Registration Dossier

Administrative data

Description of key information

In a combined OECD 422/408 study at 1000 mg/kg bw/d decreased T4 levels (for females in combination with increased incidence of follicular cell hypertrophy in the thyroid) at 1000 mg/kg, of which adversity could not be excluded was observed.

 

In females treated at 1000 mg/kg bw/d microscopic examination revealed an increased incidence of follicular cell hypertrophy (minimal or slight) in the thyroid. This was accompanied by a decrease of thyroid hormone T4 (on average 19%). In males treated at 1000 mg/kg serum levels of thyroid hormone T4 were also decreased (on average by 57%), unaccompanied by treatment related changes in thyroid weight or morphology. For both males and females no corroborative findings were observed in TSH levels. 

 

These findings, referring to Draft Guidance for the identification of endocrine disruptors in the context of Regulations (EU) No 16 528/2012 and (EC) No 1107/2009 (December 2017), may indicate a hazard for human thyroid insufficiency in adults as well as pre- and post-natal neurological development of offspring. An OECD 426 is proposed as a follow up study to further evaluate the potential neurodevelopmental effects.

Key value for chemical safety assessment

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
Study period:
2017
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
Version / remarks:
2016
Deviations:
no
Qualifier:
according to
Guideline:
OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity in Rodents)
Version / remarks:
2008
Deviations:
no
Qualifier:
according to
Guideline:
other: EPA OPPTS 870.3650 Combined Repeated Dose Toxicity Study with the Reproduction /Developmental Toxicity Screening Test
Version / remarks:
2000
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.26 (Sub-Chronic Oral Toxicity Test: Repeated Dose 90-Day Oral Toxicity Study in Rodents)
Version / remarks:
2008
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.3100 (90-Day Oral Toxicity in Rodents)
Version / remarks:
1998
Deviations:
no
Principles of method if other than guideline:
In addition, the procedures described in this study plan essentially conform to the following guidelines:
• OECD 421, Reproduction/Developmental Toxicity Screening Test, 2016.
• EPA OPPTS 870.3550, Reproduction/Developmental Toxicity Screening Test, 2000.
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
other: Crl:WI(Han) (outbred, SPF-Quality)
Details on species / strain selection:
Untreated, nulliparous, non-pregnant females and untreated males were used at the initiation of the study.
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Deutschland, Sulzfeld, Germany
- Females (if applicable) nulliparous and non-pregnant: yes
- At initiation of dosing males and females were 6 weeks old, males weighed between 154 and 187 g and females weighed between 120 and 145 g.
- Fasting period before study: no
- Housing: Pretest and pre-mating: animals were housed in groups of 5 animals/sex/cage in Macrolon plastic cages (MIV type, height 18 cm); Mating: Males and females were cohabitated on a 1:1 basis in Macrolon plastic cages (MIII type, height 18 cm); Post-mating: Males were housed in their home cage (Macrolon plastic cages, MIV type, height 18 cm) with a maximum of 5 animals/cage. Females were individually housed in Macrolon plastic cages (MIII type, height 18 cm); Lactation: Females were housed in Macrolon plastic cages (MIII type, height 18 cm). Pups were housed with the dam, except during locomotor activity monitoring of the dams, when the pups were kept warm in their home cage using bottles filled with warm water. In order to avoid hypothermia of pups, pups were not left without their dam or a bottle filled with warm water for longer than 30-40 minutes.
- Diet: Free access to pelleted rodent diet (SM R/M-Z from SSNIFF® Spezialdiäten GmbH, Soest, Germany) (During motor activity measurements, animals did not have access to food for a maximum of 2 hours)
- Water: Free access to tap-water (During motor activity measurements, animals did not have access to water for a maximum of 2 hours)
- Acclimation period: for 7 days prior to start of treatment;

DETAILS OF FOOD AND WATER QUALITY:
Diet, water, bedding and cage-enrichment/nesting material evaluation for contaminants and/or nutrients was performed according to facility standard procedures. There were no findings that could interfere with the study.

ENVIRONMENTAL CONDITIONS
(set to maintain)
- Temperature (°C): 20 to 22
- Humidity (%): 43 to 66
- Air changes (per hr): at least 10
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 26 April 2017 To: 02 August 2017
Route of administration:
oral: gavage
Vehicle:
polyethylene glycol
Remarks:
specific gravity 1.036
Details on oral exposure:
- PREPARATION OF DOSING SOLUTIONS:
Formulations (w/w) were prepared daily as solution (low and mid dose) or suspension (high dose) within 5 hours prior to dosing and were homogenized to a visually acceptable level. Test item dosing formulations were kept at room temperature until dosing. If practically possible, the dosing formulations and vehicle were continuously stirred until and during dosing.
Adjustment was made for specific gravity of the vehicle. No correction was made for the purity/composition of the test item.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Analyses of samples of formulations taken in week 1, 5 and 13 during the treatment phase were analysed according to a validated method. The samples were analyzed for homogeneity (highest and lowest concentration) and accuracy of preparation (all concentrations). The accuracy of preparation was considered acceptable if the mean measured concentrations were in agreement with the target concentrations (i.e. 90-110% for the low and mid dose and 85-115% for high dose). Homogeneity was demonstrated if the coefficient of variation was ≤ 10%.
Stability of formulations over 5 hours at room temperature under normal laboratory light conditions (concentration range 1-200 mg/mL) was determined as part of the analytical method development and validation study.
Duration of treatment / exposure:
Males were treated for 92 days, i.e. 8 weeks prior to mating, during mating, and up to the day prior to scheduled necropsy.
Females that delivered were treated for 93-98 days, i.e. during 8 weeks prior to mating, the variable time to conception, the duration of the pregnancy and 14-15 days after delivery up to and including the day before scheduled necropsy.
Females which failed to deliver healthy offspring were treated for 85-96 days.
Routinely, females that are littering are left undisturbed. The omission of one day of dosing over a period of several weeks was considered not to affect the toxicological evaluation.
Pups were not treated directly but were potentially exposed to the test item in utero, via maternal milk or from exposure to maternal urine/feces.
Frequency of treatment:
Once daily for 7 days per week
Dose / conc.:
100 mg/kg bw/day (actual dose received)
Dose / conc.:
300 mg/kg bw/day (actual dose received)
Dose / conc.:
1 000 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle
Details on study design:
Dose selection rationale: Based on the results of a 14-day dose range finding study the dose levels for this combined 90-day oral gavage study with reproduction/developmental toxicity screening test were selected to be 100, 300 and 1000 mg/kg bw/day. In this DRF no mortality or no abnormalities were noted at 500 and 1000 mg/kg bw/day.
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: At least twice daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: At least once daily from start of treatment onwards up to the day prior to necropsy

BODY WEIGHT: Yes
- Time schedule for examinations: Males and females were weighed on the first day of treatment (prior to first dosing) and weekly thereafter. Mated females were weighed on Days 0, 4, 7, 11, 14, 17, 19 and 20 postcoitum and during lactation on PND 1, 4, 7 and 13.

FOOD CONSUMPTION: Yes
- Weekly, except for males and females which were housed together for mating and for females without evidence of mating. Food consumption of mated females was measured on Days 0, 4, 7, 11, 14, 17 and 20 post-coitum and during lactation on PND 1, 4, 7 and 13

WATER CONSUMPTION: Yes
Subjective appraisal was maintained during the study, but no quantitative investigation was introduced as no treatment related effect was suspected.

OPHTHALMOSCOPIC EXAMINATION: Yes
The eyes were examined using an ophthalmoscope after application of a mydriatic agent prior to initiation of treatment in all animals, and towards the end of the treatment period in Week 13 in all males and during lactation in all females.


HAEMATOLOGY: Yes
- Time schedule for collection of blood: Blood of F0-animals was collected on the day of scheduled necropsy. Blood of F1 was collected on PND 4 and PND 13-15 if possible.
- Anaesthetic used for blood collection: Yes (isoflurane)
- Animals fasted: Yes for F0, o/n (with a maximum of 24h)
- How many animals: all F0 animals; 2 pups per litter of F1
- Parameters checked according to Guidelines

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Blood of F0-animals was collected on the day of scheduled necropsy. Blood of F1 was collected on PND 4 and PND 13-15 if possible.
- Anaesthetic used for blood collection: Yes (isoflurane)
- Animals fasted: Yes for F0,, o/n (with a maximum of 24h)
- How many animals: all F0 animals; 2 pups per litter of F1
- Parameters checked according to Guidelines. Included thyroid hormone analyzed (T4 and TSH).

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: Yes, FOB
- Time schedule for examinations:
The selected males were tested during Week 13 of treatment and the selected females were tested once during the last week of lactation (e.g. PND 8-12). These tests were performed after observation for clinical signs (incl. arena observation, if applicable).
- Dose groups that were examined:
5 animals/sex/group
- Battery of functions tested: hearing ability, pupillar reflex and static righting reflex, fore- and hind-limb grip strength, locomotor activity.

IMMUNOLOGY: No

OTHER: reproduction and developmental parameters, plasma thyroid hormone analysis, see 7.8
Sacrifice and pathology:
GROSS PATHOLOGY: Yes, according to Guidelines
HISTOPATHOLOGY: Yes, according to Guoidelines
Other examinations:
Organ weights according to Guidelines
Statistics:
All statistical tests were conducted at the 5% significance level. All pairwise comparisons were conducted using two sided tests and were reported at the 1% and or 5% levels.
Numerical data collected on scheduled occasions for the listed variables were analyzed as indicated according to sex and occasion. Descriptive statistics number, mean and standard deviation (or % CV or SE when deemed appropriate) were reported whenever possible. Inferential statistics were performed according to the comparison matrix below when possible, but excluded semi-quantitative data, and any group with less than 3 observations.
The following pairwise comparisons were made:
Group 2 vs. Group 1
Group 3 vs. Group 1
Group 4 vs. Group 1
Datasets with at least 3 groups (the designated control group and 2 other groups) were compared using Dunnett-test (many-to-one-t-test).
Datasets with at least 3 groups were compared using a Steel-test (many-to-one rank test).
The motor activity data set was compared using an overall Kruskal-Wallis.
An overall Fisher’s exact test was used to compare all groups at the 5% significance level. The above pairwise comparisons were conducted using Fisher’s exact test whenever the overall test is significant.
Clinical signs:
effects observed, non-treatment-related
Description (incidence and severity):
No treatment-related clinical signs were noted during daily detailed clinical observations or during weekly arena observations.
Salivation seen among animals treated with the test item was considered to be a physiological response rather than a sign of systemic toxicity considering its slight severity and the time of occurrence (i.e. after dosing).
Any other clinical signs noted during the treatment 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.
Mortality:
mortality observed, non-treatment-related
Description (incidence):
Mortality occurred in two incidences only; the premature death of male no. 26 (300 mg/kg) and female no. 80 (1000 mg/kg) was considered unrelated to treatment with the test item.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Compared to controls, body weight gain of male rats was reduced, to about the same extent, at 300 and 1000 mg/kg from start treatment onwards, resulting in about 10% lower mean body weights from treatment Day 36 onwards. Statistical significance was achieved in most weeks of the pre-mating period at 1000 mg/kg and occasionally at 300 mg/kg. To a lesser extent, a similar trend was noted in males treated at 100 mg/kg.
Body weights and body weight gain of females were considered not to be affected by treatment. The slightly higher mean body weights noted in 1000 mg/kg females at most time points (up to 6%, not statistically significant) were considered to represent normal biological variation and, therefore, regarded as unrelated to treatment.
Food consumption and compound intake (if feeding study):
effects observed, non-treatment-related
Description (incidence and severity):
Food consumption (before and after correction for body weight) was considered not to be affected by treatment.
For females at 1000 mg/kg a statistically significant decreased relative food consumption was noted on Days 4-7 of lactation. This was caused by the lower food consumption of female no. 75, which was related to her small litter size (i.e. one pup).
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
effects observed, non-treatment-related
Description (incidence and severity):
No ophthalmology findings were noted that were considered to be related to treatment.
The nature and incidence of ophthalmology findings noted during pretest 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, treatment-related
Description (incidence and severity):
A statistically significantly lower mean number of platelets was noted at 1000 mg/kg in males (relative difference from controls 17%). Individual values in 1000 mg/kg males generally remained within the historical control range .
There were no treatment-related changes in red and white blood cell parameters. The few statistically significant variations noted in females (higher mean values for total white blood cells, lymphocytes and red blood cell distribution width at 100 or 300 mg/kg) were considered unrelated to treatment due to the lack of a dose-related response.

A statistically significantly higher mean prothrombin time (PT) was noted at 1000 mg/kg in males (relative difference from controls 5%). Values in 1000 mg/kg males remained within the historical control range (and mostly also in the concurrent control range). There were no treatment-related changes in activated partial thromboplastin time (APTT).

Historical control data for the number of platelets (10E9/L) in male Wistar Han rats
Mean: 741, P5- 95: 566–900 (n=91; period 2014 - July 2016)
Historical control data for prothrombin time (s) in male Wistar Han rats
Mean: 17.1, P5-P95: 15.6-18.9 (n=91, period 2014 - July 2016)

Clinical biochemistry findings:
effects observed, treatment-related
Description (incidence and severity):
The following statistically significant changes, all at 1000 mg/kg in males, distinguished treated animals from control animals. Relative changes in mean values compared to the control group are indicated between parentheses.
• Higher (26%) plasma activity of aspartate aminotransferase (ASAT)
• Higher (55%) plasma activity of alkaline phosphatase (ALP).
• Higher albumin (7%)
• Higher total bilirubin (50%)
• Lower cholesterol (31%).
Mean values for ASAT, ALP and bilirubin in treated males remained within the historical control ranges, those for albumin and cholesterol were slightly out of these ranges .
The other (mostly statistically significant) variations noted in clinical biochemistry values were considered unrelated to treatment due to the lack of a dose-related response (albumin, creatinine and sodium in 100 and/or 300 mg/kg treated males) or an abnormal value in a single animal (high bile acids in male no. 40 at 1000 mg/kg; values for other liver-related clinical chemistry parameters were also relatively high in this male). Lower ASAT levels in treated females (all dose groups) was related to the relatively high mean control value, due to an abnormal high values in two control females (no. 45 and 49).

Historical control data for male Wistar Han rats (n=94, period 2014 – July 2016)
ASAT (U/L) mean: 75.2, P5-P95: 63.3-92.5
ALP (U/L) mean: 137, P5-P95: 82-246
Albumin (g/L) mean: 32.1, P5-P95: 30.8-33.9
Total bilirubin (umol/L) mean: 2.2, P5-P95: 1.6-3.0
Cholesterol (mmol/L) mean: 1.98, P5-P95: 1.57-2.41

Serum T4 levels in F0-males and F0-females were statistically significantly reduced (by 57% and 19%, respectively) at 1000 mg/kg. The statistically significantly lower (about 20%) mean T4 value noted in males at 100 mg/kg was considered unrelated to treatment due to the lack of a dose response (mean T4 at 300 mg/kg was similar to the control mean). Moreover, individual T4 values at 100 mg/kg generally remained within the concurrent and historical control range .
There were no treatment-related changes in serum TSH levels of both F0-males and F0-females. Overall, mean values of males and females at 100 and 300 mg/kg and males at 1000 mg/kg appeared higher than concurrent control mean values. Due to the relatively high standard deviations of these groups and a lack of a dose response, the higher mean values of the treated animals were considered as not toxicologically relevant. The statistically significantly higher (about four times higher than the control value) mean TSH value noted at 300 mg/kg was considered unrelated to treatment as the increase was caused by abnormal values in two single males (nos. 22 and 29).

Historical control data for T4 values in Wistar Han rats from OECD 422 studies
Males mean: 4.65, P5-P95: 2.970-6.440 (n=448; period 2015-2017)
Females mean: 3.57, P5-P95: 2.183-5.250 (n=40; period 2015-2017)
Note: males within historical control data were 14 weeks, whereas males within the current study were about 19 weeks old at time of blood sampling.
Urinalysis findings:
not examined
Behaviour (functional findings):
effects observed, non-treatment-related
Description (incidence and severity):
Hearing ability, pupillary reflex and static righting reflex were normal in all examined animals.
Grip strength was considered not to be affected by treatment. The statistically significant differences noted in hind limb grip strength (lower mean values in males at 100 and 1000 mg/kg and in females at 300 mg/kg) showed no clear dose-related response and were not accompanied by corroborative changes in other measures in the neuromuscular domain (including fore limb grip strength, gait, static righting reflex and motor activity). Moreover, the difference from controls was slight and all values remained within normal limits. Therefore, these findings were regarded as unrelated to treatment.
The variation in motor activity did not indicate a relation with treatment. All groups showed a similar 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):
Organ weights were considered not to be directly affected by treatment.
The statistically significant organ weight differences noted in 1000 mg/kg males (13% higher relative testes weight and 15% lower absolute prostate weight) were considered secondary to the (test item-related) lower body weights of these males (10% lower mean terminal body weight). Body weights of 300 mg/kg males were decreased to a similar degree but the changes in their testes and prostate weights (which were slightly smaller than those at 1000 mg/kg) were not statistically significant.
The other statistically significant variations noted in organ weight values for males (brain, thymus and adrenals in males at 100 and/or 300 mg/kg) were considered unrelated to treatment due to the lack of a dose-related response.
No treatment-related effects were observed in organ weights of females up to 1000 mg/kg.
Gross pathological findings:
effects observed, non-treatment-related
Description (incidence and severity):
There were no test item-related gross observations.
A slightly higher incidence of irregular surface of the stomach was noted in 3/10, 2/10, 4/10 and 1/9 females in the control, 100, 300 and 1000 mg/kg groups, respectively. This finding was considered unrelated to treatment due to the lack of a dose response.
All of the remaining recorded macroscopic findings were within the range of background gross observations encountered in rats of this age and strain. These findings were therefore considered to be unrelated to treatment.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
In the thyroid gland, an increased incidence of follicular cell hypertrophy was present in 1000 mg/kg females (up to slight).
In the urinary bladder, an increased incidence of hyperplasia/hypertrophy of the urothelium was present in 1000 mg/kg females (minimal). The urothelial change was characterized by a mixture of hyperplasia of the basal cell layer and/or hypertrophy of the dome-shaped surface cells (umbrella cells). The cells of the urothelium formed a uniform thickness (diffuse) without prominent focal outward or inward growth and without cellular atypia. There was no inflammation or cell death involved in this process.
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.
Histopathological findings: neoplastic:
effects observed, non-treatment-related
Description (incidence and severity):
A finding of note was observed in an ovary of 1000 mg/kg/day treated female (no.78) consisting of a tubular adenoma (also known as tubulostromal adenoma). Since there were no other test item-related proliferative findings observed in the ovaries, this incidental finding was considered as unrelated to the test item.
Other effects:
not examined
Details on results:
In females treated at 1000 mg/kg microscopic examination revealed an increased incidence of follicular cell hypertrophy (minimal or slight) in the thyroid. This was accompanied by a decrease of thyroid hormone T4 (on average 19%). In males treated at 1000 mg/kg serum levels of thyroid hormone T4 were also decreased (on average by 57%), unaccompanied by treatment related changes in thyroid weight or morphology. For both males and females no corroborative findings were observed in TSH levels. Based on the minimal or slight increase in follicular cell hypertrophy in combination with a decrease in T4 levels in 1000 mg/kg treated females and the magnitude of decreased T4 levels in 1000 mg/kg treated males, adversity could not be excluded.
Exposure to the test item up to 1000 mg/kg was not associated with obvious toxicity in the remaining parental parameters examined in this study (i.e. mortality, clinical appearance, functional tests, ophthalmoscopy, body weight, food consumption, clinical pathology, macroscopic examination and organ weights). Other changes noted in treated rats, mostly at 1000 mg/kg, were considered non-adverse as discussed below.
Slight salivation observed after dosing among treated animals (all dose levels) was considered a physiological response rather than a sign of systemic toxicity.
Male rats administered the test item showed reduced body weight gain from start treatment onwards, which was not accompanied by reduced food consumption. This occurred to a similar degree at 300 and 1000 mg/kg, and to a lesser degree at 100 mg/kg. The resulting reduction in mean body weights was modest (about 10% from Day 36 onwards at 300 and 1000 mg/kg) and the growth rate of treated males had returned to control levels after five weeks of treatment. Therefore, the decreased body weights of treated male rats were regarded as non-adverse within the context of this study.
A further treatment-related morphological change observed in females treated at 1000 mg/kg consisted of an increased incidence of minimal hyperplasia/hypertrophy of the urothelium of the urinary bladder. There was no inflammation or cell death involved in this process. Therefore, this minimal change was considered as non-adverse (Frazier et al., 2012).
Clinical pathology showed several changes in males treated at 1000 mg/kg: higher values for aspartate aminotransferase (ASAT), alkaline phosphatase (ALP), total bilirubin and albumin; and lower values for cholesterol. Additionally, the increased prothrombin time (PT) for males at 1000 mg/kg could be related to the decrease in the number of platelets. These changes were not accompanied by adverse anatomic pathology alterations and the mean values for these parameters in 1000 mg/kg males generally remained within the range considered normal for rats of this strain and age (see results section). As such, these clinical pathology changes were regarded as non-adverse (Palazzi et al., 2016).

Frazier, KS, Seely, JC, Hard, GC, Betton, G, Burnett, R, Nakatsuji, S, Nishikawa, A, Durchfeld-Meyer, B, Bube A (2012). Proliferative and Non-proliferative Lesions of the Rat and Mouse Urinary System. Toxicological Pathology 40 (4) suppl., 54S-55S.

Palazzi X, Burkhardt JE, Caplain H, Dellarco V, Fant P, Foster JR, Francke S, Germann P, Gröters S, Harada T, Harleman J, Inui K, Kaufmann W, Lenz B, Nagai H, Pohlmeyer-Esch G, Schulte A, Skydsgaard M, Tomlinson L, Wood CE, Yoshida M (2016). Characterizing "Adversity" of Pathology Findings in Nonclinical Toxicity Studies: Results from the 4th ESTP International Expert Workshop. Toxicological Pathology 44(6), 810-24.
Key result
Dose descriptor:
NOAEL
Effect level:
300 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
female
Basis for effect level:
clinical biochemistry
histopathology: non-neoplastic
Key result
Critical effects observed:
yes
Lowest effective dose / conc.:
1 000 mg/kg bw/day (actual dose received)
System:
endocrine system
Organ:
thyroid gland
Treatment related:
yes
Dose response relationship:
not specified
Relevant for humans:
yes
Conclusions:
Based on decreased T4 levels (for females in combination with increased incidence of follicular cell hypertrophy in the thyroid) at 1000 mg/kg, of which adversity could not be excluded, a NOAEL of 300 mg/kg bw/day is derived for parental toxicity.
Executive summary:

Wistar Han rats were treated with Ethylene glycol dibenzoate by daily oral gavage at dose levels of 100, 300 and 1000 mg/kg (10 rats/sex/dose level). Concurrent controls (10 rats/sex) received the vehicle, polyethylene glycol 400, alone. Males were treated for 8 weeks prior to mating, during mating, and up to termination (for 92 days). Females that delivered offspring were treated for 8 weeks prior to mating, during mating, duringpost-coitum, and 14-15 days of lactation (for 93-98 days). Females without offspring were treated for 85 days (two non-pregnant females) or 96 days (one female without evidence of mating).

Formulation analysis showed that the formulations were prepared accurately and homogeneously.

Parental results

In females treated at 1000 mg/kgmicroscopic examination revealed an increased incidence of follicular cell hypertrophy (minimal or slight) in the thyroid. This was accompanied by a decrease of thyroid hormone T4 (on average 19%). In males treated at 1000 mg/kg serum levels of thyroid hormone T4 were also decreased (on average by 57%), unaccompanied by treatment related changes in thyroid weight or morphology. For both males and females no corroborative findings were observed in TSH levels. Based on the minimal or slight increase in follicular cell hypertrophy in combination with a decrease in T4 levels in 1000 mg/kg treated femalesand the magnitude of decreased T4 levels in 1000 mg/kg treated males, adversity could not be excluded.

Exposure to the test item up to 1000 mg/kg was not associated with obvious toxicity in the remaining parental parameters examined in this study (i.e. mortality, clinical appearance, functional tests, ophthalmoscopy, body weight, food consumption, clinical pathology, macroscopic examination and organ weights). Other changes noted in treated rats, mostly at 1000 mg/kg, were considered non-adverse as discussed below.

Slight salivation observed after dosing among treated animals (all dose levels) was considered a physiological response rather than a sign of systemic toxicity.

Male rats administered the test item showed reduced body weight gain from start treatment onwards, which was not accompanied by reduced food consumption. This occurred to a similar degree at 300 and 1000 mg/kg, and to a lesser degree at 100 mg/kg. The resulting reduction in mean body weights was modest (about 10% from Day 36 onwards at 300 and 1000 mg/kg) and the growth rate of treated males had returned to control levels after five weeks of treatment. Therefore, the decreased body weights of treated male rats were regarded as non-adverse within the context of this study.

A further treatment-related morphological change observed in females treated at 1000 mg/kg consisted of anincreased incidence of minimal hyperplasia/hypertrophy of the urothelium of the urinary bladder. There was no inflammation or cell death involved in this process. Therefore, this minimal change was considered as non-adverse.

Clinical pathology showed several changes in males treated at 1000 mg/kg: higher values for aspartate aminotransferase (ASAT), alkaline phosphatase (ALP), total bilirubin and albumin; and lower values for cholesterol. Additionally, the increased prothrombin time (PT) for males at 1000 mg/kg could be related to the decrease in the number of platelets. These changes were not accompanied byadverse anatomic pathology alterations and the mean values for these parameters in 1000 mg/kg males generally remainedwithin the range considered normal for rats of this strain and age (see results section). As such, these clinical pathology changes were regarded as non-adverse.

Based on decreased T4 levels (for females in combination with increased incidence of follicular cell hypertrophy in the thyroid) at 1000 mg/kg, of which adversity could not be excluded, a NOAEL of 300 mg/kg bw/day is derived for parental toxicity.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEL
300 mg/kg bw/day
Study duration:
subchronic
Quality of whole database:
A GLP and OECD guideline K1 study is available.
System:
endocrine system
Organ:
thyroid gland

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

Justification for classification or non-classification

Based on the available information the substance is not classified as for repeated dose toxicity.