Registration Dossier

Administrative data

Key value for chemical safety assessment

Effects on fertility

Description of key information

The reproductive/developmental toxicity potential of disodium persulfate was assessed by a read-across approach using results of a reproduction/developmental toxicity tests obtained with diammonium persulfate, a substance of the Persulfate Category, as this substance shows similar toxicological properties.

Diammonium persulfate was tested for oral reproductive/developmental toxicity in a screening test with rats according to OECD guideline 421. No test substance related effects were observed in P and F1 generations. A NOAEL value of 250 mg/kg/day for parental toxicity, reproduction parameters and developmental toxicity was determined. As all substances of the Persulfate Category share the same anionic persulfate moiety and similar toxicological properties a read across approach was applied for dipotassium persulfate and disodium persulfate using results obtained with diammonium persulfate.

An extended one-generation reproductive toxicity study is already initiated and will be performed in 2020. This study is part of an ECHA compliance check decision that is due in September 2020. The study will be included in the dossier as soon as it is available.

Link to relevant study records

Referenceopen allclose all

Endpoint:
extended one-generation reproductive toxicity - basic test design (Cohorts 1A, and 1B without extension)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2020-06-23 to 2021-XX-XX
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 443 (Extended One-Generation Reproductive Toxicity Study)
Version / remarks:
June 25, 2018
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Limit test:
no
Justification for study design:
SPECIFICATION OF STUDY DESIGN FOR EXTENDED ONE-GENERATION REPRODUCTION TOXICITY STUDY WITH JUSTIFICATIONS:

- Premating exposure duration for parental (P0) animals : 10 weeks
- Basis for dose level selection: DRF (on the basis of OECD 421) was performed (rf. to study entry for study number 552-421-5133 and 552-421-5133R)
- Inclusion/exclusion of extension of Cohort 1B: not yet possible
- Termination time for F2 : not yet possible
- Inclusion/exclusion of developmental neurotoxicity Cohorts 2A and 2B : not yet possible
- Inclusion/exclusion of developmental immunotoxicity Cohort 3 : not yet possible
- Route of administration : gavage; A 14 d palatability pre-test has been performed with APS in diet at doses of 250 and 500 mg/kg bw/d (please refer to IUCLID section 7.5.1). The test substance proved to be instable in diet (analytical study not reported in the dossier).
Species:
rat
Strain:
Wistar
Details on species / strain selection:
The rat is regarded as suitable species for reproduction studies and the test guideline is designed to use the rat. The Wistar rat was selected due to a wide range of experience with this strain of rat in reproduction toxicity studies and well-known fertility parameters.
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Toxi-Coop Zrt., Cserkesz u. 90., 1103 Budapest, Hungary
- Females nulliparous and non-pregnant: yes
- Age at study initiation: (P) not older than 9 wks; (F1) not older than 9 wks
- Weight at study initiation: (P) Males: x-x g; Females: x-x g; (F1) Males: x-x g; Females: x-x g
- Housing:
Before mating: 2 animals of the same sex/cage
Mating: 1 male and 1 female/cage
Mated females: individually
Males after mating: 2 animals/cage
F1 offspring (after weaning): 2 or three animals of the same sex/cage
- Diet: ad libitum, ssniff® SM R/M-Z+H "Autoclavable complete feed for rats and mice – breeding and maintenance" (ssniff Spezialdiäten GmbH, D-59494 Soest, Germany)
- Water: ad libitum, tap water
- Acclimation period: 12 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 30 - 70
- Air changes (per hr): > 10
- Photoperiod (hrs dark / hrs light): 12/12
Route of administration:
oral: gavage
Vehicle:
water
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
The test item will be formulated in the vehicle. Formulations will be prepared in the formulation laboratory of Test Facility and used within 3 days.

VEHICLE
- Concentration in vehicle: 10, 20, 40 mg/mL
- Treatment Volume: A constant treatment volume of 5 mL dose preparation/kg body weight was administered in all groups.
Details on mating procedure:
- M/F ratio per cage: 1/1
- Length of cohabitation: max. 14 days
- Proof of pregnancy: vaginal plug or sperm in vaginal smear referred to as day 0 of pregnancy
- After 14 days of unsuccessful pairing replacement of first male by another male with proven fertility if less than 20 pregnant females per group have been achieved.
- Further matings after two unsuccessful attempts: no
- After successful mating each pregnant female was caged (how): individually
- Any other deviations from standard protocol: none
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Analysis of formulations will be performed in the Analytical Laboratory of Test Facility. Five samples will be taken from different places from each concentration (Groups 2, 3 and 4) and at least three replicates are measured on 5 occasions. Similarly, five samples will be taken from the control solution (Group 1) from different places and analyzed.
Duration of treatment / exposure:
Parental male and female animals: 18 weeks
F1 (cohort 1A): 12 weeks
F1 (cohort 1B): 13 weeks
Frequency of treatment:
daily
Details on study schedule:
- Age at mating of the mated animals in the study: 19 weeks
Dose / conc.:
50 mg/kg bw/day
Remarks:
10 mg/mL
Dose / conc.:
100 mg/kg bw/day
Remarks:
20 mg/mL
Dose / conc.:
200 mg/kg bw/day
Remarks:
40 mg/mL (dose was reduced back to 180 mg/kg bw/d due to moribund condition of male and female animals)
No. of animals per sex per dose:
24
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: The dose setting is based on findings obtained from previous studies with Diammonium Peroxodisulphate in rats (Pre-Tests for OECD 443 Study with Diammonium Peroxodisulphate in the Rat, study nos. 552-421-5133 and 552-421-5133R) and in agreement with the Sponsor. The high dose was chosen with the aim of inducing toxic effects but no mortality or severe suffering of animals. The low dose was chosen to induce no toxic effect. In case of severe signs of toxicity, the high dose will be reduced in the course of the study.
- Fasting period before blood sampling for clinical biochemistry: yes, 16 hours
Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily
- Cage side observations checked in table [No.?] were included.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: same as weighing

BODY WEIGHT: Yes
- Time schedule for examinations:
Parental males will be weighed on the first day of dosing (day 0) and weekly thereafter.
Parental females will be weighed on the first day of dosing (Day 0) then weekly, on gestation days 0, 2, 4, 7, 9, 11, 14, 16, 18, 21 and on post-partum days 0 (within 24 hours after parturition), 4, 7, 14 and 21. Body weight of the pregnant female animals will only be evaluated statistically on gestation days 0, 7, 14 and 21. Body weight data will be reported individually for adult animals.

F1 animals selected for follow-up examinations will be weighed on post-natal day 22, then twice a week during the two weeks following weaning, and once weekly thereafter.
For selected F1 offspring, the body weight will be recorded on the day when they attain puberty (completion of balano-preputial separation or vaginal patency).

Fasted body weight will be measured on the day of necropsy for all animals (P and F1).

FOOD CONSUMPTION AND COMPOUND INTAKE: Yes
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes

WATER CONSUMPTION AND COMPOUND INTAKE: Yes
- Time schedule for examinations: daily by visual inspection
Oestrous cyclicity (parental animals):
Estrous cycle will be monitored by examining vaginal smears from each parental female animal daily for two weeks before the mating starts. Vaginal smears will also be prepared and estrous cycle will be monitored daily during the mating period until evidence of copulation. Vaginal smear will also be prepared on the day of the necropsy of parental animals. Vaginal smears will be examined for all F1 Cohort 1A females selected for follow-up examinations after the onset of vaginal patency until the first cornified smear is recorded thus determining the time interval between these events.
Estrous cycle of F1 adult female animals will be examined for a period of two weeks commencing on PND89 and PND96 in Cohort 1A and Cohort 1B, respectively, including necropsy days. Vaginal smears will be stained with 1 % aqueous methylene blue solution. After drying, the smears will be examined with a light microscope.
Sperm parameters (parental animals):
Parameters examined in all control and high dose P/F1 (cohort 1A) male parental generations:
testis weight, epididymis weight, enumeration of cauda epididymal sperm reserve, sperm motility, sperm morphology
Litter observations:
STANDARDISATION OF LITTERS
- Performed on day 4 postpartum: yes
- If yes, maximum of 10 pups/litter (5/sex/litter as nearly as possible); excess pups were killed and discarded.

PARAMETERS EXAMINED
The following parameters were examined in F1 offspring:
number and sex of pups, stillbirths, live births, postnatal mortality, presence of gross anomalies, weight gain, physical or behavioural abnormalities, anogenital distance (AGD), pup weight on the day of AGD, presence of nipples/areolae in male pups, other. Particular attention should be paid to the external reproductive genitals which should be examined for signs of altered development; gross evaluation of external genitalia

GROSS EXAMINATION OF DEAD PUPS:
yes, for external abnormalities; possible cause of death was determined for pups born or found dead, if it can be identified.

ASSESSMENT OF DEVELOPMENTAL NEUROTOXICITY: No

ASSESSMENT OF DEVELOPMENTAL IMMUNOTOXICITY: No
Postmortem examinations (parental animals):
SACRIFICE
- Male animals: All surviving animals [after the optionally extended post-mating period]
- Maternal animals: All surviving animals [on post-partum day 22 or shortly after]

GROSS NECROPSY
- Gross necropsy consisted of external and internal examinations including the cervical, thoracic, and abdominal viscera.

HISTOPATHOLOGY / ORGAN WEIGHTS
The following tissues were prepared for microscopic examination (only in control and high dose animals) and weighed, respectively:
All parental animals and all adult F1 animals of Cohort 1A:
- uterus (with oviducts and cervix)
- ovaries
- testes
- epididymides
- prostate (dorsolateral and ventral parts combined)
- seminal vesicles with coagulating glands as one unit (with their fluids)
- brain,
- liver,
- kidneys,
- spleen,
- thymus,
- pituitary,
- thyroid glands (post-fixation)
- adrenal glands

In animals of Cohort 1B, the weight of following organs will be determined:
- uterus (with oviducts and cervix)
- ovaries
- testes
- epididymides
- prostate (dorsolateral and ventral parts combined)
- seminal vesicles with coagulating glands as one unit (with their fluids)
- brain
- pituitary
Postmortem examinations (offspring):
SACRIFICE
- The F1 offspring not selected as parental animals were sacrificed at 22 days of age, and all F2 offspring on PND 4 or shortly thereafter.
- These animals were subjected to postmortem examinations macroscopic and microscopic examination as follows:

GROSS NECROPSY
- Gross necropsy consisted of external and internal examinations including the cervical, thoracic, and abdominal viscera.

HISTOPATHOLOGY / ORGAN WEIGTHS
The tissues indicated in Table [#] were prepared for microscopic examination and weighed, respectively.

IMMUNOTOXIC EXAMINATION

At termination, weighing of the lymph nodes and splenic lymphocyte subpopulation analysis will be performed in 10 male and 10 female Cohort 1A animals from each group (1 male or 1 female per litter; all litters represented by at least 1 pup; randomly selected) as follows:
- weighing of the lymph nodes associated with and distant from the route of exposure (submandibular and popliteal lymph nodes);
- splenic lymphocyte subpopulation analysis (CD4+ and CD8+ Tlymphocytes, B lymphocytes and natural killer cells) by flow cytometry method using one half of the spleen
Statistics:
The statistical evaluation of appropriate data will be performed with the statistical program package SPSS PC+4.0.
Reproductive indices:
Copulatory Index (Measure of animals ability to mate):
Males: Number of males with confirmed mating / Total number of males cohabited x 100
Females: Number of sperm positive females / Total number of females cohabited x 100

Fertility Index (Measure of male’s ability to produce sperm that can fertilize eggs and measure of female’s ability to become pregnant):
Males: Number of males impregnating a females / Total number of males with confirmed mating x 100
Females: Number of pregnant females / Number of sperm positive females x 100

Gestation Index (Measure of pregnancy that provides at least one live pup):
Number of females with live born pups / Number of pregnant females x 100
Offspring viability indices:
Post-implantation mortality: Number of implantations – Number of liveborns / Number of implantation x 100

Post-natal mortality: Number of liveborns – Number of live pups on PND 13 / Number of liveborns x 100

Survival Index: Number of live pups on PND 13 / Number of liveborns x 100

Sex ratio: Number of pups examined – Number of pups males (females) / Number of pups examined x 100
Key result
Dose descriptor:
NOAEL
Remarks on result:
other: Will be completed when study has been finalised.
Key result
Dose descriptor:
NOAEL
Remarks on result:
other: Will be completed when study has been finalised.
Key result
Reproductive effects observed:
not specified
Endpoint:
screening for reproductive / developmental toxicity
Remarks:
DRF for OECD 443
Type of information:
experimental study
Adequacy of study:
supporting study
Study period:
2019-11-20 to 2020-03-16
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
comparable to guideline study
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 421 (Reproduction / Developmental Toxicity Screening Test)
Version / remarks:
29th July 2016
Deviations:
yes
Remarks:
DRF
GLP compliance:
no
Remarks:
This study was not performed according to GLP compliances, however, the principles of GLP were followed and all data were recorded and retained.
Limit test:
no
Species:
rat
Strain:
Wistar
Details on species / strain selection:
The rat is regarded as suitable species for toxicity and reproduction toxicity studies and the test guidelines were designed to use the rat. The Wistar rat was selected due to a wide range of experience with this strain of rat in toxicity and reproduction toxicity studies and wellknown fertility parameters.
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Toxi-Coop Zrt., Cserkesz u. 90., 1103 Budapest, Hungary
- Females nulliparous and non-pregnant: yes
- Age at study initiation: 13-14 weeks
- Weight at study initiation: Males: 354-422 g; Females: 204-251 g
- Housing:
Before mating: 2 animals of the same sex/cage
Mating hours: 1 male and 1 female/cage
Pregnant females were housed individually
Males after mating: 2 animals/cage

- Diet: ad libitum, ssniff® SM R/M-Z+H "Autoclavable complete feed for rats and mice – breeding and maintenance" (ssniff Spezialdiäten GmbH, D-59494 Soest, Germany)
- Water: ad libitum, tap water
- Acclimation period: 27 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 30 – 70
- Air changes (per hr): above 10
- Photoperiod (hrs dark / hrs light): 12/12
Route of administration:
oral: gavage
Vehicle:
water
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: The test item was formulated in vehicle in concentrations of 10 mg/mL, 20 mg/mL and 36/50 mg/mL in the formulation laboratory of Test Facility not longer than for three days before the use. The concentration of the high dose was reduced from 50 mg/mL to 36 mg/mL according to the dose reduction on Day 6. High dose formulation was not administered on Days 4 and 5 (washout days).
Details on mating procedure:
- M/F ratio per cage: 1/1
- Length of cohabitation: until copulation occurred.
- Proof of pregnancy: vaginal plug / sperm in vaginal smear referred to as day 0 of pregnancy
- After successful mating each pregnant female was caged (how): individually
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Analysis of formulations was performed in the Analytical Laboratory of Test Facility twice during the treatment period. Five samples were taken (from different places) from each concentration (Groups 2, 3 and 4) and measured. Similarly, five samples were taken from the control solution (Group 1).
Date of sampling: December 03, 2019 and January 08, 2020;
Date of analysis: December 04, 05, 2019 and January 09, 10, 2020;
The concentration of dosing formulations was between the range of 98 and 108 % of the nominal concentrations.
Duration of treatment / exposure:
Males: 48 days (14 days pre-mating period, mating period and post-mating period)
Females: 58-65 days (depending on day of mating)
Frequency of treatment:
daily
Dose / conc.:
50 mg/kg bw/day
Remarks:
10 mg/mL
Dose / conc.:
100 mg/kg bw/day
Remarks:
20 mg/mL
Dose / conc.:
250 mg/kg bw/day
Remarks:
50 mg/mL (Dose was reduced to 180 mg/kg bw/d after deaths occurred in male animals)
No. of animals per sex per dose:
12
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: The dose setting with 50, 100 and 250 mg/kg bw/day was based on results obtained in previous studies performed with Diammonium Peroxodisulphate in the rat.
Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily (morbidity and mortality were checked twice daily)
- Cage side observations included check of skin, fur, eyes and mucous membranes, autonomic activity (lachrymation, piloerection, pupil size, respiratory pattern, occurrence of secretions and excretions), circulatory and central nervous system, somatomotor activity and behavior pattern, changes in gait, posture and response to handling. Special attention was directed towards the observation of tremors, convulsions, salivation, diarrhea, lethargy, sleep and coma.

BODY WEIGHT: Yes
- Time schedule for examinations:
Parental males were weighed on the first day of dosing (Day 0) then weekly thereafter and on the day of necropsy (fasted body weight).
Parental females were weighed on the first day of dosing (Day 0) then weekly, on gestation days 0, 7, 14 and 21 and on post-partum days 0 (within 24 hours after parturition), 4, 7, 14 and 21.

FOOD CONSUMPTION:
- Time schedule: weekly except mating phase: premating Days 7 and 13 and by weekly interval during post-mating period for male animals; premating Days 0, 7 and 13, gestation days 0, 7, 14 and 21, lactation days 0, 7, 14 and 21 for female animals.

OTHER:
DELIVERY PROCESS
- Time schedule: from gestational day 21 onwards
- Observations included: nest preparation from the bedding material and nursing of the new-borns or not. The sucking success was monitored by the presence of milk in the pups' stomach. All observations were recorded individually for each dam. The duration of gestation was recorded and was calculated from day 0 of pregnancy.
Oestrous cyclicity (parental animals):
Estrous cycle was monitored by examining vaginal smears before the treatment started from each animal being considered for study for two weeks. Vaginal smears were also prepared and estrous cycle was monitored daily from the beginning of the treatment period (two weeks pre-mating period) and during the mating period until evidence of copulation. Vaginal smear was also prepared on the day of the necropsy.
Sperm parameters (parental animals):
Not examined
Litter observations:
STANDARDISATION OF LITTERS
- Performed on day 4 postpartum: yes
- If yes, maximum of 8 pups/litter (4sex/litter as nearly as possible); excess pups were killed and discarded.

PARAMETERS EXAMINED
The following parameters were examined in F1 offspring:
number and sex of pups, stillbirths, live births, postnatal mortality, presence of gross anomalies, weight gain, physical or behavioural abnormalities, anogenital distance (AGD), pup weight on the day of AGD, presence of nipples/areolae in male pups. Particular attention was paid to the external reproductive genitals.

GROSS EXAMINATION OF DEAD PUPS:
yes, for external and internal abnormalities; possible cause of death was determined for pups born or found dead

Postmortem examinations (parental animals):
SACRIFICE
- Male animals: All surviving animals on Day 48
- Maternal animals: All surviving animals on post-partum days 22-24 (Days 58 - 65)

GROSS NECROPSY
- Gross necropsy consisted of external and internal examinations including the cervical, thoracic, and abdominal viscera.

HISTOPATHOLOGY / ORGAN WEIGHTS
Detailed histological examinations were performed on all preserved organs and tissues of dead animals and on ovaries, uterus, vagina, testes, epididymides, prostate and seminal vesicles with coagulating gland in the control and high dose groups with special emphasis on stages of spermatogenesis in the male gonads and histopathology of interstitial testicular cell structure; on the ovaries covering the follicular, luteal, and interstitial compartments of the ovary as well as the epithelial capsule and ovarian stroma.
In addition, histological examinations were performed in organs of animals showing macroscopic findings at necropsy as follows:
- kidneys: 1/12 male and 3/12 dams in control group; 1/12 dams at 50 mg/kg bw/day; 2/12 male and 2/12 dams at 100 mg/kg bw/day; 3/10 dams at 180/250 mg/kg bw/day;
- skin: 1/12 female in control group
Organ weights: Brain weight and weight of the testes and epididymides as well as prostate and seminal vesicles with coagulating glands as a whole of adult male animals were determined.


Postmortem examinations (offspring):
SACRIFICE
- The F1 offspring were sacrificed at 22 days of age.
- These animals were subjected to postmortem examinations (macroscopic examination).

GROSS NECROPSY
- Gross necropsy consisted of external examinations. Particular attention was paid to the external reproductive genitals.
Statistics:
The statistical evaluation of appropriate data were performed with the statistical program package SPSS PC+4.0. The homogeneity of variance between groups was checked by Bartlett’s homogeneity of variance test. Where no significant heterogeneity was detected a one-way analysis of variance (ANOVA) was carried out. If the obtained result was significant Duncan Multiple Range test was used to access the significance of inter-group differences. Getting significant result at Bartlett’s test the Kruskal-Wallis analysis of variance was used and the inter-group comparisons were performed using Mann-Whitney U-test. Chi2 test was performed if feasible. Frequency of toxic response, pathological and histopathological findings by sex and dose were calculated. Results were evaluated in comparison with values of control group (i.e. control value).
Reproductive indices:
Copulatory Index, Fertility Index and Gestation Index
Offspring viability indices:
Survival Index
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Both male and female animals were normal in the control, 50 and 100 mg/kg bw/day during the entire observation period (pre-mating, mating, post-mating, gestation and lactation).
Clinical signs related to the test item were observed in male and female animals at 250 mg/kg bw/day. Signs included piloerection (8/8 male, 12/12 female), decreased activity (8/8 male, 12/12 female), decreased body tone (1/8 male, 1/12 female), diarrhea (1/8 male) and hunched back (1/8 male) between Days 1 and 5. After the dose reduction to 180 mg/kg bw/day, these animals became normal.
Mortality:
mortality observed, treatment-related
Description (incidence):
250 mg/kg bw/day dose of Diammonium Peroxodisulphate caused death of four male animals (4/12) after four-day administration on Days 4 or 5. Macroscopic and histopathological examinations revealed changes in the gastro-intestinal tract, referring to disturbances in electrolyte and water household and circulatory system along with signs of acute stress reaction in the adrenal gland and thymus due to agony.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
The body weight decreased in male animals and the body weight development was depressed in female animals at 180/250 mg/kg bw/day during the first week of the treatment (Days 0-7). After the dose reduction, the body weight gain was normalized in male and female animals but the mean body weight of male animals remained slightly below the control value up to the end of the treatment period.
Slightly reduced body weight gain at 100 mg/kg bw/day did not result in significant changes in the mean body weight during the treatment period in male animals and during the pre-mating period in female animals. In the male animals, the mean body weight and mean body weight gain were comparable in the control and 50 mg/kg bw/day during the entire observation period. In the female animals, the body weight development was comparable in the control and 50 mg/kg bw/day group during the pre-mating and gestation periods.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
The mean daily food consumption was lowered in male and female animals at 180/250 mg/kg bw/day during the first week of the treatment (between Days 0 and 7).
Organ weight findings including organ / body weight ratios:
no effects observed
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Histological examination revealed lesions in the gastro-intestinal tract in dead male animals at 250 mg/kg bw/day – acute inflammation in the mucous membrane of stomach, and acute catarrh in the small and large intestines. Additionally, congestion in the liver and lungs, increased vacuolation of cortical cell in the adrenal glands and lymphocyte depletion in the spleen and thymus were also detected.
Diammonium Peroxodisulphate did not cause any toxic or other test item related lesions detectable by histological examination of the selected reproductive organs of experimental male and female animals at 180/250 mg/kg bw/day.
Other effects:
effects observed, treatment-related
Description (incidence and severity):
Serum thyroid hormones
The FT4 and TSH levels were not adversely affected in the parental male animals or in PN22 offspring at any dose levels.

Delivery data of dams
The percentage and mean number of the pre-natal (post-implantation) loss was higher in female animals at 180/250 mg/kg bw/day than in the control group and the historical control value. Although, there were no statistically significant differences between the control and 180/250 mg/kg bw/day administered group, a test item influence cannot be excluded and verified with the results of this study.
Reproductive function: oestrous cycle:
no effects observed
Description (incidence and severity):
The examined parameters of the estrous cycle were not adversely by the test item (50, 100 or 180/250 mg/kg bw/day). Higher mean numbers of days of diestrous at 180/250 mg/kg bw/day might be indicative of test item influence but might also refer to the biological variation.
Reproductive performance:
no effects observed
Description (incidence and severity):
The examined parameters of reproductive performance were not adversely affected by the treatment with the test item in male and female animals at 50, 100 or 180/250 mg/kg bw/day.
Key result
Dose descriptor:
NOAEL
Remarks on result:
not determinable because of methodological limitations
Remarks:
DRF
Key result
Critical effects observed:
no
Clinical signs:
no effects observed
Mortality / viability:
no mortality observed
Body weight and weight changes:
no effects observed
Anogenital distance (AGD):
no effects observed
Nipple retention in male pups:
no effects observed
Key result
Dose descriptor:
NOAEL
Remarks on result:
not determinable because of methodological limitations
Remarks:
DRF
Key result
Critical effects observed:
no
Key result
Reproductive effects observed:
no
Conclusions:
Based on the above findings the suggested dose levels for the main study (OECD 443) are as follows:
Group 1: 0 mg/kg bw/day
Group 2: 50 mg/kg bw/day
Group 3: 100 mg/kg bw/day
Group 4: 200 mg/kg bw/day (Please refer to Study No. 552-421-5133R for additional information)
Executive summary:

The subject of this study was the Pre-Test for OECD 443 Study with Diammonium Peroxodisulphate (APS) in the rat on the basis of OECD 421. The guideline is designed for use with the rat, which is the preferred rodent species for reproduction toxicity testing. The purpose of this study was to determine the dose levels for the main study (OECD 443) and obtain information on the possible effects of the test item on reproduction and development when repeatedly administered orally (by gavage) to rats at doses of 50, 100 and 180/2501 mg/kg bw/day compared to control animals receiving vehicle only. Four groups of Han:WIST rats (n=12/sex/group) were administered with the test item orally (by gavage) once a day at 0 (vehicle), 50, 100 and 180/250 mg/kg bw/day doses corresponding to concentrations of 0, 10, 20 and 36/50 mg/mL. The application volume was 5 mL/kg bw. Control animals received the vehicle, distilled water. The suitability of the vehicle at the intended concentrations of the test item was analytically verified up front (concentration and homogeneity). The concentration of the test item in the dosing formulations used for animal’s treatment was checked two times during the study. Concentration of APS in the dosing formulations varied between the range of 98 % and 108 % of the nominal values and formulations were homogenous thereby confirming the proper dosing. All animals of the parent (P) generation were dosed prior to mating (14 days) and throughout mating. In addition, males received the test item or vehicle after mating up to the day before the necropsy (altogether for 48 days, except for 4 dead animals). Dams were additionally exposed through the gestation period and up to lactation days 21-23 (altogether for 58 - 65 days). Non-pregnant female animals and one dam, for which no living pups remained, were administered for 48 days. Observations included mortality, clinical signs, body weight, food consumption, mating, pregnancy and delivery process, as well as development of offspring. Estrous cycle was monitored by examining vaginal smears for two weeks before the treatment (pre-experimental period) and for two weeks from the beginning of the treatment period (two weeks pre-mating period) and during the mating period until evidence of mating and on the day of the necropsy. The dams were allowed to litter and rear their offspring up to day 21 postpartum or shortly thereafter. Litters were weighed and offspring were observed for possible abnormalities and were euthanized on post-natal day 22. Blood samples were collected for possible determination of serum levels of thyroid hormones (FT4, TSH) from 2-6 pups per litter (except for litters with 9 or less pups) on post-natal day 4, from all dams and from 2 pups per litter on post-partum/post-natal day 22 and from all parent male animals at termination. All parental animals were subjected to gross pathology one day after the last treatment. The body weight, brain weight, weight of the testes, epididymides and prostate and seminal vesicles with coagulating glands as a whole of all adult male animals were determined.

Histopathology examination was performed on reproductive organs (testes, epididymides, prostate, and seminal vesicles with coagulating glands, uterus with oviduct, cervix, vagina and ovaries) in the control and high dose groups. Full histopathology examinations were performed on the organs and tissues of dead male animals. In addition, histopathology examinations were performed on organs showing macroscopic changes (kidneys, skin) at necropsy.

 

Results

Mortality

250 mg/kg bw/day dose of APS caused death of four male animals (4/12) after four-day administration on Days 4 or 5. Piloerection, decreased activity, decreased body tone, diarrhea and narrow eye aperture were noted for these animals during the preceding days (from Day 1 to Days 3 or 4). Changes in the gastro-intestinal tract referred to disturbances in electrolyte and water household and circulatory system along with signs of acute stress reaction in the adrenal gland and thymus due to agony.

 

Clinical observation

Test item related clinical signs were observed in male and female animals at 250 mg/kg bw/day: piloerection, decreased activity, decreased body tone, diarrhea and narrow eye aperture were observed in dead animals from Day 1 up to and including Days 3 or 4 and animals were found dead on Days 4 or 5. After the dose reduction to 180 mg/kg bw/day, these animals became normal.

 

Body weight and body weight gain

The body weight development was depressed in male and female animals at 180/250 mg/kg bw/day during the first week of the treatment (Days 0-7). After the dose reduction, the body weight gain was normalized in male and female animals but the mean body weight of male animals remained slightly below the control value up to the end of the treatment period. Slight reduction in body weight gain at 100 mg/kg bw/day did not result in significant changes in the mean body weight during the treatment period in male animals and during the pre-mating period in female animals.

 

Food consumption

The mean daily food consumption was lowered in male and female animals at 180/250 mg/kg bw/day during the first week of the treatment (between Days 0 and 7).

 

Estrous cycle

The examined parameters of the estrous cycle were not adversely by the test item (50, 100 or 180/250mg/kg bw/day). Higher mean numbers of days of diestrous at 180/250 mg/kg bw/day might be indicative of test item influence but might also refer to the biological variation.

 

Delivery data of dams

The percentage and mean number of the pre-natal (post-implantation) loss was higher in female animals at 180/250 mg/kg bw/day than in the control group and the historical control value. Although, there were no statistically significant differences between the control and 180/250 mg/kg bw/day administered group, a test item influence cannot be excluded and verified with the results of this study.

 

Reproductive performance

The examined parameters of reproductive performance were not adversely affected by the treatment with the test item in male and female animals at 50, 100 or 180/250 mg/kg bw/day. Test item influence on the slightly lower fertility index in the female animals at 180/250 mg/kg bw/day cannot be excluded entirely, although similar findings are also seen in untreated rats of this strain.

 

Serum thyroid hormones

The FT4 and TSH levels were not adversely affected in the parental male animals or in PN22 offspring at any dose levels.

 

Necropsy

Macroscopic alterations related to the effect of the test item were detected in the gastro-intestinal tract (hemorrhage, dilatation, liquid content) and on the hairs around the anus (sign of diarrhea) in dead male animals administered with 250 mg/kg bw/day at the necropsy examinations. Gross necropsy did not reveal any macroscopic changes which were related to treatment with the test item in surviving animals at 50, 100 or 180/250 mg/kg bw/day.

 

Organ weight

The weights of examined organs (absolute, relative to body and brain weights) were not adversely affected by the test item at 50, 100 or 180/250 mg/kg bw/day.

 

Histopathology

Histological examination revealed lesions in the gastro-intestinal tract in dead male animals at 250 mg/kg bw/day – acute inflammation in the mucous membrane of stomach, and acute catarrh in the small and large intestines. Additionally, congestion in the liver and lungs, increased vacuolation of cortical cell in the adrenal glands and lymphocyte depletion in the spleen and thymus were also detected. APS did not cause any toxic or other test item related lesions detectable by histological examination of the selected reproductive organs of experimental male and female animals at 180/250 mg/kg bw/day.

 

Offspring

The offspring’s development was not adversely influenced at any dose level. No effects on the mortality, clinical signs, body weight development, anogenital distance (male and female) or nipple retention (male) were detected.

 

Conclusion

Under the conditions of the present study, APS caused death (male), clinical signs (male and female), depressed body weight development and food consumption (male and female), macroscopic and histopathology changes in stomach and intestines (male) in Han:WIST rats at 250 mg/kg bw/day by oral gavage as investigated in this study. After the dose reduction to 180 mg/kg bw/day, clinical signs ceased after some days and there were no adverse effects in the examined parameters. Nonetheless the body weight of the males had not recovered by the end of the study. There were no test item related adverse effects at 50 or 100 mg/kg bw/day.

APS did not adversely influence the reproductive performance (gonad function, mating behavior, conception, parturition) in parental male and female animals at 50, 100 or 180/250 mg/kg bw/day.

The development of the F1 offspring was not impaired from birth to post-natal day 21 after repeated oral administration of dams at 50, 100 or 180/250 mg/kg bw/day.

The suggested dose levels for the main study (OECD 443) are as follows:

Group 1: 0 mg/kg bw/day

Group 2: 50 mg/kg bw/day

Group 3: 100 mg/kg bw/day

Group 4: 200 mg/kg bw/day (Please refer to Study No. 552-421-5133R for additional information)

Endpoint:
screening for reproductive / developmental toxicity
Remarks:
DRF for OECD 443
Type of information:
experimental study
Adequacy of study:
supporting study
Study period:
2020-04-21 to 2020-05-05
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
comparable to guideline study
Qualifier:
no guideline followed
Principles of method if other than guideline:
In the DRF for OECD 443 effects at 180 mg/kg bw/d were considered as not severe enough to define it as a high dose in the main test. Thus, an additional 14 day DRF with 200 mg/kg bw/d was performed.
GLP compliance:
no
Remarks:
This study was not performed according to GLP compliances, however, the principles of GLP were followed and all data were recorded and retained.
Limit test:
yes
Species:
rat
Strain:
Wistar
Route of administration:
oral: gavage
Vehicle:
water
Duration of treatment / exposure:
14 days
Frequency of treatment:
daily
No. of animals per sex per dose:
6
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: Effects at 180 mg/kg bw/d in the DRF for the OECD 443 were not severe enough. Thus 200 mg/kg bw/d was tested additionally.
Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Detailed observation, once daily; Mortality/ morbidity: twice a day throughout the study

BODY WEIGHT: Yes
- Time schedule for examinations: Just before the first administration and on Days 0, 3, 7, 10 and 13.

FOOD CONSUMPTION: Yes
- Time schedule for examinations: on Days 0, 7, and 13
Postmortem examinations (parental animals):
SACRIFICE
- all animals: on study day 14

GROSS NECROPSY
- Gross necropsy consisted of external and internal examinations including gastro-intestinal tract, eyes and mucous membranes, that were observed with particular attention.
Statistics:
Statistical analysis was done with SPSS PC+ software package for the body weight, body weight gain and food consumption data.
Frequency of clinical signs and pathological findings by sex and dose was calculated.
Clinical signs:
no effects observed
Description (incidence and severity):
The general physical condition and behavior of animals were considered to be normal throughout the entire observation period. No clinical signs were detected in the animals (male and female) in the control or 200 mg/kg bw/day groups at the daily observations.
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
Description (incidence and severity):
The body weight development was not adversely affected by the treatment with 200 mg/kg bw/day throughout the 14-day treatment period.
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
The mean daily food consumption was not adversely affected at 200 mg/kg bw/day (male and female) during the 14-Day observation period.
Key result
Dose descriptor:
NOAEL
Remarks on result:
not determinable because of methodological limitations
Remarks:
DRF
Key result
Critical effects observed:
no
Key result
Dose descriptor:
NOAEL
Remarks on result:
not determinable because of methodological limitations
Remarks:
F1 not included in this DRF
Critical effects observed:
not specified
Reproductive effects observed:
not specified
Conclusions:
Diammonium Peroxodisulphate caused no signs of toxicity (clinical signs, body weight and body weight gain, food consumption, necropsy findings) in male or female Han:WIST rats after the consecutive 14-day oral (by gavage) administration of 200 mg/kg bw/day.
Executive summary:

The purpose of this study was to obtain information on the effect of the test item when administered by gavage to male and female Han: WIST rats at 200 mg/kg bw/day dose for 14 days compared to control animals receiving vehicle (distilled water) only and to determine the highest dose level for the main study (OECD 443).

The dose setting of 200 mg/kg bw/day was based on the results of previously performed study with the test item “Pre-Test for OECD 443 Study with Diammonium Peroxodisulphate in the Rat” (Study no. 552-421-5133).

The test item was administered by gavage to Han:WIST rats (n=6 animals/ sex/ group) once a day at 0 (vehicle control) and 200 mg/kg bw/day corresponding to concentrations of 0 and 40 mg/mL for 14 days. The application volume was 5 mL/kg bw.

Detailed clinical observations were made on all animals once a day, after treatment at approximately the same time. Body weight was determined twice a week and food consumption was measured weekly. Gross pathology examinations were conducted one day after the last administration (Day 14).

The results of this study were summarised as follows:

No animal died during the study.

The control (6/6 male and 6/6 female) and test item treated animals (5/6, male and 6/6 female) showed no clinical signs during the treatment period. Diarrhea was noted for one male animal (1/6) transiently on Day 0. The behavior and the general state of the test animals were considered to be normal during the study.

The mean body weight, body weight gain and the mean daily food consumption were comparable to their control in male and female animals at 200 mg/kg bw/day during the 14-day observation period.

Specific macroscopic alterations related to treatment with the test item were not observed during the terminal necropsy of treated animals.

Conclusion

Diammonium Peroxodisulphate caused no signs of toxicity (clinical signs, body weight and body weight gain, food consumption, necropsy observations) in male or female Han:WIST rats after the consecutive 14-day oral (by gavage) administration of 200 mg/kg bw/day. Thus 200 mg/kg bw/day was used as highest dose in the main study of OECD 443.

Endpoint:
one-generation reproductive toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2004-01-30 to 2004-12-13
Reliability:
1 (reliable without restriction)
Qualifier:
according to guideline
Guideline:
OECD Guideline 421 (Reproduction / Developmental Toxicity Screening Test)
Deviations:
no
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
other: Crl:CD (SD)IGS BR
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories; Raleigh; North Caroline (USA)
- Age at study initiation: (P) 11 wks
- Weight at study initiation: (P) Males: 359 - 436 g; Females: 212 - 280 g
- Fasting period before study: no
- Housing: Males and females were housed individually (except during mating)
- Diet: Powdered PMI Certified Rodent Diet #5002 (PMI Feeds, Inc.; St Louis, Missouri) ad libitum
- Water: ad libitum
- Acclimation period: at least one week

ENVIRONMENTAL CONDITIONS
- Temperature: 18 - 26 °C
- Humidity: 30 - 70 % R.H.
- Air changes: a minimum of 10 air changes per hour
- Photoperiod: 12 hrs dark/12 hrs light
Route of administration:
oral: feed
Vehicle:
unchanged (no vehicle)
Details on exposure:
Study design:
Male and female Crl:CD (SD)IGS BR rats were assigned to 4 groups (12/sex/group). Each group received a control diet consisting of powdered PMI Certified Rodent diet #5002 (PMI Feeds Inc., St. Louis, Missouri) or the same diet containing diammonium persulfate at concentrations designed to provide target doses of 40, 100 250 mg/kg/day based on mean weekly body weight and mean weekly food consumption data for each sex at each dose level.
Dose administration:
Males were administered with test material in the diet for three weeks prior to mating, during the mating period (four days) and approximately five weeks after mating. Females were dosed throughout the 3-week pre-mating period, during the 4-day mating period, throughout gestation (22 days) and during the first four days of lactation. Depending on what day the female was confirmed pregnant, the total days of dose administration varied from 47 to 51 days. The animals were dosed until the day of sacrifice (prior to removal from the room).
Details on mating procedure:
Breeding:
Animals from respective groups were mated by placing one female in the breeding cage of a male from the same dose group. A record of mating pairs was maintained. Once mating occurred, the males and females were separated. The maximum mating period was 14 days. Females for which no evidence of mating was detected after a total of 14 days were placed in nesting boxes.
Confirmation of mating:
During mating, a daily inspection was be made for the presence of a retained copulatory plug, multiple copulatory plugs on the cage tray, or vaginal sperm. The day sperm or a plug was observed was designed to be day 0 of gestation.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Concentration of diammonium persulfate in rodent feed samples were determined using the measurement of the exothermic enthalpy delta H (J/g) by DSC of the test samples and comparison with a standard calibration curve. The method was evaluated and validated and found to be suitable for the analytical determination of diammonium persulfate in rodent diet.

Experimental procedures:
Sample preparation:
About 1 g to 10 g of sample was taken with a spatula from the container and ground for 3 minutes in a cleaned mortar and pestle. About 10 - 30 mg of the ground sample was transferred into a tared round DSC sample pan. By using a small steel cylinder, which was the same size as the pan, the powdery sample in the pan was compressed. The sample was weighed to ± 0.01 mg with an Autobalance AD 6. The weight of the sample was recorded on a sample weighing sheet. The sample pan containing the sample was then transferred into the sample holder of the DSC system for analysis immediately.
Instrumentation and Analysis:
DSC System:
Perkin Elmer Pyris 1 Differential Scanning Calorimeter; Serial Number: S73N0061703;
Balance:
PE AD 6 Autobalance; Balance Serial #: 655C04;
Sample Analysis:
DSC Mode used: Ice Bath;
Sample atmosphere: Nitrogen, 20 mL/min;
Sample pan: Aluminium, open pan;
Temperature Program: Hold at 30 °C for 1 minute; Heat from 30 °C to 220 °C with a rate of 10 °C/min;
Calculation:
Exothermic enthalpy delta H (J/g) of each sample in temperature range between 160 °C to 210 °C was calculated with Pyris Software provided by the DSC system.
A five point standard curve of ammonium persulfate concentration (ppm) versus delta H (J/g) was developed in a separate method development/validation procedure. The following equation, which was obtained with the five point standard curve, was used to calculate the ammonium persulfate concentration of each sample:
Concentration (ppm) = (delta H + 0.0052671)/0.0007350
where:
0.0052671 = Intercept of the standard curve
0.0007350 = Slope of the standard curve
Duration of treatment / exposure:
Not applicable
Frequency of treatment:
Not applicable
Details on study schedule:
The purpose of the study was to provide information regarding possible effects on mating, reproduction, and/or development in male and female rats. Males were administered with test substance in the diet three weeks prior to mating, during the mating period (four days) and approximately five weeks after mating.
Females were dosed throughout the 3-week pre-mating period, during the 4-day mating period, throughout gestation (22 days) and during the first four days of lactation. The total length of exposure was approximately 51 days for females (depending on when the female was confirmed pregnant) and 56 days for males.
Dose / conc.:
0 mg/kg bw/day (nominal)
Remarks:
in diet
Dose / conc.:
40 mg/kg bw/day (nominal)
Remarks:
in diet
Dose / conc.:
100 mg/kg bw/day (nominal)
Remarks:
in diet
Dose / conc.:
250 mg/kg bw/day (nominal)
Remarks:
in diet
No. of animals per sex per dose:
12 animals per sex and per dose group
Control animals:
yes, plain diet
Details on study design:
Dietary formulations:
Diets were mixed at least weekly according to the mixing procedure approved by the sponsor. For each high-dose formulation prepared (whether for one or both sexes, depending on the phase of the study), the total amount of test substance needed for that high-dose stock formulation was weighed and the finely ground with a mortar and pestle. The finely ground test article was gradually mixed into approximately 200 g of the PMI powdered diet. This preliminary premixture of test article and powdered diet was then placed in a Hobart mixer, and the remaining powdered diet needed to achieve the high-dose stock formulation was gradually added to the test article and feed mixture. After completion of the additional powdered diet, the high dose stock formulation was mixed for at least 30 minutes. Once this mix was completed, the low and mid levels were prepared by taking the appropriate amount of the high-dose stock formulation and diluting it to the appropriate dietary concentration needed for each dose level by gradually adding powdered feed to achieve the correct precalculated weight. Calculated diet concentrations were based on historical body weight and food consumption data for Days 0 to 7 and based on current food consumption and body weight data for the appropriate sex after Day 7. All diet formulations were stored at room temperature until dosed. Dose concentrations were based on test article as supplied (i.e., no correction factor was used when calculating the amount of test article to be use).
Positive control:
Not indicated
Parental animals: Observations and examinations:
Assessment of toxicity was based on mortality, clinical observations, body weights and weight change (during premating and mating for both sexes, during gestation and lactation for females, and postmating for males), food consumption (during premating for both sexes, during gestation and lactation for females, and postmating for males), pregnancy rates and maternal data, litter data (litter size, sex and weight of pups, live and dead pups), anatomic pathology, organ weights, histopathology of selected tissues, and staging of spermatogenesis.
Oestrous cyclicity (parental animals):
Not indicated
Sperm parameters (parental animals):
The following parameters were examined:
- Normal progression of spermatogenesis at necropsy.
- Stages of the spermatogenic cycle.
- Testicles examination.
Litter observations:
Litter size, sex and weight of pups, live and dead pups were examined.
Postmortem examinations (parental animals):
The necropsy included an examination of the external features of the carcass, external body orifices, the abdominal, thoracic, and cranial cavities, and organs/tissues.
At scheduled necropsy, the following organs were weighed: brain; liver; kidneys; prostate with seminal vesicles including secretions; testes, epididymides; ovaries; uterus (with cervix).
The following tissues from adult animals were preserved: cervix, caecum, coagulating gland, colon, duodenum; epididymides, oesophagus, ileum, jejunum, lesions, ovary with oviducts, prostate, rectum, seminal vesicle, stomach, testis, uterus, vagina.
Postmortem examinations (offspring):
At Day 4: Litter size and sex, weight, and abnormal observations of individual offspring were recorded. All dead pups were examined for cervical, thoracic and abdominal viscera abnormities and preserved in alcohol. Live pups were sacrificed, examined for gross external abnormalities, and discarded without necropsy.
Statistics:
Appropriate maternal and litter data were analyzed using analysis of variance (ANOVA) techniques; Levene's test was employed to analyze homogeneity of variances and Dunnett's t-test served as the post-hoc group comparison test. Calculated indices were analyzed using Fisher-Irwin exact test. Pup weights were analyzed by analysis of covariance (ANCOVA) techniques using the litter size as the covariate. Group comparisons were evaluated at the 5 % and 1 % two-tailed probability levels for all statistic analyses; a one-tailed analysis was used to determine significance for trend where appropriate.
Reproductive indices:
Fertility indices:
Female fertility = (number of females pregnant/total number females mated) x 100
Male fertility = (no. of males shown to be fertile/total number of males mated) x 100
Gestation index = (number of live litters born/number of pregnancies) x 100.
Offspring viability indices:
Viability index = (number of pups surviving to Day 4/number of pups born alive) x 100
Percent males (Days 0 and 4) = (number of males/total number of pups) x 100.
Clinical signs:
no effects observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
no effects observed
Organ weight findings including organ / body weight ratios:
no effects observed
Histopathological findings: non-neoplastic:
not specified
Other effects:
no effects observed
Reproductive function: oestrous cycle:
not specified
Reproductive function: sperm measures:
no effects observed
Reproductive performance:
no effects observed
There was no mortality. All animals survived to terminal sacrifice with the exception of two non pregnant females dosed at 100 and 250 mg/kg/day, respectively, which were sacrificed on day 26 due to their failure to deliver.
There were no treatment-related clinical signs of toxicity observed in F0 parents of either sex or in F1 pups at any treatment level. Remarkable clinical signs in the F0 parents and F1 pups were not attributed to treatment with diammonium persulfate, as they occurred sporadically, were of short duration, and did not demonstrate a dose response.
Normal progression of spermatogenesis was observed in male parental animals at necropsy. All stages of the spermatogenic cycle were present. In the testicles examination there was no indication of any test substance effects.
Key result
Dose descriptor:
NOAEL
Effect level:
>= 250 mg/kg bw/day
Sex:
male/female
Basis for effect level:
other: male and female toxicity
Key result
Dose descriptor:
NOAEL
Effect level:
>= 250 mg/kg bw/day
Sex:
male/female
Basis for effect level:
other: male and female fertility performance
Clinical signs:
no effects observed
Mortality / viability:
no mortality observed
Body weight and weight changes:
no effects observed
Sexual maturation:
not specified
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Histopathological findings:
no effects observed
There were no treatment-related clinical signs of toxicity observed in F1 pups at any treatment level. Remarkable clinical signs in the F1 pups were not attributed to treatment with diammonium persulfate, as they occurred sporadically, were of short duration, and did not demonstrate a dose response.
Key result
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
>= 250 mg/kg bw/day
Sex:
male/female
Basis for effect level:
other: pup viability and development
Key result
Reproductive effects observed:
not specified
Conclusions:
The NOAEL for male and female toxicity, for male and female fertility performance and for embryo/foetal viability and development was >= 250 mg/kg/day.
Executive summary:

Diammonium persulfate was tested for oral reproductive/developmental toxicity in a screening test with rats according to OECD guideline 421.The purpose of this study was to obtain initial information on the possible effects of the test item on reproduction and development when administered orally in the diet to Crl:CD (SD)IGS BR rats at doses of 40, 100 and 250 mg/kg bw/day compared to control animals (plain diet only).

There were no treatment-related clinical signs of toxicity observed in F0 parents of either sex or in F1 pups at any treatment level. Remarkable clinical signs in the F0 parents and F1 pups were not attributed to treatment with diammonium persulfate, as they occurred sporadically, were of short duration, and did not demonstrate a dose response. No significant changes were observed in male and female reproductive performance such as gonadal function, mating behaviour, conception, pregnancy, parturition and in development of the F1 offspring from conception to day 4 postpartum. In conclusion, under the conditions of this study, the NOAEL for male and female toxicity, the NOAEL for male and female fertility performance and the NOAEL for F1 viability and development was >= 250 mg/kg/day.

Endpoint:
one-generation reproductive toxicity
Type of information:
read-across based on grouping of substances (category approach)
Adequacy of study:
key study
Reason / purpose for cross-reference:
read-across source
Key result
Dose descriptor:
NOAEL
Remarks on result:
other: Will be completed when study has been finalised.
Key result
Dose descriptor:
NOAEL
Remarks on result:
other: Will be completed when study has been finalised.
Key result
Critical effects observed:
not specified
Key result
Reproductive effects observed:
not specified
Endpoint:
one-generation reproductive toxicity
Type of information:
read-across based on grouping of substances (category approach)
Adequacy of study:
supporting study
Reason / purpose for cross-reference:
read-across source
Key result
Dose descriptor:
NOAEL
Remarks on result:
not determinable because of methodological limitations
Remarks:
DRF
Key result
Critical effects observed:
no
Key result
Dose descriptor:
NOAEL
Remarks on result:
not determinable because of methodological limitations
Remarks:
DRF
Key result
Critical effects observed:
no
Key result
Reproductive effects observed:
no
Endpoint:
one-generation reproductive toxicity
Type of information:
read-across based on grouping of substances (category approach)
Adequacy of study:
supporting study
Reason / purpose for cross-reference:
read-across source
Key result
Dose descriptor:
NOAEL
Remarks on result:
not determinable because of methodological limitations
Remarks:
DRF
Critical effects observed:
not specified
Key result
Dose descriptor:
NOAEL
Remarks on result:
not determinable because of methodological limitations
Remarks:
F1 not included in this DRF
Critical effects observed:
not specified
Key result
Reproductive effects observed:
not specified
Endpoint:
one-generation reproductive toxicity
Type of information:
read-across based on grouping of substances (category approach)
Adequacy of study:
supporting study
Reason / purpose for cross-reference:
read-across source
Key result
Dose descriptor:
NOAEL
Effect level:
>= 250 mg/kg bw/day
Sex:
male/female
Basis for effect level:
other: male and female toxicity
Key result
Dose descriptor:
NOAEL
Effect level:
>= 250 mg/kg bw/day
Sex:
male/female
Basis for effect level:
other: male and female fertility performance
Key result
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
>= 250 mg/kg bw/day
Sex:
male/female
Basis for effect level:
other: pup viability and development
Key result
Reproductive effects observed:
not specified
Effect on fertility: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
250 mg/kg bw/day
Study duration:
subacute
Effect on fertility: via inhalation route
Endpoint conclusion:
no study available
Effect on fertility: via dermal route
Endpoint conclusion:
no study available
Additional information

OECD 421

Diammonium persulfate was tested for oral reproductive/developmental toxicity in a screening test with rats according to OECD guideline 421. The purpose of this study was to obtain initial information on the possible effects of the test item on reproduction and development when administered orally in the diet to Crl:CD (SD)IGS BR rats at doses of 40, 100 and 250 mg/kg bw/day compared to control animals (plain diet only). There were no treatment-related clinical signs of toxicity observed in F0 parents of either sex or in F1 pups at any treatment level. Remarkable clinical signs in the F0 parents and F1 pups were not attributed to treatment with diammonium persulfate, as they occurred sporadically, were of short duration, and did not demonstrate a dose response. No significant changes were observed in male and female reproductive performance such as gonadal function, mating behaviour, conception, pregnancy, parturition and in development of the F1 offspring from conception to day 4 postpartum. In conclusion, under the conditions of this study, the NOAEL for male and female toxicity, the NOAEL for male and female fertility performance and the NOAEL for F1 viability and development was 250 mg/kg/day.

OECD 443

The final decisions on a compliance check were received on 8 September 2016 for diammonium peroxodisulphate (APS), dipotassium peroxodisulphate (KPS) and disodium peroxodisulphate (SPS). A sequential testing had been agreed for the extended one-generation study and both pre-natal developmental toxicity studies. The prenatal developmental toxicity study in rats was performed first followed by the prenatal developmental toxicity study in rabbits and the extended one generation study in rats. All studies required pre-test for dose setting. The following table shows the initiation dates of the different studies performed:

Study

Start

Finalisation

IUCLID section / study no.

14 d DRF for prenatal developmental toxicity (rats)

02 August 2017

05 July 2018

7.8.2 / 552-410-3000

Prenatal developmental Toxicity rats

28 March 2018

24 July 2018

7.8.2 / 552-414-3001

14 d DRF for prenatal developmental toxicity (rabbits)

17 August 2018

27 December 2018

7.8.2 / 552-410-4018

Prenatal developmental toxicity rabbits

12 October2018

 09 September 2020

7.8.2 / 552-414-4019

14-Day Palatability Pre-study in the rats (APS in feed)

19 June 2019

16 July 2019

7.5.1 / 552-400-4688

Validation of the Analytical Method for the Determination of Diammonium Peroxodisulphate (APS) in Rodent Diet

16 September 2019

23 June 2020

Not included in IUCLID

DRF for Extended One Generation Reproduction Toxicity study (gavage)

20 November 2019

23 June 2020

7.8.1 / 552-421-5133

Second 14 d DRF for Extended One Generation Reproduction Toxicity study (one additional dose of 200 mg/kg bw/d) (gavage)

17 April 2020

02 June 2020

7.8.1 / 552-421-5133R

Extended One Generation Reproduction Toxicity study in rats (gavage)

23 June 2020

Expected Q2 2021

7.8.1 / 552-443-5437

 

The study package was initiated end of January 2017. Unforeseen problems regarding analytics, the stepwise testing approach and organizational reasons led to a delay of the complete study package at the laboratory. The high workload of the laboratory due to the last REACH deadline in May 2018 and the Covid-19 pandemic also contributed to the delay.

Furthermore additional studies had to be considered before performing the main study for the extended one generation reproduction toxicity (EOGRT). According to OECD guideline 443 the test protocol for EOGRT study was designed for administration of the test substance in diet which is the preferred route. The available dietary studies (with APS and SPS please refer to IUCLID sections 7.5.1 and 7.8.1) both emphasized palatability problems due to the treated diet. Thus, a 14-day palatability pre-test was performed at 250 and 500 mg/kg bw/d of APS in diet to check for possible palatability issues for the main study. Animals did not show any severe signs of toxicity and also no palatability issues. Thus, analytical method development was initiated. However the method validation showed that the substance seemed to be unstable in the diet. For diet preparation water had to be added by the feed producer. APS seem to react with the diet in presence of water. This led to a very low recovery rate. Therefore, it was judged that application via the diet is not possible and gavage was chosen as main application route for the EOGRT study.

The determination of the correct dosing levels for the main EOGRT study also took longer than expected. As no data for the gavage application was available, a DRF (similar to OECD 421) for the OECD 443 was performed with 50, 100 and 250 mg/kg bw/d. At 250 mg/kg bw/d four male animals died on Days 4 and 5 (macroscopic and histopathology changes in stomach and intestines were observed during necropsy). Depressed body weight/food consumption and clinical signs were observed for all surviving male and female animals. Therefore the high dose was lowered to 180 mg/kg bw/d. At this dose the animals recovered completely. Only the body weight development of the male animals stayed slightly behind the control animals until the end of the DRF. An additional DRF study for 14 days was performed using 200 mg/kg bw/d in order to find a dose that causes toxicological effects but not severe suffering as required by the guideline. At this dose level the animals showed no signs of toxicity within the 14 days study period. Due to the steep dose response curve (no effects at 200 mg/kg bw/d and mortality at 250 mg/kg bw/d). It was decided to use 200 mg/kg bw/d as the highest dose in the EOGRT study. This dose had to be reduced again in the meantime to 180 mg/kg bw/d due to severe clinical signs and body weight loss within first week of the main study.

The main study started in June 2020. A confirmation of commissioned and running studies was provided by Toxicoop and is attached to the study record of the OECD 443 entry. The draft report was announced to be available in Q2 2021. The dossier will be updated as soon as the final report is available.

Effects on developmental toxicity

Description of key information

Of the Persulfate Category, only diammonium persulfate was tested for developmental toxicity in tests with rats and rabbits according to OECD guideline no. 414.

 

OECD 414, rat

Oral treatment of pregnant Han: Wistar rats from gestation day 5 up to day 19 (the day before Caesarean section) with Diammonium peroxodisulphate (APS) at the dose levels of 10, 30 and 100 mg/kg bw/day did not cause death, or revealed clinical signs and treatment related macroscopic changes at gross pathology. Slightly reduced food intake on GD 5 to 8 and 11 to 14 and slightly lower body weight gain on GD 5 to 8, 17 to 20 and 0 to 20, as well as slightly lower corrected body weight/gain were attributed to the test item at the dose level of 100 mg/kg bw/day. Diammonium peroxodisulphate (APS) did not reveal any adverse effect on the pre- and post-implantation loss, number of implantation, sex distribution, body and placental weight, external, visceral and skeletal development of the fetuses. Based on these observations the No Observed Adverse Effect Level (NOAEL) was determined as follows:

NOAEL (maternal toxicity): 30 mg/kg bw/day

NOAEL (developmental toxicity including teratogenicity): 100 mg/kg bw/day

 

OECD 414, rabbit

Oral (gavage) treatment of pregnant New Zealand White rabbits from gestation day 6 up to day 27 (the day before Caesarean section) with Diammonium peroxodisulphate (APS) at the dose level of 100 mg/kg bw/day induced maternal toxicity manifest as initial weight loss and subsequently reduced body weight gain together with reduced food consumption. The NOAEL for maternal toxicity is 30 mg/kg bw/day.

The severity of the maternal toxicity at 100 mg/kg bw/day was considered to impact fetal viability and growth and to slightly delay ossification. This dose of APS did not induce fetal malformations. The NOAEL for developmental toxicity is 30 mg/kg bw/day.

Effect on developmental toxicity: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
30 mg/kg bw/day
Study duration:
subacute
Species:
rabbit
Quality of whole database:
Two studies according to GLP and OECD 414 guideline are available in the rat and in the rabbit. Based on NOAELs obtained, the rabbit being more sensitive than the rat is considered to reflect the worst case.
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

OECD 414 in the rat

Diammonium peroxodisulphate (APS) was examined for its possible prenatal developmental toxicity in accordance with OECD guideline 414. Groups of 26 sperm-positive female Han: Wistar rats were treated with APS by oral administration daily at three dose levels of 10, 30 and 100 mg/kg bw/day respectively from day 5 up to and including day 19 post coitum. A control group of 26 sperm positive females was included and the animals were given the vehicle water (aqua purificata). The treatment volume was 5 mL/kg bw.

A sufficient stability and homogeneity in the chosen vehicle was verified over the range of relevant concentrations at the appropriate frequency of preparation. Diammonium peroxodisulphate (APS) in water was stable at room temperature and in the refrigerator (5 ± 3 °C) for three days at the concentrations of ca. 0.1 and 200 mg/mL. Analytical control of dosing solutions was performed during the first and last week of treatment. Concentrations of the test item in the dosing formulations varied in the acceptable range between 95 and 104 % of nominal concentrations at both analytical occasions confirming proper dosing.

During the study, mortality was checked and clinical observations were performed. Body weight and food consumption of the dams were also recorded. The day, when sperm was detected in the vaginal smear, was regarded as day 0 of gestation. Caesarean section and gross pathology were performed on gestational day (GD) 20. The number of implantations, early and late resorptions, live and dead fetuses in each uterine horn and the number of corpora lutea were recorded. Each fetus was weighed and examined for sex and gross external abnormalities. The placentas were weighed and examined externally. About half of each litter was preserved for visceral examination and the other half of the litters were preserved for skeletal evaluation. At visceral examination the bodies were micro dissected by means of a dissecting microscope. The heads were examined by Wilson's free-hand razor blade method. After cartilage-bone staining the skeletons were examined by means of a dissecting microscope. All abnormalities found during the fetal examinations were recorded.

In total, on gestation day 20 there were dams with 23, 20, 16 and 16 implantation sites in the control, 10, 30 and 100 mg/kg bw/day group, respectively. None of the females died before scheduled necropsy and there were no test item related clinical signs recorded. No treatment related necropsy findings were observed. Slight, statistically significant decreases of food consumption were indicated in the 100 mg/kg bw/day dose group on GD 5 to 8 and 11 to 14. Slightly lower body weight gain was observed between GD 5 and 8 and 17 and 20. A slightly lower corrected body weight/gain was observed between GD 0 to 20 without a statistical significance.

These differences were in association with the results of the DRF study (Dose Range Finding Prenatal Developmental Toxicity Study with Diammonium Peroxodisulphate (APS) in the Rat by Oral (Gavage) Administration Toxi-Coop study number: 552-410-3000) and were therefore attributed to the test item.

The CH square test indicated a statistical significance in the increase of pre-implantation loss in all test item treated groups. A statistical significant increase of the total intrauterine mortality was observed in the 100 mg/kg bw/day dose group. The mean percentage pre-implantation loss was between the historical control range or slightly above (30 mg/kg bw/day). The mean percentage intrauterine mortality was also between the historical background values. Thus, this was not attributed to treatment. Moreover the mean number of implantations was highest in the 100 mg/kg bw/day dose. There were no dose related increases seen in the post-implantation loss and sex distribution of the fetuses was not influenced by the treatment.

There were no test item related adverse effects on the fetal- and placental weight. There were no test item related external malformations and variations found. The visceral malformations were not attributed to the treatment. The skeletal malformations were found in three fetuses with a statistical significance. However the incidence was low and the type of alterations less severe and partially different. There was no dose related increase seen in external and skeletal variations. Based on these observations the No Observed Adverse Effect Level (NOAELs) were determined as follows: NOAEL (maternal toxicity): 30 mg/kg bw/day, NOAEL (developmental toxicity): 100 mg/kg bw/day, NOAEL (teratogenicity): 100 mg/kg bw/day (high dose).

 

OECD 414 in the rabbit

Diammonium peroxodisulphate (APS) was examined for its possible prenatal developmental toxicity. Groups of 25 (low and mid dose) and 26 (high dose) inseminated New Zealand White rabbits were treated with Diammonium peroxodisulphate (APS) by oral (gavage) administration daily at three dose levels of 10, 30 and 100 mg/kg bw/day respectively from day 6 up to and including day 27 post insemination. A control group of 25 inseminated females was included and the animals were given the vehicle water (aqua purificata). The treatment volume was 2 mL/kg bw.

Diammonium peroxodisulphate (APS) was proved to be stable in water formulations at 0.1 mg/mL and 200 mg/mL concentration levels for three days at room temperature and in the refrigerator (5 ± 3°C) according to the analytical method validation (Study no.: (Toxi-Coop study no. 552-100-3004)) at Toxi-Coop Zrt. The suitability of the chosen vehicle for the test item was analytically proven. Analytical control of dosing solutions was performed during the first and last week of treatment. The mean of the test item concentrations of the test item in the dosing formulations varied in the acceptable range between 96 and 99 % of nominal concentrations at both analytical occasions confirming proper dosing.

During the study, mortality was checked and clinical observations were performed. Body weight and food consumption of the does were recorded. The day of insemination was regarded as day 0 of gestation. Caesarean section and gross pathology were performed on gestational day 28. The number of implantations, early and late resorptions, live and dead fetuses in each uterine horn and the number of corpora lutea were recorded. Each fetus was weighed, measured and examined for gross external abnormalities. The placentas were weighed and examined externally.

Fresh visceral examination of each fetus was performed including the determination of the gender.

Head of about 50 % of each litter was removed, fixed in modified Sanomiya solution and washed in 90 % isopropanol. Examination of the heads was done by Wilson's free-hand razor blade method. All skeletons were examined by means of a dissecting microscope after cartilage-bone staining.

There was no test item related mortality, moribund state or abortion observed. In total, on gestation day 28 there were 22, 23, 21 and 20 evaluated litters in the control, 10, 30 and 100 mg/kg bw/day group respectively. There were no test item related clinical signs and pathological macroscopic findings observed.

Treatment with the test item at 100 mg/kg bw/day induced maternal toxicity manifest as an initial weight loss and subsequent reduction in body weight gain (77% between GD 6-28). Corrected body weight and corrected body weight gain clearly reflected the effect at 100 mg/kg bw/day. The reduction in body weight correlated with a reduction in food consumption, observed from the start of the treatment. Treatment with the test item at 30 mg/kg bw/day did not induce maternal toxicity. Variations in weight gain were not statistically significant during the study. Variations in the food consumption were not statistically significant at 30 mg/kg bw/day except for GD 18-21. This did not result in statistically lower body weight gain. There was no effect of 10 mg/kg bw/day on maternal body weight or food consumption.

There was evidence of an increase in early embryonic death/post-implantation loss/total intrauterine mortality and a slightly lower mean number of viable fetuses (without a statistical significance) in the 100 mg/kg bw/day dose group. This outcome was considered to be related to the severity of the maternal toxicity induced.

There was no effect of treatment at 10 or 30 mg/kg bw/day on intrauterine mortality.

Significantly lower fetal weight and crown-rump length were observed in the 100 mg/kg bw/day dose group. These smaller fetuses showed evidence of delayed ossification (e.g. larger or slightly larger anterior fontanelle, reduced or asymmetric ossification of the bones of the digits (including pollex) or small hole in xiphoid cartilage. These effects were considered to be a consequence of the maternal toxicity induced. There was no evidence of treatment-related malformation at 100 mg/kg bw/day. There was no effect of treatment at 10 or 30 mg/kg bw/day on fetal growth or development.

Oral (gavage) treatment of pregnant New Zealand White rabbits from gestation day 6 up to day 27 (the day before Caesarean section) with Diammonium peroxodisulphate (APS) at the dose level of 100 mg/kg bw/day induced maternal toxicity manifest as initial weight loss and subsequently reduced body weight gain together with reduced food consumption. The NOAEL for maternal toxicity is 30 mg/kg bw/day.

The severity of the maternal toxicity at 100 mg/kg bw/day was considered to impact fetal viability and growth and to slightly delay ossification. This dose of Diammonium peroxodisulphate (APS) did not induce fetal malformation.

The NOAEL for developmental toxicity is 30 mg/kg bw/day.

Justification for classification or non-classification

Based on the results obtained, substances of the Persulfate Category were not classified and labelled for reproduction/developmental toxicity according to Regulation (EC) No 1272/2008, as amended for the fifteenth time in Regulation (EU) 2020/1182.

Additional information