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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:
other: Data is from peer-reviewed journal

Data source

Reference
Reference Type:
publication
Title:
Effects of tartrazine on exploratory behavior in a three-generation toxicity study in mice.
Author:
Toyohito Tanaka∗, Osamu Takahashi, Shinshi Oishi, Akio Ogata
Year:
2008
Bibliographic source:
Reproductive Toxicology 26 , 156–163

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
other: OECD Guideline 426 (Developmental Neurotoxicity Study)
Principles of method if other than guideline:
Three-generation reproductive and neurobehavioral toxicity study of Tartrazine in male and female mice
GLP compliance:
not specified
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Test material form:
solid: particulate/powder
Remarks:
migrated information: powder
Details on test material:
- Name of test material (as cited in study report):Tartrazine
- Molecular formula (if other than submission substance):C16H12N4O9-S2.3Na
- Molecular weight (if other than submission substance):534.36 g/mole
- Substance type:Organic
- Physical state:Liquid , dye
- Analytical purity:85.0%
- Impurities (identity and concentrations):No data available.

Test animals

Species:
mouse
Strain:
Crj: CD(SD)
Details on test animals and environmental conditions:
Details on test animal
TEST ANIMALS
- Source: Charles River Japan Inc., Kanagawa, Japan
- Age at study initiation: 5 weeks of age of the F0 generation to 9 weeks of age of the F2 generation.
- Weight at study initiation: (P) Males: x-x g; Females: x-x g; (F1) Males: x-x g; Females: x-x g
- Fasting period before study: No data available.
- Housing: They were housed individually in polycarbonate
Solid-floored cages with wood flakes.
- Diet (e.g. ad libitum): Basal diets (Nihon Clea, CE-2)
ad libitum.
- Water (e.g. ad libitum): They were given water ad libitum.
- Acclimation period: 1 week

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 25±1 ◦C
- Humidity (%):50±5%.
- Air changes (per hr): No data available
- Photoperiod (hrs dark / hrs light): 12 h light/12 h dark cycle

Administration / exposure

Route of administration:
oral: feed
Vehicle:
other: Basal diets (Nihon Clea, CE-2)
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:

DIET PREPARATION
- Rate of preparation of diet (frequency): The experimental diets were prepared bimonthly (three times) in the laboratory.
- Mixing appropriate amounts with (Type of food): After mixing tartrazine with the powdered diet (basal Diets, Nihon Clea, CE-2), pellets were formed and fed to mice. Tartrazine was stable in the pellets during the experimental period when previously measured by
HPLC. The homogeneity of the test compound was ensured by the preparation procedures of the experimental diets in the laboratory when previously measured by HPLC. The concentration and homogeneity of the test compound in the diet was not tested during the experimental period.

- Storage temperature of food: No data available.



Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Useing HPLC
Details on mating procedure:
- M/F ratio per cage:1:1
- Length of cohabitation: 5 days
- Proof of pregnancy: [vaginal plug / sperm in vaginal smear] referred to as [day 0 / day 1] of pregnancy-No data available.
- 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):No data available.
- Any other deviations from standard protocolNo data available.
Duration of treatment / exposure:
199 days (approx 200)

Frequency of treatment:
Daily
Duration of test:
199 days (approx 200)
No. of animals per sex per dose:
Total no. of animals-80
0 mg/kg bw/day -10 male and 10 female
75 mg/kg bw/day -10 male and 10 female
225 mg/kg bw/day -10 male and 10 female
675 mg/kg/day -10 male and 10 female
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale:
Micewere assigned to the groups by the stratified randomization method.

Examinations

Maternal examinations:
Parental animal: observation and examination for F0 generation - The animals were weighed individually on experimental days 0, 2, 4, 7, 14, 21, 28, and 30 during the preconception period. Food intake and chemical intake was also observed during the preconception period. No. of females examined, No. of pregnant females, No. of litters No. of offspring, Average litter size, Average litter weight, Total sex ratio (male/female) and Average sex ratio (male %) were also observed. Exploratory behavior of mice was measured in an animal movement analyzing system ANIMATE AT- at 8 weeks of in the F0.
Parental animal: observation and examination for F0 generation
For F1 generation body weight was observed at 4 weeks, 5 weeks, 6 weeks, 7 weeks, 8 weeks, and 9 weeks of age after weaning. Different neurobehavioral test like Surface righting,
Ovaries and uterine content:
No data available.
Fetal examinations:
Litter observation- For F1 generation offspring were weighed individually on PNDs 0, 4, 7, 14, and 21 during the lactation period. The survival indices were calculated as (live offspring at each period)/ (live and dead offspring at birth) ×100%. Negative geotaxis, Cliff avoidance, Swimming behavior and olfactory orientation were observed on PND 4-14 .Food intake and chemical intake was observed during Preconception, mating, gestation and
Lactation.
For F2 generation offspring were weighed individually on PNDs 0, 4, 7, 14, and 21 during the lactation period. The survival indices were calculated as (live offspring at each period)/ (live and dead offspring at birth) ×100%. The offspring were weaned when they were 4 weeks of age, and one male and one female were selected at random from each litter to continue treatment. The animals were weighed individually at 4 weeks, 5 weeks, 6 weeks, 7 weeks, 8 weeks, and 9 weeks of age after weaning.
Animals at 7weeks of age in the F1 and F2 generation each performed 1 trial per day for 3 days in a Biel-type multiple-T water maze adapted for mice. Exploratory behavior of mice was measured in an animal movement analyzing system ANIMATE AT- at 8 weeks of in the F1 and F2.
Statistics:
Food intake, litter size, litter weight, and body weight were assessed with Bonferroni’s multiple comparison tests after the analysis of variance (ANOVA) or the Kruskal–Wallis test. Sex ratio, survival and behavioural developmental data were assessed with the X2-test or with Fisher’s exact test of frequency analysis. Movement activity data were assessed with the Steel–Dwass test of non-parametric methods. Multiple-T water maze performance data were assessed with the Sign–Wilcoxon test for trials and assessed with the Steel–Dwass test within each treatment group. Dose-response effects were assessed with the Jonckheere test for ordered alternatives or the cumulative -test (multi) for frequency data.
Indices:
Fertility index and viability index of offspring were opbserved.
Historical control data:
No data available.

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:
Maternal toxic effects:no effects

Details on maternal toxic effects:
Body weight -No significant change was observed in the male and female mice during the preconception or mating periods in F0 generation compare to control.

Food consumption:No significant effect on food consumption were observed in treated mice in F0 generation as compared to control.

Test substance intake- No significant change was observed in the chemical intake of male and female mice during the preconception ,mating periods, gestation and lactation period in F0 generation as compared to control.

Reproductive performance- No significant adverse effect was observed on survival, litter size, litter weight, or sex ratio at birth were observed in F0 generation as compared to control.

Effect levels (maternal animals)

Dose descriptor:
NOAEL
Effect level:
643.8 mg/kg bw/day
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:
Viability: No significant adverse effect were observed on viability of treated mice were observed in F1 and F2 generation: as compared to control.

Body weight -No significant change was observed in the male and female mice during the preconception or mating periods in all treated groupcompare to control in F1 and F2 generation.

Food consumption- No significant change was observed in the food consumption of F1 male and female mice during the preconception ,mating periods, gestation and lactation period in all treated group compare 0, 0.05%, 0.15%, and 0.45% to control.

Reproductive performance: No significant adverse effect was observed on litter size, litter weight, or sex ratio at birth of treated mice were observed in F1 and F2 generation: as compared to control.

Neurological behaviour-
F1 generation: At 225 mg/kg bw/day, significantly accelerated swimming direction in male offspring and Surface righting in female offspring at PND 7 were observed. In male offspring, movement time (s), total distance (cm), average distance (cm) and number of turning were significantly affected at 3 weeks of age and no significant adverse effect was observed on movement activity and bodyweight of treated F1 offspring.

F2 generation: At 75 mg/kg bw/day, significant increase in body weight of female offspring at PNDs 14 and 21, and in male offspring at PNDs 7, 14 and 21. Significant acceleration of swimming direction at PND 7 at 675 mg/kg bw/day and time taken of olfactory orientation at PND 14 and Surface righting at PND 7 in 225 mg/kg bw/day and time taken of olfactory orientation at PND 14 in 675 mg/kg bw/day were observed as compared to control. In male offspring, significant tendency to affected total distance (cm), average distance (cm) and average speed (cm/s) at 3 weeks and 8 week of age were observed at 675 mg/kg bw/day.

Effect levels (fetuses)

Dose descriptor:
NOAEL
Effect level:
643.8 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: No significant effect were observed onthe litter size or sex ratio at birth,litter weight and viability index of offspring

Fetal abnormalities

Abnormalities:
not specified

Overall developmental toxicity

Developmental effects observed:
not specified

Applicant's summary and conclusion

Conclusions:
NOAEL was found to be 643.8 mg/kg/day (0.45%) for Tartrazine in both F1and F2generation (Crlj: CD1 male and female mice when their dams were exposed to test chemical by oral diet.
Executive summary:

In a three-generation reproductive and neurobehavioral toxicity study, Crlj: CD1 male and female mice were treated with Tartrazine in the concentration of 0, 75, 225 and 675 mg/kg bw/day orally in feed. In F0 generation, No significant effect on body weight, food consumption and exploratory behaviour of treated mice were observed as compared to control. No significant effect on birth rate of offspring was observed as compared to control. In F1 generation, No significant adverse effect was observed on survival, litter size, litter weight, or sex ratio at birth. Significant increase in body weight was observed in 75, 225 and 675 mg/kg bw/day but not significant as compared to control. At 225 mg/kg bw/day, significantly accelerated swimming direction in male offspring and Surface righting in female offspring at PND 7 were observed. In male offspring, movement time (s), total distance (cm), average distance (cm) and number of turning were significantly affected at 3 weeks of age and no significant adverse effect was observed on movement activity and bodyweight of treated F1 offspring. In F2 generation, no significant adverse effects were observed on survival indices, litter size, litter weight, or sex ratio at birth as compared to control. At 75 mg/kg bw/day, significant increase in body weight of female offspring at PNDs 14 and 21, and in male offspring at PNDs 7, 14 and 21. Significant acceleration of swimming direction at PND 7 at 675 mg/kg bw/day and time taken of olfactory orientation at PND 14 and Surface righting at PND 7 in 225 mg/kg bw/day and time taken of olfactory orientation at PND 14 in 675 mg/kg bw/day were observed as compared to control. In male offspring, significant tendency to affected total distance (cm), average distance (cm) and average speed (cm/s) at 3 weeks and 8 week of age were observed at 675 mg/kg bw/day. No significant effects on reproduction were observed in treated male and female offspring were observed as compared to control. Few adverse effects on several behavioral parameters were observed in 675 mg/kg bw/day which is in excess of the ADI of tartrazine (0–7.5 mg/kg bw) unlikely to produce any adverse effects in humans. Therefore, NOAEL was considered to be 675 mg/kg bw/day for F0, F1 and F2 generation when Crlj: CD1 male and female mice were treated with Tartrazine orally in feed approx. 200 days.