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

Repeated dose toxicity: oral

Administrative data

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:

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

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP study with no significant 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:

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

Deviations:

no

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:

UVCB substance
Description: Beige-brown crystalline powder with lumps
Batch: 278-123

Test animals

Species:

rat

Strain:

Wistar

Sex:

male/female

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.

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:

corn oil

Details on oral exposure:

Initial doses (0, 15, 50, 150 mg/kg respectively) were based on an estimated purity of 90%. After completion of the in-life phase the substance was classified as UVCB with a purity of 100%. Therefore, actual doses were 10% higher, i.e. 17, 55 and 165 mg/kg respectively.

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.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Analyses were conducted according to a validated method (Project 498817). 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 was considered acceptable if the mean measured concentrations were 85-115% of the target concentration. Homogeneity was demonstrated if the coefficient of variation was ≤ 10%. Formulations were considered stable if the relative difference before and after storage was maximally 10%.

Duration of treatment / exposure:

Males were exposed for 31 days, i.e. 2 weeks prior to mating, during mating, and up to the day prior to scheduled necropsy. Females were exposed for 41-52 days, i.e. during 2 weeks prior to mating, during mating, during post-coitum, and during at least 4 days of lactation (up to the day prior to scheduled necropsy). Female nos. 44 (Group 1), 65, (Group 3) and 77 (Group 4) were not dosed during littering.

Frequency of treatment:

Once daily

Doses / concentrations

Remarks:

Doses / Concentrations:
Oral gavage in corn oil
Basis:
actual ingested

No. of animals per sex per dose:

10

Details on study design:

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.

Positive control:

No

Examinations

Observations and examinations performed and frequency:

The following observations and examinations were evaluated: mortality / viability, clinical signs (daily),
functional observations and locomotor activity (Week 4 (males); end of lactation (females)), body
weight and food consumption (at least at weekly intervals), clinical pathology (Week 4 (males); end of
lactation (females)), macroscopy at termination, organ weights and histopathology on a selection of
tissues, and reproduction/developmental parameters, consisting 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.

Sacrifice and pathology:

All males and the selected 5 females/group (see Allocation) were deprived of food overnight (with a
maximum of approximately 20 hours) prior to planned necropsy, but water was provided. Non-selected
females were not deprived of food.

Animals surviving to scheduled necropsy and the moribund animal were deeply anaesthetized using
isoflurane (Abbott B.V., Hoofddorp, The Netherlands) and subsequently exsanguinated.
All animals were subjected to macroscopic examination of the cranial, thoracic and abdominal tissues
and organs, with special attention being paid to the reproductive organs. Descriptions of all
macroscopic abnormalities were recorded.
The numbers of former implantation sites and corpora lutea were recorded for all paired females.

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 (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 (many-to-one rank test) was applied if the data could not be assumed to
follow a normal distribution.
- The Fisher Exact-test was applied to frequency data.
The following additional methods of statistical analysis were used:
Motor activity data was subjected to the Kruskal-Wallis nonparametric ANOVA test to
determine intergroup differences followed by the Wilcoxon test to compare the treated groups
to the control group.
All tests were two-sided and in all cases p < 0.05 was accepted as the lowest level of significance.
Group means were calculated for continuous data and medians were calculated for discrete data
(scores) in the summary tables. Test statistics were calculated on the basis of exact values for means
and pooled variances. Individual values, means and standard deviations may have been rounded off
before printing. Therefore, two groups may display the same printed means for a given parameter, yet
display different test statistics values.

Results and discussion

Results of examinations

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:

not examined

Haematological findings:

no effects observed

Clinical biochemistry findings:

no effects observed

Urinalysis findings:

not examined

Behaviour (functional findings):

no effects observed

Description (incidence and severity):

The lower motor activity of females at 150 mg/kg was temporary 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

Organ weight findings including organ / body weight ratios:

no effects observed

Description (incidence and severity):

.

Gross pathological findings:

no effects observed

Description (incidence and severity):

Scattered effects but not apparent dose trend. See attached data tables.

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

Tubular crystals were noted in the kidneys of one male and one female at 150 mg/kg (up to slight degree). This finding was accompanied in the female by tubular dilatation, epithelial hypertrophy and granular cast(s).

Histopathological findings: neoplastic:

no effects observed

Details on results:

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.
Tubular crystals were noted in the kidneys of one male and one female at 150 mg/kg (up to slight
degree). Crystal deposition can lead to obstructive nephropathy. In this study, this finding was
accompanied in the female by tubular dilatation, epithelial hypertrophy and granular cast(s) (up to
slight degree) in the immediate vicinity as tubular crystals were noted, and is suggestive for findings
secondary to obstruction. Therefore, the presence of tubular crystals is considered to be adverse.
The increased severity of congestion/erythrophagocytosis in the mesenteric lymph nodes of females
treated at 15 mg/kg, 50 mg/kg and 150 mg/kg (up to moderate degree, and corresponding to dark red
foci/reddish discolouration of the mesenteric lymph node in some females at 15 and 150 mg/kg) was
considered to be non-adverse given the absence of any other indicators of cellular damage, e.g.
inflammation or necrosis. The increased incidence and severity of hyperplasia/hypertrophy of the
follicular epithelium of the thyroid gland in females treated at 150 mg/kg was subtle in nature and
occurred in the absence of a dose-related trend, and was therefore considered to be non-adverse.
There were no morphological findings in the reproductive organs of either sex which could be
attributed to the test item.
No toxicologically significant changes were noted in any of the remaining parental parameters
investigated in this study (i.e. clinical appearance, functional observations, body weight, food
consumption, clinical laboratory investigations, macroscopic examination and organ weights).

Effect levels

Dose descriptor:

NOAEL

Effect level:

55 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

histopathology: non-neoplastic

Target system / organ toxicity

Critical effects observed:

yes

Lowest effective dose / conc.:

150 mg/kg bw/day (actual dose received)

System:

urinary

Organ:

kidney

Treatment related:

yes

Any other information on results incl. tables

The concentrations analysed in the formulations of Group 2 (15 mg/kg) and Group 3 (50 mg/kg) were in agreement with target concentrations (i.e. mean accuracies between 90% and 110%). For the formulation of Group 4 (150 mg/kg)

prepared for use during treatment, the mean accuracy was below the target concentration (i.e. 89% of target).

No test substance was detected in the Group 1 (control, 0 mg/kg) formulation.

The formulation of Group 2 was homogeneous (i.e. coefficient of variation ≤ 10%). The formulation of Group 4 showed a coefficient of variation exceeding 10% (i.e. 16%) and was therefore not homogeneous. The level of inhomogeneity measured for Group 4 formulations was similar to that observed for procedural recovery samples prepared at a slightly higher concentration level.

Formulations appeared visually homogenous.

Formulations at the entire range were stable when stored at room temperature under normal laboratory light conditions for at least 6 hours.

Applicant's summary and conclusion

Conclusions:

Based on these results, the No Observed Adverse Effect Levels (NOAEL) for Parental animals is 50 mg/kg (55 mg/kg as UVCB)

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. This GLP study followed the OECD 422 guideline. Dose levels were based on the results of a 10-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. This was based on an estimated purity of 90%. After completion of the testing, the substance was determined to be UVCB with a purity of 100%. Based on the UVCB purity, the doses are considered to be 17, 55 and 165 mg/kg/day.

 

After acclimatization, four groups of ten male and ten female Wistar Han rats were exposed by oral gavage to the test substance. 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, duringpost-coitum, and during at least 4 days of lactation.

 

Results/discussion

 

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.

 

Tubular crystals were noted in the kidneys of one male and one female at 150 mg/kg (up to slight degree).Crystal deposition can lead to obstructive nephropathy. In this study, this finding was accompanied in the female by tubular dilatation, epithelial hypertrophy and granular cast(s) (up to slight degree) in the immediate vicinity as tubular crystals were noted, and is suggestive for findings secondary to obstruction. Therefore, the presence of tubular crystals is considered to be adverse.

 

The increased severity of congestion/erythrophagocytosis in the mesenteric lymph nodes of females treated at 15 mg/kg, 50 mg/kg and 150 mg/kg (up to moderate degree, and corresponding to dark red foci/reddish discolouration of the mesenteric lymph node in some females at 15 and 150 mg/kg) was considered to be non-adverse given the absence of any other indicators of cellular damage, e.g. inflammation or necrosis. The increased incidence and severity of hyperplasia/hypertrophy of the follicular epithelium of the thyroid gland in females treated at 150 mg/kg was subtile in nature and occurred in the absence of a dose-related trend, and was therefore considered to be non-adverse.

 

There were no morphological findings in the reproductive organs of either sex which could be attributed to the test item.

 

No toxicologically significant changes were noted in any of the remaining parental parameters investigated in this study (i.e. clinical appearance, functional observations, body weight, food consumption, clinical laboratory investigations, macroscopic examination and organ weights).

Based on these results, the repeat dose No Observed Adverse Effect Level (NOAEL) is 50 mg/kg (55 mg/kg - as UVCB).