Registration Dossier

Administrative data

Description of key information

The NOAEL derived from the key study is:
NOAEL (oral, rat) >1000 mg/kg bw/day, according to OECD 408, Shell, 2013,

The key subchronic oral toxicity study was performed in rats by gavage at 10, 100 and 1000 mg/kg bw/day. There were no toxicologically significant effects up to 1000 mg/kg bw/day, therefore the ‘No Observed Adverse Effect Level’ (NOAEL) was 1000 mg/kg bw/day. However, the study is reliable with restrictions as TEGME (2-(2-(2-methoxyethoxy)ethoxy)ethanol, CAS 112-35-6) has been used as vehicle. TEGME itself has a subchronic repeated dose oral NOAEL of 400 mg/kg bw in rats.

The NOAEL in a 28 day rat study was 170 mg B-TEGME/kg bw/day or greater, based on a toxicity study with a brake fluid given orally up to 1000 mg/kg bw/day. No adverse findings were observed up to the highest dose level.

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:
2013
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
TEGME (2-(2-(2-methoxyethoxy)ethoxy)ethanol, CAS 112-35-6) has been used as vehicle and is not a guideline or standard vehicle. TEGME itself has a subchronic oral NOAEL of 400 mg/kg bw in rats.
Qualifier:
according to
Guideline:
OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity in Rodents)
Version / remarks:
21 September 1998
Deviations:
yes
Remarks:
Futher information on deviations are given under Principles of method if other than guideline.
Principles of method if other than guideline:
There were two main deviations from the guideline OECD 408 or study plan:

1) A fire at the site of the test facility (on 20 February 2013) did not affect the animal facility housing but influenced the daily schedule due to restricted access:
- data for male food consumption could not be collected
- to calculate the dosing volume that day animal body weights were used only
- data were recorded manually and retrospectively entered as the data capture system was offline due to the fire
- delay in dosing resulted in skipping the 5 hours of observation; observations were performed earlier and in parallel with the final mobility checks (in consistency with weekend/public holiday procedures)
It is considered that these modifications to the intended procedure outlined in the Study Plan will have no adverse impact on the scientific integrity of the study.

2) A non-standard/non-guideline vehicle with a different toxicological profile is used. However, the results or extent of effect in the study will not be influenced.
GLP compliance:
yes (incl. certificate)
Limit test:
no
Specific details on test material used for the study:
- Physical state: Clear colourless liquid
- Analytical purity: 89%
- Composition of test material, percentage of components: confidential details
- Purity test date: 31 August 2012
- Lot/batch No.: DEG4131277
- Expiration date of the lot/batch: 27 July 2014
- Stability under test conditions: yes
- Storage condition of test material: Room temperature in the dark
Species:
rat
Strain:
Wistar
Remarks:
Wistar Han:RccHan
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan Laboratories U.K. Ltd., Oxon, UK
- Age at study initiation: Approximately six to eight weeks old

- Weight at study initiation: At the start of treatment the males weighed 190 to 224g, the females weighed 149 to 174g.
- Housing: In groups of three or four by sex in solid floor polypropylene cages with stainless steel mesh lids and softwood flake bedding (Datesand Ltd., Cheshire, UK). Environmental enrichment was provided in the form of wooden chew blocks and cardboard fun tunnels (Datesand Ltd., Cheshire, UK).
- Diet: ad libitum; pelleted diet (Rodent 2014C Teklad Global Certified Diet, Harlan Laboratories U.K. Ltd., Oxon, UK.)
- Water: ad libitum
- Acclimation period: 6 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 ± 2 °C
- Humidity (%): 55 ± 15%
- Air changes (per hr): At least fifteen air changes per hour
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 23 January 2013 (first day of treatment) To: 24 April 2013 (final day of necropsy)

Route of administration:
oral: gavage
Vehicle:
other: TEGME (2-(2-(2-methoxyethoxy)ethoxy)ethanol, CAS 112-35-6)
Remarks:
The vehicle is a non-standard vehicle.
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS:
For the purpose of this study the test item was prepared at the appropriate concentrations as a solution in TEGME. The stability and homogeneity of the test item formulations were determined by Harlan Laboratories Ltd., Shardlow, UK, Analytical Services. Formulations were prepared weekly and stored at approximately 4 °C in the dark.
Samples of test item formulations were taken and analysed for concentration of B-TEGME at Harlan Laboratories Ltd., Shardlow, UK, Analytical Services. The results indicate that the prepared formulations were within ± 7% of the nominal concentration.
The volume of test and control item administered to each animal (4mL/kg) was based on the most recent scheduled body weight and was adjusted at weekly intervals.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Samples of test item formulations were taken and analysed for concentration of B-TEGME at Harlan Laboratories Ltd., Shardlow, UK, Analytical Services. The results indicate that the prepared formulations were within ± 7% of the nominal concentration.
Duration of treatment / exposure:
90 days
Frequency of treatment:
Daily
Dose / conc.:
10 mg/kg bw/day (actual dose received)
Dose / conc.:
100 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 (40 males and 40 females in total), for details refer to table 1
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale:
The dose levels were chosen based on the results of previous toxicity work (Harlan Study Number 41204722). The oral route was selected as the most appropriate route of exposure, based on the physical properties of the test item, and the results of the study are believed to be of value in predicting the likely toxicity of the test item to man.
- Rationale for animal assignment: The animals were allocated to treatment groups using a stratified body weight randomisation procedure and the group mean body weights were then determined to ensure similarity between the treatment groups. The cage distribution within the holding rack was also randomised.

Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: All animals were examined for overt signs of toxicity, ill-health or behavioural change immediately before dosing, up to thirty minutes post dosing and one and five hours after dosing during the working week. Animals were observed immediately before and after dosing and one hour after dosing at weekends and public holidays. All observations were recorded.

DETAILED CLINICAL OBSERVATIONS: Yes
Prior to the start of treatment and at weekly intervals thereafter, all animals were observed for signs of functional/behavioural toxicity. During Week 12 functional performances tests were also performed on all animals together with an assessment of sensory reactivity to different stimuli.

BODY WEIGHT: Yes
- Time schedule for examinations: Individual body weights were recorded on Day 1 (prior to dosing) and at weekly intervals thereafter. Body weights were also recorded at terminal kill.

FOOD CONSUMPTION: Yes
- Food consumption calculated as g food/animal/day
Food consumption was recorded for each cage group at weekly intervals throughout the study.

WATER CONSUMPTION: Yes
- Water intake was observed daily, for each cage group, by visual inspection of the water bottles for any overt changes.

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: The eyes of all control and high dose animals were examined pre-treatment and before termination of treatment (during Week 12). Examinations included observation of the anterior structures of the eye, pupillary and corneal blink reflex. Following pupil dilation with 0.5% Tropicamide solution (Mydriacyl® 0.5%, Alcon Laboratories (UK) Ltd., Pentagon Park, Boundary Way, Hemel Hampstead, Hertfordshire), detailed examination of the internal structure of the eye using a direct ophthalmoscope was performed.
- Dose groups that were examined: All control and high dose animals.

HAEMATOLOGY: Yes
- Time schedule for collection of blood: Haematological and blood chemical investigations were performed on all animals from each test and control group at the end of the study (Day 90). Blood samples were obtained from the lateral tail vein. Where necessary repeat samples were obtained by cardiac puncture prior to necropsy on Day 91.
- Anaesthetic used for blood collection: No
- Animals fasted: No. Animals were not fasted prior to sampling.
- How many animals: All animals from each test and control group at the end of the study (Day 90).
- Parameters checked:
Haemoglobin (Hb), Erythrocyte count (RBC), Haematocrit (Hct), Mean corpuscular haemoglobin (MCH), Mean corpuscular volume (MCV), Mean corpuscular haemoglobin concentration (MCHC), Total leucocyte count (WBC), Differential leucocyte count:(neutrophils (Neut) - lymphocytes (Lymph) - monocytes (Mono) - eosinophils (Eos) - basophils (Bas), Platelet count (PLT), Reticulocyte count (Retic) - Methylene blue stained slides were prepared but reticulocytes were not assessed.
Prothrombin time (CT) was assessed by ‘Innovin’ and Activated partial thromboplastin time (APTT) was assessed by ‘Actin FS’ using samples collected into sodium citrate solution (0.11 mol/L).

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Haematological and blood chemical investigations were performed on all animals from each test and control group at the end of the study (Day 90). Blood samples were obtained from the lateral tail vein. Where necessary repeat samples were obtained by cardiac puncture prior to necropsy on Day 91.
- Animals fasted: No. Animals were not fasted prior to sampling.
- How many animals: All animals from each test and control group at the end of the study (Day 90).
- Parameters checked:
Urea, Glucose, Total protein (Tot.Prot.), Albumin, Albumin/Globulin (A/G) ratio (by calculation), Sodium (Na+), Potassium (K+), Chloride (Cl-), Calcium (Ca++), Inorganic phosphorus (P), Aspartate aminotransferase (ASAT), Alanine aminotransferase (ALAT), Alkaline phosphatase (AP), Creatinine (Creat), Total cholesterol (Chol), Total bilirubin (Bili), Bile acids

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: end of study
- Battery of functions tested:

1. Behavioural Assessment
Detailed individual clinical observations were performed for each animal using a purpose built arena. The following parameters were observed:
Gait Hyper/Hypothermia, Tremors Skin colour, Twitches Respiration, Convulsions Palpebral closure, Bizarre/Abnormal/Stereotypic behaviour Urination, Salivation Defecation, Pilo-erection Transfer arousal, Exophthalmia Tail elevation, Lachrymation
This test was developed from the methods used by Irwin (1968) and Moser et al (1988). The scoring system used is outlined in The Key to Scoring System and Explanation for Behavioural Assessments and Sensory Reactivity Tests.

2. Functional Performance Tests
A. Motor Activity.
Twenty purpose built 44 infra-red beam automated activity monitors were used to assess motor activity. Animals of one sex were tested at each occasion and were randomly allocated to the activity monitors. The tests were performed at approximately the same time each occasion (at least two hours after dosing), under similar laboratory conditions. The evaluation period was one hour for each animal. The time in seconds each animal was active and mobile was recorded for the overall one hour period and also during the final 20% of the period (considered to be the asymptotic period, Reiter and Macphail 1979).
B. Forelimb/Hindlimb Grip Strength. An automated grip strength meter was used. Each animal was allowed to grip the proximal metal bar of the meter with its forepaws. The animal was pulled by the base of the tail until its grip was broken. The animal was drawn along the trough of the meter by the tail until its hind paws gripped the distal metal bar. A record of the force required to break the grip for each animal was made. Three consecutive trials were performed for each animal. The assessment was developed from the method employed by Meyer et al (1979).

3. Sensory Reactivity
Each animal was individually assessed for sensory reactivity to auditory, visual and proprioceptive stimuli. This assessment was developed from the methods employed by Irwin (1968) and Moser et al (1988). The following parameters were observed:
Grasp response Touch escape, Vocalisation Pupil reflex, Toe pinch Blink reflex, Tail pinch Startle reflex, Finger approach
Sacrifice and pathology:
GROSS PATHOLOGY: Yes
On completion of the dosing period all animals were killed by intravenous overdose of sodium pentobarbitone followed by exsanguination.
All animals were subjected to a full external and internal examination, and any macroscopic abnormalities were recorded.

ORGAN WEIGHTS:
The following organs, removed from animals that were killed at the end of the study, were dissected free from fat and weighed before fixation:
Adrenals, Brain, Epididymides, Testes, Heart, Kidneys Liver, Ovaries, Spleen, Thymus, Uterus

HISTOPATHOLOGY: Yes
Samples of the following tissues were removed from all animals and preserved in buffered 10% formalin, except where stated:
Adrenals, Aorta (thoracic), Bone & bone marrow (femur including stifle joint)•, Bone & bone marrow (sternum) Prostate, Brain (including cerebrum, cerebellum and pons), Caecum, Colon, Duodenum, Epididymides ♦, Eyes*, Gross lesions, Heart, Ileum (including Peyer’s patches), Jejunum, Kidneys, Liver, Lungs (with bronchi) #, Lymph nodes (mandibular and mesenteric), Mammary glands, Muscle (skeletal)•, Oesophagus, Ovaries, Pancreas, Pituitary, Rectum, Salivary glands (submaxillary), Sciatic nerve, Seminal vesicles, Skin (hind limb), Spinal cord (cervical, mid-thoracic and lumbar), Spleen, Stomach, Testes ♦, Thymus, Thyroid/Parathyroid, Tongue•, Trachea, Urinary bladder, Uterus

• Retained only and not processed
* Eyes fixed in Davidson’s fluid
♦ Preserved in Bouin’s fluid and then transferred to Industrial Methylated Spirits (IMS) approximately 48 hours later
# Lungs were inflated to approximately normal inspiratory volume with buffered 10% formalin before immersion in fixative

All tissues were despatched to the Test Site (TUPI Manufacturing) for processing (Principal Investigator: R Trindall). All tissues from control and 1000 mg/kg bw/day dose group animals were prepared as paraffin blocks, sectioned at a nominal thickness of 5 μm and stained with Haematoxylin and Eosin for subsequent microscopic examination. Any macroscopically observed lesions were also processed.
Statistics:
Data were processed to give summary incidence or group mean and standard deviation values where appropriate. All data were summarized in tabular form.
Where considered appropriate, quantitative data was subjected to statistical analysis to detect the significance of intergroup differences from control; statistical significance was achieved at a level of p<0.05. Statistical analysis was performed on the following parameters:
Grip Strength, Motor Activity, Body Weight Change, Haematology, Blood Chemistry, Absolute Organ Weights, Body Weight-Relative Organ Weights.

Data were analysed using the decision tree from the ProvantisTM Tables and Statistics Module as detailed as follows:
Where appropriate, data transformations were performed using the most suitable method. The homogeneity of variance from mean values was analysed using Bartlett’s test. Intergroup variance were assessed using suitable ANOVA, or if required, ANCOVA with appropriate covarities. Any transformed data were analysed to find the lowest treatment level that showed a significant effect using the Williams Test for parametric data or the Shirley Test for nonparametric data. If no dose response was found but the data shows non-homogeneity of means, the data were analysed by a stepwise Dunnett’s (parametric) or Steel (non-parametric) test to determine significant difference from the control group. Where the data were unsuitable for these analyses, pair-wise tests were performed using the Student t-test (parametric) or the Mann- Whitney U test (non-parametric).

Probability values (p) are presented as follows:
p<0.01 **
p<0.05 *
p>0.05 (not significant)
Clinical signs:
no effects observed
Description (incidence and severity):
There were no substance related clinical signs of toxicity detected in treated animals.
Mortality:
no mortality observed
Description (incidence):
There were no unscheduled deaths during the study. There were no substance related clinical signs of toxicity detected in treated animals.
Body weight and weight changes:
no effects observed
Description (incidence and severity):
No adverse effects were detected in body weight gain in treated animals when compared to controls.
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
No adverse effect in overall food consumption or food efficiency was detected in treated animals when compared to controls.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
no effects observed
Description (incidence and severity):
There were no treatment related effects detected in water consumption.
Ophthalmological findings:
no effects observed
Description (incidence and severity):
There were no treatment related ocular effects detected.
Haematological findings:
no effects observed
Description (incidence and severity):
There were no toxicologically significant effects detected in the haematological parameters examined.
Clinical biochemistry findings:
no effects observed
Description (incidence and severity):
There were no toxicologically significant effects detected in the blood chemical parameters examined.
Urinalysis findings:
not examined
Behaviour (functional findings):
no effects observed
Description (incidence and severity):
There were no treatment-related changes in behavioural parameters measured. There were no toxicologically significant changes in functional performance. There were no treatment related changes in sensory reactivity.
Organ weight findings including organ / body weight ratios:
no effects observed
Description (incidence and severity):
No toxicologically significant effects were detected in the organ weights examined.
Gross pathological findings:
no effects observed
Description (incidence and severity):
There were no toxicologically significant macroscopic abnormalities detected.
Histopathological findings: non-neoplastic:
no effects observed
Description (incidence and severity):
No treatment related microscopic abnormalities were detected.
Details on results:
CLINICAL SIGNS AND MORTALITY
Animals of either sex from all treatment groups showed episodes of increased salivation during the treatment period. The number of animals, the number of incidences and the onset of this observation followed a true dose related response. Isolated incidences of noisy respiration were
also noted in animals of either sex treated with 1000 and 100 mg/kg bw/day and in control animals. One male treated with 1000 mg/kg bw/day showed hunched posture on Day 32 only. Observations of this nature are commonly observed following the oral administration of an unpalatable formulation or the dosing procedure, and in the absence of any associated changes are considered not to be of toxicological importance. Scab formation was evident in one male treated with 100 mg/kg bw/day between Days 87 and 91. Observations of this nature are commonly observed in group housed animals and are considered not to be related to treatment. Staining of the ano-genital region was evident in one control female between Days 63 and 91. In the absence of treatment, this was considered to be a low incidence finding occasionally observed in laboratory maintained rats.

BODY WEIGHT AND WEIGHT GAIN
There were no toxicologically significant effects detected in body weight development.
Males treated with 1000 mg/kg bw/day showed a statistically significant increase in body weight gain during Week 6. Females from this treatment group also showed a statistically significant increase in body weight gain during Week 7 of treatment. An increase in body weight gain is not considered to represent an adverse effect of treatment.

FOOD CONSUMPTION
No adverse effect in overall food consumption or food efficiency was detected in treated animals when compared to controls.

WATER CONSUMPTION
There were no treatment related effects detected in water consumption.

OPHTHALMOSCOPIC EXAMINATION
There were no treatment related ocular effects detected.

HAEMATOLOGY
There were no toxicologically significant effects detected in the haematological parameters examined.
Animals of either sex treated with 1000 mg/kg bw/day showed a statistically significant increase in prothrombin time. Males from this treatment group also showed a statistically significant reduction in haematocrit. The majority of individual values were within normal ranges for rats of the strain and age used and in the absence of any associated correlates the intergroup differences were considered not to be of toxicological importance.
Females treated with 1000 mg/kg bw/day showed a statistically significant reduction in mean corpuscular haemoglobin and mean corpuscular haemoglobin concentration. Although the majority of individual values were outside of the background control ranges for these parameters, individual control values for mean corpuscular haemoglobin concentration were also outside of the background ranges. Therefore in the absence of any associated histology correlates the intergroup differences were considered not to be of toxicological importance.

CLINICAL CHEMISTRY
There were no toxicologically significant effects detected in the blood chemical parameters examined.
Males treated with 1000 mg/kg bw/day showed a statistically significant increase in chloride concentration and a statistically significant reduction in cholesterol. Females treated with 1000 mg/kg bw/day also showed a statistically significant increase in albumin/globulin ratio.
The majority of individual values were within normal ranges for rats of the strain and age used and in the absence of any associated histology correlates the intergroup differences were considered not to be of toxicological importance.

NEUROBEHAVIOUR
Weekly open field arena observations did not reveal any treatment-related effects for treated animals when compared to controls.
All inter and intra group differences in urination, defecation and transfer arousal scores were considered to be a result of normal variation for rats of the strain and age used, and the differences were of no toxicological importance.

There were no toxicologically significant changes in functional performance.
Males from all treatment groups showed a statistically significant reduction in mean hind limb grip strength and overall activity. Males treated with 1000 and 100 mg/kg bw/day also showed a statistically significant reduction in overall mobility whilst males treated with 10 mg/kg bw/day also showed a statistically significant reduction in the final 20% of activity. In the absence of a true dose related response or any supporting clinical observations to suggest an effect of neurotoxicity, the intergroup differences were considered not to be of toxicological significance.
Females treated with 1000 and 100 mg/kg bw/day showed a statistically significant reduction in mean fore limb grip strength. In the absence of a true dose related response or any supporting clinical observations to suggest an effect of neurotoxicity, the intergroup differences were considered not to be of toxicological significance.

There were no treatment-related changes in sensory reactivity.

ORGAN WEIGHTS
There were no toxicologically significant effects detected in the organ weights measured.
Males treated with 1000 mg/kg bw/day showed a statistically significant reduction in thymus weight both absolute and relative to terminal body weight. In the absence of any associated histopathology correlates the intergroup difference was considered not to be of toxicological
importance.

GROSS PATHOLOGY
There were no toxicologically significant macroscopic abnormalities detected.
Two males treated with 1000 mg/kg bw/day had small and flaccid testes. One of these males also had small epididymides. A further male treated with 1000 mg/kg bw/day had a small left testis. One male treated with 10 mg/kg bw/day had small testes and another male from this treatment group had a small right testis. In the absence of any treatment related histology correlates the intergroup differences were considered not to be of toxicological importance. One male treated with 100 mg/kg bw/day had increased renal pelvic space in the right kidney. This kidney was also fluid filled. These effects are considered to be a congenital abnormality and unrelated to treatment. One control female and two females treated with 100 mg/kg bw/day had reddened lungs at necropsy. In the absence of a dose related response or the administration of test item in the case of the control female the intergroup differences were considered to be incidental.

HISTOPATHOLOGY
There were no treatment related microscopic abnormalities detected.
The findings recorded were within the range of normal background lesions which may be recorded in rats of the strain and age used.
Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day (actual dose received)
Based on:
test mat.
Remarks:
B-TEGME (test material) + TEGME (vehicle)
Sex:
male/female
Remarks on result:
other: No adverse effects observed.
Critical effects observed:
no
Conclusions:
The oral administration of B-TEGME to rats by gavage, at dose levels of 10, 100 and 1000 mg/kg bw/day, did not result in any toxicologically significant effects. The ‘No Observed Adverse Effect Level’ (NOAEL) was therefore considered to be 1000 mg/kg bw/day.
Executive summary:

In a sub-chronic, repeated dose toxicity study according to OECD 408, 10 male and 10 female Han Wistar rats were administered B-TEGME (tris{2-[2-(2-methoxyethoxy)ethoxy]ethyl}borate) orally by gavage 10, 100 or 1000 mg/kg bw/day for 90 consecutive days.

A control group of ten males and ten females was dosed with vehicle alone TEGME (2-(2-(2-methoxyethoxy)ethoxy)ethanol).

Clinical signs, functional observations, body weight change, dietary intake and water consumption were monitored during the study. Haematology and blood chemistry were evaluated for all animals at the end of the study. Ophthalmoscopic examination was also performed on control group and high dose animals. All animals were subjected to gross necropsy examination and histopathological evaluation of selected tissues from high dose and control animals was performed.

There were no unscheduled deaths nor any treatment related clinical signs during the study. There were no treatment-related changes in behavioural parameters, functional performance and sensory reactivity. No adverse effects were detected in body weight (gain), food consumption and water consumption. There were no treatment related ocular effects detected. There were no toxicologically significant effects detected in the haematological and clinical chemistry parameters, organ weights, macroscopic and microscopic examination.

The oral administration of B-TEGME to rats by gavage, at dose levels of 10, 100 and 1000 mg/kg bw/day, did not result in any toxicologically significant effects.

The ‘No Observed Adverse Effect Level’ (NOAEL) was therefore considered to be 1000 mg/kg bw/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 with restrictions as TEGME (2-(2-(2-methoxyethoxy)ethoxy)ethanol, CAS 112-35-6) has been used as vehicle. TEGME itself has a subchronicrepeated dose oral NOAEL of 400 mg/kg bw in rats.

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

In a key oral subchronic toxicity study (90 days), the test item was administered by gavage to three groups, each of ten male and ten female Wistar Han:RccHan:WIST strain rats at dose levels of 10, 100 and 1000 mg/kg bw/day (Shell, 41204723, 2013). A control group of ten males and ten females was dosed with vehicle alone (TEGME, CAS 112-35-6). Clinical signs, functional observations, body weight change, dietary intake and water consumption were monitored during the study. Haematology and blood chemistry were evaluated for all animals at the end of the study. Ophthalmoscopic examination was also performed on control group and high dose animals. All animals were subjected to gross necropsy examination and histopathological evaluation of selected tissues from high dose and control animals was performed.

There were no unscheduled deaths, nor any treatment related clinical signs during the study. There were no treatment-related changes in behavioural parameters, functional performance and sensory reactivity. No adverse effects were detected in body weight (gain), food consumption and water consumption. There were no treatment related ocular effects detected. There were no toxicologically significant effects detected in the haematological and clinical chemistry parameters, organ weights, macroscopic and microscopic examination.

The oral administration of B-TEGME to rats by gavage, at dose levels of 10, 100 and 1000 mg/kg bw/day, did not result in any toxicologically significant effects. The 'No Observed Adverse Effect Level' (NOAEL) was therefore considered to be 1000 mg/kg bw/day. However, the study is regarded to be reliable with restrictions as TEGME (CAS 112-35-6) has been used as vehicle at concentrations from 3200 mg/kg bw (high dose group) up to 4200 mg/kg bw (control/vehicle group). TEGME itself has a subchronic repeated dose oral NOAEL of 400 mg/kg bw/d (see attached OECD SIDS for TEGME, CAS 112-35-6).

An oral subacute toxicity study (28 days) was performed with a brake fluid containing 17% B-TEGME, which was administered to rats at doses of 4, 25 and 170 mg B-TEGME/kg bw/day, based upon doses of 25, 150 and 1000 mg brake fluid/kg bw/day (Shell, SGBR.92.180, 1993). This study was conducted according to OECD 407 guideline and in compliance with GLP, therefore it was considered to be adequate, reliable and relevant. As it was performed with a brake fluid containing 17% B-TEGME and other borated and non-borated components, it was not considered as a key study, but a supporting study for the 90-day repeated dose toxicity study. Dosing did not result in treatment related clinical signs, effects on body weight, clinical chemistry, haematology, gross pathology, organ weights and urinalysis. The only finding related to treatment was a very slight hepatocellular centrilobular hypertrophy at histology in males and some females at 170 mg B-TEGME/kg/day or 1000 mg brake fluid/kg bw/day. Liver weight increases at this high dose compared to controls were approximately 7% in males and 3% in females. Considering the mild nature of the findings, and in the absence of any other relevant treatment related effect, this was not considered to constitute an adverse effect. The NOAEL was therefore considered to be at least 170 mg B-TEGME/kg bw/day or 1000 mg brake fluid/kg bw/day. The other borated and non-borated glycol ethers and components present in the brake fluids also might have contributed to the findings.

Liver effects such as liver weight increase and histopathological hepatocellular hypertrophy in experimental animals have been described in literature (Andrew, 2005) as frequently observed and adaptive effects, often related to enzyme induction and metabolic breakdown of the products in the liver. A liver weight increase of <10% is generally considered to be non-adverse, while increases of 10% or more are interpreted as being potentially adverse. Liver enlargement associated with an increase in the activity of xenobiotic metabolising enzymes is an adaptation to increased functional load and is therefore considered to be a physiological rather than a pathological response or adverse effect.

Justification for classification or non-classification

The available experimental test data are reliable and suitable for classification purposes under Regulation 1272/2008. As a result, the substance is not considered to be classified for repeated dose toxicity under Regulation (EC) No. 1272/2008, as amended for the tenth time in Regulation (EU) No 2017/776.