Oligomerisation products of beta-pinene

Endpoint summary

Description of key information

A reliable sub-acute (28-day) oral toxicity test (OECD 407) in the rat found only minor liver changes (organ enlargement and minimal centrilobular hepatocyte enlargement) after dosing at 1000 mg/kg/day. These changes were considered to represent adaptive changes related to hepatic metabolism of the test substance and the NOAEL was set as 1000 mg/kg/day.

Further to this, a reliable sub-chronic (90-day) oral toxicity test (OECD 408) indicated the substance was well tolerated in rats at levels of 1000 mg/kg/day. non-adverse alterations were observed in a few hematology and clinical chemistry parameters, in the thyroid and the liver. The NOAEL was also set as 1000 mg/kg/day.

 

Key value for chemical safety assessment

Toxic effect type:

dose-dependent

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Reference

Endpoint:

sub-chronic toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.26 (Sub-Chronic Oral Toxicity Test: Repeated Dose 90-Day Oral Toxicity Study in Rodents)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.3100 (90-Day Oral Toxicity in Rodents)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Species:

rat

Strain:

Wistar

Remarks:

Crl: WI(Han)

Details on species / strain selection:

Standard species/strain for toxicology testing.

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Deutschland, Sulzfeld, Germany
- Females (if applicable) nulliparous and non-pregnant: yes
- Age at study initiation: 8 to 9 weeks.
- Weight at study initiation: Males weighed between 225 to 305 g and females 154 to 198 g.
- Fasting period before study: Not applicable.
- Housing: Polycarbonate cages containing sterilized sawdust as bedding material equipped with water bottles. 5 animals of the same sex and same dosing group together. During locomotor activity monitoring, animals are housed individually in a Hi-temp polycarbonate cage without cage-enrichment, bedding material, food and water. The room(s) in which the animals were kept was documented in the study records. Cages were arranged on the racks according to a Latin-square model.
- Diet (e.g. ad libitum): SM R/M-Z from SSNIFF pelleted diet provided ad libitum except during designated procedures.
- Water (e.g. ad libitum): Municipal tap water provided ad libitum except during designated procedures. :

DETAILS OF FOOD AND WATER QUALITY: With regards to food, results of analysis for nutritional components and environmental contaminants were provided by the supplier and are on file at the Test Facility. It is considered that there were no known contaminants in the feed that would interfere with the objectives of the study. Periodic analysis of the water was performed, and results of these analyses are on file at the Test Facility. It is considered that there were no known contaminants in the water that could interfere with the outcome of the study.
- Acclimation period: The animals were allowed to acclimate to the Test Facility toxicology accommodation for 28 days before the commencement of dosing.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 to 21°C
- Humidity (%): 46 to 58%
- Air changes (per hr): At least 10 air changes per hour
- Photoperiod (hrs dark / hrs light): 12 hours light/dark cycle (except during designated procedures).

IN-LIFE DATES: From: To: 05 December 2019 to 06 March 2020.

Route of administration:

oral: gavage

Details on route of administration:

Standard as per OECD guidelines.

Vehicle:

corn oil

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS: Test item dosing formulations (w/w) were homogenized to visually acceptable levels at appropriate concentrations to meet dose level requirements. For Weeks 1-2, the dosing formulations were prepared daily as a solution and dosed within 6 hours after preparation of the formulation. From Week 3 onwards, the dosing formulations were prepared weekly, filled out in daily portions and stored in the refrigerator protected from light and stirred for at least 30 minutes before dosing. A factor of 0.935 was used to correct for the specific gravity of the test item. If practically possible, the dosing formulations and vehicle were continuously stirred until and during dosing.

VEHICLE
- Justification for use and choice of vehicle (if other than water): Corn oil is a standard vehicle for repeated dose toxicity studies and was deemed to be suitable for the test substance based on trial formulation and analytical work.
- Concentration in vehicle: 0, 20, 60 and 200 mg/mL.
- Amount of vehicle (if gavage): 5 mL/Kg

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Analysis was performed using a validated analytical procedure.

Accuracy and homogeneity were determined for formulations prepared for use in Week 1, Week 6, Week 9 and Week 13.

The chemical analyses were conducted as specified below. Samples and remaining formulation were stored in normal glassware causing the samples stored at room temperature to be exposed to normal laboratory light conditions.

Duplicate samples (approximately 500 or 630 mg), which were taken from the formulations using a pipette, were accurately weighed into volumetric flasks of 50 mL. For determination of accuracy, samples were taken at middle position (50% height) or at top, middle and bottom position (90%, 50% and 10% height). The samples taken at 90%, 50% and 10% height were also used for the determination of the homogeneity of the formulations. The volumetric flasks were filled up to the mark with tetrahydrofuran. When necessary, the solutions were further diluted with 1% corn oil in tetrahydrofuran to obtain concentrations within the calibration range.

Results:

Accuracy: A small response at the retention time of the test item was observed in the chromatograms of the Group 1 formulation prepared for use in Week 6. The maximum contribution to the Group
2 samples was 11%. Therefore, an additional analysis of Group 1 was performed in Week 9. In all other formulations of Group 1, no test item was detected. The concentrations analyzed in the formulations of Group 2, Group 3 and Group 4 were in agreement with target concentrations (i.e. mean accuracies between 90% and 110%).

Homogeniety: The formulations of Group 2 and Group 4 were homogeneous (i.e. coefficient of variation ≤ 10%).

Duration of treatment / exposure:

7 days a week for a minimum of 13 weeks.

Frequency of treatment:

Once daily

Dose / conc.:

0 mg/kg bw/day (nominal)

Dose / conc.:

100 mg/kg bw/day (nominal)

Dose / conc.:

300 mg/kg bw/day (nominal)

Dose / conc.:

1 000 mg/kg bw/day (nominal)

No. of animals per sex per dose:

10 males and 10 females per dose.

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: The dose levels were selected based on information provided from a 28 day repeated dose toxicity study with oral exposure in Sprague Dawley rats, and in an attempt to produce graded responses to the test item. In the 28-day study the NOAEL was set on 1000 mg/kg/day and the NOEL on 150 mg/kg/day. The high-dose level should produce some toxic effects, but not excessive lethality that would prevent meaningful evaluation. The mid-dose level was expected to produce minimal to moderate toxic effects. The low-dose level should produce no observable indications of toxicity.
- Rationale for animal assignment (if not random): Animals were randomly assigned to groups. Males and females were randomized separately.
- Fasting period before blood sampling for clinical biochemistry: Yes (overnight with a maximum of 24 hours).

Positive control:

Not a requirement of the test guideline

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: At least once daily starting from Day 1 at 0 to 1-hour post-dose during dosing. Animals were observed within their cage unless necessary for identification or confirmation of possible findings.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Weekly; from at least Week 1 and throughout the study, and on the day of necropsy. Animals were removed from the cage.

BODY WEIGHT: Yes
- Time schedule for examinations: Weekly; from at least Day 1 and throughout the study. Fasted weight on the day of necropsy.

FOOD CONSUMPTION:
- Time schedule for examinations: Weekly; from at least Day 1 and throughout the study. Quantitatively measured per cage.

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): Yes
- Time schedule for examinations: Regular basis throughout the study. Water consumption was monitored by visual inspection of the water bottles.

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: Pretreatment Period - All Study animals once (including spare animals) Dosing Period - All Group 1 and 4 Study animals during Week 13. The eyes were examined using an ophthalmoscope after application of a mydriatic agent.

HAEMATOLOGY: Yes
- Time schedule for collection of blood: End of treatment
- Anaesthetic used for blood collection: Yes (isoflurane)
- Animals fasted: Yes (overnight with a maximum of 24 hours)
- How many animals: All animals
- Parameters examined: White blood cell count, neutrophils, lymphocytes, monocytes, easinophils, basophils, large unstained cells, red blood cell count, reticulocytes, red blood cell distribution width, hemoglobin, hematocrit, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration and platelets. Coagulation parameters including prothrombin time and activated partial thromboplastin time were also included.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: End of treatment
- Animals fasted: Yes (overnight with a maximum of 24 hours)
- How many animals: All animals
- Parameters examined: Alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, total protein, albumin, total bilirubin, urea, creatinine, glucose, cholesterol, triglycerides, HDL and LDL cholesterol, sodium, potassium, chloride, calcium, inorganic phosphate, Triiodothyronine, thyroxine, and thyroid stimulating hormone.

URINALYSIS: Yes
- Time schedule for collection of urine: Urine was collected into a specimen vial from animals housed in individual metabolism cages overnight (approximately 15-20 hours)
- Metabolism cages used for collection of urine: Yes
- Animals fasted: Yes (overnight with a maximum of 24 hours)
- Parameters examined: Volume, specific gravity, clarity, color, pH, blood, leukocyte esterase, bilirubin, urobilinogen, protein, ketones, glucose and nitrite.

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: Once during the Dosing Period. The first 5 animals per sex per group during Week 12-13.
- Dose groups that were examined: All groups
- Battery of functions tested: Hearing, pupillary reflex and static righting reflex, fore and hind limb grip strength and locomotor activity using a computerized monitoring system.

IMMUNOLOGY: No

ESTROUS STAGE DETERMINATION: Yes
- Time schedule for examinations: End of Treatment - on the day of necropsy, a vaginal smear was taken to determine the stage of estrus from all Study animals. This was done for all females.
- Parameters examined: Estrous stage

Sacrifice and pathology:

GROSS PATHOLOGY: Study animals were subjected to a complete necropsy examination, which included evaluation of the carcass and musculoskeletal system; all external surfaces and orifices; cranial cavity and external surfaces of the brain; and thoracic, abdominal, and pelvic cavities with their associated organs and tissues.

ORGAN WEIGHTS: The following organs were weighed (paired organs were weighed together): Brain, epididymis, adrenal gland, pituitary gland, prostate gland, seminal vesicle gland, thyroid, heart, kidney, liver, ovary, spleen, testis, thymus, uterus/cervix. Organ to body weight ratio (using the terminal body weight) were calculated.

HISTOPATHOLOGY: The following were examined by a pathologist: Aorta, sternum, brain, cervix, epididymis, esophagus, eye, adrenal gland, mammary gland, parathyroid, pituitary, prostate gland, seminal vesicle, gross lesions/masses, heart, kidney, cecum, colon, rectum, liver, lung, mandibular and mesenteric lymph node, skeletal muscle, optic nerve, sciatic nerve, ovary, pancreas, ski, duodenum, ileum, jejunum, spinal cord, spleen, stomach, testis, thymus, trachea, urinary bladder, uterus and vagina.

Statistics:

Variables for inferential analysis: Body weight, body weight gains, hematology variables, coagulation variables, clinical chemistry variables, urinalysis variables, organ weights and organ weight relative to body weight.

Parametric/Non-parametric: Levene’s test was used to assess the homogeneity of group variances. The groups were compared using an overall one-way ANOVA F-test if Levene’s test was not significant or the Kruskal-Wallis test if it was significant. If the overall F-test or Kruskal-Wallis test was found to be significant, then pairwise comparisons were conducted using Dunnett’s or Dunn’s test, respectively.

ANCOVA: The data corresponding to a response variable of interest and to a related covariate were submitted to an analysis of covariance (ANCOVA), including only groups with at least three non-missing paired values and if found to be significant, then pairwise comparisons were conducted using Dunnett’s test.

Parametric: Datasets with at least 3 groups (the designated control group and 2 other groups) were compared using Dunnett-test (many-to-one-t-test).
For the motor activity data set (at least 3 groups) parametric (ANOVA) tests on group means were applied with Bonferroni correction for multiple testing. Mixed modelling techniques, comparing six different covariance structures, were used in order to select the best fitting statistical model.

Non-parametric: Datasets with at least 3 groups were compared using a Steel-test (many-to-one rank test).

Incidence: An overall Fisher’s exact test was used to compare all groups. The above pairwise comparisons were conducted using Fisher’s exact test whenever the overall test was significant.

Clinical signs:

no effects observed

Description (incidence and severity):

No relevant test item-related clinical signs were observed in the treated groups.

Salivation after dosing was observed throughout the study period in males at 100, 300 and 1000 mg/kg/day and in females at 300 and 1000 mg/kg/day at a dose-related response. The salivation was considered not toxicologically relevant, taking into account the nature and minor severity of the effect and its time of occurrence (i.e. after dosing). This sign was considered to be a physiological response related to taste of the test item rather than a sign of systemic toxicity.

In two males at 1000 mg/kg/day, labored breathing, abnormal breathing sounds, hunched posture, erected fur and/or red colored discharge from the muzzle was observed on Day 24. As this only occurs on one day, it was considered to be not test item-related and likely to be caused by the method of dosing.

Any other clinical signs noted during the Dosing Period occurred within the range of background findings to be expected for rats of this age and strain which are housed and treated under the conditions in this study and did not show any apparent dose-related trend. At the incidence observed, these were considered to be unrelated to treatment with the test item.

Mortality:

no mortality observed

Description (incidence):

No mortality occured during the study period.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

Body weights and body weight gain of treated males remained in the same range as controls over the study period.

Any statistically significant changes in body weight gains in males and females were considered to be unrelated to treatment with the test item, since no trend was apparent regarding dose and time.

Food consumption and compound intake (if feeding study):

no effects observed

Description (incidence and severity):

No effects on food consumption were observed in any of the test item-treated groups.

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

no effects observed

Description (incidence and severity):

There were no test item-related ophthalmology findings in Week 13.

The nature and incidence of ophthalmology findings noted during the Pre-treatment Period and in Week 13 was similar among the groups and occurred within the range considered normal for rats of this age and strain. These findings were therefore considered to be unrelated to treatment with the test item.

Haematological findings:

effects observed, non-treatment-related

Description (incidence and severity):

Hematology:
No test item-related hematology parameter changes were observed in males at 100 and 300 mg/kg/day and in females at 100 mg/kg/day.

In females at 300 and 1000 mg/kg/day, a dose-dependent increase in red blood cell distribution width (RDWG; 1.04x and 1.07x of controls, respectively) was observed. These changes remained within the historical control data. In addition in females at 1000 mg/kg/day, a decrease in mean corpuscular volume (MCV; 0.97x of controls) and mean corpuscular hemoglobin (MCH; 0.96x of controls) was seen.

In females at 1000 mg/kg/day, an increase in neutrophil counts were observed (1.67x of controls). As this change remained well within the historical control data , it was considered to be not toxicologically relevant.

In males at 1000 mg/kg/day, an increase in white blood cell, neutrophil, lymphocyte and monocyte counts were observed in all tests item-groups. At the severity observed and in absence of a dose-related response, these changes were considered to be not test item-related. In one male at 300 mg/kg/day, moderate echinocytes and severe platelet clumps and in one male at 1000 mg/kg/day severe platelet clumps were also observed. As this was only seen in individual animals and lacked a dose-related response, these findings were considered to be not test item-related.

Other values in treated males and females achieving a level of statistical significance, when compared to controls, were considered to have arisen as a result of slightly high or low control values, occurred in the absence of a dose-related distribution and/or were, given the magnitude of change, considered to be of no toxicological significance.

Coagulation:
No test item-related effects on coagulation parameters were observed.
The statistically significant lower prothrombin time (PT) of females treated at 100 and 1000 mg/kg/day (0.95x and 0.94x of control, respectively) was considered to be of no toxicological relevance as no clear dose-related response was observed.

Clinical biochemistry findings:

effects observed, non-treatment-related

Description (incidence and severity):

No test item-related effects on clinical chemistry parameters were observed in males up to 1000 mg/kg/day.

No test item-related effects on clinical chemistry parameters were observed in males up to 1000 mg/kg/day.

In females total protein (TPROT) and albumin (ALB) concentrations were increased while total bilirubin (TBIL) concentration was decreased in all test item treated groups at a dose-related response. Furthermore, an increase in LDL cholesterol was observed in females at 1000 mg/kg/day. Calcium (CA) concentration was also increased in females at 300 and 1000 mg/kg/day.

For thyroid hormones an increase in TSH concentration was noted in females at 300 and 1000 mg/kg/day.

For Triiodothyronine (T3) only two results were available in males of the control group, as values of the diluted samples were below the reportable range and due to hemolysis. In order to have a proper evaluation, historical control data was used to complete the evaluation. For males at 1000 mg/kg/day, T3 results were available for five out of ten males. As these results were within the historical control range and also sufficient data was available on thyroxine (T4) and thyroid-stimulating hormone (TSH), a complete evaluation could still be made. No effects on T3, T4 and TSH were seen in males at 1000 mg/kg/day.

Other values achieving a level of statistical significance, when compared to controls, were considered to have arisen as a result of slightly high or low control values, occurred in the absence of a dose-related distribution and/or were, given the magnitude of change, considered to be of no toxicological significance.

Urinalysis findings:

no effects observed

Description (incidence and severity):

No test item-related effects on urine parameters were observed in any of the test item-treated groups.

Behaviour (functional findings):

no effects observed

Description (incidence and severity):

Hearing ability, pupillary reflex and static righting reflex were normal in all examined animals. Grip strength and motor activity was similar between control and high dose animals. All groups showed a similar motor activity habituation profile with a decreasing trend in activity over the duration of the test period.

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

Statistically significant, test item-related higher liver weights (absolute and/or relative to body weights) were noted in males and females starting at 100 mg/kg/day and higher thyroid gland weights (absolute and relative to body weights) were noted in males at 1000 mg/kg/day.

The microscopic correlate for the increased liver weights in both sexes at 1000 mg/kg/day was considered centrilobular hepatocellular hypertrophy. The microscopic correlate for the increased thyroid gland weights in males at 1000 mg/kg/day was considered follicular cell hypertrophy.

The statistically significant higher absolute kidney weights in females at 100 and 1000 mg/kg/day was considered to be of no toxicological significance as the kidney relative to body weight was unchanged, it occurred in the absence of a dose-related trend and remained within the range considered normal for rats of this age and strain. There were no other test item-related organ weight changes.

Gross pathological findings:

no effects observed

Description (incidence and severity):

There were no test item-related gross observations.

All the recorded macroscopic findings were within the range of background gross observations encountered in rats of this age and strain.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Histopathological findings: neoplastic:

not examined

Description (incidence and severity):

Test item-related microscopic findings were noted in both sexes in the liver starting at 1000 mg/kg/day and in the thyroid glands at 100 mg/kg/day.

In the liver, minimal centrilobular hepatocellular hypertrophy was recorded in 6/10 males and 6/10 females at 1000 mg/kg/day at minimal degree.

In the thyroid gland, an increased incidence and/or severity (up to mild degree) of hypertrophy of follicular cells was recorded in males and females starting at 100 mg/kg/day, which was in males at 300 mg/kg/day and in males and females at 1000 mg/kg/day combined with increased incidence and severity (up to mild degree) of colloid alteration.

The minimal follicular cell hypertrophy as recorded in the control groups and the colloid alteration in control and 100 mg/kg/day in both sexes and in females 300 mg/kg/day was considered within background pathology for rats of this age and strain.

There were no other test item-related histologic changes. The remainder of the recorded microscopic findings were within the range of background pathology encountered in rats of this age and strain. There was no test item related alteration in the prevalence, severity, or histologic character of those incidental tissue alterations.

Other effects:

not examined

Details on results:

Test formulations prepared were considered homogeneous at the concentrations tested and analysis of the accuracy revealed acceptable levels. A small response at the retention time of the test item was observed in the chromatograms of the Group 1 formulation prepared for use in Week 6. As the concentration measured was only very minimal (3.1% of the mean concentration of Group 2) and was not observed in Weeks 1 and 13, this Week 6 result had no impact on the study.

At hematology, a higher red blood cell distribution width was observed in females at 300 and 1000 mg/kg/day in combination with a lower mean corpuscular volume and mean corpuscular hemoglobin in females at 1000 mg/kg/day. As these changes were only slight, remained within historical control range and lacked a histopathological correlation, they were considered to be not adverse.

At clinical chemistry, an increase in total protein and albumin concentration and a decrease in total bilirubin concentration in females at all treated groups were observed, at a dose-related response. Furthermore, an increase in LDL cholesterol concentration in females at 1000 mg/kg/day and in calcium concentration in females at 300 and 1000 mg/kg was seen. In absence of a histopathological correlation, these changes were considered to be not adverse.

For thyroid hormones an increase in TSH concentration was noted in females at 300 and 1000 mg/kg/day, which might correlate to the microscopically hypertrophy of follicular cells in the thyroid gland.
At histopathological examination test item-related effects were observed in the liver and thyroid gland.

In the liver, centrilobular hepatocellular hypertrophy (minimal degree) was noted in both sexes at 1000 mg/kg/day, which correlated with higher liver weights. The hypertrophy was without any changes in liver enzyme activity or significant morphologic degenerative changes microscopically, and was therefore interpreted to be non-adverse.

In the thyroid gland, an increased incidence and/or severity (up to mild degree) of hypertrophy of follicular cells was recorded in males and females starting at 100 mg/kg/day, which was in males at 300 mg/kg/day and males and females at 1000 mg/kg/day combined with increased incidence and severity (up to mild degree) of colloid alteration. This finding correlated with the higher thyroid gland weight in male at 1000 mg/kg/day. The increased incidence and severity in test item-treated animals compared to control animals was in absence of any inflammatory or degenerative changes and were therefore considered non-adverse at the severities (minimal or mild degree) recorded.

There were no test item-related or toxicologically significant changes noted in any of the remaining parameters investigated in this study (i.e. clinical signs, body weight, food consumption, functional observations, ophthalmoscopy, coagulation parameters, urinalysis and gross pathology).

Dose descriptor:

NOAEL

Effect level:

1 000 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Remarks on result:

not determinable due to absence of adverse toxic effects

Critical effects observed:

no

Full study summary reports can be found in the attachment in 'Attached background material'.

Ratio clinical chemistry differences from control groups

Groups	2	3	4
Dose (mg/kg/day)	100	300	1000
Sex	F	F	F
TPROT	1.07	1.07	1.12
ALB	1.06	1.07	1.12
TBIL	0.83	0.68	0.75
LDL	1.31	1.29	1.54
CA	1.03	1.06	1.11
TSH	1.00	2.77	3.90

F = Females, TPROT= total protein, ALB = albumin, TBIL= total bilirubin, LDL = LDL cholesterol, CA = calcium, TSH = thyroid stimulating hormone.
Numerical values indicate fold change of the treated group mean value relative to the control group mean value. Bolded values indicate the mean value was statistically different from controls at P</=0.05 or P</=0.01.

Historical control data for thyroid hormones in Wistar Han rats (males): T3 (ng/dL) : Mean 0.64; P5 to P95 = 0.41 to 0.95 (n=60)

 

Mean percent organ weight differences fron control groups

 

	Males			Females
Dose level (mg/kg/day):	100	300	1000	100	300	1000
LIVER
Absolute	4	17	19	20	23	39
Relative to body weight	8	18	20	15	21	37
THYROID GLAND
Absolute	13	30	41	-	-	-
Relative to body weight	18	32	43	-	-	-
Bolded values indicate the mean value was statistically different from controls at P</=0.05 or P</=0.01. - : No statistically significant change

 

Summary of test-item related microscopic findings:

	Males				Females
Dose level (mg/kg/day):	0	100	300	1000	0	100	300	1000
LIVER*	10	10	10	10	10	10	10	10
Hypertrophy, hepatocellular,            centrilobular
Minimal	-	-	-	6	-	-	-	6
THYROID GLANDS*	10	10	10	10	10	10	10	10
Hypertrophy follicular cell
Minimal	3	6	6	5	1	4	5	6
Mild	-	1	1	5	-	-	2	4
Alteration colloid
Minimal	1	3	3	2	1	2	2	4
Mild	-	-	2	3	-	-	-	3

* = Number of tissues examined from each group.

Conclusions:

Administration of the test substance by once daily oral gavage was well tolerated in rats at levels of 1000 mg/kg. Non-adverse alterations were observed in a few hematology and clinical chemistry parameters, in the thyroid gland and in the liver.

From the results a No Observed Adverse Effect Level (NOAEL) for the substance was at least 1000 mg/kg/day.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

1 000 mg/kg bw/day

Study duration:

subchronic

Species:

rat

Quality of whole database:

Reliable guideline studies available

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - local effects

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Comparison of the acute oral toxicity of pinene oligomers to rats via different routes of administration suggests no clear difference in toxicity, and based on toxicokinetic data for close chemical analogues extensive absorption from the gastro-intestinal tract followed by relatively rapid hepatic metabolism and bioelimination is expected. The evidence of adaptive hepatic change in rats dosed at 1000 mg/kg/day for 28 or 90 days supports this. From the available subchronic study data, a NOAEL value of 1000 mg/kg/day was found.

Justification for classification or non-classification

Low toxicity has been demonstrated for pinene oligomers with NOAEL values from 28 and 90-day repeated dose studies set as the limit dose of 1000 mg/kg/day. Accordingly, no classification for specific target organ toxicity (STOT) repeated-dose toxicity applies.