Naphthalenesulfonic acid, di-C9-rich C8-10-branched alkyl derivs., barium salts

Developmental toxicity / teratogenicity

Administrative data

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

07 March 2012 to 11 June 2013 (end of in-life phase)

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP study without deficiencies

Cross-reference

Reason / purpose for cross-reference:

reference to other study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2013

Report date:

2013

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

other: OECD 422

Deviations:

yes

Remarks:

Minor deviations occurred. The study integrity was not adversely affected by the deviations.

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

barium bis(di C8-C10, branched, C9 rich, alkylnaphthalenesulphonate)

EC Number:

939-718-2

Cas Number:

1474044-80-8

Molecular formula:

C28H44O3S.1/2Ba

IUPAC Name:

barium bis(di C8-C10, branched, C9 rich, alkylnaphthalenesulphonate)

Test material form:

solid: particulate/powder

Remarks:

migrated information: powder

Details on test material:

Description: Beige-brown crystalline powder with lumps
Batch: 278-123
Purity: UVCB (treated as 100% pure)

Test animals

Species:

rat

Strain:

Wistar

Details on test animals or test system and environmental conditions:

Source F0: Charles River Deutschland, Sulzfeld, Germany.
Age at start F0-treatment: Approximately 11 weeks.
Number of F0-animals: 40 females and 40 males.
Acclimatization F0: At least 5 days prior to start of treatment.
Health inspection F0: Upon receipt of the animals.
Identification F0: Earmark and tattoo.
Randomization F0: Prior to commencement of treatment, by computer-generated random algorithm according to body weight, with all animals within ± 20% of the sex mean.

Environmental controls for the animal room were set to maintain 18 to 24°C, a relative humidity of 40 to 70%, approximately 15 room air changes/hour, and a 12-hour light/12-hour dark cycle. Any variations to these conditions were maintained in the raw data and had no effect on the outcome of the study.

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

unchanged (no vehicle)

Details on exposure:

Method: Oral gavage, using a plastic feeding tube. Formulations were placed on a magnetic stirrer during dosing.
Dose volume: 5 mL/kg body weight. Actual dose volumes were calculated according to the latest body weight.
Frequency: Once daily for 7 days per week, approximately the same time each day with a maximum of 6 hours difference between the earliest and latest dose. Animals were dosed up to the day prior to scheduled necropsy.
Males were exposed for 29 days, i.e. 2 weeks prior to mating, during mating, and up to termination.
Females were exposed for 42-55 days, i.e. during 2 weeks prior to mating, during mating, during
post-coitum, and during at least 4 days of lactation.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Analyses were conducted according to a validated method. These analyses were conducted after the in-life phase as no suitable analytical method was available at an earlier stage. Samples of formulations were analyzed for homogeneity (highest and lowest concentration) and accuracy of preparation (all concentrations). Stability in vehicle over 6 hours at room temperature was also determined (highest and lowest concentration).

The accuracy of preparation of the formulation at 15 and 50 mg/kg (Groups 2 and 3,
respectively) was within specifications (i.e. mean accuracies between 90% and 110%), and the
formulation at 15 mg/kg was homogeneous. The mean accuracy of preparation of the 150 mg/kg
formulation (Group 4) was slightly outside specifications (i.e. 89% vs. an accepted mean accuracy
between 90% and 110%). Although formulations appeared visually homogenous, the formulation at
150 mg/kg was not homogenous analytically (coefficient of variation: 16% vs. an accepted coefficient
of variation of 10%). It cannot be excluded that the actual dose administered to Group 4 animals was
variable over the treatment period. However, the level of inhomogeneity of Group 4 formulations was
similar to that observed for procedural recovery samples prepared at a slightly higher concentration
level. the mean accuracy of preparation was only 1% below the target range, and the purity data
supplied by the sponsor after completion of the in-life phase indicated that purity was 10% higher than
accounted for when preparing formulations. Therefore, the mean dose administered over the
treatment period was considered to be sufficiently close to the target dose of 150 mg/kg to allow a
valid conclusion to be drawn from the results.

Details on mating procedure:

Following a minimum of 14 days of exposure for the males and females, one female was cohabitated with one male of the same treatment group, avoiding sibling mating. Detection of mating was confirmed by evidence of sperm in the vaginal lavage or by the appearance of an intravaginal copulatory plug. This day was designated Day 0 post-coitum. Once mating occurred, the males and females were separated. A maximum of 14 days was allowed for mating, after which females who had not shown evidence of mating were separated from their males. Female nos. 74 and 75 were re-mated with a proven male of the same dose group since male nos. 34 and 35 were sacrificed in extremis on Day 5 and 2 of the mating period respectively.

Duration of treatment / exposure:

After acclimatization, four groups of ten male and ten female Wistar Han rats were exposed by oral
gavage to the test substance at 0, 15, 50 and 150 mg/kg. Males were exposed for 29 days, i.e. 2
weeks prior to mating, during mating, and up to termination. Females were exposed for 42-55 days,
i.e. during 2 weeks prior to mating, during mating, during post-coitum, and during at least 4 days of
lactation.

Frequency of treatment:

Rat were dosed daily. Males were exposed for 29 days, i.e. 2 weeks prior to mating, during mating, and up to termination.
Females were exposed for 42-55 days, i.e. during 2 weeks prior to mating, during mating, during post-coitum, and during at least 4 days of
lactation.

No. of animals per sex per dose:

10 per sex per dose level including control

Examinations

Maternal examinations:

Mortality / Viability:
At least twice daily. Animals showing pain, distress or discomfort, which was considered not transient in nature or was likely to become more severe, were sacrificed for humane reasons based on OECD guidance document on humane endpoints (ENV/JM/MONO/ 2000/7).

Clinical signs:
Daily detailed clinical observations were made in all animals immediately (0-15 min) after dosing. Once prior to start of treatment and at weekly intervals during the treatment period this was also performed outside the home cage in a standard arena.
The time of onset, grade and duration of any observed sign was recorded. Signs were graded for severity and the maximum grade was predefined at 3 or 4. Grades were coded as slight (grade 1), moderate (grade 2), severe (grade 3) and very severe (grade 4). For certain signs, only its presence (grade 1) or absence (grade 0) was scored. In the data tables, the scored grades were reported, as well as the percentage of animals affected in summary tables.

Ovaries and uterine content:

Pregnant females were examined to detect signs of difficult or prolonged parturition, and cage debris of pregnant females was examined to detect signs of abortion or premature birth.

Fetal examinations:

Each litter was examined to determine the following, if practically possible -

Mortality / Viability: The numbers of live and dead pups on Day 1 of lactation and daily thereafter were determined. If possible, defects or cause of death were evaluated.
Clinical signs: At least once daily, detailed clinical observations were made for all animals.
Body weights: Live pups were weighed on Days 1 and 4 of lactation.
Pups surviving to planned termination were killed by decapitation on Days 5-7 of lactation.

All pups were sexed and descriptions of all external abnormalities were recorded. The stomach was examined for the presence of milk. If possible, defects or cause of death were evaluated.

Statistics:

The following statistical methods were used to analyze the data:
- If the variables could be assumed to follow a normal distribution, the Dunnett-test (Ref. 2; many-to-one t-test) based on a pooled variance estimate was applied for the comparison of the treated groups and the control groups for each sex.
- The Steel-test (Ref. 3; many-to-one rank test) was applied if the data could not be assumed to follow a normal distribution.
- The Fisher Exact-test (Ref. 4) was applied to frequency data.

Indices:

see table attached.

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:

Maternal toxic effects:no effects

Details on maternal toxic effects:
The lower motor activity of females at 150 mg/kg was temporary in nature since at commencement
and the end of the measurement interval motor activity appeared similar to control levels. Moreover,
the lower motor activity was not associated with concurrent changes in clinical appearance or changes
during functional observation tests. Therefore, this change in motor activity was considered to be of no
toxicological relevance.

Effect levels (maternal animals)

Dose descriptor:

NOAEL

Effect level:

> 150 mg/kg bw/day (nominal)

Based on:

test mat.

Basis for effect level:

other: developmental toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
No toxicologically significant changes were noted in any of the developmental parameters investigated
in this study (i.e. gestation index and duration, parturition, maternal care and early postnatal pup
development consisting of mortality, clinical signs, body weight and macroscopy) up to the highest
dose level tested (150 mg/kg).

Effect levels (fetuses)

Remarks on result:

not determinable due to absence of adverse toxic effects

Remarks:

Embryotoxic / teratogenic effects:no effects Details on embryotoxic / teratogenic effects: No toxicologically significant changes were noted in any of the developmental parameters investigated in this study (i.e. gestation index and duration, parturition, maternal care and early postnatal pup development consisting of mortality, clinical signs, body weight and macroscopy) up to the highest dose level tested (150 mg/kg).

Fetal abnormalities

Abnormalities:

not specified

Overall developmental toxicity

Developmental effects observed:

not specified

Applicant's summary and conclusion

Conclusions:

No toxicologically significant changes were noted in any of the developmental parameters investigated
in this study (i.e. gestation index and duration, parturition, maternal care and early postnatal pup
development consisting of mortality, clinical signs, body weight and macroscopy) up to the highest
dose level tested (150 mg/kg).

Executive summary:

 

A Combined 28-day repeated dose toxicity study with the reproduction/developmental toxicity screening test was conducted with Barium bis( di c8-c10, branched, c9 rich, alkylnaphthalene sulphonate) in rats by oral gavage. The study was based on the OECD 422, Combined Repeated Dose Toxicity Study with the Reproduction/Developmental Toxicity Screening Test. Based on the results of a 15-day dose range finding study, the dose levels for this combined 28-day oral gavage study with reproduction/developmental toxicity screening test were selected to be 15, 50 and 150 mg/kg.

 

After acclimatization, four groups of ten male and ten female Wistar Han rats were exposed by oral gavage to the test substance at 0, 15, 50 and 150 mg/kg. Males were exposed for 29 days, i.e. 2 weeks prior to mating, during mating, and up to termination. Females were exposed for 42-55 days, i.e. during 2 weeks prior to mating, during mating, during post-coitum, and during at least 4 days of

lactation.

 

Evaluated reproduction/developmental parameters consisted of mating, fertility and conception indices, precoital time, number of corpora lutea and implantation sites, gestation index and duration, parturition, maternal care, sex ratio and early postnatal pup development (mortality, clinical signs,

body weights and macroscopy). Formulations were analyzed once during the study to assess accuracy, homogeneity and stability.

 

Results/discussion

 

Formulation analysis showed that the formulations were stable for at least 6 hours at room temperature and that dose levels were within planned limits.

 

Parental results:

There were no observable signs of maternal toxicity.The lower motor activity of females at 150 mg/kg was temporary in nature since at commencement and the end of the measurement interval motor activity appeared similar to control levels. Moreover, the lower motor activity was not associated with concurrent changes in clinical appearance or changes during functional observation tests. Therefore, this change in motor activity was considered to be of no toxicological relevance. Parental toxicity at 150 mg/kg, consisting of histopathological findings in the kidneys of the males and females is described in the endpoint study record -Repeat dose toxicity: oral.001.

 

Reproductive results:

No toxicologically significant changes were noted in any of the reproductive parameters investigated in this study (i.e. mating, fertility and conception indices, precoital time, and numbers of corpora lutea and implantation sites) up to the highest dose level tested (150 mg/kg).

 

Developmental results:

No toxicologically significant changes were noted in any of the developmental parameters investigated in this study (i.e. gestation index and duration, parturition, maternal care and early postnatal pup development consisting of mortality, clinical signs, body weight and macroscopy) up to the highest dose level tested (150 mg/kg).

 

Conclusion

Treatment of male and female Wistar Han rats with Barium bis( di c8-c10, branched, c9 rich, alkylnaphthalene sulphonate) by oral gavage at dose levels of 15, 50 and 150 mg/kg revealed parental toxicity at 150 mg/kg, consisting of histopathological findings in the kidneys of the males and females. No reproduction and developmental toxicity was observed for treatment up to 150 mg/kg.

 

Based on these results, the following No Observed Adverse Effect Levels (NOAEL) were derived:

Parental NOAEL: 50 mg/kg (55 mg/kg*)

Reproduction NOAEL: at least 150 mg/kg*

Developmental NOAEL: at least 150 mg/kg*

 

*Information supplied by the sponsor after completion of the in-life phase indicated that the purity of the test substance was

100% instead of an estimated purity of 90%. Although for 150 mg/kg formulations this would result in an actual dose of 165

mg/kg, formulation analyses showed that accuracy of preparation of was approximately 10% below target.