N-[[3,5-dichloro-2-fluoro-4-(1,1,2,3,3,3-hexafluoropropoxy)phenyl]carbamoyl]-2,6-difluorobenzamide

Basic toxicokinetics

Administrative data

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

19 March 2003 - 19 October 2005

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

test procedure in accordance with generally accepted scientific standards and described in sufficient detail

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2005

Report date:

2005

Materials and methods

Objective of study:

distribution

Test guideline

Qualifier:

no guideline followed

Principles of method if other than guideline:

The purpose of the study was to determine the levels of the test material in Sprague-Dawley dams’ plasma and milk, in the plasma of foetuses and in the plasma of nursing pups. Milk samples were collected from the dams exposed to 100 mg/kg/day of the test material in diet for 28 weeks as part of a one-generation reproduction toxicity study. Milk and blood samples were collected from the nursing rats on lactation day 9 - 10. Blood samples were also collected from the postnatal day 4 pups and on gestational day 21 foetuses and dams. Both blood and milk samples were analysed for the test material using gradient high performance liquid chromatography - negative ion electrospray ionisation – mass spectrometry.

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

1-[3,5-dichloro-2-fluoro-4-(1,1,2,3,3,3-hexafluoropropoxy)phenyl]-3-(2,6-difluorobenzoyl)urea

EC Number:

601-779-5

Cas Number:

121451-02-3

Molecular formula:

C17H7Cl2F9N2O3

IUPAC Name:

1-[3,5-dichloro-2-fluoro-4-(1,1,2,3,3,3-hexafluoropropoxy)phenyl]-3-(2,6-difluorobenzoyl)urea

Test material form:

solid: particulate/powder

Details on test material:

- Physical state: White powder

Radiolabelling:

no

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Age at study initiation: Approximately 6 weeks
- Fasting period before study: No
- Housing: Animals were housed one per cage (pre-breeding) or two per cage (one male and one female during breeding) in stainless steel cages. Dams were housed one per cage (with their litter) in plastic cages provided with corn cob nesting material from approximately day 19 of gestation and throughout the lactation phase of the study. Cages had wire-mesh floors and were suspended above catch pans. Cages contained feed containers and pressure activated, nipple-type watering systems.
- Diet: ad libitum
- Water: municipal water provided ad libitum
- Acclimation period: Approximately 2 weeks

ENVIRONMENTAL CONDITIONS
- Temperature: 19 - 25 °C
- Humidity: 40 - 70 % (relative)
- Air changes: Approximately 12 - 15 times/hour
- Photoperiod: A 12-hour light/dark photocycle was maintained with lights on at 06:00 and off at 18:00

Administration / exposure

Route of administration:

oral: feed

Vehicle:

unchanged (no vehicle)

Details on exposure:

DIET PREPARATION
- Prepared using a dietary pre-mix.

Duration and frequency of treatment / exposure:

Dams were exposed daily for 28 weeks

Doses / concentrations

Dose / conc.:

100 mg/kg diet

No. of animals per sex per dose / concentration:

Not reported

Control animals:

yes, plain diet

Details on study design:

- Dose selection rationale: The dose administered was the highest dose provided in feed in a one-generation developmental toxicity study.

Details on dosing and sampling:

- Milk samples: Milk samples were collected on gestation day 9 - 10 from dams that had been administered 100 mg/kg/day in a one generation reproductive toxicity study.
- Blood samples: Blood samples were collected on gestation day 9 - 10. They were also obtained from nursing dams on gestational day 21 and postnatal day 4. Blood samples were also collected from gestational day 21 foetuses.
- Sample preparation: Blood samples were centrifuged to obtain plasma. All samples were stored at -80 °C until analysis.
- Sample analysis: Both blood and milk were analysed for the parent compound only; the test material is known to be negligibly metabolised by rats. Milk and plasma samples were removed from the freezer and allowed to thaw. The vials were vortex mixed (~30 seconds) and an aliquot of each sample (100 µL) was transferred to a high-recovery autosampler vial and fortified with radiolabelled test material (¹³C and ¹⁵N, internal standard). 200 µL of acetonitrile was added to each vial, and vortex mixed for ~1 minute. The vials were centrifuged for 10 minutes at 1322 x g for 10 minutes and the liquid was transferred to a clean vial for analysis by gradient high performance liquid chromatography - negative ion electrospray ionisation - mass spectrometry.

ANALYTICAL CONDITIONS
> HPLC-NESI-MS QUANTITATIVE ANALYSIS CONDITIONS
- HPLC System: Hewlett Packard 1100
- Injection Volume: 25 μL
- Guard Column: YMC ODS-AQ, 5 μm, cartridge
- Analytical Column: YMC ODS-AQ, 5 μm, 50 x 3 mm
- HPLC Eluent: A = MilliQ water + 0.1 % acetic acid; B = Acetonitrile + 0.1 % acetic acid
- Gradient: at time 0.00: 90 % A, 10 % B; at time 4.5 and 6.5: 0 % A, 100 % B; at time 7.5 and 10.5: 90 % A, 10 % B. The flow was 0.800 mL/min
- Eluent Split: 15/85 to MS/UV
- UV 1100 VWD: 254 nm
- 1100 MSD Mode: Negative ESI
- Drying Gas: N₂ at 7 L/min and 350 °C
- Nebulizer Pressure: 30 psig
- Capillary Voltage: 4000 V
- Fragmentor: 70 V
- EMV: ~2300 V; EMV Gain: 22
- SIM Ions: 527, 533
- Actual Dwell for all ions: 589 msec

>HPLC-NESI-MS QUALITATIVE (SCAN) ANALYSIS CONDITIONS
Same as above with the following exceptions:
- Guard Column: YMC ODS-AQ, 5 μm, cartridge (20 mm)
- Analytical Column: YMC ODS-AQ, 5 μm, 250 x 4.6 mm
- Fragmentor - 90 V
- EMV: ~2300 V; EMV Gain: 10
- Scan Range: m/z 100 to 800
- Threshold: 20
- Step Size: 0.10

Statistics:

Descriptive statistics were used (mean ± standard deviation).

Results and discussion

Toxicokinetic / pharmacokinetic studies

Transfer into organs

Key result

Test no.:

#1

Transfer type:

blood/placenta barrier

Observation:

slight transfer

Details on excretion:

The circulating plasma levels of the test material in the dams during pregnancy on gestational day 21 was 18 ± 5 µg/mL, which was 3-fold higher than the levels found in the foetuses (6 ± 1 µg/mL). Concentration in the dams’ plasma dropped by almost 3-fold during nursing (7 ± 1 µg/mL) during lactation days 9-10 after delivering the pups, which was attributed to lactation. The concentration found in milk was 23-fold higher (169 ± 35 µg/mL). Most of the test material was found to transfer to the pups through nursing. The plasma levels found in pups at postnatal day 4 were 8-9 fold higher than the foetuses at gestational day 21. There was no gender difference observed in the plasma levels of the pups. The concentration of the test material in the postnatal day 4 male and female pups was 48 ± 4 and 52 ± 9 µg/mL, respectively.

Metabolite characterisation studies

Metabolites identified:

not measured

Any other information on results incl. tables

Table 1: Results of Analysis of Milk and Plasma Samples

Sample Time	Sample	Matrix Type	Concentration of Test Material µg/mL
Gestation day 21	2968 Dam	Plasma	16.6
	2968 Foetus	Plasma	5.45
	2972 Dam	Plasma	24.7
	2972 Foetus	Plasma	8.16
	2975 Dam	Plasma	13.3
	2975 Foetus	Plasma	6.06
	2979 Dam	Plasma	16.0
	2979 Foetus	Plasma	5.15
Postnatal day 4	2978 male pup	Plasma	43.7
	2978 female pup	Plasma	42.5
	2984 male pup	Plasma	52.1
	2984 female pup	Plasma	60.0
	2986 male pup	Plasma	45.7
	2986 female pup	Plasma	59.1
	2993 male pup	Plasma	50.0
	2993 female pup	Plasma	45.0
Lactation day 9-10	2983 Dam	Plasma	7.33
		Milk	216
	2984 Dam	Plasma	8.38
		Milk	176
	2986 Dam	Plasma	7.11
		Milk	137
	2992 Dam	Plasma	6.34
		Milk	148

 

Table 2: Summary of results

Sample	Concentration of Test Material (µg/mL) in Sample
	Mean	SD
Dam Plasma Gestation Day 21	17.65	4.91
Foetus Plasma Gestation Day 21	6.21	1.36
Pup Plasma Postnatal Day 4 - Males	47.88	3.85
Pup Plasma Postnatal Day 4 - Females	51.65	9.19
Dam Plasma Lactation Day 9-10	7.29	0.84
Dam Milk Lactation Day 9-10	169.25	35.23

 

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information): bioaccumulation potential cannot be judged based on study results
The data indicate that the exposure of the test material to foetuses through the placenta is limited and the majority of exposure occurs postnatally through nursing. The systemic exposure of neonates through milk surpassed the systemic exposure to non-lactating dams by approximately 3-fold.

Executive summary:

The purpose of the study was to determine the levels of the test material in Sprague-Dawley dams’ plasma and milk, in the plasma of foetuses and in the plasma of nursing pups. Milk samples were collected from the dams exposed to 100 mg/kg/day of the test material in diet for 28 weeks as part of a one-generation reproduction toxicity study. Milk and blood samples were collected from the nursing rats on lactation day 9-10. Blood samples were also collected from the postnatal day 4 pups and on gestational day 21 foetuses and dams. Both blood and milk samples were analysed for the test material using gradient high performance liquid chromatography - negative ion electrospray ionisation – mass spectrometry.

The circulating plasma levels of the test material in the dams during pregnancy on gestational day 21 was 18 ± 5 µg/mL, which was 3-fold higher than the levels found in the foetuses (6 ± 1 µg/mL). Concentration in the dams’ plasma dropped by almost 3-fold during nursing (7 ± 1 µg/mL) during lactation days 9-10 after delivering the pups, which was attributed to lactation. The concentration found in milk was 23-fold higher (169 ± 35 µg/mL). Most of the test material was found to transfer to the pups through nursing. The plasma levels found in pups at postnatal day 4 was 8-9 fold higher than the foetuses at gestational day 21. There was no gender difference observed in the plasma levels of the pups. The concentration of the test material in the postnatal day 4 male and female pups was 48 ± 4 and 52 ± 9 µg/mL, respectively.

The data indicate that the exposure of the test material to foetuses through the placenta is limited and the majority of exposure occurs postnatally through nursing. The systemic exposure of neonates through milk surpassed the systemic exposure to non-lactating dams by approximately 3-fold.