Ethylenebis(oxyethylene) bis[3-(5-tert-butyl-4-hydroxy-m-tolyl)propionate]

Basic toxicokinetics

Administrative data

Endpoint:

basic toxicokinetics

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From Apr. 18, 1989 to Aug. 29, 1990

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

comparable to guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1990

Report date:

1990

Materials and methods

Objective of study:

absorption

distribution

excretion

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

other: US Food and Drug Administration, Bureau of Food, 1982

Deviations:

not specified

Test guideline 2

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 417 (Toxicokinetics)

Version / remarks:

1984

Deviations:

yes

Remarks:

- No metabolism studied (only absorption, distribution, and excretion tested)

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Ethylenebis(oxyethylene) bis[3-(5-tert-butyl-4-hydroxy-m-tolyl)propionate]

EC Number:

253-039-2

EC Name:

Ethylenebis(oxyethylene) bis[3-(5-tert-butyl-4-hydroxy-m-tolyl)propionate]

Cas Number:

36443-68-2

Molecular formula:

C34H50O8

IUPAC Name:

ethane-1,2-diylbis(oxyethane-2,1-diyl) bis[3-(3-tert-butyl-4-hydroxy-5-methylphenyl)propanoate]

Details on test material:

- Physical state: Solid.
- Storage condition of test material: Refrigerator, at 4°C.
- Stability under test conditions: The test article proved to be sufficiently stable as shown by TLC-analysis of the radioactivity in the formulation after the administration of the high dose level (purity ≥99.2%, mean of two determinations) as compared to the radioactivity in the stock solution (purity ≥ 98.3%).

Radiolabelling:

yes

Remarks:

Both labelled and unlabelled substances were used.

Test animals

Species:

rat

Strain:

Wistar

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: BRL Biological Research Laboratories Ltd. Wölferstrasse 4 CH-4414 Fuellinsdorf/Swltzerland.
- Weight at study initiation: 156-176g.
- Fasting period before study: Prior to dosing with 14C-IRGANOX 245 (14C-TK 12627), rats were fasted overnight.
- Housing: Groups of 2-3 rats in Makrolon cages with standard soft wood bedding during at least 2 days (Test 1 and 2). Animals of Test 2 were held individually in Makrolon cages with a stainless steel grill floor. During the metabolism studies (Test 1) rats were held individually in all-glass metabolism cages. Metabolism cages were ventilated with about 600 mL air per minute.
- Individual metabolism cages: yes.
- Diet (e.g. ad libitum): Pelleted 343-Kliba rat maintenance diet (Klingentalmushle AG, CH-4303 Kaiseraugst/Switzerland), ad libitum.
- Water (e.g. ad libitum): Tap water, ad libitum.
- Acclimation period: At least 5 days to laboratory environment incl. 3 days to metabolism cages (Test 1) or 2 days to Makrolon cages (Test 2).

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3.
- Humidity (%): 40-70.
- Air changes (per hr): 10-15.
- Photoperiod (hrs dark / hrs light): 12 / 12.

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

other: 0.5% carboxymethylcellulose in 0.9% NaCl.

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
The original viscous radioactive material was quantitatively transferred and made up to 10 mL with acetone. The addition of 1 mL chloroform to the suspension obtained did not result in a better dissolution of the radioactive material. The organic solvents were evaporated at room temperature under a gentle stream of nitrogen and the residue taken up in 50 ml of chloroform.
A clear solution (stock solution I) was obtained. As determined by liquid scintillation counting (LSC) stock solution I contained 182.01 mg of 14C-IRGANOX 245 (14C-TK 12627) with a specific radioactivity of 47.78 µCi/mg.
Based on the target dose levels and intended specific radioactivity, the following stock solutions were prepared:

Stock Solution C (Groups 1 and 3 ):
Low target dose level: 1 mg/kg (spec. radioactivity: 50 µCi/mg)
An aliquot of stock solution I, i.e. 3.0 mL corresponding to about 10.92 mg 14CIRGANOX 245 (14C-TK 12627) were made up to 10 mL with chloroform. As determined by LSC and based on a specific radioactivity of 47.78 µCi/mg, this stock solution (stock solution C) contained 11.11 mg of 14C-IRGAN0X 245 (14C-TK 12627).

Stock Solution D (Groups 2 and 4 ):
High target dose level: 10 mg/kg (spec. radioactivity: 5 µCi/mg)
Due to the specific radioactivity of about 5 µCi/mg intended for the high target dose level, an aliquot of stock solution C (spec. radioactivity: 47.78 µCi/mg) was diluted about 10 times. For this purpose, a weighed aliquot of 48.08 mg unlabelled IRGANOX 245 (TK 12627) was placed into a volumetric flask and made up to 5 mL with stock solution C. This stock solution (stock solution D) contained 53.88 mg of 14C-IRGAN0X 245 (14C-TK 12627) with a final specific radioactivity of 5.15 µCi/mg.

Formulation: Based on the average body weight of the rats, intended target dose levels of 1 mg/kg or 10 mg/kg and an excess of about 10%, appropriate aliquots (160-170 µL) of the respective stock solutions were evaporated at room temperature under a gentle stream of nitrogen, taken up in about 170-180 mg CREMOPHOR EL (BASF, Ludwigshafen/FRG) thoroughly mixed with 170-180 µL acetone and the solutions stored at -20°C until administration. Before administration, each solution was made up with 0.5% CMC (carboxymethylcellulose) in 0.9% NaCl to 1.0 mL/100 g body weight and thereafter directly administered.

VEHICLE
- Justification for use and choice of vehicle (if other than water): No Data.
- Concentration in vehicle: Refer to above description on preparation of dosing solution.
- Amount of vehicle (if gavage): 0.5% carboxymethylcellulose and 0.9% NaCl.

HOMOGENEITY AND STABILITY OF TEST MATERIAL: The test article proved to be sufficiently stable as shown by TLC-analysis of the radioactivity in the formulation after the administration of the high dose level (purity ≥99.2%, mean of two determinations) as compared to the radioactivity in the stock solution (purity ≥ 98.3%).

Duration and frequency of treatment / exposure:

Single administration.

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

1 mg/kg bw/day

Doses / concentrations 2

Dose / conc.:

10 mg/kg bw/day

No. of animals per sex per dose / concentration:

5/group (males only).

Control animals:

yes, historical

Positive control reference chemical:

Not available.

Details on study design:

- Dose selection rationale: No data.
- Rationale for animal assignment (if not random): Since the study was performed in various separate experiments, animal numbers were given randomly at each batch.

Details on dosing and sampling:

PHARMACOKINETIC STUDY (Absorption, distribution, excretion)

- Tissues and body fluids sampled:
For Experiment 1 (balance study): Urine, faeces, blood, heart, lung, liver, stomach, spleen, kidney, muscle, bones (femur), brain, fat, pancreas, intestinal tract with contents, adrenal glands, testicles, thyroid gland, skin (back region) and residual carcass. The content of the stomach was added to the carcass.
For Experiment 2 (balance study): Blood, urine, faeces, cage wash, intestinal tract/carcass/bones, organs/tissues, blood/plasma.

- Time and frequency of sampling:
For Experiment 1 (balance study): Urine/feces: Urine was taken 8, 24, 48, 72, 96, 120, 144, and 168 hours after the administration. Additionally, at sampling intervals of 8 and 24 hours metabolism cages were rinsed with a small amount of bidistilled water (about 2 mL) to remove droplets of urine. Feces were collected at room temperature 24, 48, 72, 96, 120, 144, and 168 hours after administration.
Blood/organ/tisues/intestinal tract/carcass: After 168 hours, animals were killed and blood was collected and organs and tissues were taken.
Cage wash: At the end of the study, the cages were washed first with water and liquid soap followed by acetone.
For Experiment 2 (balance study): Blood: Blood samples (100-200 microlitres) were withdrawn retroorbitally from the individual rats 0 (prior to dosing), 0.25, 0.5, 1, 2, 4, 8, 12, 24, 48, 72, 96, 120, 144 and 168 hours after the administration.
Organs/tissues/blood/plasma: The radioactivity in organs and tissues was determined after digestion of subsamples with tissue solubilizer. The radioactivity in blood and plasma was determined by digestion of subsamples followed by liquid scintillation counting.
Urine: The radioactivity in all urine samples was determined by liquid scintillation counting of subsamples. Thereafter, the rest of the urine was stored at -20°C.
Feces: To check for volatile radioactivity, a selected sample of feces was divided in two aliquots. One aliquot was homogenized wet by addition of water (about 2+l, w/w), the other aliquot was homogenized after lyophilization. Radioactivity of the respective homogenates was determined by combustion of subsamples. Since similar amounts of radioactivity were recovered in the lyophilized feces as compared to the wet feces, all feces samples were lyophilized. Feces was homogenized in a Waring Blendor (Bender & Hobein, Zürich/Switzerland) equipped with a mini-beaker (12-37 mL working volume) and a cross-knife of 26 mm diameter. In case of too small amounts, feces was directly homogenized in a mortar.
Cage Wash: Solid materials (feed, feces, etc) were homogenized by means of an Ultra-Turrax (Bender & Hobein, Zürich/Switzerland).

Statistics:

No data.

Results and discussion

Preliminary studies:

Not applicable.

Toxicokinetic / pharmacokinetic studies

Details on absorption:

The test article was absorbed in considerable amounts from the gastro-intestlnal tract. Absorption at both dose levels was rapid since the maximal blood/plasma values were reached one hour after administration.

Details on distribution in tissues:

Low dose level: 1.023 mg/kg:
At 168 hours, highest residual radioactivity levels were found in the excretory organs kidney (0.320 µg/g) and liver (0.150 µg/g). Lower but still significant amounts were found in blood (0.009 µg/g), plasma (0.009 µg/g), intestinal tract (0.009 µg/g), muscle (0.007 µg/g), skin back region (0.006 µg/g) and carcass (0.004 µg/g). In stomach and in femur, residual radioactivity levels were about the limit of quantification. In all other organs/tissues, radioactivity levels were at or below the limit of quantification.

High dose level: 10.312 mg/kg:
Again at 168 hours, highest residual radioactivity levels were found in kidney (1.033 µg/g) and liver (0.807 µg/g). In blood (0.073 µg/g), plasma (0.061 µg/g), intestinal tract (0.073 µg/g), skin back region (0.052 µg/g), muscle (0.051 µg/g), carcass (0.040 µg/g) and femur (0.027 µg/g), radioactivity levels amounted to about 1.2 to 5.2 fold the limit of quantification. In all other organs/tissues, radioactivity levels were below or about the limit of quantification. As compared to the low dose level, at about 10 times higher dose levels radioactivity levels increased about 7 - 12 fold in most organs/tissues. In the excretory organs kidney and liver, residual radioactivity levels increased only about 3.2 and 5.4 times, respectively, reflecting an efficient elimination of the test article.

Details on excretion:

Total excretion of radioactivity via the urine represented, on average, 35.01% and 33.49% of the radioactivity administered for the low and high dose level, respectively. Already 24 hours (low dose level) and 48 hours (high dose level) after the administration, urinary excretion accounted for 28.97% and 30.79% at the low and high dose levels, respectively.
Excretion via the feces accounted, on average, for 57.30% (low dose level) and 54.22% (high dose level). Again, the major amount (44.47% and 48.77%) was excreted within the first 24 hours (low dose level) and 48 hours (high dose level), respectively.
Additional radioactivity (1.48%) resulting from excretion via urine and feces was found in the cage wash at both dose levels. Total excreted radioactivity accounted, on average, for 93.79% and 89.20% at the low and high dose level, respectively.
Low amounts of radioactivity were found at 168 hours after the administration in organs/tissues (0.53 % to 1.24 %), intestinal tract (0.07% to 0.09%), blood (0.04% to 0.05%) and residual carcass (0.33% to 0.38%). Recoveries of radioactivity accounted, on average, for 95.56% and 90.23% of the radioactivity administered at the low dose and high dose levels, respectively. Based on the amounts of radioactivity recovered, excretion via expired air can not be excluded.
In conclusion, after single oral administration of 14C-test substance to male rats, the test article was absorbed in considerable amounts based on urinary excretion and thereafter rapidly excreted within the used dose range.

Toxicokinetic parameters

Toxicokinetic parameters 1

Test no.:

#2

Toxicokinetic parameters:

Cmax: 0.818 µg/g at 1 hr (plasma; dose of 1 mg/kg)

Toxicokinetic parameters 2

Test no.:

#2

Toxicokinetic parameters:

C(time): 0.036 µg/g at 48 hrs (plasma; dose of 1 mg/kg)

Toxicokinetic parameters 3

Test no.:

#2

Toxicokinetic parameters:

C(time): 2.2 fold the limit of quantification at 168 hrs (plasma; dose of 1 mg/kg)

Toxicokinetic parameters 4

Test no.:

#2

Toxicokinetic parameters:

Cmax: 0.528 µg/g at 1 hr (blood; dose of 1 mg/kg)

Toxicokinetic parameters 5

Test no.:

#2

Toxicokinetic parameters:

C(time): 0.031 µg/g at 48 hrs (blood; dose of 1 mg/kg)

Toxicokinetic parameters 6

Test no.:

#2

Toxicokinetic parameters:

C(time): 0.012 µg/g at 168 hrs (blood; dose of 1 mg/kg)

Toxicokinetic parameters 7

Test no.:

#2

Toxicokinetic parameters:

Cmax: 12.947 µg/g at 1 hr (plasma; dose of 10 mg/kg)

Toxicokinetic parameters 8

Test no.:

#2

Toxicokinetic parameters:

C(time): 0.338 µg/g at 48 hrs (plasma; dose of 10 mg/kg)

Toxicokinetic parameters 9

Test no.:

#2

Toxicokinetic parameters:

C(time): 2 fold the limit of quantification at 168 hrs (plasma; dose of 10 mg/kg)

Toxicokinetic parameters 10

Test no.:

#2

Toxicokinetic parameters:

Cmax: 7.874 µg/g at 1 hr (blood; dose of 10 mg/kg)

Toxicokinetic parameters 11

Test no.:

#2

Toxicokinetic parameters:

C(time): 0.286 µg/g at 48 hrs (blood; dose of 10 mg/kg)

Toxicokinetic parameters 12

Test no.:

#2

Toxicokinetic parameters:

C(time): 2.4 fold limit of quantification at 168 hrs (blood; dose of 10 mg/kg)

Toxicokinetic parameters 13

Test no.:

#2

Toxicokinetic parameters:

half-life 1st: 3.0 hrs (plasma; dose of 1 mg/kg)

Toxicokinetic parameters 14

Test no.:

#2

Toxicokinetic parameters:

half-life 1st: 2.9 hrs (blood; dose of 1 mg/kg)

Toxicokinetic parameters 15

Test no.:

#2

Toxicokinetic parameters:

half-life 1st: 7.2 hrs (plasma; dose of 10 mg/kg)

Toxicokinetic parameters 16

Test no.:

#2

Toxicokinetic parameters:

half-life 1st: 8.6 hrs (blood; dose of 10 mg/kg)

Toxicokinetic parameters 17

Test no.:

#2

Toxicokinetic parameters:

AUC: 9.69 µg/g x h (plasma; dose of 1 mg/kg)

Toxicokinetic parameters 18

Test no.:

#2

Toxicokinetic parameters:

AUC: 6.97 µg/g x h (blood; dose of 1 mg/kg)

Toxicokinetic parameters 19

Test no.:

#2

Toxicokinetic parameters:

AUC: 176.35 µg/g x h (plasma; dose of 10 mg/kg)

Toxicokinetic parameters 20

Test no.:

#2

Toxicokinetic parameters:

AUC: 125.52 µg/g x h (blood; dose of 10 mg/kg)

Metabolite characterisation studies

Metabolites identified:

not measured

Details on metabolites:

Not determined.

Any other information on results incl. tables

The pharmacokinetic study with 14C-labeled test material has shown that after single oral administration to male rats, the test article was absorbed in considerable amounts from the gastro-intestinal tract and thereafter rapidly excreted within the used dose range. Based on urinary excretion, the amount of absorbed radioactivity accounted for a minimum of 35.01 % and 33.49% for the low and high dose level, respectively. Excretion via the feces amounted, on average, to 57.30% (low dose level) and 54.22% (high dose level). Absorption at both dose levels was rapid since the maximal blood/plasma values were reached one hour after administration. The highest blood/plasma levels and AUC-values Increased about 15-16 and 18 fold, respectively, at about 10 times higher dose levels. These results indicate that the process of absorption was not saturated at the high dose level and that the test article was more efficiently absorbed at 10 mg/kg as compared to 1 mg/kg. Radioactivity from plasma and blood was eliminated with half-lives ranging from 2.9 hours to 8.6 hours at both dose levels. Taking into account the lower concentrations in blood as compared to the ones in plasma, these results indicate that residual radioactivity in blood was not associated with blood cells. At both dose levels, highest residual radioactivity levels at 168 hours after the administration were found in kidney and liver. These results support the function of these organs as excretory organs for the test substance. At 10 fold higher dose levels, residual radioactivity levels at 168 hours in the excretory organs kidney and liver increased only about 3.2 to 5.4 times, respectively. Taking into account the higher absorption of the test article at the higher dose level, the results implicate a more efficient elimination of the test article at 168 hours after administration of the high dose level as compared to the low dose level.