N-(1,1,3,3-tetramethylbutyl)acrylamide

Developmental toxicity / teratogenicity

Administrative data

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2004-02-16 to 2008-09-25

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

This is a modern GLP compliant screening study which follows the OECD422 guideline including the well defined test substance, N-(1,1,3,3-tetramethylbutyl)acrylamide CAS 4223-03-4. This study would eb expected to detect any clear developmental toxicity or teratogenic effects as both sexes are treated with the test substance for two weeks prior to mating.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2008

Report date:

2008

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

other: OECD422

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

N-(1,1,3,3-tetramethylbutyl)acrylamide

EC Number:

224-169-7

EC Name:

N-(1,1,3,3-tetramethylbutyl)acrylamide

Cas Number:

4223-03-4

Molecular formula:

C11H21NO

IUPAC Name:

N-(1,1,3,3-tetramethylbutyl)acrylamide

Test material form:

solid: particulate/powder

Remarks:

migrated information: powder

Details on test material:

Name of test material in report: Octylamcrylamide 27-8098
Analytical purity: 99.7%
Impurities: Polymer 0.08%. Sulfuric acid 0.011%, Water 0.20%

Test animals

Species:

rat

Strain:

other: Crl:WI(Glx/BRL/Han)BR

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River (UK) Ltd, Margate
- Age at study initiation: 13 to 15 weeks old
- Weight at study initiation: from 286.4 to 466.3 g (males) and from 200.9 to 253.4 g (females)
- Fasting period before study: no data
- Housing: in groups of up to five (pre- and post-pairing)
- Diet (e.g. ad libitum): SQC Rat and Mouse Breeder Diet No 3, Expanded. (Special Diets Sevices Ltd. Witham), ad libitum
- Water (e.g. ad libitum): Mains water ad libitum
- Acclimation period: at least 10 days prior to the start of treatment

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19-25
- Humidity (%): 40-70
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

other: 5% v/v ethanol in corn oil

Details on exposure:

Males: for 2 weeks prior to pairing, during the pairing period and until the day before necropsy
Females: for 2 weeks prior to pairing, during the pairing period and until day 4 post-partum, inclusive

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

HPLC: analysis was performed at the beginning and at the end of the dosing period

Details on mating procedure:

After the two week pre-mating dosing the rats were mated one male with each female, daily vaginal smears were taken. The appearance of a positive vaginal smear was taken as an indication of successful mating

Duration of treatment / exposure:

Males: for 2 weeks prior to pairing, during the pairing period and until the day before necropsy
Females: for 2 weeks prior to pairing, during the pairing period and until day 4 post-partum, inclusive

Frequency of treatment:

daily

Duration of test:

Males at least 28 days
Females ca. 40 days (until after day 4 post partum.

Doses / concentrations

Doses / concentrations 1

Remarks:

Doses / Concentrations:
70 mg/kg bw/day
Basis:
actual ingested

Doses / concentrations 2

Remarks:

Doses / Concentrations:
155 mg/kg bw/day
Basis:
actual ingested

Doses / concentrations 3

Remarks:

Doses / Concentrations:
350 mg/kg bw/day
Basis:
actual ingested

No. of animals per sex per dose:

10

Control animals:

yes, concurrent vehicle

Details on study design:

Dose selection rationale: the high dose was based on the results of the rat Maximum Tolerated Dose assay where a dose level of 300 mg/kg/day was the no-observedadverse-effect-level. Dose levels of 400 mg/kg/day or above caused neurological signs such as ataxia, prostration, spasmodic twitching and convulsions in some females and males. The low dose level of 70 mg/kg/day was not expected to elicit parental toxicity and the intermediate dose level of 155 mg/kg/day is the geometric mean of these values.
- Rationale for selecting satellite groups: no data
- Post-exposure recovery period in satellite groups: no data
- Section schedule rationale (if not random): no data

Examinations

Maternal examinations:

HISTOPATHOLOGY / ORGAN WEIGHTS
Listed tissues from 5 selected adult terminal animals per sex Group 1 and 4adults examined:
Femur+marrow, sciatic nerve, liver, spleen, mesenteric LN, stomach, duodenum, jejunum, ileum, caecum, colon, adrenal, kidney, ovary, urinary bladder, uterus, vagina, mandibular LN, thymus, lung, heart, trachea,
pituitary, brain, spinal cord, skin+subcutis, pancreas

Ovaries and uterine content:

Ovaries were examined during the post mortem examination and the numbers of implantation sites counted, to allow comparison with the number of offspring seen at day 1 post partum.

Fetal examinations:

Pups were weighed and examined on day 1 post partum for any obvious external abnormalities. They were then observed daily until day 4 when they were again weighed to allow an examination of their weight gain. An assessment was also made of pup survival based on the comparison of the number of pups in each litter as day 1 and Day 4 post partum. Any pups found dead and all pups on day 5 post partum were given a gross post mortem examination for the presence of any abnormalities.

Statistics:

Body weight gains, necropsy body weights, food consumption, haematology, clinical chemistry, locomotor and functional observation
data were analysed using one-way analysis of variance (ANOVA), separately for each sex. Levene's test for equality of variances
among the groups was performed. Where this showed no evidence of heterogeneity (P≥0.01). pairwise comparisons with control
were made using Dunnett's test. A linear contrast was used to determine whether there was a relationship between increasing dose
and response. A significant trend (P<0.05) was reported only where none of the pairwise comparisons was significant. The numbers
of implantation sites and pups born, the percentage of male pups on Day 1 and the mean pup weights were analysed using nonparametric
methods. The non-parametric methods employed were the Kruskal-Wallis ANOVA, the Terpstra-Jonckheere test for a
dose related trend and the Wilcoxon rank sum test for pairwise comparisons. Where the Kruskal-Wallis ANOVA was not significant,
the pairwise comparisons were not reported in order to protect the Type I error. The non-parametric methods were used in place of
the one-way ANOVA for clinical chemistry parameters with values above or below the l imit of the assay. Organ weights were
analysed using Analysis of Covariance (ANCOVA) and Dunnett's test, for each sex separately, using the necropsy body weight as
covariate. This analysis depends on the assumption that the relationship between the organ weights and the covariate is the same for
all groups and the validity of this assumption was tested. Where the test for equality of slopes failed (male heart weights; P<0.01), the
organ was analysed using one-way ANOVA on absolute organ weights and organ to necropsy body weight ratios. Levene's test for
equality of variances across the groups was also performed for all organs and, in all cases, this showed no evidence of heterogeneity
(P≥0.01).

Indices:

For each litter to day 4 post partum: number of pups born (live and dead); daily live litter size and sex, daily clinical observations,
individual pup weights on days 1 and 4 post partum

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:

Maternal toxic effects:yes. Remark: reduced food intake and effects in the lungs

Details on maternal toxic effects:
No mortality occurred. One high dose group female (number 79) showed signs of subdued bahviour, twitching and piloerection
after dosing on Days 4 and 5, was not dosed on Day 6, staggered after dosing on Day 7 but not on subsequent days.

At histopathological examination, there were several findings, these included in the liver, centrilobular hypertrophy was for two intermediate dose group females and the majority of high dose group females. Centrilobular hypertrophy was characterised by enlarged hepatocytes with a pale or more eosinophilic cytoplasm. In the lungs, there was a minor increase in the level of foamy histiocytes in high dose group females. Foamy histiocytes were characterised by small collections of alveolar histiocytes with abundant foamy cytoplasm.

Effect levels (maternal animals)

Dose descriptor:

NOAEL

Effect level:

155 mg/kg bw/day (actual dose received)

Based on:

test mat.

Basis for effect level:

other: Reduced litter size at 350 mg/kg bw/day

Maternal abnormalities

Abnormalities:

not specified

Results (fetuses)

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
The number of implantations and the litter size and therefore litter weight were reduced at the highest dose of 350mg/kg bw/day and the NOAEL for developmental effects is 155 mg/kg bw/day. Malformed/shortened limbs occurred in one pup in the intermediate and high dose in the absence in any increase in incidence and severity of this finings with increased dose, these findings are considered to be incidental and not clearly related to treatment

Effect levels (fetuses)

Dose descriptor:

NOAEL

Effect level:

155 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: Slightly reduced pup survival at 350 mg/kg bw/day

Fetal abnormalities

Abnormalities:

not specified

Overall developmental toxicity

Developmental effects observed:

not specified

Applicant's summary and conclusion

Conclusions:

Oral administration of the test substance dissolved in 5 % v/v ethanol in corn oil to female and male rats at doses of 70, 155, 350 mg/kg bw/day for 14 days prior to mating, during mating period, gestation and four days post partum resulted in limited evidence of developmental toxicity with the number of implantations and the litter size being reduced at the highest dose in the presence of maternal toxic effects, the NOAEL for reproductive effects is 155 mg/kg bw/day.

Executive summary:

The purpose of this study was to provide preliminary information on the possible effects of the test article t-octyl acrylamide following repeated administration in the rat following OECD Guideline 422. Male and female Crl:WI(Glx/BRL/Han)BR rats were assigned to four groups (10 animals/sex/group). Each treated group received dose preparations containing the vehicle (5% v/v ethanol in corn oil) or 70, 155 or 350 mg of test article/kg of body weight/day (mg/kg/day) at a dose volume of 5 mL/kg. Animals were treated for two weeks prior to mating, during mating period of 15 days and in case of females during gestation and for 4 days post partum. Oral administration of t-octyl acrylamide at a dose level of 350 mg/kg/day elicited adult toxicity in the form of reduced food intakes, a transient reduction in body weight gain, reduced forelimb grip strength in males and microscopic findings in the liver, kidneys and lungs. At a dose level of 155 mg/kg/day liver weights in male rats was increased with microscopic liver changes seen in some males and females. This change is considered adaptive rather than adverse, therefore the NOAEL for parental toxicity was considered to be 155 mg/kg/day. Oral administration of the test article had no effects on mating or fertility indices. However, the number of implantations and the litter size were reduced at the highest dose and the NOAEL for reproductive effects is 155 mg/kg bw/day.