Registration Dossier

Administrative data

Key value for chemical safety assessment

Effects on fertility

Link to relevant study records
Reference
Endpoint:
toxicity to reproduction
Remarks:
other: subacute study
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: GLP study
Reason / purpose:
reference to same study
Qualifier:
according to
Guideline:
other: OECD TG 407
GLP compliance:
yes (incl. certificate)
Species:
rat
Strain:
Wistar
Sex:
male/female
Route of administration:
oral: gavage
Vehicle:
water
Details on exposure:
The application volume was 10 mL/kg bw.
Details on mating procedure:
not applicable because it is a subacute study
Analytical verification of doses or concentrations:
no
Details on analytical verification of doses or concentrations:
The test item AAS-Loesung is a 48.8% solution in water. Dilution in water is therefore expected not to influence the stability of the test item. The stability of the formulations was declared by the sponsor as at least 8 days.
Duration of treatment / exposure:
29 days
Frequency of treatment:
once daily
Remarks:
Doses / Concentrations:
0, 100, 300, 1000 mg/kg bw and day
Basis:
other: solid content of AAS-Loesung (48.8%)
Remarks:
Doses / Concentrations:
0, 205, 615, 2050 mg/kg bw and day
Basis:
other: AAS-Loesung
No. of animals per sex per dose:
5
Control animals:
yes, concurrent vehicle
Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day (actual dose received)
Based on:
other: solid content of AAS-Loesung (48.8%)
Sex:
male/female
Basis for effect level:
other: no adverse effects on reproductive organs or tissues
Remarks on result:
other: Generation: not applicable because it is a subacute study (migrated information)
Reproductive effects observed:
not specified
Conclusions:
No adverse effects on reproductive organs or tissues were observed in a subacute oral study with rats up to and including 1000 mg/kg and bw.
Executive summary:

In a repeated dose toxicity study in rats (Wistar, OECD TG 407) the substance was administered via gavage to 5 rats/sex/dose at 0, 100, 300, 1000 mg/kg bw in demineralized water for 4 weeks. Up to and including 1000 mg/kg bw no mortality, treament related clinical signs or other toxicological relevant effects occured. Neither gross pathological nor histopathological changes of organs, including the male and female reproductive organs (testes, epididymides, prostate, seminal vesicles incl. coagulation glands, ovaries with oviducts, uterus with cervix, vagina, mammary glands) were reported at any exposure concentration, including the limit dose of 1000 mg/kg bw. Overall, based on the subacute toxicity study there is no indication of a reproductive toxicity potential of AAS and the NOAEL for reproductive effects for male and female rats can be considered as 1000 mg AAS/kg bw.

Effect on fertility: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
1 000 mg/kg bw/day
Study duration:
subacute
Species:
rat
Quality of whole database:
No two-generation reproductive toxicity study is required because a 28-day study indicates no adverse effects on reproductive organs or tissue up to and including the limit dose of 1000 mg/kg bw and day. Data waiver is claimed.
Effect on fertility: via inhalation route
Endpoint conclusion:
no study available
Effect on fertility: via dermal route
Endpoint conclusion:
no study available
Additional information

The dossier contains a subacute repeated oral dose toxicity study in rats performed according to OECD TG 407 with AAS doses of up to and including the limit dose of 1000 mg/kg bw and day. No effects on the reproductive organs or tissues were observed at any dose. The NOAEL for adverse effects on reproductive organs or tissues can therefore be determined with 1000 mg/kg bw.

Although studies on fertility, respectively multigeneration studies are not available for AAS further testing is considered to be of low priority. In accordance to REACH Annex XI, section 1.2. there is sufficient weight of evidence to conclude that AAS is not a reproductive toxicant and therefore further testing on vertebrate animals for that property shall be omitted (for details see waiver for fertility/multigeneration study). In line with REACH Annex XI, section 1.1.2 the available information on the reproductive toxicity potential of AAS is adequate for the purpose of classification and labeling and/or risk assessment.


Short description of key information:
28 day oral toxicity study: rat (Wistar) male/female, gavage (OECD 407): NOAEL for effects on reproductive organs 1000 mg/kg bw and day (male/female)

Effects on developmental toxicity

Description of key information
Developmental toxicity study (OECD 414) in rats: the maternal and developmental NOAEL was established with 1000 mg/kg bw/day
Link to relevant study records
Reference
Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study
Qualifier:
according to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Version / remarks:
(2001)
Qualifier:
according to
Guideline:
EU Method B.31 (Prenatal Developmental Toxicity Study)
Qualifier:
according to
Guideline:
EPA OPPTS 870.3700 (Prenatal Developmental Toxicity Study)
GLP compliance:
yes (incl. certificate)
Limit test:
no
Species:
rat
Strain:
Wistar
Details on test animals and environmental conditions:
TEST ANIMALS
- Strain: Crl:WI(Han) (outbred, SPF-Quality)
- Source: Charles River Deutschland, Sulzfeld, Germany
- Age at delivery: 10-14 weeks
- Housing: individually in Makrolon cages (MIII type) on sterilized sawdust as bedding material.
- Diet and water: ad libitum
- Acclimation period: at least 5 days prior to treatment


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18 - 24°C
- Humidity (%): approximately 40-70 %
- Air changes (per hr): > 10 per hour
- Photoperiod (hrs dark / hrs light): 12/12
Route of administration:
oral: gavage
Vehicle:
water
Remarks:
(Elix, Milipore)
Details on exposure:
Administration volume: 10 mL/kg bw; actual dose volumes were calculated according to the latest body weight;
Appearance of formulations: clear solutions
PREPARATION OF DOSING SOLUTIONS: Formulations (w/w) were prepared daily within 5 hours prior to dosing and were homogenized to a visually acceptable level. Adjustment was made for density of the test substance. Correction was made for the purity/composition of the test substance.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Accuracy of dose formulations, homogeneity and stability of the test substance in formulations were confirmed.
No test substance was detected in the Group 1 formulations (control group). The concentrations analysed in the formulations of Groups 2, 3, and 4 were in agreement with the target concentrations (i.e. mean accuracies between 90% and 110%).
The formulations of Group 2 and Group 4 were homogeneous (i.e. coefficient of variation ≤ 10%).
Formulations at the entire range were stable when stored at room temperature under normal laboratory light conditions for at least 5 hours (i.e. relative difference ≤ 10%). The long term storage samples were stable at ≤-70°C for at least 23 days.
Details on mating procedure:
Untreated females were mated at the Supplier, and were at Day 0 or 1 post-coitum on arrival at the Test Facility (Day 0 post-coitum is the day of
successful mating; confirmed by vaginal plug).
Duration of treatment / exposure:
Days 6 - 20 p. c., inclusive
Frequency of treatment:
Once daily for 7 days per week, approximately the same time each day with a maximum of 6 hours difference between the earliest and latest
dose.
Duration of test:
from day 0 or day 1 post-coitum to necropsy at day 21 p.c.
Remarks:
Doses / Concentrations:
100, 300, 1000 mg/kg bw
Basis:
actual ingested
No. of animals per sex per dose:
22 females
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: In order to set the dose levels for the main teratology study, a non-GLP dose range finding study was performed. Four groups of 6 females were exposed to 200, 500, or 1000 mg/kg bw/day for Days 6 to 20 post-coitum inclusive by oral gavage. These dose levels were based on a 28-day oral toxicity study (OECD 407) with AAS-Solution in which male and female rats received the test substance by oral gavage at dose levels of 0, 100, 300 or 1000 mg/kg bw/day. In this study no toxicologically significant effects were noted up to 1000 mg/kg bw/day. Thus, the NOAEL for repeated dose toxicity was determined with 1000 mg/kg bw, corresponding to 2050 mg active ingredient/kg bw.
Maternal examinations:
CLINICAL EXAMINATIONS: Yes
- Time schedule: At least once daily from Day 2 post-coitum onwards up to the day prior to necropsy.

BODY WEIGHT: Yes
- Time schedule for examinations: Days 2, 6, 9, 12, 15, 18 and 21 post-coitum.

FOOD CONSUMPTION: Yes
- Days 2-6, 6-9, 9-12, 12-15, 15-18 and 18-21 post-coitum.

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

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 21 with subsequently exsanguinated and subjected to an external, thoracic and abdominal examination, with special attention being paid to the reproductive organs.
Ovaries and uterine content:
Each ovary and uterine horn of animals surviving to planned necropsy was dissected and examined as quickly as possible to determine:
- The number of corpora lutea.
- The weight of the (gravid) uterus.
- The number and distribution of live and dead fetuses.
- The number and distribution of embryo-fetal deaths (early and late resorptions).
- The weight of each fetus.
- The sex of each fetus from the ano-genital distance (during necropsy) and also from gonadal
inspections (during further fetal examination).
- Externally visible macroscopic fetal abnormalities.
Fetal examinations:
External, visceral, and skeletal findings were recorded as developmental variations (alterations in anatomic structure that are considered to have no significant biological effect on animal health or body conformity and/or represent slight deviations from normal) or malformations (those structural
anomalies that alter general body conformity, disrupt or interfere with normal body function, or may be incompatible with life).
External:
Each viable fetus was examined in detail, weighed and sexed. All live fetuses were euthanized by administration of approximately 0.05 mL (=10mg) of sodium pentobarbital into the oral cavity using a small flexible plastic or metal feeding tube.
For late resorptions a gross external examination performed and thereafter resorptions were discarded.
Visceral (Internal):
Approximately one-half of the fetuses (live and dead) in each litter (all groups)were examined for visceral anomalies by dissection in the fresh (non-fixed) state. The thoracic and abdominal cavities were opened and dissected. This examination included the heart and major vessels. Fetal kidneys were examined and graded for renal papillae development. The sex of all fetuses was confirmed by internal examination.
The heads were removed from these fetuses and placed in Bouin's solution. Tissues were then transferred to a 70% aqueous ethanol solution for subsequent processing and soft-tissue examination of all groups using the Wilson sectioning technique. After examination, the tissues were stored in 10% buffered formalin. Any remaining tissues (from the fetuses used for fresh visceral examination) was discarded. The carcasses were processed and stained with Alizarin Red S (as described below), but not examined in first instance.
Skeletal:
From the other one-half of the fetuses (live and dead) in each litter (all groups), the sex was confirmed by internal examination. All fetuses were eviscerated, fixed in 96% aqueous ethanol, macerated in potassium hydroxide and stained with Alizarin Red S. Skeletal examination was done for one-half of the fetuses (i.e. the fetuses with heads). The specimens were archived in glycerin with bronopol as preservative. A few bones were not available for skeletal examination because they were accidentally damaged or lost during processing. The missing bones were listed in the raw data; evaluation by the fetal pathologist and study director determined there was no influence on the outcome of the individual or overall skeletal examinations, or on the integrity of the study as a whole.
Statistics:
The following statistical methods were used to analyze the data:
- If the variables could be assumed to follow a normal distribution, the Dunnett-test (many-to one t-test) based on a pooled variance estimate was applied for the comparison of the treated groups and the control group.
- The Steel-test (many-to-one rank test) was applied if the data could not be assumed to follow a normal distribution.
- The Fisher Exact-test was applied to frequency data.
- The Mann Whitney test was used to compare mean litter proportions (percent of litter) of the number of viable and dead fetuses, early and late resorptions, total resorptions, pre- and postimplantation loss, and sex distribution.
- Mean litter proportions (percent per litter) of total fetal malformations and developmental variations (external, visceral and skeletal), and each particular external, visceral and skeletal malformation or variation were subjected to the Kruskal-Wallis nonparametric ANOVA test to determine intergroup differences. If the ANOVA revealed statistically significant (p<0.05) intergroup variance, Dunn’s test was used to compare the compound-treated groups to the control group.
All tests were two-sided and in all cases p < 0.05 was accepted as the lowest level of significance.
Group means were calculated for continuous data and medians were calculated for discrete data (scores) in the summary tables. Test statistics were calculated on the basis of exact values for means and pooled variances. Individual values, means and standard deviations might be rounded off before printing. Therefore, two groups might display the same printed means for a given parameter, yet display different test statistics values.
No statistics were applied for data on maternal survival, pregnancy status, group mean numbers of dead fetuses, early and late resorptions, and pre- and post-implantation loss.
Indices:
For each litter the following calculations were performed:
Percent Pre-implantation loss, percent Post-implantation loss
The fetal developmental findings were summarized by: 1) presenting the incidence of a given finding both as the number of fetuses and the number of litters available for examination in the group; and 2) considering the litter as the basic unit for comparison, calculating the number of affected fetuses as a
mean litter proportion on a total group basis, i.e. percent Viable fetuses affected/litter
The following definitions were applicable for implantation data:
- Fetal (late) resorptions were defined as a dead fetus with external degenerative changes and presence of distinguishable features such as head or limbs.
- Embryonic (early) resorptions were defined as evidence of implantation without presence of distinguishable features such as head or limbs.
- Dead fetus was defined as a non-viable fetus without external degenerative changes and presence of distinguishable features such as head or limbs.
- Post-implantation loss included embryonic (early) resorptions and fetal (late) resorptions.
Details on maternal toxic effects:
Maternal toxic effects:no effects

Details on maternal toxic effects:
No treatment related clinical signs were observed the animals at any of the dose levels 100, 300 or 1000 mg/kg bw/day or vehicle controls.
Body weights and food consumption was unaffected by treatment.
All females, except two females treated at 300 mg/kg bw/day, at scheduled necropsy were pregnant and had litters with viable fetuses.
Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day (actual dose received)
Based on:
act. ingr.
Basis for effect level:
other: developmental toxicity
Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day (actual dose received)
Based on:
act. ingr.
Basis for effect level:
other: maternal toxicity
Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
There were no treatment related effects on litter size, sex ratio, fetal body weights, fetal morphological examination, fetal external, visceral and sceletal morphology.
Skeletal malformations occurred in two fetuses from Group 3 and three control fetuses and not in fetuses from the low and high dose groups. The malformations noted in Group 3 fetuses were vertebral anomaly with or without associated rib anomaly (fetus A063-03) and vertebral centra anomaly (fetus A059-10). Because these malformations occurred singly in Group 3, they were not considered to be treatment related. Besides, there were two control fetuses with a vertebral anomaly with or without associated rib anomaly, namely fetus A002-02 and A015-01. The first one (no. A002 - 02, which had exencephaly and open eye externally) had bent scapulae as well. The third skeletally malformed control fetus (no. A007-09) had a skull anomaly.
Skeletal variations were observed for 76.4%, 74.3%, 76.3% and 76.8% of fetuses per litter in Groups 1, 2, 3 and 4, respectively. The variations noted were not considered treatment related as they occurred in the absence of a dose-related incidence and/or occurred infrequently.
Abnormalities:
not specified
Developmental effects observed:
not specified
Executive summary:

In a prenatal developmental toxicity study performed according to OECD TG 414 mated female Wistar rats were assigned to four dose groups, each containing twenty-two animals. The test item was administered once daily by gavage from Day 6 to 20 post-coitum at doses of 100, 300 and 1000 mg/kg bw/day. The rats of the control group received the vehicle, water, alone.

No maternal toxicity and no treatment-related effects on the fetal development were observed in all treatment groups. In detail, litter size, sex-ratio, fetal body weight, and fetal morphological examinations (external, visceral and skeletal) were not affected by the treatment. In conclusion, based on the results of this prenatal developmental toxicity study the No Observed Adverse Effect Level (NOAEL) for maternal and developmental effects of the test item was established as being 1000 mg/kg bw/day.

Effect on developmental toxicity: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
1 000 mg/kg bw/day
Study duration:
subacute
Species:
rat
Quality of whole database:
GLP and Guideline compliant developmental toxicity study
Effect on developmental toxicity: via inhalation route
Endpoint conclusion:
no study available
Effect on developmental toxicity: via dermal route
Endpoint conclusion:
no study available
Additional information

In a prenatal developmental toxicity study performed according to OECD TG 414 mated female Wistar rats were assigned to four dose groups, each containing twenty-two animals. The test item was administered once daily by gavage from Day 6 to 20 post-coitum at doses of 100, 300 and 1000 mg/kg bw/day. The rats of the control group received the vehicle, water, alone.

No maternal toxicity and no treatment-related effects on the fetal development were observed in all treatment groups. In detail, litter size, sex-ratio, fetal body weight, and fetal morphological examinations (external, visceral and skeletal) were not affected by the treatment. In conclusion, based on the results of this prenatal developmental toxicity study the No Observed Adverse Effect Level (NOAEL) for maternal and developmental effects of the test item was established as being 1000 mg/kg bw/day.

Justification for classification or non-classification

There is conclusive data for the assessment of the test substance with regard to fertility and developmental toxicity. The substance is not classified for both endpoints in accordance to Regulation (EC) No 1272/2008.