2,2'-oxybis[5,5-dimethyl-1,3,2-dioxaphosphorinane] 2,2'-disulphide

Endpoint summary

Description of key information

The submission substance did not produce adverse effects in a reliable combined repeated dose oral toxicity study with a reproduction / developmental toxicity screening test (NOAEL >/= 1000 mg/kg bw/day).

No reliable standard sub-chronic toxicity study (90-day) is available as required in REACH Annex IX columns 1 and 2, under section 8. Thus, a sub-chronic toxicity study (90-day) according to OECD 408 is proposed by the registrant to fill the data gap. In accordance with column 2 of REACH Annexes VIII and IX, studies with inhalation or dermal exposure are not necessary.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Reference

Reference 1

Endpoint:

sub-chronic toxicity: oral

Type of information:

experimental study planned

Justification for type of information:

TESTING PROPOSAL ON VERTEBRATE ANIMALS

NON-CONFIDENTIAL NAME OF SUBSTANCE:
- Name of the substance on which testing is proposed to be carried out:
2,2'-oxybis[5,5-dimethyl-1,3,2-dioxaphosphorinane]2,2'disulphide
- Synonym: 1,3,2-Dioxaphosphorinane, 2,2'-oxybis 5,5-dimethyl-, 2,2'-disulfide
- Commercial name: Exolit 5060
- CAS No.: 4090-51-1

CONSIDERATIONS THAT THE GENERAL ADAPTATION POSSIBILITIES OF ANNEX XI OF THE REACH REGULATION ARE NOT ADEQUATE TO GENERATE THE NECESSARY INFORMATION
- Available GLP studies:
Regarding IUCLID endpoint 7.5.1 “Repeated Dose Toxicity”, there is no reliable standard sub-chronic toxicity study (90-day) nor a short-term toxicity study (28-day) available. Instead, there is a guideline conform OECD 422 study, dated 2013, available. As this study is a combined repeated dose toxicity with reproduction/ developmental screening test, the relevant data for repeated oral application of the test item are available and discussed in the dossier. The test item was orally administered repeatedly to male (28 days) and female (maximum 54 days) Wistar rats at dosages of 100, 300 and 1000 mg/kg body weight/ day. In conclusion, the repeated dose administration of the test substance to the male (28 days) and female (maximum 54 days) Wistar rats revealed neither mortalities nor findings of toxicological relevance in male and female animals. There were also no toxicologically relevant findings noted for reproductive and developmental parameters.

- Available non-GLP studies:
There is a non-GLP dose-range-finding study for the OECD 422 study, dated 2013, available, which is presented and discussed in IUCLID section 7.8.1 “Toxicity to Reproduction”
- Historical human/control data: n.a.
- (Q)SAR: No validated (Q)SAR models exist for this endpoint for this substance. Based on the absence of significant toxicity in the OECD 422 study (including a 28-day repeated dose toxicity part), no indication of a specific mode of action for this substance exist.”

- In vitro methods: Currently there are no sufficient in vitro methods available to receive the required information from sub-chronic repeated oral application over a period of 90 days.
- Weight of evidence: There are no sufficient analogue substances available for a weight of evidence approach to get the required information from sub-chronic repeated oral application over a period of 90 days.
- Grouping and read-across: There are no suitable analogue substances available for a read across strategy.
- Substance-tailored exposure driven testing [if applicable]: n.a.
- Approaches in addition to above [if applicable]: n.a.
- Other reasons [if applicable]: n.a.

CONSIDERATIONS THAT THE SPECIFIC ADAPTATION POSSIBILITIES OF ANNEXES VI TO X (AND COLUMN 2 THEREOF) OF THE REACH REGULATION ARE NOT ADEQUATE TO GENERATE THE NECESSARY INFORMATION:
- In accordance with column 2 of Reach Regulation (EC) No 1907/2006 Annex VIX, amended 17.June 2021 (EU) by 2021/979 of EU Commission, the need to perform a study on sub-chronic toxicity (90-day) is not needed if … ”- a reliable short-term toxicity study (28 days) is available showing severe toxicity effects according to the criteria for classifying the substance, as STOT RE (Cat 1 or 2) …”. This requirement is not fulfilled. Although a short-term toxicity study as part of the OECD 422 study is available for the test substance, no severe toxicologically relevant findings were noted. Thus, a sub-chronic toxicity study is proposed to fill the data gap.

FURTHER INFORMATION ON TESTING PROPOSAL IN ADDITION TO INFORMATION PROVIDED IN THE MATERIALS AND METHODS SECTION:
- Details on study design/methodology proposed:
Sub-chronic 90-day toxicity study in rats (oral gavage) according OECD 408 testing guideline. At least 20 animals (ten female and ten male) should be used for each dose level. An additional satellite group of at least ten animals (five per sex) in the control and in the top dose group for observation after the treatment period, for the potential reversibility or persistence of any toxic effects.

Qualifier:

according to guideline

Guideline:

OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)

Critical effects observed:

not specified

Reference 2

Endpoint:

short-term repeated dose toxicity: oral

Remarks:

combined repeated dose and reproduction / developmental screening

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2012-08-03 to 2013-03-25

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP Guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

(Bayerisches Landesamt für Gesundheit und Lebensmittelsicherheit, München, Germany)

Limit test:

no

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

Test System
Species/strain: Wistar rats, Crl: WI(Han) (Full Barrier)
Source: Charles River, 97633 Sulzfeld, Germany
Sex: male and female; the female animals were non-pregnant and nulliparous.
Age at the start of the treatment period: males: 9-10 weeks old, females: 9-10 weeks old.
Body weight at the allocation of the animals to the experimental groups: males: 236 - 263 g; (mean: 252.08 g, ± 20% = 201.66 – 302.49 g)
females: 162 - 188 g; (mean: 176.35 g, ± 20% = 141.08 – 211.62 g)
The animals were derived from a controlled full-barrier maintained breeding system (SPF). According to Art. 9.2, No. 7 of the German Act on Animal Welfare the animals were bred for experimental purposes.

Housing and Feeding Conditions
- Full barrier in an air-conditioned room
- Temperature: 22 +/- 3°C
- Relative humidity: 55 +/- 10%
- Artificial light, sequence being 12 hours light, 12 hours dark
- Air change: 10 x / hour
- Free access to Altromin 1324 maintenance diet for rats and mice (lot no. 0939)
- Free access to tap water, sulphur acidified to a pH of approximately 2.8 (drinking water, municipal residue control,
microbiological controls at regular intervals)
- The animals were kept individually in IVC cages (except during the mating period when one female will be paired with one male),
type III H, polysulphone cages on Altromin saw fibre bedding (lot no. 300512)
- Certificates of food, water and bedding are filed at BSL BIOSERVICE
- Adequate acclimatisation period (at least 5 days) under laboratory conditions

Preparation of the Animals
Prior to the start of the treatment period a detailed clinical observation outside the home cage was made.
Before the first administration all animals used for the study were weighed and assigned to the experimental groups
with achieving a most homogenous variation in body weight throughout the groups of males and females.

Route of administration:

oral: gavage

Vehicle:

corn oil

Details on oral exposure:

The test item was weighed into a tarred plastic vial on a precision balance.
The dose formulations were prepared by adding the required volume of corn oil and further vortexing it for 2-3 minutes.
The vehicle was selected based on the test item’s characteristics. The test item formulations were prepared freshly on each administration day
before the administration procedure. The time of preparation and time of dosing was recorded for all dosing formulations.
The homogeneity was guaranteed by intensive stirring during application.

The following doses were evaluated:
Control: 0 mg/kg body weight
Low Dose: 100 mg/kg body weight
Medium Dose: 300 mg/kg body weight
High Dose: 1000 mg/kg body weight

The highest dose level was chosen with the aim of inducing toxic effects, but no death or severe suffering.
Thereafter, a descending sequence of dose levels was selected with a view to demonstrate any dosage related response and NOAEL.
The animals in the control group were handled in an identical manner to the test group subjects and received the vehicle
using the same dose volume.

Dose volumes were adjusted individually based on weekly body weight measurements. The administration volume was 5 mL/kg body weight.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Each dosing concentration was analyzed for nominal concentration.
Stability and homogeneity of the test item in the vehicle was analyzed for the LD, MD and HD dosing formulation.
Samples for the nominal concentration verification was taken in study week 1 (first week of pre mating period), 3 (first week of mating),
5 (gestation) and 7 (gestation/lactation) of control and all treatment groups.
Samples for homogeneity were taken from the top, middle and bottom of HD, MD, and LD preparation in study week 1 and 5.
All formulation samples were preserved at -20 oC until the analysis depending on the procedure. The samples were analyzed at
BSL BIOSERVICE Scientific Laboratories GmbH. The procedure followed for the sample analysis was mentioned in phase plan
(BSL phase study Nr. 122500).

Duration of treatment / exposure:

The animals were treated with the test item formulation or vehicle on 7 days per week for a period of 54 days,
i.e. during 14 days of pre-mating and 14 days of mating in both males and females, during the gestation period and
up to post-natal day 3 in females. Males were dosed after the mating period until the minimum total dosing period of 28 days was completed.

Frequency of treatment:

Daily

Remarks:

Doses / Concentrations:
0, 100, 300, 1000 mg/kg bw/day
Basis:
actual ingested

No. of animals per sex per dose:

80 animals (40 males and 40 females) were included in the study (10 male and 10 female animals per group).

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: The highest dose level was chosen with the aim of inducing toxic effects, but no death or severe suffering.
Thereafter, a descending sequence of dose levels was selected with a view to demonstrate any dosage related response and NOAEL.
The animals in the control group were handled in an identical manner to the test group subjects and received the vehicle
using the same dose volume.

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: daily
Once before the first exposure, and once a week thereafter, detailed clinical observations were made in all animals outside the home cage in a standard arena. Multiple detailed behavioural observations were made in the week before the first treatment and during the last week of the treatment in 5 randomly selected males and on lactation days in 5 randomly selected females (only lactating females were evaluated) outside the home cage using a functional observational battery of tests.

BODY WEIGHT: Yes
- Time schedule for examinations: The body weight was recorded once before the assignment to the experimental groups, on the first day of administration and weekly during the treatment period as well as at the end of the study. During pregnancy, females were weighed on gestation days (GD) 0, 7, 14 and 20 and within 24 hours of parturition (day 0 post-partum) as well as day 4 post-partum along with pups.

FOOD CONSUMPTION:
- Food consumption was measured weekly on the corresponding days of the body weight measurements after the beginning of the
dose administration. Food consumption was not measured during the mating period in males and females and the post-mating period in males.

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: Yes

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No

OPHTHALMOSCOPIC EXAMINATION: yes

HAEMATOLOGY: Yes
- Time schedule for collection of blood: at the end of the treatment period
- Anaesthetic used for blood collection: Yes (ketamine/xylazin, 3:1)
- Animals fasted: No
- How many animals: five randomly selected males and females of each group
- Parameters examined: haematocrit value (Hct), haemoglobin content (Hb), red blood cell count (RBC), mean corpuscular volume (MCV), mean corpuscular haemoglobin (MCH), mean corpuscular haemoglobin concentration (MCHC), reticulocytes (Re), platelet count (PLT),
white blood cells (WBC), neutrophils (Neu), lymphocytes (Lym), monocytes (Mono), eosinophils (Eos), basophils (Baso), prothrombin time (PT),
activated partial thromboplastin time (aPTT)



CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: at the end of the treatment period
- Animals fasted: No
- How many animals: five randomly selected males and females of each group
- Parameters examined: alanine aminotransferase (ALAT), aspartate-aminotransferase (ASAT), alkaline phosphatase (AP),
creatinine (Crea), total protein (TP), albumin (Alb), urea, total bile acids (TBA), total cholesterol (Chol), glucose (Gluc), sodium (Na),
potassium (K), Calcium (Ca), Phosphorus (PHS)


URINALYSIS: Yes
- Time schedule for collection of urine: 5 randomly selected males at necropsy.
- Metabolism cages used for collection of urine: No
- Animals fasted: No
- Parameters were examined: specific gravity, nitrite, ph-value (pH), protein, glucose, ketone bodies (ketones), urobilinogen (ubg),
bilirubin, blood, leukocytes


NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: in the week before the first treatment and during the last week of the treatment
- Dose groups that were examined: all
- Battery of functions tested: sensory activity / grip strength / motor activity

OTHER:

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
All male animals were sacrificed after the completion of the mating period (total dosing period: 28/ 29 days) on study day 29/ 30,
while female animals were sacrificed on post-natal day 4 using an anaesthesia (ketamine/xylazin, 3:1, medistar Arzeneimittel, lot no: 00212,
expiry date: 03/2014, Actavis, lot no: 146066, expiry date: 10/2014, Serumwerk, lot no: 00711, expiry date: 08/2013 and Narcoren®,
Merial; lot no.: 221022; expiry date: 28/02/2015) was used. Pups were killed on PND 4 by decapitation.
Pups sacrificed on day 4 post-partum were carefully examined externally for gross abnormalities.
Females (animals 50, 60, 70 and 73) showing no sign of pregnancy was sacrificed 26 days after the last day of the mating period.
All animals were subjected to a detailed gross necropsy which includes careful examination of the external surface of the body,
all orifices and the cranial, thoracic and abdominal cavities and their contents.
The number of implantation sites and corpora lutea was recorded for each parental female at necropsy.

The wet weight of the organs (liver, kidneys, adrenals, testes, epididymides, prostate, seminal vesicles and coagulating glands,
ovaries, uterus with cervix, thymus, thyroid/parathyroid glands, spleen, brain, pituitary gland, heart) of 5 males and 5 females randomly
selected from each group was recorded as soon as possible. Paired organs were weighed separately. In addition reproductive organs
of all animals were weighed.

The following tissues (brain (cerebrum, cerebellum and pons), ovaries (females), spinal cord, uterus with cervix (females),
liver, vagina (females), kidneys, testes (males), adrenal glands, epididymides (males), stomach, prostate and seminal vesicles
with coagulating glands as a whole (males), small and large intestines (including Peyer´s patches), urinary bladder,
thymus, lymphnodes (mesentric and axillary), Thyroid, peripheral nerve (e.g. sciatic nerve) with skeletal muscle,
spleen, bone with bone marrow (sternum), lung and trachea pituitary gland, mammary glands, oesophagus, heart, gross lesions)
of the same selected animals from each group were preserved in 10% neutral buffered formalin except eyes, testes and epididymides
that were fixed in Modified Davidson’s Fixative for approximately 24 hours before they were transferred to 10% neutral buffered formalin.

HISTOPATHOLOGY: Yes

All organs and tissues listed above were evaluated from five randomly selected males and females of the control and high dose group:
Males Nos.: 2, 3, 4, 8, 9, 34, 35, 36, 38, 40; Females Nos.: 41, 42, 43, 45, 46, 72, 74, 77, 79, 80.
Reproductive organs (ovary, uterus, cervix, vagina, testis, epididymis, prostate gland, seminal vesicle and coagulating gland)
and macroscopic changes were evaluated in all study animals. For the testes, a detailed qualitative examination was made; taking
into account the tubular stages of the spermatogenic cycle for the evaluation of additional hematoxylin-PAS (Periodic Acid Schiff) stained slides.
Histological processing of tissues to microscope slides was performed at the GLP-certified contract laboratory Propath UK Ltd.
(test site for tissue processing), Willow Court, Netherwood Road, Hereford HR2 6JU, England. Histopathological evaluation was
performed at the GLP-certified contract laboratory KALEIDIS – Consultancy in Histopathology (test site for histopathology),
6 rue du Gers, 68300 Saint-Louis, France. Blocking, embedding, cutting, H&E staining and scientific slide evaluation were
performed according to the corresponding SOP’s of the test sites.

Statistics:

A statistical assessment of the results of the body weight, food consumption, parameters of haematology,
blood coagulation and clinical biochemistry and absolute and relative organ weights were performed for each gender by comparing
values of dosed with control animals of the main groups using a one-way ANOVA and a post-hoc Dunnett Test.
These statistics were performed with GraphPad Prism 5.01 software (p<0.05 was considered as statistically significant).

Clinical signs:

no effects observed

Mortality:

no mortality observed

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

no effects observed

Food efficiency:

not examined

Water consumption and compound intake (if drinking water study):

not examined

Ophthalmological findings:

no effects observed

Haematological findings:

no effects observed

Clinical biochemistry findings:

no effects observed

Urinalysis findings:

no effects observed

Behaviour (functional findings):

no effects observed

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

no effects observed

Details on results:

Mortality
There was no mortality noted in treated and control groups during the study period.

Clinical Observations
There were clinical signs recorded in male and female animals of treated groups, which were isolated and transient.
These findings were not considered to be of toxicological relevance.
During the weekly detailed clinical observation, no significant changes or differences between the groups were found.
There were no ophthalmoscopic findings in any of the animals of this study.

Functional Observations
No relevant effects were observed in any of the parameters of the functional observation battery before and at the end of the treatment period.
There were no biologically relevant differences in body temperature between the groups.

Body Weight Development
In both males and females, no treatment related changes were noted for body weight and body weight gain during the study period.
Statistically there were no significant diferences noted between the treated and corresponding controls.

Food Consumption
In both males and females, no treatment related changes were noted for treated group when compared to corresponding control.
The statistical evaluation of data revealed significant increase in food intake in male HD group.
This change was not considered to have toxicological relevance.

Haematology and Coagulation
There were no statistically significant changes noted for haematological and coagulation parameters in male and female treated groups
when compared to the corresponding control. However, there was slight decrease in mean monocyte and eosinophil values in male HD group
and slight increase in mean WBC value in female HD group. In the absence of statistical significance and the values being within the historical
control range the changes were not considered to have toxicological relevance.

Clinical Biochemistry
There were no treatment related changes considered for measured clinical biochemistry parameters of male and female treated groups
when compared to corresponding control. However, there was statistically significant increase noted for mean TP value female LD and HD groups.
In the absence of dose response pattern no relevance to treatment was considered.
There was also increase in mean ALP values in male treated groups (LD, MD and HD groups) and female HD group; increase in mean GLU values
in male MD and HD groups and increase in mean Crea values in male HD group. These changes did not show statistical significance and in
addition the changes noted for GLU and Crea of male and ALP of female lacked dose response pattern. These changes were not of toxicological
relevance. The changes noted for ALP values in males of HD group were not associoated with test item due to absence of relevant
histological changes.

Urinalysis
The urinalysis performed in male animals revealed no treatment related effect in treated groups when compared to the control group.

Pathology
One control female (No. 50), one female LD group (No. 60), one female of MD group (No. 70) and one female of HD group (No. 73) were
found not to be pregnant at terminal sacrifice, but in view of the group distribution this was not considered to be treatment-related.
Other macroscopic organ findings noted were very few, and all of them were considered to be incidental and not to be test item-related.
This included a yellow spot on the epididymis in one control and two treated males, which were confirmed histologically to correspond
to spermatic granuloma, a change observed spontaneously in male rats of this strain and age.

Organ Weight
There were no changes noted for organ weight in both males and females considered to be related to treatment when compared to corresponding
control. However, the statistical analysis of data revealed significant increase in relative (to terminal body weight) of thyroid (+parathyroid)
gland in male LD group. In the absence of dose response this change was not considered to be related to treatment.

Histopathology
Reproductive organs
There was no indication of test item-related histopathological findings in reproductive organs of male or female rats of this study.
In the males, some minor degenerative testicular and associated epididymal changes were seen in two rats of MD group, but were not
considered test item-related as they lacked dose relationship. Spermatic granuloma(s) were noted in a low number of treated rats.
They were considered to be spontaneous in nature as they are occasionally observed in untreated male rats of this strain and age.
Reproductive organs of most female study animals showed typical post-partum histomorphology without any relevant inter-group difference.
There was a tendency towards a lower number of large corpora lutea in the ovary of females of HD group, but in the absence of any effect on
litter size this was not considered a test item-related effect.
The reproductive organs of the control and treated females found not to be pregnant at terminal sacrifice showed normal reproductive sexual cycle.The female treated at 100 mg/kg/day had a mild mixed cell endometritis of the uterus which might have contributed to the lack of gravidity in
this animal but was not considered treatment-related in view of its lack of dose relationship and isolated occurrence in this study.
Other organs
No test item-related histopathological findings were noted in the other organs evaluated in this study.
All histopathological changes seen at terminal sacrifice were considered to be incidental in origin and/or within the range of expected
changes for rats of this age and strain kept under laboratory conditions.

Dose descriptor:

NOAEL

Effect level:

>= 1 000 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: no effects adverse observed at the highest dose

Critical effects observed:

not specified

Concentration analysis of formulation samples was determined in study week 1, 3, 5, and 7 for all dose groups. The mean recoveries observed in LD, MD and HD groups were 105.1%, 100.0%, and 94.6% of the nominal concentration, respectively. Homogeneity of formulation samples was determined in study week 1 and 5 for all dose groups. The mean recoveries observed for LD group were 105.3% and 106.1%, for MD group 97.4% and 98.9%, and for HD group 94.3% and 95.7% of the nominal value. The coefficients of variation of the different sampling locations (top, middle, bottom) were in LD group 0.5% and 4.6%, in MD group 1.0% and 2.5%, and in HD group 0.7% and 0.7%.

Conclusions:

In conclusion, the repeated dose administration of the test item to the male (28 days) and female (maximum 54 days)
Wistar rats at dosages of 100, 300 and 1000 mg/kg body weight/ day revealed neither mortalities nor findings of toxicological relevance in male and female animals. There were also no toxicologically relevant findings noted for reproductive and developmental parameters.
Based on the data generated from this “Combined Repeated Dose Oral Toxicity Study with the Reproduction/ Developmental Toxicity Screening
Test the no observed adverse effect level (NOAEL) for general and reproductive and developmental toxicity is considered to be 1000 mg/kg body weight/ day.

Executive summary:

The aim of this study was to assess the possible effects of the test item on toxicity after repeated exposure, male and female fertility and embryofetal development in Wistar rats (10 animals per sex and group). The test item was administered daily in graduated doses to 3 groups of test animals, one dose level per group for a treatment period of approx. 54 days. Animals of control group were handled identically as the dose groups but received same dose volume of the vehicle used in this study. The groups received the test item in concentrations of 0, 100, 300 and 1000 mg/kg body weight/day

After 14 days of treatment to both male and female, animals were mated (1:1) for a maximum of 14 days. The males were sacrificed after completion of the mating period on day 29 and the females along with their pups were sacrificed on post natal day 4. Non-pregnant females were sacrificed on GD 26. Pups were sacrificed on postnatal day 4. A full histopathological evaluation of the tissues was performed on high dose and control animals. Organs showing gross alterations were also examined histopathologically. Reproductive organs were evaluated in all study animals.

No mortality occurred in the control or any of the dose groups during the treatment period of this study. There were no clinical signs considered to be of toxicological relevance recorded in male and female animals of treated groups. During the weekly detailed clinical observation, no significant changes or differences between the groups were found. There were no ophthalmoscopic findings in any of the animals of this study.

No relevant effects were observed in any of the parameters of the functional observation battery before and at the end of the treatment period.

There were no biologically relevant differences in body temperature between the groups. In both males and females, no treatment related changes were considered for body weight, body weight change and food consumption in treated groups when compared to control.

No treatment-related changes were noted for number of still births, number of runts, total number of pups born on PND 0 and number of male and female pups, sex ratio, live pups on PND 0 and PND 4. No treatment related changes were noted for the mean litter weight, total litter weight, male and female litter weight on PND 0 and 4 in treated groups when compared to corresponding control. No treatment related changes were noted for the precoital interval and duration of gestation in treated groups when compared to control. All pregnancies resulted in normal births. No treatment related changes were noted for number of corpora lutea, number of implantation sites, number of live pups born on PND 0 and percentage of pre- and post-implantation loss in treated groups when compared to control. There were no treatment related changes noted for copulation index (%), fertility index (%) and delivery index (%) in treated groups when compared to corresponding control group. No treatment related changes were noted for survival of the pups from PND 0 to PND 4 treated group when compared to controls. No treatment related changes were considered for viability index (%). No treatment-related gross external findings were observed in any of the treated groups.

There were no statistically significant changes considered to be of toxicological relevance noted for haematological, clinical biochemistry and coagulation parameters in male and female treated groups when compared to the corresponding control. The urinalysis performed in male animals revealed no test- item related effect in any of the treated groups when compared to control. One control female, one female of the low dose group, one female of mid dose group and one female of high dose group were found not to be pregnant at terminal sacrifice, but in view of the group distribution this was not considered to be treatment-related.

The macroscopic organ findings noted were very few, and all of them were considered to be incidental and not to be test item-related. There were no changes noted for organ weight in both males and females considered to be related to treatment when compared to corresponding control. No test item-related histopathological findings were noted in the reproductive organs and in the other organs evaluated in this study.

The no observed adverse effect level (NOAEL) for general and reproductive and developmental toxicity of this study is considered to be 1000 mg/kg body weight/day.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

1 000 mg/kg bw/day

Study duration:

subchronic

Species:

rat

Quality of whole database:

Guideline study, no toxic effects observed at highest dose, NOAEL >/= 1000 mg/kg bw/day

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - local effects

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Oral exposure

The submission substance was tested in a combined repeated dose oral toxicity study with a reproduction / developmental toxicity screening test according to OECD guideline 422 (RL1). No adverse effects were observed up to the highest dose tested (1000 mg/kg bw/day) with respect to parental animals, reproduction and offspring. No toxic response was evident after exposure to doses up to 1000 mg/kg bw/day after a total exposure duration of 54 days for parental females, and there are no other indications of systemic toxicity in all other tests performed.

However, with regard to IUCLID endpoint 7.5.1 “Repeated Dose Toxicity”, there is no reliable standard sub-chronic toxicity study (90-day) available as required in REACH Annex IX columns 1 and 2, under section 8.6. Thus, a sub-chronic toxicity study (90-day, oral, rat) according to OECD 408 is proposed by the registrant to fill the data gap. 

 

Inhalation exposure

This information is not available. In accordance with column 2 of REACH Annexes VIII and IX, the repeated dose toxicity study, as required in section 8.6.1 of Annex VIII and in section 8.6.2 of Annex IX, does not need to use the inhalation route because exposure of humans via inhalation is considered unlikely taking into account the vapour pressure of the substance and/or the possibility of exposure to aerosols, particles or droplets of an inhalable size.

 

Dermal exposure

This information is not available.In accordance with column 2 of REACH Annexes VIII and IX, the repeated dose toxicity study, as required in section 8.6.1 of Annex VIII and in section 8.6.2 of Annex IX, does not need to use the dermal route because

- the physicochemical, toxicokinetic and toxicological properties do not suggest potential for a significant rate of absorption through the skin,

- based on a less reliable study (RL3), the toxicity after acute dermal application is not considered higher than that after acute oral administration,

- no systemic effects were observed in reliable skin irritation studies (RL2) as well as in an LLNA (RL1).

 

Justification for classification or non-classification

Based on the findings of a repeated dose guideline study, the submission substance has not to be classified for effects of repeated exposure according to Regulation (EC) No 1272/2008.