Phenol, sulfonated

Endpoint summary

Key value for chemical safety assessment

Effects on fertility

Description of key information

OECD 443 (GLP; Sprague Dawley rats; 20, 60, 180 mg/kg bw/day):

- increased kidney weights, enlarged kidneys and histopathological findings in the kidney at 180 mg/mg bw/d

- no adverse substance-related effects on fertility and reproductive performance

- NOAEL (general, systemic toxicity) = 60 mg/kg bw/d; NOAEL (fertility and reproductive performance) = 180 mg/kg bw/day

Effect on fertility: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

180 mg/kg bw/day

Study duration:

subchronic

Species:

rat

Effect on fertility: via inhalation route

Endpoint conclusion:

no study available

Effect on fertility: via dermal route

Endpoint conclusion:

no study available

Additional information

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. The test substance was administered to groups of 24 male and 24 female healthy young Sprague-Dawley rats for test groups 00 - 03 as an aqueous preparation by stomach tube at different dosages (0, 20, 60 and 180 mg/kg body weight/day [mg/kg bw/d]). F0 animals were treated at least for 10 weeks prior to mating to produce a litter (F1 generation). Mating pairs were from the same dose group. Pups of the F1 litter were selected (F1 rearing animals) and assigned to 5 different cohorts which were subjected to specific postweaning examinations. Cohort 1B (= F1 generation parental animals) were selected to produce F2 pups. F1 animals selected for breeding were continued in the same dose group as their parents. Groups of 24 males and 24 females, selected from F1 pups to become F1 parental generation, were offered an aqueous preparation by stomach tube at different dosages (0, 20, 60 and 180 mg/kg bw/d) of the test substance post weaning, and the breeding program was repeated to produce a F2 litter. The study was terminated with the terminal sacrifice of the F2 weanlings and F1 parental animals. Administration period for F0 parental animals was four months in total, F1B parental animals approximately four months, cohort F1A two months, cohort 2A one and a half months, and cohort 3 just over one month in total. Control animals were dosed daily with the vehicle (0.5% Sodium carboxymethyl cellulose [CMC] suspension in drinking water).

The parents' and the pups' state of health was checked each day, and parental animals were examined for their mating and reproductive performances.

Water consumption of the F0 and F1 parents and F1 rearing animals was determined regularly once weekly and weekly during gestation days (GD) 0-1, 3-4, 7-8, 10-11, 14-15, 17-18, 19-20 and lactation days (PND) 1-2, 4-5, 7-8, 10-11, 14-15, 17-18 and 20-21. Food consumption of the F0 and F1 parents and F1 rearing animals was determined regularly once weekly and weekly during GD 0-7, 7-14, 14-20 and PND 1-4, 4-7, 7-10, 10-14, 14-17, 17-19 and 19-21.

In general, body weights of F0 and F1 parents and F1 rearing animals were determined once weekly. However, during gestation and lactation F0/F1 females were weighed on GD 0, 7, 14 and 20 and on PND 1, 4, 7, 10, 14, 17, 19 and 21.

A detailed clinical observation (DCO) was performed in all F0 parents and F1 animals in cohorts 1A, 1B, 2A and 3 at weekly intervals.

Estrous cycle data were evaluated for F0 and cohort 1B (= F1 generation) females over a three weeks period prior to mating until evidence of mating occurred. In all cohort 1A females, vaginal smears were collected after vaginal opening until the first cornified smear (estrous) was recorded. The estrous cycle also was evaluated in cohort 1A females for 2 weeks around PND 75. Moreover, the estrous stage of each female was determined on the day of scheduled sacrifice.

An auditory startle response test was carried out in all animals of cohort 2A on PND 24.

Learning and memory was tested using a Morris water maze in all animals of cohort 2A from PND 60 onwards.

A functional observational battery (FOB) was performed in all animals of cohort 2A on PND 75.

Motor activity was measured in all animals of cohort 2A on PND 75.

The F1 and F2 pups were sexed on the day of birth (PND 0) and were weighed on the first day after birth (PND 1) as well as on PND 4, 7, 14 and 21. Their viability was recorded. At necropsy, all pups were examined macroscopically (including weight determinations of brain, spleen and thymus in one pup/sex/litter).

Date of sexual maturation, i.e. day of vaginal opening (females) or balanopreputial separation (males), of all F1 pups selected to become F1 parental generation was recorded.

All surviving pups were examined for the presence or absence of nipple/areola anlagen on PND 13. If nipple/areola anlagen were recorded, all surviving male pups were carefully reexamined one PND 20 (F1) or PND 21 (F2).

Anogenital distance (defined as the distance from the anus [center of the anal opening] to the base of the genital tubercle) measurements were conducted in a blind randomized fashion, using a measuring ocular on all live F1 and F2 pups on PND 1.

Blood samples for clinical pathological investigations were withdrawn from 10 selected F0 and cohort 1A animals per sex and group.

Further blood samples were taken from 10 surplus (culled) PND 4 pups per sex and group as well as from 10 surplus PND 22 pups per sex and group. Various sperm parameters (motility, sperm head count, morphology) were assessed in the F0 and 1A males at scheduled sacrifice or after appropriate staining.

All F0 and F1 parental animals were assessed by gross pathology (including weight determinations of several organs) and subjected to an extensive histopathological examination; special attention being paid to the organs of the reproductive system. A quantitative assessment of primordial and growing follicles in the ovaries was performed for all control and high-dose F1 parental females. All F1 rearing animals were assessed by different pathological, neuro- and histopathological examinations.

 

Test substance-related adverse effects/findings were noted in the F0 parental animals at 180 mg/kg bw/day regarding kidneys. Increased absolute (+16%) and relative (+20%) weights of the kidneys in males, macroscopically enlarged kidneys in 6/24, increased mineralization in the kidneys in the transition from medulla to cortex ranging from minimal to severe in 21/24 males, nuclear crowding in the kidneys ranging from minimal to moderate in 22/24 males, and tubular dilation in the kidneys ranging from minimal to slight in 13/24 males were observed. F1 pups in the 180 mg/kg bw/day showed a decreased number of liveborn pups subsequent to higher postimplantation loss. Cohort F1A animals in the 180 mg/kg bw/day had increased water consumption in females during major parts of the study period (up to 25% above control) and increased food consumption in females during major parts of the study period (up to 21% above control). In addition, increased body weights in females on study days 14 and 28 (up to 7% above control, respectively) and increased body weight change in females during study days 7 - 14 (about 19% above control, respectively) were observed. Furthermore, prolonged prothrombin time in males and increased total protein, albumin and calcium values in females were reported. In males, increased absolute (+12%) and relative (+17%) weights of the kidneys as well as increased mineralization in the kidneys in the transition from medulla to cortex in 7/20 male animals (minimal to moderate), nuclear crowding in the kidneys in 6/20 male animals (minimal to slight), tubular dilation in the kidneys in 7/20 male animals (minimal to slight), and increased incidence of atrophy in the mammary gland and mammary fat pad were observed.

In F1 parental animals (cohort 1B) at 180 mg/kg bw/day, increased water consumption in females during major parts of the premating period (up to 26% above control) and increased food consumption in females during premating days 28 - 49 and 0 - 70 (up to 22% and 13% above control, respectively) were reported. Additionally, increased body weights in females during major parts of the premating period (up to 10% above control) and increased body weight change in females during premating days 0 - 21 and 0 - 70 (up to 18% and 10% above control, respectively) were observed. In females, increased postimplantation loss (24.6% vs. 6.4% in control) was revealed. Males showed increased absolute (+26%) and relative (+32%) weights of the kidneys and macroscopically enlarged kidneys in 10/24 animals.

In F2 pups, decreased mean pups delivered were subsequent to higher postimplantation loss in F1B maternal animals.

Adverse effects noted in F0 parental animals at 60 mg/kg bw/day were increased water consumption in females during GD 17 - 20 (up to 22% above control), increased food consumption in females during premating days 0 - 7, 28 - 35 and during GD 14 - 20 (up to 9%, 14% and 6% above control, respectively), and increased postimplantation loss (9.4% vs. 3.1% in control).

Cohort F1A animals showed increased water consumption in females during study days 7 - 17 (up to 20% above control), increased body weights in females on study days 14 and 28 (up to 7% above control, respectively), and increased body weight change in females during study days 0 - 7 (about 9% above control, respectively) at 60 mg/kg bw/day.

At 60 mg/kg bw/day F1B parental animals showed increased water consumption in females during premating days 14 - 17 and 42 - 45 (about 14% and 15% above control, respectively) and increased food consumption in females during premating days 7 - 14 (about 13% above control). In addition, increased body weights in females during major parts of the premating period and on PND 14 (up to 9% and 6% above control, respectively) and increased body weight change in females during premating days 0 - 21 (up to 20% above control) were observed. Increased absolute (+13%) and relative (+10%) weights of the kidneys in males were noted.

No treatment-related adverse findings were observed at 20 mg/kg bw/day in any treatment group, either parental or progeny.

Under the conditions of the present modified extended 1-generation reproduction toxicity study the NOAEL (no observed adverse effect level) for general, systemic toxicity is 60 mg/kg bw/d for the F0 and F1 parental as well as adolescent animals, based on evidence for kidney toxicity, as well as corresponding effects on clinical-pathological parameters, which were observed at the LOAEL (Lowest Observed Adverse Effect Level) of 180 mg/kg bw/d. The NOAEL for fertility and reproductive performance for the F0 and F1 parental rats is 180 mg/kg bw/d, the highest dose tested.

Effects on developmental toxicity

Description of key information

- increased kidney weights, enlarged kidneys and histopathological findings in the kidney at 180 mg/mg bw/d

- increased post-implantation loss in F0 and F1B parental animals at 60 and 180 mg/kg bw/d

- NOAEL (general, systemic toxicity) = 60 mg/kg bw/d; NOAEL (developmental toxicity) = 20 mg/kg bw/day; NOAEL (developmental neurotoxicity) = 180 mg/kg bw/d; NOAEL (developmental immunotoxicity) = 180 mg/kg bw/d

Effect on developmental toxicity: via oral route

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEL

20 mg/kg bw/day

Study duration:

subchronic

Species:

rat

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

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.

The test substance was administered to groups of 24 male and 24 female healthy young Sprague-Dawley rats for test groups 00 - 03 as an aqueous preparation by stomach tube at different dosages (0, 20, 60 and 180 mg/kg bw/d). F0 animals were treated at least for 10 weeks prior to mating to produce a litter (F1 generation). Mating pairs were from the same dose group. Pups of the F1 litter were selected (F1 rearing animals) and assigned to 5 different cohorts which were subjected to specific postweaning examinations. Cohort 1B (=F1 generation parental animals) were selected to produce F2 pups. F1 animals selected for breeding were continued in the same dose group as their parents. Groups of 24 males and24 females, selected from F1 pups to become F1 parental generation, were offered an aqueous preparation by stomach tube at different dosages (0, 20, 60 and 180 mg/kg bw/d) of the test substance post weaning, and the breeding program was repeated to produce a F2 litter. The study was terminated with the terminal sacrifice of the F2 weanlings and F1 parental animals. Administration period for F0 parental animals was four months in total, F1B parental animals approximately four months, cohort F1A two months, cohort 2A one and a half months, and cohort 3 just over one month in total.

Control animals were dosed daily with the vehicle (0.5% Sodium carboxymethyl cellulose [CMC] suspension in drinking water).

 

Test substance-related adverse effects/findings were noted in the F0 parental animals at 180 mg/kg bw/day regarding kidneys. Increased absolute (+16%) and relative (+20%) weights of the kidneys in males, macroscopically enlarged kidneys in 6/24, increased mineralization in the kidneys in the transition from medulla to cortex ranging from minimal to severe in 21/24 males, nuclear crowding in the kidneys ranging from minimal to moderate in 22/24 males, and tubular dilation in the kidneys ranging from minimal to slight in 13/24 males were observed. F1 pups in the 180 mg/kg bw/day showed a decreased number of liveborn pups subsequent to higher postimplantation loss.

Cohort F1A animals in the 180 mg/kg bw/day had increased water consumption in females during major parts of the study period (up to 25% above control) and increased food consumption in females during major parts of the study period (up to 21% above control). In addition, increased body weights in females on study days 14 and 28 (up to 7% above control, respectively) and increased body weight change in females during study days 7 - 14 (about 19% above control, respectively) were observed. Furthermore, prolonged prothrombin time in males and increased total protein, albumin and calcium values in females were reported. In males, increased absolute (+12%) and relative (+17%) weights of the kidneys as well as increased mineralization in the kidneys in the transition from medulla to cortex in 7/20 male animals (minimal to moderate), nuclear crowding in the kidneys in 6/20 male animals (minimal to slight), tubular dilation in the kidneys in 7/20 male animals (minimal to slight), and increased incidence of atrophy in the mammary gland and mammary fat pad were observed.

In F1 parental animals (cohort 1B) at 180 mg/kg bw/day, increased water consumption in females during major parts of the premating period (up to 26% above control) and increased food consumption in females during premating days 28 - 49 and 0 - 70 (up to 22% and 13% above control, respectively) were reported. Additionally, increased body weights in females during major parts of the premating period (up to 10% above control) and increased body weight change in females during premating days 0 - 21 and 0 - 70 (up to 18% and 10% above control, respectively) were observed. In females, increased postimplantation loss (24.6% vs. 6.4% in control) was revealed. Males showed increased absolute (+26%) and relative (+32%) weights of the kidneys and macroscopically enlarged kidneys in 10/24 animals.

In F2 pups, decreased mean pups delivered were subsequent to higher postimplantation loss in F1B maternal animals.

Effects noted in F0 parental animals at 60 mg/kg bw/day were increased water consumption in females during GD 17 - 20 (up to 22% above control), increased food consumption in females during premating days 0 - 7, 28 - 35 and during GD 14 - 20 (up to 9%, 14% and 6% above control, respectively), and increased postimplantation loss (9.4% vs. 3.1% in control).

Cohort F1A animals showed increased water consumption in females during study days 7 - 17 (up to 20% above control), increased body weights in females on study days 14 and 28 (up to 7% above control, respectively), and increased body weight change in females during study days 0 - 7 (about 9% above control, respectively) at 60 mg/kg bw/day.

At 60 mg/kg bw/day F1B parental animals showed increased water consumption in females during premating days 14 - 17 and 42 - 45 (about 14% and 15% above control, respectively) and increased food consumption in females during premating days 7 - 14 (about 13% above control). In addition, increased body weights in females during major parts of the premating period and on PND 14 (up to 9% and 6% above control, respectively) and increased body weight change in females during premating days 0 – 21 (up to 20% above control) were observed. Increased absolute (+13%) and relative (+10%) weights of the kidneys in males were noted.

No treatment-related adverse findings were observed at 20 mg/kg bw/day in any treatment group, either parental or progeny.

 

Under the conditions of the present modified extended 1-generation reproduction toxicity study the NOAEL (no observed adverse effect level) for general, systemic toxicity is 60 mg/kg bw/d for the F0 and F1 parental as well as adolescent animals, based on evidence for kidney toxicity, as well as corresponding effects on clinical-pathological parameters, which were observed at the LOAEL (Lowest Observed Adverse Effect Level) of 180 mg/kg bw/d. The NOAEL for developmental toxicity in the F1 and F2 progeny is 20 mg/kg bw/d, based on increased postimplantation loss in P0 and P1 animals, which was observed at the LOAEL (Lowest Observed Adverse Effect Level) of 60 mg/kg bw/d. The NOAEL for developmental neurotoxicity for the F1 progeny is 180 mg/kg bw/d, the highest dose tested. The NOAEL for developmental immunotoxicity for the F1 progeny is 180 mg/kg bw/d, the highest dose tested. Lower mean and median anti-SRBC IgM antibody titers of the positive control group (4.5 mg/kg bw/d cyclophosphamide, oral) demonstrated that the test system worked properly.

Justification for classification or non-classification

Under the conditions of the present modified extended 1-generation reproduction toxicity study, there were no indications from clinical examinations as well as gross and histopathology, that the test substance adversely affected the fertility or reproductive performance of the F0 or F1 parental animals up to and including the highest administered dose of 180 mg/kg bw/d.

The main findings were increases of mean number of resorptions and of postimplantation loss in the high- and mid-dose groups. These findings were noted in both F0 and F1 parental animals.

In addition, estrous cycle length was increased in the high-dose group in the F0 and F1 generations: the increase in the F0 generation was statistically significant with 4.1 days vs. 3.9 days in the control group, in the F1 generation the increase was not significant with 4.1 days vs. 3.9 days in the control group .

No evidence of immunotoxicity or neurotoxicity was noted. 

The NOAEL (no observed adverse effect level) for general, systemic toxicity is 60 mg/kg bw/d for the F0 and F1 parental as well as adolescent animals, based on evidence for kidney toxicity, as well as corresponding effects on clinical-pathological parameters, which were observed at the LOAEL (Lowest Observed Adverse Effect Level) of 180 mg/kg bw/d.

The NOAEL for fertility and reproductive performance for the F0 and F1 parental rats is 180 mg/kg bw/d, the highest dose tested.

The NOAEL for developmental toxicity was established at 20 mg/kg (based on increased postimplantation loss in F1 progeny).

In accordance with the outcome of the RAC discussions (adopted 10 December 2020) and the CLH proposal submitted by the Belgium FPS Public Health, Food Chain Safety and Environment in October 2019 (“CLH report - Proposal for Harmonised Classification and Labelling of 4,4’-sulphonyldiphenol (CAS 80-09-1)”), classification and labelling as Repr.Cat 1B H360FD is warranted.