Document ID: EPA-HQ-OPP-2007-0431-0009
Agency: epa
Document Type: Supporting & Related Material
Title: 
Posted Date: 2007-06-20T04:00Z

SEQ CHAPTER \h \r 1 



   

UNITED STATES ENVIRONMENTAL PROTECTION AGENCY

					WASHINGTON, D.C.  20460

	

																OFFICE OF PREVENTION PESTICIDES AND

                                                			TOXIC SUBSTANCES

							

		

TXR:		0054614

DATE: 	January 31, 2007

SUBJECT:	Mefluidide:   SEQ CHAPTER \h \r 1 Toxicology Section for the
Reregistration Eligibility Decision Document (RED)

		  SEQ CHAPTER \h \r 1 Reregistration Case #: 2370

		PC Code: 114001

		DP Code D334513

FROM:	Abdallah Khasawinah, Ph.D.

		Reregistration Branch 4

		Health Effects Division (7509P)

		

THROUGH:	Susan Hummel

		Reregistration Branch 4

		Health Effects Division (7509P)

				

TO:		Yan Donovan

		Reregistration Branch 4

		Health Effects Division (7509P)

  SEQ CHAPTER \h \r 1   SEQ CHAPTER \h \r 1 Action Requested:  

  SEQ CHAPTER \h \r 1 Prepare a toxicology section for mefluidide
re-registration eligibility decision. 

4.0	Hazard Characterization/Assessment 

	4.1 Hazard characterization

Mefluidide has shown low acute toxicity by the oral, dermal and
inhalation routes (Toxicity Category III and IV).  It was a weak eye or
dermal irritant (Toxicity Category III and IV).  However, the precursor
of mefluidide (S-15733) caused eye irritation (Toxicity Category II). 
Mefluidide did not cause dermal sensitization in the guinea pig.  In
rats and rabbits, critical effects of acute oral toxicity occurring at
doses of 100 mg/kg/day and above were tremors, hunched posture,
salivation, reduced body weight and body weight gain.

Mefluidide and its diethanolamine salt subchronic and chronic toxicity
are manifested by decreased body weight and body weight gain in several
species tested (rats, rabbits and dogs). Dogs are most sensitive to
these effects, which occur at doses as low as 15 mg/kg/day in diets fed
for one year. In addition, dogs fed mefluidide for one year exhibited
chronic cortical nephrosis at doses of 150 mg/kg/day. Increased
incidence of liver hyperplastic nodules in both sexes was observed in
mice fed mefluidide at doses of 270 mg/kg/day and higher, but there was
no oncogenic response in mice at doses as high as 900 mg/kg/day.  Rats
fed mefluidide at doses up to 300 mg/kg/day did not exhibit any
carcinogenic response either.  Based on lack of carcinogenic response in
both rats and mice, mefluidide is considered as not likely to be
carcinogenic to humans. 

Mefluidide exhibited a negative response in various genotoxicity
screening assays (bacterial reverse mutation, in vitro mouse lymphoma
gene mutation, in vitro mammalian chromosome aberration, in vivo sister
chromatid exchange, unscheduled DNA synthesis).

Mefluidide and its DEA salt were not dermally toxic when tested in
rabbits at limit doses of 1000 mg/kg/day for 21 days. Effects were
limited to slight erythema at the application site at the 1000 mg/kg/day
dose.

Mefluidide or its DEA salt has not been tested for subacute or
subchronic inhalation toxicity.  However, both of them are in category
IV for acute inhalation toxicity.

  

Developmental effects of Mefluidide in rats included increased number of
early resorptions and mean postimplantation loss.  These effects were
observed at the same dose that caused maternal toxicity indicating there
was no increased susceptibility to fetuses.  The maternal toxicity
included tremors, decreased body weight, weight gain and mortality.  In
rabbit, the LOAEL/NOAEL for developmental toxicity were above the
highest dose tested (60 mg/kg/day).  In the 2-generation reproduction
toxicity study, the offspring toxicity was characterized by decreased
body weights in both sexes and both litters in all generations.  The
reproductive LOAEL was not observed (NOAEL = 346/604 mg/kg bw/day). 

The toxicology profile of mefluidide does not indicate a potential
concern for estrogens, androgen and/or thyroid mediated toxicity.

Mefluidide was almost completely absorbed following oral ingestion
(approximately 96%) and rapidly eliminated within 24 hours.  A majority
of dose was eliminated in urine (86-89 %) and remainder in feces after a
single oral dose in 24 hours. Residue consisted of mefluidide (97%) and
2 unidentified metabolites (1.2% and 0.5%) and unidentified polar
material (0.7%).  Excretion of the radioactivity in expired air was not
detected.  The chemical is unlikely to accumulate in body since it was
excreted almost completely within 24 hrs and steadily declined
thereafter. 

The toxicology profile of mefluidide and its DEA salt is adequate for
the purposes of hazard and dose response assessment.



4.2 Hazard considerations For Women and Children  tc \l1 "2.2	Food
Quality Protection Act (FQPA) HAZARD CONSIDERATIONS 

	4.2.1. Adequacy of the Toxicity Database

 tc \l2 "1. Adequacy of the Toxicity Database 

The toxicology database for mefluidide is considered adequate.  The
following acceptable studies are available:

- Developmental toxicity studies in rats

- Developmental toxicity studies in rabbits

- Two-generation reproduction study in rats

	4.2. tc \l2 "2.2.  2. Evidence of Neurotoxicity

Acute and subchronic neurotoxicity studies were not performed.  Clinical
signs of neurotoxicity (such as tremors, ataxia, atonia, decreased limb
tone, salivation) were seen in several studies (14-day oral in rabbit at
or above 200 mg/kg/day, demyelination study in chickens at 1000
mg/kg/day and two developmental toxicity studies in rats at 115
mg/kg/day.  Edema and swelling with myelin loss in sciatic nerve was
observed in a dermal toxicity study in rabbits at doses of 720 mg/kg and
above. However, these effects were not seen in an additional dermal test
of similar duration using a 58.2% mefluidide formulation or
diethanolamine salt of mefluidide 28.8%.

	4.2.3. Developmental Toxicity Study Conclusions tc \l2 "2.2.3.
Developmental Toxicity Study Conclusions 

Developmental Toxicity Study - Rabbits:

In a developmental toxicity study (MRIDs 00047139 and 00047138),
technical MBR 12325 (Lot #9) in 4% gum acacia was administered to 16-20
New Zealand White rabbits/dose group via gavage at dose levels of 0, 15,
30, or 60 mg/kg bw/day from gestation days (GD) 6-18.

There were no treatment-related effects on survival, clinical signs,
body weight, food consumption, or cesarean parameters.

The maternal LOAEL was not observed.  The maternal NOAEL is 60 mg/kg
bw/day (the highest dose tested).

There were no effects of treatment on the numbers of litters, live
fetuses, dead fetuses, or resorptions, or on fetal body weights, sex
ratio, or post-implantation loss.  There were no treatment-related
external, visceral, or skeletal variations or malformations.

The developmental LOAEL was not observed.  The developmental NOAEL is 60
mg/kg bw/day (the highest dose tested).

This developmental toxicity study in rabbits has a number of
deficiencies:  a LOAEL was not observed; test material purity was not
provided; no information on dose formulation preparation or storage was
provided; and no analyses of homogeneity, stability, or concentration
were reported. However, when combined with the 14-day oral gavage study
in rabbits (MRID 00047138), where a LOAEL of <100 mg/kg bw/day based on
mortality and tremor was established, this developmental toxicity study
is considered acceptable and satisfies the guideline for a developmental
toxicity study (OPPTS 870.3700b; OECD 414) in rabbits.

Developmental Toxicity Study - Rats:

In a developmental toxicity study (MRID 42026102), Diethanolamine salt
of Mefluidide (28.78% a.i. Lot # JB0624) in distilled water was
administered to pregnant Sprague Dawley Crl:CD BR VAF/Plus (25/dose) by
gavage at dose levels of 0, 50, 200 or 400 mg/kg bw/day (adjusted doses
for 100 % purity were 0, 14, 58, or 115 mg/kg/day, respectively) from
days 6 through 15 of gestation.  

Animals were checked daily for clinical signs, mortality.  Body weights
were measured on gestation day 0, 6, 9, 12, 16 and 20.  Unscheduled
deaths, scheduled sacrifice and c-sections were subjected to gross
necropsy examination. Each fetus was examined for
external/visceral/skeletal anomalies, sexed and then weighed.

Evidence of maternal toxicity included transient clinical signs
(tremors, dark material around the nose, few feces, urine stain and
reddish vaginal discharge), decreased body weight gain (11-61%),
decreased food consumption and mortality (2/25 females found dead on GD
11 and 16) observed at the 400 mg/kg/day levels.  At the 400 mg/kg dose,
the clinical signs of toxicity appeared within 2 days after dosing in
few animals, and after few days of dosing in some others and more than
half of the animals at this dose were free from clinical signs of
toxicity. No external malformations or developmental variations were
observed associated with any fetus. Fetal toxicity was manifested by
increase in the number of early resorptions which resulted in increase
in mean postimplantation loss at 400 mg/kg/day dose. 

After adjusting to the pure active ingredient, the maternal NOAEL is 58
mg/kg/day and the LOAEL is 115 mg/kg/day based on clinical signs
(tremors, dark material around the nose, urine stain and reddish vaginal
discharge), decreased body weight gain, decreased food consumption and
mortality (2/25 females).   The developmental toxicity NOAEL is also 58
mg/kg/day, the LOAEL is 115 mg/kg/day based on increase in the number of
early resorptions and increase in mean postimplantation loss. 

	This developmental toxicity study is classified acceptable/Guideline
and it does satisfy the guideline requirement for a developmental
toxicity study (OPPTS 870.3700; OECD 414) in the rat.

	4.2.4. Reproductive Toxicity Study

In a three-generation reproduction study (MRID 00082748), MBR 12325
(Mefluidide; 93% a.i., Lot #25) was administered in the diet to 20 male
and 40 female Charles River CD® rats/dose group at dose levels of 0,
600, 1800, or 6000 ppm (equivalent to Males/Females - 0/0, 34/60,
102/183, and 346/604 mg/kg bw/day).  When approximately 100 days old,
the P generation animals were mated (1 male: 2 females) for up to 15
days to produce the F1a litter.  Following weaning of the F1a litters,
50 F1a offspring/sex/dose were selected for a 2-year chronic feeding
study, and the remaining F1a offspring were discarded.  The P generation
was reduced to 10 males/20 females per dose group.  After a 10-day
post-weaning rest period, these P animals were mated again to produce
the F1b litter.  Upon weaning, 10 male and 20 female F1b offspring/dose
group were selected to be parents of the F2 generation.  This study
design was continued for three generations with 2 litters per
generation.

There were no effects on food consumption, organ weights, gross
pathology, or histopathology.

Numerous absolute and relative (to bw) organ weights in the 6000 ppm
parents were significantly (p<0.05) different from the controls,
however, none of these differences were corroborated by any macroscopic
or microscopic findings indicating these decreases were most likely not
related to treatment.  Thus, it is likely that they were attributable to
decreased body weights at this dose. 

The only deaths included one 6000 ppm F1 female, one 6000 ppm F2 male,
and one 1800 ppm F2 female.  It was stated that macroscopic and
microscopic findings in these animals were unremarkable.  Therefore,
these deaths were considered incidental and were not treatment related.

At 6000 ppm, body weights were decreased by 1-8% in males and 1-12% in
females throughout the study in the P generation, attaining significance
(p<0.05) at Week 18 in the males and Weeks 8, 18, 19, and 27 in the
females.  In the F1 generation at this dose, body weights were decreased
throughout the study in the males (decr. 13-21%) and females (decr.
10-21%), attaining significance (p<0.01) at Weeks 27, 37, and 56 in both
sexes.  Similarly in the F2 generation, body weights were decreased
throughout the study in the 6000 ppm males (decr. 14-21%) and females
(decr. 11-23%), attaining significance (p<0.01) at Weeks 57, 66, and 85
in both sexes.

At 1800 ppm, only minor and infrequent decreases in body weights were
noted.  There were no treatment-related findings at 600 ppm.

The parental systemic LOAEL is 6000 ppm (346/604 mg/kg bw/day in
males/females), based on decreased body weights in both sexes in all
generations.  The parental systemic NOAEL is 1800 ppm (102/183 mg/kg
bw/day in males/females). 

There were no effects of treatment on post-natal survival (i.e.,
viability and lactation) indices in the pups at any dose.  There were no
treatment-related findings at 600 or 1800 ppm.

At 6000 ppm, body weights were decreased by up to 27% compared to
controls throughout the post-natal period in both litters in each
generation (i.e., F1a, F1b, F2a, F2b, F3a, and F3b litters). These
decreases attained significance in both sexes at PND 21.

The offspring LOAEL is 6000 ppm (346/604 mg/kg bw/day in males/females),
based on decreased body weights in both sexes and both litters in all
generations.  The offspring NOAEL is 1800 ppm (102/183 mg/kg bw/day in
males/females). 

There were no effects of treatment on male or female fertility indices
or gestation survival index.

The reproductive LOAEL was not observed.  The reproductive NOAEL is 6000
ppm (346/604 mg/kg bw/day in males/females). 

This study is acceptable/guideline and satisfies the guideline
requirement for a three-generation reproductive study (OPPTS 870.3800;
OECD 416) in rats.

	4.2.5. Additional Information from Literature sources

There was no published information on this subject.

	4.2.6. Pre-and/or Postnatal Toxicity tc \l2 "5. Pre-and/or Postnatal
Toxicity 

		4.2.6.1. Determination of Susceptibility:

There is no evidence of increased pre-natal susceptibility for
mefluidide from in utero exposure in rats.  Developmental effects of
mefluidide in rats included increased number of early resorptions and
mean postimplantation loss. These effects were observed at the same dose
that caused maternal toxicity indicating there was no increased
susceptibility to the fetuses.  The maternal toxicity included tremors,
decreased body weight, weight gain and mortality.  In rabbit, the NOAEL
for developmental toxicity were above the highest dose tested (60
mg/kg/day).  In the 3-generation reproduction toxicity study, no
treatment related reproductive effects were reported.  The offspring
toxicity (decreased pup body weights) was observed at the highest dose
tested (346 mg/kg/day) that also produced maternal toxicity indicating
there was no increased post-natal susceptibility for the mefluidide.

		4.2.6.2 Degree of Concern analysis and Residual Uncertainities for
Pre- and 			/or Post –natal Susceptibility

There is no evidence of increased pre- or post-natal susceptibility in
the developmental study or in the multi-generation reproduction study in
rat.  Although the LOAEL/NOAEL for developmental toxicity in the rabbits
were not established, the concern is low for the increased
susceptibility to the rabbit fetuses since the developmental effects
were not seen at the highest dose tested (60 mg/kg/day) which is above
the developmental NOAEL in rat (58 mg/kg/day) and well above (40X) the
dose that is used to establish chronic RfD (1.5 mg/kg/day).  Therefore,
there is no residual uncertainty for pre- and/or post natal
susceptibility.

4.3. Hazard Identification and Toxicity Endpoint Selection

 tc \l2 "2.4	Hazard Identification and Toxicity Endpoint Selection 

4.3.1. Acute Reference Dose (aRfD)

Females age 13-49 : Acute dietary endpoint for child bearing females
(females 13+ years old) was determined from the developmental toxicity
study in rat (MRID 42026102).  A NOAEL of 58 mg/kg/day was derived based
on developmental toxicity (increased number of early resorptions and
mean post-implantation loss ) at a LOAEL of 115 mg/kg/day.  An UF of
100X (10-fold for inter-species extrapolation, 10-fold for intra-species
variability) was applied to the NOAEL of 58 mg/kg/day to derive the
aRfD.  

 

Acute Reference Dose (aRfD) - General Population

The acute RfD for the general population including infants and children
was determined from the developmental toxicity study in rat (MRID
42026102).  A NOAEL of 58 mg/kg/day was derived based on maternal
toxicity (clinical signs: tremors)  at a LOAEL of 115 mg/kg/day.  An UF
of 100X (10- fold for inter-species extrapolation, 10-fold for
intra-species variability) was applied.  The selected endpoint of
toxicity is appropriate for this exposure since clinical signs of
toxicity occurred within two days of dosing.

4.3.2. Chronic Reference Dose (cRfD)  

The cRfD of 0.015 mg/kg/day was determined on the basis of the Chronic
Oral Feeding study in dogs (MRID 00132995).  A NOAEL of 1.5 mg/kg/day
was selected based on chronic toxicity (decreased body weight (15%) and
body weight gain (50%) in the males) occurring at a LOAEL of 15.0
mg/kg/day.  This was the most sensitive endpoint.  An UF of 100X
(10-fold for interspecies extrapolation, 10-fold for intraspecies
variability) was applied to the NOAEL of 1.5 mg/kg/day to derive the
cRfD to give and RfD of 0.015 mg/kg/day. 

	4.3.3. Incidental Oral Exposure (Short-and Intermediate-term durations:
1 day – 6 months)  

Points of departure for these scenarios were based on the rat
developmental study (MRID 42026102).  NOAEL = 58 mg/kg bw/day, LOAEL =
115 mg/kg bw/day based on mortality and clinical signs.  These data were
also supported by the rabbit developmental study (NOAEL = 60 mg/kg
bw/day) and rabbit 14 day oral study (LOAEL = 100 mg/kg bw/day based on
mortality).  The level of concern for residential exposure is for MOEs =
100 and for occupational exposure is for MOEs =100.  

	4.3.4. Dermal Absorption Factor 

A dermal penetration study is not available.  A dermal absorption factor
is derived by extrapolation from the rabbit 21-day dermal (MRID
41972901) and rabbit 14 day oral (MRID 00082073) studies.  The dermal
systemic NOAEL in the 21-day study is 1000 mg/kg/day based on minor
increases in liver enzymes.   In the 14 day rabbit oral study(MRID
00047138), the LOAEL is less than 100 mg/kg/day based on mortality and
clinical signs (tremors) and the NOAEL is <100 mg/kg/day, therefore, the
calculated dermal absorption factor would at the most be (100/1000) x
100 = 10%. 

	4.3.5. Dermal Exposure (Short and Intermediate:  (1-30 days and 30
d-180 days) 

Three subacute (21-day) dermal toxicity studies were considered. Only
one study (MRID 00082073) with 24% active ingredient showed toxic
effects.  The other two more recent studies (MRID 42029601 and 41972901)
showed no systemic effects at the limit dose.  These effects were not
seen in additional GLP dermal tests of similar duration using a 58.2%
mefluidide formulation (MRID 42029601) or diethanolamine salt of
mefluidide 28.8% (MRID 41972901).  The risk assessment team determined
that no quantitative dermal assessment is needed due to the following:

1) Two 21-day dermal toxicity studies with rabbits indicated no dermal
systemic toxicity at 1000 mg/kg/day (the highest dose tested), 

2) The rat developmental study indicated no developmental concern
(developmental NAOEL equals to maternal NOAEL),

3) The acute dermal toxicity of mefluidide, where the acute dermal LD50
is >4000 mg/kg, it is not a skin irritant and is not a dermal
sensitizer.  

	4.3.6. Inhalation (Short- and Intermediate-Term) 

Endpoint for this scenario was determined from the rat developmental
study.  NOAEL = 58 mg/kg bw/day, LOAEL = 115 mg/kg bw/day based on
mortality and clinical sings.  These data were also supported by the
rabbit developmental study (NOAEL = 60 mg/kg bw/day) and rabbit 14 day
oral study (LOAEL = 100 mg/kg bw/day based on mortality).  Since oral
study was selected for inhalation exposure assessment an
inhalation-absorption factor of 100% oral equivalent should be used.   

 	4.3.7. Margins of Exposure

These are summarized in the following table:

Route

                                    	       Duration	Short-Term

(1-30 Days)	Intermediate-Term

(1 - 6 Months)

 

Occupational (Worker) Exposure

Dermal	NA	NA

Inhalation	100	100

Residential (Non-Dietary) Exposure

Oral	100	100

Dermal	NA	NA

Inhalation	100	100

 	

4.3.8. Classification of Carcinogenic Potential 

Mefluidide was negative for carcinogenicity in mouse (MRID 00082747) and
rat (MRID 00061930 7 00082737) bioassays.  It was also evaluated for
genotoxicity in several tests and found negative. It is unlikely that
mefluidide will pose a cancer risk to humans.

Table 4.3.  Summary of Toxicological Dose and Endpoints for Mefluidide
and its salt (114001, 114002, 114003) Used in Human Risk Assessment

Exposure

Scenario	

Point of Departure 	

Uncertainty Factors	

Level of Concern for Risk Assessment	

Study and Toxicological Effects

Acute Dietary (general population)	

NOAEL = 58 mg/kg/day 

	UFA = 10X

UFH = 10X	

Acute RfD = 

Maternal NOAEL   

UncertaintyFactor

= 0.58 mg/kg /day	

MRID 42026102

Developmental toxicity - rat;

LOAEL= 115.0 mg/kg/day based on mortality(within 5 days of dosing) and
clinical signs (within 2 days of dosing), and the NOAEL of 58 mg/kg/day.

Acute Dietary (Females 13+)	

NOAEL = 58 mg/kg/day 

	UFA = 10X

UFH = 10X	

Acute RfD =

Develop. NOAEL   

UncertaintyFactor

= 0.58 mg/kg	

MRID 42026102

Developmental toxicity - rat;

LOAEL= 115.0 mg/kg/day based on increased number of early resorptions
and mean postimplantation loss. NOAEL = 58 mg/kg/day

Chronic Dietary

(All populations)	

NOAEL = 1.5 mg/kg/day

	UFA = 10X

UFH = 10X	

Chronic RfD = 

NOAEL__ 

Uncertainty Factor

= 0.015 mg/kg/day	

MRID 00132995

Chronic Oral Feeding - dog;

LOAEL= 15.0 mg/kg/day based on decreased body weight (15%) and body
weight gain (50%) in the males at 15 mg/kg/day.

Short-Term 

Incidental Oral (1-30 days)	

NOAEL = 58 mg/kg/day 

	UFA = 10X

UFH = 10X	

Residential LOC for MOE = 100

	MRID 42026102

Developmental toxicity - rat;

NOAEL = 58 mg/kg bw/day, LOAEL = 115 mg/kg bw/day based on mortality and
clinical sings.  These data were also supported by the rabbit
developmental study (MRID 00047139) (NOAEL = 60 mg/kg bw/day) and rabbit
14 day oral study (LOAEL = 100 mg/kg bw/day based on mortality). 

Intermediate-Term 

Incidental Oral (1- 6 months)

Short-Term Dermal (1 to 30 days)

	

Dermal  NOAEL = 1000 mg/kg/day 

	UFA = 10X

UFH = 10X	

No quantitative dermal assessment is needed.

	

Three subacute (21-day) dermal toxicity studies were considered. The
risk assessment team determined that no quantitative dermal assessment
is needed due to the following:

1) Two 21-day dermal toxicity studies with rabbits indicated no dermal
systemic toxicity at 1000 mg/kg/day (the highest dose tested), 

2) the rat developmental study indicated no developmental concern
(developmental NOAEL = maternal NOAEL),  

3) acute toxicity of mefluidide, where acute dermal LD50 is >4000 mg/kg,
not a skin irritant and is not a dermal sensitizer.

Intermediate-Term

Dermal (1 to 6 months)

Short-Term Inhalation (1 to 30 days)	

Oral NOAEL = 58 mg/kg/day

(inhalation- absorption rate = 100% oral equivalent)	UFA = 10X

UFH = 10X	

Residential LOC for MOE = 100;

Occupational LOC for MOE = 100

	

MRID 42026102

Developmental toxicity - rat;

NOAEL = 58 mg/kg bw/day, LOAEL = 115 mg/kg bw/day based on mortality and
clinical sings.  These data were also supported by the rabbit
developmental study (NOAEL = 60 mg/kg bw/day) and rabbit 14 day oral
study (LOAEL = 100 mg/kg bw/day based on mortality).

Intermediate-Term Inhalation (1 to 6 months)

Cancer	Mefluidide was negative for carcinogenicity in mouse (MRID
00082747) and rat (MRID 00061930 7 00082737) bioassays.  It was also
evaluated for genotoxicity in several tests and found negative.

Point of Departure (POD) = a data point or an estimated point that is
derived from observed dose-response data and used to mark the beginning
of extrapolation to determine risk associated with lower environmentally
relevant human exposures.  UF = uncertainty factor, UFA = extrapolation
from animal to human (intraspecies), UFH = potential variation in
sensitivity among members of the human population (interspecies), NOAEL
= no observed adverse effect level, LOAEL = lowest observed adverse
effect level, RfD = reference dose (a = acute, c = chronic), MOE =
margin of exposure, LOC = level of concern, NA = Not Applicable. Safety
Factor = UF = 100.

4.4. Cumulative tc \l1 "9.0 Cumulative  Risk Characterization/Assessment

Unlike other pesticides for which EPA has followed a cumulative risk
approach based on a common mechanism of toxicity, EPA has not made a
common mechanism of toxicity finding as to Mefluidide and its salts. 
Mefluidide does not appear to produce a toxic metabolite produced by
other substances.  For the purposes of this action, therefore, EPA has
not assumed that mefluidide has a common mechanism of toxicity with
other substances.  For information regarding EPA’s efforts to
determine which chemicals have a common mechanism of toxicity and to
evaluate the cumulative effects of such chemicals, see the policy
statements released by EPA’s Office of Pesticide Programs concerning
common mechanism determinations and procedures for cumulating effects
from substances found to have a common mechanism on EPA’s website at  
HYPERLINK "http://www.epa.gov/pesticides/cumulative/" 
http://www.epa.gov/pesticides/cumulative/ .

Appendix A:  Toxicology Assessment  TC \l1 "Appendix A:  Toxicology
Assessment 

A.1	Toxicology Data Requirements TC \l2 "A.1  Toxicology Data
Requirements  

The requirements (40 CFR 158.340) for food and non food use for
Mefluidie are in Table 1. Use of the new guideline numbers does not
imply that the new (1998) guideline protocols were used. 

Test 

	Technical

	Required	Satisfied

870.1100    Acute Oral Toxicity	

870.1200    Acute Dermal Toxicity	

870.1300    Acute Inhalation Toxicity	

870.2400    Primary Eye Irritation	

870.2500    Primary Dermal Irritation	

870.2600    Dermal Sensitization		yes

yes

yes

yes

yes

yes	yes

yes

yes

yes

yes

no

870.3100    Oral Subchronic (rodent)	

870.3150    Oral Subchronic (nonrodent)	

870.3200    21-Day Dermal	

870.3250    90-Day Dermal	

870.3465    90-Day Inhalation		yes

yes

yes

-

-	yes

yes

yes

-

-

870.3700a  Developmental Toxicity (rodent)	

870.3700b  Developmental Toxicity (nonrodent)	

870.3800    Reproduction		yes

yes

yes	yes

yes

yes

870.4100a  Chronic Toxicity (rodent)	

870.4100b  Chronic Toxicity (nonrodent)	

870.4200a  Oncogenicity (rat)	

870.4200b  Oncogenicity (mouse)	

870.4300    Chronic/Oncogenicity		yes

yes

yes

yes

yes	yes

yes

yes

yes

yes

870.5100    Mutagenicity—Gene Mutation - bacterial	

870.5300    Mutagenicity—Gene Mutation - mammalian	

870.5375    Mutagenicity—Structural Chromosomal Aberrations	

870.5550    Mutagenicity—Unscheduled DNA synthesis		yes

yes

yes

yes	yes

yes

yes

yes

870.6100a  Acute Delayed Neurotox. (hen)	

870.6100b  90-Day Neurotoxicity (hen)	

870.6200a  Acute Neurotox. Screening Battery (rat)	

870.6200b  90-Day Neuro. Screening Battery (rat)	

870.6300    Develop. Neuro		no

no

no

no

no	-

-

-

-

-

870.7485    General Metabolism	

870.7600    Dermal Penetration		yes

-	yes*

Special Studies for Ocular Effects

Acute Oral (rat)	

Subchronic Oral (rat)	

Six-month Oral (dog)		

no

no

no	

-

-

-

* Non-guideline studyA.2. Toxicology Profile

Table A.2.1.  Acute Toxicity of Mefluidide and its salts (114001,
114002, 114003)

Guideline

 No.	

Study Type	

MRID	Results (LD50/LC50)	

Toxicity Category

870.1100

(81-1)	Acute Oral (female rat)

Mefluidide tech	>4000 mg/kg

MRID 00047118	

III

870.1100

(81-1)	Acute Oral (mouse) Mefluidide tech	1920.2 mg/kg

MRID 00047117 	

III

870.1100

(81-1)	Acute Oral (mouse) Mefluidide tech	829.8 mg/kg

MRID 00047116 	

III

870.1100

(81-1)	Acute Oral (dog)

Mefluidide tech	Not established

MRID 00049627;  emesis precluded evaluation at 100, 500, 2000 mg/kg
doses	

III

870.1200

(81-2)	Acute Dermal (female rabbit)

Mefluidide tech	>4000 mg/kg

MRID 00047122 & 00049628 & 00083817	

IV

870.1300

(81-3)	Acute inhalation – rat 

DEA salt of Mefluidide       	>5.2 mg/L

MRID 41888801	IV

870.1300

(81-3)	Acute inhalation – rat 

Mefluidide tech.  	>5.4 mg/L

MRID 41964601	IV

870.2400

(81-4)	Primary Eye Irritation (rabbit) Mefluidide tech	minimal
irritation

MRID 00047126, 00049630	

III

870.2400

(81-4)	Primary Eye Irritation (rabbit) DEA Mefluidide	minimal irritation

MRID43481203	III

870.2500

(81-5 )	

Primary Skin Irritation (rabbit), Mefluidide tech	

Not a dermal irritant

MRID 00047124, 00049629, 00083819	

IV

87.2600

(81-6)	Dermal Sensitization (guinea pig), Mefluidide 	Not a dermal
sensitizer

MRID 41887701	

N/A

87.2600

(81-6)	Dermal Sensitization (guinea pig), Mefluidide 	Not a dermal
sensitizer

MRID 00082076	

N/A



Table A.2.2.  Toxicity Profile of Mefluidide and its salts (114001,
114002, 114003)

Guideline No./ Study type	

MRID No.(year)/Doses/ classification	

Results

Non-guideline

21-day Oral - dog	00047137, (1975)

0, 1000, 3000, 10000 ppm Vistar tech, 93% a.i./d (0, 25, 75, 250
mg/kg/d)

One dog/sex/dose

range finding

Acceptable/non-guideline

	LOAEL = not established.

NOAEL > 250 mg/kg/d, 

Non-guideline

5-week - mouse	00082072, (1976)

0, 1800, 6000 ppm Vistar tech, 93% a.i./d (0, 270, 900 mg/kg/d)

(Dietary 5/sex/dose)

range finding

Acceptable/non-guideline	LOAEL = not established

NOAEL  = 900 mg/kg/d, 

None-guideline

28-Day oral dietary [rat]	00047135, (1973), 

0, 1000, 3000 or 10000 ppm Vistar tech, 93% a.i. (0, 100, 300, 1000
mg/kg/d)

 (Dietary 5 rats/sex/dose)

range finding

Acceptable/non-guideline

	LOAEL = not established.

NOAEL > 1000 mg/kg/d, 

870.3100

(82-1a) 

90-Day oral dietary [rat]	00047136, (1975), 

0, 300, 1000 or 6000 ppm Vistar tech, 93% a.i. (0, 15, 50, 300 mg/kg/d)
(10 rats/sex/dose)

00047140 (1975)

0, 300, 1000, 3000 ppm (0, 15, 50, 150 mg/kg/day). (10 females/dose)

Acceptable/Guideline

	

LOAEL = 300 mg/kg/d, based on decreased body weight, body weight gain
and food consumption in the females.

NOAEL = 150 mg/kg/d (in conjunction with MRID # 00047140), 

870.3150

82-1(b) 

90-Day oral dietary [dog]	00047141, (IBT Study, 1977), 

0, 300, 1000 or 6000 ppm Vistar tech, 93% a.i. (0, 7.5, 25, 150 mg/kg/d)

(4/sex/dose)

Unacceptable/guideline (LOAEL was not observed)

	LOAEL = not established.

NOAEL = 150 mg/kg/d.  

870.3200

82-2 

21-Day  Dermal toxicity - rabbit	00082073, (1977)

0, 1, 3, 10 ml of 2S formulation/kg/day (Formulation containing 24%
a.i., equivalent to  0, 240, 720, or 2400 mg mefluidide/kg/day)

(4 rabbits/sex/dose)

Acceptable/Non-guideline

(NOAEL was not observed)

	Dermal LOAEL = 240 mg/kg/day, based on irritation, inflammation and
necrosis at test sites.

Systemic LOAEL = 240 mg/kg/day, based on clinical chemistry (increased
alkaline phosphatase and alanine aminotransferase) and organ weights
(decreased spleen weight in females and increased liver weights in
males).  Edema and swelling with myelin loss in sciatic nerve was seen
in 720 and 2400 mg/kg/day dose group. Dehydration observed at 2400
mg/kg/day dose. 

Dermal and systemic NOAELs were not established.

870.3200

82-2 

21-Day  Dermal toxicity - rabbit	42029601 (1991)

0, 100, 500 or 1000 mg a.i./kg/d

Mefluidide 58.2% a.i.

(5/rabbits/sex/dose)

Acceptable/guideline

	Dermal toxicity LOAEL = 1000 mg/kg/day based on erythema at the test
site.

Dermal toxicity NOAEL = 500 mg/kg/day

Systemic toxicity LOAEL was not established.  Minor hematological and
clinical chemistry findings at 1000 mg/kg/day dose reported, that were
within normal biological variation and did not correlate to
histopathological findings.

Systemic toxicity NOAEL = 1000 mg/kg/day

870.3200

82-2 

21-Day  Dermal toxicity - rabbit	41972901 (1991)

0, 100, 500 or 1000 mg a.i./kg/d

Mefluidide DEA salt 28.78% a.i.

(5/rabbits/sex/dose)

Acceptable/ guideline

	Dermal and systemic toxicity LOAEL was not established.  Minor
incidences of erythema at 500 mg and 1000 mg/kg/day dose. Increased
liver weight (absolute and relative) noted at 1000 mg/kg dose but no
correlating histopathological findings. Minor statistical increases in
liver enzymes AST and ALT.

Dermal and systemic NOAEL = 1000 mg/kg/day

Non-guideline

1-year feeding (Rat)	00132993, (1981)

0, 60, 200, 600 ppm Vistar tech, 93% a.i. (0, 3, 10, 30 mg/kg/d)

(20 rats/sex/dose)

Addendum to 2-year feeding study.

Acceptable/non-guideline

	LOAEL = not established

NOAEL = 30 mg/kg/d,

870.4100b 

83-1b 

Chronic Oral Feeding [dog]	00132995, (1982)

0, 60, 600, 6000 ppm Vistar tech, 93% a.i. (0, 1.5, 15, 150 mg/kg/d)

(6 dogs/sex/dose)

Acceptable/guideline

	LOAEL = 15 mg/kg/d, based on decreased body weight (15%) and body
weight gain (50%) in the males. Chronic cortical nephrosis was observed
at 150 mg/kg/day dose.

NOAEL = 1.5 mg/kg/d,

870.4100b 

83-2b

 Carcinogenicity Dietary [mouse]	00082747, (1979)

0, 600, 1800, 6000 ppm Vistar tech, 93% a.i. (0, 90, 270, 900 mg/kg/d)

(60 mice/sex/dose)

Acceptable/guideline

	LOAEL = 270 mg/kg/day, based on increased incidence of liver
hyperplastic nodules in both sexes.  

NOAEL = 90 mg/kg/day. 

No oncogenicity up to and including the highest dose tested.

870.4300

83-5

 2-year feeding/carcinogenicity [rat]	00061930, 00082737 (1979)

0, 600, 1800, 6000 ppm Vistar tech, 93% a.i. (0, 30, 90, 300 mg/kg/d)

(50 rats/sex/dose)

Acceptable/guideline

	LOAEL = 30 mg/kg/d, based on body weight loss.

NOAEL < 30 mg/kg/d,  

No oncogenicity up to and including the highest dose tested.

870.3700a 

83-3(a) 

Developmental Toxicity, gavage [rat]	

00132992, (1981)

0, 15, 30, 60 mg/kg/d

Unacceptable/guideline

(LOAEL was not observed)

	

Maternal LOAEL = not established.

Maternal NOAEL > 60 mg/kg/day, 

 

Developmental NOAEL > 60 mg/kg/day, Developmental LOAEL = not
established.

Non-guideline

Range Finding Developmental Toxicity , gavage[rat]	42026101, (1991)

0, 100, 200, 400, 600or 800 mg diethanolamine salt of mefluidide
(28.78%)/kg/day

Doses adjusted for 100 % purity were 0, 29, 58, 115, 173, 230 mg/kg/day

(6 female rats/dose)

Acceptable/non-guideline

	Maternal LOAEL: 115 mg a.i./kg/day based on clinical signs (tremors,
hunched posture, and salivation), maternal body weight gain and food
consumption.

Maternal NOAEL: 58 mg a.i./kg/day; 

Developmental LOAEL: 230 mg a.i. /kg/day based on significantly
decreased fetal body weight.

Developmental NOAEL: 173 mg a.i. /kg/day, 

The dosage levels of 0, 50, 200 and 400 mg of the 28.78%
formulation/kg/day were selected for the definitive developmental study.

870.3700a 

83-3(a) 

Developmental Toxicity, gavage [rat]	42026102, (1991)

0, 50, 200 400 mg diethanolamine salt of mefluidide (28.78%)/kg/d

(25 females/dose)

Doses adjusted for 100 % purity were 0, 14, 58, or 115 mg/kg/day. 

Acceptable/guideline	Maternal LOAEL = 115 mg a.i./kg/day based on
mortality, clinical signs (tremors, stained nose, urine and vaginal
discharge), decreased body weight and weight gain.

Maternal NOAEL = 58 mg a.i./kg/day, 

Developmental LOAEL = 115 mg a.i./kg/day based on increased number of
early resorptions and mean post-implantation loss.

Developmental NOAEL: 58mg a.i./kg/day  

Non-guideline

14-Day Oral gavage [rabbit]	00047138, (1975)

0, 100, 200, 400, 800 mg/kg/d Vistar tech, 93% a.i.

4 females/dose

range finding

Acceptable/non-guideline	LOAEL = < 100 mg/kg/day (females), based on
mortality (1/3 deaths) at 100 mg/kg/d.  Tremors and 100% mortality were
noted at the levels of 200 mg/kg/d and above. Histopathology not
reported.

NOAEL: not established, 

870.3700b

83-3(b) Developmental Toxicity, gavage [rabbit]

	00047139, (1975)

0, 15, 30, 60 mg technical MBR 12325/kg/d (purity not reported).

Unacceptable by itself, however, if combined with the 14-day oral study
(00047138), it is acceptable.	Maternal LOAEL = not established.

Maternal NOAEL = 60 mg/kg/day, 

Developmental LOAEL = not established.

Developmental NOAEL = 60 mg/kg/day, 

870.3800

(83-4 )

3-generation reproduction [rat]	

00082748, (1979)

0, 600, 1800, 6000 ppm, 93% a.i. (M/F: 0/0, 34/60, 102/183, 346/604
mg/kg/d)

Acceptable/guideline	

The parental systemic LOAEL = 346/604 mg/kg bw/day (M/F), based on
decreased body weights.  

The parental systemic NOAEL = 102/183 mg/kg bw/day in males/females.

The offspring LOAEL = 346/604 mg/kg bw/day in males/females, based on
decreased body weights in both sexes and both litters in all
generations.  The offspring NOAEL = 102/183 mg/kg bw/day in
males/females. 

The reproductive LOAEL was not observed.  

The reproductive NOAEL = 346/604 mg/kg bw/day in males/females. 

870.5100

84-2 

Bacterial reverse mutation 	00132996, (1983)

EL-565 (Lily compound 151065: mefluidide technical) tested at 0.1 –
1000 µg/ml 

Acceptable/Guideline	No reverse mutations were noted in any of 8 tester
strains of Salmonella typhimurium and two tryptophan autotrophs of E.
coli with or without metabolic activation 

870.5100

84-2 

Bacterial reverse mutation 	41888804, (1991)

0, 100, 333, 667, 1000, 3330, or 5000 µg/plate diethanolamine salt of
mefluidide (28.78%)/

Acceptable/guideline	DEA mefluidide did not increase the number of
histidine revertants per plate in any of the tester strains with or
without metabolic activation.

870.5300

84-2 

In-vitro Mouse lymphoma - gene mutation 	00132996, (1983)

EL-565 (Lily compound 151065: mefluidide technical)0, 1, 25, 50, 100,
250, 500, 750 or 1000 µg/ml 

Acceptable/guideline	There was no evidence of mutation in the presence
or absence of metabolic activation.

870.5375

84-2 

In-vitro mammalian chromosome aberration test	41888803, (1991)

Diethanolamine salt of mefluidide (28.8%)

500,- 5010 µg a.i. /ml (without S9 mix) or 500, - 5000 (with S9 mix)

Acceptable/guideline 	No significant increase in structural chromosomal
aberration with or without metabolic activation was seen, however, the
results were considered equivocal.

(84-2 In-vitro mammalian chromosome aberration test	(1992)

concentrations of 1250 to 5000 µg/ml (w S9 mix) or 200-1600 µg/ml (wt
S9 mix)

	Not mutagenic in Chinese Hamster Ovary cells

870.5550

84-2 

Unscheduled DNA Synthesis	41888802, (1991)

Diethanolamine salt of mefluidide (28.8%)

Concentrations of 100, 250, 500, 1000, 2000, 3000 µg/ml in trial 1;
1000, 1500, 2000, 3000, 3500 µg/ml in trial 2.

Acceptable/guideline	No unscheduled DNA synthesis response in the
absence of moderate to severe cytotoxicity.

870.5550

84-2

Unscheduled DNA Synthesis	00132996, (1983)

EL-565 (Lily compound 151065: mefluidide technical)

Tested at 0.5 to 1000 nmoles/mL	No indication of DNA repair synthesis
was observed in cultured rat heapatocytes treated with the test material
(EL-565 (Lily compound 151065: mefluidide technical))

870.5915

84-2 

In-vivo Sister Chromatid Exchange	00132996, (1983)

EL-565 (Lily compound 151065: mefluidide technical)

(0, 12.5, 25, 50, or 100 mg/kg.

Acceptable/Guideline	Negative in sister chromatid exchange in in-vivo
bone marrow of Chinese hamster assay.

870.5915

85-1 

Metabolism- male rat	

MRID is not known: Steifer, LJ (1978). 3M Company Report Number 852
(1-26-78)

Dose: 1 or 10 mg/kg of C-14, labeled mefluidide

Acceptable/None-Guideline 

	

By 24 hrs of post-treatment, 86-89% of the dose was found in urine with
the remainder in the feces.  Residue consisted of mefluidide (97%) and 2
unidentified metabolites (1.2% and 0.5%) and unidentified polar material
(0.7%).

870.6200

81-8

 Demyelination study – Female Chicken

	

00082075 (1977)

S-12325-2S  formulation

Single doses at 1000, 3000, 5000, 10,000 and 20,000 mg/kg/day.

TOCP as positive control

5-10 hens/dose

Non-Acceptable/Non-Guideline	

LD50 was 8500 mg/kg 

At 1000 mg/kg/day: clinical signs (hypoactivity, ataxia, tremors,
lethargy and dyspnea) that were subsided by 48 hrs following dosing.  

The test material did not induce delayed neurotoxicity in hens

M = Males; F = Females

 PAGE   

Page   PAGE  17  of   NUMPAGES  17 

Acute RfD (general population)  =      58 mg/kg (NOAEL)   = 0.58 mg/kg

		    	100 (UF)

Acute RfD (Females 13-50 years old)  =      58 mg/kg (NOAEL)   = 0.58
mg/kg

		    	         100 (UF)