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Basic toxicokinetics

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

Data source

Reference
Reference Type:
secondary source
Title:
Absorption and metabolism of [Hexyl-2-14C] tri-(2-ethylhexyl)trimellitate in the rat
Author:
Enriquez PM, Giordano CJ and DiVincenzo GD
Year:
1984
Bibliographic source:
NTIS OTS0507501 Document ID 40-8465021

Materials and methods

Objective of study:
absorption
metabolism
Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 417 (Toxicokinetics)
Deviations:
yes
Remarks:
Only a single dose level investigated
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): tri-(2-ethylhexyl)trimellitate
- Analytical purity: 91.7%
- Impurities (identity): di-(2-ethylhexyl)terephthalate
- Radiochemical purity (if radiolabelling): > 99%
- Specific activity (if radiolabelling): 4.82 mCi/mmole
- Locations of the label (if radiolabelling): [Hexyl 2-14C] tri(2-ethylhexyl)trimellitate
- Stability under test conditions: Stable
Radiolabelling:
yes
Remarks:
[Hexyl 2-14C] tri(2-ethylhexyl)trimellitate

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Inc, Wilmington, MA
- Weight at study initiation: 200 - 300 g
- Fasting period before study: 16 hours
- Housing:
- Individual metabolism cages: yes
- Diet: Purina laboratory rodent chow Ref. 5001 available ad libitum immediately following dosing
- Water: ad libitum

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
corn oil
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: Labeled substance mixed with non-labelled substance and dissolved/diluted in corn oil

VEHICLE
- Justification for use and choice of vehicle (if other than water): Commonly used vehicle for poorly water soluble substances
- Concentration in vehicle: Not reported
- Amount of vehicle (if gavage): Not reported

HOMOGENEITY AND STABILITY OF TEST MATERIAL: Not reported
Duration and frequency of treatment / exposure:
Dosed once only
Doses / concentrations
Remarks:
Doses / Concentrations:
100 mg/kg body weight (16-18 microCi/animal)
No. of animals per sex per dose:
4
Control animals:
yes
Details on study design:
Rats were dosed once only, by gavage, with 100 mg/kg TOTM. Following dosing animals were placed in separate glass metabolism chambers to permit separation of expired air, urine and faeces. Expired air was passed through two silica gel traps to remove volatile organic compounds and then through two NaOH traps to remove CO2. The silica gel traps were changed daily and the NaOH traps were changed at intervals up to 144 hours post dose. Urine and faeces were collected at 4, 8, 12, 24, 36, 48, 72, 96, 120 and 144 hours post dose an radioactivity measured. At each collection period the metabolism cages were rinsed with water. On termination following 144 hours cages were also washed with methanol followed by methylene chloride. Rats were killed 144 hours post dose and brain, heart, lungs, liver, spleen, kidneys, small and large intestines, testes and abdominal fat were removed.

Radioactivity Measurements - A Liquid Scintillation Spectrometer was used for measurement of radioactivity with efficiency corrections made by external standardisation. Most samples were counted in a naphthalene-dioxane based scintillator fluid.

Processing of Tissues and Excreta - Tissues were extracted twice with 95% aqueous acetone. Combined extracts were counted in scintillation fluid. Residues were dried, powdered, combusted and counted. Carcasses were shredded in a blender with acetone, the mixture refluxed overnight and then filtered. The residue was then refluxed with n-hexane and filtered. The acetone and n-hexane were then counted in scintillation fluid. Solid residues were powdered and counted for radioactivity. Faeces were homogenised in acetone. Solids were separated by centrifugation and residues extracted twice with acetone. Combined extracts were counted in scintillation fluid. Residues were then further extracted with methylene chloride and the extracts counted in scintillation fluid.

Analysis of TOTM and its metabolites in excreta - Radioactive components in faecal extracts were separated using HPLC utilising both spectrophotometric (uv/vis) and radiochemical detection. Standards used for metabolite characterisation were prepared from [Hexyl 2-14C] tri(2-ethylhexyl)trimellitate and from synthetic mixtures prepared by the incomplete esterification of trimellitic acid with 2-ethylhexanol. These mixtures were formed from 1:1 and 2:1 molar ratios of 2-ethyhexanol and trimellitic acid and were regarded as being mono- and di-esters, mono(2-ethylhexyl)trimellitate and di(2-ethylhexyl)trimellitate. Faecal extracts were analysed directly using HPLC. Urinary metabolites were analysed by GC/MS following solvent extraction in diethyl ether.

Analysis of Kinetic Data - Excretion half-lives were estimated from the rates of excretion of radioactivity in urine and in expired air.
Details on dosing and sampling:
PHARMACOKINETIC STUDY (Absorption, distribution, excretion)
- Tissues and body fluids sampled: urine, faeces, expired air, cage washes, tissues (brain, heart, lungs, liver, kidneys, small & large intestines, testes and abdominal fat)
- Time and frequency of sampling: 4, 8, 12, 24, 36, 48, 72, 96, 120 and 144 hours post-dose - tissues at 144 hours only

METABOLITE CHARACTERISATION STUDIES
- Tissues and body fluids sampled: urine, faeces
- Time and frequency of sampling: 4, 8, 12, 24, 36, 48, 72, 96, 120 and 144 hours post-dose
- From how many animals: 4 animals. No details on pooling of samples
- Method type(s) for identification: GC-MS, HPLC-UV-vis
- Limits of detection and quantification: Not reported
:
Statistics:
Students' t-test. Values of p < 0.05 regarded as significant

Results and discussion

Main ADME resultsopen allclose all
Type:
absorption
Results:
Half-life - 0.7 hours
Type:
excretion
Results:
Half-life - 42 hours

Toxicokinetic / pharmacokinetic studies

Details on absorption:
The excretion of radioactivity following administration of a single oral dose occurred mainly in the faeces, this accounting for approximately 75% of the administered dose with 16.3% found in the urine and 1.9% in expired air.
Two peaks in the plot of the rate of excretion of 14CO2 in expired air were observed for each rat in the study. The first was observed 2-3 hours post dose and the second 8-12 hours post dose, the authors suggesting this to be a result of metabolism of 2-ethylhexanol occurring from its release by hydrolysis from the tri-ester followed by a further hydrolysis step releasing 2-ethyl hexanol from the di-ester. The half-life for initial absorption was estimated to be approximately 0.7 hours.
Details on distribution in tissues:
Residual radioactivity in the carcass 144 hours post dose was <0.6% of the administered dose. Tissues analysed for radioactivity revealed only liver (5x) and adipose tissue (3x) containing levels of radioactivity greater than the carcass average.
Details on excretion:
Radioactivity in the faecal extracts was identified as 86% TOTM, 7% di-(2-ethylhexyl)trimellitate and 1% mono-(2-ethylhexyl)trimellitate. Only one of the three possible mono-ester isomers was found.
Analysis of urine by GC/MS revealed the presence of 2-ethylhexanol, 2-ethylhexanoic acid, 2-heptanone and mono-(2-ethylhexyl)trimellitate. HPLC retention times indicated that the mono-ester was the same isomer found in faecal extracts. No isomers of di-(2-ethylhexyl)trimellitate were found. Urinary elimination of radioactivity was bi-phasic with half-lives of 3.1 and 42 hours

Metabolite characterisation studies

Metabolites identified:
yes
Details on metabolites:
Radioactivity in the faecal extracts was identified as 86% TOTM, 7% di-(2-ethylhexyl)trimellitate and 1% mono-(2-ethylhexyl)trimellitate. Only one of the three possible mono-ester isomers was found.
Analysis of urine by GC/MS revealed the presence of 2-ethylhexanol, 2-ethylhexanoic acid, 2-heptanone and mono-(2-ethylhexyl)trimellitate. HPLC retention times indicated that the mono-ester was the same isomer found in faecal extracts. No isomers of di-(2-ethylhexyl)trimellitate were found.

Any other information on results incl. tables

Disposition of Radioactivity -The excretion of radioactivity following administration of a single oral dose occurred mainly in the faeces, this accounting for approximately 75% of the administered dose with 16.3% found in the urine and 1.9% in expired air. Residual radioactivity in the carcass 144 hours post dose was <0.6% of the administered dose. Tissues analysed for radioactivity revealed only liver (5x) and adipose tissue (3x) containing levels of radioactivity greater than the carcass average. Recovery was 94.1% of the administered dose.

 

Characterisation of Faecal and Urinary metabolites -The radioactive components of faecal extracts were characterised by HPLC. Using the prepared standards it was possible to identify three monoesters and two diesters while evidence for a third diester was obtained by UV spectral analysis. Radioactivity in the faecal extracts was identified as 86% TOTM, 7% di-(2-ethylhexyl)trimellitate and 1% mono-(2-ethylhexyl)trimellitate. Only one of the three possible mono-ester isomers was found.

Analysis of urine by GC/MS revealed the presence of 2-ethylhexanol, 2-ethylhexanoic acid, 2-heptanone and mono-(2-ethylhexyl)trimellitate. HPLC retention times indicated that the mono-ester was the same isomer found in faecal extracts. No isomers of di-(2-ethylhexyl)trimellitate were found.

 

Kinetics -Two peaks in the plot of the rate of excretion of14CO2in expired air were observed for each rat in the study. The first was observed 2-3 hours post dose and the second 8-12 hours post dose, the authors suggesting this to be a result of metabolism of 2-ethylhexanol occurring from its release by hydrolysis from the tri-ester followed by a further hydrolysis step releasing 2-ethyl hexanol from the di-ester. The half-life for initial absorption was estimated to be approximately 0.7 hours. The die-away of14CO2was bi-phasic with half-lives of 4.3 and 31 hours. Urinary excretion of radioactivity was also bi-phasic with half-lives of 3.1 and 42 hours.

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information): low bioaccumulation potential based on study results
TOTM is only partially hydrolysed in the gastro-intestinal tract to 2-ethylhexanol and the corresponding di-ester and, following further hydrolysis, the mono-ester. Only 2-ethylhexanol and a single isomer of mono-(2-ethylhexyl)trimellitate appear to be absorbed. Following absorption, 2-ethylhexanol was extensively metabolised with metabolites eliminated in the urine and as expired 14CO2.There was no evident metabolism of mono-(2-ethylhexyl)trimellitate, this being eliminated unchanged.
Executive summary:

The absorption, distribution, metabolism and elimination of TOTM have been investigated in the rat following oral administration of a single dose. Findings indicate that TOTM is only partially hydrolysed in the gastro-intestinal tract to 2-ethylhexanol and the corresponding di-ester and, following further hydrolysis, the mono-ester. Only 2-ethylhexanol and a single isomer of mono-(2-ethylhexyl)trimellitate appear to be absorbed. Following absorption, 2-ethylhexanol was extensively metabolised with metabolites eliminated in the urine and as expired14CO2. There was no evident metabolism of mono-(2-ethylhexyl)trimellitate, this being eliminated unchanged.