2-tert-butyl-4-methoxyphenol

Repeated dose toxicity: oral

Administrative data

Endpoint:

sub-chronic toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Justification for type of information:

Data is from peer reviewed journal

Data source

Reference

Reference Type:

publication

Title:

Repeated dose oral toxicity study of the test chemical

Author:

Liang et al

Year:

2014

Bibliographic source:

The Journal of Toxicological Sciences

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

other: Refer below principle

Principles of method if other than guideline:

Repeated dose oral toxicity study was performed to determine the toxic nature of the test chemical

GLP compliance:

not specified

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

2-tert-butyl-4-methoxyphenol

EC Number:

204-442-7

EC Name:

2-tert-butyl-4-methoxyphenol

Cas Number:

121-00-6

Molecular formula:

C11H16O2

IUPAC Name:

2-tert-butyl-4-methoxyphenol

Test material form:

solid: bulk

Details on test material:

- Name of test material: tert-butyl-4-methoxyphenol
- Common name: Phenol, (1,1-dimethylethyl)-4-methoxy-
- Molecular formula: C11H16O2
- Molecular weight: 180.2454 g/mol
- Smiles notation: COc1ccc(O)c(c1)C(C)(C)C
- InChI=1S/C11H16O2/c1-11(2,3)9-7-8(13-4)5-6-10(9)12/h5-7,12H,1-4H3
- Substance type: Organic

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

male

Details on test animals or test system and environmental conditions:

Details on test animal
TEST ANIMALS
- Source: Chongqing Medical University Lab Animal Center (Chongqing, China)
- Age at study initiation: 8 weeks
- Weight at study initiation: No data available
- Fasting period before study: No data available
- Housing: Animals were evenly distributed into 16 cages in controlled environment and marked with a number on their ears using non-toxic ink. All care and experimental procedures for the rats were conducted in accordance with guidelines published in the Guide for the Care and Use of Laboratory Animals of Chongqing Medical University.
- Diet (e.g. ad libitum): Standard laboratory rodent food, ad libitum (?)
- Water (e.g. ad libitum): Tap water, ad libitum (?)
- Acclimatization period: One week

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22°C (± 2°C)
- Humidity (%): 50% (± 20%)
- Air changes (per hr): No data available
- Photoperiod (hrs dark / hrs light): 12-hr light/dark cycle

IN-LIFE DATES: From: To: No data available

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

peanut oil

Details on oral exposure:

Details on oral exposure
PREPARATION OF DOSING SOLUTIONS: BHA is poorly water soluble and to make it soluble, BHA was mixed with peanut oil to provide consistent absorption and was then subjected to sonication for 30 min at 40°C. This was performed on a weekly basis. The daily reagent dosage (50 mg/kg/day) was determined individually for each rat based on body weight.

DIET PREPARATION
- Rate of preparation of diet (frequency): No data available
- Mixing appropriate amounts with (Type of food): No data available
- Storage temperature of food: No data available

VEHICLE
- Justification for use and choice of vehicle (if other than water): Peanut oil (20 mg/mL)
- Concentration in vehicle: BHA: 50 mg/kg/day, and B(a)P-BHA combination group: B(a)P 2 mg/kg/day + BHA 50 mg/kg/day.
- Amount of vehicle (if gavage): No data available
- Lot/batch no. (if required): No data available
- Purity: No data available

Analytical verification of doses or concentrations:

not specified

Duration of treatment / exposure:

90 days

Frequency of treatment:

Daily

Doses / concentrations

Remarks:

Doses / Concentrations:
0, 50 BHA or 2 B(a)P +50 BHA mg/kg/day
Basis:
no data

No. of animals per sex per dose:

Total: 72
0 mg/kg/day : 24 male
50 mg/kg/day : 24 male
2 (B(a)P) + 50 (BHA) mg/kg/day: 24 male

Control animals:

yes, concurrent vehicle

Details on study design:

No data

Positive control:

No data

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: No data available
- Time schedule: No data available
- Cage side observations: No data available

DETAILED CLINICAL OBSERVATIONS: Yes, swimming ability of each rat was observed.
- Time schedule: Prior to the Morris Water Maze (MWM) test.

BODY WEIGHT: Yes
- Time schedule for examinations: No data available

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): No data available
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: No data available
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No data available

FOOD EFFICIENCY: No data available
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: No data available

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No data available
- Time schedule for examinations: No data available

OPHTHALMOSCOPIC EXAMINATION: Yes, visual ability was observed.
- Time schedule for examinations: Prior to the MWM test.
- Dose groups that were examined: All dose groups were examined.

HAEMATOLOGY: No data available
- Time schedule for collection of blood: No data available
- Anaesthetic used for blood collection: No data available
- Animals fasted: No data available
- How many animals: No data available
- Parameters checked in table [No.?] were examined. No data available

CLINICAL CHEMISTRY: No data available
- Time schedule for collection of blood: No data available
- Animals fasted: No data available
- How many animals: No data available
- Parameters checked: No data available

URINALYSIS: No data available
- Time schedule for collection of urine: No data available
- Metabolism cages used for collection of urine: No data available
- Animals fasted: No data available
- Parameters checked: No data available

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: Prior to the MWM test.
- Dose groups that were examined: All 72 dose groups animals were examined.
- Battery of functions tested: sensory activity / grip strength / motor activity / other: Yes, motor activity was tested.

OTHER: No data available

Sacrifice and pathology:

GROSS PATHOLOGY: No data available

HISTOPATHOLOGY: No data available

Other examinations:

Following the WMW test, malonaldehyde (MDA) content and superoxide dismutase (SOD) activity were measured to determine the oxidative damage induced by B(a)P and the protective effect of BHA.

The activity of Na+-K+-ATPase and Ca2+-Mg2+-ATPase was also measured after the MWM to determine the energy metabolism damage induced by B(a)P and the protective effect of BHA.

In addition, following the WMW test, the Ca2+-content was measured to detect the damage in the hippocampus induced by B(a)P and the protective effect of BHA.

Statistics:

All data analyses were performed with SPSS v.20.0 (SPSS, Inc., Chicago, IL, USA) and the quantitative data were expressed as mean ± S.D. The data were analyzed using analysis of variance (ANOVA). Data obtained over training days from the hidden platform trial were analyzed by ANOVA for repeated measurement. The remaining data were analyzed by one-way ANOVA. When appropriate, post hoc comparisons were assessed using the Least
Significant Difference (LSD). A probability of P < 0.05 was considered significant for all analyses.

Results and discussion

Results of examinations

Clinical signs:

not specified

Mortality:

not specified

Body weight and weight changes:

not specified

Food consumption and compound intake (if feeding study):

not specified

Food efficiency:

not specified

Water consumption and compound intake (if drinking water study):

not specified

Ophthalmological findings:

no effects observed

Description (incidence and severity):

No change were observed in visual ability of treated rats as compare to control.

Haematological findings:

not specified

Clinical biochemistry findings:

not specified

Urinalysis findings:

not specified

Behaviour (functional findings):

no effects observed

Description (incidence and severity):

No change were observed in motor activity of treated rats as compare to control.

Immunological findings:

not specified

Organ weight findings including organ / body weight ratios:

not specified

Gross pathological findings:

not specified

Neuropathological findings:

not specified

Histopathological findings: non-neoplastic:

not specified

Histopathological findings: neoplastic:

not specified

Other effects:

not specified

Details on results:

BHA was chosen to prevent the neurotoxicity on oxidative stress caused by B(a)P.
In the B(a)P-BHA-combination group, the rats had a better performance significantly in MWM compared to the rats in the B(a)P group. This demonstrates that in behavior tests BHA has protective action against the harmful effect of B(a)P.
The MDA level of rats in B(a)P-BHA-combination group is lower, and SOD activity is higher than in the B(a)P-group. This is a further sign of the positive effect against B(a)P damage.

The activity of ATPase has a significant improvement, which indicates that BHA also plays a positive role in energy metabolism.
There is no significant difference of Ca2+ content between the B(a)P group and the B(a)P-BHA-combination group.

Effect levels

Dose descriptor:

NOAEL

Effect level:

50 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male

Basis for effect level:

other: Effect on motor activity, and levels of MDA and SOD, ATPase activity and Ca2+ concentration in rat brain.

Target system / organ toxicity

Critical effects observed:

not specified

Applicant's summary and conclusion

Conclusions:

The no observed adverse effect level (NOAEL) was considered to be 50 mg/kg body weight/ day in male Sprague-Dawley rat when exposed to the test chemical.

Executive summary:

In a 90 days repeat dose toxicity study, male Sprague-Dawley rats were exposed to the test chemical by oral gavage in the concentrations of 0 or 50 mg/kg/day. The results showed only effect in Morris Water Maze (Hidden platform MWM and Probe trial), MDA content and SOD activity when exposed to 50 mg/kg/day the test chemical alone. In combination with 2 mg/kg/day B(a)P, the results showed an improved performance in MWM compared to the rats in the B(a)P group, demonstrating that the test chemical has a protective action against the harmful effect of B(a)P. The MDA level of rats in B(a)P-test chemical-was lower, and SOD activity was higher than in the B(a)P-group, thus also showing positive effect against B(a)P damage. The activity of ATPase improved significantly in B(a)P-test chemical-treated rats, which indicates that the test chemical plays a positive role in energy metabolism. Therefore, no observed adverse effect level (NOAEL) was considered to be 50 mg/kg body weight/ day in male Sprague-Dawley rat when exposed to the test chemical by oral gavage on a daily basis for 90 days.