Registration Dossier

Administrative data

Link to relevant study record(s)

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Endpoint:
basic toxicokinetics in vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Unspecified (results initially published in 2000)
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
test procedure in accordance with generally accepted scientific standards and described in sufficient detail
Objective of study:
toxicokinetics
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 417 (Toxicokinetics)
GLP compliance:
not specified
Radiolabelling:
yes
Remarks:
[14C]-p-tert-butylcatechol
Species:
mouse
Strain:
B6C3F1
Sex:
male
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Inc. (Raleigh, NC, and Portage, MI)
- Age at study initiation: 9 to 10 weeks old
- Weight at study initiation: No data
- Fasting period before study: No data
- Housing: Polycarbonate cages
- Individual metabolism cages: yes (from the day before dosing)
- Diet: Purina Rodent Chow No. 5002
- Water: ad libitum
- Acclimation period: 1 week


ENVIRONMENTAL CONDITIONS
Not specified


IN-LIFE DATES: Not specified
Route of administration:
dermal
Vehicle:
other: acetone
Details on exposure:
TEST SITE
- Area of exposure: 1 cm²
- % coverage: Not specified
- Type of wrap if used: metal tissue capsule glued over the dosing site with QuicktiteTM Super Glue Gel
- Time intervals for shavings or clipplings: Clipping approx. 24 hours before dose application


REMOVAL OF TEST SUBSTANCE
- Washing (if done): No
- Time after start of exposure: up to 72 hours after dosing


TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 2.5 to 22.2 μCi [14C]-radiolabel and appropriate amount of unlabeled p-tert-butylcatechol
- concentration (if solution): Not applicable


VEHICLE
- Justification for use and choice of vehicle (if other than water): Not specified
- Amount(s) applied (volume or weight with unit): approximately 25 μL
- Concentration (if solution): Not applicable
- Lot/batch no. (if required): Not specified
- Purity: Not specified


USE OF RESTRAINERS FOR PREVENTING INGESTION: No data
Duration and frequency of treatment / exposure:
single exposure
Remarks:
Doses / Concentrations:
0.04 or 4 mg/animal
No. of animals per sex per dose:
4 males
Control animals:
no
Details on study design:
- Dose selection rationale: dermal doses for mice selected to provide approximately equal concentrations of p-tert-butylcatechol per square centimeter of skin as the 0.6 and 60 mg/kg doses administered to rats.
Details on dosing and sampling:
PHARMACOKINETIC STUDY (Absorption, distribution, excretion)

- Tissues and body fluids sampled: urine, faeces.
- Time and frequency of sampling: once after 72h. Radioactivity was measured in urine collected 6, 24, 48 and 72h after  dosing and feces collected 24, 48 and 72h after dosing (stored in dark at  -20°C until analysis). Skin wash and digests of the application site skin and carcass were  analysed by LSS.

METABOLITE CHARACTERISATION STUDY: no.
Details on absorption:
Most of the dose was absorbed by 72 hours following application (72% for the 0.04 mg dose and  86% for the 4 mg dose).
Mice applied 4 mg showed signs of local inflammation at the site of application.
Details on distribution in tissues:
Global distribution of radioactivity in tissues, at the application site, in urine and feces (% of dose) is detailed below.
Details on excretion:
Cumulative excretion of radioactivity is detailed below.
Metabolites identified:
not measured

Cumulative excretion of radioactivity (cumulative % of dose, mean +/- standard deviation):

Dose (mg) Time (h) Urine (%) Feces (%) Total (%)
0.04 4.3 +/- 5.2  - (not collected) 4.3 +/- 5.2 
  24 32.1 +/- 9.6 7.45 +/- 5.39  39.6 +/- 13.6
  48 47.1 +/- 10.7 8.15 +/- 5.31  55.2 +/- 15.3
  72  51.7 +/- 8.9 12.4 +/- 7.7  64.1 +/- 15.7
4 6 11.9 +/- 14.0  - (not collected) 11.9 +/- 14.0
  24  54.3 +/- 10.5 2.58 +/- 1.30 56.9 +/- 10.3
  48  68.3 +/- 3.9 3.48 +/- 1.51 71.8 +/- 3.0 
  72  71.4 +/- 3.5 5.42 +/- 2.55  76.8 +/- 1.4

Distribution of radioactivity at 72 hours (% of dose, mean +/- standard deviation):

 Dose (mg/kg)  Tissues (%)  Application site (%)  Feces (%)  Urine (%)  Total absorbed dose (%)  Total unabsorbed dose (%)
 0.04 11.6 +/- 8.3 1.51 +/- 0.43  12.4 +/- 7.7 57.0 +/- 7.5 72.2 +/- 14.2 11.6 +/- 8.3
 4 0.76 +/- 0.30 6.18 +/- 0.52 5.4 +/- 2.6 73.7 +/- 3.2 86.0 +/- 2.7 2.25 +/- 0.81


Conclusions:
Bioaccumulation potential cannot be judged based on study results
The absorption, distribution and excretion of 4-tert butylpyrocatechol were studied in mice following a single dermal application of radiolabeled compound.
Executive summary:

Groups of 4 mice were administered a single dermal application of 0.04 or 4 mg/kg 14C-4-tert butylpyrocatechol (TBC).

At 72 hours following application, the total unabsorbed dose was 11.6% or 2.25% from total radioactivity, at 0.04 or 4 mg, respectively.

Most of the excreted radioactivity was retrieved from urine (up to 73.7%).

Endpoint:
basic toxicokinetics in vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Unspecified (results initially published in 2000)
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
test procedure in accordance with generally accepted scientific standards and described in sufficient detail
Objective of study:
toxicokinetics
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 417 (Toxicokinetics)
GLP compliance:
not specified
Radiolabelling:
yes
Remarks:
[14C]-p-tert-butylcatechol
Species:
mouse
Strain:
B6C3F1
Sex:
male
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Inc. (Raleigh, NC, and Portage, MI)
- Age at study initiation: 9 to 10 weeks old
- Weight at study initiation: No data
- Fasting period before study: No data
- Housing: Polycarbonate cages
- Individual metabolism cages: yes (from the day before dosing)
- Diet: Purina Rodent Chow No. 5002
- Water: ad libitum
- Acclimation period: 1 week


ENVIRONMENTAL CONDITIONS
Not specified


IN-LIFE DATES: Not specified
Route of administration:
intravenous
Vehicle:
other: isotonic saline or 10% Emulphor in B6C3F1 mouse plasma
Details on exposure:
Doses for intravenous injection contained 6.3 to 23 μCi [14C]-radiolabel and an appropriate amount of unlabeled p-tert-butylcatechol.

The initial set of doses for mice were formulated in isotonic saline; doses for a repeat study were formulated in 10% Emulphor in B6C3F1 mouse plasma. The dosing volume for mice was 2 mL/kg.

The doses were injected into a lateral tail vein.
Duration and frequency of treatment / exposure:
single administration
Remarks:
Doses / Concentrations:
3 mg/kg
No. of animals per sex per dose:
4 males (initial study) + 2 males (repeat study)
Control animals:
no
Details on study design:
- Dose selection rationale: Not specified
- Rationale for animal assignment (if not random): No data
Details on dosing and sampling:
PHARMACOKINETIC STUDY (Absorption, distribution, excretion):
- Tissues and body fluids sampled: adrenal gland, blood, brain, kidney,  liver, lung, prostate gland, spleen, testis with seminal vesicle, as well  as three samples each of adipose tissue, muscle and skin (analysed for  carbon-14 content). For some animals, the cecum, large and small  intestine, and stomach were also analysed.
Radioactivity was measured in urine collected 6, 24, 48 and 72h after  dosing and feces collected 24, 48 and 72h after dosing (stored in dark at  -20°C until analysis).
Details on distribution in tissues:
As for rats, no tissue showed high concentrations of TBC residues.

Because the tissue-to-blood ratio favored lung over liver by a factor of 10 for mice but was approximately equal for rats, a repeat study was conducted for mice with the dose formulated in a mixture of Emulphor in plasma as for rats. The percentage of dose excreted was similar between vehicles, and the tissue-to-blood ratio in the repeat study was only slightly greater for lung than for liver. Therefore it was speculated that the dose for the initial study (formulated in isotonic saline) contained particles of TBC large enough to lodge in the lung following injection; the dose for the repeat study contained no particles of this size.
Details on excretion:
Excretion of radioactivity was slower in mice than in rats, with greater concentrations excreted in feces (28% for mice versus 6% for rats). However, the mouse feces samples were often contaminated with urine-soaked feed pellets.
Metabolites identified:
not measured

Cumulative excretion of radioactivity (cumulative % of dose, mean +/- standard deviation):

Time (h) Urine (%) Feces (%) Total (%)
Initial assay
6 13.3 +/- 937 - (not collected) 13.3 +/- 9.7
24 37.3 +/- 17.6 16.4 +/- 13.4 53.7 +/- 21.7
48 43.2 +/- 19.9 21.0 +/- 15.5 64.3 +/- 20.3
72 59.9 +/- 22.3 28.1 +/- 17.2 88.0 +/- 8.3
Repeat assay
6  8.26 +/- 7.74   - (not collected) 8.26 +/- 7.74 
24 28.7 +/- 14.5 13.1 +/- 13.1 41.8 +/- 27.7
48  37.6 +/- 22.8 24.5 +/- 0.8 62.1 +/- 22.0
72 40.8 +/- 21.2 25.7 +/- 1.1 66.6 +/- 20.0
Conclusions:
Bioaccumulation potential cannot be judged based on study results
The distribution and excretion of 4-tert butylpyrocatechol were studied in mice following a single intravenous injection of radiolabeled compound during two distinct assays with different vehicles.
Executive summary:

Groups of 2 or 4 mice were administered a single intravenous injection of 3 mg/kg14C-4 -tert butylpyrocatechol (TBC) in isotonic saline or in 10% Emulphor in mouse plasma.

As for rats, no tissue showed high concentrations of TBC residues.

 

Because the tissue-to-blood ratio favored lung over liver by a factor of 10 for mice but was approximately equal for rats, a repeat study was conducted for mice with the dose formulated in a mixture of Emulphor in plasma as for rats. The percentage of dose excreted was similar between vehicles, and the tissue-to-blood ratio in the repeat study was only slightly greater for lung than for liver. Therefore it was speculated that the dose for the initial study (formulated in isotonic saline) contained particles of TBC large enough to lodge in the lung following injection; the dose for the repeat study contained no particles of this size.

 

Excretion of radioactivity was slower in mice than in rats, with greater concentrations excreted in feces (28% for mice versus 6% for rats). However, the mouse feces samples were often contaminated with urine-soaked feed pellets.

Endpoint:
basic toxicokinetics in vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Unspecified (results initially published in 2000)
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
test procedure in accordance with generally accepted scientific standards and described in sufficient detail
Objective of study:
toxicokinetics
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 417 (Toxicokinetics)
GLP compliance:
not specified
Radiolabelling:
yes
Remarks:
[14C]-p-tert-butylcatechol
Species:
mouse
Strain:
B6C3F1
Sex:
male
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Inc. (Raleigh, NC, and Portage, MI)
- Age at study initiation: 9 to 10 weeks old
- Weight at study initiation: No data
- Fasting period before study: No data
- Housing: Polycarbonate cages
- Individual metabolism cages: yes (from the day before dosing)
- Diet: Purina Rodent Chow No. 5002
- Water: ad libitum
- Acclimation period: 1 week


ENVIRONMENTAL CONDITIONS
Not specified


IN-LIFE DATES: Not specified
Route of administration:
oral: gavage
Vehicle:
other: 20% Emulphor in water
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
- Vehicle: 20% Emulphor in water
- Dosing volume: 5 mL/kg
Duration and frequency of treatment / exposure:
single administration
Remarks:
Doses / Concentrations:
300 mg/kg bw
No. of animals per sex per dose:
4 males
Control animals:
no
Details on study design:
- Dose selection rationale: Not specified
- Rationale for animal assignment (if not random): No data
Details on dosing and sampling:
PHARMACOKINETIC STUDY (Absorption, distribution, excretion):
- Radioactivity was measured in urine collected 6, 24, 48 and 72h after  dosing and feces collected 24, 48 and 72h after dosing (stored in dark at  -20°C until analysis).
- Tissues and body fluids sampled: adrenal gland, blood, brain, kidney,  liver, lung, prostate gland, spleen, testis with seminal vesicle, as well  as three samples each of adipose tissue, muscle and skin (analysed for  carbon-14 content). For some animals, the cecum, large and small  intestine, and stomach were also analysed.
Details on distribution in tissues:
No tissue showed high concentrations of TBC residues.
The total dose in tissues was 0.234% at 72 hours after dosing.
Details on excretion:
Approximately half of the administered dose was excreted by 24 hours and 90% by 72 hours, mostly in urine.
The relatively high concentration of radioactivity recovered in feces may have been due to urine-soaked feed pellets present in the fecal samples.
Metabolites identified:
not measured

Cumulative excretion of radioactivity (cumulative % of dose, mean +/- standard deviation):

Time (h) Urine (%) Feces (%) Total (%)
6 21.3 +/- 15.5 - (not collected) 21.3 +/- 15.5
24 40.2 +/- 18.1 14.2 +/- 6.2 54.4 +/- 15.4
48 47.6 +/- 18.6 20.5 +/- 6.2 68.1 +/- 12.8
72 50.1 +/- 18.1 25.6 +/- 9.5 75.7 +/- 10.3
Conclusions:
Bioaccumulation potential cannot be judged based on study results
The distribution and excretion of 4-tert butylpyrocatechol were studied in mice following a single oral administration of radiolabeled compound.
Executive summary:

A group of 4 mice was administered a single oral dose of 300 mg/kg14C-4-tert butylpyrocatechol (TBC) by gavage.

No tissue showed high concentrations of TBC residues.

Approximately half of the administered dose was excreted by 24 hours and 90% by 72 hours, mostly in urine. The relatively high concentration of radioactivity recovered in feces may have been due to urine-soaked feed pellets present in the fecal samples.

Endpoint:
basic toxicokinetics in vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Unspecified (results initially published in 2000)
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
test procedure in accordance with generally accepted scientific standards and described in sufficient detail
Objective of study:
toxicokinetics
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 417 (Toxicokinetics)
GLP compliance:
not specified
Radiolabelling:
yes
Remarks:
[14C]-p-tert-butylcatechol
Species:
rat
Strain:
Fischer 344
Sex:
male
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Inc. (Raleigh, NC, and Portage, MI)
- Age at study initiation: 13 to 15 weeks old
- Weight at study initiation: No data
- Fasting period before study: No data
- Housing: Polycarbonate cages
- Individual metabolism cages: yes (from the day before dosing)
- Diet: Purina Rodent Chow No. 5002
- Water: ad libitum
- Acclimation period: 1 week


ENVIRONMENTAL CONDITIONS
Not specified


IN-LIFE DATES: Not specified
Route of administration:
dermal
Vehicle:
other: acetone
Details on exposure:
TEST SITE
- Area of exposure: 4 cm²
- % coverage: Not specified
- Type of wrap if used: protective foam appliances glued onto the rats’ backs with Hollister’s medical adhesive; nonocclusive cloth cover attached to the foam appliance, and protective metal mesh cover secured over the appliance with elastoplast adhesive bandage after dosing
- Time intervals for shavings or clipplings: Clipping approx. 24 hours before dose application


REMOVAL OF TEST SUBSTANCE
- Washing (if done): No
- Time after start of exposure: up to 72 hours after dosing


TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 13.5 to 23.1 μCi [14C]-radiolabel and appropriate amount of unlabeled p-tert-butylcatechol
- concentration (if solution): Not applicable


VEHICLE
- Justification for use and choice of vehicle (if other than water): Not specified
- Amount(s) applied (volume or weight with unit): approximately 200 μL
- Concentration (if solution): Not applicable
- Lot/batch no. (if required): Not specified
- Purity: Not specified


USE OF RESTRAINERS FOR PREVENTING INGESTION: No data
Duration and frequency of treatment / exposure:
single exposure
Remarks:
Doses / Concentrations:
0.6, 6 or 60 mg/kg
No. of animals per sex per dose:
4 males
Control animals:
no
Details on study design:
- Dose selection rationale: Not specified
- Rationale for animal assignment (if not random): No data
Details on dosing and sampling:
PHARMACOKINETIC STUDY (Absorption, distribution, excretion)
- Tissues and body fluids sampled: urine, faeces.
- Time and frequency of sampling: radioactivity was measured in urine collected 6, 24, 48 and 72h after dosing and feces collected 24, 48 and  72h after dosing (stored in dark at -20°C until analysis).
- The skin at the site of application was excised with the foam appliance still attached. The appliance was removed and analysed by LSS.

METABOLITE CHARACTERISATION STUDIES:
- Urine collected during the first 24 h after dosing from rats  administered 60 mg/kg was analysed for urinary metabolites.
- Blood samples were collected in rats administered 60 mg/kg at 15 and 30  minutes and 1, 2, 4, 8 and 24 hours after dosing. Plasma concentrations  of [14C]-TBC were determined.
- Method type for identification: HPLC, GC/MS with electron impact ionisation and NMR spectroscopy.
Details on absorption:
Absorption increased with increasing dose:
- Approximately two-thirds of the 60 mg/kg dose was absorbed by 24 hours after application and more  than 80% by 72 hours.
- The 0.6 mg/kg dose was far less well absorbed.
The severity of local inflammation at the site of application increased with increasing dose; this increase in blood flow to the area may have  been responsible for the dose-related increase in absorption.

The percentage of the dose recovered in the foam appliance was greatest for the 0.6 mg/kg dose and decreased with increasing dose. The cloth appliance cover in the 6 mg/kg group contained 1.5% to 5.4% of the dose (data not shown).
Details on distribution in tissues:
Global distribution of radioactivity in tissues, at the application site, in urine and feces (% of dose) is detailed below.
Details on excretion:
Cumulative excretion of radioactivity is detailed below.
Metabolites identified:
yes
Details on metabolites:
- Urine:
HPLC analyses of urine collected from the 0.6 and 60 mg/kg groups from 0  to 6 hours after dosing indicated only polar metabolites; no parent  compound was detected in the urine.
TBC was excreted as TBC sulfate and other polar metabolites that included  predominantly sulfate conjugates. One of the metabolites was determined to  be an O-methyl-O'-sulfate of TBC.

- Plasma:
Peak plasma concentrations (in rats of the 60 mg/kg group) were observed 1 hour after application. No parent compound was detected in plasma extracts.

Cumulative excretion of radioactivity (cumulative % of dose, mean +/- standard deviation):

Dose (mg/kg) Time (h) Urine (%) Feces (%) Total (%)
0.6 1.45 +/- 1.82  - (not collected)  1.45 +/- 1.82
  24 16.3 +/- 2.4  0.363 +/- 0.135  16.7 +/- 2.6
  48 24.6 +/- 5.3  1.11 +/- 0.77  25.7 +/- 5.6
  72  31.7 +/- 7.8  1.34 +/- 0.88  33.1 +/- 8.2
6 6  7.46 +/- 2.16  - (not collected)  7.46 +/- 2.16
  24  27.4 +/- 5.7  0.641 +/- 0.221  28.1 +/- 5.9
  48  38.9 +/- 6.4  1.04 +/- 0.30  39.9 +/- 6.7
  72  46.8 +/- 6.8  1.69 +/- 0.83  48.5 +/- 6.8
60 6  12.2 +/- 16.5  - (not collected)  12.2 +/- 16.5
  24  64.3 +/- 4.3  2.43 +/- 0.67  66.7 +/- 4.2
  48  73.2 +/- 5.0  3.37 +/- 0.87  76.6 +/- 4.6
  72  79.5 +/- 5.2  4.09 +/- 1.41  83.6 +/- 4.2

Distribution of radioactivity at 72 hours (% of dose, mean +/- standard deviation):

 Dose (mg/kg)  Tissues (%)  Application site (%)  Feces (%)  Urine (%)  Total absorbed dose (%)  Total unabsorbed dose (%)
 0.6  0.35 +/- 0.11  10.2 +/- 4.3  1.34 +/- 0.88  31.7 +/- 7.8  43.7 +/- 9.8  40.4 +/- 9.8
 6  0.52 +/- 0.19  8.24 +/- 2.48  1.69 +/- 0.83  46.8 +/- 6.8  57.2 +/- 4.7  28.9 +/- 6.1
 60  0.995 +/- 0.444  2.84 +/- 0.17  4.09 +/- 1.41  79.5 +/- 5.2  87.4 +/- 4.3  2.74 +/- 0.76
Conclusions:
Bioaccumulation potential cannot be judged based on study results
The absorption, distribution, metabolism and excretion of 4-tert butylpyrocatechol were studied in rats following a single dermal application of radiolabeled compound.
Executive summary:

Groups of 4 rats were administered a single dermal application of 0.6, 6 or 60 mg/kg 14C-4-tert butylpyrocatechol (TBC).

Absorption increased with increasing dose. Approximately two-thirds of the 60 mg/kg dose was absorbed by 24 hours after application and more than 80% by 72 hours. The 0.6 mg/kg dose was far less well absorbed. The severity of local inflammation at the site of application increased with increasing dose; this increase in blood flow to the area may have been responsible for the dose-related increase in absorption.

The percentage of the dose recovered in the foam appliance was greatest for the 0.6 mg/kg dose and decreased with increasing dose. The cloth appliance cover in the 6 mg/kg group contained 1.5% to 5.4% of the dose.

At 72 hours following application, the total unabsorbed dose increased with increasing dose: from 43.7% (at 0.6 mg/kg) to 87.4% (at 60 mg/kg). Most of the excreted radioactivity was retrieved from urine (up to 79.5%).

HPLC analyses of urine collected from the 0.6 and 60 mg/kg groups from 0 to 6 hours after dosing indicated only polar metabolites; no parent compound was detected in the urine. TBC was excreted as TBC sulfate and other polar metabolites that included predominantly sulfate conjugates. One of the metabolites was determined to be an O-methyl-O'-sulfate of TBC.

Peak plasma concentrations (in rats of the 60 mg/kg group) were observed 1 hour after application. No parent compound was detected in plasma extracts.

Endpoint:
basic toxicokinetics in vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Unspecified (results initially published in 2000)
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
test procedure in accordance with generally accepted scientific standards and described in sufficient detail
Objective of study:
toxicokinetics
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 417 (Toxicokinetics)
GLP compliance:
not specified
Radiolabelling:
yes
Remarks:
[14C]-p-tert-butylcatechol
Species:
rat
Strain:
Fischer 344
Sex:
male
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Inc. (Raleigh, NC, and Portage, MI)
- Age at study initiation: 13 to 15 weeks old
- Weight at study initiation: No data
- Fasting period before study: No data
- Housing: Polycarbonate cages
- Individual metabolism cages: yes (from the day before dosing)
- Diet: Purina Rodent Chow No. 5002
- Water: ad libitum
- Acclimation period: 1 week


ENVIRONMENTAL CONDITIONS
Not specified


IN-LIFE DATES: Not specified
Route of administration:
intravenous
Vehicle:
other: 10% Emulphor in F344 rat plasma.
Details on exposure:
Doses for intravenous injection contained 6.3 to 23 μCi [14C]-radiolabel and an appropriate amount of unlabeled p-tert-butylcatechol.

The doses for rats were formulated in 10% Emulphor in F344 rat plasma at a dosing volume of 1 mL/kg.

The doses were injected into a lateral tail vein.
Duration and frequency of treatment / exposure:
single administration
Remarks:
Doses / Concentrations:
3 mg/kg
No. of animals per sex per dose:
4 males
Control animals:
no
Details on study design:
- Dose selection rationale: Not specified
- Rationale for animal assignment (if not random): No data
Details on dosing and sampling:
PHARMACOKINETIC STUDY (Absorption, distribution, excretion)
- Tissues and body fluids sampled: adrenal gland, blood, brain, kidney,  liver, lung, prostate gland, spleen, testis with seminal vesicle, as well  as three samples each of adipose tissue, muscle and skin (analysed for  carbon-14 content). For some animals, the cecum, large and small  intestine, and stomach were also analysed.
- Time and frequency of sampling: once after 72h.
Radioactivity was measured in urine collected 6, 24, 48 and 72h after  dosing and feces collected 24, 48 and 72h after dosing (stored in dark at  -20°C until analysis).
Details on distribution in tissues:
Radioactivity was measured in small amounts in various tissues . The total dose in tissues was 0.283% of injected radioactivity.
Details on excretion:
After intravenous injection, the radioactivity was rapidly excreted in  urine, with 54% of the administrated radioactivity recovered in the first  6 hours after dosing.
After 72 hours, approximately 90% of the dose had been recovered in the  urine and 6% in the feces.
Metabolites identified:
not measured
Details on metabolites:
No tissue developed high tissue/blood ratios of TBC equivalents; this  finding is consistent with the production of very polar metabolites.

Cumulative excretion of radioactivity (cumulative % of dose, mean +/- standard deviation):

Time (h) Urine (%) Feces (%) Total (%)
53.7 +/- 1.8  - (not collected) 53.7 +/- 1.8 
24 81.0 +/- 2.4 4.45 +/- 3.31 85.5 +/- 3.6
48 86.3 +/- 1.9 5.69 +/- 3.02 91.9 +/- 1.9
72 89.8 +/- 1.9 5.97 +/- 2.98 95.8 +/- 1.5
Conclusions:
Bioaccumulation potential cannot be judged based on study results
The distribution and excretion of 4-tert butylpyrocatechol were studied in rats following a single intravenous injection of radiolabeled compound.
Executive summary:

A group of 4 rats were administered a single intravenous injection of 3 mg/kg 14C-4-tert butylpyrocatechol (TBC).

Radioactivity was measured in small amounts in various tissues. The total dose in tissues was 0.283% of injected radioactivity.

After intravenous injection, the radioactivity was rapidly excreted in urine, with 54% of the administrated radioactivity recovered in the first 6 hours after dosing.

After 72 hours, approximately 90% of the dose had been recovered in the urine and 6% in the feces.

Endpoint:
basic toxicokinetics in vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Unspecified (results initially published in 2000)
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
test procedure in accordance with generally accepted scientific standards and described in sufficient detail
Objective of study:
toxicokinetics
Qualifier:
according to
Guideline:
OECD Guideline 417 (Toxicokinetics)
GLP compliance:
not specified
Radiolabelling:
yes
Remarks:
[14C]-p-tert-butylcatechol
Species:
rat
Strain:
Fischer 344
Sex:
male
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Inc. (Raleigh, NC, and Portage, MI)
- Age at study initiation: 13 to 15 weeks old
- Weight at study initiation: No data
- Fasting period before study: No data
- Housing: Polycarbonate cages
- Individual metabolism cages: yes (from the day before dosing)
- Diet: Purina Rodent Chow No. 5002
- Water: ad libitum
- Acclimation period: 1 week


ENVIRONMENTAL CONDITIONS
Not specified


IN-LIFE DATES: Not specified
Route of administration:
oral: gavage
Vehicle:
other: water or 20-80% Emulphor in water
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
- Vehicle: the 2 mg/kg dose was formulated in water, the 200 mg/kg dose  was formulated in 20% Emulphor in water, and the 1000 mg/kg dose was  formulated in 80% Emulphor in water
- Dosing volume: 5 mL/kg
Duration and frequency of treatment / exposure:
single administration
Remarks:
Doses / Concentrations:
2, 200 or 1000 mg/kg
No. of animals per sex per dose:
4 males
Control animals:
no
Details on study design:
- Dose selection rationale: Not specified
- Rationale for animal assignment (if not random): No data
Details on dosing and sampling:
PHARMACOKINETIC STUDY (Absorption, distribution, excretion):
- Radioactivity was measured in urine collected 6, 24, 48 and 72h after  dosing and feces collected 24, 48 and 72h after dosing (stored in dark at  -20°C until analysis).
- Tissues and body fluids sampled (except rats administered 1000 mg/kg):  adrenal gland, blood, brain, kidney, liver, lung, prostate gland, spleen, testis with seminal vesicle, as well as three samples each of adipose  tissue, muscle and skin (analysed for carbon-14 content). For some  animals, the cecum, large and small intestine, and stomach were also  analysed.

DETERMINATION OF PLASMA CONCENTRATION OF [14C]-TBC:
4 rats were administered a single oral dose of 200 mg/kg. Blood samples (0.3 mL) were collected before dosing and at 15 and 30  minutes and 1, 2, 4, 8 and 24 hours after dosing. The samples were  centrifuged to separate the plasma, which was analysed by liquid  scintillation spectrometry for total radioactivity.
Details on absorption:
Approximately 90% of the radiolabel was recovered, indicating a very high rate of oral absorption.
Peak plasma concentrations were observed 30 minutes after gavage.
Details on distribution in tissues:
Similarly to rats dosed intravenously, the tissues of rats given 2 or 200 mg/kg orally did not show significant concentrations of TBC-derived residues; also, very little radioactivity remained in the gastrointestinal tract tissues or contents.
The total dose in tissues was 0.18% or 0.16% at 2 or 200 mg/kg, respectively.
Details on excretion:
The overall rates and routes of excretion were similar between the 2 and 200 mg/kg doses 6 hours after dosing.
The doses were excreted primarely in urine.
For the 1000 mg/kg dose, the rate of excretion was slower than for lower doses, and a greater percentage of the radioactivity (25%) was excreted in feces.
Metabolites identified:
yes
Details on metabolites:
TBC was excreted primarily as conjugates and other polar metabolites. Incubation with beta-glucuronidase or sulfatase hydrolyzed the conjugates to parent compound and a less polar analyte. HPLC analysis of urine samples treated with purified beta-glucuronidase or purified sulfatase demonstrated that sulfatase liberated a greater proportion of the polar metabolites, indicating that sulfate conjugates constituted the majority of the polar metabolites.
No parent compound was detected in plasma extracts.
Conclusions:
Bioaccumulation potential cannot be judged based on study results
The absorption, distribution, metabolism and excretion of 4-tert butylpyrocatechol were studied in rats following a single oral administration of radiolabeled compound.
Executive summary:

Groups of 4 rats were administered a single oral dose of 2, 200 or 1000 mg/kg14C-4-tert butylpyrocatechol (TBC) by gavage.

Approximately 90% of the radiolabel was recovered, indicating a very high rate of oral absorption. Peak plasma concentrations were observed 30 minutes after gavage.

Similarly to rats dosed intravenously, the tissues of rats given 2 or 200 mg/kg orally did not show significant concentrations of TBC-derived residues; also, very little radioactivity remained in the gastrointestinal tract tissues or contents.

The overall rates and routes of excretion were similar between the 2 and 200 mg/kg doses 6 hours after dosing. The doses were excreted primarely in urine. For the 1000 mg/kg dose, the rate of excretion was slower than for lower doses, and a greater percentage of the radioactivity (25%) was excreted in feces.

 

TBC was excreted primarily as conjugates and other polar metabolites. Incubation with beta-glucuronidase or sulfatase hydrolyzed the conjugates to parent compound and a less polar analyte. HPLC analysis of urine samples treated with purified beta-glucuronidase or purified sulfatase demonstrated that sulfatase liberated a greater proportion of the polar metabolites, indicating that sulfate conjugates constituted the majority of the polar metabolites. No parent compound was detected in plasma extracts.

Endpoint:
dermal absorption in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
report date: 2005-06-24
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 428 (Skin Absorption: In Vitro Method)
Deviations:
no
Qualifier:
according to
Guideline:
other: EPA Guideline "In vitro Dermal Absorption Rate Testing of Certain Chemicals of Interest to the Occupational Safety and Health Administration", Federal Registrer: April26, 2004 (Volume 69, Number 80)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Radiolabelling:
yes
Species:
human
Sex:
male/female
Type of coverage:
occlusive
Vehicle:
unchanged (no vehicle)
Duration of exposure:
6 hours
Details on study design:
DOSE PREPARATION
- Method for preparation of dose suspensions:
FORMULATION OF [14C] 4-TERT-BUTYLPYROCATECHOL
Preparation of the Stock Solution I:
About 8 mg [14C] 4-tert-butylpyrocatechol were dissolved in 2.5 mL diethyl ether.
The concentration of radioactivity in the stock solutions was determined by Liquid Scintillation Counting (LSC).

Preparation of the Stock Solution II:
33.2 mg unlabeled 4-tert-butylpyrocatechol were dissolved in 1 mL ethanol.

Formulation of the administration solution:
A volume of the stock solution I (250 µL) containing 0.7 mg [14C] 4-tert-butylpyrocatechol and a volume of the stock solution II (100 µL) containing 3.32 mg unlabeled 4-ter-butylpyrocatechol were placed in a flask. The organic solvent was removed by a gentle stream of nitrogen.This dilution with unlabeled 4-tert-butylpyrocatechol yielded [14C] 4-tert-butylpyrocatechol with a new specific radioactivity of 775 kBq/mg. The dry test item was thereafter dissolved in 4 mL water (MilliQ) leading to the target concentration of 1 mg 4-tert-butylpyrocatechol / mL administration solution.

ANALYSIS
- Method type(s) for identification: The purity of the formulated test item was determined by HPLC at the time of each application. And the skin membrane rinse pools of each subgroup were analyzed by HPLC.
HPLC analysis were carried out on a Merck HPLC system of a solvent-pump (Merck-Hitachi L-7100), an Autosampler (Merck-Hitachi L-7200), an UV-detector (Merck-Hitachi L-4000). The radioactivity signal was monitored with a Radiomatic 500TR radioactivity flow monitor (Packard Instrument Company inc.) connected to a personal computer. The flow monitor operated with a 500 µL liquid cell and continuous mixing of the eluent with 3 mL/min scintillation liquid (Flo-Scint A13).
Column: Phenomenex Luna C18 5 µm, 250 mm x 4.6 mm
Mobile phase: Eluent A (Water MilliQ), Eluent B (Acetonitrile)
Flow: 1 mL/min
Gradient: 0 - 25 min, 40% B (isocratic)

- Limits of quantification: The limit of quantification (LOQ) in the perfusate was calculated according to Currie: LOQ accounted for 0.0019 - 0.0021 [µg 4-TBC equivalents/cm²]
Details on in vitro test system (if applicable):
SKIN PREPARATION
- Source of skin: Human cadaver skin form Caucasian donors was obtained from "Institut für Pathologie, Kantonsspital Basel", Basel, Switzerland.
- Ethical approval: yes
- Type of skin: upper leg dorsal (Q1A1, Q1A1a, Q1A2, Q2A1) and abdominal (Q1A1a, Q1A2a, Q2A1)
- Thickness of skin: 200 µm
- Membrane integrity check: yes
- Storage conditions: Upon receipt the skin was stored at about -20°C until preparation of the skin membranes. All skin samples used were almost hair-free and in good condition as visually observed.
- Justification of species, anatomical site and preparative technique: The upper leg area for withdrawal of the skin samples is the common standard at the institute of pathology. To get skin samples of the abdominal area is very difficult due to ethical aspects. However the anatomical properties for the 200 µm skin membranes were equal for both used body areas.

PRINCIPLES OF ASSAY
An automated flow-through cell system was used. Seven flow-through diffusion cells (Aesculap) designated cell 1 through cell 7 were placed in one aluminium manifold (Permegear) connected to a water bath (Digitana Ltd.) to maintain the temperature of the skin membranes at 32°C. The diffusion cells of the second part of the system with the same setup were designated cell 8 through cell 14. Each diffusion cell consisted of a donor and receptor chamber. The area of skin membrane exposed to the donor chamber was 0.64 cm². The receptor chambers were connected to a multi-channel peristaltic pump, model IPC-16 (Ismatec Ltd.). The pump speed was adjusted to about 3 mL/h. During the given time intervals, the effluent from the cell was collected directly into vials on a fraction collector, model ISCO retriever IV (IG Instrumenten Gesellschaft).

EXPERIMENTAL PROCEDURE
Pieces of the skin membranes (approximately 1.8 x 1.8 cm) were cut and mounted in the diffusion cells between the donor and receptor chamber. The cells were placed in the manifolds and connected to the peristaltic pump. For an equilibration period of 1 hour, saline (0.9% NaCl w/v) was pumped through the receptor chamber at a flow rate of about 3 mL/h.

Test of Skin Membrane Integrity:
The integrity of the skin membranes was checked by applying 50 μL tritium water (about 200,000 dpm) to the skin membrane surface. The donor chamber was covered with adhesive tape (occluded conditions). The cumulative penetration was determined over 6 hours by collecting hourly fractions. The permeability coefficient (Kp) of each skin membrane was calculated for the 3 - 6 hours interval. Skin membranes with Kp > 2.5x10-3 cm/h were excluded from the subsequent experiment.
After 6 hours, adhesive tape was removed from the donor cell chamber and the cells left open overnight with the saline flowing through the receptor chamber.

APPLICATION OF DOSE
A 64 μL aliquot of the application solution was applied manually to each skin membrane preparation using a μL-syringe. The amount applied to each cell was checked by determination of the radioactivity content of three control doses, taken prior to the first, in the middle, and after the last administration for each dose group. The doses applied and the concentration of the application solution are given in the table below:

Objective Subgroup Applied dose Concentration
µg/cell µg/cm² Dpm/cell mg/cm3
Short term penetration Q1A1 64.4 100.6 2994982 1.006
Q1A1a 63.5 99.2 2953104 0.992
Q1A2 64.4 100.7 2996111 1.007
Q1A2a 63.4 99.1 2950257 0.991
Permeability constant Q2A1 64.1 100.01 2979176 1.000

Application of the Test Item:
Those cells with skin membranes of acceptable Kp values in the integrity test were arranged on one manifold and aliquots of 64 μl dose solution were applied to the surface of each of the skin membrane. The donor chambers were covered by an adhesive tape (occluded conditions). The receptor fluid, i.e. aqueous saline (0.9% NaCl w/v), was delivered at a flow rate of about 3 mL/h during the testing period. The perfusates were collected at ambient temperature in time intervals as follows:

Objective Subgroup Sampling interval
Short term penetration Q1A1 0 – 10 minutes
Q1A1a 0 – 10 minutes
Q1A2 0 – 60 minutes
Q1A2a 0 – 60 minutes
Permeability constant Q2A1 0 – 6 hours: 1 hour intervals (6 intervals)
0 – 24 hours: 2 hour intervals (9 intervals)

End of the experimental period:
After the corresponding exposure period the cover tape was removed from the donor chamber and retained for radiometry. Thereafter test item which was still present on the skin membrane was removed from the skin surface by rinsing three times with about 0.5 ml of a mild shower gel solution (Nivea Douche Fitness) (1 % in tap water) followed by three times with 0.5 ml water (MilliQ).
The skin membrane rinse was collected for each cell for the determination of radioactivity by Liquid Scintillation Counting (LSC). The skin membranes were then removed from the diffusion cell digested in tissue solubilizer (Solvable), and the radioactivity was determined by LSC.
Finally the cells were washed with water (150 ml) and the radioactivity in the cell wash was determined by LSC.

Total recovery:
INTEGRITY TEST:
Prior to each individual application of [14C] 4-tert-butylpyrocatechol the integrity of the human skin membranes was checked using tritiated water. All skin membrane preparations were selected showing a permeability coefficient (Kp) of tritiated water < 2.5 cm/h x 10-3. For the first experiments of the short-term penetration test (Q1A1 and Q1A2) two and three cells, respectively, did not pass the integrity test, therefore additional replicates were performed designated as Q1A1a and Q1A2a.

SHORT TERM PENETRATION
After a 10-minute exposure with aqueous [14C] 4-tert-butylpyrocatechol solution only 0.04% of the applied dose penetrated through the human skin membrane. The bulk of applied test item could be removed from the skin membrane after the exposure period, accounting for 96.80% of the dose. Additional 2.14% and 0.27% of the dose were found in/on the skin membrane and in the cell wash. The total recovery amounted to 99.25% of the dose.

During the exposure time of 60 minutes a mean of 2.22% of the applied [14C] 4-tert-butylpyrocatechol penetrated through the human skin membranes. The variation of the different subjects ranged from 0.86% to 3.23% of the dose. Again the bulk of applied test item could be washed off after the end of the exposure period accounting for 85.18%. A somewhat higher amount, i.e. 6.30% of the dose, was found in/on the skin membranes after the washing procedure. The total recovery after 60 minutes of exposure accounted for 95.11% of the dose.

DETERMINATION OF THE PERMEABILITY CONSTANT (Kp)
Within an exposure period of 24 hours a mean of 90.28% of the applied [14C] 4-tert-butylpyrocatechol penetrated through human skin membranes. After a lag time of about 0.5 hours 4-tert-butylpyrocatechol penetrated with a flux (penetration rate at steady state) of 6.836 µg/cm²/h. This steady state was achieved, on average, between 1 and 10 hours after start of exposure. The variation of the individual subjects ranged from 1-8 up to 1-18 hours. After the steady state period more than 50% the applied dose were penetrated through the skin membranes and therefore a infinite dose situation was no longer given. Based on the calculated flux of 6.836 µg/cm²/h and the concentration of 4-tert-butylpyrocatechol in the administration solution of 993 µg/cm3 the permeability constant (Kp) of 4-tert-butylpyrocatechol was calculated to be 6.881 x 10-3 [cm/h]. The calculated permeability constant (Kp) showed a standard deviation of 31% for the four subjects used.



Dose:
0.10 mg/cm²
Parameter:
percentage
Absorption:
0.04 %
Remarks on result:
other: 10 minutes
Dose:
0.10 mg/cm²
Parameter:
percentage
Absorption:
2.22 %
Remarks on result:
other: 60 minutes
Dose:
0.10 mg/cm²
Parameter:
percentage
Absorption:
90.28 %
Remarks on result:
other: 24 hours
Conclusions:
[14C] 4-tert-butylpyrocatechol, applied as aqueous solution at an infinite dose of 1 mg/cm3 to human split thickness skin membranes, showed a fast penetration through the skin membranes. After a lag time of 0.5 hours 4-tert-butylpyrocatechol penetrated wih a flux of 6.836 µg/cm²/h corresponding to a permeability constant (Kp) of 6.881 x 10-3 [cm/h].
Executive summary:

The percutaneous penetration of 4 -tert-butylpyrocatechol (4 -TBC), formulated as aqueous solution, was determined in vitro using split-thickness skin membranes from human skin.

The skin membranes were set up in flow-through diffusion cells and [14C] 4 -tert-butylpyrocatechol was applied onto the skin membranes at a concentration of 1 mg/cm3. Aliquots of 64 µL of the administration solution were applied to a skin membrane area of 0.64 cm² corresponding to a infinite dose of 0.1 mg 4 -TBC/cm². The in vitro percutaneous penetration was investigated in two groups, i.e. Groups Q1 and Q2. The Group Q1 was designed to investigate the short term dermal penetration and Group Q2 was designed to determine the permeability constant (Kp) for dermal penetration of 4 -tert-butylpyrocatechol. For each objective, i.e. short term penetration (10 and 60 min) and determination of the permeability constant, duplicates of valid skin membranes from at least 3 different individuals were used. The integrity of each skin membrane preparation was checked with tritiated water.

Objective                      Subgroup       Sampling interval

Short term penetration    Q1A1              0 - 10 minutes

Q1A1a           0 - 10 minutes

Q1A2 0 - 60 minutes

Q1A2a 0 - 60 minutes

Permeability constant Q2A1 0 - 6 hours: 1 hour interval (6 intervals)

0 - 24 hours: 2 hours interval (9 intervals)

The results of the short term experiments revealed that 4 -TBC penetrated to a moderate extent through human skin membranes within the first hour of exposure. Only 0.04% and 2.22% of the applied dose penetrated through the skin membrane within 10 and 60 minutes of exposure, respectively. The bulk of test item could be washed off at the end of the exposure period amounting to 96.80% and 85.18% of the dose. After skin membrane rinse 2.14% and 6.30% of the dose remained in/on the skin membranes after 10 and 60 minutes of exposure.

Within a exposure period of 24 hours in mean 90.28% of the applied [14C] 4 -TBC penetrated through human skin membranes. After a lag time about 0.5 hours 4 -TBC penetrated with a flux (penetration rate at steady state) of 6.836 µg/cm²/h through human skin membranes. The corresponding permeability constant Kp for 4 -TBC was calculated to be 6.881 x 10 -3 cm/h with a standard deviation of 31% (range 4.75 to 9.01) for the subjects used (n = 4).

HPLC analysis of the skin membrane rinse pools revealed that 4 -TBC remained unchanged on the skin membranes during the exposure period.

In conclusion, 4 -tert-butylpyrocatechol, applied as aqueous solution to human skin membranes, penetrated very fast with a penetration rate of 6.836 µg/cm²/h after a lag time of 0.5 hours.

The permeability constant Kp was calculated to be 6.881 x 10 -3 cm/h.

Description of key information

Key value for chemical safety assessment

Additional information

The absorption, distribution, metabolism, and excretion of 4-tert-butylpyrocatechol following intravenous injection, gavage dosing, or dermal application were determined in male F344/N rats and B6C3F1 mice using radiolabeled compound. The skin absorption properties of 4 -tert-butylpyrocatechol, including Kp (permeability constant) determination, were also determined in an in vitro study on human skin.

The absorption of14C-4 -tert-butylpyrocatechol (TBC) following gavage dosing or dermal application was high. The percent absorption following dermal application increased with increasing dose. Peak concentrations of14C-TBC equivalents in plasma were reached 1 hour after gavage dosing (200 mg/kg) and 2 hours after dermal application (60 mg/kg); no parent compound was detected in the plasma extracts.

Regarding administration by dermal route, three doses were tested: 0.6, 6 and 60 mg/kg. The severity of local inflammation at the site of application increased with increasing dose. The increase in blood flow to the area may have been responsible for the dose-related increase in absorption.

Regardless of route of administration, TBC-derived radioactivity was readily excreted in the urine and was markedly nonpersistent in the tissues.

TBC was excreted as 4 -tert-butylpyrocatechol sulfate and other polar metabolites that included predominately sulfate conjugates; it was not excreted as the parent compound. One metabolite was determined to be an O-methyl-O'-sulfate of TBC.

In vitro, 4 -tert-butylpyrocatechol applied as an aqueous solution to human skin membranes, penetrated very fast with a penetration rate of 6.836 µg/cm²/h after a lag-time of 0.5 hours. The permeability constant Kp was calculated to be 6.881 x 10 -3 cm/h.