Strontium 4-[(5-chloro-4-methyl-2-sulphonatophenyl)azo]-3-hydroxy-2-naphthoate (1:1)

Toxicity to reproduction

Administrative data

Endpoint:

screening for reproductive / developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1993

Report date:

1993

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Combined Repeated Dose and Reproductive / Developmental Toxicity Screening Test (Precursor Protocol of GL 422)

Deviations:

no

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Calcium 3-hydroxy-4-[(4-methyl-2-sulphonatophenyl)azo]-2-naphthoate

EC Number:

226-109-5

EC Name:

Calcium 3-hydroxy-4-[(4-methyl-2-sulphonatophenyl)azo]-2-naphthoate

Cas Number:

5281-04-9

Molecular formula:

C18H14N2O6S.Ca

IUPAC Name:

calcium 3-hydroxy-4-[(4-methyl-2-sulfonatophenyl)diazenyl]-2-naphthoate

Test material form:

solid: nanoform

Details on test material:

- Purity: approximately 98% [ Containing as impurities : NaCl, CaCl2, Ca(OH)2, and water ]
- Lot/Batch No : T-1322-2
- Storage condition of test material : Stored in cold-dark conditions
- Stability in the vehicle: Stable for 4 hours in the vehicle at the concentrations ranging between 0.925 and 50.0 mg/ml, according to the stability test performed by the laboratory.

Test materials used in this dossier are all considered to fall under the definition of nano-materials according to the European Commission Recommendation 2011/696/EU as the synthesis and manufacturing of this pigment always yields particulate material with a fine particle size distribution.

Specific details on test material used for the study:

-Analytical purity : 98 % w/w
-Lot/batch No.: T-1322-2
-Appearance : Red powder

Test animals

Species:

rat

Strain:

Crj: CD(SD)

Sex:

male/female

Details on test animals or test system and environmental conditions:

- Sources:Charles River Laboratories Japan (Atsugi Breeding Center)
- Age at study initiation : 7 weeks
- Weight at study initiation : females 196.5-227.4 g, males: 235.5-299.6 g
- Housing : Individually housed in stainlelss suspended cages
- Diet : CA-1 (CLEA Japan, Inc.) ad libitum
- Water : Tap water, ad libitum
- Acclimation period : 1 week for quarantine and acclimaition

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

other: 5% gum arabic solution

Details on mating procedure:

- M/F ratio per cage: 1:1

Analytical verification of doses or concentrations:

yes

Duration of treatment / exposure:

Males: Total of 42 days ( 14 days before mating, 14 days for mating, and 14 days after mating)
Females Total of 41-50 days (14 days before mating, during mating, pregnancy and up to lactation day 3)

Frequency of treatment:

daily

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

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

Doses / concentrations 2

Dose / conc.:

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

Doses / concentrations 3

Dose / conc.:

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

No. of animals per sex per dose:

13/sex/dose

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: Doses were selected on the basis of a 14-day preliminary repeat dose study at 1000 mg/kg bw/day. No deaths and clinical signs observed. 3 cases of necrosis or regeneration of tubular epithelium were seen in males.

Positive control:

not used

Examinations

Parental animals: Observations and examinations:

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Males and Females - on each administration day.

BODY WEIGHT: Yes
- Time schedule for examinations: Males - measured once a week on administration days 0, 7, 14, 21, 28, 35, and 42; Females - measured once a week on administration days 0, 7, 14, and 21, during pregnancy on days 0, 7, 14, and 20, during lactation on day 0 and 4.

FOOD CONSUMPTION AND COMPOUND INTAKE :
- Food consumption for each animal determined: Yes
- Time schedule for examinations : Males - measured once a week on administration days 0, 7, 14, 21, 28, 35, and 42; Females - measured once a week on administration days 0, 7, 14, and 21, during pregnancy on days 0, 7, 14, and 20, during lactation on day 0 and 4.

HAEMATOLOGY: Yes
- Time schedule for collection of blood: Males - day after final administration day
- Anaesthetic used for blood collection: Yes (pentobarbital)
- Animals fasted: No data
- How many animals: 13 animals per male per group
- Parameters checked in table 1 were examined.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Males - day after final administration day
- Anaesthetic used for blood collection: Yes (pentobarbital)
- Animals fasted: No data
- How many animals : 13 animals per male per group
- Parameters checked in table 2 were examined.

Litter observations:

STANDARDISATION OF LITTERS
not applicable


PARAMETERS EXAMINED
The following parameters were examined in offspring: number and sex of pups, stillbirths, live births, postnatal mortality, presence of gross anomalies, weight gain, physical or behavioural abnormalities


GROSS EXAMINATION OF DEAD PUPS: yes, for external and internal abnormalities; possible cause of death was not determined for pups born or found dead.

Postmortem examinations (parental animals):

SACRIFICE
- Male animals: All surviving animals
- Maternal animals: All surviving animals


GROSS NECROPSY
- yes, see entry for repeated dose toxicity for details


HISTOPATHOLOGY / ORGAN WEIGHTS
- yes, see entry for repeated dose toxicity for details

Postmortem examinations (offspring):

SACRIFICE
- The F1 offspring was sacrificed at 4 days of age.


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

Statistics:

x squared test was conducted for copulation and fertility indices. For all the other parameters, Bartlett's test was used to examine uniformity of distribution in each group, and one-way statistical analysis was performed when the distribution is uniform. Where significant differences were observed, Duneett's test or Scheff's test was performed to examine differences in averages between each test group and control groups. Kruskal-Wallis's test was conducted when the distribution was not uniform. Significant testing was carried out at the 5% and the 1% levels.

Reproductive indices:

yes

Offspring viability indices:

yes

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

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:

effects observed, treatment-related

Description (incidence and severity):

Males - 1 day after termination of administration period: Significantly lower values of MCH (p<0.05) were seen in the 300 and 1000 mg/kg bw/day groups compared to the controls. Dose-dependent decrease in WBC was seen in the the 300 and 1000 mg/kg bw/day groups, but the effects were not significant. In the absence of historical control data, no corresponding histopathology findings and other affected parameters, the findings are considered of no biological relevance.

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

Males - 1 day after termination of administration period: Significantly lower levels of inorganic phosphorus and Calcium were seen in the 300 mg/kg bw/day group. Significant low levels of Total cholesterol and pottasium and significant high values of Chloride and GOT were noted in the 1000 mg/kg bw/day group.

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

MALES - 1 day after termination of administration period:
Thymus: Two cases of hemorrhage were seen in the 1000 mg/kg bw/day group, one case in the control group. In the absence of a dose-response relationship, this is considered incidental.
Heart: Small areas of myocardial degeneration were seen in 2 cases in the control group. No other abnormalities were observed.
Liver: No treatment-related findings were observed. Ten cases of very slight microgranuloma were seen respectively in the control and the 1000 mg/kg bw/day groups. Slight fatty changes in peripheral zone were seen in a case of the control group and in four cases of the 1000 mg/kg bw/day group. Therefore, there was no significant difference in severity and incidence of the findings between the control and the 1000 mg/kg bw/day groups. A case of very slight focal necrosis was seen in the 1000 mg/kg bw/day group.
Kidney: Large numbers of regenerating tubular epithelium were observed in three cases in the 300 mg/kg bw/day group and twelve cases in the 1000 mg/kg bw/day group compared to the control groups. The severity was augmented in the 1000 mg/kg bw/day group. The regenerating epithelial cells were found predominantly in convoluted proximal tubules, showing increased cell density, slightly enlarged nucleoli, and slightly bright or basophilic cytoplasm. In most cases, slightly-yellowish debris was noted in the tubular lumen. A single case of cast in tubular lumen was found in the 100 mg/kg bw/day group. In all groups, including the control group, eosinophilic bodies were observed with no significant differences in incidence and severity among the groups.
Adrenal cortex : Brown pigment deposits were observed both in the 1000 mg/kg bw/day and the control groups but there is no significant difference in incidence and severity.
Spleen: Brown pigment deposits and extramedullary hematopoiesis were found both in the 1000 mg/kg bw/day and the control groups, with no significant difference in incidence and severity.
Testis: Atrophy of tubule was found in 2 cases in the control group and 3 cases in the 1000 mg/kg bw/day group. One of the cases in the 1000 mg/kg bw/day group had calcification in the tubule.
Epididymis: Decreased numbers of sperm were noted in each one case in the control and the 1000 mg/kg bw/day groups. Both animals had atrophy of the tubule.

FEMALES
Thymus: A case of very slight and a case of slight involution were found in the control group. In the 1000 mg/kg bw/day group, increased numbers of involution were found with a case rated "very slight", 3 cases rated "slight", and 2 cases rated "moderately slight".
Liver: Very slight microgranuloma was seen in a case of the control and 2 cases of the 1000 mg/kg bw/day group sacrificed at Day 4 of lactation, and one non-pregnant animal in the control group sacrificed at Day 25 of pregnancy. Very slight to slight fatty changes in peripheral zone were seen in 3 cases in the control group, and very slight to moderate fatty changes in 2 cases in the 1000 mg/kg bw/day group. There were no significant differences in severity and incidence between the control and the 1000 mg/kg bw/day groups.
Kidney: In all treatment groups sacrificed at Day 4 of lactation, increased numbers of incidences of regenerating tubular epithelium were observed. The findings are accompanied with foamy epithelial cells and vacuolar degeneration predominantly in convoluted proximal tubule. In many cases, necrotized epithelial cells were noted, and eosinophilic necrotized cells and yellowish debris were contained in the tubular lumen. In tubular basement of these animals, regenerating epithelial cells included large-sized basophilic cytoplasma. The incidence was similar in all dose groups; the severity of this lesion was slightly increased in the high-dose group.
In all dose groups, there were some animals completely free of kidney findings. Among those were a non-delivering animal (total implantation loss) in the 100 mg/kg bw/day group, 2 non-pregmant animals in the 300 mg/kg bw/day group, and a non-copulated animal in the 1000 mg/kg bw/day group.

Other kidney findings that are considered incidental included each one case of cast in the tubular lumen in the control and the 1000 mg/kg bw/day groups, each one case of slight vacuolar degeneration in the control and the 100 mg/kg bw/day groups, and each one case of focal dilatation of tubule in the 100 mg/kg bw/day and the 1000 mg/kg bw/day groups. The case of cast of the animal in the control group was considered be associated with the very slight incidence of chronic nephropathy.

Adrenal cortex: A case of very slight brown pigment deposit and a case of focal necrosis were observed in the 1000 mg/kg bw/day group.
Spleen: Brown pigment deposits and extramedullary hematopoiesis were observed in 11 animals in the control group and 12 animals in the 1000 mg/kg bw/day group. Two cases were also found in non-pregnant animals in the control group, sacrificed at Day 25 of pregnancy. There were no clear differences in incidence and severity between the treated and the control groups.
Ovary: No abnomarities were found with non-pregnant animals (two in the control and two in the 300 mg/kg bw/day groups) and in the non-copulated animal (one in the 1000 mg/kg bw/day group).

Histopathological findings: neoplastic:

no effects observed

Effect levels (P0)

Dose descriptor:

NOEL

Effect level:

< 100 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: Adverse effects on kidneys of parental animals observed at all dose groups.

Target system / organ toxicity (P0)

Critical effects observed:

yes

Lowest effective dose / conc.:

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

System:

urinary

Organ:

kidney

Treatment related:

yes

Results: F1 generation

Details on results (F1)

No treatment-related adverse effects on the offspring was observed.

Effect levels (F1)

Dose descriptor:

NOAEL

Generation:

F1

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 on offspring observed

Target system / organ toxicity (F1)

Critical effects observed:

no

Overall reproductive toxicity

Reproductive effects observed:

no