Amines, C12-18-alkyldimethyl

Endpoint summary

Key value for chemical safety assessment

Effects on fertility

Description of key information

Reliable data from guideline studies on reproduction/developmental toxicity are available for C10-DMA, C12-14-DMA, and C16-DMA. These studies were performed according to OECD TG 421/422. For C18-DMA a study according to OECD TG 422 is ongoing.

In addition, the results are supported by read-across data from several DMAOs. The two-generation fertility study and three studies according to OECD TG 422 did not reveal any effects related to reproductive toxicity. No classification according to Regulation (EC) No. 1272/2008 is warranted for the DMA category members. As key value the most conservative value for the category members is selected.

Link to relevant study records

Reference

Reference 1

Endpoint:

two-generation reproductive toxicity

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Justification for type of information:

For details on endpoint specific justification please see read-across report in section 13 or find a link in cross-reference “assessment report”.

Reason / purpose for cross-reference:

assessment report

Reason / purpose for cross-reference:

read-across source

Dose descriptor:

NOAEL

Effect level:

> 43 mg/kg bw/day (nominal)

Based on:

other: Read-across from C12 DMAO; value converted based on MW

Sex:

male/female

Remarks on result:

other: Original data for the source: >40 mg/kg bw/d

Dose descriptor:

NOAEL

Generation:

F1

Effect level:

> 43 mg/kg bw/day (nominal)

Based on:

other: Read-across from C12 DMAO; value converted based on MW

Sex:

male/female

Remarks on result:

other: Original data for the source: >40 mg/kg bw/d

Dose descriptor:

NOAEL

Generation:

F2

Effect level:

> 43 mg/kg bw/day (nominal)

Based on:

other: Read-across from C12 DMAO; value converted based on MW

Sex:

male/female

Remarks on result:

other: Original data for the source: >40 mg/kg bw/d

Reproductive effects observed:

not specified

Executive summary:

The study used as source investigated reproductive toxicity in rats. The study results of the source compound were considered applicable to the target compound. Justification and applicability of the read-across approach (structural analogue) is outlined in the read-across report in section 13 or find a link in cross reference “assessment report”.

Reference 2

Endpoint:

two-generation reproductive toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From NOV 1979 to NOV 1980

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Reason / purpose for cross-reference:

reference to same study

Principles of method if other than guideline:

Two-generation study in rats

GLP compliance:

no

Remarks:

This study was conducted prior to the adoption of GLP compliance standards. However, it was conducted in accordance with the Japanese Ministry of Health and Welfare Guidelines, and was found to be acceptable by the laboratory´s Quality Assurance Unit.

Limit test:

no

Species:

rat

Strain:

other: Charles River CD

Sex:

male/female

Details on test animals or test system and environmental conditions:

no data

Route of administration:

oral: feed

Vehicle:

not specified

Details on mating procedure:

no data

Analytical verification of doses or concentrations:

not specified

Duration of treatment / exposure:

approximately 4 months

Frequency of treatment:

daily

Details on study schedule:

no data

Remarks:

Doses / Concentrations:
0, 750, 1500, 3000 ppm for 6.5 weeks, doses were reduced to 0, 188, 375 and 750 ppm. 750 ppm corresponds to 40 mg /kg / day, 375 ppm corresponds to 20 mg/ kg / day, 188 ppm corresponds to 11 mg/ kg / day
Basis:
nominal in diet

No. of animals per sex per dose:

15 males, 30 females

Control animals:

yes, plain diet

Details on study design:

- Dose calculation according to SIDS.

Positive control:

no data

Parental animals: Observations and examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: no data

BODY WEIGHT: Yes
- Time schedule for examinations:
F0:males: weekly, females: weekly until mating, then weighed an days 1, 3, 7, 14, 21 post coitum and on days 1, 4, 11, 18 and 25 post partum
F1: days 1, 4, 11, 18, 25 post partum

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes
- 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: Yes

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No data

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day: no
- Organs examined: adrenals, all tumors, brain, bone marrow, cecum, duodenum, epididymides, eye and optic nerve, heart, ileum, kidneys, liver, lungs, lymph nodes, mammary glands, esophagus, ovaries, pancreas, pituitary, prostate, seminal vesicles, spleen, stomach, testes, thymus, thyroid, urinary bladder and uterus

Oestrous cyclicity (parental animals):

no data

Sperm parameters (parental animals):

no data

Statistics:

Multiple t-test; Mann-Whitney U-test; x2-test; or Fisher´s Exact Probability test.

Reproductive indices:

no data

Offspring viability indices:

Details for first generation (F1) and second generation (F2)given.

Clinical signs:

no effects observed

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

slight reductions in weight gain of both parents and offspring, but was without adverse effect on their mating performance and fertility

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

slight reductions in weight gain of both parents and offspring, but was without adverse effect on their mating performance and fertility

Organ weight findings including organ / body weight ratios:

not specified

Histopathological findings: non-neoplastic:

not specified

Other effects:

no effects observed

Reproductive function: oestrous cycle:

no effects observed

Reproductive function: sperm measures:

not specified

Reproductive performance:

no effects observed

Dose descriptor:

NOAEL

Effect level:

> 40 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: slight reduction in weight gain of both parents. Reductions in weight gain did not exceed 10%. The general condition of the animals throughout the study was unaffected by treatment.

Clinical signs:

no effects observed

Mortality / viability:

mortality observed, treatment-related

Description (incidence and severity):

There was a slight reduction in the number of F2 offspring born at the 750 ppm level.

Body weight and weight changes:

no effects observed

Sexual maturation:

no effects observed

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

no effects observed

Histopathological findings:

no effects observed

Dose descriptor:

NOAEL

Generation:

F1

Effect level:

> 40 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: Mating performance, fertility, and conception rate were not affected by treatment in either generation. Gestation and parturition proceeded normally.

Dose descriptor:

NOAEL

Generation:

F2

Effect level:

> 40 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: see 'Remark'

Reproductive effects observed:

not specified

Conclusions:

Under the conditions of the test the NOAEL for the parent generation was established as >40 mg/kg bw/day. NOAEL values for F1 and F2 offsprings were >40 mg/kg bw/day as well. Slight reduction in weight gain of both parents was observed. Reductions in weight gain did not exceed 10%. The general condition of the animals throughout the study was unaffected by the treatment. Mating performance, fertility, and conception rate were not affected by treatment in either generation. Gestation and parturition proceeded normally. There was a slight reduction in the number of F2 offsprings born at the 750 ppm level, however there were no adverse effects of treatment on litter size at birth, live birth index and birth weight in either generation. Therefore, this was not considered an adverse effect.

Executive summary:

15 Male and 30 female Charles River CD rats per dose group were exposed to feed containing 0, 750, 1500, 3000 ppm test substance for 6.5 weeks, doses then were reduced to 0, 188, 375 and 750 ppm. This corresonds to: 0, 11, 20 and 40 mg/kg / day ad libitum for 101 days before mating, throughout mating, gestation and lactation.

Slight reduction in weight gain of both parents was observed. Reductions in weight gain did not exceed 10%. The general condition of the animals throughout the study was unaffected by the treatment. Mating performance, fertility, and conception rate were not affected by treatment in either generation. Gestation and parturition proceeded normally. There was a slight reduction in the number of F2 offsprings born at the 750 ppm level, however there were no adverse effects of treatment on litter size at birth, live birth index and birth weight in either generation. Therefore, this was not considered an adverse effect.

Under the conditions of the test the NOAEL for the parent generation was established as >40 mg/kg bw/day. NOAEL values for F1 and F2 offsprings were >40 mg/kg bw/day as well.

Effect on fertility: via oral route

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEL

150 mg/kg bw/day

Study duration:

subchronic

Species:

rat

Quality of whole database:

good

Effect on fertility: via inhalation route

Endpoint conclusion:

no study available

Effect on fertility: via dermal route

Endpoint conclusion:

no study available

Additional information

Reliable combined repeated dose toxicity studies with the reproduction/developmental toxicity screening test according to OECD guideline 422 are available for C10-DMA, C12-14 DMA, and for C16-DMA. For C18-DMA guideline a study according to OECDT TG 422 is ongoing. For C12-14-DMA a supporting reproduction/developmental toxicity screening test (OECD guideline 421) is also available.

In the two newly available gavage studies performed in Wistar rats according to OECD TG 422 with C10-DMA, and C12-14 DMA the NOAEL for both, parental and F1 animals was 150 mg/kg bw/d, the highest dose tested. The same NOAEL was derived in the third OECD TG 422 with C16-DMA for females, while the NOAEL for males was 100 mg/(kg bw x d) based on reduced body weight and weight gain (no effects on reproductive parameters observed). No further treatment related adverse effects on systemic, reproduction and fertility parameters up to and including the highest dose could be observed in any of these studies.

In the supporting reproduction/developmental toxicity screening test (OECD guideline 421) for C12-14-DMA, Sprague-Dawley rats were dosed with 0, 50, 150, 300 and 450 mg/kg bw/d by gavage. The NOEL for both parent animals and their pups was 50 mg/kg bw/d. 150 mg/kg bw/d were in the beginning lethal range for pregnant rats and in the lethal range for embryos/fetuses/pups. No effects were observed on estrous cycle and reproductive performances. However, at 150 mg/kg bw/d parturition of two dams was influenced. In addition, as outlined in the section on repeated dose toxicity, at 150 mg kg bw/d lethality was already reported (2/10 females died). The NOEL for fertility is 50 mg/kg bw/d.

Overall, in all available studies, no indication of any effect on fertility, reproduction, gestation, partition, growth of offspring or lactation was found beyond the dose levels of apparent maternal toxicity.

Based on these observations, the DMAs of this category do not have to be classified for effects on fertility according to Regulation (EC) No 1272/2008.

Effects on developmental toxicity

Description of key information

Reliable data from guideline studies on reproduction/developmental toxicity are available for C10-DMA, C12-14-DMA and C16-DMA. These studies were performed according to OECD TG  422. No indication of developmental toxicity was found. For C18-DMA a study according to OECD TG 422 is ongoing. And either for C12-14 DMA or C16-DMA a OECD TG 414 is planned. In addition, the unlikeliness of developmental toxicity for DMA is supported by read-across data from several DMAOs. A developmental toxicity study with C12-14 DMAO is currently used to cover the information requirements for this endpoint. The three new studies according to OECD TG 422 did not reveal any effects related do developmental toxicity. No classification according to Regulation (EC) No. 1272/2008 is warranted for the DMA category members.

Link to relevant study records

Reference

Reference 1

Endpoint:

developmental toxicity

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Justification for type of information:

For details on endpoint specific justification please see read-across report in section 13 or find a link in cross-reference “assessment report”.

Reason / purpose for cross-reference:

assessment report

Reason / purpose for cross-reference:

read-across source

Species:

rabbit

Dose descriptor:

NOAEL

Effect level:

> 171 mg/kg bw/day (actual dose received)

Based on:

other: Read-across from C12 DMAO; value converted based on MW

Remarks on result:

other: Original data for the source: >160 mg/kg bw/d

Dose descriptor:

NOAEL

Effect level:

> 171 mg/kg bw/day (actual dose received)

Based on:

other: Read-across from C12 DMAO; value converted based on MW

Remarks on result:

other: Original data for the source: 160 mg/kg bw/d

Abnormalities:

not specified

Developmental effects observed:

not specified

Executive summary:

The study used as source investigated developmental toxicity in rabbits. The study results of the source compound were considered applicable to the target compound. Justification and applicability of the read-across approach (structural analogue) is outlined in the read-across report in section 13 or find a link in cross reference “assessment report”.

Reference 2

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From MAY 1980 to OCT 1980

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Principles of method if other than guideline:

Test for developmental toxicity.

GLP compliance:

no

Remarks:

This study was conducted prior to adoption of GLP compliance standards. However, it was conducted in accordance with the Japanese Ministry of Health and Welfare Guidelines, and was found to be acceptable by the laboratory´s Quality Assurance Unit

Limit test:

no

Species:

rabbit

Strain:

New Zealand White

Details on test animals or test system and environmental conditions:

no data

Route of administration:

oral: gavage

Vehicle:

not specified

Details on exposure:

no data

Analytical verification of doses or concentrations:

not specified

Details on mating procedure:

Animals were artificially inseminated using pooled semen from New Zealand White bucks.

Duration of treatment / exposure:

days 6-18 (inclusive) of gestation

Frequency of treatment:

daily

Duration of test:

approximately 5 months

No. of animals per sex per dose:

14 female animals per dose group

Control animals:

yes, concurrent vehicle

Details on study design:

no data

Maternal examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily

DETAILED CLINICAL OBSERVATIONS: No data

BODY WEIGHT: Yes
- Time schedule for examinations: daily, reported on days 0, 6, 8, 10, 12, 14, 16, 18, 23, 28.

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

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 29
- Organs examined: no data

Ovaries and uterine content:

The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: No
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
- Other:

Fetal examinations:

- External examinations: Yes
- Soft tissue examinations: No
- Skeletal examinations: Yes: (all)
- Head examinations: No

Statistics:

Multiple ´t´-test, ´t´-test, Mann-Whitney U-test, Chi-squared test and Fisher´s Exact Probability test (Armitage modification).

Indices:

no data

Historical control data:

no data

Details on maternal toxic effects:

Maternal toxic effects:yes. Remark: details see below

Details on maternal toxic effects:
Maternal condition was generally unaffected by treatment with the test material; however, maternal bodyweight gain was depressed in all treated groups, although at 40 mg/kg/day terminal bodyweights similar to those of controls were achieved. Three females receiving 80 mg/kg/day and three females receiving 160 mg/kg/day died or were killed in extremis, but no direct involvement of the test substance was apparent. Food intake, when compared with pre-treatment values, was reduced during the second half of the treatment period in groups receiving 40 and 80 mg/kg/day, and from the commencement of treatment in animals receiving 160 mg/kg/day. Water intake was also decreased in all treated groups. Reductions in body weight gain, food intake and water intake did not exceed 10%.

Dose descriptor:

NOAEL

Effect level:

> 160 mg/kg bw/day (actual dose received)

Based on:

test mat.

Basis for effect level:

other: maternal toxicity

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
Litter response was unaffected by any treatment with the test material. No adverse effects upon litter responses and development were recorded. No teratogenic responses were observed.

Dose descriptor:

NOAEL

Effect level:

> 160 mg/kg bw/day (actual dose received)

Based on:

test mat.

Basis for effect level:

other: teratogenicity

Abnormalities:

not specified

Developmental effects observed:

not specified

Conclusions:

Under the conditions of the test, the maternal animals were generally unaffected by treatment with the test material; however, maternal bodyweight gain was depressed in all treated groups, although at 40 mg/kg/day terminal bodyweights similar to those of controls were achieved. Food intake, when compared with pre-treatment values, was reduced during the second half of the treatment period in groups receiving 40 and 80 mg/kg/day, and from the commencement of treatment in animals receiving 160 mg/kg/day. Water intake was also decreased in all treated groups. Reductions in body weight gain, food intake and water intake did not exceed 10%. The NOAEL for maternal toxicity was established as > 160 mg/kg bw/day
Litter response was unaffected by any treatment with the test material. No adverse effects upon litter responses and development were recorded. No teratogenic responses were observed. The NOAEL for teratogenicity was established as > 160 mg/kg bw/day

Executive summary:

14 New Zealand white rabbits per dose group were exposed to 0, 40, 80 or 160 mg test substance/kg bw/day during days 6 -18 og gestation.

Under the conditions of the test, the maternal animals were generally unaffected by treatment with the test material; however, maternal bodyweight gain was depressed in all treated groups, although at 40 mg/kg/day terminal bodyweights similar to those of controls were achieved. Food intake, when compared with pre-treatment values, was reduced during the second half of the treatment period in groups receiving 40 and 80 mg/kg/day, and from the commencement of treatment in animals receiving 160 mg/kg/day. Water intake was also decreased in all treated groups. Reductions in body weight gain, food intake and water intake did not exceed 10%.

Litter response was unaffected by any treatment with the test material. No adverse effects upon litter responses and development were recorded. No teratogenic responses were observed.

The NOAEL for maternal toxicity was established to be > 160 mg/kg bw/day.

The NOAEL for teratogenicity is > 160 mg/kg bw/day.

Reference 3

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

N/A

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Guideline study (EPA OTS) and according to GLP.

Qualifier:

according to guideline

Guideline:

EPA OTS 798.4900 (Prenatal Developmental Toxicity Study)

GLP compliance:

yes

Limit test:

no

Species:

rat

Strain:

Sprague-Dawley

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Inc., Raleigh, North Carolina.
- Age at study initiation: Females: 65 days; Males: 42 days.
- Weight at study initiation: Females: 218 - 262 g; Males: 506 - 969 g.
- Housing: Rats were individual housed except during cohabitation period. During cohabitation, each pair of male and female rats was housed in the male rat's cage.
- Diet (ad libitum): Certified Rodent Diet #5002 (Purina Nutrition International, St. Louis, Missouri) in individual feeders.
- Water (ad libitum): Local water that had been processed by passage through a reverse osmossis membrane (R.O. water) was available to the rats from an automatic watering system and/or individual water bottles. Chlorine was added to the processed water as a bacteriostat.
- Acclimation period: N/A


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18 to 26
- Humidity (%): 30 to 70
- Air changes (per hr): 10/hr
- Photoperiod (hrs dark / hrs light): 12/12


IN-LIFE DATES: From: N/A To: N/A

Route of administration:

oral: gavage

Vehicle:

other: Sterile Water

Details on exposure:

PREPARATION OF DOSING SOLUTIONS: Solutions of the test substances were prepared weekly at the Testing Facility. Dosage calculations were adjusted for the 32% (w/v) concentration of the test substance. Prepared formulations were stored at room temperature and stirred continuously (magnetic stir plate with stir bar) during dosage administration.


DIET PREPARATION
- Rate of preparation of diet (frequency): Weekly
- Storage temperature of food: Room temperature.


VEHICLE
- Justification for use and choice of vehicle (if other than water): Sterile Water
- Amount of vehicle (if gavage): 5 mL/kg

Analytical verification of doses or concentrations:

not specified

Details on mating procedure:

- Impregnation procedure: cohoused
- If cohoused:
- M/F ratio per cage: 1/1
- Length of cohabitation: 5 days
- Proof of pregnancy: sperm in vaginal smear and/or a copulatory plug observed in situ will be referred to as day 0 of pregnancy

Duration of treatment / exposure:

Days 6 through 19 of presumed gestation.

Frequency of treatment:

Daily

Duration of test:

Approximately 4 weeks.

No. of animals per sex per dose:

25

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: Dosages were selected on the basis of a dosage-range study (Angus Research Laboratories, Inc., Protocol 916-025P), in which dosage levels of 0, 32.5, 100, 325 and 650 mg/kg/day were evaluated.
Dosages of 325 and 650 mg/kg/day were excessively toxic to the dams and conceptuses. At the 32.5 and 100 mg/kg/day dosage levels, no developmental toxicity was observed. Excess salivation was observed at the 100 mg/kg/day dosage level, and slight, non-statistically significant decreases in maternal body weight gains and feed consumption values were observed at the 32.5 and 100 mg/kg/day dosage levels. Doses of 200, 100, and 25 mg akly dimethyl amine oxide/kg/day were chosen.
- Rationale for animal assignment (if not random): Unpon arrival, rats were assigned to individual housing on the basis of computer-generated random units. Female rats were assigned to four dosage groups (Groups I through IV), twentry-five rats per dosage group, using a computer-generated (weight-ordered) randomization procedure based on body weights recorded on DG 0.

Maternal examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice a day
- Cage side observations checked were: mortality, moribundity, pertinent behavioral changes and other signs of overt toxicity.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Weekly and Rats were also examined for clinical observations immediately before and approximately 60 minutes after dosage and on the day of sacrifice (DG 20).

BODY WEIGHT: Yes
- Time schedule for examinations: Weekly during acclimation, on DG 0, daily during the dosage period and on DG 20.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes
- 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: Yes

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No
- Time schedule for examinations: N/A

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day # 20

Ovaries and uterine content:

The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
- Other: A late resorption was defined as one in which the occurrence of organogenesis was grossly evident. A live fetus was defined as a term fetus that responded to stimuli. Nonresponding term fetuses are considered to be dead (there were no dead fetuses). Dead fetuses and late resorptions are differentiated by the degree of autolysis present; marked to extreme autolysis indicated that the fetuses was a late resorption.

Fetal examinations:

- External examinations: Yes: half per litter
- Soft tissue examinations: Yes: half per litter
- Skeletal examinations: Yes: half per litter
- Head examinations: Yes: half per litter

Statistics:

Clinical observation and other proportion data were analyzed using the Variance Test for Homogeneity of the Bionomial Distribution.
Continuous data (e.g., body weights, body weight changes, feed consumption values, organ weights and litter averages for percent male fetuses, percent resorbed conceptuses, fetal body weights and feal anomaly data) were analyzed using Bartletts' Test of Homogeneity of Variances and the Analysis of Variance, when appropriated [i.e., Bartlett's Test was not significant (pCount data obtained at Caesarean-sectioning of the dams were evaluated using the procedures described above for the Kruskal-Wallis Test.

Details on maternal toxic effects:

Maternal toxic effects:yes. Remark: details see below

Details on maternal toxic effects:
MORTALITY:
Two rats in the 200 mg/kg/day dosage group died; one of these deaths was attributed to the test substance, the other was the result of an intubation error. All other rats survived until scheduled sacrifice.
A single rat in the 200 mg/kg/day dosage group was found dead on day 19 of gestation (DG 19) after 13 daily dosages. This dam lost body weight after DG 9 and had reduced feed consumption values throughout the study. Adverse clinical observations included excessive salivation (DGs 8 to 9, 14 and 17 to 18), gasping (DGs 9 to 11 and 17 to 18), labored breathing (DGs 10 to 13 and 16 to 18), ungroomed coat (DGs 10 to 13), brown perianal substances (DGs 12 to 13), urine-stained abdominal fur (DGs 12 to 14 and 16 to 18), brown perivaginal substance (DGs 12 to 15), chromorhinorrhea (DG 15), rales (DG 16) and emaciation (DGs 17 to 18). No gross lesions were revealed by necropsy; all tissues appeared normal for moderate degree autolysis. There were 17 early resorptions in utera. This death was attributed to effects of the test substance because similar observations occurred in surviving rats in this dosage group.
A single rat in the 200 mg/kg/day dosage group was found dead on DG 18 after 12 daily dosages. This death was attributed to an intubation accident. This dam lost body weight after DG 13 and had reduced feed consumption values throughout the study. Adverse clinical observations included rales (DGs 10 to 11 and 13), excessive salivation (DGs 13 to 14), gasping (DGs 13 to 14), labored breathing (DGs 13 to 17), red perioral substance (DGs 15 to 17), urine-stained abdominal fur (DGs 15 to 17), dehydration (DG 16) and emaciation (DG 17). External observations at necropsy included red substance on fur of the nose, forepaws and forelimbs. Gross necropsy revealed a tear in the esophagus; all other tissues appeared normal for slight degree of autolysis. There were 14 fetuses in utero. The viability of the fetuses could not be determined because of the maternal death.

CLINICAL OBSERVATIONS:
Adverse clinical observations occurred at increased incidences in the 100 and 200 mg/kg/day dosage group rats. Excessive salivation, rales, urine-stained abdominal fur, brown or red perioral substance, labored breathing and gasping occurred in 100 mg/kg/day dosage group rats and in significantly increased (pAll other clincal observations were considered unrelated to the test substance because: 1) the incidences were not dosage-dependent; and/or 2) the observations occurred in only one or two rats. These clinical observations included one 200 mg/kg/day dosage group dam with an axillary mass attributed to an intubation error, and localized alopecia (limbs and underside) in one or two rats in the 0 (Vehicle), 25, and 200 mg/kg/day dosage groups.

NECROPSY OBSERVATIONS:
All necropsy observations were considered unrelated to the test substance because they were single events. These observations included slight dilation of the pelvis of the right kidney of one vehicle group dam and one 200 mg/kg/day dosage group dam. A single dame also had a distended urinary bladder with ten calculi and thickened and red bladder walls. One 100 mg/kg/day dosage group dam had large adrenals, four dark red areas on the fundic mucosa and numerous raised tan areas on the pyloric mucosa of the stomach, the intestines and stomach were distended with gas and the left lateral lobe of the liver was mottled. One 200 mg/kg/day dosage dam had a mass in the left axilla in-life; at necropsy, a clear tan gelatinous fluid was located subcutaneously in the area of the ventral neck, left axilla and leateral chest, the esophagus had a thickened area and the plyoric folds of the stomach were thickened. These observation were presumed to be the sequelae of a presumed intubation error.

MATERNAL BODY WEIGHTS, GRAVID UTERINE WEIGHTS AND BODY WEIGHT CHANGES:
Rats in the 100 and 200 mg/kg/day dosage groups had significantly reduced (pThe gravid uterine weight and the corrected DG 20 maternal body weight (DG 20 body weight minus the gravid uterine weight) were significantly reduced (pBody weights and body weights gains were unaffected at dosages of 25 mg/kg/day of the test substance. Gravid uterine weights were unaffected by the 25 and 100 mg/kg/day dosages of the test substance.

MATERNAL ABSOLUTE (g/day) and RELATIVE (g/kg/day) FEED CONSUMPTION VALUES:
Absolute (g/day) feed consumption values for the entire dosage period (calculated as DGs 6 to 20) and the entire gestation period (DGs 0 to 20) were significantly reduced (p
CAESAREAN-SECTIONING:
Caesarean-sectioning observations were based on 24, 25, 24 and 22 pregnant rats in the 0 (vehicle), 25, 100 and 200 mg/kg/dosage groups, respectively. Male and female fetal body weights were significantly reduced (pNo other Caesarean-sectioning or litter parameters were affected by dosages of the test substance as high as 200 mg/kg/day. The litter averages for corpora lutea, implantations, late resorptions, percent resorbed conceptuses (calculated excluding the completely resorbed litter in the 200 mg/kg/day dosage group), and percent live male fetuses were comparable among the four dosage groups and did not significantly differ. There were no dead fetuses.

Dose descriptor:

NOAEL

Effect level:

25 mg/kg bw/day

Based on:

test mat.

Basis for effect level:

other: maternal toxicity

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:yes. Remark: details see below

Details on embryotoxic / teratogenic effects:
FETAL ALTERATIONS:
Fetal alterations were defined as: 1) malformations (irreversible changes that occur at low incidences in this species and strain); or 2) variations (common findings in this species and strain and reversible delays or accelerations in development). Litter averages were calculated for specific fetal ossification sites as part of the evaluation of the degree of fetal ossification.
Fetal evaluations were based on 339, 375, 348, and 297 Ceasarean-delivered live fetuses in 24, 25, 24, and 21 litters in the 0 (vehicle), 25, 100 and 200 mg/kg/day dosage groups, respectively. Each fetus was examined for gross external alterations, approximately one half of the fetuses in each litter were examined for soft tissue alterations and the remaining fetuses were examined for skeletal alterations and the number of ossification sites.
The percentages of fetuses and litters with alterations in the 200 mg/kg/day dosage group were significantly increased (pAll other gross external, soft tissues or skeletal alterations (malformations or variations) were considered unrelated to the test substance because 1) the litter and fetal incidences were not dosage-dependent; 2) the alteration occurred in only one fetus; or 3) the incidences were within the ranges observed historically at the Test Facility.

FETAL ALTERATIONS:
In groups I through IV, litters with fetuses with alterations numbered 8 (33.3%), 11 (44.0%), 8 (33.3%), and 15 (71.4%), respectively. The numbers of fetuses with any alterations were 11 (3.2%), 25 (6.7%), 14 (4.0%) and 32 (10.8%) respectively. The percentages of fetuses with any alteration per litter were 3.6%, 6.4%, 4.5% and 11.5% in the four respective dosage groups.
The significant increases in the percentages of fetuses and litters with alterations in the 200 mg/kg/day dosage group were considered to reflect delays in skeletal ossification, related to the significantly reduced (p
FETAL GROSS EXTERNAL ALTERATIONS:
One vehicle group fetus had whole body edema (anasarca); this fetus also had an absent innominate artery at soft tissue examination. One 25 mg/kg/day fetus had a thread-like tail; at skeletal evaluation, this fetus had fused arches of the 3rd sacral vertebra and no caudal vertebrae. One 200 mg/kg/day group fetus had a kinked tail, the 4th and 5th digits of the lift hindlimb were fused and a skin tab located to the left of its tail. At skeletal evaluation of this fetus, further evaluation of the skin tab revealed two bones (possibly a femur and fibula) fused together on the left side of the pelvis. An extra paw appeared to be attached to the underside of the left paw; also the centra of the 6th and 11th thoracic vertebrae were bifid. One 200 mg/kg/day fetus had a micrognathia; soft tissue evaluation of this fetus revealed a small tongue.

MALFORMATIONS - SOFT TISSUE EVALUATION:
One 200 mg/kg/day fetus had a small tongue; micrognathia was identified at gross external evaluation.

VARIATIONS - SOFT TISSUE EVALUATION:
One control group fetus and one 25 mg/kg/day dosage group fetus had an absent innominate vessel; whole body edema (anasarca) was identified at gross external evaluation for the control fetus, also an additional fetus had the umbilical artery descending to the left of the urinary bladder.

MALFORMATION - SKELETAL ALTERATIONS:
One control group fetus had fused arches of the 4th cervical vertebra; additional variations in ossification occurred in this fetus (unossified 1st sternal centra, incompletely ossified pubes).
One 25 mg/kg/day dosage group fetus had fused arches of the 3rd sacral vertebra; this fetus also had no caudal vertebrae (thread-like tail was noted at gross external examination).

VARIATIONS - SKELETAL ALTERATIONS:
A cervical rib was present at the 7th vertebra in one 25 mg/kg/day fetus. This fetus had no other skeletal alterations.
A bifid centrum in the thoracic vertebrae occurred in 1, 6, 5 and 9 fetuses from 1, 3, 5 and 8 litters in the 0 (vehicle), 25, 100 and 200 mg/kg/day dosage groups respectively. One fetus in the 25 mg/kg/day dosage group also had incompletely ossified 1st and 2nd sternal centra, a fetus in the 100 mg/kg/day dosage group also had a bifid centrum of the 1st lumbar vertebra and a fetus in the 200 mg/kg/day dosage group also had incompletely ossified pubes.
An incompletely ossified arch in the 6th lumbar vetrebra occurred in one 100 mg/kg/day dosage group fetus; this fetus also had incompletely ossified ischia and pubes.
The significant increase (pDelayed sternal ossification (incompletely ossified and/or not ossified 1st and/or 2nd sternebrae) occurred in 4, 8, 3 and 11 fetuses from 4, 6, 2 and 7 litters in the 0 (vehicle), 25, 100 and 200 mg/kg/day dosage groups, respectively. The fetal incidence of only incompletely ossified 1st sternal centra was also significantly increased (pThe significant increases (pThe pubes and/or ischia were incompletely ossified in 8, 12, 7 and 16 fetuses from 6, 4, 3 and 6 litters in the 0 (vehicle), 25, 100 and 200 mg/kg/day dosage groups, respectively. The fetal incidenc of only incompletely ossified pubes was significantly increased (pThe significant increase (pThe litter averages for ossified caudal vertebrae, sternal centers and metacarpals per fetus were significantly decreased (pAnalyses of the average numbers of fetal ossification sites per fetus did not reveal any other statistically significant differences among the four dosage groups. Ossification of the hyoid, vertebrae (cervical, thoracic, lumbar, and sacral) ribs, sternum (manubrium and xiphoid), forelimbs (carpals and phalanges) and hindlimbs (tarsals, metatarsals and phalanges) occurred at similar incidences in litter in all dosage groups.

Dose descriptor:

NOAEL

Effect level:

25 mg/kg bw/day

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: delays in skeletal ossification

Abnormalities:

no effects observed

Developmental effects observed:

yes

Lowest effective dose / conc.:

100 mg/kg bw/day

Treatment related:

yes

Relation to maternal toxicity:

developmental effects as a secondary non-specific consequence of maternal toxicity effects

Dose response relationship:

yes

Relevant for humans:

no

          

Conclusions:

On the basis of these data, the maternal no-observable-adverse-effect-level (NOAEL) for the test substance (alkyl dimethyl amine oxide) is 25 mg/kg/day. The 200 mg/kg/day dosage caused mortality in one dam, the 100 and 200 mg/kg/day dosages caused adverse clinical observations, reductions in body weight gain and reduced feed consumption values. Reductions in relative feed consumption values were also noted in the 25 mg/kg/day dose group; however, no concomitant adverse effects were noted at this dosage level. The developmental NOAEL was 25 mg/kg/day; the 200 mg/kg/day dosage caused reduced fetal body weights and associated delays in skeletal ossification and the 100 mg/kg/day dosage also caused delays in skeletal ossification.

Executive summary:

One-hundred Crl:CDBR VAF/Plus presumed pregnant female rats were randomly assigned to four dosage groups (Groups 1 through IV), 25 rats per group. The test substance preparations for dosing were corrected for purity. The test substance, was administered orally (via gavage) once daily to these female rats on days 6 through 19 of presumed gestation (DGs 6 through 19), at dosages of 0 (Vehicle), 25, 100, and 200 mg alkyl dimethyl amine oxide/kg/day. The dosage volume was 5 mL/kg, adjusted daily on the basis of the individual body weights recorded before intubation. The rats were intubated at approximately the same time each day.

The rats were observed for viability at least twice a day and for general appearance weekly during acclimation and on DG 0. The rats were also examined for clinical observations of effects of the test substance, abortions, premature deliveries and deaths immediately before and approximately 60 minutes after dosage and on the day of sacrifice (DG 20). Body weights were recorded weekly during acclimation, on DG 0, daily during the dosage period and on DG 20. Feed consumption values were recorded on DGs 0, 6, 9, 12, 15, 18, and 20.

All surviving rats were sacrificed on DG 20, and a gross necropsy of the thoracic, abdominal and pelvic viscera was performed. Caesarean-sectioning and subsequent fetal observations were conducted without knowledge of dosage group in order to minimize bias. The number of corpora lutea in each ovary was recorded. The uterus of each rat was excised and examined for pregnancy, number and distribution of implantations, live and dead fetuses and early and late resorptions. The gravid uterus was weighed. Each fetus was removed from the uterus and subsequently weighed and examined for sex and gross external alterations. Approximately one-half of the fetuses in each litter were examined for soft tissue alterations using a variation of the microdissection techniques. The remaining fetuses in each litter were examined for skeletal alterations.

Two rats in the 200 mg/kg/day dosage group died; one of these deaths was attributed to the test substance, the other was the result of an intubation error. All other rats survived until scheduled sacrifice.

Excessive salivation, rales, urine-stained abdominal fur, brown or red perioral substance, labored breathing and gasping occurred in 100 mg/kg/day dosage group rats and in significantly increased numbers of 200 mg/kg/day dosage group rats. Additionally, chromorhinorrhea occurred in one and two rats in these two dosage groups, respectively. Observations of brown or red perivaginal substance, emaciation, brown perianal or perinasal substance, dehydration, ungroomed coat and soft or liquid feces occurred in one or two rats in the 200 mg/kg/day dosage group. All necropsy observations were considered unrelated to the test substance.

Rats in the 100 and 200 mg/kg/day dosage groups had significantly reduced body weight gains for the entire dosage period (calculated as days 6 to 20 of gestation) and body weight gains were significantly reduced in the 200 mg/kg/day dosage group for the entire gestation period (DGs 0 to 20). Body weights were significantly reduced in the 200 mg/kg/day dosage group on DGs 11 through 20. The gravid uterine weight and the corrected DG 20 maternal body weight (DG 20 body weight minus the gravid uterine weight) were significantly reduced in the 200 mg/kg/day dosage group.

Absolute (g/day) and relative (g/kg/day) feed consumption values for the entire dosage period (calculated as DGs 6 to 20) and the entire gestation period (DGs 0 to 20) were significantly reduced in the 200 mg/kg/day dosage group. Relative feed consumption values for the entire dosage period were also significantly reduced in the 100 mg/kg/day dosage group, while values for the gestation period were significantly reduced in the 25, 100 and 200 mg/kg/day dosage groups.

Male and female fetal body weights were significantly reduced in the 200 mg/kg/day dosage group. Live litter size was decreased and the number of early resorptions was increased in the 200 mg/kg/day dosage group, but apparently as the result of one dam in this dosage group that had a litter consisting of 16 early resorptions.

The percentages of fetuses and litters with alterations in the 200 mg/kg/day dosage group were significantly increased and reflected delays in skeletal ossification related to the significantly reduced fetal body weights in this group. These delays in ossification included significant increases in the fetal and/or litter incidences of bifid thoracic vertebrae centra, incompletely and/or not ossified 1st or 2nd sternal centra, incompletely ossified pubes and significant decreases in the numbers of ossified caudal vertebrae, sternal centers and metacarpals. Additionally, delays in ossification occurred in the 100 mg/kg/day dosage group and included a significant increase in the litter incidence of bifid thoracic vertebrae centra.

NOAEL for both maternal toxicity and developmental toxicity is 25 mg/kg/day.

Reference 4

Endpoint:

developmental toxicity

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Justification for type of information:

For details on endpoint specific justification please see read-across report in section 13 or find a link in cross-reference “assessment report”.

Reason / purpose for cross-reference:

assessment report

Reason / purpose for cross-reference:

read-across source

Species:

rat

Dose descriptor:

NOAEL

Effect level:

26 mg/kg bw/day

Based on:

other: Read-across from C12-14 DMAO; value converted based on MW

Remarks on result:

other: Original data for the source: 25 mg/kg bw/d

Dose descriptor:

NOAEL

Effect level:

26 mg/kg bw/day

Based on:

other: Read-across from C12-14 DMAO; value converted based on MW

Basis for effect level:

other: Original data for the source: 25 mg/kg bw/d

Abnormalities:

no effects observed

Developmental effects observed:

yes

Lowest effective dose / conc.:

104 mg/kg bw/day

Treatment related:

yes

Relation to maternal toxicity:

developmental effects as a secondary non-specific consequence of maternal toxicity effects

Dose response relationship:

yes

Relevant for humans:

no

Overall developmental toxicity: Value reported above is based on read-across from C12-14 DMAO, the value is converted based on molecular weight (value for the source substance: 100 mg/kg bw/d).

Executive summary:

The study used as source investigated developmental toxicity in rats. The study results of the source compound were considered applicable to the target compound. Justification and applicability of the read-across approach (structural analogue) is outlined in the read-across report in section 13 or find a link in cross reference “assessment report”.

Effect on developmental toxicity: via oral route

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEL

25 mg/kg bw/day

Study duration:

subacute

Species:

rat

Quality of whole database:

good

Effect on developmental toxicity: via inhalation route

Endpoint conclusion:

no study available

Effect on developmental toxicity: via dermal route

Endpoint conclusion:

no study available

Additional information

For C10-DMA, C12-14 DMA and C16-DMA reliable combined repeated dose toxicity studies with the reproduction/developmental toxicity screening test according to OECD guideline 422 are available. For for C18-DMA an OECD TG 422 is ongoing. Either for C12-14 DMA or C16-DMA an OECD TG 414 is planned. For C12-14-DMA a supporting reproduction/developmental toxicity screening test (OECD guideline 421) is also available.

Currently the information requirements for this endpoint can be fulfilled by read across from DMAOs, for which a qualified and reliable OECD Guideline 422 study (C12-18-DMAO) and a reliable developmental toxicity study according to EPA OTS 798.4900 (C12-14-DMAO) are available.

In addition, two developmental toxicity studies (one in rats for C10-16-DMAO, and one in rabbits for C10-16-DMAO) as well as a two- and a three-generation study for DMAO, are reported.

In the reproduction/developmental toxicity screening test (OECD guideline 422) for C10-DMA, C12-14 DMA and C16-DMA Wistar rats were exposed via gavage to 15, 50, 100 or 150 mg/kg bw/d. No treatment-related effects on implantation and mean litter size were observed. In addition, no external abnormalities in live or dead pups, no treatment-related changes in the ano-genital distance, ano-genital ratio or pup body weights were observed in any of the dose groups. The male pups did not exhibit areola/nipple retention on LD 13 at any of the doses tested. Therefore, the NOAEL for developmental toxicity in these studies is ≥ 150 mg/kg bw/d, the highest dose tested.

In the reproduction/developmental toxicity screening test (OECD guideline 421) for C12-14-DMA, rats were dosed with 0, 50, 150, 300 and 450 mg/kg bw/d by gavage. The NOEL for both parent animals and their pups was 50 mg/kg bw/d. 150 mg/kg bw/d were in the beginning lethal range for pregnant rats and in the toxic range for embryos/fetuses/pups. At 150 mg/kg bw/d, the number of pups alive was reduced. However, effects on pups were observed in a dose range where maternal toxicity was already evident.. Regarding developmental toxicity the effects on the pubs were clearly secondary to maternal toxicity in the OECD 421 study.

In the reliable developmental toxicity study with C12-14-DMAO (reliability 1, according to EPA OTS 798.4900) a lower NOAEL of 25 mg/kg bw/d for maternal and developmental toxicity based on adverse clinical observations, reduction in body weight gain and reduced feed consumption in dams as well as reduced fetal body weights and associated delays in skeletal ossification in the higher dose groups (100 and 200 mg/kg bw/d) in pups is reported.

In the supporting study with C12-18-DMAO (OECD TG 422), a developmental NOEL of 100 mg/kg bw/d due to pre-implantation loss in the higher dose group (200 mg/kg bw/d) was determined. No other effects on development of pups were observed. The NOEL for maternal toxicity was 40 mg/kg bw/d, due to local and systemic toxicity at 100 mg/kg bw/d or higher doses.

Two additional studies on DMAOs support the data presented so far with NOAEL values for developmental toxicity of 100 mg/kg bw/d (rats, Lion Corporation, 1979B) and >160 mg/kg bw/d (rabbits, Lion Corporation, 1980):

In the study by Lion Corporation, (1979B), rats were exposed to 0, 50, 100 or 200 mg C10-16-DMAO/kg bw/d from day 7 to 17 of gestation. In pups from females killed on day 20 of gestation, mean fetal weight was depressed at the highest dose level, and this was associated with a slight retardation of fetal ossification. No other effects were observed. The NOAEL for maternal toxicity and teratogenicity was 100 mg/kg bw/d.

In a study on developmental toxicity in rabbits (Lion Corporation, 1980), animals were exposed to 0, 40, 80 or 160 mg C10-16-DMAO/kg bw/d during days 6-18 of gestation. Litter response was unaffected by any treatment with the test material. No adverse effects upon litter responses and development were recorded. No teratogenic effects and no maternal toxicity were observed. The NOAEL for maternal toxicity and teratogenicity in this study was 160 mg/kg bw/d.

The results of the two- and the three-generation studies support the findings reported above. In the two-generation study, rats were exposed to 0, 11, 20 and 40 mg C10-16-DMAO/kg bw/d. The NOAEL values for the parent, the F1 and the F2 generation were 40 mg/kg bw/d. In the three-generation study, rats were exposed to 0, 0.005, 0.02 and 0.1% C10-16-DMAO in diet. No adverse effects were observed at any dose level, resulting in a NOEL of 0.1%, the highest concentration tested (40 mg/kg bw/d for males and 50 mg/kg bw/d for females).

In all available studies no indication of developmental toxicity was found beyond the dose levels of apparent maternal toxicity.

Based on these observations, the DMAs of this category do not have to be classified for developmental effects according to Regulation (EC) No 1272/2008.

Justification for classification or non-classification

Based on the observation that effects on fertility and developmental toxicity were found beyond the dose levels of apparent maternal toxicity, the DMAs of this category do not have to be classified for reproductive toxicity and effects on or via lactation according to Regulation (EC) No 1272/2008.