2,4,6-tris(2,4,6-tribromophenoxy)-1,3,5-triazine

Basic toxicokinetics

Administrative data

Endpoint:

basic toxicokinetics

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP and appropriate guidelines

Cross-reference

Cross-reference 1

Reason / purpose for cross-reference:

reference to same study

Cross-reference 2

Reason / purpose for cross-reference:

reference to other study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2006

Report date:

2006

Materials and methods

Objective of study:

other: Absorption, excretion and distribution analysis.

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 417 (Toxicokinetics)

Version / remarks:

Annex V (Toxicokinetics)

Principles of method if other than guideline:

not relevant

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

2,4,6-tris(2,4,6-tribromophenoxy)-1,3,5-triazine

EC Number:

426-040-2

EC Name:

2,4,6-tris(2,4,6-tribromophenoxy)-1,3,5-triazine

Cas Number:

25713-60-4

Molecular formula:

C 21 H 6 Br 9 N 3 O 3

IUPAC Name:

tris(2,4,6-tribromophenoxy)-1,3,5-triazine

Constituent 2

Reference substance name:

4260402

IUPAC Name:

4260402

Details on test material:

Radiolabelled material:
Name: FR-245
Physical state (20°C) White powder
Solubility: less than 0.001 mg/l in water at 20°C
Batch number: RDB/DSV014/11
Specific activity: 107.6 µCi/mg, 114.8 mCi/mmol
Radiochemical purity: > 97%
Storage: <-15°C in the dark

Non-Radiolabelled material:
Lot number: 059317
Purity: 99.9%
Storage: at ambient temperature
Exp. date: May 2007

Radiolabelling:

yes

Remarks:

Radiolabelled in the [U-14C] phenyloxy rings

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

Strain: Spargue-Dawley (Crl: CD R BR)
Age: 6-9 weeks
Weight at study initiation: 197-239 gr
Room temperature: 21+ 2oC
Relative humidity: 55+ 15%
Light/dark cycle: Alternating 12-hour light/dark
Air changes: Approx. 15 per hour
Diet: The animals were provided with food and mains tap water ad libitum
Individual metabolism cages: yes

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

other: 0.5% carboxymethylcellulose

Details on exposure:

Dose formulations were prepared freshly on the day of administration for each group of animals.
Radiolabeled and non-radiolabelled FR-245 were mixed together in dichloromethane to achieve the desired specific activity for the dosed material. The solution was dispensed in portions onto a mortar and the solvent was removed under a stream of nitrogen. To aid crystallisation diethyl ether and then hexane were added onto the solid residue and the solvent removed under nitrogen. Aqueous carboxymethylcellulose (0.5%, 0.1 ml) was added and the mixture was ground to produce a suspension of fine particles. Further carboxymethylcellulose was added in stages to a total volume of 1 ml with grinding after each addition. This was then transferred by pipette into a flask. This process was repeated until all of the FR-245 had been transferred to the flask. Additional vehicle was added to give the final dose fonnulation at the target concentration [10 mg/ml (low dose) or 200 mg/ml (high dose). If required the dose solution was ultra-sonicated using a Labsonic µ sonic probe (Braun Biotech Ltd) for 1 minute followed by 2 x 2 minutes. Throughout the preparation the dose formulation was mixed to ensure a good suspension and once prepared the formulation was continuously stirred during dosing.
For oral dosing, animals were dosed by gastric intubation at a rate of ca 5 ml formulation/kg bodyweight. The quantity of radioactivity administered to each animal was determined from the weight of the administered fonnulation (based on the weight of the dosing syringe before and after dosing) and the measured concentration of radioactivity in the fonnulation. Further weighed portions of the dose fonnulation were dispensed into 10 ml volumetric flasks. These were made up to volume with acetone and triplicate aliquots of each solution taken to confirm the homogeneity of the dose formulation and for purity measurement.
The specific activity of the 14C-FR-245 in each formulation was determined from the mass of radiolabeled and non-radiolabelled FR-245 by calculation.
Dose preparation (non-radioactive Group 3)
The procedure as described for radioactive doses was followed except only non-radiolabelled material was used. The dose was prepared at a target concentration of 200 mg/ml.

Duration and frequency of treatment / exposure:

single dose administration

Doses / concentrations

Remarks:

Doses / Concentrations:
Based on an acute oral toxicity test and a 28-day repeat dose oral toxicity study, 50 and 1000mg/kg/day of radiolabelled FR-245 were administrated orally to the rats. The absorption, distribution and excretion of the substance were studied up to 168 hours after administration.

No. of animals per sex per dose / concentration:

Male: 4 animals at 50 mg/kg
Male: 12 animals at 50 mg/kg
Male: 2 animals at 1000 mg/kg
Male: 4 animals at 1000 mg/kg
Male: 12 animals at 1000 mg/kg
Female: 4 animals at 50 mg/kg
Female: 12 animals at 50 mg/kg
Female: 2 animals at 1000 mg/kg
Female: 4 animals at 1000 mg/kg
Female: 12 animals at 1000 mg/kg

Control animals:

no

Positive control reference chemical:

no

Details on study design:

14C-FR-245 radiolabelled in the [U-14] phenyloxy rings and suspended in 0.5% carboxymethylcellulose was administered orally to groups of rats either 50 mg/kg (low) or 1000mg/kg (high) body weight.

Details on dosing and sampling:

Excretion-balance experiments (Groups 1 and 4)
Urine was collected separately from each animal into containers cooled in solid carbon dioxide at 0-6 hours, 6-24 hours and at 24-hour intervals up to 168 hours after dosing. Faeces were collected separately from each animal into containers cooled in solid carbon dioxide at 24-hour intervals up to 168 hours after dosing. Expired air was passed through two sequential traps containing 2-ethoxyethanol/ethanolamine (3:1, v/v). These solvents were exchanged after 6 and 24 hours, and then at 24-hour intervals up to 48 hours after dosing. The interior of each metabolism cage was washed with ultra-pure water at 24-hour intervals and these washings were retained for analysis.
The animals were killed by cervical dislocation at 168 hours after dosing. Following sacrifice, the gastrointestinal tract (including contents) and the remaining carcass were taken for analysis.
Plasma and whole-blood kinetics experiments (Groups 2 and 5)
In each experiment, animals were divided into three groups of eight (four of each sex). Blood samples (ca 0.4 ml) were taken from a tail vein into heparinised tubes at the following times from each group:
Subgroup 1: Pre-dose, 1, 4, 24, 96 hours. Subgroup 2: 0.25, 2, 6, 48, 120 hours. Subgroup 3: 0.5, 3, 12, 72, 168 hours.
The final blood sample in each group was taken under anaesthesia at sacrifice. Plasma was separated from the blood cells by centrifugation at 3500 revs per minute for 15 minutes.
Tissue distribution experiments (Group 6)
A group of twenty four rats (12 male and 12 female) each received a single low level dose of 50mg/kg 14C-FR-245. Eight rats (4 of each sex) were sacrified at 168 hours and at times based on the observed peak and half peak plasma concentrations from the plasma kinetics experiment. The sacrifice times were:
Sub-group 1: 4 hours Sub-group 2: 24 hours Sub-group 3: 168 hours
Immediately prior to sacrifice, the animals were lightly anaesthetised with Isoflurane/oxygen and a blood sample removed from the heart by cardiac puncture. The blood was transferred to a heparinised tube and a sample (ca 1 ml) retained as whole-blood for packed cell volume (haematocrit) and radioactivity measurement, while the remainder was centrifuged to separate the plasma. Cell fractions remaining after centrifugation were discarded. The animals were then killed by cervical dislocation. Following sacrifice, the brain, epididymis (males), gastrointestinal tract (including contents), heart, kidneys, liver, lungs, ovaries (females), testes (males), uterus (females) and samples of bone, abdominal fat, skeletal muscle and the remaining carcass were taken for analysis.
All samples generated during the course of the study were stored at <-15°C prior to analysis except for whole-blood which was stored at about 4°C and volatile trapping solutions which were stored at ambient room temperature.

Statistics:

no available

Results and discussion

Toxicokinetic / pharmacokinetic studies

Details on absorption:

Total absorption was 0.2% of the administered dose. Concentrations of radioactivity in plasma and whole blood were low, reflecting the low absorption of FR-245. Peak concentration occured 4-6 hours after dose administration indicating slow absorption from the digestive tract.

Details on distribution in tissues:

Following sacrifice of the rats at the plasma Cmax (4 hours), half plasma Cmax (24 hours) and 168 hours, proportions of radioactivity were determined in tissues. The proportion of the absorbed dose in all tissues, carcasses and including blood at the time of the peak plasma concentration (4 hours) was only 0.3%. This reduced to 0.2-0.3% after 24 hours and to < 0.01-0.02% after 168 hours. The maximum proportion of the dose in whole blood was only 0.05-0.07% indicationg there was minimal absorption into the blood stream and re-circulation back to the intestine via bile.

At 4 hour sampling the highest levels were detected in liver and kidney with all other tissues concentrations representing less than 151 ng equivalents/g. At 24 hour sampling the highest levels were detected in liver, kidney, epididymis, ovaries and uterus with all other tissue concentrations less than 86.1 ng equivalent/g.
After 168 hours the highest concentrations were detected in epididymis, fat and kidney with all other tissue representing less than 14.5ng equivalent/g.

Transfer into organs

Test no.:

#1

Transfer type:

other: Minimal absorption into the blood stream and re-circulation back to the intestine via bile

Observation:

slight transfer

Remarks:

maximum proportion in whole blood 0.05-0.07%

Details on excretion:

Following a single oral dose of 14C-FR-245 at 50 mg/kg and 100 mg/kg the radioactivity was almost entirely excreted via the faeces (95.4-105%) with the majority excreted with the interval of 0- 48 hours. Only very low amounts of radioactivity were found in the urine (0.2%), expired air (<0.1%), carcass (<0.01% low dose; 0.03-1.27% high dose) and cage washes (0.01-0.26%). Total absorption was 0.2% of the administered dose. No biliary excretion was considered to have occured since the majority of faecal radioactivity was recovered within a short time period (0-48 hours) and was mainly unmetabolised parent compound. Thus, there was no significant additional contribution from entero-hepatic recirculation via the bile to the overall absorption for this compound.
The rapid excretion in faeces, low levels of radioactivity excreted via urine and low amounts of the dose present in carcass at sacrifice indicate that FR-245 passes rapidly through the rat with minimal absroption (0.2%).

Metabolite characterisation studies

Metabolites identified:

no

Details on metabolites:

The majority (81.3-93.2%) of the excreted radioactivity over 0-48 hours was confirmed by analysis to be unmetabolised FR-245 parent test substance. Several unknown components were detected, none of which represented greater than 2.6% of the dose. The minor components detected (total of <9.1%) are most likely formed as a results of the aggressive action of both gastric and intestinal secretions, including various enzymes, on FR-245 molecules during transit through the gastro-intestinal tract.

Any other information on results incl. tables

The substance was administered as a single dose.

Absorption (groups 2 and 5): The concentration of test substance in the plasma was greatest at 4h (0.039-0.043%)
and 6h (0.06%) following administration of 50 and 1000 mg/kg, respectively. Levels were no longer detectable in the
plasma of animals treated with 50 mg/kg after either 48 h (males) or 72 h (females) and were undetectable after 24 h
in animals dosed with 1000 mg/kg. Terminal half-lives calculated for animals dosed with 50 mg/kg were 16.3 and 9.7
h, for males and females, respectively. Only estimations of the terminal half-lives could be provided for the 1000
mg/kg treated group. These were 11 and 6.3 h for males and females, respectively.

Following administration of 50 mg/kg, test substance levels in the whole-blood were greatest after 4 h (0.04-0.047%) and
were undetectable after 12h. For the 1000 mg/kg treated group, levels peaked at 6 h (0.056-0.06%) and were
undetectable after 24 h. The terminal half-lives for whole blood in the 50 mg/kg treatment groups was calculated as 2.9
h in females and estimated as 6.6 h in males. For the 1000 mg/kg treatment group the female half-life was calculated as
7.2 h, it was not possible to calculate the corresponding half-life for males.

Distribution (group 6): Distribution was investigated in the bone, brain, epydidymis, fat, heart, kidney, liver, lungs,
muscle, ovaries, residual carcass, testes, uterus, whole-blood, blood cells and plasma. The pattern of distribution
was investigated in rats dosed with 50 mg/kg and sacrificed at 4 (plasma Cmax), 24 (half plasma Cmax) and 168 h. At 4h,
the proportion of radioactivity incorporated as a whole was very low (0.26-0.34%). The highest incorporation was
observed in the liver (0.0624-0.049%), whole blood (0.05 -0.07%), plasma (0.05%), fat (0.1%) and muscle (0.008-0.02%).
All other tissues contained >0.01%. By 24 hours, the proportion recovered from each tissue had, in the main,
reduced: liver (0.0110- 0.0072%), whole blood (0.01-0.02%), plasma (0.01-0.02%), fat (0.01%), muscle (0.02%). By 168 h,
the amount of radioactivity had decreased to below 0.01% for all tissues (apart from male muscle 0.02%). Elimination
half-lives were in the region of 30-80 hours with the exception of muscle in the males (134h) and fat in the
females (112h).

Excretion (group 1 and 4): Within 48h, the majority of the administered dose was detected in the faeces (95.42-95.65%
at 50 mg/kg and 101.34-104.24% at 1000 mg/kg). Analysis of the faeces, showed the substance to be largely unaltered
(only 2.6% of the faecal extracts contained minor components, suggesting possible limited degeneration of the
test substance). Very low levels were detected in the urine (50 mg/kg: 0.17-0.24% and 1000 mg/kg: 0.17-0.18%), cagewash
(50 mg/kg: 0.03-0.28% and 1000 mg/kg: 0.01-0.03%) and expired air (50 mg/kg: 0.01-0.02% and 1000 mg/kg: 0.01-
0.03%). Excretion of test material in urine and faeces was essentially complete within 96 hours.

Biliary excretion is thought unlikely as the majority of the faecal radioactivity was recovered in the first 48h.

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information): no bioaccumulation potential based on study results No bioaccumulation potential based on study results.
Following oral administration of 14C-FR-245 to rats at 50mg/kg and 1000mg/kg there was very low absorption (<0.2%). The majority of radioactivity was excreted in te faeces.