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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:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment

Data source

Reference
Reference Type:
publication
Title:
Unnamed
Year:
1974

Materials and methods

Test guideline
Qualifier:
no guideline followed
Principles of method if other than guideline:
The purpose of this study was to evaluate the teratogenic potential of linear alkylbenzene sulphonate (LAS) in rats. Twenty female rats/dose were divided into 5 groups and administered 0 (water), 0.2, 2.0, 300 and 600 mg/kg bw/day of test substance by oral gavage from day 6 to 15 of gestation. Throughout experiment animals were observed daily for signs of malreaction and weighed regularly. After euthanasia on day 20, their uterine content was examined for number of implantations, viable young and embryonic deaths along with ovaries, number of corpora lutea and viable pups. Body weight, incidents of external malformation, visceral and skeletal anomalies were also recorded in pups. Non parametric methods were used for statistical analysis and on the basis of adverse effects, NOAEL value was derived for maternal toxicity and teratogenicity.
GLP compliance:
not specified
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
-Name of test material (as cited in article): LAS
-Supplied by Lion Fat & Oil Co., Tokyo, Japan.
No further details were provided in article

Test animals

Species:
rat
Strain:
other: CD rats
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Breeding Laboratories, Wilmington, Mass., U.S.A. and St-Aubin-les-Elbeuf, France.
- Housing: Female rats (5 per cage) were housed in opaque plastic cages.
- Diet: Spratt’s Laboratory Diet No. 1, ad libitum.
- Drinking water: ad libitum.

ENVIRONMENTAL CONDITIONS
- Temperature: 20 ± 1°C
- Humidity: 50 ± 5 % RH

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
water
Details on exposure:
- Preparation of dosing solutions: Doses were prepared on daily basis as series of graded aqueous solutions for oral dosing.
- Administration of dosage: Standard volume of doses was administered orally by gastric intubation. Control animals were dosed with water.
- Duration of dosage: Doses were commenced on day 6 after detection of vaginal plug in rats and continued daily up to and including day 15 of gestation.
Analytical verification of doses or concentrations:
not specified
Details on mating procedure:
Mating was confirmed by the detection of vaginal plug in rats.
Duration of treatment / exposure:
Animals were dosed for 10 days [from Day 6 (after detection of vaginal plug) to Day 15].
Frequency of treatment:
daily from day 6 after detection of the vaginal plug to till day 15
Duration of test:
21 days
Doses / concentrationsopen allclose all
Dose / conc.:
0 mg/kg bw/day
Remarks:
(control)
Dose / conc.:
0.2 mg/kg bw/day
Dose / conc.:
2 mg/kg bw/day
Dose / conc.:
300 mg/kg bw/day
Dose / conc.:
600 mg/kg bw/day
No. of animals per sex per dose:
20 female rats per dose
Control animals:
yes
Details on study design:
- Dose selection rationale: Two doses formed the basis for safety evaluation (0.2 and 2.0 mg/kg bw/day) because the likely maximum human intake of detergent from ordinary kitchen use has been estimated at 0.14 mg/kg bw/day, thus providing factors of 1-2 times the human exposure level. Two further doses (300 and 600 mg/kg bw/day) were also investigated based on previous toxicity data suggesting that these would impair maternal economy and result in obvious adverse effects.

Examinations

Maternal examinations:
All animals were observed daily for signs of malreaction and were weighed regularly throughout gestation. All animals that died, and survivors at termination, were dissected and examined for macroscopic changes.
Ovaries and uterine content:
After euthanasia, uteri were dissected and examined to determine the number of:
a) Implants
b) Viable young
c) Embryonic deaths (abortion/resorption sites)

Ovaries were examined and number of corpora lutea were also recorded.
Fetal examinations:
After weighing, fetus were examined for external malformations, visceral anomalies by free and sectioning and skeletal examination was performed by alizarin staining.
Statistics:
Differences in mean values were statistically analyzed by non-parametric method i.e. Wilcoxon test.
Historical control data:
In CD rats, percentage of major malformations, minor visceral anomalies and minor skeletal anomalies were recorded as 0.41%, 2.02% and 2.35%, respectively in 51349 examined fetuses.

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Toxic effects were generally associated with the gastrointestinal tract disturbances.
Dermal irritation (if dermal study):
not examined
Mortality:
mortality observed, treatment-related
Description (incidence):
Single mortality was observed at 600 mg/kg bw/day.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Retarded weight gain was observed on 600 mg/kg bw/day.
Food consumption and compound intake (if feeding study):
not examined
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
no effects observed
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
not examined
Histopathological findings: neoplastic:
not examined
Other effects:
not examined
Details on results:
Retarded weight gain was observed on 600 mg/kg bw/day. No other significant maternal toxicity was observed at all doses when compared to the control group. Mating index was also comparable across all doses when compared to the control group.

Maternal developmental toxicity

Number of abortions:
no effects observed
Pre- and post-implantation loss:
no effects observed
Total litter losses by resorption:
not specified
Early or late resorptions:
not specified
Dead fetuses:
no effects observed
Changes in pregnancy duration:
not specified
Changes in number of pregnant:
not specified
Other effects:
not specified
Details on maternal toxic effects:
Body weight: Moderate maternal toxicity was observed in 600 mg/kg bw/day on the basis of retarded weight gain.

Litter loss: Only single litter was lost at 0.2 mg/kg bw/day but not significant as no dose response relationship was followed.

Pre-implantation embryonic losses were 18.1%, 23.2%, 36.9%, 15.6% and 20.3% at 0, 0.2, 2, 300 and 600 mg/kg bw/day, respectively.

Post-implantation embryonic losses were 2.8%, 12.4%, 5.9%, 3.6% and 3.1% at 0, 0.2, 2, 300 and 600 mg/kg bw/day, respectively.

Embryonic deaths were 0.3, 0.7, 0.6, 0.4 and 0.4 at 0, 0.2, 2, 300 and 600 mg/kg bw/day, respectively.

Effect levels (maternal animals)

Key result
Dose descriptor:
NOAEL
Effect level:
300 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
other: mortality and retarded weight gain at 600 mg/kg bw/day

Maternal abnormalities

Abnormalities:
not examined

Results (fetuses)

Fetal body weight changes:
no effects observed
Reduction in number of live offspring:
no effects observed
Changes in sex ratio:
not specified
Changes in litter size and weights:
no effects observed
Changes in postnatal survival:
not specified
External malformations:
no effects observed
Skeletal malformations:
no effects observed
Visceral malformations:
no effects observed
Other effects:
not specified
Details on embryotoxic / teratogenic effects:
EMBRYO TOXICITY
Fetal weights: Although the fetal weights differed significantly at 0.2 and 2 mg/kg bw/day, these changes does not represent an adverse change because there was no consistent dose relationship and were invariably due to higher mean pub weight in the test group.

Fetal loss: No test substance related effect or statistically significant differences were observed.

Litter size and weight: Litter size and weights were comparable across all doses when compared to the control group.

MALFORMATIONS:
No test substance related or statistically differences in the incidence of major malformations observed even a maternally toxic dosage but minor visceral anomalies were increased at 600 mg/kg bw/day (not significant statistically) but same incident was also observed in control group.

DEVELOPMENTAL VARIATIONS:
Incidences of skeletal abnormalities were comparable across all doses when compared to the control group.

Effect levels (fetuses)

Key result
Dose descriptor:
NOAEL
Effect level:
300 mg/kg bw/day
Based on:
test mat.
Sex:
not specified
Basis for effect level:
other: marginal retardation of sternebral ossification at the highest dose

Overall developmental toxicity

Key result
Developmental effects observed:
yes
Lowest effective dose / conc.:
600 mg/kg bw/day (nominal)
Treatment related:
not specified
Relation to maternal toxicity:
developmental effects as a secondary non-specific consequence of maternal toxicity effects

Any other information on results incl. tables

Formula for calculation of mating index:

Mating index: (number of confirmed mating/ number of animals cohabitated) X 100

Table 1: Mating index of the maternal animals (Palmer et.al., 1975)

Dose 

Number of confirmed mating

Number of animals cohabitated

 
Mating index %  
0 15 20 75
0.2 15 20 75
2 18 20 90
300 16 20 80
600 16 19 84

Table 2: Summary of adult performance (Palmer et.al., 1975)

Observations  Number of animals at dosage (mg/kg bw/day)
0 0.2 2 300 600
Mated  20 20 20 20 20
Died  0 0 0 0 1
Non-pregnant 5 5 2 4 3
Total litter loss 0 1 0 0 0
With viable young  15 14 18 16 16
Body weight change  - - - - retarded gain 

Table 3: Group mean litter data (Palmer et.al., 1975)

Dosage (mg/kg bw/day)

Number of litters  Litter size viable young  Embryogenic deaths  Implantations  Corpora lutea  Embryonic loss %  Litter weight (g) Fetal weight (g)
Pre-implantation  Post-implantation 
0 15 (A & B) 9.9 0.3 10.3 12.5 18.1 2.8 36.15 3.66
0.2 15 (A) 9.1 0.7 9.8 12.7 23.2 12.4 - -
14 (B) 9.8 0.6 10.4 12.8 18.4 6.1 37.14 3.84a
2 18 (A & B) 8.2 0.6 8.8 13.9 36.9 5.9 31.62 3.94b
300 16 (A & B) 9.6 0.4 10 12.4 15.6 3.6 35.72 3.78
600 16 (A & B) 10.4 0.4 10.8 13.9 20.3 3.1 37.49 3.63

A: Mean value includes all surviving animals showing evidence of implantation, including those with total litter loss

B: Mean value includes only animals bearing viable young at termination

Difference from controls statistically significant at Wilcoxon test: aP < 0.05 and bP < 0.001.

Table 4:

a)       Group mean incidence for major and minor malformations (Palmer et.al., 1975)

Dosage (mg/kg bw/day) Number of young
Major malformations  Minor malformations*
Gross or visceral Skeletal
Examined  Affected  Examined  Affected  Examined  Affected 
Total number  Mean (%)  Total number  Mean (%)  Total number  Mean (%) 
0 149 0 0 33 1 2.2 116 2 2
0.2 137 0 0 26 0 0 111 2 1.7
2 147 0 0 22 0 0 125 5 5.5
300 153 1 0.6 30 0 0 122 4 3.6
600 166 0 0 34 1 6.7 132 2 1.8

b) Group mean incidence for pups with extra ribs (Palmer et.al., 1975)

Dosage (mg/kg bw/day) Mean percent incidences of pups with extra lumber ribs*
0 18.6
0.2 33.1
2 21.3
300 12.9
600 15.3

* Young showing major malformations excluded

Applicant's summary and conclusion

Executive summary:

A pre-natal developmental toxicity study is conducted with C10-13LAS, sodium salt in pregnant female CD rats. The test substance was administered to groups of 20 pregnant female rats via oral gavage at doses of 0, 0.2, 2, 300 and 600 mg/kg bw/day from gestation day 6 (GD 6) till GD 15. All animals were observed daily for clinical signs of toxicity and were weighed regularly throughout the gestation. On GD 17, animals were euthanised and uteri were examined for number of implantations, viable youngs and embryonic deaths (abortion or resorption sites). Ovaries were examined and number of corpora lutea were also recorded. Individual foetus was weighed and examined for external malformations. About one third of the foetuses were examined for visceral anomalies and remaining two thirds for skeletal anomalies. Moderate maternal toxicity was observed at the highest dose due to retarded weight gain and mortality. These effects were associated with the gastrointestinal disturbances. No instances of major, minor and skeletal malformations were observed up to the highest dose of 600 mg/kg bw/day with the exception of a marginal retardation of sternebral ossification at 600 mg/kg. Based on the results, the maternal toxicity and developmental toxicity NOAELs were determined to be 300 mg/kg bw/day based on retarded weight gain and mortality in treated females at higher dose and based on absence of major, minor and skeletal malformations at highest dose respectively (Palmer, 1975a).