TERRPLANT

Effects on non-target terrestrial plants are most likely to occur as a
result of spray drift and/or runoff from ground applications.  These are
important factors in characterizing the risk of metrafenone to
non-target plants, which is assumed to reach off-site soil.  The
TerrPlant (Ver.1.2.2) model predicts EECs for terrestrial plants located
in dry and semi-aquatic areas adjacent to the treated field.  The EECs
are based on the application rate and solubility of the pesticide in
water and drift characteristics.  The amount of metrafenone that runs
off is a proportion of the application rate and is assumed to be 1%,
based on metrafenone’s solubility of <10 ppm (i.e. 0.474 mg/L) in
water.  Drift from ground applications are assumed to be 1% the
application rate. An incorporation depth was not referenced in the label
setting the default value to 1 inch for ground applications.  For a
standard scenario on an agricultural field, the runoff scenario for
terrestrial plants inhabiting dry areas adjacent to a field is
characterized as “sheet runoff” (one treated acre to an adjacent
acre; a 1:1 ratio) and inhabiting semi-aquatic or wetland areas adjacent
to a field is characterized as “channelized runoff” (10 treated acre
to an adjacent low-lying acre; a 10:1 ratio).  The TerrPlant model EECs
are presented in Table 16.

Table 16.  Estimated Environmental Concentrations of Metrafenone for
Terrestrial Plants from Grape Use  TC \f 3 "Table IIIB-4.  Estimated
Environmental Concentrations of Imazapyr for Terrestrial Plants from All
Uses for Ecological Assessment.

Application Method	Application Rate (lbs a.i./A)	Total Loading to Dry
areas (lb/A) 1	Total Loading to Semi-Aquatic Areas (lb/A)2	Drift (lb/A)3

Ground	0.3*	0.006	0.033	0.003

Ground	1.8**	0.036	0.198	0.018

1 EEC = Sheet Runoff + Drift (1% for ground)

2 EEC = Channelized Runoff + Drift = 1% for ground

3 EEC for ground (appl rate x 1% drift)

* Maximum single application rate

**Maximum seasonal application rate

Ecological Effects Characterization

Aquatic Effects Characterization

a. Aquatic Animals

(1) Acute Effects

Freshwater Fish and Aquatic-Phase Amphibians -Technical

The freshwater fish studies on rainbow trout (Oncorhynchus mykiss, MRID
47267443) and bluegill sunfish (Lepomis macrochirus, MRID 47267442) are
classified as supplemental for several reasons. The water samples were
not centrifuged prior to analytical determination. Without
centrifugation, the amount of chemical that is freely dissolved and
bioavailable cannot be determined. Stability of exposure to the
dissolved form throughout the test is unknown. This is especially a
problem as some of the testing concentrations were conducted above the
water solubility limit of the technical grade active ingredient (0.2-0.5
mg a.i./L at 12oC). Therefore, the concentrations are reported as mg
‘total’ a.i./L and are not useful for RQ calculations, but can
contribute to risk description.

Freshwater Fish and Aquatic-Phase Amphibians – Metabolites /
Degradates

Similar to freshwater fish studies on the technical, the freshwater fish
studies on degradates of metrafenone on rainbow trout (O. mykiss MRID
47267444, 47267445) are also classified supplemental. The water
solubility limit was not given, and there was no report of whether the
test solutions were centrifuged prior to analytical determination of the
test concentrations to ensure that the measured concentrations represent
bioavailable material. For example, in the latter study (MRID 47267445),
undissolved test substance (likely the result of exceeding the limit of
water solubility of the test material) was noted at the bottom of the
test vessel in all treatment vessels throughout the definitive test.
Again, the concentrations are reported as mg ‘total’ a.i./L and are
not useful for RQ calculations, but can contribute to risk description.

Freshwater Fish and Aquatic-Phase Amphibians -Formulations

Similar to the freshwater fish technical grade and metabolite/degradate
studies, the freshwater fish study on rainbow trout (O. mykiss MRID
47267605) is also classified as supplemental. There was no report of
whether the test solutions were centrifuged prior to analytical
determination. Without centrifugation, the amount of chemical that is
freely dissolved and bioavailable cannot be determined. Stability of
exposure to the dissolved form throughout the test is unknown; neither
is the water solubility for this formulation active ingredient. Again,
the concentrations are reported as mg ‘total’ a.i./L and are not
useful for RQ calculations, but can contribute to risk description.

Table 17.  Freshwater Fish Acute Toxicity Data

Common Name	%AI	Study parameters	LC50/NOAEC/LOAEC	MRID	Classification/

Category

Technical AC 375839 (a.k.a. BAS 560F)

Rainbow trout

Oncorhynchus mykiss	97.1	96 hour flow-through study

2 reps / 10 fish per rep.

Mean-measured: <0.000498 (negative, solvent), 0.065, 0.13, 0.25, 0.43,
and 0.82 mg total a.i./L	96-hr LC50 > 0.82 mg total a.i./L

NOAEC: 0.25 mg total a.i./L

Endpoint(s) affected: mortality, sublethal effects.

Mortality

Cumulative mortality after 96 hours was 0% among all fish in the
negative and solvent control groups, and in the treatment groups exposed
to 0.13 and 0.25 mg a.i./L of AC 375839. Cumulative mortality was 5%
among fish exposed to 0.065, 0.43 and 0.82 mg a.i./L of AC 375839 after
96 hours.

Sublethal effects

No sublethal effects were observed among fish in the negative or solvent
controls, 0.065, 0.13, 0.25 or 0.43 mg a.i./L treatment groups exposed
to AC 375839 after 96 hours. Within the 0.82 mg a.i./L treatment group,
16% of fish still living were lethargic and 5% exhibited dark
discoloration after 96 hours.	47267443	Supplemental (not to be used in
risk estimation)/

At most, Highly toxic1

Bluegill sunfish Lepomis macrochirus	97.1	96 hour flow-through study

2 reps / 10 fish per rep.

Mean-measured: <0.000498 (negative, solvent), 0.066, 0.14, 0.25, 0.45,
and 0.87 mg total a.i./L	96-hr LC50 > 0.87 mg total a.i./L

NOAEC: 0.45 mg total a.i./L

Endpoint(s) affected: mortality, sublethal effects.

Mortality

Cumulative mortality after 96 hours was 0% among all fish in the
negative and solvent control groups and all treatment groups except the
0.87 mg a.i./L treatment group in which 15% of fish died.

Sublethal effects

No sublethal effects were observed among fish in the negative or solvent
controls, 0.066, 0.14, 0.25 or 0.45 mg a.i./L treatment groups exposed
to AC 375839 after 96 hours. Within the 0.87 mg a.i./L treatment group,
18% of fish still living were lethargic and 12% were lying on the bottom
of the test chamber with little motion other than gill movement after 96
hours.	47267442	Supplemental (not to be used in risk estimation)/ At
most, Highly toxic1

Metabolites/ Degradates

Rainbow trout

Oncorhynchus mykiss	99.5	Reg. No. 4074484 (Metabolite of BAS 560 F)

96 hour static study; limit test

2 reps (neg. control), 3 reps (treatment) / 10 fish per rep.

Mean-measured: <1 (negative) and 99 mg total a.i./L	96-hr LC50 > 99 mg
total a.i./L

NOAEC ≥ 99 mg total a.i./L

Endpoint(s) affected: none.	47267444	Supplemental (not to be used in
risk estimation)/

At most, Slightly toxic1

Rainbow trout Oncorhynchus mykiss	98.2	Reg. No. 4084564

(Metabolite of BAS 560F)

96 hour static study

1 rep / 10 fish per rep.

Mean-measured: <1 (negative), 4.4, 9.2, 20.3, 35.2 and 58.4 mg total
a.i./L	96-hr LC50 15.8 (9.2-35.2) mg total a.i./L

NOAEC: 9.2 mg total a.i./L

Endpoint(s) affected: mortality, sublethal effects

Mortality

Cumulative mortality at 96 hours was 80% in the 20.3 mg total a.i./L
concentration and 100% in the two highest concentrations.

Sublethal effects

Tottering and distended abdomen were observed after 4 hours in the group
exposed to 58.4 mg total a.i./L; also observed among those in the group
exposed to 20.3 mg total a.i./L between 24 and 72 hours. Apathy and
distended abdomen were observed after 24 hours among those in the 35.2
mg total a.i./L treatment group and after 96 hours among those in the
group exposed to 20.3 mg total a.i./L.	47267445	Supplemental (not to be
used in risk estimation)/ Slightly toxic1

Formulations

Rainbow trout

Oncorhynchus mykiss 	24.7% a.i.;

294 g BAS 560 00F/L	BAS 560 00F (SF 10358, RLF 12359)

96 hour static study

2 reps. / 10 fish per rep.

Mean-measured: <4.05 (neg. control), 12, 20, 34, 56, and 94 mg form/L
[<1 (neg. control), 3.0, 4.9, 8.3, 13.7, and 23.3 mg total a.i./L]	96-hr
LC50 >94 mg form/L [>23.3 mg total a.i./L]

NOAEC ≥ 94 mg form/L [23.3 mg total a.i./L]

Endpoint(s) affected: none.  	47267605	Supplemental (not to be used in
risk estimation)/ At most, Slightly toxic1

1Based on LC50 (mg/L): < 0.1 very highly toxic; 0.1-1 highly toxic;
>1-10 moderately toxic; >10-100 slightly toxic; >100 practically
nontoxic

Freshwater Invertebrates -Technical

The freshwater invertebrate study using the technical grade active
ingredient on water flea (Daphnia magna, MRID 47267437) is classified as
supplemental. The water samples were not centrifuged prior to analytical
determination. Without centrifugation, the amount of chemical that is
freely dissolved and bioavailable cannot be determined. Stability of
exposure to the dissolved form throughout the test is unknown. This is
especially a problem as some of the testing concentrations were
conducted at or above the water solubility limit of the technical grade
active ingredient (0.2-0.5 mg a.i./L at 12oC). Therefore, the
concentrations are reported as mg ‘total’ a.i./L and are not useful
for RQ calculations, but can contribute to risk description.

Freshwater Invertebrates –Metabolites / Degradates

Similar to freshwater invertebrate study on the technical, the
freshwater invertebrate studies on degradates of metrafenone on the
water flea (D. magna MRID 47267438, 47267439) is also classified
supplemental. The water solubility limit was not given, and there was no
report of whether the test solutions were centrifuged prior to
analytical determination of the test concentrations to ensure that the
measured concentrations represent bioavailable material.  Again, the
concentrations are reported as mg ‘total’ a.i./L and are not useful
for RQ calculations, but can contribute to risk description.

Freshwater Invertebrates –Formulations

Similar to the freshwater invertebrate technical grade and
metabolite/degradate studies, the freshwater invertebrate study on water
flea (D. magna MRID 47267604) is also classified as supplemental. There
was no report of whether the test solutions were centrifuged prior to
analytical determination. Without centrifugation, the amount of chemical
that is freely dissolved and bioavailable cannot be determined.
Stability of exposure to the dissolved form throughout the test is
unknown; neither is the water solubility for this formulation active
ingredient. Again, the concentrations are reported as mg ‘total’
a.i./L and are not useful for RQ calculations, but can contribute to
risk description.

Table 18.  Freshwater Invertebrate Acute Toxicity Data

Common Name	%AI	Study parameters	EC50/NOAEC/LOAEC	MRID	Classification/

Category

Technical AC 375839 (a.k.a. BAS 560F)

Water flea

Daphnia magna	97.1	48 hour static study

2 reps.; 10 inverts. per rep

Mean-measured: <0.000498 (negative, solvent), 0.059, 0.12, 0.22, 0.45,
and 0.92 mg total a.i./L 	48-hr EC50 > 0.92 mg total a.i./L

Slope:  N/A

NOAEC ≥ 0.92 mg total a.i./L

Endpoint(s) affected: none.  	47267437	Supplemental (not to be used in
risk estimation)/

At most, Highly toxic1

Metabolites/ Degradates

Water flea

Daphnia magna	99.5	CL 375816

48 hour static study

4 reps.; 5 inverts per rep.

Mean-measured: <1 (negative), 12.9, 25.8, 51.4, and 102.5 mg total
a.i./L	48-hr EC50 > 102.5 mg total a.i./L Slope:  N/A

≥  102.5 mg total a.i./L

Endpoint(s) affected: none.  	47267438	Supplemental (not to be used in
risk estimation)/ Practically non-toxic1

Water flea

Daphnia magna	98.2	CL 4084564

48 hour static study

4 reps.; 5 inverts per rep.

Mean-measured: <1 (negative), 6.4, 12.5, 23.2, 49.6, 66.4 mg a.i./L
48-hr EC50 = 50.9 (42.1-63.5) mg total a.i./L

Slope:  4.59 (2.52-6.66)

NOAEC =  23.2 mg total a.i./L

Endpoint(s) affected: immobility.

Immobility

Cumulative immobility was 0% among all daphnids exposed to 0 (negative
control), 6.4 and 12.5 mg total a.i./L of CL 4084564 (Metabolite of BAS
560 F) after 48 hours. Cumulative immobility was 5% among animals
exposed to 23.2 mg total a.i./L, 55% among daphnids exposed to 49.6 mg
total a.i./L and 65% among animals exposed to 66.4 mg total a.i./L of CL
4084564 (Metabolite of BAS 560 F) after 48 hours.

  	47267439	Supplemental (not to be used in risk estimation)/ Slightly
toxic1

Formulations

Water flea

Daphnia magna	24.7% a.i.; 294 g BAS 560 00 F/L	BAS 560 00 F (SF 10358,
RLF 12359)

48 hour static study

2 reps.; 10 inverts per rep.

Mean-measured: <0.810 (neg. control), 1.7, 3.1, 6.1, 12.2, 24.3, and
47.1 mg form/L [<0.20 (neg. control), 0.42, 0.76, 1.5, 3.0, 6.0, and
11.6 mg total a.i./L]

	48-hr EC50: 17.1 (12.8-24.3) mg form/L [4.2 (3.2-6.0) mg total a.i./L]

NOAEC: 6.1 mg form/L [1.5 mg total a.i./L]

Endpoint(s) affected: mortality and sublethal effects (immobility)

Mortality

Cumulative mortality after 48 hours was 0% among animals exposed to
mean-measured concentrations of 0 (negative control), 1.7, 3.1, 6.1 and
12.2 mg form/L. Cumulative mortality was 65% among animals exposed to
24.3 mg form/L and 90% among animals exposed to 47.1 mg form/L after 48
hours.

Immobility

Cumulative immobility aft 48 hours was 0% at mean-measured
concentrations of 1.7 and 47.1 mg form/L; 5% at neg. control, 3.1, and
6.1 mg form/L; 15% at 12.2 mg form/L; and, 25% at 24.3 mg form/L.

	47267604	Supplemental (not to be used in risk estimation) / Moderately
toxic1

1Based on EC50 (mg/L): < 0.1 very highly toxic; 0.1-1 highly toxic;
>1-10 moderately toxic; >10-100 slightly toxic; >100 practically
nontoxic

Marine/Estuarine Fish

Unlike the majority of the aquatic studies, which were supplemental on
account of not having centrifuged samples which clearly exceeded the
water solubility limit, the marine/estuarine fish study on the
sheepshead minnow (Cyprinodon variegatus, MRID 47267446) is acceptable
on account of having centrifuged the two highest concentrations, both of
which were above the solubility limit of 0.3 mg a.i./L (in saltwater).
The results reveal no effects on mortality or sublethal effects.

Table 19.  Marine/ Estuarine Fish Acute Toxicity Data

Common Name	%AI	Study parameters	LC50/NOAEC/LOAEC	MRID	Classification/

Category

Technical AC 375839 (a.k.a. BAS 560F)

Sheepshead Minnow

Cyprinodon variegatus	94.2	96 hour flow-through study

2 reps / 10 fish per rep.

Mean-measured: <0.04 (negative, solvent), 0.072, 0.13, 0.24, 0.32 (0.13
mg a.i./L based on centrifuged samples) and 0.65 (0.35 mg a.i./L based
on centrifuged samples) mg a.i./L	96-hr LC50 > 0.35 mg dissolved a.i./L

NOAEC ≥ 0.35 mg dissolved a.i./L

Endpoint(s) affected: none.	47267446	Acceptable/

At most, Highly toxic1

1Based on LC50 (mg/L): < 0.1 very highly toxic; 0.1-1 highly toxic;
>1-10 moderately toxic; >10-100 slightly toxic; >100 practically
nontoxic

Marine/Estuarine Invertebrates – Technical

The marine/estuarine invertebrate study using the technical grade active
ingredient on eastern oyster (Crassostrea virginica, MRID 47267440) is
classified as supplemental. Given that centrifugation was not performed
on all concentration test levels, it is possible that the endpoint
retrieved from this study will provide an underestimation of risk. For
example, we know that after centrifugation of the highest test level the
mean-measured concentration decreased by 42% relative to the
pre-centrifugation mean-measured concentration. The reduction in soluble
substance is likely due to the concentration being above the limit of
solubility (i.e., 0.3 mg a.i./L). Nevertheless, it is unknown from the
study results whether a similar reduction will occur at lower
concentrations, including one more above the limit of solubility and the
three levels remaining below the limit of solubility. Similarly, the
saltwater mysid study (Americamysis bahia, MRID 47267441) is classified
supplemental on account of no centrifugation of water samples at any
test concentration level prior to analytical determination.  Without
centrifugation, the amount of chemical that is freely dissolved and
bioavailable cannot be determined. Stability of exposure to the
dissolved form throughout the test is unknown. Therefore, the
concentrations are reported as mg ‘total’ a.i./L and are not useful
for RQ calculations, but can contribute to risk description.

Table 20.  Marine/ Estuarine Invertebrate Acute Toxicity Data

Common Name	%AI	Study parameters	EC50/LC50/NOAEC/LOAEC	MRID
Classification/

Category

Technical AC 375839 (a.k.a. BAS 560F)

Eastern oyster Crassostrea virginica	94.2	96 hour flow-through study

20 bivalves per level

Time-weighted average: <0.04 (negative, solvent), 0.0522, 0.104, 0.203,
0.287, and 0.573 mg total a.i./L (highest concentration centrifuged
yielded 0.33 mg dissolved a.i./L)	96-hr EC50: 0.22 (0.20-0.25) mg total
a.i./L2

NOAEC: 0.0522 mg total a.i./L

Endpoint(s) affected: shell deposition.

Shell deposition

Relative to the negative control, the mean percent reduction in shell
growth starting with the negative solvent is as follows: 17.4, 13.9,
26.3, 44.3, 84.2, and 100%, respectively. No significant differences (p=
0.05) were detected between the two controls.	47267440	Supplemental (not
to be used in risk estimation)/

Highly toxic1

Saltwater mysid Americamysis bahia	94.2	96 hour flow-through study

2 reps./ 10 mysids per rep.

Time-weighted average: <0.04 (negative, solvent), 0.0749, 0.129, 0.240,
0.416, and 0.663 mg total a.i./L

	96-hr LC50: 0.487 (0.428-0.575) mg total a.i./L2

NOAEC: 0.0749 mg total a.i./L

Endpoint(s) affected: mortality, sublethal effects.

Mortality

At 96 hours, cumulative mortality at the TWA concentrations 0.129 and
0.240 mg total a.i./L was 5%, at 0.416 mg total a.i./L was 15%, and at
0.663 mg total a.i./L was 95%.

Sublethal effects

No sub-lethal effects were observed in the controls or TWA 0.0749-0.240
mg total ai/L treatment levels.  At test termination 7 out of the
surviving 17 mysids in the 0.416 mg total ai/L level and the single
surviving mysid at the 0.663 mg total ai/L level were observed swimming
erratically.

	47267441	Supplemental (not to be used in risk estimation)/ Highly
toxic1

1Based on EC50 (mg/L): < 0.1 very highly toxic; 0.1-1 highly toxic;
>1-10 moderately toxic; >10-100 slightly toxic; >100 practically
nontoxic

2 Range is 95% confidence interval for endpoint

(2) Chronic Effects

Freshwater Fish – Technical

The freshwater fish study on fathead minnow (Pimephales promelas, MRID
47267449) is classified as supplemental. The solubility limit of the
test compound is not reported; it is possible given previous reports
that the limit was approximately 0.2-0.5 mg a.i./L at 12 oC, which
implies that the two highest test concentrations (0.421 and 0.839 mg
total a.i./L) potentially exceeded the solubility limit and yet
centrifugation (or filtration) was not mentioned as part of the protocol
prior to analytical determination. Without centrifugation, the amount of
chemical that is freely dissolved and bioavailable cannot be determined.
Stability of exposure to the dissolved form throughout the test is
unknown. Therefore, the concentrations are reported as mg ‘total’
a.i./L and are not useful for RQ calculations, but can contribute to
risk description.

Hatching occurred at all levels on Day 4, and hatching success averaged
88-94% for all levels, with no statistically significant differences
observed.  Post-hatch survival (28-days post-hatch) was
statistically-reduced compared to the negative control at the 227, 421,
and 839 μg total a.i./L levels (p≤0.05).  Post-hatch survival
averaged 96% at the negative control through 118 μg total a.i./L
levels, and 87, 86, and 11% at the 227, 421, and 839 μg total ai/L
levels, respectively.  No clinical signs of toxicity were observed
during the study in any treatment group.  Fish length was significantly
reduced relative to the average negative control length at levels above
421 μg total ai/L and wet and dry weight were significantly lower than
the negative control weights at the 839 μg total ai/L level; however,
significant impact on survival occurred at these levels.

Table 21.  Freshwater Fish Chronic Toxicity Data

Common Name	%AI	Study parameters	NOAEC/LOAEC	MRID
Classification/Category

Technical AC 375839 (a.k.a. BAS 560F)

Fathead minnow

Pimephales promelas	97.1	32-day flow-through test

80 embryos per level, split into 20 embryos per cup, 1 cup per aquarium,
4 rep. aquaria per treatment

Time-weighted average: <0.498 (negative, solvent), 57, 118, 227, 421,
and 839 µg total a.i./L	NOAEC:  0.118 mg total a.i./L

LOAEC:  0.227 mg total a.i./L

Most sensitive endpoint: post-hatch survival

Endpoint(s) affected: post-hatch survival and growth (total length, wet
and dry weight)	47267449	Supplemental (not to be used in risk
estimation)

Freshwater Invertebrates – Technical

A chronic freshwater invertebrate study (MRID 47267447) with metrafenone
technical (97.1%) was deemed invalid due to instability of the chemical
under test conditions. A non-guideline chronic midge study (Chironomus
riparius, MRID 47267501) with metrafenone technical (97.1%) is also
invalid on the basis of low negative control emergence.

Marine/Estuarine Fish

No chronic marine/estuarine fish studies were submitted for review.

Marine/Estuarine Invertebrates- Technical

The marine/estuarine invertebrate study on saltwater mysid (Americamysis
bahia, MRID 47267448) is classified as acceptable. Although the
solubility limit of the test compound is not reported; it is possible
given previous reports that the limit was approximately 0.2-0.5 mg
a.i./L at 12 oC. The mean-measured test concentrations of the total
metrafenone present do not exceed the suggested solubility limit. In
addition, the highest nominal concentration (0.05 mg a.i./L) is
significantly below solubility (i.e., ≥ 4 - 10 times); in this
particular case, not centrifuging is considered to not affect
acceptability of the study. Finally, given the dilution system design,
where the stock solution was a solvent stock solution suggests that
solubility of the compound is not an issue in this particular case.

45 μg ai/L level (1.4 versus 5.8 offspring/female).  Similarly, the
number of offspring per female per reproductive day was
statistically-reduced compared to the negative control at the 45 μg
ai/L (0.10 versus 0.44 offspring/female/day).  Although not
statistically-compared, the percentage of females producing young
averaged ≥85% at the control through 22 μg ai/L treatment levels, but
only 25% at the 45 μg ai/L level.  The NOAEC for reproduction was
reported to be 22 μg ai/L.

Table 22.  Marine/ Estuarine Invertebrate Chronic Toxicity Data

Common Name	%AI	Study parameters	NOAEC/LOAEC	MRID
Classification/Category

Technical AC 375839 (a.k.a. BAS 560F)

Saltwater mysid Americamysis bahia	94.2	28-day flow-through test

Before pairing: 60 mysids per level

After pairing: 20 mysids per level

Time-weighted average: <LOD (negative, solvent), 2.8, 6.2, 12, 22, and
44 µg total a.i./L	NOAEC:  0.022 mg a.i./L1

LOAEC:  0.044 mg a.i./L

Most sensitive endpoint: reproduction (number of offspring/female/repro.
day)

Endpoint(s) affected: reproduction (number of offspring/female/repro.
day)	47267448	Acceptable

1 Bold value is the value that will be used to calculate risk quotients

Aquatic Plants

Vascular Aquatic Plants

The vascular aquatic plant study (MRID 47267511) on duckweed (Lemna
gibba) is classified as supplemental for several reasons, which are
considered major guideline deviations. The test was conducted for 7 days
instead of the guideline prescribed 14 days. More importantly, the
dissolved or soluble concentrations (i.e., post-centrifugation) of test
material were not determined. As a result of the latter, the stability
of exposure to the dissolved form throughout the test is unknown. This
is especially a problem as some of the testing concentrations were
conducted above the solubility limit of the technical grade active
ingredient (0.457 mg/L in water; 0.3 mg/L in 20X AAP media both at pH9).
Therefore, the concentrations are reported as mg ‘total’ a.i./L and
are not useful for RQ calculations, but can contribute to risk
description.

Non-vascular Aquatic Plants - Technical

The non-vascular aquatic plant studies on cyanobacteria (Anabaena
flos-aquae; MRID 47267512), marine diatom (Skeletonema costatum, MRID
47267513), freshwater diatom (Navicula pelliculosa, MRID 47267514), and
green algae (Pseudokirchneriella subcapitata, MRID 47267515) are
classified as supplemental for several reasons. The water samples were
not centrifuged prior to analytical determination. Without
centrifugation, the amount of chemical that is freely dissolved and
bioavailable cannot be determined. Stability of exposure to the
dissolved form throughout the test is unknown. This is especially a
problem as some of the testing concentrations were conducted above the
solubility limit of the technical grade active ingredient (0.457 mg/L in
freshwater; 0.3 mg/L in saltwater). Therefore, the concentrations are
reported as mg ‘total’ a.i./L and are not useful for RQ
calculations, but can contribute to risk description.

Non-vascular Aquatic Plants – Metabolites / Degradates

Similar to the non-vascular aquatic plant technical grade studies, the
non-vascular aquatic plant studies on the degradates are classified as
supplemental for several important reasons. For example, the algae study
(Pseudokirchneriella subcapitata MRID 47267516) did not report daily
observations of test solution appearance, metabolite water solubility,
and method of filtration of test solution, which leads to uncertainty in
the measured concentrations as well as the relationship of mean-measured
and nominal concentrations relative to the metabolite water solubility
value. For this reason too, the concentrations are reported as mg
‘total’ a.i./L and are not useful for RQ calculations, but can
contribute to risk description. In addition, the other algae study
(Pseudokirchneriella subcapitata MRID 47267517) did not centrifuge the
water samples prior to analytical determination. Without centrifugation,
the amount of chemical that is freely dissolved and bioavailable cannot
be determined. Stability of exposure to the dissolved form throughout
the test is unknown; neither is the metabolite water solubility. Again,
the concentrations are reported as mg ‘total’ a.i./L and are not
useful for RQ calculations, but can contribute to risk description.

Non-vascular Aquatic Plants - Formulations

Similar to the non-vascular aquatic technical grade and
metabolite/degradate studies, the non-vascular aquatic plant study on
the green algae (Pseudokirchneriella subcapitata) is also classified as
supplemental. Although centrifugation was conducted on the water samples
in this particular case the protocol states that it was used to remove
algal cells, which suggests that the centrifugation method may not have
removed the undissolved material. Without centrifugation, the amount of
chemical that is freely dissolved and bioavailable cannot be determined.
Stability of exposure to the dissolved form throughout the test is
unknown; neither is the water solubility for this formulation active
ingredient. Again, the concentrations are reported as mg ‘total’
a.i./L and are not useful for RQ calculations, but can contribute to
risk description.

Table 23.  Aquatic Plant Toxicity Data  TC \f 3 "Table IIIC-7.
Non-target Aquatic Plant Toxicity for Imazapyr Acid and Isopropylamine
Salt of Imazapyr.

Species

	%A.I.	Study Parameters	EC50/NOAEC	MRID No.	Study Classification

Technical AC 375839 (a.k.a. BAS 560F)

Vascular Aquatic Plants

Duckweed

Lemna gibba

	94.2	Tier II study

7 day static renewal study

3 reps. / 4 plants per rep.

Mean measured: <0.04 (negative, solvent), 0.057, 0.10, 0.21, 0.41, and
0.76 mg a.i./L	Biomass (dry weight)

EC50 > 0.76 mg total a.i./L

NOAEC: 0.21 mg total a.i./L