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Description of key information

Screening study for reproductive/developmental toxicity (OECD 422, oral, rat, RL1): NOAEL reproductive toxicity 1000 mg/kg bw/day

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records
Reference
Endpoint:
short-term repeated dose toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2018-02-12 to 2018-08-14
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Reason / purpose:
reference to same study
Qualifier:
according to
Guideline:
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
Version / remarks:
29 July 2016
Deviations:
no
GLP compliance:
yes (incl. certificate)
Remarks:
Országos Gyógyszerészeti és Élelmezés-egészégügyi Intézet
Limit test:
no
Species:
rat
Strain:
other: Han:WIST of Wistar origin
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Toxi-Coop Zrt. 1103 Budapest, Cserkesz u. 90
- Females nulliparous and non-pregnant: yes
- Age at study initiation: Males: 85-90 days; Females: 85-90 days
- Weight at study initiation: Males: 315-396 g; Females: 190-230 g
- Housing: Before mating: 2 animals of the same sex/cage; Mating: 1 male and 1 female/cage; Pregnant females: individually; Males after mating: 2 animals/cage; Recovery animals: 2 or 3 animals of the same sex/cage
- Diet: ssniff® SM R/M-Z+H complete diet for rats and mice provided ad libitum
- Water: tap water provided ad libitum
- Acclimation period: 20 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 30 - 70
- Air changes (per hr): 10
- Photoperiod: Artificial light, from 6 a.m. to 6 p.m
Route of administration:
oral: gavage
Vehicle:
water
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS: Butylmonoglycol sulphate, Na-salt was formulated in the vehicle (distilled water) at concentrations of 20, 60 and 200 mg/mL in the formulation laboratory of the Test Facility daily.

VEHICLE
- Amount of vehicle: Control animals were handled in an identical manner to the test groups receiving vehicle (5 mL/kg bw).
- Lot/batch no. (if required): 1801-5512; 1801-5512; 1803-5501
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Analytical control of dosing formulations (control of concentration) was performed twice during the study.
Duration of treatment / exposure:
49 days (male animals) or 50-56 days (female animals); the day of first treatment is considered as Day 0 of examination.
Frequency of treatment:
7 days/week, every day at a similar time (±2 hours)
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:
12 animals per sex per dose
5 animals per sex for control and high dose group as recovery group
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: The dose levels were chosen by the Sponsor on the basis of the results of a preliminary dose range finding study with Butylmonoglycol sulphate, Na-salt in rats (Study no. 904-400-3039). The high and mid-high doses were chosen with the aim of inducing toxic effects but no mortality or severe suffering of animals. The low dose was chosen to induce no toxic effect. The mid dose was interpolated geometrically.
- Rationale for animal assignment: All male and female animals were sorted according to body weight and divided to weight groups aided by a computerized calculation. There were an equal number of animals from each weight group in each of the experimental groups assigned by randomization to ensure that the mean weight of animals from all test groups was as uniformly as practicable. Grouping was aided by SPSS/PC software, verifying the homogeneity and variability between the groups according to the actual body weight.
Positive control:
No
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: General clinical observations were made on parental animals once a day, after the administration at approximately the same time. Detailed examinations were made at the times of wee kly weighing, prior to and during the mating and until necropsy.
- Observations were performed on the skin, fur, eyes and mucous membranes, autonomic activity (lachrymation, piloerection, pupil size, respiratory pattern, occurrence of secretions and excretions), circulatory and central nervous system, somatomotor activity and behavior pattern, changes in gait, posture and response to handling. Special attention was directed towards the observation of tremors, convulsions, salivation, diarrhea, lethargy, sleep and coma.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Detailed examinations were made at the times of weekly weighing, prior to and duri ng the mating and until necropsy.

BODY WEIGHT: Yes
- Time schedule for examinations: Parental males were weighed on the first day of dosing (Day 0) and weekly thereafter and on the day of the necropsy (treatment and recovery periods). Parental females were weighed on the first day of dosing (Day 0) then weekly, on gestation days 0, 7, 14 and 21 and on days 0 (within 24 hours after parturition), 4 and 13 post-partum. Body weight of the female animals was additionally weighed on gestational day 10. Body weight was measured on day of necropsy for female animals subjected to organ weighing. Animals assigned to the recovery groups were weighed by weekly interval during the treatment and post-treatment period.

FOOD CONSUMPTION
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No

WATER CONSUMPTION AND COMPOUND INTAKE: No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: one day after the last treatment
- Anaesthetic used for blood collection: Yes; Isofluran anesthesia
- Animals fasted: Yes
- How many animals: 5 animals/sex/group
- Parameters checked in table [No. 1 and 2] were examined.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: one day after the last treatment
- Animals fasted: Yes
- How many animals: 5 animals/sex/group
- Parameters checked in table [No. 3 and 4] were examined.

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: the last exposure week (Day 49 (corresponding to PND 9-12 for dams) before the blood sampling
- Dose groups that were examined: 5 animals/sex/group
- Battery of functions tested: sensory reactivity to different type of stimuli (e.g. auditory, visual and proprioceptive), assessment of grip strength and motor activity

IMMUNOLOGY: No

OTHER: No
Sacrifice and pathology:
GROSS PATHOLOGY: Yes; all animals were subjected to a full detailed gross necropsy. After examination of the external appearance, the cranial, thoracic and abdominal cavities were opened and the appearance of the tissues and organs was observed, and any abnormality was recorded including details of the location, color, shape and size. Special attention was paid to the organs of the reproductive system.

HISTOPATHOLOGY: Yes; the tissues indicated in Table 5 were prepared for microscopic examination and weighed, respectively.
Statistics:
The statistical evaluation of appropriate data (marked † above) was performed with the statistical program package SPSS PC+4.0.

The homogeneity of variance between groups was checked by Bartlett’s homogeneity of variance test.

Where no significant heterogeneity was detected, a one-way analysis of variance (ANOVA) was carried out. If the obtained result was significant Duncan Multiple Range test was used to access the significance of inter-group differences. Getting significant result at Bartlett’s test, the Kruskal-Wallis an alysis of variance was used and the inter-group comparisons were performed using Mann-Whitney U-test. Chi² test was performed if feasible.

For evaluation of data obtained during the recovery period, the homogeneity of variance between groups was checked by F-test. Depending on the result pooled or separate variance estimate of the Two-Sample t-test was performed.
Clinical signs:
effects observed, non-treatment-related
Description (incidence and severity):
Alopecia on the right forelimb was observed on one female animal at 300 mg/kg bw/day (1/12) on gestation days 11-21 and for the same female alopecia on the forelimbs, thorax and abdomen was observed on lactation days 0-13. Piloerection was recorded for one female animal at 1000 mg/kg bw/day (1/12) on gestation day 23 probably due to the delivery and not related to the treatment. Because clinical signs of toxicity occurred at a low incidence and were not necessarily dose related, these effects were not considered treatment-related.

There were no clinical signs in male or female animals the weekly detailed clinical observations during the course of the recovery period.
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, non-treatment-related
Description (incidence and severity):
Statistical significance was observed at the higher mean body weight gain in male animals at 100 and 300 mg/kg bw/day between Days 41 and 48. In the male animals at 1000 mg/kg bw/day, the mean body weight gain exceeded the control value from Day 20 up to Day 48 except between Days 34 and 41 and for the study overall (between days 0 and 48). In female animals at 300 mg/kg bw/day, statistical significance with respect to the control was ob served at the higher mean body weight gain between Days 7 and 13 and between lactation days 0 and 4.

Slightly higher mean body weight gain was observed in male animals at 1000 mg/kg bw/day comparing to their control during the first week of the recovery period. The mean body weight and body weight gain were similar to that of the control group in female animals at 1000 mg/kg bw/day during the course of post-treatment period.

These slight changes in the mean body weight gain of test item treated male animals during the treatment and recovery periods and in female animals at 300 mg/kg bw/day did not result in changes in the mean body weight therefore were considered to be toxicologically not relevant.
Food consumption and compound intake (if feeding study):
no effects observed
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
effects observed, non-treatment-related
Description (incidence and severity):
In male animals, slight but statistically significant difference with respect to the control was noted for the lower mean corpuscular (erythrocyte) hemoglobin concentration (MCHC) at 1000 mg/kg bw/day.

In female animals, slightly but statistically higher values were observed in some parameters: in the mean concentration of hemoglobin (HGB) and mean hematocrit value (HCT) at 100 and 300 mg/kg bw/day, in the mean corpuscular (erythrocyte) volume (MCV) and mean corpuscular (erythrocyte) hemoglobin (MCH) at 300 mg/kg bw/day.

These differences with respect to control were judged to be toxicologically not relevant due to the minor degree or in the lack of dose relevance and values were within the historical control range (MCV, MCH) or were marginal changed (MCHC, HGB, HCT) to the historical control ranges.

The mean percentages of monocytes (MONO) and basophil granulocytes (BASO) were higher than in the control group in male animals at 1000 mg/kg bw/day at the end of the recovery period.

In the female animals at 1000 mg/kg bw/day, all examined parameters were comparable to the control at the end of the recovery period.

These differences with respect to control in male animals were judged to be indicative of biological variation and toxicologically not relevant. These differences were only observed at the end of the recovery period but not at the end of the treatment period.
Clinical biochemistry findings:
effects observed, non-treatment-related
Description (incidence and severity):
Compared to their control, the mean concentration of glucose (GLUC) was statistically significantly higher in male animals at 100, 300 and 1000 mg/kg bw/day. All individual values were well within the historical ranges and statistical significances were mainly originated from the relatively low value of the control group. Therefore, this was considered to have no toxicological relevance.

In female animals, slightly but statistically significantly higher mean concentration of sodium (Na+) was detected at 300 and 1000 mg/kg bw/day without any toxicological importance.

Statistically significant difference with respect to the control was noted for the slightly higher mean T4 thyroid hormone (free T4) level in PN13 offspring at 1000 mg/kg bw/day. This slight difference was considered to be toxicologically not relevant as all individual values were within or marginal to the historical control range.

The mean activity of alanine aminotransferase (ALT) was higher than in the control group in male animals at 1000 mg/kg bw/day at the end of the recovery period. The slight difference with respect to control in ALT activity was judged to be indicative of biological variation and toxicologically not relevant in the lack of histopathological alterations and similar finding was not detected at the end of the treatment period.
Urinalysis findings:
not examined
Behaviour (functional findings):
no effects observed
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
effects observed, non-treatment-related
Description (incidence and severity):
Statistical significance was noted for the higher mean liver weights (absolute) in male animals at 1000 mg/kg bw/day.

In the female animals at 100 mg/kg bw/day, the mean weights of liver (absolute and relative to brain weight) and adrenal glands (absolute) were slightly lower than in the control group. The mean kidney weight relative to body weight was higher than in the control group.

At 300 mg/kg bw/day, statistical significance was observed at the lower mean weights of adrenal glands (absolute and relative to body and brain weights) in female animals.

At 1000 mg/kg bw/day, statistical significance was noted for the slightly higher mean fasted body weight and at the lower mean weights of adrenal glands (absolute and relative to body and brain weights) for females.

At the end of the recovery period, the mean brain weight relative to body weight was slightly but statistically significantly lower and the mean body weight relative to brain weight was statistically significantly higher compared to the control group in male animals at 1000 mg/kg bw/day.

In the female animals of the 1000 mg/kg bw/day recovery group, the weight of the examined organs was comparable to their controls.

There were no related pathological changes (clinical chemistry, histopathology) therefore these slight changes in the liver of male animals dosed with the high dose were considered to be toxicologically not relevant.

Changes in the organ weights of female animals were of minor degree and not related to doses (liver, kidneys, adrenal glands) thus were considered to be toxicologically not relevant.

Gross pathological findings:
effects observed, non-treatment-related
Description (incidence and severity):
In the control group, yellow-brown formation (0.5 x 1 cm in size) in the abdominal cavity was observed in one male animal (1/12).

In male animals at 1000 mg/kg bw/day, smaller than normal seminal vesicle on the left side (1/12) and pyelectasia (1/12) were observed.

In female animals at 1000 mg/kg bw/day, interrupted left horn of the uterus with liquid content (1/12) was observed.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
effects observed, non-treatment-related
Description (incidence and severity):
In animals selected for full histological examinations, minimal alveolar emphysema in the lungs (1/5 control male, 1/5 male at 1000 mg/kg bw/day) was detected sporadically.

Hyperplasia of bronchus associated lymphoid tissue (BALT) in the lungs (1/5 control male; 1/5 male at 1000 mg/kg bw/day; 1/5 control female) was observed is some animal. Since the effect was observed in the control and high dose group. The observation was not judged to be treatment related.

Tissue formation (necrotic debris surrounded by fibrotic capsule) in the abdominal cavity was observe d in one control male animal (1/6). One female at 1000 mg/kg bw/day had a cyst in the ovary (one side).

The histological picture of epididymides, seminal vesicles, and coagulating glands was normal in all cases except one male animal (1/12 at 1000 mg/kg bw/day). This animal had smaller than normal seminal vesicle (one side) and histological examination revealed decreased amount of secrete in the seminal vesicle, without any other pathological lesion (inflammation, fibrosis etc.).
Histopathological findings: neoplastic:
no effects observed
Other effects:
not examined
Key result
Dose descriptor:
NOAEL
Effect level:
>= 1 000 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: no adverse effects observed up to the highest dose tested
Key result
Critical effects observed:
no
Conclusions:
In an oral gavage study conducted to OECD 422 and according to GLP (reliability score 1) the NOAEL for butylmonoglycol sulphate, Na-salt relating to repeated dose (parental systemic) toxicity was at least 1000 mg/kg bw/day, as no significant adverse effects were observed in rats during the study period.
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
1 000 mg/kg bw/day
Study duration:
subacute
Species:
rat
Quality of whole database:
The available information comprises an adequate and reliable study, and is thus sufficient to fulfil the standard information requirements set out in Annex VIII, 8.6, of Regulation (EC) No 1907/2006.

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

Mode of Action Analysis / Human Relevance Framework

not applicable

Additional information

A key study (combined repeated dose toxicity study with reproductive/developmental toxicity screening test (oral)) with butylmonoglycol sulphate, Na-salt (CAS 67656-24-0) is available and was performed according to OECD TG 422 and in compliance with GLP (Toxi-Coop, 2018). The test substance was administered daily in graduate doses (100, 300, or 1000 mg/kg bw/day) to 3 groups of test animals. A control group was evaluated concurrently. The experimental period involved 20 days of acclimatization (including 14 days for examination of estrous cycle) and 49 days (male animals) or 50-56 days (female animals) treatment/observation period (depending on the effectiveness of mating) and necropsy days. For animals allocated to recovery groups, the experimental period involved 49 days treatment/observation period and additional 14-day post-treatment (recovery) period followed by a necropsy day.

 

No mortality was observed at any dose level. Additionally, no clinical signs of toxicity or behavioral changes were observed at any dose level. Significant changes were observed in body weight gain (increased for 1000-mg/kg bw/day males and 300-mg/kg bw/day females), hematology parameters (decreased MCHC at 1000 mg/kg bw/day for males; increased HGB and HCT at 100 and 300 mg/kg bw/day for females; increased MCV and MCH at 300 mg/kg bw/day for females), clinical chemistry parameters (increased glucose levels in males at all dose levels; increased sodium at 300 and 100 mg/kg bw/day for females), and organ weights (increased absolute liver weight at 1000 mg/kg bw/day for males; decreased absolute and relative liver weight and absolute adrenal gland weight at 100 mg/kg bw/day for females; decreased absolute and relative adrenal gland weight at 300 mg/kg bw/day for females; and decreased absolute and relative to body and brain weight adrenal gland weight at 1000 mg/kg bw/day). None of these changes noted were considered treatment-related because they were minor in nature and/or were not dose related. Macroscopic findings related to the effect of the test item were not found in male and female animals at 100, 300 or 1000 mg/kg bw/day (treatment and the recovery periods). Histopathological examinations of the selected organs (ovaries, uterus, vagina, testes, epididymides, prostate and seminal vesicles with coagulating gland) did not reveal any test item related changes at 1000 mg/kg bw/day. There were no pathologic changes in the examined organs or tissues of randomly selected male or female animals in the control or 1000 mg/kg bw/day groups (treatment and the recovery periods).

 

As no adverse effects were observed up to the highest dose tested, the systemic NOAEL was at least 1000 mg/kg bw/day.

Justification for classification or non-classification

The available data on repeated dose oral toxicity of butylmonoglycol sulphate, Na-salt do not meet the criteria for classification according to Regulation (EC) No 1272/2008, and are therefore conclusive but not sufficient for classification.