Registration Dossier

Administrative data

Endpoint:
toxicity to reproduction: other studies
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study
Justification for type of information:
Data is from publication

Data source

Reference
Reference Type:
publication
Title:
Early Life Triclocarban Exposure During Lactation Affects Neonate Rat Survival
Author:
Rebekah C. M. Kennedy, Fu-Min Menn, Laura Healy, Kellie A. Fecteau, Pan Hu, Jiyoung Bae, Nancy A. Gee, Bill L. Lasley, Ling Zhao, and Jiangang Chen
Year:
2015
Bibliographic source:
Reproductive Sciences 2015, Vol. 22(1) 75-89

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
other: 414- prenatal develpmental toxicity studies
Deviations:
yes
Principles of method if other than guideline:
The experiment designed to differentiate whether the decreased neonate survival was secondary to the effect of test chemical on the reduction in the lactational capacity of the mammary glands
GLP compliance:
yes
Type of method:
in vivo

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): Triclocarban (3,4,4-trichlorocarbanilide; TCC)
- Molecular formula (if other than submission substance): C13H9Cl3N2O
- Molecular weight (if other than submission substance):315.5861g/mol
- Substance type: Organic
- Physical state: No data
- Impurities (identity and concentrations): Purity 99% and impurities 1% (Unknown)

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
female
Details on test animals and environmental conditions:
- Source: Harlan Laboratory, Dublin, Virginia
- Age at study initiation: (P) x wks; (F1) x wks (Animals were pregnant)
- Weight at study initiation: No data
- Fasting period before study: No data
- Housing: housed individually with Harlan Teklad laboratory grade 7087 soft cob bedding (Harlan Laboratories, Madison, Wisconsin) in clear plastic cages

- Use of restrainers for preventing ingestion (if dermal): yes/no
- Diet (e.g. ad libitum):
- Water (e.g. ad libitum): Ad libitum assess to water
- Acclimation period: Harlan ground 2020X chow

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20_C to 22_C
- Humidity (%):of 40% to 50%.
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): 12:12-hour photoperiod

Administration / exposure

Route of administration:
oral: feed
Vehicle:
other: feed
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
The test chemical supplement was prepared by first weighing the correct amount of test chemical and mixing the compound with small amounts of powdered chow using a mortar and pestle.

DIET PREPARATION
- Rate of preparation of diet (frequency): weekly
- Mixing appropriate amounts with (Type of food): powdered chow to obtain the required concentration.
- Storage temperature of food: no data
Analytical verification of doses or concentrations:
not specified
Duration of treatment / exposure:
GD5 to PND14
Frequency of treatment:
daily
Doses / concentrations
Remarks:
0.5% =500mg/kg
based on concentration in diet
No. of animals per sex per dose:
Control=6 pregnant female
0.5% (500mg/kg)= 3 preganant female
Control animals:
yes, plain diet
Details on study design:
Pregnant (GD5) dams were weight ranked and randomized by body weight into groups fed either rat chow (n =6) or chow supplemented with 0.5% w/w test chemical (n = 3) from GD 5 until PND 14. After delivery at PND 0, litter size was culled to 6 from all 3 of 0.5% w/w-treated dams and only 2 control dams. No culling was conducted for the rest of the control dams (n = 4) which served as reserve controls to provide healthy pups Starting on PND 1, healthy age-matched pups (n = 3) born to the 4 reserve control litters were added to replace half (n =3) the pups raised by test c hemical treated dams to maintain normal suckling activity. Therefore, by PND 1, all 0.5% w/w-treated dams carried 6 pups (3 born to 0.5% w/w test chemical-treated dams and 3 born to reserve control dams). On PND 3, the same procedure was conducted as PND 1, except 3 healthy control pups born to the 4 reserve control dams were added to treated dams to replace the pups previously transferred on PND 1 from reserve control dams. Therefore, on PND 3, all 0.5% w/w test chemical-treated dams carried 3 of their own pups and 3 new pups transferred from reserve control dams. At PND 6, the procedure was again conducted except that 3 healthy age-matched pups born to the reserve control dams were added to each treated dam to replace the remaining 3 pups originally born to 0.5% w/wtreated dams. After the above-mentioned manipulation on PND 6, pups nursed by the treated dams were all born to reserve control dams. The same substitution procedure was conducted once again on PND 9, and this time the 3 pups transferred from reserve control dams to treated dams on PND 3 were replaced. Milk band quantification comparison was only conducted between control born/raised animals and 0.5% w/w born/raised animals on PNDs 1, 3, and 6, the last day before all the pups born to test chemical-treated dams were replaced with pups born to reserve control dams.All dams were terminated on PND 14 and mammary tissue was removed for histological analysis.

Milk bands were rated as described by Ruppert and colleagues.
Briefly, 0 indicates no band visible; 1, small band visible on the side of pup; 2, small band visible across pup’s abdomen, and 3, large band visible across the pup’s abdomen
Statistics:
Data were presented as group mean + standard error of the mean. Data were analyzed using SPSS (version 20, IBM, Armonk, New York) by analysis of variance or ANOVA with repeat measurements. In addition Milk band rating was analyzed with a nonparametric Mann-Whitney U test. Statistical significance was considered P <.05. Data were transformed if either normality or the equal variance assumption was invalid. If transformation did not correct normality or equal variance assumption, Kruskal-Wallis one-way ANOVA on ranks was used.

Results and discussion

Effect levels

Dose descriptor:
dose level:
Effect level:
500 mg/kg bw/day
Based on:
test mat.
Sex:
female
Basis for effect level:
other: overtime decresed milk band size
Remarks on result:
other: Not Specified

Observed effects

The milk band size decreased over time after PND 3. On PND 6, the median milk band score was 0 in 0.5% w/w born/raised pups and 2 among pups born/raised by control dams Compared to results from control dams.

Histology evaluation revealed that mammary tissue collected from treated dams on PND 14 was not involuted when additional healthy pups were continuously provided on PNDs 3, 6, and 9 to maintain normal suckling activity

Applicant's summary and conclusion

Conclusions:
The dose level of the test chemical in pregnant Sprague Dawley rats was considered to be 500mg/kg when test chemical exposed during gestation and lactation period results in decreased milk band size overtime
Executive summary:

The  experiment designed to differentiate whether the decreased neonate survival was secondary to the effect of test chemical on the reduction in the lactational capacity of the mammary glands. Pregnant (GD5) dams were weight ranked and randomized by body weight into groups fed either rat chow (n =6) or chow supplemented with 0.5% w/w test chemical (n = 3) from GD 5 until PND 14. After delivery at PND 0, litter size was culled to 6 from all 3 of 0.5% w/w-treated dams and only 2 control dams. No culling was conducted for the rest of the control dams (n = 4) which  served as reserve controls to provide healthy pups Starting on PND 1, healthy age-matched pups (n = 3) born to the 4 reserve control litters were added to replace half (n =3) the pups raised by test c hemical treated dams to maintain normal suckling activity. Therefore, by PND 1, all 0.5% w/w-treated dams carried 6 pups (3 born to 0.5% w/w test chemical-treated dams and 3 born to reserve control dams). On PND 3, the same procedure was conducted as PND 1, except 3 healthy control pups born to the 4 reserve control dams were added to treated dams to replace the pups previously transferred on PND 1 from reserve control dams. Therefore, on PND 3, all 0.5% w/w test chemical-treated dams carried 3 of their own pups and 3 new pups transferred from reserve control dams. At PND 6, the procedure was again conducted except that 3 healthy age-matched pups born to the reserve control dams were added to each treated dam to replace the remaining 3 pups originally born to 0.5% w/wtreated dams. After the above-mentioned manipulation on PND 6, pups nursed by the treated dams were all born to reserve control dams. The same substitution procedure was conducted once again on PND 9, and this time the 3 pups transferred from reserve control dams to treated dams on PND 3 were replaced. Milk band quantification comparison was only conducted between control born/raised animals and 0.5% w/w born/raised animals on PNDs 1, 3, and 6, the last day before all the pups born to test chemical-treated dams were replaced with pups born to reserve control dams.All dams were terminated on PND 14 and mammary tissue was removed for histological analysis. Milk bands were rated as described by Ruppert and colleagues. Briefly, 0 indicates no band visible; 1, small band visible on the side of pup; 2, small band visible across pup’s abdomen, and 3, large band visible across the pup’s abdomen The result o fthe study revealed, milk band size decreased over time after PND 3. On PND 6, the median milk band score was 0 in 0.5% w/w born/raised pups and 2 among pups born/raised by control dams  Compared to results from control dams. Histology evaluation revealed that mammary tissue collected from treated dams on PND 14 was not involuted when additional healthy pups were continuously provided on PNDs 3, 6, and 9 to maintain normal suckling activity. The dose level of the test chemical in pregnant Sprague Dawley rats was considered to be 500mg/kg when test chemical exposed during gestation and lactation period results in decreased milk band size overtime.