Phenol, 2,4-bis[(dodecylthio)methyl]-6-methyl-

Repeated dose toxicity: oral

Administrative data

Endpoint:

sub-chronic toxicity: oral

Type of information:

migrated information: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: OECD guideline study under GLP. A default reliability of 2 is assigned for read-across according to ECHA guidance.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1989

Report date:

1989

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)

GLP compliance:

yes

Test material

Test material information

Constituent 1

Details on test material:

- Physical state: yellow liquid
- Lot/batch No.: EN 123 8l4.72
- Storage condition of test material: room temperature

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Centre Elevage Charles River (76410 Saint-Aubin-les-Elbeuf, France)
- Age at study initiation: approx. 6 weeks
- Weight at study initiation: mean body weight was 198 g for the males and 151 g for the females.
- Housing: in suspended wire-mesh cages and each cage contained 2 rats from the same sex and the same group.
- Diet: free access to A04 C pelleted diet (U.A.R., 91360 Villemoisson-sur-Orge, France) distributed weekly.
- Water: free access to bottles containing filtered tap water.
- Acclimation period: 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 ± 2°C
- Humidity (%): 50 ± 20%
- Air changes (per hr): about 13 cycles/hour of filtered, not recycled air
- Photoperiod (hrs dark / hrs light): 12 h dark/12 h light

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

other: the vehicle was a solution of water for injectable preparations with 0.5% carboxymethylcellulose and 0.1% Tween 80

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS: the test substance was diluted in the vehicle and homogenized by a magnetic stirrer.

Analytical verification of doses or concentrations:

not specified

Duration of treatment / exposure:

at least 92 days and at most 93 days

Frequency of treatment:

once a day, approximately at the same daily time (in the morning), 7 days per week

Doses / concentrations

Remarks:

Doses / Concentrations:
0, 10, 100, 1000 mg/kg bw
Basis:
nominal in water

No. of animals per sex per dose:

0 and 1000 mg/kg bw: 10
10 and 100 mg/kg bw: 20

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: dose levels were based on the results of a previously conducted study: the test item was administered daily by gavage for 28 days at doses of 0, 10, 50, 250 and 1000 mg/kg/day.

- Post-exposure recovery period in satellite groups: 28 days

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: during the study, clinical signs were recorded for each animal, at least once a day, at approximately the same daily time. All animals were checked at least twice a day for possible mortality.

DETAILED CLINICAL OBSERVATIONS: No data

BODY WEIGHT: Yes
- Time schedule for examinations: the bodyweight of each animal was recorded before the time of allocation of animals into groups, on the first day of treatment and then once a week until the end of the study.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
The quantity of food consumed by the animals of each cage was recorded once a week for a period of 7 days until the end of the study.

FOOD EFFICIENCY:
- Efficiency of food utilization was calculated once a week for each sex and each group using bodyweight and food consumption means until the end of the study.


OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: before the start of treatment and in week 13.
- Dose groups that were examined: all surviving males and females from the control and high dose level groups.

HAEMATOLOGY: Yes
- Time schedule for collection of blood: in week 13
- Anaesthetic used for blood collection: Yes (light ether anaesthesia)
- Animals fasted: No data
- How many animals: all surviving animals
- Parameters examined: Erythrocytes (RBC), Haemoglobin (HB), Mean Cell Volume (MCV), Packed Cell Volume (PCV), Mean Cell Haemoglobin Concentration (MCHC), Mean Cell Haemoglobin (MCH), Leucocytes (WBC), Thrombocytes (PLAT), Differential White Cell Count: neutrophils (N), eosinophils (E), basophils (B), lymphocytes (L), monocytes (M), Quick time (QT), Activated Partial Thromboplastin Time (APTT).

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: week 13 and at the end of the recovery period (week 17)
- Animals fasted: No data
- How many animals: all surviving animals
- Parameters examined: Sodium (Na+), Potassium (K+), Chloride (Cl-), Calcium (Ca++), Inorganic Phosphorus (I.PHOS), Glucose (GLUC), Urea (UREA), Creatinine (CREAT), Total bilirubin (TOT.BIL), Total proteins (PROT), Cholesterol (CHOL), Triglycerides (TRIG), Alkaline phosphatase (ALP), Aspartate aminotransferase (ASAT), Alanine aminotransferase (ALAT).

URINALYSIS: Yes
- Time schedule for collection of urine: in week 13
- Metabolism cages used for collection of urine: Yes
- Animals fasted: Yes, 18 h
- Parameters examined: volume (VOLUME), appearance (APP), pH (PH), specific gravity (SP.GRAV), urobilinogen (UROB), proteins (PROT), glucose (GLUC), ketones (CETO), bilirubin (BILI), blood (BLOOD), nitrites (NITR). Microscopy of deposit after centrifugation: investigation for leucocytes (WBC), erythrocytes (RBC), hyaline (HYAL) and granular (GRAN) cylinders, magnesium ammonium phosphate (MAM.PH.), calcium phosphate (CAL.PH) and calcium oxalate (CAL.OX) crystals, epithelial (EPITH.), bladder (BLAD) and kidney (KIDN) cells.

NEUROBEHAVIOURAL EXAMINATION: No data

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
Organ weights: For all animals, the following organs were weighed wet as soon as possible after dissection: brain, spleen, adrenals, testes, liver, kidneys, ovaries. The animals were weighed before necropsy. Paired organs were weighed separately.

Macroscopic examination: Aorta, caecum, brain including medulla/pons, cerebellar and cerebral cortex, heart, colon, duodenum, stomach, femoral bone with articulation (*),liver, mammary glands (*), submaxillary glands (*), sublingual glands (*), Harderian glands (*), pituitary, ileum, jejunum, spinal cord, cervical, thoracic and lumbar sections (*),sciatic nerve, skeletal muscle (*),lymph nodes, mandibular and mesenteric esophagus, ovaries, pancreas, skin (*), lungs with bronchia, prostate (*),spleen, rectum, kidneys, sternum with bone marrow, adrenals, testes and epididymides, thymus, thyroids and parathyroids, trachea, uterus, horns, and cervix, vagina (*), seminal vesicles (*),urinary bladder, eyes (*) with optic nerve and lacrymal glands.


HISTOPATHOLOGY: Yes
Microscopic examinations was performed on: all macroscopic lesions and tissues listed above (except those marked by (*)) in all animals from the high dose level and control groups sacrificed at the end of the treatment and recovery periods, in all animals that died.
All macroscopic lesions and lungs, liver, kidneys for all animals from the low and intermediate dose level groups.

Statistics:

The following sequence was used for the statistical tests of the clinical parameters (bodyweight, food consumption and water consumption) and for the haematological and biochemical parameters, and for organ weights: A normal distribution of values in the samples was checked by Kolmogorov-Smirnov test (1948), In the case of an abnormal distribution, this test was performed after the logarithmic transformation of the values.
If a significant heterogeneity persists after the logarithmic transformation of the values, the analysis of variances was not performed, A comparison between the treated groups and the control group in order to prove a treatment-related difference was made by using Mann-Whitney's test (1947).

In the case of the normal distribution of values according to the normal law an analysis of variances was made by the Bartlett's test (1937) (more than 2 samples) or Fisher's test (1934) (2 samples),

A comparison between the treated and the control groups in order to prove a treatment-related difference was made by either: Dunnett's test (1955) if no significant heterogeneity of the variances was established, Mann-Whitney's test (1947) in the case of significant heterogeneity of the variance.

Results and discussion

Results of examinations

Details on results:

CLINICAL SIGNS AND MORTALITY
1) Clinical signs
Hypersalivation after treatment was observed in 6/20 males and 6/20 females of the 1000 mg/kg/day group (observed daily from day 17 to day 22, except in one female, from day 31 to day 43). This clinical sign was considered treatment-related.
Additional clinical signs were noted, including:
- Area of hair loss on the head, neck or back of some animals of all groups, and swollen ear in 1 male of the control group, probably due the prehension of animals at test substance administration.
- Damaged eye in 1 male of the control group, due to a traumatism at blood sampling.
- Chromorhinorrhea (from day 7I to day 119) in 1 male of the 1000 mg/kg/day group, considered to be spontaneous without any treatment relationship,
- A palpable mass on 1 female of the 100 mg/kg/day group.

2) One male animal died in the control group on day 119, one male (after 11 days) and one female (after 36 days) died in the 100 mg/kg bw dose group. However, no clinical signs were observed preceding the death, and, at necropsy, no relevant macroscopically findings were found. The observed mortality either occurred accidentally during blood sampling or as consequence of test substance aspiration.

BODY WEIGHT AND WEIGHT GAIN
The bodyweight gain in all treated females and in males of the 10 and 100 mg/kg/day groups was similar to that of the control animals. A very slight increase in the bodyweight gain was observed in males of the 1000 mg/kg/day group, when compared with the control animals. The difference in bodyweight was in mean 6% and was not statistically significant. It was still noted at the end of the recovery period.
Although this change was very slight, the relationship to the treatment cannot be ruled out.

FOOD CONSUMPTION
The food consumption in all treated females and in males of the 10 and 100 mg/kg/day groups was comparable to that of the control animals. In males of the 1000 mg/kg/day group, it was very slightly increased throughout the treatment period. This change was in mean 3% and was occasionally statistically significant. This could be associated with the very slight increase in bodyweight gain noted in the same animals.
Throughout the recovery period, the food consumption was normal and similar to that of the control animals except in Week 17 when the mean value of food consumption in controls was lower than that recorded in the same animals on the previous weeks.

FOOD EFFICIENCY
No change of toxicological significance was observed.

OPHTHALMOSCOPIC EXAMINATION
The ophthalmological examination performed on animals of each sex from the control and high dose level groups revealed only a few minor changes often observed in the laboratory rat (cornea vacuolisation, persistence of the hyaloid vessel, diffuse opacity of the cornea and punctual opacity of the posterior cortical). Therefore, these changes were not considered to be treatment-related.

HAEMATOLOGY
No perceptible changes were found in the haematological parameters and the individual values were within the normal range of the background data for the animal species used.

CLINICAL CHEMISTRY
The variations observed in some biochemical parameters (sodium, potassium, chloride, inorganic phosphorus, creatine, total protein, cholesterol, triglyceride, transaminases and proteinogram) were considered to be of no toxicological significance, although they were sometimes statistically significant (they were minor, not dose-related and the individual values still within the normal range of the background data of the testing laboratory).

URINALYSIS
No treatment-related abnormalities were found.

ORGAN WEIGHTS
The statistical analysis of organ weights of the animals sacrificed at the end of the treatment period revealed the following changes:
- An increase in absolute {22%) and relative (14%) liver weight in the males at the high dose level (1000 mg/kg/day);
- An increase in relative liver weight in the females at the mid-dose (8%) and high dose (9%) levels.
The following statistical significant variations were found in the organ weights of the animals sacrificed at the end of the recovery period:
- An increase in absolute {22%) and relative (12%) liver weight in the test substance treated males;
- A decrease (11%) in relative testes weight in the test substance treated males,
No perceptible changes were found in the other organ weights.

GROSS PATHOLOGY
The macroscopic changes found at autopsy in the animals, at the end of the treatment and recovery periods, were recognized as those commonly recorded findings in the laboratory rat, therefore they were considered to be of no toxicological significance.

HISTOPATHOLOGY: NON-NEOPLASTIC
The microscopic findings encountered in all organs examined, at the end of the treatment and the recovery periods, were similar as those commonly recorded changes in the laboratory rat. Moreover, their incidence, severity and morphological characters were comparable between control and treated animals; therefore they bear no relationship to treatment. In the absence of any relevant histopathological findings in the liver and testes, the slight above mentioned variations in those organs weight were considered to be of no toxicological significance.

Effect levels

1

Dose descriptor:

LOEL

Remarks:

(NOAEL)

Effect level:

1 000 other: mg/kg bw/day

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: hypersalivation in some animals of the high dose level; very slight increase in the bodyweight gain related to an increase of the same intensity in the food consumption in the males of the high dose level

2

Dose descriptor:

NOEL

Effect level:

10 other: mg/kg bw/day

Target system / organ toxicity

Critical effects observed:

not specified