Poly(oxy-1,2-ethanediyl), α-[2-(dide- cylmethylammonio)ethyl]- .omega.- hydroxy-, propanoate (salt) (Bardap 26)

Toxicity to reproduction

Administrative data

Endpoint:

two-generation reproductive toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

not available

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1991

Report date:

1991

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

EPA OPP 83-4 (Reproduction and Fertility Effects)

Deviations:

no

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

DDAC

IUPAC Name:

DDAC

Constituent 2

Reference substance name:

Didecyldimethylammonium chloride

EC Number:

230-525-2

EC Name:

Didecyldimethylammonium chloride

Cas Number:

7173-51-5

Molecular formula:

C22 H48 N.Cl

IUPAC Name:

N-decyl-N,N-dimethyldecan-1-aminium chloride

Constituent 3

Reference substance name:

N,N-Didecyl-N,N-dimethylammonium Chloride

IUPAC Name:

N,N-Didecyl-N,N-dimethylammonium Chloride

Constituent 4

Reference substance name:

1-Decanaminium, N-decyl-N,N-dimethyl-, chloride

IUPAC Name:

1-Decanaminium, N-decyl-N,N-dimethyl-, chloride

Constituent 5

Reference substance name:

Badrac 2280

IUPAC Name:

Badrac 2280

Details on test material:

The test material was Bardac 2280 (Didecyldimethylammonium Chloride; DDAC, in aqueous/alcohol solution), batch number B-1889, obtained from Lonza Inc. The substance was described as a very viscous honey coloured liquid, supplied as 80.8% a.s. in aqueous/ethanol solution.

DDAC is hydrolytically and photolytically stable under the conditions of this study and has been shown to be stable in aqueous, alcohol and alcohol/aqueous solutions for extended periods, e.g. at least seven years under standard laboratory conditions.

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

The test animals were male and female Sprague-Dawley rats (Crl:CD (SD) BR), obtained from Charles River Breeding Laboratories (MI, USA). The animals were acclimatised for approximately 2 weeks. Representative animals were selected for faecal examination, serum viral antibody analysis and histological examination of selected organs. The rats were housed in pairs during quarantine, and singly thereafter (except during cohabitation and lactation) in stainless steel mesh cages. From gestational day 20 through to weening, females were housed in plastic shoebox cages with bedding.
The rats were 6 weeks old at study initiation and weighed 204.2-205.8 g (males) and 153.3-155.2 g (females). Diet (Certified Ground Rodent Chow #5002) and tap water were provided ad libitum. Room temperature and humidity were monitored, and a 12 hour light/dark cycle was maintained.

Administration / exposure

Route of administration:

oral: feed

Vehicle:

unchanged (no vehicle)

Details on exposure:

The test substance was mixed direcly into the animal diet (fed ad libitum). All weights of the test material were corrected for percent active ingredient for diet preparation, and loss of ethanol during diet preparation was also taken into account. The test diets were stored at room temperature.

Details on mating procedure:

Rats were mated one male to one female for 3 weeks.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Concentration of the test substance in the diet, homogeneity and stability were determined using gas chromatography with Nitrogen-Phosphorous detection. Homogeneity was confirmed, and the test diets were demonstrated to be stable for 21 days when stored in closed polyethylene containers and for at least 14 days when stored in open glass jars at room temperature. Periodic analyses of the test diets indicated that the mean test substance concentrations in the diet for the 300, 750 and 1500 ppm levels were 96.2-108.8%, 98.8-108.8% and 94.5-109.2% of nominal, respectively.

Duration of treatment / exposure:

F0: 27 weeks (from 1st prebreed dose to last F0 sacrifice)
F1: 32 weeks (from 1st F1B wean to last F1B sacrifice)
F2B: to weaning

Frequency of treatment:

Continuous - ad libitum dietary exposure

Details on study schedule:

After a 10-week pre-breed period, rats were mated (one male to one female) for three weeks to produce the F1A generation. The observation of a dropped copulation plug or the presence of vaginal sperm was considered evidence of successful mating. Exposures continued through mating, gestation, parturition and lactation. At least 10 days after the weaning of the F1A litters, the F0 parents were paired again to produce the F1B generation. At weaning, 28F1B weanlings/sex/group were selected to produce the F2 generation, and were then exposed to the same dietary concentrations of Didecyldimethylammonium Chloride as their parents for 10 weeks. After their pre-breed exposure, F1 animals were paired as described above to produce F2A and F2B litters.

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

300 ppm

Remarks:

Basis:
nominal in diet

Doses / concentrations 2

Dose / conc.:

750 ppm

Remarks:

Basis:
nominal in diet

Doses / concentrations 3

Dose / conc.:

1 500 ppm

Remarks:

Basis:
nominal in diet

No. of animals per sex per dose:

28/sex/group

Control animals:

yes, plain diet

Details on study design:

Initial animals (F0) were randomly assigned to groups using a computer generated stratified body weight programme. Only healthy animals with a body weight ±20% of the mean weight for each sex were included in the study.

Positive control:

Not applicable.

Examinations

Parental animals: Observations and examinations:

Detailed clinical observations were performed once each week. Observations for overt clinical signs were made twice daily on days when detailed observations were not conducted. Observations for mortality were made twice daily.
Bodyweights were recorded weekly during the pre-beeding period and during mating for both sexes. Bodyweights were recorded for females on gestational day (gd) 0, 6, 15 and 20 and on postnatal day (pnd) 0, 7, 14 and 21.
Food consumption was recorded weekly during the pre-breeding period for both sexes, and for females in 3 or 4 day intervals throughout gestation and to pnd 4.

Oestrous cyclicity (parental animals):

Not determined.

Sperm parameters (parental animals):

Not determined.

Litter observations:

All pups were weighed on postnatal day (pnd) 1, 4, 7 and 14, and at weaning (day 21).

Postmortem examinations (parental animals):

All F0 and F1 parental animals were necropsied and examined for gross lesions. Reproductive tissues and other tissues with gross lesions identified as potentially related to treatment were harvested. All these tissues from the high dose and control groups and any tissues or organs showing gross alterations from the low and mid dose groups were examined histologically. A complete gross necropsy and histopathologic examination were conducted for any parental animals dying on test.

Postmortem examinations (offspring):

Examination for gross lesions was performed on any pup appearing abnormal or dying on test. Ten F1A, F1B and F2 weanlings/sex/group were randomly selected and examined for gross lesions. Remaining nonselected F1 and F2 pups at weaning were euthanised and discarded after the necropsy of the selected pups.

Statistics:

The results of the quantitative continuous variables (e.g., body weights, food consumption, etc.) were intercompared for the three treatment groups and one control group by use of Leaven’s test for equal variances, analysis of variance and (pooled or separate variance) t-test. Non-parametric data were statistically evaluated using the Kruskal-Wallis test followed by the Mann-Whitney U test for pairwise comparisons when appropriate. Frequency data were compared using the Fisher’s exact test.

Reproductive indices:

Mating index, fertility index, gestational index, live birth index, 4-day survival index, 7-day survival index, 14-day survival index, 21-day survival index, lactation index, 28-day survival index.

Offspring viability indices:

See above.

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:

no effects observed

Dermal irritation (if dermal study):

not examined

Mortality:

no mortality observed

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

reduced at 1500 ppm a.s.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

reduced at 1500 ppm a.s.

Food efficiency:

not examined

Water consumption and compound intake (if drinking water study):

not examined

Ophthalmological findings:

not specified

Haematological findings:

no effects observed

Clinical biochemistry findings:

no effects observed

Urinalysis findings:

not specified

Behaviour (functional findings):

no effects observed

Immunological findings:

not specified

Organ weight findings including organ / body weight ratios:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

not specified

Other effects:

no effects observed

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:

not examined

Reproductive function: sperm measures:

not examined

Reproductive performance:

no effects observed

Details on results (P0)

CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS)
No significant signs of toxicity during the pre-breed, mating, gestation or lactation periods at any dose for either generation. There were no mortalities.

BODY WEIGHT AND FOOD CONSUMPTION (PARENTAL ANIMALS)
F0 and F1 10-Week Pre-mating Exposure: No treatment-related effects on body weights were observed at 300 or 750 ppm a.s.. Males and females at 1500 ppm a.s. had reductions in body weight beginning one week after treatment. Body weight gain was also reduced.
F0 and F1 Gestation/Lactation: No treatment-related effects on body weights were observed at 300 or 750 ppm a.s.. Females in the 1500 ppm a.s. group showed significant reductions in body weights but no body weight gain reductions during the first (producing the F1A and F2A litters) breeding period. Body weight but not weight gain was also reduced throughout lactation. During the second breeding (producing the F1B and F2B litters), gestational body weights and weight gains appeared to be lower than control and lactational body weights but not weight gains were reduced.

TEST SUBSTANCE INTAKE (PARENTAL ANIMALS)
F0 and F1 10-Week Pre-mating Exposure: No treatment-related effects on food consumption were observed at 300 or 750 ppm a.s.. Food consumption in males and females was reduced throughout the pre-breed periods for the 1500 ppm a.s. group.
F0 and F1 Gestation/Lactation: Food consumption during gestation and lactation for both breeding periods was unaffected by test substance treatment at any dose.

REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS)
Reproductive parameters were unaffected by treatment for all groups during the first and second breeding of both the F0 and F1 animals.

ORGAN WEIGHTS (PARENTAL ANIMALS)
No treatment-related findings were observed.

GROSS PATHOLOGY (PARENTAL ANIMALS)
No treatment-related findings were observed.

HISTOPATHOLOGY (PARENTAL ANIMALS)
No treatment-related findings were observed.

OTHER FINDINGS (PARENTAL ANIMALS)
No other findings were reported.

Effect levels (P0)

Key result

Dose descriptor:

NOAEL

Effect level:

750 ppm (nominal)

Based on:

act. ingr.

Sex:

male/female

Basis for effect level:

other: Equivalent to ca. 928 ppm test substance. Based on reduced body weights and food consumption at 1500 ppm a.s.

Results: F1 generation

General toxicity (F1)

Clinical signs:

no effects observed

Dermal irritation (if dermal study):

not examined

Mortality / viability:

no mortality observed

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

reduced at 1500 ppm a.s.

Food consumption and compound intake (if feeding study):

not specified

Food efficiency:

not examined

Water consumption and compound intake (if drinking water study):

not examined

Ophthalmological findings:

not specified

Haematological findings:

not specified

Clinical biochemistry findings:

not specified

Urinalysis findings:

not specified

Sexual maturation:

not examined

Anogenital distance (AGD):

not examined

Nipple retention in male pups:

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

no effects observed

Histopathological findings:

not examined

Other effects:

not specified

Developmental neurotoxicity (F1)

Behaviour (functional findings):

not examined

Developmental immunotoxicity (F1)

Developmental immunotoxicity:

not examined

Details on results (F1)

CLINICAL SIGNS (OFFSPRING)
There were no signs of toxicity in the F1A, F1B, F2A or F2B animals.

BODY WEIGHT (OFFSPRING)
No treatment-related effects on body weights were observed at 300 or 750 ppm. F1A litters exhibited reduced body weights and weight gains from pnd 14 through 28 at 1500 ppm. A similar effect was observed for F1B, F2A and F2B pups.

ORGAN WEIGHTS (OFFSPRING)
There were no treatment-related findings in the F1A, F1B, F2A or F2B weanling animals at necropsy.

GROSS PATHOLOGY (OFFSPRING)
There were no treatment-related findings in the F1A, F1B, F2A or F2B weanling animals at necropsy.

Effect levels (F1)

Key result

Dose descriptor:

NOAEL

Generation:

F1

Effect level:

750 ppm (nominal)

Based on:

act. ingr.

Sex:

male/female

Basis for effect level:

other: Equivalent to ca. 928 ppm test substance. Based on reduced body weights and food consumption at 1500 ppm a.s.

Results: F2 generation

Effect levels (F2)

Key result

Dose descriptor:

NOAEL

Generation:

F2

Effect level:

750 ppm (nominal)

Based on:

act. ingr.

Sex:

male/female

Basis for effect level:

other: Equivalent to ca. 928 ppm test substance. Based on reduced body weights at 1500 ppm a.s.

Overall reproductive toxicity

Key result

Reproductive effects observed:

no

Lowest effective dose / conc.:

750 ppm

Treatment related:

yes

Applicant's summary and conclusion

Conclusions:

These results indicate that continuous exposure to test item in the diet for two generations in Sprague-Dawley (CD®) rats resulted in no adverse reproductive effects. Parental toxicity was observed at 1500 ppm, limited to body weight reduction, weight gain depression and decreased food consumption. Postnatal toxicity was observed at 1500 ppm, consisted of reduced pup body weights. The adult effect levels are equivalent to 112.6 mg of test chemical per kg body weight at 1500 ppm. The "no observable effect level" (NOEL) for adults in this study was 750 ppm. The NOEL for offspring in this study was 750 ppm (males: 38 - 79 mg/kg bw/day and females: 49 - 82 mg/kg bw/day), indicating no increased risk to offspring in the absence of maternal effects.

Executive summary:

The study was carried out in accordance with EPA OPP 83-4. Sprague-Dawley rats were given diets containing Bardac 2280 ( Didecyldimethylammonium Chloride in aqueous/alcohol solution) at concentrations of 0, 300, 750 and 1500 ppm a.s.. A 10-week pre-breed exposure was used for both the F0 and F1 generations. Two litters per generation were produced. Body weights were decreased in males and females at 1500 ppm for most of the pre-breeding exposure period as well as for the F1A, F1B, F2A, and F2B offspring during lactation. Food consumption was also reduced during the pre-breeding periods for both the F0 and F1 parental animals. No other treatment-related effects were observed including on any reproductive parameters. On the basis of these results the NOAEL (parental, F1 offspring and F2 offspring) was considered to be 750 ppm a.s. (equivalent to ca. 928 ppm test substance). It was concluded that the test substance was not toxic to reproduction under the conditions of the study.