Registration Dossier

Administrative data

Key value for chemical safety assessment

Effects on fertility

Description of key information

A key study for reproductive toxicity in rats by was available from an OECD 422 study with ead across substance EC 939 -637 -2 or 'Butanedioic acid, 2(or 3)-sulfo-, 4-[2-[(1-oxo(C12-C18(even numbered) and C18unsaturated)alkyl))amino]ethyl]esters, disodium salts', at dose levels of 100, 300 and 1000 mg/kg bw/day. No reproductive toxicity effects were observed up to 1000 mg/kg bw. At the dose of 1000 mg/kg bw, decrease body weight and other systemic effects were observed, therefore NOAEL for systemic toxicity was 300 mg/kg bw/day, whereas NOAEL for reproductive toxicity was 1000 mg/kg bw/day.

Link to relevant study records
Reference
Endpoint:
screening for reproductive / developmental toxicity
Remarks:
based on test type (migrated information)
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Justification for type of information:
REPORTING FORMAT FOR THE ANALOGUE APPROACH

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
For details please refer to Read Across Justification Document, Section 13.2

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
For details please refer to Read Across Justification Document, Section 13.2

3. ANALOGUE APPROACH JUSTIFICATION
For details please refer to Read Across Justification Document, Section 13.2

4. DATA MATRIX
For details please refer to Read Across Justification Document, Section 13.2
Reason / purpose for cross-reference:
reference to same study
Reason / purpose for cross-reference:
read-across source
Frequency of treatment:
d
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Slightly increased salivationwas noted in one male rat dosed at 1000 mg/kg bw/day.
No signs of clinical toxicity were noted in all female treatment groups.
Dermal irritation (if dermal study):
not examined
Mortality:
no mortality observed
Description (incidence):
No premature deaths were noted in the male and female rats treated with 100, 300 or 1000 mg/kg bw/day.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Slight reduction of body weight and body weight gain was noted in male and female rats dosed at 1000 mg/kg bw/day.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
oral gavage study
Slight reduction in food consumption was noted in male and female rats dosed at 1000 mg/kg bw/day
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):
Increased number of eosinophils in intermediate dose females on test day 15 lacking dose dependence (p≤0.05)
Clinical biochemistry findings:
effects observed, treatment-related
Description (incidence and severity):
In the high dose group (1000 mg/kg bw/day) a statistically significant (p≤0.01) increase (males: +66%; females: +46%)) was noted for the plasma activity of ALAT.
In the male high dose group (1000 mg/kg bw/day) a decrease in albumin on test day 15 (p≤0.01) was noted but not considered test-item related (slight alteration in comparison to control animals without biological relevance).
In the female intermediate dose group (300 mg/kg bw/day) an increase in bile acids on test day 15 (p≤0.05) was noted but not considered test-item related (lacking dose dependence).
Urinalysis findings:
not examined
Behaviour (functional findings):
effects observed, non-treatment-related
Description (incidence and severity):
No test item-related influence was noted for the fore- and hindlimb grip strength in any male/female treatment group (100, 300 or 1000 mg/kg bw/day).
No test item-related influence was noted on the spontaneous motility of the rats in any of the treatment groups.
In the male low dose group (100 mg/kg bw/day) a decrease in hindlimb grip strength on test day 36 (p≤0.05) was noted but not considered test-item related (lacking dose dependence).
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
effects observed, non-treatment-related
Description (incidence and severity):
No test item-related influence in observational screening was noted for any treatment group (100, 300 or 1000 mg/kg bw/day).
In the high dose males a statistically significant decreased mean body temperature was noticed in comparison to the control group. This was considered to be not test item related.
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
No test item-related microscopic changes were seen in the reproductive organs for both males and females.
A test item-related squamous cell hyperplasia was noted in the forestomachs from 5/5 male and 5/5 female animals of the high dose group (1000 mg/kg bw/day). These test item-related stomach changes were localized in the zone adjoining the glandular stomach mucosa (forestomach or non-glandular mucosa) and attained statistical significance for both sexes (p≤0.01). Occasionally the squamous cell hyperplasia with subsequent hyperkeratinization was associated with acute inflammation of the submucosa in the non-glandular stomach (for 2/5 males and 1/5 females).
Examination of the stomachs from the animals (5 per group) of the low- and intermediate dose groups (100 and 300 mg/kg bw/day) did not reveal any test item-related changes. As humans lack a forestomach, the relevance of these changes for humans is questionable.
Histopathological findings: neoplastic:
not examined
Other effects:
no effects observed
Reproductive function: oestrous cycle:
not specified
Reproductive function: sperm measures:
effects observed, non-treatment-related
Reproductive performance:
effects observed, non-treatment-related
Description (incidence and severity):
No statistically significant differences were noted in the length of the pre-coital time between the control group and the treatment groups.
There were no statistically significant differences for the female fertility rates between the control and the treatment groups.
No test item-related influence was noted on the gestation length of the females in any of the treatment groups compared to the control group.
No statistically significant differences were noted in the number of corpora lutea between the control and the treatment groups of P0 dams.
No statistically significant differences were noted in the number of implantation sites between the control and the treatment groups of P0 dams.
No statistically significant differences were noted in the total number of born pups (alive and dead) or in the number of pups born alive between the control group and the treatment groups of P0 dams.
No test item-related differences were noted between the control group and the treatment groups of P0 dams with respect to the reproduction indices which were calculated from the above mentioned
parameters (Fertility index, Gestation index, Birth index, Live birth index, Pre-implantaion losss and Post-implantation loss).
Parameters with statistically significant differences in comparison to the control group which were considered as to be not test item related:
-Increased Birth Index in low and intermediate dose females
-Increased Pre-implantation loss in low dose females
-Decreased Post-implantation loss in low and intermediate dose females
CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS)
No premature deaths were noted in the male and female rats treated with 100, 300 and 1000 mg/kg bw/day.
No signs of clinical toxicity were noted for the male and female rats of the low and intermediate dose groups (100 and 300 mg/kg bw/day).
A slightly increased salivation was noted in one male rat, no further signs of clinical toxicity were noted for the male and female rats of the high dose group (1000 mg/kg bw/day). No test item related influence was noted for the male and female rats of all treatment groups (100, 300 and 1000 mg/kg bw/day) during the observational and functional (grip strength and spontaneous motility) neurological screenings.

BODY WEIGHT AND FOOD CONSUMPTION (PARENTAL ANIMALS)
A slight reduction in body weight was noted for the male and female rats of the high dose group (1000 mg/kg bw/day). For the male rats the reduction in body weight was noted from test day 8 (4.4%) until test day 36 (5.5%) , statistically significant (p≤0.05) in comparison to the control group on test day8. Body weight gain was accordingly reduced in the high dose group during the whole study, mostly pronounced and statistically significant at the end of the first test week on test day 8 (p≤0.01), with a reduction in body weight gain by 35.4% in comparison to the control group. During the further course of the study the differences in body weight gain between the high dose group and the control group declined, leading to a body weight gain at the end of the study of 42.1%, the control group revealed a body weight gain of 50.5%.
For the high dose female rats a slight, not statistically significant, reduction in body weight was noted of 3.7% and 3.4% was noted on test days 8 and 15 (pre-mating). Body weight gain was 6.6% on test day 8 and 10.6% on test day 15 in the high dose group in comparison to 10.7% (test day 8) and 14.6% (test day 15) in the control group. Body weight was decreased during the whole gestation period between 7.6% on gestation day 0 and 6.5% on gestation day 20, statistically significant (p≤0.05) on gestation days 7 and 14 with 9.7% and 7.9% decrease, respectively. A small reduction in body weight gain was noted in the high dose group on gestation day 0 (15% in comparison to 18%.
On gestation day 20 body weight gain was nearly identical in the high dose group (59.7%) and the control group (59.4%). During lactation a slight but not significant reduction in body weight was noted in the intermediate dose female rats by 4.4 % on lactation day 0 and 5.6% on lactation day 4. In the high dose group the body weight was statistically significantly (p≤0.05) reduced on lactation days 1 and 4 by 9.1% and 9.5%. No noticeable differences were noted in body weight gain during the lactation period with 5.6% in the high dose group and 6.1% in the control group.
A slightly statistically significant (p ≤ 0.01) increase in relative food consumption by 10.3% was noted in the high dose males during the 2nd test week. This was caused by the reduced body weight of the rats of the high dose group.
A slightly statistically significant (p≤0.05) decrease in relative food consumption by 7.4% was noted in the high dose females during the first test week.

HAEMATOLOGICAL FINDINGS
No test item-related influences were noted between the control group and the treatment groups for the haematological parameters, i.e. the haemoglobin content, the number of erythrocytes, leucocytes, reticulocytes and platelets, the haematocrit value, the thromboplastin time (TPT, aPTT), the mean corpuscular volume (MCV), the mean corpuscular haemoglobin (MCH) and the mean corpuscular haemoglobin concentration (MCHC). No test item-related changes were noted in the relative and absolute differential blood counts.
Statistically significant increased number of eosinophils (p≤0.05) was noted in the intermediate dose females on test day 15. This was considered to be not test item related ( lacking dose dependence).

CLINICAL BIOCHEMISTRY
In the high dose male and female group (1000 mg/kg bw/day) a statistically significant (p≤0.01) increase was noted for the plasma activity of ALAT.
In males no test item related influence was noted for the plasma levels of globulin, the albumin/ globulin ratio, bilirubin (total), cholesterol (total), creatinine, glucose, urea in blood, sodium, potassium, calcium, chloride, the activity of the alkaline phosphatise (aP), the activity of ASAT and the serum levels of the bile acids. Decreased albumin in the high dose males (p≤0.01) was considered to be not test item related (slight alteration in comparison to control animals without biological relevance and 4/5 individual values within range of LPT Background Data))
In females no test item related influence was noted for the plasma levels of albumin, of globulin, the albumin/globulin ratio, bilirubin (total), cholesterol (total), creatinine, glucose, protein (total), urera in blood, sodium, potassium, calcium, chloride, the activity of the alkaline phosphatase (aP) and the serum levels of the bile acids. Increased bile acids in the intermediate dose females (p≤0.05) was considered to be not test item related (lacking dose dependence).

BEHAVIOUR (FUNCTIONAL FINDINGS)
The functional neurological screenings were performed on test day 36 for the male rats and between test days 40-50 for the female rats, 2 hours after dosing.
No test item-related influence was noted for the fore- and hindlimb grip strength in any male/female treatment group (100, 300 or 1000 mg/kg bw/day).
Hindlimb grip strength in low dose males was statistically significant decreased on test day 36 (p≤0.05) but lacking dose dependence and was considered to be not test item related.
No test item-related influence was noted on the spontaneous motility of the rats in any of the treatment groups. No test item-related influence on relative and absolute organ weights was noted for the rats treated with 100, 300 or 1000 mg/kg bw/day in comparison to the control group.
Statistically significant differences in the relative organ weights compared to the control, which are not considered to be test item-related were:
- increased relative weight of left gonads in low and high dose males (p≤0.05)
- increased relative weight of right kidney in high dose females (p≤0.05)
- increased relative weight of right adrenal in high dose males (p≤0.05)
Statistically significant differences in the absolute organ weights compared to the control, which are not considered to be test item-related were:
-decreased absolute brain weight in high dose males (p≤0.05)
-decreased absolute heart weight by 16.6% in high dose females (p≤0.05)

GROSS PATHOLOGY (PARENTAL ANIMALS)
Macroscopic inspection at autopsy for the males was performed on test day 37.
No test-item related findings were noted in the low and intermediate dose group (100 and 300 mg/kg bw/day).
However, one animal (no. 3) with a reduction in the size of the testes was noted in the control group.
At the low dose group (100 mg/kg bw/day) animal no. 21 showed a partly reddened thymus and animal no. 29 a thickened right prostate.
In the intermediate dose group (300 mg/kg bw/day) one animal (no. 49) with macroscopic changes in the stomach (yellowish contents, detachment of mucosa) was found.
All changes are considered to be not test item-related but spontaneous due to the low number of occurrence.
In the high dose group (1000 mg/kg bw./day) 2 animals with macroscopic changes in the stomach were noted: Animal no. 68 showed a whitish thickening in the cardia part of the stomach and in the stomach of animal no. 69 a yellowish content was noted.
These findings were considered to be test item-related, because they were associated with microscopic changes in the forestomach from the animals of the high dose group.
Macroscopic inspection at autopsy for the females was performed between test days 43 and 54.
No test-item related macroscopic changes were noted in any treatment group (100, 300 and 1000 mg/kg bw/day) in females.
In 2 of the 3 non-pregnant rats (nos 16 and 54) from the control and the intermediate dose group (300mg/kg bw/day) a thickened uterus was noted.
These findings were considered to be not test item-related but spontaneous due to the low number of occurrence.

NEUROPATHOLOGY (OBSERVATIONAL FINDINGS)
The observational screenings were performed on test day 36 for the male rats and between test days 40-50 for the female rats, 2 hours after dosing.
No test item-related influence was noted for any treatment group (100, 300 or 1000 mg/kg bw/day).
A decrease in mean body temperature (p≤0.05) in the high dose males on test day 36 was considered to be not test item related. The slight alteration in comparison to control animals was without biological relevance (only 0.4°C difference to controls)

HISTOPATHOLOGY (PARENTAL ANIMALS)
Histopathological examination performed on one testicle and one epididymis with special emphasis on
the qualitative stages of spermatogenesis (proliferative, meiotic and spermiogenic phases) and histopathology of the interstitial testicular structure of all adult male animals of the highest dose group and the control group following H & E and PAS staining, did not reveal any test item-related effects.
No test item-related microscopic changes were seen in the reproductive organs for both males and females.
A pulmonary congestion was noted in the lungs from 4/5 male (control 0/5) and 5/5 female rats (control 4/5) of the high dose group (1000 mg/kg bw/day). As pulmonary congestion is occasionally seen in rats as a background finding, this change was not considered to be test item-related.
A test item-related squamous cell hyperplasia was noted in the fore-stomachs from 5/5 male and 5/5 female animals of the high dose group (1000 mg/kg bw/day). These test item-related stomach changes were localized in the zone adjoining the glandular stomach mucosa (forestomach or 13 the squamous cell hyperplasia with subsequent hyperkeratinization was associated with acute inflammation of the submucosa in the non-glandular stomach (for 2/5 males and 1/5 females).
Examination of the stomachs from the animals (5 per group) of the low- and intermediate dose groups (100 and 300 mg/kg bw/day) did not reveal any test item-related changes. As humans lack a fore-stomach, the relevance of these changes for humans is questionable.
All other microscopic changes observed were either coincidental, or lie within the normal background alterations which may be seen in untreated rats of this age and strain.

REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS)
All animals were successfully mated as determinated by positive sperm detection. No statistically significant differences were noted in the length of the pre-coital time between the control group and the treatment groups.
There were no statistically significant differences for the female fertility rates between the control and the treatment groups.
No test item-related influence was noted on the gestation length of the females in any of the treatment groups compared to the control group.
No statistically significant differences were noted in the number of corpora lutea between the control and the treatment groups of P0 dams.
No statistically significant differences were noted in the number of implantation sites between the control and the treatment groups of P0 dams.
No statistically significant differences were noted in the total number of born pups (alive and dead) or in the number of pups born alive between the control group and the treatment groups of P0 dams.
No test item-related differences were noted between the control group and the treatment groups of P0 dams with respect to the reproduction indices which were calculated from the above mentioned parameters (Fertility index, Gestation index, Birth index, Live birth index, Pre-implantation loss and Post-implantation loss).
Parameters with statistically significant differences in comparison to the control group which were considered as to be not test item related:
-Increased Birth Index in low and intermediate dose females (p≤0.05)
-Increased Pre-implantation loss in low dose females (p≤0.05)
-Decreased Post-implantation loss in low and intermediate dose females (p≤0.05).
Key result
Dose descriptor:
NOAEL
Remarks:
reproductive toxicity
Effect level:
>= 1 000 mg/kg bw/day (actual dose received)
Based on:
act. ingr.
Remarks:
read-across test item
Sex:
male/female
Basis for effect level:
other: no effects observed
Remarks on result:
other: highest dose tested
Dose descriptor:
NOAEL
Remarks:
general toxicity
Effect level:
300 mg/kg bw/day (actual dose received)
Based on:
act. ingr.
Remarks:
read-across test item
Sex:
male/female
Basis for effect level:
clinical signs
body weight and weight gain
food consumption and compound intake
clinical biochemistry
gross pathology
histopathology: non-neoplastic
Key result
Critical effects observed:
no
No P1 generation: screening study
Clinical signs:
not examined
Dermal irritation (if dermal study):
not examined
Mortality / viability:
mortality observed, non-treatment-related
Description (incidence and severity):
No test item-related differences were noted between the survival index of the control group and the treatment groups.
No live born pup from the control group and the low and intermediate dose groups died between lactation day 1 and 4, leading to a survival index of 100% for these groups.
In the high dose group (1000 mg/kg bw/day) 9 of 10 dams revealed a survival index of 100%, whereas all 3 live born pups from dam no. 77 were found dead without milk on lactation day 2, leading to a survival index of 0.0% for dam no. 77 and a survival index of 97.6% for the pups of the high dose group. This small decrease in the survival index of the pups of the high dose group was regarded as to be spontaneous.
Body weight and weight changes:
no effects observed
Description (incidence and severity):
No test item-related influence was noted on the mean litter weight of the pups in all treatment groups.
No test item-related influence was noted on the total litter weight of the pups in all treatment groups.
Food consumption and compound intake (if feeding study):
not examined
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Sexual maturation:
not examined
Description (incidence and severity):
Pups were sacrificed on day 4 of lactation.
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
no effects observed
Description (incidence and severity):
The external examinations of the pups at sacrifice on day 4 of lactation revealed no test item-related external visible changes or gross abnormalities after treatment of the parental animals with 100, 300 or 1000 mg/kg bw/day. No milk was noted in the stomachs of the 3 dead pups from dam no. 77. This observation was spontaneous and considered as not test-item related.
Histopathological findings:
not examined
Behaviour (functional findings):
not examined
Developmental immunotoxicity:
not examined
VIABILITY (OFFSPRING)
No test item-related influence was noted on the survival rate of the pups.
No live born pup from the control group and the low and intermediate dose groups died between lactation day 1 and 4, leading to a survival index of 100% for these groups.
In the high dose group (1000 mg/kg bw/day) 9 of 10 dams revealed a survival index of 100%, whereas all 3 live born pups from dam no. 77 were found dead without milk on lactation day 2, leading to a survival index of 0.0% for dam no. 77 and a survival index of 97.6% for the pups of the high dose group. This small decrease in the survival index of the pups of the high dose group was regarded as to be spontaneous.

BODY WEIGHT (OFFSPRING)
No test item-related influence was noted on the mean litter weight of the pups in all treatment groups.
No test item-related influence was noted on the total litter weight of the pups in all treatment groups.

GROSS PATHOLOGY (OFFSPRING)
The external examinations of the pups at sacrifice on day 4 of lactation revealed no test item-related external visible changes or gross abnormalities after treatment of the parental animals with 100, 300 or 1000 mg/kg bw/day. No milk was noted in the stomachs of the 3 dead pups from dam no. 77. This observation was spontaneous and considered as not test-item related.


Key result
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
>= 1 000 mg/kg bw/day (actual dose received)
Based on:
act. ingr.
Remarks:
read-across test item
Sex:
male/female
Basis for effect level:
other: development of F1 offspring
Remarks on result:
other: highest dose tested
Key result
Critical effects observed:
no
No F2 generation: screening study
Key result
Reproductive effects observed:
no

Table 1. Fertility and Reproductive parameters Parental generation

Parameter

Group 1

Control

Group 2

100 mg/kg

Group 3

300 mg/kg

Group 4

1000 mg/kg

No. of females evaluated for pre-coital time

10

10

10

10

Mean precoital interval (days)

4.5

3.4

3.9

2.6

No. of females evaluated for fertility

10

10

10

10

Number of pregnant dams

8

9

9

10

Fertility index (%)

80

90

90

100

No. of females evaluated for gestation length

8

9

9

10

Gestation length (days)

22.0

22.2

22.2

22.1

Number of dams with live pups

8

9

9

10

Gestation Index (%)

100

100

100

100

Corpora lutea(total)

110

137

129

142

Corpora lutea(mean)

13.8

15.2

14.3

14.2

Implantation sites (total)

110

131

128

139

Implantation sites (mean)

13.8

14.6

14.2

13.9

Number of pups at birth (total)

96

125

123

127

Number of pups at birth (mean)

12.0

13.9

13.7

12.7

Birth Index (mean %)

90.4

95.5

95.9

90.1

Birth Index (total# %)

87

951

96 1

90.1

Number of stillbirths

0

0

1

0

No. of dams with stillborn pups

0

0

1

0

Number of live born pups (total)

96

125

123

126

Number of live born pups (mean)

12.0

13.9

13.7

12.6

Live birth index (mean %)

100.0

100.0

100.0

97.5

Live birth index (total#1 %)

100

100

100

99

Pre-implantation loss (mean %)

0.0

4.4

0.7

1.9

Pre-implantation loss (total#2  %)

0.0

4.42

0.8

2.1

Post-implantation loss (mean %)

9.6

4.5

4.1

11.2

Post-implantation loss (total#3 %)

12.7

4.62

3.92

9.4

Number of runts

0

0

0

0

Number of malformed pups

0

0

0

0

# based on the total No. of implantation sites and total No. of pups at birth (alive and dead)

#1 based on the total No. live born pups and total No. of pups at birth (alive and dead)

#2 based on the total No. corpora lutea and total No. of implantation sites

#3 based on the total No. implantation sites and toal number of live born pups

1 p≤0.05 Chi2-test

2 p<0.05 Chi2-test

Conclusions:
NOAEL (no-observed-adverse-effect level) for reproductive toxicity of the read-across test item: >= 1000 mg/kg bw/day, p.o.
Executive summary:

The aim of the study was to obtain information on possible effects of the read-across test item on general toxicity, reproduction and/or development according to OECD guideline 422. The test item was administered orally by gavage to rats at dose levels of 100, 300 and 1000 mg active ingredient/kg bw/day. The application started two weeks before mating on test day one and ended on the day or one day before sacrifice. Day of sacrifice was on test day 37 for the male rats and between lactation day 4 and 7 for the female rats.

Effects on the parental generation (general toxicity)
No test item-related premature death was noted in any treatment group (100, 300 and 1000 mg test item/kg bw/day).

No signs of clinical toxicity were noted for the male and female rats of the low and intermediate dose groups (100 and 300 mg test item/kg bw/day). A slightly increased salivation was noted in one male rat, no further signs of clinical toxicity were noted for the male and female rats of the high dose group (1000 mg test item/kg bw/day). A slight reduction in body weight was noted for the male and female rats of the high dose group (1000 mg test item/kg bw/day). For the male rats the reduction in body weight was noted from test day 8 (4.4%) until test day 36 (5.5%) and for the female rats from gestation day 0 (7.6%) until lactation day 4 (9.5%). The body weight at autopsy was reduced accordingly.

Effects on reproduction parameters and organs

No test item-related influence was noted on the reproduction parameters in any treatment group (100, 300 and1000 mg/kg bw/day).

Microscopic examination revealed no changes in the reproductive organs from the male and female rats of the high dose group (1000 mg/kg bw/day).

Effects on the F0-generation

NOAEL (no-observed-adverse-effect level): 300 mg/kg bw/day, p.o.

Effects on reproductive toxicity

NOAEL (no-observed-adverse-effect level): >=1000 mg/kg bw/day, p.o.

 

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:
Reliable quality (Klimisch 1)
Effect on fertility: via inhalation route
Endpoint conclusion:
no study available
Effect on fertility: via dermal route
Endpoint conclusion:
no study available
Additional information

Read across data were available from EC 939 -637 -2 or 'Butanedioic acid, 2(or 3)-sulfo-, 4-[2-[(1-oxo(C12-C18(even numbered) and C18unsaturated)alkyl))amino]ethyl]esters, disodium salts'.

A key study for repeated dose toxicity was performed by means of an oral combined repeated dose and reproduction/development screening study according to OECD guideline 422 (Hansen, 2013b). The test item was administered orally by gavage to rats at dose levels of 100, 300 and 1000 mg act. ingr./kg bw/day for at least 28 days in male rats and at least 39 days in females. No test item-related premature death was noted in any treatment group. No signs of clinical toxicity were noted for the male and female rats of the low and intermediate dose groups (100 and 300 mg/kg bw/day), whereas slightly increased salivation was noted in one male rat as the only finding at 1000 mg/kg bw/day. A slight reduction in body weight was noted for the male and female rats dosed at 1000 mg/kg bw/day. Other parameters such as neurological observations, haematology and serum chemistry are discussed in the repeated dose toxicity section.

No test item-related influence was noted on the reproduction toxicity parameters in any treatment group (100, 300 and1000 mg/kg bw/day).

Microscopic examination revealed no changes in the reproductive organs from the male and female rats of the high dose group (1000 mg/kg bw/day). NOAEL for systemic toxicity was 300 mg/kg bw/day, whereas NOAEL for reproductive toxicity was >= 1000 mg/kg bw.

 

Conclusion

An oral gavage reproductive screening study with read across substance EC 939 -637 -2 or 'Butanedioic acid, 2(or 3)-sulfo-, 4-[2-[(1-oxo(C12-C18(even numbered)andC18unsaturated)alkyl)) amino]ethyl]esters, disodium salts' showed NOAEL of 300 mg/kg bw for paternal/maternal toxicity, whereas 1000 mg/kg bw was NOAEL for reproductive and developmental toxicity.

Effects on developmental toxicity

Description of key information

Developmental toxicity was not observed in the oral combined repeated dose and reproduction/development screening study according to OECD guideline 422, in which rats were dosed at 100, 300 and 1000 mg act. ingr./kg bw/day with the read-across substance EC 939 -637 -2 or 'Butanedioic acid, 2(or 3)-sulfo-, 4-[2-[(1-oxo(C12-C18(even numbered) and C18unsaturated)alkyl))amino]ethyl]esters, disodium salts'. No test item related influence was noted on the survival rate and the mean and total body weights of the pups. External examination of the pups revealed no visible changes related to the test item. NOAEL for systemic toxicity was 300 mg/kg bw/day, whereas NOAEL for reproductive and developmental toxicity was >= 1000 mg/kg bw.

A key prenatal developmental toxicity study in female rats at dose levels of 100, 300 or 1000 mg/kg b.w./day of the read-across substanceEC 939 -637 -2 or 'Butanedioic acid, 2(or 3)-sulfo-, 4-[2-[(1-oxo(C12-C18(even numbered) and C18unsaturated)alkyl))amino]ethyl]esters, disodium salts'.

Maternal changes at 1000 mg/kg bw/day included a reduction of the body weight, body weight gain, the food consumption (transient), carcass weight, and pathologic changes in the forestomach, of which the latter is not considered relevant for humans. There were no changes in reproductive parameters or fetal findings. The NOAEL was 300 mg act. ingr./kg b.w./day for the dams and above 1000 mg act. ingr./kg b.w./day for the fetuses.

Link to relevant study records
Reference
Endpoint:
developmental toxicity
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Justification for type of information:
REPORTING FORMAT FOR THE ANALOGUE APPROACH

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
For details please refer to Read Across Justification Document, Section 13.2

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
For details please refer to Read Across Justification Document, Section 13.2

3. ANALOGUE APPROACH JUSTIFICATION
For details please refer to Read Across Justification Document, Section 13.2

4. DATA MATRIX
For details please refer to Read Across Justification Document, Section 13.2
Reason / purpose for cross-reference:
read-across source
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
(No toxicologically relevant effects). No adverse influences on the behaviour, external appearance or faeces were noted for the treatment groups (100, 300 or 1000 mg act. ingr./kg bw/day). However, at 1000 mg act. ingr./kg bw/day, salivation was noted for 16 of 20 females. This was not considered to be an adverse effect.
Dermal irritation (if dermal study):
not examined
Mortality:
mortality observed, non-treatment-related
Description (incidence):
No test item-related premature deaths were noted in the dose groups (100, 300 or 1000 mg act. ingr./kg bw/day).
In the intermediate dose group (300 mg act. ingr./kg bw/day), female no. 67 was prematurely sacrificed on its gestation day 12 due to animal welfare reasons. Female no. 67 was noted with clinical observations in form of piloerection and slightly reduced motility on GD 11. Also, a reduced food consumption from GD 7 onwards and a reduced body weight from GD 8 onwards were noted. Necropsy revealed macroscopic changes in form of a haemorrhagic nose and canthus of the left eye, dark red discoloured lungs and a stomach ulcus. The death of the animal was regarded to be due to a misgavage.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
At 1000 mg act. ingr./kg bw/day, a reduced body weight and a reduced body weight gain were noted from GD 16 until study termination on GD 21 (at maximum 6.1% below the value of the control group for the body weight and 22% below the value of the control group from GD 6 to GD 21 for the body weight gain).
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
At 1000 mg act. Ingr./kg bw/day, a test item-related transient decrease in food consumption was noted between GD 6 and GD 10.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Description (incidence and severity):
No test item-related differences were noted.
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
no effects observed
Description (incidence and severity):
Thyroid hormone concentration: No toxicologically relevant changes for serum T3, T4 and TSH levels were noted.
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Description (incidence and severity):
Uterus and carcass weights: No test item-related differences were noted for the gravid uterus weight. In the high dose group (1000 mg act. ingr./kg bw/day), a reduction was noted for the carcass weight (7.0% below the value of the control group, p ≤ 0.01).
Thyroid weights: No test item-related differences were noted.
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
Macroscopic inspection of the dams at necropsy revealed no adverse effects at 100 and 300 mg act. ingr./kg bw/day, but test item-related changes in the cardiac region of the stomach in form of a (partly) whitish thickening or white deposits for 18 of 20 high dose animals at 1000 mg act. ingr./kg bw/day. However, due to the absence of a forestomach in humans this finding is of no noteworthy relevance for humans.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
no effects observed
Description (incidence and severity):
Histopathologic examination of the thyroids revealed no test item-related changes.
Histopathological findings: neoplastic:
no effects observed
Details on results:
- Behaviour, external appearance, faeces:
No changes in behaviour, the external appearance or the faeces were noted in the control group and the low dose group (100 mg act. ingr./kg bw/day) and no toxicologically relevant changes were noted for the intermediate and high dose groups (300 or 1000 mg act. ingr./kg bw/day).
In the intermediate dose group female no. 67 was prematurely sacrificed on its gestation day 12 due to animal welfare reasons; two further animals (nos. 59 and 60) were noted with clinical observations. However, as only 3 of 22 animals including prematurely sacrificed animal no. 67 were affected and the clinical signs were noted only for a few days , the clinical signs were considered to be not toxicologically relevant. In addition, animal no. 59 was identified with 19 dead fetuses at laparotomy. However, it is unknown whether the fetuses died first and were the cause for the observations noted for animal no. 59 or the death of all fetuses was caused by the poor health of the dam. Nevertheless, this animal can be considered as an outlier.
In the high dose group (1000 mg act. ingr./kg b.w./day), clinical observations were noted in form of piloerection, salivation and increased water consumption. As piloerection and increased water consumption were noted only for a few group 4 females, these two clinical observations were not considered to be an adverse effect. Salivation was noted for 16 of 20 animals. However, as animals were affected not more than two days each and salivation lasted only shortly, the observations of salivation are regarded to be test item-related but not considered to be an adverse effect.
- Mortality:
No test item-related premature deaths were noted in the dose groups (100, 300 or 1000 mg act. ingr./kg bw/day). In the intermediate dose group (300 mg act. ingr./kg bw/day), female no. 67 was prematurely sacrificed on its gestation day 12 due to animal welfare reasons. Female no. 67 was noted with clinical observations in form of piloerection and slightly reduced motility on GD 11. Also, a reduced food consumption from GD 7 onwards and a reduced body weight from GD 8 onwards were noted. Necropsy revealed macroscopic changes in form of a haemorrhagic nose and canthus of the left eye, dark red discoloured lungs and a stomach ulcus. The death of the animal was regarded to be due to a misgavage.
- Body weight and body weight gain:
No test item-related differences in body weight were noted between the dams of the control group and low and intermediate dose groups (100 or 300 mg act. ingr./kg bw/day).
In the intermediate dose group (300 mg act. ingr./kg bw/day), a reduced but not statistically significant body weight was noted from GD 18 until GD 21 (at maximum 5.9% below the value of the control group). However, as this was mainly due to the two females nos. 59 (animal with 19 dead fetuses) and female no. 60 that were in a poor health conditions; the reduced body weight was considered to be not test item-related. In the high dose group (1000 mg act. ingr. /kg bw/day), a decreased body weight was noted from GD 16 until study termination on GD 21 (at maximum 6.1% below the value of the control group on GD 21, statistically significant for GD 20 and GD 21 at p ≤ 0.05 or 0.01) for nearly all animals. This constantly decreased body weight was considered to be test item-related.
No test item-related difference between the control group and the animals treated with 100 or 300 act. ingr./kg bw/day was noted for the body weight gain from GD 0 to 21. At 300 mg act. ingr./kg bw/day, a slight reduction was noted for the body weight gain (statistically significant for the body weight gain of the treatment period from GD 6 to GD 21, p ≤ 0.01). However, this was mainly due to only two females (nos. 59 and 60). Animal no. 59 delivered 19 dead fetuses with distinctly reduced body weights and also the fetuses of animal no. 60 had a lower body weight due to the poor health of this dam. The decreased body weights of the fetuses led to decreased maternal body weights and therefore, the reduced body weight gain was considered to be not test item-related. At 1000 mg act. ingr./kg bw/day, a statistically significant reduction was noted for the body weight gain for the whole study period (GD 0 to GD 21) and for the treatment period (GD 6 to GD 21) at p ≤ 0.01 for nearly all animals. As this was in accordance with the test item-related reduction in the body weight, also the reduced body weight gain was considered to be test item-related.
There were no test item-related influences on the gravid uterus weight. Therefore, the gravid uterus weight had the same influence on the body weight gain for all groups
- Food and drinking water consumption:
No test item-related difference was noted between the control group and the treatment groups (100, 300 or 1000 mg act. ingr./kg bw/day). In the high dose group, a statistically significantly (at p ≤ 0.05 or 0.01) decreased food consumption was noted between GD 6 to GD 10 (at maximum 25.8% below the value of the control group on GD 7 to GD 8). It reflects the decreased body weight and body weight gain observed in this group. As the food consumption recovered thereafter, this transient reduction in food consumption was considered to be test item-related. Furthermore, a statistically significant increase in food consumption was noted for the high dose group (GD 2 to GD 3) and a statistically significant decrease was noted for the low dose group (GD 12 to GD 13). However, as these differences were either before start of treatment (high dose group) or lasted for only one day (low dose group) these differences were considered to be spontaneous.
Although female no. 86 of the high dose group was noted with increased water consumption on gestation day 20 and 21, no test item-related changes in drinking water consumption were noted between the dams of the control group and the dams of the treatment groups by visual appraisal.
- Thyroid hormone concentration:
No test item-related differences were noted for the serum concentration of T4 in all dose groups (100, 300 or 1000 mg act. ingr./kg bw/day) and for T3 and TSH in the low dose group.
In the intermediate and high dose group (300 or 1000 mg act. ingr./kg bw/day), a dose-related increase was noted for the serum concentration of T3 (64.4% and 92.2% above the value of the control group, both statistically significant at p < 0.01) and TSH (8.8% and 19.8% above the value of the control group, not statistically significant). However, no difference was noted for the thyroid weights and the histopathological examination of the thyroids revealed no pathologic changes. Therefore, as it is known that histopathological examination of the thyroids is usually more sensitive than hormone levels (Beekhuijzen et al., 2016) the increases in serum levels of T3 and TSH were considered to be of no toxicological relevance.
- Necropsy findings:
No test item-related observations were noted for the dams of the low and intermediate dose groups (100 or 300 mg act. ingr./kg bw/day) during the macroscopic inspection of the organs and tissues.
In the intermediate dose group, macroscopic changes were noted for three females: No. 59 that delivered only dead fetuses revealed changes in form of a haemorrhagic nose and snout, rough fur , an anus that was soiled with faeces and inflated intestines filled with yellow liquid. Female no. 60 was noted with rough fur11 and few haemorrhagic foci in the stomach and the prematurely sacrificed female no. 67 was noted with a nose and the canthus of the left eye that was haemorrhagic, dark-red discoloured lungs and a stomach ulcus.
All findings were regarded to be spontaneous (animals no. 59 and 60) or related to the misgavage (animal no. 67). It is unknown whether the dead fetuses of animal no. 59 were the cause or the result of the bad health condition of the dam. Although the occurrence of dead fetuses is a very rare incident, this incident with all fetuses being dead is considered to be not test item-related as it was observed in only 1 animal of the intermediate dose group and therefore regarded to be an outlier.
In the high dose group (1000 mg act. ingr./kg bw/day), necropsy revealed for single animals changes in the stomach in form of ulcers (no. 80) and detachment of the mucosa (no. 93). Furthermore, a (partly) whitish thickening of the cardiac region of the stomach was noted for 18 of 20 examined animals.
Due to the high incidence of stomach observations, the changes related to the cardiac region of the stomach observed in the high dose animals were considered to be test item-related. However, due to the absence of a forestomach in humans this finding is of no noteworthy relevance for humans.
- Histopathology:
No test item-related morphological lesions were noted during histopathologic examination of the thyroids of the dose groups (100, 300 or 1000 mg act. ingr./kg bw/day).
In all groups, histopathologic examination revealed microfollicular structures with cells in the follicular lumen of the thyroids and squamous cell cysts of the thyroid follicle. However, these findings were considered to be coincidental and not test item-related.
- Thyroid weights
No test item-related differences to the control group were noted for the thyroid weights of the dose groups (100, 300 or 1000 mg act. ingr./kg bw/day).
- Gravid uterus weight, carcass weight and body weight gain from day 6:
No test item-related differences were noted between the gravid uterus weight of the control dams and the dams of the treatment groups (100, 300 or 1000 mg act. ingr./kg bw/day).
For the carcass weight, a statistically significant decrease was noted for the high dose animals (7.0% below the value of the control group, p ≤ 0.01). As this was in accordance with the lower body weight, the reduced carcass weight was considered to be test item-related.
No test item-related differences were noted for the net body weight gain from GD 6 to 21 between the dams of the control group and the dams of the low and intermediate dose groups (100 or 300 mg act. ingr./kg bw/day).
In the intermediate dose group, a statistically significant decrease (p ≤ 0.01) was noted for the net body weight gain. However, as also the low net body weight gain was mainly due to the animals nos. 59 and 60, the reduction was considered to be not test item-related.
At 1000 mg act. ingr./kg bw/day, the distinctly decreased net body weight gain (9.2 g compared to 37.0 g in the control group, p ≤ 0.01) was considered to be test item-related as it was in accordance with the reduced carcass weight and the reduced weight of the whole body (gravid uterus weight + carcass weight).
Number of abortions:
no effects observed
Pre- and post-implantation loss:
effects observed, non-treatment-related
Description (incidence and severity):
In the intermediate dose group, a statistically significantly increased value was noted for the post-implantation loss (p ≤ 0.01). As this was mainly due to one animal that delivered only dead fetuses (animal no. 59), the increased post-implantation loss was considered to be spontaneous and not test item-related.
Total litter losses by resorption:
no effects observed
Early or late resorptions:
effects observed, non-treatment-related
Description (incidence and severity):
In the intermediate dose group, a statistically significantly increased value was noted for the post-implantation loss (p ≤ 0.01). As this was mainly due to one animal that delivered only dead fetuses (animal no. 59), the increased post-implantation loss was considered to be spontaneous and not test item-related.
Dead fetuses:
effects observed, non-treatment-related
Description (incidence and severity):
In the intermediate dose group, one animal that delivered only dead fetuses (animal no. 59), which was considered to be spontaneous and not test item-related.
Changes in pregnancy duration:
not examined
Changes in number of pregnant:
no effects observed
Other effects:
no effects observed
Details on maternal toxic effects:
No test item-related influence on the reproductive parameters (number of implantation sites, fetuses, resorptions and the index of pre- and post-implantation loss) was noted between the dams of the control group and the dams of the treatment groups (100, 300 or 1000 mg act. ingr./kg bw/day).
In the intermediate dose group, a statistically significantly increased value was noted for the post-implantation loss (p ≤ 0.01). As this was mainly due to one animal that delivered only dead fetuses (animal no. 59), the increased post-implantation loss was considered to be spontaneous and not test item-related.
Key result
Dose descriptor:
NOAEL
Effect level:
300 mg/kg bw/day (actual dose received)
Based on:
act. ingr.
Remarks:
read-across test item
Basis for effect level:
body weight and weight gain
food consumption and compound intake
organ weights and organ / body weight ratios
Key result
Abnormalities:
effects observed, treatment-related
Localisation:
other: reduced body weight
Fetal body weight changes:
no effects observed
Description (incidence and severity):
Body weight of the fetuses and the placentae: No test item-related differences were noted between the control group and the dose groups.
Reduction in number of live offspring:
no effects observed
Description (incidence and severity):
No dead fetus was noted in the control group and in the low and high dose group (100 or 1000 mg act. ingr./kg bw/day).
In the intermediate dose group (300 mg act. ingr./kg bw/day), dam no. 59 delivered only dead fetuses (11 male and 8 female fetuses). External inspection revealed one female fetus (no. 59-08) with a malformation in form of an anencephaly. However, as no other influences on the reproduction were noted and no dead fetuses were observed in the high dose group, the occurrence of one litter with only dead fetuses was considered to be not test item-related.
Changes in sex ratio:
no effects observed
Description (incidence and severity):
No test item-related differences between the ratio of male and female fetuses were noted between the control group and the dose groups (100, 300 or 1000 mg act. ingr./kg bw/day).
Changes in litter size and weights:
no effects observed
Changes in postnatal survival:
not examined
External malformations:
no effects observed
Description (incidence and severity):
No test item-related malformations were noted during the macroscopic examinations at laparotomy (external inspection and inspection of the organs and tissues for gross lesions).
Skeletal malformations:
no effects observed
Description (incidence and severity):
No test item-related malformations were noted during the skeletal examination according to DAWSON.
Visceral malformations:
no effects observed
Description (incidence and severity):
No test item-related malformations were noted during the soft tissue examination according to WILSON.
Other effects:
no effects observed
Description (incidence and severity):
Mortality: No test item-related death of fetuses was noted.
Variations: The macroscopic examinations at laparotomy, the skeletal examination according to DAWSON and the soft tissue examination according to WILSON revealed no test item-related variations
Retardations: No test item-related retardations (delays in ossification) were noted during the skeletal examination according to DAWSON.
Details on embryotoxic / teratogenic effects:
- Mortality:
No dead fetus was noted in the control group and in the low and high dose group (100 or 1000 mg act. ingr./kg bw/day).
In the intermediate dose group (300 mg act. ingr./kg bw/day), dam no. 59 delivered only dead fetuses (11 male and 8 female fetuses). External inspection revealed one female fetus (no. 59-08) with a malformation in form of an anencephaly. However, as no other influences on the reproduction were noted and no dead fetuses were observed in the high dose group, the occurrence of one litter with only dead fetuses was considered to be not test item-related.
- Sex distribution of the fetuses:
No test item-related differences between the ratio of male and female fetuses were noted between the control group and the dose groups (100, 300 or 1000 mg act. ingr./kg bw/day).
- Weight of placentae and fetuses:
The placental and fetal weights showed no test item-related differences between the control group and the treatment groups (100, 300 or 1000 mg act. ingr./kg bw/day).
- Number of runts:
No runts were noted for the control group and the intermediate and high dose groups (300 or 1000 mg act. ingr./kg bw/day).
In the low dose group (100 mg act. ingr./kg bw/day), two runts (nos. 26-15 and 36-11) were noted. The occurrence of two runts in the low dose group is within the normal range of biological variability and therefore, was considered to be not test item-related.
- Ano-genital distance:
No test item-related differences to the control group were noted for the fetal ano-genital distance of the dose groups (100, 300 or 1000 mg act. ingr./kg bw/day).
- Macroscopic inspection of the fetuses at laparotomy:
1) External inspection at laparotomy
No macroscopically visible external malformations were noted for the fetuses of the low and high dose group (100 or 1000 mg act. ingr./kg bw/day) during the external inspection at laparotomy.
In the intermediate dose group (300 mg act. ingr./kg bw/day), one fetus (no. 59 08, dead at birth) was noted with a malformation in form of an anencephaly and one fetus (no. 54-19) was noted with multiple malformations in form of a bifurcated tail, supernumerary extremities at the back (presumably a conjoined twin), a cleft palate and an anencephaly. The occurrence of two fetuses with malformations was considered to be spontaneous and not test item-related.
2) Gross inspection of the organs and tissues at laparotomy
The macroscopic inspection of the organs and tissues for gross alterations at laparotomy revealed no malformations or variations for the fetuses of the control group and the fetuses of the dose groups (100, 300 or 1000 mg act. ingr./kg bw/day).
3) Testicular development
No cryptorchidism and no testicular malposition was noted during assessment of the testicular development of the male fetuses of the control group and the dose groups (100, 300 or 1000 mg act. ingr./kg bw/day).
- Skeletal examination according to DAWSON:
1) Skeletal malformations
No skeletal malformations were noted for the fetuses of the control group and the test item-treated groups (100, 300 or 1000 mg act. ingr./kg bw/day) during the skeletal examination according to DAWSON.
2) Skeletal variations
Skeletal variations were noted for the ribs (ribs isolated, ribs short or ribs wavy) and the sternum (bipartite or misaligned to a slight degree).
No test item-related difference in the incidence of the observed skeletal variations in comparison to the control group was noted for the fetuses of the treatment groups (100, 300 or 1000 mg act. ingr./kg bw/day).
3) Skeletal retardations
Retardations (delayed ossifications) were related to the skull (incomplete ossification of frontal, parietal, interparietal and/or supraoccipital areas), the hyoid (unossified), the sternum (sternebra(e) incompletely ossified, reduced in size or unossified), the thoracic vertebral bodies (bipartite, reduced in size or dumbbell-shaped), the caudal vertebral bodies (only one body ossified or all bodies unossified), the lumbar vertebral bodies (unossified), the sacral vertebral bodies (unossified), the lumbar vertebral arches (incompletely ossified), the os ischii (unossified or incompletely ossified), the os pubis (unossidied or incompletely ossified) and the metacarpalia/metatarsalia (absence of ossification in metacarpalia/metatarsalia 2 to 5).
No test item-related increase in the incidence of skeletal retardations at 100, 300 or 1000 mg act. ingr./kg bw/day was noted during skeletal examination according to DAWSON.
In the intermediate dose group, statistically significantly increased incidences were noted for skeletal retardations that were outside the range of the LPT background data (all caudal vertebral bodies unossified, os ischii unossified, os pubis unossified and sacral vertebral bodies unossified). However, the majority of the mentioned retardations were noted for dam no. 59 that littered only dead fetuses. Therefore, the retardations were considered to be not test item-related.
- Soft tissue examination according to WILSON:
1) Malformations
No malformations were noted for the fetuses of the control group and the fetuses of the low and high dose groups (100 or 1000 mg act. ingr./kg bw/day) during the soft tissue examination according to WILSON.
In the intermediate dose group (300 mg act. ingr./kg bw/day), one fetus (no. 59 8) was noted an anencephaly and one fetus (no. 54 19) was a conjoined twin with two spinal cords, extremities in the back region, a bifurcated tail, anencephaly, an open eye, a cleft palate and a missing anus.
However, as no malformations were noted in the high dose group, the occurrence of two fetuses with malformations in the intermediate dose group was considered to be spontaneous and not test item-related.
2) Variations
During the examination of the organs and tissues according to WILSON, variations were noted for the brain (dilatation of the 4th cerebral ventricle), the kidneys (uni- or bilateral dilatation of the renal pelvis or malpositioned) and the liver (haemorrhagic focus/foci).
No test item-related differences and no statistically significant differences in the incidences of the observed variations were noted between the control group and the treatment groups (100, 300 or 1000 mg act. ingr./kg bw/day).
3) Unclassified observations
No unclassified observations were noted for the control group and for the low and high dose group (100 or 1000 mg act. ingr./kg bw/day).
An unclassified observation in form of a thoracic cavity filled with blood was noted for two fetuses (nos. 60-1 and 60-2) of the intermediate dose group (300 mg act. ingr./kg bw/day). This observation was considered to be a preparation-induced artefact and not test item-related.
Key result
Dose descriptor:
NOAEL
Effect level:
> 1 000 mg/kg bw/day (actual dose received)
Based on:
act. ingr.
Remarks:
read-across test item
Sex:
male/female
Basis for effect level:
other: no developmental effect observed
Remarks on result:
other: highest tested dose
Key result
Abnormalities:
no effects observed
Key result
Developmental effects observed:
no
Conclusions:
Under the present test conditions, the no-observed-adverse-effect level (NOAEL) of the read-across test item was 300 mg act. ingr./kg bw/day for the dams.
The no-observed-adverse-effect level (NOAEL) for the fetal organism was above 1000 mg act. ingr./kg bw/day.
Under the conditions of the study, read-across test item Butanedioic acid, 2(or 3)-sulfo-,4-[2-[(1-oxo(C12-C8 (even numbered) and C18 (unsaturated)alkyl) amino]ethyl]esters, disodium salts did not show any teratogenic potential in rats
Executive summary:

The aim of the study was to obtain information of the influence of the read-across test item on the pregnant rat and the fetus when administered orally during the critical period of organogenesis and the fetal development (6th to 20th day of gestation) according to OECD Guideline 414. The read-across test item was administered orally by gavage to rats at dose levels of 100, 300 and 1000 mg active ingredient/kg bw/day starting from the 6th and lasting until the 20th day of pregnancy. Their development was observed during the gestation period (day of mating is day 0 of pregnancy). On gestation day 21 (one day before the calculated date of parturition), the dams were laparotomised and examined for corpora lutea, implantation sites and resorptions in the uterus and for the condition of the fetuses.

 

Examination of the dams:

No test item-related premature deaths were noted in any of the test groups.

No adverse influences on the behaviour, external appearance or faeces were noted for the treatment groups (100, 300 or 1000 mg act. ingr./kg bw/day). However, at 1000 mg act. ingr./kg bw/day, salivation was noted for 16 of 20 females.

At 1000 mg act. ingr./kg bw/day, a reduced body weight and a reduced body weight gain were noted from GD 16 until study termination on GD 21 (at maximum 6.1% below the value of the control group for the body weight and 22% below the value of the control group from GD 6 to GD 21 for the body weight gain).

At 1000 mg act. ingr./kg bw/day, a test item-related transient decrease in food consumption was noted between GD 6 and GD 10.

No test item-related differences were noted in drinking water consumption.

Macroscopic inspection of the dams at necropsy revealed no adverse effects at 100 and 300 mg act. ingr./kg bw/day, but test item-related changes in the cardiac region of the stomach in form of a (partly) whitish thickening or white deposits for 18 of 20 high dose animals at 1000 mg act. ingr./kg bw/day. However, due to the absence of a forestomach in humans this finding is of no noteworthy relevance for humans.

No test item-related differences were noted in thyroid weights.

No toxicologically relevant changes for serum T3, T4 and TSH levels were noted.

Histopathologic examination of the thyroids revealed no test item-related changes.

No test item-related differences were noted for the gravid uterus weight.

In the high dose group (1000 mg act. ingr./kg bw/day), a reduction was noted for the carcass weight (7.0% below the value of the control group, p ≤ 0.01).

No test item-related influence was noted on the reproductive parameters (number of implantation sites, resorptions and fetuses).

 

Examination of the fetus:

No test item-related death of fetuses was noted.

No test item-related differences in body weight of the fetuses and the placentae were noted between the control group and the dose groups.

 

Fetal alterations:

No test item-related malformations were noted during the macroscopic examinations at laparotomy (external inspection and inspection of the organs and tissues for gross lesions), and no malformations were noted during the skeletal examination according to DAWSON and the soft tissue examination according to WILSON.

The macroscopic examinations at laparotomy, the skeletal examination according to DAWSON and the soft tissue examination according to WILSON revealed no test item-related variations.

No test item-related retardations (delays in ossification) were noted during the skeletal examination according to DAWSON.

 

Analysis of test item formulations:

The measured actual concentrations of the read-across test item in the test item vehicle mixtures were between 98.5% and 101.6% of the nominal concentrations, indicating correctly prepared and homogeneous formulations.

 

Conclusion

Under the present test conditions, the no-observed-adverse-effect level (NOAEL) of the read-across test item was 300 mg act. ingr./kg bw/day for the dams.

The no-observed-adverse-effect level (NOAEL) for the fetal organism was above 1000 mg act. ingr./kg bw/day.

Under the conditions of the study, read-across test item Butanedioic acid, 2(or 3)-sulfo-,4-[2-[(1-oxo(C12-C8 (even numbered) and C18 (unsaturated)alkyl) amino]ethyl]esters, disodium salts did not show any teratogenic potential in rats.

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:
Reliable (Klimisch 2)
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

Supporting data for absence of developmental toxicity were available from an oral combined repeated dose and reproduction/development screening study according to OECD guideline 422 (Hansen, 2013b) with the read-across substance EC 939-637-2 or 'Butanedioic acid, 2(or 3)-sulfo-, 4-[2-[(1-oxo(C12-C18(even numbered) and C18unsaturated)alkyl))amino]ethyl]esters, disodium salts'. The test item was administered orally by gavage to rats with a formulation containing 41.5% active ingredient at dose levels of 100, 300 and 1000 mg act. ingr./kg bw/day for for at least 28 days in male rats and at least 39 days in females. No test item-related premature death was noted in any treatment group. No signs of clinical toxicity were noted for the male and female rats of the low and intermediate dose groups (100 and 300 mg/kg bw/day), whereas slightly increased salivation was noted in one male rat as the only finding at 1000 mg/kg bw/day. A slight reduction in body weight was noted for the male and female rats dosed at 1000 mg/kg bw/day. Other parameters such as neurological observations, haematology and serum chemistry are discussed in the repeated dose toxicity section.

No test item-related influence was noted on the developmental toxicity parameters in any treatment group (100, 300 and 1000 mg/kg bw/day). Microscopic examination revealed no changes in the reproductive organs from the male and female rats of the high dose group (1000 mg/kg bw/day). No test item related influence was noted on the survival rate and the mean and total body weights of the pups. External examination of the pups revealed no visible changes related to the test item. NOAEL for paternal/maternal toxicity was 300 mg/kg bw/day, whereas NOAEL for developmental toxicity was >= 1000 mg/kg bw.

A key prenatal developmental toxicity study with the read-across substance EC 939-637-2 or 'Butanedioic acid, 2(or 3)-sulfo-, 4-[2-[(1-oxo(C12-C18(even numbered) and C18unsaturated)alkyl))amino]ethyl]esters, disodium salts' was performed female rats at dose levels of 100,300 or 1000 mg/kg bw/day from the 6th to 20th day of pregnancy (Hansen, 2020).

No test item-related premature death was noted for any dose group. No test item-related changes in behaviour, external appearance, or faeces were noted for the treatment groups except for salivation in 16 of 20 animals dosed with 1000 mg act. ingr./kg bw/day. However, this was not considered to be an adverse effect. No toxicological relevant changes in the serum levels of thyroid hormones were noted for any dose group. Furthermore, no difference was noted for the thyroid weights, and histopathological examination of the thyroids revealed no pathological changes.

A test item-related reduction of the body weight, the body weight gain, the food consumption (transient) and the carcass weight was noted for the females dosed with 1000 mg/kg bw/day. In the high dose group (1000 mg./kg bw/day), pathologic changes were noted for the cardiac region of the stomach (non-glandular stomach, i.e. the forestomach). However, due to the absence of a forestomach in humans this finding is of no noteworthy relevance for humans.

The reproductive parameters (number of implantation sites, number of resorptions and number of fetuses) were not influenced by the test item.

No test item-related deaths of fetuses and no test item-related malformations, variations or retardations were noted.

Under the present test conditions, the NOAEL was 300 mg/kg bw/day for the dams. The NOAEL for the fetal organism was above 1000 mg/kg bw/day.

Mode of Action Analysis / Human Relevance Framework

There is no evidence for species specific effects of the substance. Therefore the results of the studies are regarded as relevant for humans.

Justification for classification or non-classification

Based on these results and according to the CLP (No. 1272/2008 of 16 December 2008), the test item does not have to be classified and has no obligatory labelling requirement for reproductive and development toxicity.

Additional information