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Toxicological information

Toxicity to reproduction

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Administrative data

Endpoint:
toxicity to reproduction
Remarks:
other: sub chronic
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:
Reproductive Toxicology of Tartrazine (FD and C Yellow No. 5) in Swiss Albino Mice
Author:
N. Mehedi, S. Ainad-Tabet, N. Mokrane, S. Addou, C. Zaoui, O. Kheroua and D. Saidi
Year:
2009
Bibliographic source:
American Journal of Pharmacology and Toxicology 4 (4): 130-135

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
other: as below
Principles of method if other than guideline:
To evaluate the testicular toxicity of Tartrazine in male swiss albino mice and when they were mated with untreated female mice.
GLP compliance:
not specified
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Test material form:
other: Liquid
Details on test material:
- Name of test material (as cited in study report):Tartrazine
- Molecular formula (if other than submission substance):C16H12N4O9S2.3Na
- Molecular weight (if other than submission substance):534.36 g/mole
- Substance type:Organic
- Physical state:Liquid , dye
- Analytical purity:86.7%
- Impurities (identity and concentrations):No data available.

Test animals

Species:
mouse
Strain:
Swiss
Sex:
male
Details on test animals and environmental conditions:
Details on test animal
TEST ANIMALS
- Source: No data available
- Age at study initiation: (P) x wks;- 4 weeks old (F1) x wks- No data available.
- Weight at study initiation: (P) Males: x-x g; - 20±2.01 g - Fasting period before study: No data available
- Housing: No data available
- Diet (e.g. ad libitum): Standard food pellets diet was
given ad libitum
- Water (e.g. ad libitum): water was given ad libitum
- Acclimation period: No data available

ENVIRONMENTAL CONDITIONS
They were kept under conditions of ambient room temperature and relative humidity.

Administration / exposure

Route of administration:
oral: drinking water
Vehicle:
water
Details on exposure:
Details on exposure
PREPARATION OF DOSING SOLUTIONS:

DIET PREPARATION
- Rate of preparation of diet (frequency):
- Mixing appropriate amounts with (Type of food):
- Storage temperature of food:

VEHICLE
- Justification for use and choice of vehicle (if other than water):
- Concentration in vehicle: The average (±SE) tartrazine intake calculated from liquid consumption, in mg kg-1 day-1, was (173.9±0.25), (1767.8±0.32), (5541.4±0.47) for 0.1, 1 and 2.5% Tartrazine groups, respectively.
- Amount of vehicle (if gavage):
- Lot/batch no. (if required):
- Purity:
Details on mating procedure:
- M/F ratio per cage:1:1 male and female
- Length of cohabitation:1 week
- Proof of pregnancy: [vaginal plug / sperm in vaginal smear] referred to as [day 0 / day 1] of pregnancy: Groups of Tartrazine treated mice, six males per dose group, were mated 1:1 with untreated females for 1 week. Females were then separated and allowed to gestate to term. For females that failed to deliver a litter, this was considered as a sign of male infertility whereas litter delivery indicated male fertility.
- 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 protocol:No data available.
Analytical verification of doses or concentrations:
not specified
Duration of treatment / exposure:
13 weeks
Frequency of treatment:
Daily
Details on study schedule:
0, 0.1%, 1% and 2.5% Groups of Tartrazine treated mice, six males per dose group, were mated 1:1 with untreated females for 1 week. Females were then separated and allowed to gestate to term. For females that failed to deliver a litter, this was considered as a sign of male infertility whereas litter delivery indicated male fertility. Litter size and weight after 7, 14 and 28 days of growth were examined.
Doses / concentrations
Remarks:
Doses / Concentrations:
0, 0.1%, 1% and 2.5% (173.9±0.25,1767.8±0.32and 5541.4±0.47 mg/kg/day )
Basis:

No. of animals per sex per dose:
Total: 24
0 mg/kg/day- 6 male mice
173.9 mg/kg/day- 6 male mice
1767.8 mg/kg/day-6 male mice
5541.4 mg/kg/day-6 male mice

Control animals:
yes, concurrent vehicle
Details on study design:
No data available.
Positive control:
No data available.

Examinations

Parental animals: Observations and examinations:
BODY WEIGHT: Yes
- Time schedule for examinations: Weekly

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No data

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): Yes
- Time schedule for examinations: Daily
Oestrous cyclicity (parental animals):
No data available.
Sperm parameters (parental animals):


Parameters examined in [all/P/F1/F2] male parental generations: For P generation
[testis weight, epididymis weight, daily sperm production, sperm count in testes, sperm count in epididymides, enumeration of cauda epididymal sperm reserve, sperm motility, sperm morphology, other:]-- Sperm motility, morphology and sperm production was observed.
Litter observations:


STANDARDISATION OF LITTERS
- Performed on day 4 postpartum: [yes/no]- No
- If yes, maximum of [...] pups/litter ([...]/sex/litter as nearly as possible); excess pups were killed and discarded.- No data available

PARAMETERS EXAMINED
The following parameters were examined in [F1 / F2 / F3] offspring:F1,Litter size and weight after 7, 14 and 28 days of growth were examined.
[number and sex of pups, stillbirths, live births, postnatal mortality, presence of gross anomalies, weight gain, physical or behavioural abnormalities, other:]

GROSS EXAMINATION OF DEAD PUPS:
[no / yes, for external and internal abnormalities; possible cause of death was/was not determined for pups born or found dead.]-No data available.
Postmortem examinations (parental animals):
After mating, male mice were killed by cervical dislocation. Testes, epididymides were weighed immediately. The left epididymis was excised. The tail region tissue was minced with scalpels for approximately 1min and placed in a 37°Cincubator for 15 min, prior to determining sperm motility.
•Histologic examination: Histologic examination of testis was performed. The left testis was fixed in formalin-buffer. Six microns thick paraffin sections were stained with hematoxylin and eosin and examined by light microscopy.
Postmortem examinations (offspring):
No data available.
Statistics:
The data is expressed as mean ± SE. Statistical test one way ANOVA was applied to find significant difference between values of various parameters recorded for control and treated animals. p<0.05 was considered statistically significant.
Reproductive indices:
Mating index was observed.
Offspring viability indices:
Yes

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:
not specified
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Other effects:
no effects observed

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:
not specified
Reproductive function: sperm measures:
effects observed, treatment-related
Reproductive performance:
effects observed, treatment-related

Details on results (P0)

CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS)- No data available.

BODY WEIGHT AND FOOD CONSUMPTION (PARENTAL ANIMALS)-Body weight gain was significantly increased in 1767.8 mg/kg/day Tartrazine (p<0.05). This increased bodyweight gain is not evidence related dose.

FOOD and liquid CONSUMPTION (PARENTAL ANIMALS)- Significant decrease in the food consumption was observed at all dose level 173.9, 1767.8 and 5541.4 mg/kg day treated group compare to control.
Significant increase in the water consumption was observed at all dose level 173.9, 1767.8 and 5541.4 mg/kg day in treated group compare to control.
(Data not shown).

TEST SUBSTANCE INTAKE (PARENTAL ANIMALS)- No data available

REPRODUCTIVE FUNCTION: ESTROUS CYCLE (PARENTAL ANIMALS)No data available

REPRODUCTIVE FUNCTION: SPERM MEASURES (PARENTAL ANIMALS)- Total number of spermatids count was reduced significantly in the mice administered 5541.4 mg/kg/day tartrazine (p<0.01).
Sperm concentration in epididymides was reduced in all treated groups 173.9, 1767.8 and 5541.4 mg/kg day but sperm epididymis reserves were reduced significantlyonly in mice treated with 5541.4 mg/kg/day tartrazine (p<0.01).
The percentage motility was reduced in 1767.8 and 5541.4 mg/kg/day treated groups (p<0.01).
The percentage morphologically normal spermatozoa were significantly affected in 5541.4 mg/kg/day dose level of tartrazine (p<0.01).


REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS)- Male mating index was decreased in the 5541.4 mg/kg/day treated groups compared to the control values.

ORGAN WEIGHTS (PARENTAL ANIMALS)- A decrease of relative testis and seminal vesicles weight were observed in all treated groups 173.9, 1767.8 and 5541.4 mg/kg day compared to control but not statistically significant. However, their absolute weight did not change.

GROSS PATHOLOGY (PARENTAL ANIMALS)-No data available.

HISTOPATHOLOGY (PARENTAL ANIMALS)- Histological examination revealed that semniferous tubules were not identical with conjunctive tissue dystrophy in animals testes treated with 173.9 mg/kg/day tartrazine.
Intercellular connections were reduced and imperfect and dilation in some semniferous tubules of testis mice treated 1767.8 mg/kg/day tartrazine.
Significant damage was observed in testis mice treated with 5541.4 mg/kg/day tartrazine; extensive disruption in semniferous tubules, widening of the interstitial spaces and loss leydig cells.
Spermatogenic cells are affected and then depleted with absence of spermatozoa in the Lumen .


OTHER FINDINGS (PARENTAL ANIMALS)

Effect levels (P0)

Dose descriptor:
NOAEL
Effect level:
173.9 mg/kg bw/day
Based on:
test mat.
Sex:
male
Basis for effect level:
other: No significant change were observed in the body weight ,reproductve fuction , Reproductive performance ,organ weight and histopathology ,litter size and weight.

Results: F1 generation

General toxicity (F1)

Clinical signs:
effects observed, treatment-related
Mortality / viability:
mortality observed, treatment-related
Body weight and weight changes:
effects observed, treatment-related
Sexual maturation:
not specified
Organ weight findings including organ / body weight ratios:
not specified
Gross pathological findings:
not specified
Histopathological findings:
not specified

Details on results (F1)

VIABILITY (OFFSPRING)- Yes

CLINICAL SIGNS (OFFSPRING)- There was significant decrease in the litter sizes in 1767.8 and 5541.4 mg/kg/day treated rats as comparison to litters sired from control males.


BODY WEIGHT (OFFSPRING)-There was significant decrease in the litter weight at treated dose group 173.9, 1767.8 and 5541.4 mg/kg/day comparison to litters sired from control males during lactation periiod.

Effect levels (F1)

Remarks on result:
not measured/tested

Overall reproductive toxicity

Reproductive effects observed:
not specified

Any other information on results incl. tables

This study attributed that the differences of offspring weight between the control and low-dosed groups were caused not by tartrazine treatment but by the body size at birth.

Applicant's summary and conclusion

Conclusions:
NOAEL was found to be 173.9±0.25 mg/kg/day (0.1%) for Tartrazine in male swiss albino mice for 13 weeks by oral drinking water administration .
Executive summary:

A sub chronic study was conducted for Tartrazine in male swiss albino mice for 13 weeks.They were administrated Tartrazine at the concentration of 0, 0.1%, 1% and 2.5% (173.9 ±0.25, 1767.8±0.32and 5541.4±0.47 mg/kg/day ) by oral drinking water. Groups of Tartrazine treated mice, six males per dose group, were mated 1:1 with untreated females for 1 week. Females were then separated and allowed to gestate to term. For females that failed to deliver a litter, this was considered as a sign of male infertility whereas litter delivery indicated male fertility. Food and water consumption, body weight, reproductve fuction, Reproductive performance, organ weight and histopathology were observed. Decrase in food consumption and increase in water consumption were observed in 173.9, 1767.8 and 5541.4 mg/kg/day treated male rats as compared to control. Average (±SE) tartrazine intake calculated from liquid consumption, in mg kg-1day-1, was (173.9±0.25), (1767.8±0.32), (5541.4±0.47) for 0.1, 1 and 2.5% tartrazine groups, respectively. Significantly increased body weight gain was observed in 1767.8 mg/kg/day and decrease in relative testis and seminal vesicles weight in 173.9, 1767.8 and 5541.4 mg/kg/day treated male rats, but the effect were not statistically significant and no effect on absolute organ weight were observed in treated male rats as compared to control. Decreased in male mating index and body weight and litter sizes were observed in 5541.4 mg/kg/day treated male rats as compared to control. Similarly, significant decreased in Total number of spermatids count, Sperm concentration in epididymis reserves and percentage motility in 5541.4 mg/kg/day, percentage motility and Sperm concentration in epididymides in 1767.8 mg/kg/day and Sperm concentration in epididymides in 173.9 mg/kg/day treated male rats as compared to control. Sperm head (amorphous, macro-or microcephaly) and the sperm flagellum (entangled, twisted, coiled, with ANSA) morphological abnormalities and significantly affected percentage morphologically normal spermatozoa were observed in 5541.4 mg/kg/day treated male rats as compared to control. In addition, extensive disruption in semniferous tubules, widening of the interstitial spaces and loss leydig cells, Spermatogenic cells are affected and then depleted with absence of spermatozoa in the lumen at 5541.4 mg/kg/day, decreased intercellular connections and imperfect and dilation in some semniferous tubules of testis at 1767.8 mg/kg/day and emniferous tubules were not identical with conjunctive tissue dystrophy in testes at 173.9 mg/kg/day treated male rats as compared to control. Therefore NOAEL was considered to be173.9 mg/kg/day (0.1%) when swiss albino male mice treated with tartrazine orally by drinking water for 13 weeks.