Registration Dossier

Administrative data

Key value for chemical safety assessment

Effects on fertility

Description of key information

A reproduction/developmental toxicity screening test was performed with 4,4`-sulphonylphenol according to OECD 421. The NOAEL for parental and reproductive toxicity was considered to be 60 mg/kg bw/day. An Extended One Generation Reproductive Toxicity Study is currently ongoing.

Link to relevant study records

Referenceopen allclose all

Endpoint:
reproductive toxicity, other
Remarks:
Reproduction/Developmental Toxicity Study in Sprague Dawley Rats; Range-Finding Study
Type of information:
experimental study
Adequacy of study:
supporting study
Study period:
05.04.2017-31.07.2017
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
Version / remarks:
There are no guidelines for a range-finding study. This study is being conducted in preparation for a definitive Extended One-Generation Reproductive Toxicity Study (OECD TG 443).
Deviations:
yes
GLP compliance:
yes (incl. certificate)
Limit test:
no
Specific details on test material used for the study:
Name of test substance: 4,4'-dihydroxydiphenylsulfone
Test substance No.: 05/0066-8
Batch No.: 03508136W0
CAS No.: 80-09-1
Purity: 4,4-DHDPS: 99.87 area-% (Analysis)
Homogeneity: Given
Storage stability: Expiry date: 23 Nov 2019
Species:
rat
Strain:
Sprague-Dawley
Details on species / strain selection:
The rat is the preferred animal species for developmental and reproductive toxicity studies according to the various test guidelines.
Based on a regulatory decision on substance evaluation persuant to Article 46(1) of Regulation (EC) No 1907/2006 (CoRAP) SD rats were chosen as rat test strains.
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Research Models and Services, Germany GmbH/ Charles River Laboratories, UK
- Age at study initiation: About 9 weeks (male) / about 10 weeks (female)
- Housing: Type of cage: Polysulfonate cages Typ 2000P (H-Temp) 2-3 animals. During mating, gestation, lactation and females after weaning: Polycarbonate cages type III., 1 animal. Exceptions: during mating: 1 male/1 female per cage, during rearing up to PND 21: 1 dam with her litter
- Diet (e.g. ad libitum): Ground Kliba maintenance diet mouse/rat “GLP”, Provimi Kliba SA, Kaiseraugst, Switzerland; ad libitum
- Water (e.g. ad libitum): Drinking water ad libitum
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24°C
- Humidity (%): 30-70%,
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12
Route of administration:
oral: gavage
Vehicle:
CMC (carboxymethyl cellulose)
Details on exposure:
Dose volume: 10 mL/kg body weight
Preparation: For the test substance preparations, the specific amount of test substance was weighed, topped up with 0.5% Carboxymethylcellulose suspension in drinking water in a calibrated beaker and intensely mixed with a homogenizer.
Before and during administration, the preparations were kept homogeneous with a magnetic stirrer.
Preparation frequency: The test substance preparations was prepared at intervals which guarantee that the test substance concentrations in the vehicle will remain stable.
Details on mating procedure:
F0 generation parental animals and formation of F1 pups:
Male and female rats, aged about 8 weeks (male) and about 9 weeks (female) when supplied, were used as F0 generation parental animals. F0 animals were mated. The female F0 animals were allowed to deliver and rear their pups (F1 generation pups) until postnatal days (PND) 4 or 21.

Mating of the F0 generation parental animals:
After the premating period, the male and female parental animals were mated overnight in a 1:1 ratio until there was evidence of copulation or the maximum period has elapsed. Throughout the mating period, each female was mated with a predetermined male.
A vaginal smear was prepared for each pair after each mating and examined for sperm. If sperm were detected, mating of the pair was discontinued. The day on which sperm was detected is referred to as gestation day (GD) 0 and the following day as GD 1.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The analytical investigations of the test substance preparations were carried out at the Analytical Chemistry Laboratory of Experimental Toxicology and Ecology of BASF SE, 67056 Ludwigshafen, Germany, as a part of this study.
The analytical investigations were performed according to the most recently authorized version of the control procedure 05/0066_01.
The stability of the test substance in 1% Carboxymethylcellulose suspension in drinking water over a period of 4 days at room temperature and over a period of 7 days in a refrigerator has been verified prior to the start of the study in a comparable batch (Project No.: 01Y0066/05Y009).
Duration of treatment / exposure:
All animals, with the exception of the controls, received the test substance daily by gavage. For males, the administration period was 10 weeks, for females it continued through premating, mating, gestation, lactation, post-lactation until one day before necropsy.
Frequency of treatment:
daily
Dose / conc.:
30 mg/kg bw/day
Dose / conc.:
100 mg/kg bw/day
Dose / conc.:
300 mg/kg bw/day
No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle
Parental animals: Observations and examinations:
Clinical examinations of parental animals
Mortality: A check for moribund and dead animals was made twice daily from Mondays to Fridays and once daily on Saturdays, Sundays and public holidays.
Clinical signs: cageside examination were conducted at least once daily for any signs of morbidity, pertinent behavioral changes and/or signs of overt toxicity.
During the administration period all animals were checked daily for any abnormal clinically signs before the administration as well as within 2 hours and within 5 hours after the administration. The parturition and lactation behavior of the dams was evaluated in the morning in combination with the daily clinical inspection of the dams.

Food consumption: determined once a week for the male and female parental animals
Water consumption: determined twice a week for the male and female parental animals
Body weights: determined once a week at the same time of the day (in the morning).

Clinical Pathology: Blood samples were taken from fasted animals by puncturing the retrobulbar venous plexus under Isoflurane anesthesia. Blood sampling and examination was carried out in a randomized sequence.
Hematology: Leukocytes, Erythrocytes, Hemoglobin, Hematocrit, Mean corpuscular volume (MCV), Mean corpuscular hemoglobin (MCH), Mean corpuscular hemoglobin concentration (MCHC), Platelets, Differential blood count, Reticulocytes, Prothrombin time,
Clinical chemistry: Alanine aminotransferase, Aspartate aminotransferase, Alkaline phosphatase, Serum gamma glutamyl transferase, Sodium, Potassium, Chloride, Inorg. phosphate, Calcium, Urea, Creatinine, Glucose, Total bilirubin, Total protein, Albumin, Globulins, Triglycerides, Cholesterol.

Necropsy:
After the end of the administration period all surviving parental males were sacrificed and examined.
All surviving parental females were sacrificed after weaning and examined.
all animals were sacrificed by decapitation under isoflurane anesthesia. The exsanguinated animals were necropsied and assessed by gross pathology, special attention being given to the reproductive organs.
Organ weights: the following weights were determined in all animals sacrificed on schedule: Anesthetized animals, Adrenal glands, Epididymides, Kidneys, Liver, Ovaries, Prostate, Seminal vesicles with coagulating gland, Spleen, Testes, Thymus, Uterus with cervix,
Organ / Tissue fixation: the following organs or tissues were fixed in 4% buffered formaldehyde solution or in modified Davidson’s solution: All gross lesions, Adrenal glands, Cecum, Cervix, Coagulating gland, Colon, Duodenum, Epididymides, Ileum, Jejunum (with Peyer’s patches), Kidneys, Liver, Mammary gland (male + female) incl. inguinal lymph nodes, right, Mammary gland fat pad (male + female) incl. inguinal lymph nodes, left (Carnoy’s solution), Ovaries, Parathyroid glands, Pituitary gland, Prostate, Rectum, Seminal vesicles, spleen, Stomach (forestomach and glandular stomach), Testes (modified Davidson’s solution), Thymus, Thyroid glands, Uterus, Vagina
Histopathology: was performed on all gross lesions, Adrenal glands, Cecum, Kidneys, Liver, Mammary gland (males and females), right, Mammary gland fat pad (males and females), left, Spleen, Uterus, Vagina
Oestrous cyclicity (parental animals):
In all parental females in the premating phase, estrous cycle length and normality were evaluated by preparing vaginal smears during a minimum of 3 weeks prior to mating and throughout cohabitation until there is evidence of sperm in the vaginal smear.
Additionally, on the day of scheduled sacrifice, the estrous status was determined in all parental female rats.
Litter observations:
Pup status and litter size after birth: The status (sex, live-born or stillborn) and number of all pups delivered from the parents was determined as soon as possible after birth. At the same time, the pups were examined for gross-morphological changes.
Pup viability/mortality: In general, a check was made for any dead or moribund pups twice daily on workdays.
Clinical signs: All live pups were examined daily for clinical symptoms (including gross-morphological findings) during the clinical inspection of the dams.
Body weights: pups were weighed
Postmortem examinations of pups: sacrificed pups were examined externally, eviscerated and their organs were assessed macroscopically.
Statistics:
Food consumption, water consumption, body weight and body weight change, gestation days - DUNNETT test (two-sided)
Male and female mating indices, male and female fertility indices, females mated, females delivering, gestation index (females with liveborn pups), females with stillborn pups, females with all stillborn pups - FISHER'S EXACT test (one-sided)
Mating days until day 0 pc, %postimplantation loss, pups stillborn, %perinatal loss - WILCOXON test (one-sided+) with BONFERRONI-HOLM
Implantation sites, pups delivered, pups liveborn, live pups day x, viability Index, survival index - WILCOXON test (one-sided-) with BONFERRONI-HOLM
% live male day x, %live female day x - WILCOXON test (two-sided)
Number of cycles and cycle length, clinical pathology parameters - KRUSKAL-WALLIS test (two-sided) and WILCOXON test (two-sided)
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
clinical signs in high dose
salivation:
in life males 7/10; females 6/10
mating males 7/10; females 3/10
gestation females 5/10
lactation females 4/10

Water consumption:
In F0 males water consumption was decreased at the dose level of 30 mg/kg bw (-16.6%, low dose) and slightly increased from days 6 to 24 at 300 mg/kg bw (+7.6%, high dose).
In F0 females water consumption increased in a dose related manner (9.8%, 11.3% and 18.3% in the 30, 100 and 300 mg/kg bw dose groups, respectively).
In lactating females water consumption was increased in a dose related manner by 1.8%, 5.3%, and 14.3%, in th low, mid and high dose groups, respectively.
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
In F0 males and females body weights were decreased at the high dose level (-7% in males; -6% in females at 300 mg/kg bw, respectively).
Similarly, decreases in body weights were observed in the high dose group (300 mg/kg bw) in mating males (-8.3%), mating females (-4.6%) and females during gestation (-14.6%).
Body weight changes were less prominent in females during lactation.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
In F0 males food consumption was decreased by 7.7 % in the low dose group (30 mg/k bw) and -9.2% in the high dose group (300 mg/kg bw). A slight increase of 4.6% was noted at the end of the pre-mating period in males of the mid dose group (100 mg/kg bw).
In F0 females an increase in food consumtion of 13%, 16% and 16.2% was identified in the low, mid and high dose group, respectively.
Variations in food consumtion were less prominent in females during gestation and lactation.
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Description (incidence and severity):
No biologically relevant observations in clinical chemistry and haematology.
Clinical biochemistry findings:
no effects observed
Description (incidence and severity):
No biologically relevant observations in clinical chemistry and haematology.
Urinalysis findings:
not examined
Behaviour (functional findings):
no effects observed
Immunological findings:
no effects observed
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
In the organs assessed in this study statistically significant increases in relative organ weights were observed in kidneys of males of the mid and high dose group (11.5% and 35%, respectively) and the livers of high dose males (11%).
See below for details.
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
An significant incease in gross lesions in parental animals was observed in the cecum (dilation), kidneys (enlarged, discoloration) and livers (enlarged) of males in the 300 mg/kg bw dose group (high dose).
See below for details.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Males:
In the cecum a dose related increase in incedence of wall thickening and apoptosis was observed in the mid and high dose.
In the kidneys a dose related increase in incedence in degeneration/regeneration, mineralization and tubular dilation was observed in the mid and high dose animals.
In addition a diffuse atrophy of the mammary gland was described in high dose males.

Females:
Histopathological findings were restricted to the kidneys of the high dose group (mineralization, lymphoid infiltration).
See below for details.
Histopathological findings: neoplastic:
no effects observed
Other effects:
not examined
Reproductive function: oestrous cycle:
effects observed, treatment-related
Description (incidence and severity):
High-dose F0 parental females (300 mg/kg bw/d) had a distinctly prolonged estrous cycle.
See below for details.
Reproductive function: sperm measures:
not examined
Reproductive performance:
effects observed, treatment-related
Description (incidence and severity):
See below for details.
Dose descriptor:
NOAEL
Effect level:
100 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male
Basis for effect level:
organ weights and organ / body weight ratios
Dose descriptor:
NOAEL
Effect level:
100 mg/kg bw/day
Based on:
test mat.
Sex:
female
Basis for effect level:
reproductive performance
Clinical signs:
no effects observed
Mortality / viability:
no mortality observed
Body weight and weight changes:
no effects observed
Description (incidence and severity):
A mild statistically significant increase in bw was observed in male pups of the low dose at PND21 (+6.6%).
Gross pathological findings:
no effects observed

Table 1: Absolute body weights and relative organ weights in parental animals-

sex

Males (n=10)

Females (n=10)

group

0

30

100

300

0

30

100

300

Terminal bw (absolute)

548.6

+/-35.7

530.2

+/-43.6

546.3

+/-33.3

497.5

+/-38*

304.5

+/-22.8

301.8

+/-23.8

292.6

+/-22.9

286.6

+/-30.1

adrenals

0.014

+/-0.002

0.014

+/-0.002

0.014 +/-0.002

0.015 +/-0.002

0.027 +/-0.003

0.029 +/-0.003

0.03 +/-0.006

0.03 +/-0.006

kidneys

0.662

+/-0.055

0.692

+/-0.051

0.748 +/-0.049**

1.013 +/-0.114**

0.722 +/-0.039

0.731 +/-0.064

0.762 +/-0.075

0.752 +/-0.068

Liver

2.375

+/-0.148

2.378

+/-0.176

2.519 +/-0.302

2.668 +/-0.186**

2.846 +/-0.154

2.938 +/-0.395

3.297 +/-0.677

2.927+/-0.189

Prostate

0.302 +/-0.038

0.303 +/-0.042

0.278 +/-0.042

0.297 +/-0.035

-

-

-

-

Seminal vesicle

0.357 +/-0.042

0.366 +/-0.04

0.336 +/-0.056

0.348 +/-0.051

-

-

-

-

Spleen

0.147 +/-0.017

0.186 +/-0.103

0.155 +/-0.028

0.164 +/-0.02

0.186 +/-0.023

0.179 +/-0.016

0.186 +/-0.019

0.179 +/-0.025

testes

0.685 +/-0.063

0.663 +/-0.08

0.663 +/-0.047

0.734 +/-0.089

-

-

-

-

thymus

0.054 +/-0.013

0.048 +/-0.008

0.048 +/-0.011

0.044 +/-0.014

0.075 +/-0.015

0.079 +/-0.012

0.072 +/-0.016

0.076 +/-0.017

ovaries

-

-

-

-

0.035 +/-0.001

0.035 +/-0.004

0.037 +/-0.004

0.034 +/-0.002

uterus

-

-

-

-

0.197 +/-0.026

0.224 +/-0.088

0.224 +/-0.099

0.307 +/-0.152

*P<=0.05; **p<=0.01

Table 2: Incidence of gross lesions in parental animals

sex

Males (n=10)

Females (n=10)

group

0

30

100

300

0

30

100

300

 

Cecum dilation

 

 

 

3

 

 

 

 

 

Glandular stomach focus

1

 

2

1

 

1

 

 

 

Kidneys -dilation

1

 

 

1

 

 

 

 

 

     -discoloration

 

 

 

8

 

 

 

 

 

-enlarged

 

1

1

9

 

 

 

 

 

Liver -enlarged

 

1

2

3

 

 

 

 

 

-focus

 

 

1

 

 

 

 

 

 

Spleen -enlarged

 

1

1

 

 

 

 

 

 

-focus

 

 

 

 

 

 

 

 

 

Testes -focus

 

1

 

1

 

 

 

 

 

Table 3: Selected incidence of microscopic lesions in parental animals

sex

Males (n=10)

Females (n=10)

group

0

30

100

300

0

30

100

300

Cecum dilation

 

 

 

3

 

 

 

 

Cecum, thickening of wall

 

 

5

9

 

 

 

 

Cecum, increased apoptosis

 

 

3

9

 

 

 

 

Spleen, haematopoiesis, extramedullar

8

1

2

10

 

 

 

3

 

 

 

 

 

 

 

 

 

Kidneys -degeneration/regeneration

 

 

6

10

 

 

 

 

Kidneys - mineralization      

 

 

2

2

1

 

 

4

Kidneys - tubular dilation

 

 

5

10

 

 

 

 

Kidneys – tubular cast 

 

 

1

1

2

 

 

 

Kidneys – infiltration, lymphoid

 

 

 

 

2

 

 

6

Kidneys – mineralization medulla

 

 

 

 

1

 

 

4

 

 

 

 

 

 

 

 

 

Liver – infiltration lymphoid

10

1

2

10

10

 

 

10

Liver – Necrosis multifocal

1

 

1

1

 

 

 

 

Mammary gland, diffuse atrophy

 

 

 

10

 

 

 

 

Mammary gland, fatpat diffuse atrophy

 

 

 

10

 

 

 

 

 

Table 4: Reproductive performance parameters

mg/kg bw

0

30

100

300

dams

 

 

 

 

pregnant

fertility index [%]

10/10

100

9/10

90

10/10

100

6/10

60

without implantation sites

0/10

0/10

0/10

2/10

Mean implantation sites

15.8

15

15.5

10.4**

% implantation loss

3.6

5.2

6.5

34.6*

mean gestation days

22

22.1

22

22

with stillborn pups

2

1

2

2

with all pups stillborn

0

0

0

0

Estrus cycles lengthy (days)

4.02

3.97

4.01

5.16**

pups

 

 

 

 

Pups delivered

152

127

145

65

Mean pups delivered

15.2

14.1

14.5

10.8**

stillborn

2

1

3

3

liveborn

150

126

142

62

Mean perinatal loss [%]

1.3

0.6

2

5.3

Executive summary:

In a Reproduction/Developmental Toxicity Screening Study which was performed as a Range-Finding Study for an Extended One Generation Reproductive Toxicity study (EOGRTS), 4,4'-sulphonyldiphenol was orally administered via gavage to groups of 10 male and 10 female Sprague-Dawley rats (F0 animals) at doses of 0, 30, 100 and 300 mg/kg bw/d (BASF, 2017). The duration of treatment covered a 6-week premating and a 14-days mating period in both sexes, approximately 4 weeks post-mating in males, the entire gestation and lactation periods in F0 females and their pups, as well as about one week after weaning in selected F1 offspring.

Clinical signs of parental toxicity included salivation in F0 males and females at 300 mg/kg bw. Food consumption was mildly increased in F0 females of all dose groups (13-16%). No clear trend could be observed for food consumtion of F0 males. In F0 males and females at the high dose level body weights were decreased by 7 and 6% at the end of the pre-mating period, respectively. 

In males, dose related target organ specific toxicity was identified by macroscopic and histopathological examinations in the cecum, kidneys and the liver. Histopathology further identified a diffuse atrophy in mammary glands of the high dose group.

Indications for target organ specific toxicity were less pronounced in females. In the high dose group, histopathological examinations identified substance related effects in the kidney (lymphoid infiltration, medullar mineralization) and liver (lymphoid infiltration).

High-dose F0 parental females (300 mg/kg bw/d) had a distinctly prolonged estrous cycle. All 8 pregnant females had a significantly lower average number of implants compared to the concurrent control (10.4 vs. 15.8). In addition, post-implantation loss was significantly increased (34.6 vs 3.6%) and 2 of the 8 pregnant females had complete intrauterine litter losses. These effects resulted in a significantly lower live litter size (10.3 vs. 15.0). Newborn pups developed normally.

Endpoint:
screening for reproductive / developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Qualifier:
according to
Guideline:
OECD Guideline 421 (Reproduction / Developmental Toxicity Screening Test)
Deviations:
yes
Remarks:
only seven pregnant females in the high dose group
GLP compliance:
yes
Remarks:
(Mitsubishi Chemical Safety Institute Ltd.)
Limit test:
no
Species:
rat
Strain:
Crj: CD(SD)
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Strain as specified in the report: Crj:CD(SD)IGS
- Source: Tsukuba Breeding Center, Charles River Laboratories Japan
- Age at study initiation: 9 weeks
- Body weight ranges at study initiation: 329 to 374 g in males and from 206 to 251 g in females
- Housing: Each one male and female in mating period, a litter of pups in lactation period and one animal during gestation period
- Diet: Autoclave sterilized CRF-1, Oriental Yeast Co., Ltd, ad libitum
- Water: Tap water filtrated with 5 µm pore size filter followed by ultra-violet radiation, ad libitum
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 2
- Humidity (%): 55 ± 15
- Air changes (per hr): 12
- Photoperiod (hrs dark / hrs light): 12 / 12
Route of administration:
oral: gavage
Vehicle:
other: 0.5% aqueous sodium carboxymethylcellulose solution with 0.1% Tween 80
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
The test substance was individually weighed for each dose and suspended using an agate mortar in 0.5% CMC-Na aqueous solution with 0.1% Tween 80 added. The administration solution was prepared once a week and kept in under dark and cold conditions to be used within 8 days after preparation. Before start of administration, the stability of the test substance in the administration solution of 1 and 200 mg/mL stored under cold and dark conditions was assured. In addition, the administration solutions for respective dose groups were analyzed at the time of first preparation to confirm that they were within +- 10% of the prescribed concentrations.

VEHICLE
- Justification for use and choice of vehicle (if other than water): The test substance is sparingly soluble in water.
- Amount of vehicle (if gavage): 5 mL/kg bw
Details on mating procedure:
- M/F ratio per cage: 1 : 1
- Length of cohabitation: 7 days
- Proof of pregnancy: vaginal plug or sperm in vaginal smear referred to as day 0 of gestation
- After successful mating each pregnant female was caged: individually
Analytical verification of doses or concentrations:
not specified
Details on analytical verification of doses or concentrations:
The administration solutions for respective dose groups were analyzed at the time of first preparation to confirm that they were within +- 10% of the prescribed concentrations (data not available).
Duration of treatment / exposure:
males in total 45 days (including 14 days pre-mating period), females in total 40 to 46 days from mating through gestational period and parturition to day 3 of lactation. Animals without delivery were administered until day 25 after confirmation of copulation.
Frequency of treatment:
daily
Dose / conc.:
10 mg/kg bw/day (actual dose received)
Dose / conc.:
60 mg/kg bw/day (actual dose received)
Dose / conc.:
300 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
12
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: In a preliminary test, the test substance was administered repeatedly for 14 days to each 3 male and female rats per group at doses of 0, 100, 300 and 1000 mg/kg bw., respectively. As a result, suppressed gain or decrease in body weight, decreased or low food intake and reduced weight of seminal vesicle were observed in males and females of the 300 and 1000 mg/kg bw groups. In addition, caecum abdominal distension was observed in the males of the 300 mg/kg bw group and in males and females of the 1000 mg/kg bw group. Furthermore, decreased weights of thymus and prostate gland were observed in males of the 1000 mg/kg group. Considering these results and the administration period in the main test, the high dose was determined as 300 mg/kg bw where clear toxicity is expected, and 3 doses in total including 60 and 10 mg/kg bw were set up at a common ratio of 5.
Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice a day (before and after administration)

BODY WEIGHT: Yes
- Time schedule for examinations: on the day 1, 3, 7, and 14 of test substance administration and once a week thereafter; additionally, females with successful copulation were weighed on day 0, 7, 14 and 20 of gestation period and on day 0 and 4 of lactation period. The body weight gain was calculated in males based on the body weight on day 1 and in females based on day 1 of test substance administration, and on day 0 of gestation as well as on day 0 of lactation, respectively, in pre-mating, gestation and lactation periods.

FOOD CONSUMPTION: Yes
- Mean food intake per animal per day was calculated for both males and females by measuring gross weight on the first day of gavage as well as on day 3, 7, and 14 of the treatment period, and for males once a week excluding mating period thereafter as well as females on day 0, 7, 14 and 20 of gestation period and day 0 and 4 of lactation period.

OTHER:
- The starting day of administration was counted as day 0 of pre-mating period, the day of successful mating as day 0 of pregnancy and the day of parturition as day 0 of lactation, respectively.
- Gestation status was observed twice a day from day 21 to 25 of parturition.
Oestrous cyclicity (parental animals):
Vaginal smears of females were collected every day in the morning starting with the first day of treatment to the day of confirmed copulation and subjected to eosin-thionine staining (with Hemacolor multi-purpose quick staining kit) to inspect their estrus cycle and calculate the number of mean estrus cycle days until successful copulation.
Females whose estrus cycle was not in a range from 4 to 6 days were considered as those of irregular estrus cycle.
Sperm parameters (parental animals):
Parameters examined in male parental animals:
testis weight, epididymis weight
Litter observations:
PARAMETERS EXAMINED
The following parameters were examined in F1 offspring:
- Total number of offspring at birth (number of live offspring at birth, number of dead offspring at birth), sex, external abnormalities were checked on day 0 of lactation. Thereafter, general conditions, death etc. were observed every day until day 4 of lactation.
- Body weight: Live offspring was individually measured on day 0 and 4 of lactation. In addition, body weight gain was calculated based on the body weight on day 0 of lactation.
- Anogenital distance: Anogenital distance (AGD) of all the live offspring was measured.
Furthermore, relative distance divided by cube root of body weight on day 4 of lactation (AGD / ∛body weight) was calculated to compensate fluctuation of measured values due to body weight balance among individual offspring.
- Neonates were lactated for 4 days after birth (day 4 of lactation period) to observe every day lactation conditions including lactation, nest-building, eventual cannibalism and etc.
Postmortem examinations (parental animals):
SACRIFICE
- Male animals: All surviving animals [day 45]
- Maternal animals: All surviving animals [day 4 of lactation]
- Animals without delivery were brought to necropsy on day 26 after confirmation of copulation.
GROSS NECROPSY
- Gross necropsy consisted of external and internal examinations including the cervical, thoracic, and abdominal viscera.
- Ovaries and uterus of dams were removed on necropsy to check the number of corpora lutea and number of implantation sites.

HISTOPATHOLOGY / ORGAN WEIGHTS
- In all animals, organs and tissues from liver, thymus, testes, epididymis, seminal vesicle, prostate, uterus, ovaries, pituitary, vagina and gross abnormal lesions were fixed with 10% neutral phosphate buffer formalin solution to be stored.
- In addition, abnormalities were found in caecum of 60 and 300 mg/kg bw groups and hence, caecum was taken as reference from all the males and one female of the control groups, while their testes and epididymis were fixed with Bouin liquid.
- In the histopathological test, hematoxylin and eosin-stained samples were prepared based on usual technique from the said organs and tissues stored of all the males and females of the control groups and 300 mg/kg bw group and subjected to microscopic examination. As a result, changes in liver of males and females caused by the test substance were observed and hence, livers of all the males and females in 10 and 60 mg/kg bw groups were examined.
Postmortem examinations (offspring):
SACRIFICE
- The F1 offspring was sacrificed at 4 days of age.
- These animals were subjected to postmortem examinations (macroscopic and/or microscopic examination) as follows:

GROSS NECROPSY
- After inspecting external appearance including oral cavity of all the live offspring on day 4 of lactation, they were euthanized similarly to their parent animals, followed by necropsy. After fixation in 10% neutral phosphate buffer formalin solution, animals were subjected to necropsy using a stereomicroscopy.
Statistics:
The measured data were subjected to Bartlett test for homogeneity of variances and they were subjected to one-way analysis of variances when variances were equal and to Kruskal-Wallis test when variances were not equal. In the case that significant differences were found among the groups, the data were subjected to multiple comparison according to Dunnett test. Some items were first subjected to Kruskal-Wallis test and also to multiple comparison of Dunnett type when significant differences were found among the groups. The data obtained in the histopathological test were subjected to x2 test for (a x b) or to Amitage x2 test when significant differences were found among the groups. Furthermore, Fischer’s exact test was performed. The significant level for each test was set as 5%. The target items for statistical analysis are as follows and the statistical analysis was not applied for general conditions and necropsy examination;
* Multiple comparison test: Body weight, body weight gain, food intake, organ weight, number of corpora lutea, number of implantation sites, number of offspring at birth, and anogenital distance
* Multiple comparison of Kruskal-Wallis and Dunnet type: Number of days required for copulation, number of estrus cycles which elapsed till successful copulation, mean number of days of estrus cycle, gestation period, implantation index, delivery index, live birth index, and viability index of offspring on day 4 after birth
* x2 test and Amitage x2 test: Histopathological test
Fischer’s exact test: Incidence index of females with irregular estrus cycle, copulation index, fertility index, gestation index, and sex (male/female)
Reproductive indices:
Copulation index (%): (Number of copulated animals / number of animals housed together) x 100
Fertility index (%): (Number of pregnant females / number of copulated animals) x 100
Gestation period: number of days from day 0 gestation to the day when the delivery was assured
Delivery index (%): (number of pregnant females delivered live offspring / number of pregnant females) x 100
Implantation index (%): (number of implantation sites / number of corpora lutea) x 100
Gestation index (%): (total number of offspring delivered / number of implantation sites) x 100
Offspring viability indices:
Live birth index (%): (Number of live offspring at birth / Total number of offspring at birth) x 100
Viability index (%) of offspring on day 4 after birth: (Number of live offspring on day 4 of lactation / number of offspring at birth) x 100
CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS)
- Salivation was observed immediately before or immediately after gavage in 7 males of the 300 mg/kg bw group from day 22 after start of administration to day 54 of administration and in one female from day 17 after start of administration to day 15 of gestation, but all of them recovered in approx. 30 minutes after administration.
- No abnormalities were found in any males and females of the 10 and 60 mg/kg bw groups.

BODY WEIGHT AND FOOD CONSUMPTION (PARENTAL ANIMALS)
- Body weight gain was significantly suppressed from day 3 (males, day 3: 85%, day 7: 62%, day 14: 40%, day 21: 32%, day 28: 23%, day 35: 21%) or day 7 (females, before mating: day 7: 75%, day 14: 39%, during gestation period: day 7: 20%, day 14: 22%, day 20: 23%) after start of administration and decrease in food consumption was observed in males and females of the 300 mg/kg bw group. Furthermore, significant difference from the control group was found in body weight in females on day 4 of lactation in 60 mg/kg bw group and low food intake were found on day 4 of lactation in 10 and 60 mg/kg bw groups.

REPRODUCTIVE FUNCTION: ESTROUS CYCLE (PARENTAL ANIMALS)
- In the estrus cycle test, extended cycles were observed in 300 mg/kg bw group and 5/12 animals showed a continued diestrus for 6 to 10 days, but all the cases succeeded in copulation and no significant differences from the control group were observed in terms of copulation index, number of days required for copulation and number of estrus cycles which elapsed till successful copulation. However, 4 out of the animals showing a continued diestrus in 300 mg/kg bw group did not conceive and revealed a declined fertility index (58.3%).
The animals of the 10 and 60 mg/kg bw groups showed no significant differences from the control group in every inspection item and index.

REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS)
- No changes attributable to the compound were observed in parameters such as mating index, delivery index, gestation index, number of corpora lutea, gestation length, parturition state, or lactation behaviour.

ORGAN WEIGHTS (PARENTAL ANIMALS)
- Relative weights of liver (12%) and pituitary gland (19%) were increased and absolute weight of seminal vesicles (14%) were decreased in males of 300 mg/kg bw group (significantly different from the control group).
No significant differences from the control group were found in males and females of the 10 and 60 mg/kg bw groups.

GROSS PATHOLOGY (PARENTAL ANIMALS)
- Distension of the caecum was observed at each one male and female of the 60 mg/kg bw group and all males (12/12) and 4/12 females of the 300 mg/kg bw group.

HISTOPATHOLOGY (PARENTAL ANIMALS)
- Changes to be attributable to the test substance were observed in caecum and liver of males and females of all test groups. Mucosal epithelium diffusion in caecum was observed in 1/12 females of the 60 mg/kg bw group and 11/12 males and 4/12 females of the 300 mg/kg bw group, and hyperplasia and diffusion of absorptive epithelium were significant, and single cell necrosis of absorptive epithelium was found in 5/12 males and 1/12 females of the 300 mg/kg bw group. No clear change was observed in one male of the 60 mg/kg bw group in which distension of the caecum was found at necropsy.
- In liver, hypertrophy of centrolobular hepatocytes was observed in 5/12 males and 3/12 females without pregnancy of the 300 mg/kg bw group, and extramedullary hematopoiesis was observed in males and females of each group including the control group.
- Furthermore, inflammatory cell infiltration, micro-granuloma, focal necrosis and single cell necrosis in liver, thymus atrophy, pyelectasis, sperm-granuloma in epididymis, and lymphocyte infiltration in prostate were occasionally observed in each group including the control group, but no clear tendency to increasing abnormalities in 300 mg/kg bw group was found and thus, the effects were assumed not to be test substance related.
In addition, no histological abnormalities were found in the pituitary gland exhibiting an enhanced relative weight and in the seminal vesicle with a decreased absolute weight in males.
- No abnormalities were found in genitals of infertile males and females.

OTHER FINDINGS (PARENTAL ANIMALS)
- No change in the number of corpora lutea was observed in the 300 mg/kg bw group but the number of implantation sites tended to decrease, showing a significant decrease in the implantation index (64.89%) from the control group (95.80%). In addition, the total number of offspring (9.1) in the 300 mg/kg bw group was not significantly different but it was lower than the control group (14.3).
- Furthermore, gestation index in each group including the control group was 100%, and no significant differences from the control group were observed in terms of gestation period and delivery index.
- No abnormalities was observed in lactation behaviour of each maternal animal including the control group.
Dose descriptor:
NOAEL
Remarks:
for parental toxicity
Effect level:
10 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
gross pathology
histopathology: non-neoplastic
Dose descriptor:
NOAEL
Remarks:
for reproductive toxicity
Effect level:
60 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
female
Basis for effect level:
other: estrous cycles, fertility index, number of implantations, implantation index, number of live births and number of live offspring
VIABILITY (OFFSPRING)
- Total number of offspring at birth, number of live offspring at birth and number of live offspring on day 4 of lactation were decreased in the 300 mg/kg bw group at systemic parental toxicity.
- In addition, no significant difference in live birth index and viability index on day 4 was observed between the test substance administered group and the control group, and no abnormalities in external appearance and general conditions were found in offspring of each group including the control group.
- No changes attributable to the compound were observed in parameters including the sex ratio, the live birth index, and viability index on day 4.

BODY WEIGHT (OFFSPRING)
- No significant differences in male/female body weight and body weight gain were observed between the control group and the treatment groups.

SEXUAL MATURATION (OFFSPRING)
- No significant differences in male/female anogenital distance were observed between the control group and the treatment groups.

GROSS PATHOLOGY (OFFSPRING)
- No abnormalities in each group including the control group were observed in dead offspring on day 0 to 4 of lactation and live offspring on day 4 of lactation.
Key result
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
60 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: total number of offspring
Reproductive effects observed:
not specified

Table 1 Body weight of rats during gavage of the test substance (Body weight gain in brackets)

Dose

[mg/kg bw]

 

Day of treatment

 

Males

0

3

7

14

21

28

35

42

 

 

 

No.

Mean body weight [g ± SD]

 

 

 

0

12

351.9 ± 14.3

373.8 ± 15.5

(21.9 ± 4.8)

395.9 ± 16.8

(44.0 ± 7.9)

435.9 ± 24.4

(84.0 ± 18.6)

457.8 ± 27.1

(105.9 ± 22.1)

482.1 ± 31.0

(130.2 ± 25.5)

499.8 ± 33.9

(147.9 ± 29.3)

511.8 ± 33.1

(159.9 ± 28.4)

 

 

10

12

354.3 ± 12.5

373.5 ± 14.9

(19.2 ± 5.0)

397.4 ± 16.9

(43.1 ± 6.6)

435.9 ± 20.8

(81.6 ± 11.9)

458.7 ± 27.4

(104.3 ± 18.9)

482.7 ± 27.3

(128.3 ± 18.0)

500.4 ± 31.4

(146.1 ± 23.1)

514.5 ± 36.4

(160.2 ± 28.2)

 

 

60

12

352.5 ± 10.6

373.7 ± 11.7

(21.2 ± 6.8)

396.8 ± 13.1

(44.3 ± 8.9)

437.9 ± 19.9

(85.4 ± 15.9)

458.2 ± 24.4

(105.7 ± 20.1)

486.2 ± 28.7

(133.7 ± 22.9)

507.3 ± 31.1

(154.8 ± 24.8)

523.5 ± 34.0

(171.0 ± 28.1)

 

 

300

12

354.2 ± 11.1

357.5 ± 14.4*

(3.3 ± 12.7**)

370.8 ± 23.4**

(16.6 ± 20.8**)

404.5 ± 30.6**

(50.3 ± 26.7**)

426.6 ± 32.4*

(72.4 ± 28.3**)

454.2 ± 37.6

(100.0 ± 32.9*)

471.3 ± 45.4

(117.1 ± 40.5*)

486.0 ± 54.3

(131.8 ± 49.2)

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Premating period

Day of gestation

Day of lactation

 

Females

0

3

7

14

0

7

14

20

0

4

 

No.

Mean body weight [g ± SD]

 

 

 

0

12/11/11

229.1 ± 9.5

236.0 ± 11.4

(6.9 ± 7.7)

244.9 ± 12.3

(15.8 ± 6.6)

264.2 ± 11.9

(35.1 ± 6.9)

272.8 ± 13.5

 

315.8 ± 14.6

(43.0 ± 5.4)

352.6 ± 14.9

(79.8 ± 5.5)

436.5 ± 22.4

(163.6 ± 14.4)

325.5 ± 22.0

360.1 ± 17.1

(34.5 ± 13.9)

10

12/11/11

228.4 ± 11.6

237.3 ± 13.1

(8.8 ± 6.1)

245.8 ± 13.9

(17.4 ± 6.4)

263.8 ± 17.2

(35.4 ± 9.8)

277.1 ± 19.6

 

320.7 ± 18.4

(43.6 ± 5.8)

358.5 ± 21.8

(81.4 ± 10.9)

433.1 ± 36.7

(156.0 ± 30.1)

327.5 ± 31.2

354.1 ± 20.9

(26.5 ± 17.2)

60

12/12/12

228.2 ± 9.2

233.5 ± 13.2

(5.3 ± 9.4)

241.6 ± 12.8

(13.4 ± 7.8)

262.3 ± 16.3

(34.1 ± 10.9)

266.8 ± 12.9

 

304.9 ± 16.0

(38.2 ± 6.9)

338.7 ± 18.2

(71.9 ± 10.3)

418.6 ± 21.9

(151.8 ± 13.2)

314.3 ± 25.2

333.0 ± 24.2*

(18.7 ± 16.2)

300

12/7/7

230.3 ± 12.5

228.8 ± 15.7

(-1.5 ± 10.5)

234.3 ± 15.9

(4.0 ± 10.4**)

251.7 ± 16.1

(21.4 ± 10.0**)

264.4 ± 16.9

 

298.9 ± 19.6

(34.4 ± 6.5*)

326.9 ± 19.9*

(62.4 ± 7.2**)

390.4 ± 29.2**

(126.0 ± 17.9 **)

316.0 ± 15.0

338.6 ± 20.0

(22.6 ± 8.8)

Significantly different from control group (*: p<0.05; **: p<0.01)

Table 2 Reproductive performance

Dose

[mg/kg bw]

Number of animals

Mean oestrus cycle (± SD)

Incidence of females with irregular oestrus cycle

Copulation index (%)

Fertility index (%)

0

12

4.08 ± 0.21

0/12

100.0

91.7

10

12

4.01 ± 0.11

0/12

100.0

91.7

60

12

4.14 ± 0.34

1/12

100.0

100.0

300

12

5.57 ± 1.81**

5/12*

100.0

58.3

Significantly different from control group (*: p<0.05; **: p<0.01)

 

Table 3 Delivery data

Dose

[mg/kg bw]

Number of animals

Number of corpora lutea

Number of implantation sites

(Mean ± SD)

Total number of offspring

(Mean ± SD)

Implantation index (%)

Delivery

index (%)

0

11

16.6 ± 2.0

15.9 ± 1.9

14.3 ± 2.1

95.80 ± 5.75

90.03 ± 10.08

10

11

15.9 ± 3.6

13.3 ± 5.3

12.5 ± 5.0

80.84 ± 26.01

94.60 ± 8.19

60

12

17.3 ± 2.1

14.8 ± 2.0

13.5 ± 1.8

86.15 ± 7.76

91.22 ± 7.71

300

7

15.7 ± 2.8

10.7 ± 5.5

9.1 ± 4.2

64.89 ± 26.57**

89.75 ± 12.11

Significantly different from control group (*: p<0.05; **: p<0.01)

 

Table 4 Litter size (Sex ratio in brackets)

Dose

[mg/kg bw]

Number of animals

Total number of offspring at birth

(Mean ± SD)

Number of live offspring at birth

(Mean ± SD)

Number of live offspring on day 4

(Mean ± SD)

males

females

total

males

females

total

total

0

11

6.6 ± 1.3

7.6 ± 2.7

14.3 ± 2.1

(73/84)

6.5 ± 1.3

7.6 ± 2.7

14.2 ± 2.1

(72/84)

14.1 ± 2.2

(72/83)

10

11

7.3 ± 3.3

5.2 ± 3.3

12.5 ± 5.0

(80/57*)

7.3 ± 3.3

5.2 ± 3.3

12.5 ± 5.0

(80/57*)

12.4 ± 5.2

(79/57*)

60

12

6.8 ± 1.2

6.8 ± 1.7

13.5 ± 1.8

(81/81)

6.7 ± 1.2

6.8 ± 1.7

13.4 ± 1.7

(80/81)

13.3 ± 1.6

(80/80)

300

7

4.1 ± 1.7*

5.0 ± 2.7

9.1 ± 4.2

(29/35)

4.1 ± 1.7*

5.0 ± 2.7

9.1 ± 4.2

(29/35)

9.1 ± 4.2

(29/35)

Significantly different from control group (*: p<0.05; **: p<0.01)

Conclusions:
Based on the test results, the NOAEL for parental and reproductive toxicity was considered to be 60 mg/kg bw/day.
Executive summary:

In the repeated dose oral toxicity study according to GLP and OECD 421 (Reproduction/Developmental Toxicity Screening Test), rats (12/sex/dose) were administered a repeated oral dose (gavage) of the test item (analytical purity: 99.87%) at dose levels of 10, 60 and 300 mg/kg body weight. The test item was suspended in vehicle (0.5% aqueous sodium carboxymethylcellulose solution with 0.1% Tween 80) and administered at a volume of 5 mL/kg bw.

The animals were observed daily. Body weight measurements and determination of reproductive parameters as well as pathological examinations were performed in parental animals. Viability, body weight, sexual maturation were examined and after sacrifice on day 4 of lactation offspring was subjected to gross pathology.

Salivation was observed in the high dose group immediately following test substance application. Reduced food intake and body weight gain from day 3 after start of administration to the time of necropsy were observed in males and females of the high dose group. At necropsy, abdominal distension of the caecum was observed in males and females of the mid and high dose group, mostly with mucoepithelial cell necrosis, which might be a reaction of the absorptive epithelium to physical stimulus. Similar results were found in the 28 -day repeated dose oral toxicity study in rats, thereof indicating that the effects might be similar changes caused by the test substance. Furthermore, increased relative liver weight observed in the males of the high dose group as well as centrolobular hypertrophy of hepatocytes found in both the males and females, may indicate enzyme induction, which is often observed after administration of chemical substances causing induction of drug-metabolizing liver enzymes. Hypertrophy of hepatocytes was only found in non-pregnant females, and it is assumed that hepatocytes of delivering females are enlarged due to lactation and therefore conceal the test substance induced effects.

With respect to cecum effects in rats the relevance for humans is unclear. Significant anatomical and functional differences of the cecum exists for rodents and humans. In rodents this anatomical structure is large and in comparison to humans has a significant function in digestion. Nevertheless these disturbances in the content and structure of the gastrointestinal tract in rats may have a significant impact e.g. on body weight development.

No abnormalities were observed in parental animals of the high dose group with regard to copulation index, parturition index, delivery index, number of corpora lutea, gestation period, parturition status and lactation behaviour, but extended oestrus cycle, increased number of animals showing irregular oestrus cycle, tendency to decreased number of implantation sites, and significantly decreased implantation index were observed. Most of the females exhibiting continued dioestrus were not fertilized and tendency to decreased fertility index was noticed. Since no effects on delivery index and live birth index were observed in the high dose group, the decrease in fertility index and total number of offspring at birth may partly be attributable to the increased pre-implantation loss. No histopathological abnormalities were found in pituitary glands in males of the high dose group, exhibting increased relative weight and in seminal vesicles with decreased absolute weight. Moreover, no abnormalities possibly causing infertility were found in histological examinations of genitals of both males and females and thus, the reasons for decreased reproductive performance in paternal animals remained unclear.

No effects were observed with respect to growth of the offspring after birth, as no abnormalities were found in sex ratio (male/female), body weight, viability index of pups on day 4 after birth, anogenital distance, general conditions, external appearance and necropsy findings.

Since the relevance of rat cecum effects is questionable for humans and no other adverse efects occurred at 60 mg/kg bw, the dose is considered to be a NOAEL for parental toxicity.

Based on the distension in caecum and histological changes in caecum observed in males and females of the mid and high dose group, the no-observed adverse effect level regarding repeated oral administration of the test substance is10 mg/kg bw/day for both, males and females.

Based on extended oestrus cycle, increased number of animals showing continued dioestrus, decreased fertility index, number of implantation sites, implantation index, and total number of live offspring at birth observed in the high dose group, the no-observed adverse effect level for reproductive development is 60 mg/kg bw/day for parental animals and pups.

Effect on fertility: via oral route
Dose descriptor:
NOAEL
60 mg/kg bw/day
Effect on fertility: via inhalation route
Endpoint conclusion:
no study available
Effect on fertility: via dermal route
Endpoint conclusion:
no study available
Additional information

There is one study available (OECSEH, 2001) which was performed according to OECD guideline 421 (Reproduction / Developmental Toxicity Screening Test), however with limited documentation. 12 Crj:CD (SD) rats per sex per dose were orally administered with the test substance 4,4`-sulphonyldiphenol at daily dose levels of 0, 10, 60 and 300 mg/kg bw.

Male rats were treated from 14 days before mating to the day before necropsy (including the mating period; 45 days in total) and female rats from 14 days before mating to day 3 of lactation (including the mating period, gestation period, and delivery; a total of 40 to 46 days). The animals were terminally sacrificed on day 45 (males) and on day 4 of lactation (females).

Distinct indications of maternal toxicity were observed in the mid and high dose group (60, 300 mg/kg bw). A suppression in body weight gain in the high dose group is resulting from a decrease in food consumption. Furthermore, cecum distention including histopathological lesions in the cecum epithelium occurred in the mid and high dose and a liver enlargement accompanied by centrilobular hypertrophy was identified in the high dose group.

With respect to the observed cecum effects in rats and in accordance to the subchronic assays, the relevance for humans is unclear. Significant anatomical and functional differences of the cecum exist for rodents in comparison to humans. In rodents this anatomical structure is large and in comparison to humans has a significant function in digestion. Nevertheless, it is apparent, that these disturbances in the content and structure of the gastrointestinal tract in rats have a significant impact e.g. on body weight development.

In parent animals in the 300 mg/kg bw/day group, there were no test article-related changes in the copulation index, delivery index, gestation index, number of corpora lutea, length of gestation period, delivery condition, or lactation behavior. However, at a systemic toxic dose (300 mg/kg bw/day) there were significant prolongation of estrous cycle, increase in the number of animals that showed prolonged diestrus period, and decrease in the implantation index at 300 mg/kg bw/day. A tendency toward decrease in the fertility index was also observed at 300 mg/kg bw/day.

In offspring in the 300 mg/kg bw/day group, the total number of offspring delivered, number of live offspring and the number of offspring alive on day 4 of lactation tended to be low, but these effects were considered to be due to decreased implantation index. In the 300 mg/kg bw/day group, there were no test article-related changes in the live-birth index, sex ratio, body weight, viability index on day 4 after birth, anogenital distance, general condition, external findings or necropsy findings. In the 10 and 60 mg/kg bw/day groups, there were no test article-related changes in parent animals or offspring.

Since the relevance of rat cecum effects is questionable for humans and no further adverse effect was observed at 60 mg/kg bw, this dose is considered to be a NOAEL for parental toxicity. Based on a prolongation of estrous cycle and decreased implantation index the NOAEL for reproductive toxicity was also considered to be 60 mg/kg bw/day.

 

The final Decision on substance evaluation acknowledges the possibility of strain specificities, as“…the same rat strain used for the OECD 421 (Sprague Dawley) shall be tested … to avoid any difference in sensitivity between rat strains”. Due to the change in the testing rat strain prescribed in the Decision on substance evaluation pursuant to Article 46(1) of Regulation (EC) No 1907/2006, a 28-day range finding experiment (bridging study) was performed. For 4,4`-sulphonyldiphenol no control data in non-mated and mated SD rats was available in the executing laboratory. In addition, rat strain or source specific differences in sensitivity could not be excluded based on the existing data, and the existing data for SD rats from an unknown source is of limited transferability to the present testing campaign. For a reliable identification of the dose levels and an unambiguous interpretation of the test results from a definitive study it appears inevitable to generate lab internal control data for 4,4`-sulphonyldiphenol in non-mated and mated SD rats from the specific supplier.

 

Therefore, a Repeated-dose 28-day toxicity study in SD rats (bridging study) was performed.

Administration by gavage

5 male and female Sprague Dawley rats were dosed at 0, 100, 300 and 600 mg/kg bw/ day by gavage. A 0.5% Carboxymethylcellulose suspension in drinking water was used as vehicle. The following parameters were investigated: detailed clinical examinations, food and drinking water consumption, body weight, organ weights and organ/tissue fixation of the target organs adrenal glands, kidneys, liver, all other organs were weighted and fixed (brain, epididymides, heart, ovaries, prostate, seminal vesicles with coagulating glands, spleen, testes, thymus, thyroid glands, uterus with cervix), histopathological assessment of gastro-intestinal tract incl. caecum, liver, kidneys and adrenal glands. Salivation was observed in all test groups treaded with test substance (21/30 animals). Clinical chemistry revealed no relevant findings. Changes in body and organ weights were the following:

-    dose dependent and statistically significant decrease of terminal body weight in males (-9 at 300mg/kg bw, -12% at 600mg/kg bw).

-    increase of mean relative weights of the kidneys in males (+33% at 300 and 600 mg/kg bw, statistically significant at 600 mg/kg bw) and females (+12% at 300 mg/kg bw and 600 mg/kg bw, statistically significant at 600 mg/kg bw).

-    not statistically significant increase in relative weights of the adrenal glands in males (+18 and +39% at 300 and 600 mg/kg bw, respectively)

-    dose dependent and statistically significant increase of the mean relative weights of the liver in females (+9 and 12% at 300 and 600 mg/kg bw, respectively).

-    decrease in relative weights of prostate and seminal vesicles at 600 mg/kg bw but not to a statistical significance (-15% and -16%, respectively).

-    dose dependent but not statistically significant decrease in thymus weights in males and females

Enlarged kidneys in 4/5 male animals at 600 mg/kg bw and 3/5 male animals at 300 mg/kg bw as well as cecum dilation in in 2/5 male animals at 600 mg/kg bw were observed. The following histopathology findings were observed:

-    tubular degeneration/regeneration in the kidney of all male animals at 300 and 600 mg/kg bw graded minimal to moderate and in 2/5 male animals at 100 mg/kg bw graded minimal or slight.

Tubular hypertrophy in the kidney in 5/5 males at 600 mg/kg bw graded mostly moderate, in 5/5 males at 300 mg/kg bw graded mostly minimal and in 1/5 male at 100 mg/kg bw graded mostly minimal.

-    minimal hypertrophy/hyperplasia in the adrenal gland cortex in 3/5 male animals at 600 mg/kg bw

-    centrilobular hypertrophy of the liver

at 600 mg/kg bw in all male animals graded up to moderate and in all female animals graded mostly slight

at 300 mg/kg bw in 4/5 male animals graded slight,

at 100 mg/kg bw in 1/5 male animal graded minimal.

-    dilation of the cecum in 5/5 male and 2/5 female animals at 600 mg/kg bw

-    diffuse atrophy of the mammary gland in 4/5 male animals at 600 mg/kg bw and in 3/5 male animals at 300 mg/kg bw.

The target organs identified in the existing repeated dose studies for 4,4`-sulphonyldiphenol were confirmed (28 day gavage study, Reproduction / Developmental Toxicity Screening Test in SD rats, OECSEH 1999, 2001; 90 day gavage study in Wistar rats, BASF, 2015). The kidney appears to be more susceptible to DHDPS related effects in SD-rats compared to Wistar rats.

 

In a Reproduction/Developmental Toxicity Screening Study which was performed as a Range-Finding Study for an Extended One Generation Reproductive Toxicity study (EOGRTS), 4,4'-sulphonyldiphenol was orally administered via gavage to groups of 10 male and 10 female Sprague-Dawley rats (F0 animals) at doses of 0, 30, 100 and 300 mg/kg bw/d (BASF, 2017). The duration of treatment covered a 6-week premating and a 14-days mating period in both sexes, approximately 4 weeks post-mating in males, the entire gestation and lactation periods in F0 females and their pups, as well as about one week after weaning in selected F1 offspring.

Clinical signs of parental toxicity included salivation in F0 males and females at 300 mg/kg bw. Food consumption was mildly increased in F0 females of all dose groups (13-16%). No clear trend could be observed for food consumption of F0 males. In F0 males and females at the high dose level body weights were decreased by 7 and 6% at the end of the pre-mating period, respectively. 

In males, dose related target organ specific toxicity was identified by macroscopic and histopathological examinations in the cecum and kidneys. Liver enlargement correlated with an increase in liver weight. Histopathology further identified a diffuse atrophy in mammary glands of the high dose group.

Indications for target organ specific toxicity were less pronounced in females. In the high dose group, histopathological examinations identified substance related effects in the kidney (lymphoid infiltration, medullar mineralization) and liver (lymphoid infiltration).

High-dose F0 parental females (300 mg/kg bw/d) had a distinctly prolonged estrous cycle. All 8 pregnant females had a significantly lower average number of implants compared to the concurrent control (10.4 vs. 15.8). In addition, post-implantation loss was significantly increased (34.6 vs 3.6%) and 2 of the 8 pregnant females had complete intrauterine litter losses. These effects resulted in a significantly lower live litter size (10.3 vs. 15.0). Newborn pups developed normally.

 

According to the final Decision on substance evaluation an Extended One Generation Reproductive Toxicity Study in Sprague-Dawley rats, oral route, according to test method OECD 443, with the developmental neurotoxicity and immunotoxicity (DNT/DIT) cohorts and with the conditional extension of Cohort 1B to mate the F1 animals to produce an F2 generation has been conducted.

24 Sprague-Dawley rats per sex per dose were orally administered with the test substance 4,4`-sulphonyldiphenol at daily dose levels of 0, 20, 60 and 180 mg/kg bw (F0 generation parental animals). These animals were mated to produce F1 generation pubs. Pubs of the F1 litter (F1 rearing animals) were orally dosed with 0, 20, 60 and 180 mg/kg bw/day test substance and assigned to 5 different cohorts which were subjected to specific postweaning examinations. Cohort 1B (=F1 generation parental animals) were selected to produce F2 pubs. The in-life phase of the study iscompleted, however histopathological evaluationis still ongoing. As the OECD 443 is a modular study, e.g. certain histopathological findings triggered additional histopathological investigations. Results of the study will be reported in a dossier update as soon as all the results and the final report will be available.

Effects on developmental toxicity

Description of key information

No effects on the development of rats were identified in a prenatal developmental toxicity study performed with 4,4’-sulfonyldiphenol in accordance to the OECD 414 guideline as well as in reproduction/developmental toxicity screening tests performed according to OECD 421 or similar to OECD 422 guideline.


Link to relevant study records
Reference
Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
10 Sep 2013 - 28 Feb 2014
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.3700 (Prenatal Developmental Toxicity Study)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Limit test:
no
Species:
rat
Strain:
Wistar
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Research Models and Services, Germany GmbH
- Age at study initiation: about 11-13 weeks
- Weight at study initiation:
- Housing: Makrolon cages type M III (1 animal per cage)
- Diet (e.g. ad libitum): Ground Kliba maintenance diet mouse/rat "GLP", Provimi Kliba SA, Kaiseraugst, Switzerland; ad libitum
- Water (e.g. ad libitum): yes
- Acclimation period: from GD 0 (day of supply) to the beginning of administration (GD 6), the animals were accustomed to the environmental conditions and to the diet.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24°C;
- Humidity (%): 30-70%
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12
Route of administration:
oral: gavage
Vehicle:
CMC (carboxymethyl cellulose)
Remarks:
1 % suspension in drinking water
Details on exposure:
Gavage exposure from gestation day (GD) 6 through GD 19.

PREPARATION OF DOSING SOLUTIONS:
10 ml/kg body weight were administered. Test substance preparations were suspended in 1 % Carboxymethylcellulose in drinking water.
The aqueous test substance preparations were prepared at the beginning of the administration period and thereafter at maximum intervals of 7 days, which took into account the period of established stability. The preparations were kept in a refrigerator.
For the test substance preparations, the specific amount of test substance were weighed, topped up with 1% Carboxymethylcellulose suspension in drinking water in a calibrated beaker and intensely mixed with a homogenizer.
During administration the preparations were kept homogeneous with a magnetic stirrer.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Analytical investigations of the test substance preparations, and verification of stability of the test substance in 1 % Carboxymethylcellulose suspension in drinking water over a period of a maximum of 7 days prior to the start of the study (Project No.: 01Y0066/05Y009).
Details on mating procedure:
The animals were paired by the breeder (time-mated animals) and were supplied at noon on the day of evidence of mating. This day is referred to as GD 0 and the following day as GD 1.
Duration of treatment / exposure:
Gavage exposure from gestation day (GD) 6 through GD 19.
On GD 20, blood samples were obtained from dams by retrobulbar venous puncture.
Following blood sampling on GD 20, all dams were sacrificed and examined. Fetuses were removed from the opened uterus.
Frequency of treatment:
daily
Dose / conc.:
30 mg/kg bw/day (actual dose received)
Dose / conc.:
100 mg/kg bw/day (actual dose received)
Dose / conc.:
300 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
25 females
Control animals:
yes, concurrent vehicle
Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes, at least once daily for any signs of morbidity, pertinent behavioral changes and/or signs of overt toxicity.

BODY WEIGHT: Yes
- Time schedule for examinations: on GD 0, 1, 3, 6, 8, 10, 13, 15, 17, 19 and 20.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes, for GD 0-1 , 1-3, 3-6, 6-8, 8-10, 10-13, 13-15, 15-17,
17-19 and 19-20.

POST-MORTEM EXAMINATIONS: Yes, on GD 20, following anesthetisia with isoflurane, sacrificion by decapitation.
Fetuses were be removed from the uterus.
- Organs examined: Adrenal glands, Kidneys, Liver, Spleen. Organ / Tissue fixation for all gross lesions and organs examined.

HEMATOLOGY /CLINICAL CHEMISTRY: Yes
Blood samples were taken from animals by puncturing the retrobulbar venous plexus following isoflurane anesthesia.
Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Site of implantations in the uterus
Fetal examinations:
- Weight of each fetus
- Sex
- Weight of the placentas
- Gross-pathological examination of the fetuses after dissection from the uterus (including abnormalities of the fetal membranes, placentas, amniotic fluid and umbilical cord);
- fetuses wer sacrificed by subcutaneous injection of pentobarbital, and about half of the fetuses of each dam were skinned, fixed in ethyl alcohol and, after fixation, stained according to a modified method (KIMMEL and TRAMMELL) to show the skeleton and the cartilage. The other half of the fetuses of each dam were fixed in Harrison's fluid.
- examination of skeletons/cartilage and soft tissues.
Statistics:
Food consumption, body weight, organ weights - DUNNETT's test.
Implantations, resorptions and live fetuses - DUNNETT's test.
Number of pregnant animals, mortality and number of litters with fetal findings - FISHER's exact test.
Proportion of fetuses with findings per litter - WILCOXON test.
Clinical pathology parameters - KRUSKAL-WALLIS and WILCOXON test.
Details on maternal toxic effects:
Maternal toxic effects:yes

Details on maternal toxic effects:
There were no test substance-related or spontaneous mortalities in any females of all test groups.
Seven (out of 25) high-dose females showed transient salivation during major parts of the treatment period. Salivation persisted in the respective animals only for some minutes after daily gavage dosing (i.e. up to 20 minutes) and was initially observed on GD 10.
No clinical signs or changes of general behavior, which may be attributed to the test substance, were detected in any female of the low- and the mid-dose groups during the entire study period.

The mean food consumption of the high-dose dams was statistically significantly reduced at the beginning of the treatment period (GD 6-13; up to 15% below control), but recovered afterwards.
If calculated for the entire treatment period (GD 6-19) or the entire study period (GD 0-20), the high-dose dams consumed 8% or 6%, respectively, less food in comparison to the concurrent control group.
The mean food consumption of the dams in test groups 1 and 2 (30 or 100 mg/kg bw/d) was generally comparable to the concurrent control throughout the entire study period. The only exception was a slightly, but statistically significantly lower mean food consumption of the low- and high-dose dams on GD 0-1, which was a solitary event and considered to be accidental.

The mean body weights of the high dose dams were in general comparable to the controls throughout the entire study period.
The body weight change of the high-dose dams was statistically significantly reduced on GD 8-10 (approx. 29% below control) and if calculated for the entire treatment period (GD 6-19; 8% below control).
The mean body weights and the average body weight gains of the low- and mid-dose dams were in general comparable to the controls throughout the entire study period.
The corrected body weight gain (terminal body weight on GD 20 minus weight of the unopened uterus minus body weight on GD 6) of high dose was clearly lower than the concurrent control value (approx. 10% below control), but without attaining statistical significance.
The corrected body weight gain of low and mid dose groups revealed no difference of any biological relevance to the corresponding control group.
Moreover, mean carcass weights of all test groups remained unaffected by the treatment.

Mean gravid uterus weights of the animals of all test groups were not influenced by the test substance.

No necropsy findings which could be attributed to the test substance were seen in any dam.
In the high dose spontaneous findings were noted in individual females of the high dose group (a diaphragmatic hernia (this female was not pregnant), a dilated renal pelvis, and a hemometra).

The conception rate varied between 96% in the high dose group 3 and 100% in the other test groups. With these rates, a sufficient number of
pregnant females were available for the purpose of the study.
There were no test substance-related and/or biologically relevant differences in conception rate, in the mean number of corpora lutea and implantation sites or in the values calculated for the pre- and the postimplantation losses, the number of resorptions and viable fetuses.
Dose descriptor:
NOAEL
Effect level:
100 mg/kg bw/day (nominal)
Based on:
test mat.
Basis for effect level:
other: maternal toxicity
Dose descriptor:
NOAEL
Effect level:
300 mg/kg bw/day (nominal)
Based on:
test mat.
Basis for effect level:
other: developmental toxicity
Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:yes

Details on embryotoxic / teratogenic effects:
The sex distribution of the fetuses in all dose groups was comparable to the control fetuses.
The mean placental weights of all dose groups were comparable to the corresponding control group.
The mean fetal weights of all dose groups were not influenced by the test substance and did not show any biologically relevant differences in
comparison to the control group.

No soft tissue malformations were recorded.
Some soft tissue variations were detected in all test groups including the control, i.e. short innominate, dilated renal pelvis and dilated ureter. The incidences of these variations were neither statistically significantly different from control nor dose-dependent and therefore, not considered biologically relevant. Most of them can be found in the historical control data at comparable incidences.
No unclassified soft tissue observations were recorded.

One mid-dose fetus with multiple external malformations was seen (misshapen head and absent face). The overall incidences of external malformations were comparable to those found in the historical control data. An association of these findings to the treatment is not assumed.
No external variations were recorded.
One unclassified external observation, i.e. blood coagulum around placenta, was recorded in two fetuses of the high-dose group. This finding was not considered biologically relevant.

Skeletal malformations were detected in all test groups exept low dose group (0, 100 and 300 mg/kg bw/d) affecting the skull, sternum and forelimbs.
One fetus of the mid dose group (same animal as external malformation) was multiple-malformed (malformations affected the skull, vertebral column, ribs, pelvic girdle and forelimbs) and had associated external findings. The average rate of affected fetuses per litter showing skeletal malformations was statistically significantly increased in the high-dose group. However, each of the findings leading to that increased rate ist present in the historical control data and no abnormality pattern became obvious. The total incidences of skeletal malformations in the low- and mid-dose groups were comparable to the concurrent control group. These scattered observations in individual fetuses of these groups are are common for this rat strain.

For all test groups, skeletal variations of different bone structures were observed, with or without effects on corresponding cartilages. The observed skeletal variations were related to several parts of fetal skeleton and appeared without a relation to dosing. The overall incidences of skeletal variations were comparable to the historical control data, and no dose dependency was observed.
Additionally, some isolated cartilage findings without impact on the respective bony structures, which were designated as unclassified cartilage observations, occurred in all test groups. The observed unclassified cartilage findings were related to the skull, the sternum and ribs and did not show any relation to dosing. The overall incidences of skeletal unclassified cartilage observations in the substance-treated groups did not differ significantly from the concurrent control group.
Key result
Dose descriptor:
NOAEC
Effect level:
300 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: no observed effects at highest dose level
Abnormalities:
not specified
Developmental effects observed:
not specified

Table 1: Terminal body weights, food consumption and relative organ weights [g].

dose [mg/kg bw/day]

0

30

100

300

food consumption (0-20)

19.3

19.1

19.3

18.2

absolute terminal bw

295.9

302.4

297.8

291

corrected bw gain

40.9

43.7

40

36.9

carcass weight

236.8

242.8

239.1

235.2

bw change 6 -19

85.2

89.8

84.3

78.6*

 

Conclusions:
Under the conditions of this prenatal developmental toxicity study, the oral administration of DHDPS to pregnant Wistar rats from implantation to one day prior to the expected day of parturition (GD 6-19) at a dose of 300 mg/kg bw/d caused evidence of maternal toxicity, such as reduced food consumption and (net) body weight gain. In conclusion, the no observed adverse effect level (NOAEL) for maternal toxicity is 100 mg/kg bw/d.
There were no toxicologically relevant adverse fetal findings evident. Thus, the no observed adverse effect level (NOAEL) for prenatal developmental toxicity is 300 mg/kg bw/d.
Executive summary:

In a prenatal developmental toxicity study the test substance DHDPS was administered to pregnant Wistar rats daily by gavage from implantation to one day prior to the expected day of parturition (GD 6-19) to evaluate its potential maternal and prenatal developmental toxicity.

Analyses confirmed the correctness of the prepared concentrations, the homogeneous distribution and the stability of the test substance in the vehicle.

The test substance caused no mortality nor clinical symptoms of systemic toxicity in any of the exposed groups receiving 30, 100 or 300 mg/kg bw/d DHDPS. Some females (7 out of 25) of the high-dose group (300 mg/kg bw/d) showed transient salivation after treatment. This salivation persisted in the respective females for a few minutes immediately after each administration. It is considered to be treatment-related, likely as a result of the bad taste of the test substance/vehicle preparation or due to local irritation of the upper digestive tract. It is not considered to be a sign of systemic toxicity. The high-dose of the test substance (300 mg/kg bw/d) caused a significant decrease in food consumption (mainly at the beginning of treatment) and body weight gain as well as a distinct decrease in the corrected (net) body weight gain. These effects are considered to be treatment-related and adverse. No toxicologically relevant effect on food consumption and body weight gain was noted for the animals exposed to 30 or 100 mg/kg bw/d DHDPS.

No differences of toxicological relevance between the control and the treated groups (30, 100 or 300 mg/kg bw/d) were determined for any reproductive parameters, such as conception rate, mean number of corpora lutea, mean number of implantations, as well as pre- and postimplantation loss. Similarly, no influence of the test substance on fetal weight and sex distribution of the fetuses was noted at any dose. Overall, there was no evidence for toxicologically relevant adverse effects of the test substance on fetal morphology at any dose.

Effect on developmental toxicity: via oral route
Dose descriptor:
NOAEL
300 mg/kg bw/day
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

A prenatal developmental toxicity study was performed with 4,4’-sulfonyldiphenol in accordance to the OECD 414 guideline. The test substance was administered to pregnant Wistar rats by daily gavage from implantation to one day prior to the expected day of parturition (GD 6-19) (BASF, 2014).

The test substance caused no mortality nor clinical symptoms of systemic toxicity in any of the exposed groups receiving 30, 100 or 300 mg/kg bw/d 4,4’-sulfonyldiphenol. Signs of maternal toxicity included transient salivation after treatment of the high dose (300 mg/kg bw/d). In addition a significant decrease in food consumption (mainly at the beginning of treatment) and a decrease in body weight gain as well as a distinct decrease in the corrected (net) body weight gain were observed in the high dose group. These effects are considered to be treatment-related and adverse. No toxicologically relevant effect on food consumption and body weight gain was noted for the animals exposed to 30 or 100 mg/kg bw/d DHDPS.

No differences of toxicological relevance between the control and the treated groups (30, 100 or 300 mg/kg bw/d) were determined for any reproductive parameters, such as conception rate, mean number of corpora lutea, mean number of implantations, as well as pre- and postimplantation loss. Similarly, no influence of the test substance on fetal weight and sex distribution of the fetuses was noted at any dose. Overall, there was no evidence for toxicologically relevant adverse effects of the test substance on fetal morphology at any dose.

 

A reproduction/ developmental toxicity screening test was performed with 4,4´-sulphonyldiphenol according to OECD 421. 12 rats per sex per dose were orally administered with the test substance at daily dose levels of 0, 10, 60 and 300 mg/kg bw for 45 days in males and from 14 days before mating to day 3 of lactation in females. The animals were terminally sacrificed on days 45 (males) and on day 4 of lactation (females).

Distinct indications of maternal toxicity were observed in the mid and high dose group (60, 300 mg/kg bw). A suppression in body weight gain in the high dose group is resulting from a decrease in food consumption. Furthermore cecum distention including histopathological lesions in the cecum epithelium occurred in the mid and high dose and a liver enlargement accompanied by centrilobular hypertrophy was identified in the high dose group.

With respect to cecum effects in rats the relevance for humans is unclear. Significant anatomical and functional differences of the cecum exist for rodents and humans. In rodents this anatomical structure is large and in comparison to humans has a significant function in digestion. Nevertheless, these disturbances in the content and structure of the gastrointestinal tract in rats may have a significant impact e.g. on body weight development. Therefore the NOAEL for parental toxicity (repeated dose) was considered to be 60 mg/kg bw/day.

In parent animals in the 300 mg/kg bw/day group, there were no test article-related changes in the copulation index, delivery index, gestation index, number of corpora lutea, length of gestation period, delivery condition, or lactation behavior. However, at a severely systemic toxic dose (300 mg/kg bw/day) there were significant prolongation of estrous cycle, increase in the number of animals that showed prolonged diestrus period, and decrease in the implantation index at 300 mg/kg bw/day. A tendency toward decrease in the fertility index was also observed at 300 mg/kg bw/day.

In offspring in the 300 mg/kg bw/day group, the total number of offspring delivered, number of live offspring and the number of offspring alive on day 4 of lactation tended to be low, but these effects were considered to be due to decreased implantation index. In the 300 mg/kg bw/day group, there were no test article-related changes in the live-birth index, sex ratio, body weight, viability index on day 4 after birth, anogenital distance, general condition, external findings or necropsy findings. In the 10 and 60 mg/kg bw/day groups, there were no test article-related changes in parent animals or offspring. Based on a prolongation of estrous cycle and decreased implantation index the NOAEL for reproductive toxicity was considered to be 60 mg/kg bw/day.

Toxicity to reproduction: other studies

Description of key information

Effects on fertility were observed reproduction/ developmental toxicity screening studies.

Justification for classification or non-classification

For 4,4'-sulphonyldiphenol adverse effects on sexual function were described in Reproduction/Developmental Toxicity Screening Studies, by an increased incidence of irregular estrus cycles. Adverse effects on fertility were described by a decrease in fertility index, a decrease in the number of implantation sites and post-implantation loss.

This evidence was consistently made in two studies with rats of good quality, at equivalent dose levels and with a statistical significance.

Based on the shortage in currently available toxicokinetic or mechanistic information a relevance for humans can neither be excluded, nor reliably be established. 

Effects on fertility occurred together with signs of maternal toxicity in dams (e.g. salivation, GI-tract- and kidney-toxicity). However, and even though e.g. maternal stress is a known non-specific secondary mechanism, a causal relationship between mild maternal toxicity and toxicity to fertility observed in the available rat studies can neither be established nor refuted.

Substances are classified in Category 2 for reproductive toxicity (Suspected human reproductive toxicant), when there is some evidence from humans or experimental animals, of an adverse effect on sexual function and fertility and where the evidence is not sufficiently convincing to place the substance in Category 1 (known or presumed human reproductive toxicant).

As the current data was derived from screening studies and in absence of human information, classification of the substance as reproductive toxicant Category 2 is indicated. Toxicokinetic and mechanistic information to evaluate human relevance are currently being generated.