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Toxicological information

Developmental toxicity / teratogenicity

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Administrative data

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
weight of evidence
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
data from handbook or collection of data
Justification for type of information:
Data is from peer reviewed journal

Data source

Reference
Reference Type:
publication
Title:
A TERATOGENICITY STUDY OF PHLOXINE B IN ICR MICE
Author:
M. SENO, S. FUKUDA and H. UMISA
Year:
1984
Bibliographic source:
Fd Chem. Toxic. Vol. 22, No. 1 pp. 55-60, 1984

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
other: as below
Principles of method if other than guideline:
In teratogenicity study the toxic effect of PHLOXINE B was observed in mice during 6-16 days of gestation.
GLP compliance:
not specified
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
3,4,5,6-tetrachloro-2-(1,4,5,8-tetrabromo-6-hydroxy-3-oxoxanthen-9-yl)benzoic acid
EC Number:
242-355-6
EC Name:
3,4,5,6-tetrachloro-2-(1,4,5,8-tetrabromo-6-hydroxy-3-oxoxanthen-9-yl)benzoic acid
Cas Number:
18472-87-2
Molecular formula:
C20H4Br4Cl4O5.2Na
IUPAC Name:
2',4',5',7'-tetrabromo-4,5,6,7-tetrachloro-3',6'-dihydroxy-3H-spiro[2-benzofuran-1,9'-xanthen]-3-one
Test material form:
solid: particulate/powder
Details on test material:
- Name of test material (as cited in study report):PHLOXINE B
- Molecular formula :C20H4Br4Cl4O5.2Na
- Molecular weight : 829.64 g/mol
- Substance type: Organic
- Physical state:Red-brown powder
- Impurities (identity and concentrations): < 5 ( weight loss on drying < 10%; chloride and sulphate < 5% )
Specific details on test material used for the study:
- Name of test material (as cited in study report):PHLOXINE B
- Molecular formula :C20H4Br4Cl4O5.2Na
- Molecular weight : 829.64 g/mol
- Substance type: Organic
- Physical state:Red-brown powder
- Impurities (identity and concentrations): < 5 ( weight loss on drying < 10%; chloride and sulphate < 5% )

Test animals

Species:
mouse
Strain:
other: Jcl:ICR
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Nippon CLEA Co., Ltd, Tokyo.
- Sex: Male and female
- Age at study initiation: 8-9 wk old
- Weight at study initiation: 26--31 g- female, male - No data available.
.- Fasting period before study: No data available.
- Housing: Males were housed singly and females four to a cage.
- Diet (e.g. ad libitum): commercial diet (Nippon CLEA, CE-2) ad lib.
- Water (e.g. ad libitum): tap-water ad lib.
- Acclimation period: 1 week.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 23 + I°C
- Humidity (%):55 + 5%
- Air changes (per hr): No data available.
- Photoperiod (hrs dark / hrs light): 12-hr light/dark cycle

IN-LIFE DATES: From: To No data available.

Administration / exposure

Route of administration:
oral: feed
Vehicle:
other: Nippon CLEA, CE-2 commercial diet
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: No data available

DIET PREPARATION
- Rate of preparation of diet (frequency): No data available
- Mixing appropriate amounts with (Type of food): Nippon CLEA, CE-2 commercial diet
- Storage temperature of food: No data available

VEHICLE
- Justification for use and choice of vehicle (if other than water): No data available
- Concentration in vehicle: 0, 1.0, 3.0 and 5.0%
- Amount of vehicle (if gavage): No data available
- Lot/batch no. (if required): No data available
- Purity: No data available
Analytical verification of doses or concentrations:
not specified
Details on mating procedure:
- M/F ratio per cage: 1:1
- Length of cohabitation: Overnight.
- Proof of pregnancy: [vaginal plug / sperm in vaginal smear] referred to as [day 0 / day 1] of pregnancy Females with vaginal plugs were considered to be in day 0 of pregnancy.
- After … days of unsuccessful pairing replacement of first male by another male with proven fertility. No data available
- Further matings after two unsuccessful attempts: [no / yes (explain)] No data available
- After successful mating each pregnant female was caged (how): Females were housed individually.
- Any other deviations from standard protocol: No data available
Duration of treatment / exposure:
11 day
Frequency of treatment:
Daily
Duration of test:
Upto day 18 of gestation
Doses / concentrations
Remarks:
0, 2000,6000 and 10000 mg/kg/day (0, 1.0, 3.0 and 5.0 %)
No. of animals per sex per dose:
Total No – 84 Female
0 % (0mg/kg/day)22 females
1%(2000 mg/kg/day)21 females
3%(6000 mg/kg/day)20 females
5% (10000 mg/kg/day)21 females
Control animals:
yes, concurrent vehicle
Details on study design:
Further details on study design
- Dose selection rationale: No data available
- Rationale for animal assignment (if not random): Pregnant dams were divided into experimental and control groups such that body-weight differences between groups were minimized.
- Other: No data available

Examinations

Maternal examinations:
Survival, clinical sign, body weight, food consumption and food efficiency, Absolute and Relative organ weight were observed.
Ovaries and uterine content:
Uterine horn count, number of females with live fetuses, corpora lutea, implantations, resorptions, number of dead fetuses and total resorptions were examined.
Fetal examinations:
Foetal body weight, soft tissue abnormalities and skeletal anomalies were observed.
Statistics:
The litter was considered to be the experimental unit for the analysis of data on embryo/foetal toxicity and teratogenicity. Statistical significance of the differences between groups was determined by the Mann-Whitney U test (Siegel, 1956). Two-tailed tests were performed and P < 0.05 was selected as the level of statistical significance.
Indices:
Number of females with live fetuses, resorptions, Number of dead fetuses and total resorptions were examined.
Historical control data:
No data available

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:
not specified
Dermal irritation (if dermal study):
not specified
Mortality:
mortality observed, treatment-related
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
not specified
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
not specified
Haematological findings:
not specified
Clinical biochemistry findings:
not specified
Urinalysis findings:
not specified
Behaviour (functional findings):
not specified
Immunological findings:
not specified
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
not specified
Neuropathological findings:
not specified
Histopathological findings: non-neoplastic:
not specified
Histopathological findings: neoplastic:
not specified
Other effects:
not specified

Maternal developmental toxicity

Number of abortions:
not specified
Pre- and post-implantation loss:
not specified
Total litter losses by resorption:
no effects observed
Early or late resorptions:
not specified
Dead fetuses:
no effects observed
Changes in pregnancy duration:
not specified
Description (incidence and severity):
Migrated Data from removed field(s)
Field "Effects on pregnancy duration" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsMaternalAnimals.MaternalDevelopmentalToxicity.EffectsOnPregnancyDuration): not specified
Changes in number of pregnant:
not specified
Other effects:
not specified
Details on maternal toxic effects:
Maternal toxic effects:yes

Details on maternal toxic effects:
Mortality:
In 10,000 mg/kgbw/day, 2 dams died on days 16 and 17. Another female in this group
aborted on day 17.

These three animals were excluded from all computations.

Clinical signs:
No overt signs of toxicity were observed in treated dams.

Body weight:
Significantly decreased in Maternal body-weight gains for days 6-16 of gestation were observed in 2000, 6000 and 10,000 mg/kg bw/day treated dams as compared to control.

Food consumption:
Average food intakes of the groups were approximately equal both in treated and control group.

Reproductive performance:
The numbers of corpora lutea, implantations or live foetuses in all of the treated groups were slightly decreased, but not significantly in comparison with those of the control.

No effect were observed on Number of dead fetuses, Total resorptions and Number of live fetuses of treated rats as compared to control.

Organ weights:
In 10,000 mg/kgbw/day treated dams, absolute and relative liver weight were significantly incerased as compared to control.

Effect levels (maternal animals)

Dose descriptor:
NOAEL
Effect level:
2 000 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
other: maternal toxicity

Maternal abnormalities

Abnormalities:
not specified

Results (fetuses)

Fetal body weight changes:
not specified
Description (incidence and severity):
Migrated Data from removed field(s)
Field "Fetal/pup body weight changes" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsFetuses.FetalPupBodyWeightChanges): not specified
Reduction in number of live offspring:
not specified
Changes in sex ratio:
not specified
Changes in litter size and weights:
not specified
Changes in postnatal survival:
not specified
External malformations:
effects observed, treatment-related
Skeletal malformations:
effects observed, treatment-related
Visceral malformations:
not specified
Other effects:
not specified
Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:yes

Details on embryotoxic / teratogenic effects:
Mortality:
No effect were observed on number of live fetuses in treared rats

Clinical signs:
No data available

Gross pathology:
Foetuses with open eyelids were observed in all treated dams.

Exencephaly, significantly increased incidence of cleft palate were observed in 10,000 mg/kgbw/day treated fetuses as compaed to control.

These types of external abnormalities have been seen spontaneously in this strain.

Histopathology:
Significantly decreased numbers of ossified caudal vertebrae and phalanges, slightly reduced incidence of accessory sternebrae and significantly increased numbers of foetuses with a fourteenth rib or with an extra rib (at least one fourteenth rib that was half or greater than half the length of the preceding rib)
were observed in 10,000 mg/kgbw/day dose group.

Increase in fourteenth or extra ribs might be taken as an indicator of the possible teratogenic potential of a chemical.

Slight retardation of ossification were observed in 6000 mg/kgbw/day treaed fetuses as compared to control.

Splitting of the cervical vertebral arches were observed in 1000 mg/kg bw/day treatet fetouses.

Effect levels (fetuses)

Key result
Dose descriptor:
LOAEL
Effect level:
2 000 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
not specified
Basis for effect level:
other: Adverse effect on gross pathology and histopathology

Fetal abnormalities

Abnormalities:
not specified

Overall developmental toxicity

Developmental effects observed:
yes
Lowest effective dose / conc.:
2 000 mg/kg bw/day
Treatment related:
not specified
Relation to maternal toxicity:
not specified
Dose response relationship:
not specified
Relevant for humans:
not specified

Applicant's summary and conclusion

Conclusions:
NOAEL was considered to be 2000 mg/kg body weight/day (1%) for F0 generation and LOAEL was considered to be 2000 mg/kg body weight/day (1%) for F1 genration when Jcl:ICR female mice treated with Phloxine B.
Executive summary:

In a one generation Teratogenicity study, Jcl:ICR female mice treated with Phloxine B in the concentration of 0, 2000,6000 and 10000 mg/kg/day (0, 1.0, 3.0 and 5.0 %)from the morning of day 6 through day 16 of pregnancy by oral feed.2 dams died ondays 16 and 17. Another female in this group aborted on day 17.Significant decreased inbody-weight gains form days 6-16 of gestation wereobserved in 2000, 6000 and 10,000 mg/kg bw/day treated dams as compared to control. Similarly,The numbers of corpora lutea, implantations or live foetuses in all of the treated groups were slightly decreased, but not significantly in comparison with the control. Absolute and relative liver weight were significantly incerased in 2000 mg/kg bw/day dose group as compared to control in F0 generation. In addition,Foetuses with open eyelidswere observed in all treated dams. Exencephaly, significantlyincreased incidence of cleft palate and Significantly decreased numbers of ossified caudal vertebrae and phalanges, slightly reduced incidence of accessory sternebrae and significantly increased numbers of foetuses with a fourteenth rib or with an extra rib (at least one fourteenth rib that was half or greater than half the length of the preceding rib) were observed in 10,000 mg/kgbw/day treated fetuses as compaed to control. Slight retardation of ossification were observed in 6000 mg/kgbw/day treaed fetuses as compared to control. Splitting of the cervical vertebral arches were observed in 1000 mg/kg bw/day treated fetouses as compared to control. Hencse, dose response was seen in the total incidence of skeletal abnormalities,suggested a teratogenic effect.

Therefore, NOAEL was considered to be 2000 mg/kg body weight/day (1%) for F0 generation and LOAELwas considered to be 2000 mg/kg body weight/day (1%) forF1 genrationwhen Jcl:ICR female mice were treated with Phloxine B orally by feed from day 6 to 16 of gestation.