N-[3-(dimethylamino)propyl]docosanamide

Endpoint summary

Key value for chemical safety assessment

Toxic effect type:

dose-dependent

Effects on fertility

Description of key information

- oral reproduction / developmental toxicity screening test, 10 Wistar rats/sex/dose by gavage at dose levels of 0, 20, 70 and 200 mg/kg bw/d; Males were exposed for 28 days, Females were exposed for 41 – 54 days, according to OECD guideline 421, GLP; NOAEL(development) = 200 mg/kg; NOAEL(parental toxicity) = 70 mg/kg bw/d (reduced body weight gain/food consumption at 200 mg/kg bw/d); NOAEL(male fertility) = 200 mg/kg bw/d; NOAEL(female fertility) = 70 mg/kg bw/d (reduced number of implantation sites at 200 mg/kg bw/d); read-across: Stearic acid 3-(dimethylaminopropyl)amide

- Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test, 10 Wistar rats/sex/dose by gavage at dose levels of 0, 5, 15 and 45 mg/kg bw/d; Males were exposed for 28 days, Females were exposed for 54 days, according to OECD guideline 422, GLP; NOAEL(reproduction/development) = 45 mg/kg; NOAEL(parental toxicity) = 15 mg/kg bw/d (increased salivation, reduced body weight gain at 45 mg/kg bw/d); read-across: N-[3-(dimethylamino)propyl] C12-C18 alkylamide

Link to relevant study records

Reference

Endpoint:

screening for reproductive / developmental toxicity

Type of information:

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

Adequacy of study:

key study

Justification for type of information:

REPORTING FORMAT FOR THE ANALOGUE APPROACH

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
This read-across is based on the hypothesis that source and target substances have similar toxicological properties because
• they are manufactured from similar or identical precursors under similar conditions
• they share structural similarities with common functional groups: tertiary amines, amides, and fatty acid chains with comparable length and degree of saturation (corresponding to scenario 2 of the read-across assessment framework)

The read-across hypothesis is based on structural similarity of target and source substances. The target and source chemicals have a very similar structure in that they are comprised of a hydrophobic (alkyl) and hydrophilic (amine headgroup) end. Due to this motif they form micelles (colloidal dispersions) and have surfactant properties.
Based on available experimental data, including key physicochemical properties and data from genotoxicity studies, the read-across strategy is supported by a similar toxicological profile of all substances.

Therefore, read-across from the existing toxicity studies conducted with the source substances is considered as an appropriate adaptation to the standard information requirements of the REACH Regulation for the target substance, in accordance with the provisions of Annex XI, 1.5 of the REACH Regulation.

A justification for read-across is attached to IUCLID section 13.

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
See justification for read-across attached to IUCLID section 13.

3. ANALOGUE APPROACH JUSTIFICATION
See justification for read-across attached to IUCLID section 13.

4. DATA MATRIX
See justification for read-across attached to IUCLID section 13.

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across: supporting information

Reason / purpose for cross-reference:

read-across source

Species:

rat

Strain:

other: Crl:WI(Han)

Sex:

male/female

Route of administration:

oral: gavage

Vehicle:

propylene glycol

Organ weight findings including organ / body weight ratios:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Reproductive function: oestrous cycle:

not examined

Reproductive function: sperm measures:

no effects observed

Reproductive performance:

effects observed, treatment-related

CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS)
- No test substance-related mortality occurred during the study period.
Two males at 200 mg/kg bw/d (nos. 31 and 37) and one female at 70 mg/kg bw/d(no. 66) were euthanized in extremis on Days 11 (nos. 37 and 66) or Day 20 (no. 31): macroscopic and microscopic examinations suggested that gavage trauma was the cause of morbidity for these animals; the deaths were not substance-related

- no toxicologically relevant clinical signs were noted
- 200 mg/kg bw/d: hunched posture was noted among all males primarily during the second week of treatment, and at lower incidence, rales, piloerection and lean appearance were noted among some males. These findings had resolved for most animals as treatment progressed.
- clinical signs noted for the animals euthanized in extremis (nos. 31, 37 and 66) included (but were not limited to) hunched posture, rales, gasping, abdominal swelling, piloerection, lethargy and laboured respiration and chromodacryorrhoea, and were considered to be due to gavage trauma.
- salivation noted at 70 and 200 mg/kg bw/d was considered to be a physiological response rather than a sign of systemic toxicity considering the nature and minor severity of the effect and its time of occurrence (i.e. after dosing). This sign may be related to the taste of the test substance. No toxicological relevance was ascribed to these changes.

Incidental findings: included rales, alopecia and scabs; the incidence remained within the range of background findings to be expected for rats of this age and strain housed and treated under the conditions in this study; these were not considered to be toxicologically relevant


BODY WEIGHT AND FOOD CONSUMPTION (PARENTAL ANIMALS)
200 mg/kg bw/d
- males showed weight loss up to 15% of day 1 weight during the first 2 weeks of treatment, which largely recovered during the treatment period
- mean body weight and body weight gain remained statistically significantly lower throughout treatment, but body weight gain exceeded that of controls during the mating period
- females showed minor (statistically significant) reduced body weight gain during the first two weeks of treatment
- at start of post-coitum, mean body weight was similar to control levels, but body weight (gain) was lower during the post coitum phase
- absolute and relative food consumption was reduced for males during the premating period, and for females during the first week of the premating period
- for males, food consumption had recovered to control levels during the mating period, while for females food consumption remained slightly lower throughout the post-coitum and lactation period

70 mg/kg bw/d
- slightly lower (but statistically significant) body weight gain was noted for females during the last week of the post coitum phase
- lower absolute and relative food consumption for females at 70 mg/kg bw/d throughout the post-coitum and lactation period (statistically significant on most occasions)


REPRODUCTIVE FUNCTION: SPERM MEASURES (PARENTAL ANIMALS)
- spermatogenic staging profiles were normal for males examined
- testes and epididymides weights were unaffected by treatment


REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS)
- significantly lower number of implantation sites at 200 mg/kg bw/d
- attributable to low numbers for female nos. 79 and 80; upon exclusion of the values for these two females, the mean was similar to that of the 70 mg/kg bw/d group

- mating, fertility and conception indices, precoital time, and number of corpora lutea were unaffected by treatment
- in one female the number of pups born was slightly higher than the number of implantations and corpora lutea recorded; this was considered to be caused by normal resorption of these areas as these enumerations were performed on Day 7 of lactation

- No toxicologically relevant effects on the gestation index and duration, parturition, maternal care and early postnatal pup development (mortality, clinical signs, body weight and macroscopy)

- No signs of difficult or prolonged parturition were noted among the pregnant females.
Examination of cage debris of pregnant females revealed no signs of abortion or premature birth and no deficiencies in maternal care were observed.

- significantly lower mean number of living pups at first litter check at 70 and 200 mg/kg bw/d
- at 200 mg/kg bw/d, 4/9 females had litter sizes of 3-9 pups (the lowest litter size was found for the two females with a low number of implantation sites)
- at 70 mg/kg bw/d, 5/9 females had litter sizes of 7-9 pups
- in the control group, 1/10 female had a litter size of 8 pups, while all other females had litter sizes of 12-15
- the historical control mean values for litter size in this lab is 11.8 (st.dev.= 2.47), n=588 litters (min=2, max=18), 95% confidence interval: 7-15.


ORGAN WEIGHTS (PARENTAL ANIMALS)
- Testes and epididymides weights were unaffected by treatment.
- lower terminal body weights for males at 200 mg/kg bw/d were in line with the lower in-life body weight noted for these animals


GROSS PATHOLOGY (PARENTAL ANIMALS)
Macroscopic observations at necropsy did not reveal any alterations that were attributable to treatment with the test substance.

Perforation of the esophagus was noted for two animals (male no. 37 (200 mg/kg bw/d) and female no. 66 (20 mg/kg bw/d)) that were euthanised in extremis. This was considered direct evidence that these deaths were considered due to gavage trauma. For the other male at 200 mg/kg bw/d euthanised in extremis (no. 31) macroscopic findings were not directly indicative of gavage trauma, but based on histopathological assessment this death was also ascribed to a gavage-related incident. Other macroscopic findings noted for these deaths included emaciated appearance, gastro-intestinal tract distended with gas, red foci on the lungs, irregular surface of the forestomach, reddish discoloration of the mesenteric lymph node or the stomach glandular mucosa, reduced size of the spleen and/or thymus.

The incidence of other necropsy findings noted for control and/or treated animals remained within the background range of findings encountered among rats of this age and strain, and did not show a dose-related trend. They were not considered to be toxicologically relevant.


HISTOPATHOLOGY (PARENTAL ANIMALS)
- no test item related microscopic findings
- no findings in the reproductive organs for animals that failed to sire or deliver healthy offspring that were outside the range of normal background pathology
- spermatogenic staging profiles were normal for males examined

Dose descriptor:

NOAEL

Remarks:

general toxicty

Effect level:

70 mg/kg bw (total dose)

Based on:

act. ingr.

Sex:

male/female

Basis for effect level:

other: body weight; food consumption

Dose descriptor:

NOAEL

Remarks:

fertility

Effect level:

70 mg/kg bw/day

Based on:

act. ingr.

Sex:

female

Basis for effect level:

other: lower number of implantation sites

Dose descriptor:

NOAEL

Remarks:

fertility

Effect level:

200 mg/kg bw (total dose)

Based on:

act. ingr.

Sex:

male

Basis for effect level:

other: no adverse effects observed

Critical effects observed:

no

Sexual maturation:

not examined

Anogenital distance (AGD):

not examined

Nipple retention in male pups:

not examined

Organ weight findings including organ / body weight ratios:

not examined

Histopathological findings:

not examined

Behaviour (functional findings):

not examined

Developmental immunotoxicity:

not examined

The number of dead pups at first litter check, postnatal loss, viability index and sex ratio were unaffected by treatment, and clinical signs, body weight and external macroscopy did not reveal treatment-related findings.

VIABILITY (OFFSPRING)
One pup in the control, 70 and 200 mg/kg bw/d groups went missing during lactation. These pups were most likely cannibalised. No toxicological relevance was attributed to these dead/missing pups since the mortality incidence did not show a dose-related trend and remained within the range considered normal for pups of this age. No pups died or went missing at 20 mg/kg bw/d.

CLINICAL SIGNS (OFFSPRING)
Scabs on the left foreleg were noted for two pups at 200 mg/kg bw/d. The nature and incidence of this finding remained within the range considered normal for pups of this age, and was therefore not considered to be toxicologically relevant.

BODY WEIGHT (OFFSPRING)
Body weights of pups were unaffected by treatment up to 200 mg/kg bw/d.

GROSS PATHOLOGY (OFFSPRING)
Scabs on the left foreleg were noted for two pups at 200 mg/kg bw/d were incidental in nature. The nature and incidence of this finding remained within the range considered normal for pups of this age, and was therefore not considered to be toxicologically relevant.

Dose descriptor:

NOAEL

Remarks:

development

Generation:

F1

Effect level:

200 mg/kg bw/day

Based on:

act. ingr.

Sex:

male/female

Basis for effect level:

other: no adverse effects observed

Remarks on result:

not measured/tested

Reproductive effects observed:

no

Conclusions:

Based on the results obtained with the source substance Stearic acid 3-(dimethylaminopropyl)amide, the following NOAELs were derived for DIMAPDO:
parental NOAEL: 70 mg/kg
fertility NOAEL, females: 70 mg/kg
fertility NOAEL, males: 200 mg/kg
developmental NOAEL: 200 mg/kg

Effect on fertility: via oral route

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEL

70 mg/kg bw/day

Study duration:

subacute

Species:

rat

Effect on fertility: via inhalation route

Endpoint conclusion:

no study available

Effect on fertility: via dermal route

Endpoint conclusion:

no study available

Additional information

Experimental data on reproductive toxicity is not available for the target substance DIMAPDO. For the assessment of effects of DIMAPDO to reproduction results from the following studies are taken into consideration:

- an oral reproduction / developmental toxicity screening test in rats according to OECD Guideline 421 with the source substance Stearic acid 3-(dimethylaminopropyl)amide

- a Combined Repeated Dose Toxicity Study with the Reproduction/Developmental Toxicity Screening Test in rats according to OECD Guideline 422 with the source substanceN-[3-(dimethylamino)propyl] C12-C18 alkylamide

A justification for read-across is attached to IUCLID section 13.

 

In a Reproduction/Developmental Toxicity Screening Test according to OECD guideline 421 (July 1995) Stearic acid 3-(dimethylaminopropyl)amide (100% a.i.) was administered to groups of 10 Wistar rats/sex/dose by gavage at dose levels of 0, 20, 70 and 200 mg/kg bw/d. 

Males were exposed for 28 days, i.e. 2 weeks prior to mating, during mating, and up to termination. Females were exposed for 41 – 54 days, i.e. during 2 weeks prior to mating, during mating, duringpost-coitum, and during at least 4 days of lactation.

At 200 mg/kg bw/d, males showed weight loss up to 15% of day 1 weight during the first 2 weeks of treatment, which largely recovered during the treatment period. The mean body weight and body weight gain remained statistically significantly lower throughout treatment. Females of the same dose group showed statistically significant reduced body weight gain during the first two weeks of treatment, as well as during pregnancy. Food intake was reduced for males during the premating period, and for females during the first week of the premating period; for females food intake remained slightly lower throughout pregnancy and lactation.

No treatment-related changes were noted in any of the remaining parental parameters investigated in this study (i.e. macroscopic examination, organ weights, and microscopic examination).

The mean number of corpora lutea was slightly lower in the 70 and 200 mg/kg bw/d dose groups compared with the control animals, however, this was not statistically significant.

A statistically significant lower number of implantation sites were noted for females at 200 mg/kg bw/d. This was attributable to extremely low numbers of implantation sites in two females (3 and 6, respectively); upon exclusion of the values for these two females, the mean number of implantation sites was similar to that of the 70 mg/kg bw/d group, which showed also a slight, but not statistically significant reduction of implantations when compared to control animals.

A statistically significant lower number of living pups was noted in the 70 and 200 mg/kg bw/d dose groups. However, as the lower litter size correlated with lower number of implantation sites also when regarding single animals, this was considered to be a consequence of the reduced number of implantation sites.

No treatment-related changes were noted in any of the remaining reproductive parameters investigated in this study (i.e. mating, fertility and conception indices and precoital time, testes and epididymides weights, spermatogenic staging profiles).

Due to the remarkable effects on body weight /body weight gain and food consumption, the observed fertility effects – reduced number of implantation sites and subsequently lower litter size – are considered to be a consequence of general parental toxicity.

Based on these results, the following NOAELs were derived from this study:

parental NOAEL: 70 mg/kg

fertility NOAEL, females: 70 mg/kg

fertility NOAEL, males: 200 mg/kg

developmental NOAEL: 200 mg/kg

 

 

Discussion of effects concerning fertility (reproduction/developmental toxicity screening test)

Reduced number of implantation sites

In the reproduction / developmental toxicity screening test a reduction in the mean number of implantation sites was noted in females of the 200 mg/kg bw/d dose group (mean number of implantation sites 9.8). However, this reduction was attributed to only 2/9 animals which had very low numbers of implantation sites. When considering these two animals as outliers and not taking them into account for the calculations, the new calculated mean would be 11.3 (compared to 13.5, 13.2 and 11.4 in the control, 20 and 70 mg/kg bw/d groups).

 

The mean number of corpora lutea was also slightly lower in the 70 and 200 mg/kg bw/d dose groups compared with the control animals, however, this was not statistically significant. The two females of the high dose group with extremely low implantation sites had also the lowest numbers of corpora lutea (9 compared to 11-20 in the other 7 animals of the high dose group).

 

In this study the following signs of general toxicity were noted:

In females of the 200 mg/kg bw/d dose group, a statistically significant reduction in body weight gain was noted during the first two weeks of treatment, as well as during pregnancy. Also in males effects on body weight were noted in the high dose group: most males showed weight loss (up to 15% of day 1 values) during the first two weeks of treatment. Although body weights largely recovered as treatment progressed, this effect was considered to be of toxicological relevance.

The absolute and relative food consumption was reduced for males at 200 mg/kg bw/d during the premating period (31 g/kg bw/d vs. 61 g/kg bw/d for the control animals in week 1, 46 g/kg bw/d vs. 63 g/kg bw/d for the control animals in week 2), and for females at during the first week of the premating period (48 g/kg bw/d vs. 68 g/kg bw/d for the control animals). The relative food consumption of females of the 70 and 200 mg/kg bw/d dose groups were significantly lower throughout pregnancy.

 

This is also supported by the findings in the14 day dose range finding study, where all animals treated with 500 mg/kg bw/d were sacrificed for humane reasons between days 6 and 8. Animals showed lethargy, hunched posture, laboured respiration, abdominal swelling, piloerection, chromodacryorrhoea, a lean appearance and/or ptosis from day 4 of treatment onwards. This shows, that there is a steep dose-response-curve for the tested substance and that the dose of 200 mg/kg bw/day is most probably the highest tolerable dose with already some signs of general toxicity.

There are no data gaps for effects on fertility. No human data are available. However, there is no reason to believe that these results from rat would not be applicable to humans.

 

Discussion of effects concerning development (reproduction/developmental toxicity screening test)

Reduced litter size

A statistically significant lower number of living pups was noted in the 70 and 200 mg/kg bw/d dose groups (mean numbers of living pups 12.9, 12.5, 10.0* and 8.8** at 0, 20, 70 and 200 mg/kg bw/d, respectively; * p<0.05, ** p<0.01).

As the lower litter size of the 200 mg/kg bw/d dose group was predominantly the consequence of the lower number of implantation sites, the two dams with the extremely low implantation sites were not taken into account for calculation, resulting in mean number of living pups of 10.0 for the 200 mg/kg bw/d dose group. This was still lower than the number of living pups of the control animal, but within the range of historical control data of the laboratory for animals of that strain. It has also to be taken into account that the mean litter size of the control group in this experiment was rather high in comparison to historical control values of the performing laboratory (mean = 11.8, SD = 2.47, n = 588 litters, min=2, max=18, 95% confidence interval: 7-15).

 

Nature of the effect

A reduced litter size may be a developmental effect. However, in this study the lower mean litter size is accompanied by a lower number of implantation sites. This can also be observed at the level of individual animals as demonstrated in the table below.

In the mid dose group the animals with relatively low litter size (7 to 9) had in general also lower implantation sites (9 to 11). In the high dose group the animals which delivered a very low number of living pups (6 or 3) had the same number of implantation sites.

 

Conclusion

Based on the data of the reproduction/developmental toxicity screening test with the source substance Stearic acid 3-(dimethylaminopropyl)amide, the NO(A)EL fertility was 70 mg/kg bw/d (based on reduced implantation sites at 200 mg/kg bw/d) and the NO(A)EL development was 200 mg/kg bw/d. The lower litter size is considered to be secondary to the reduction of implantation sites. Therefore, the reduced litter size is judged to be not a developmental effect.

 

A Combined Repeated Dose Toxicity Study with the Reproduction/Developmental Toxicity Screening Test in accordance with OECD TG 422 was performed with the source substanceN-[3-(dimethylamino)propyl] C12-C18 alkylamide.The test item was administered by gavage to three groups, each of ten male and ten female Wistar rats, for up to eight weeks (including a two week maturation phase, pairing, gestation and early lactation for females), at dose levels of 5, 15 and 45 mg/kg/day. A control group of ten males and ten females was dosed with vehicle alone (Corn oil). Clinical signs, behavioural assessments, body weight change and food and water consumption were monitored during the study. Extensive functional observations were performed on five selected males from each dose group after the completion of the mating phase, and for five selected parental females from each dose group on Day 4 postpartum. Haematology and blood chemistry were evaluated prior to termination on five selected males and females from each dose group. Males were terminated on Day 43, followed by the termination of all surviving females and offspring on Day 5 post partum. All animals were subjected to a gross necropsy examination and histopathological evaluation of selected tissues was performed.

There were no unscheduled deaths that were related to treatment. Episodes of increased salivation were evident in animals of either sex treated with 45 mg/kg/day throughout the treatment period. An isolated incident of noisy respiration was also evident in one male treated with 45 mg/kg/day on Day 4. No toxicologically significant effects were detected in animals of either sex treated with 15 or 5 mg/kg/day. There were no treatment-related changes in the behavioural parameters measured, no toxicologically significant changes in functional performance and no treatment-related changes in sensory reactivity. Females treated with 45 mg/kg/day showed a reduction in body weight gain during the premating period, with four females showing actual body weight losses during the first week of treatment whereas this was the case for two animals of the control group. However, this difference was not statistically significant, nor was there any difference in body weight visible during the remainder of the study. No body weight effects were detected in males treated with 45 mg/kg/day or animals of either sex treated with 15 or 5 mg/kg/day. No adverse effect on food or water consumption or food efficiency was detected.

There were no treatment-related effects on mating or conception rates for treated animals. There were no differences in gestation lengths. The distribution for treated females was comparable to controls. Of the litters born, litter size at birth and subsequently on Days 1 and 4 postpartum were comparable to controls. Offspring bodyweight gain and litter weights at birth and subsequently on Days 1 and 4 postpartum were comparable to controls. The 'No Observed Adverse Effect Level' (NOAEL) for systemic toxicity was therefore considered to be 15 mg/kg/day. No treatment related effects were detected in the reproductive parameters examined therefore the 'No Observed Effect Level' (NOEL) for reproductive/developmental toxicity was considered to be 45 mg/kg/day.

Effects on developmental toxicity

Description of key information

Prenatal Developmental Toxicity Test; oral (gavage), rat (Sprague-Dawley); 22 pregnant females/dose, 0, 15, 40 and 100 mg/kg bw/d, OECD TG 414, GLP; NOAEL(development)=100 mg/kg bw/d (no developmental effects at highest dose level); NOAEL(maternal) = 40 mg/kg bw/day (based on rales, piloerection, and reduction in body weight gain at 100 mg/kg bw/day); read-across:N-[3-(dimethylamino)propyl] C12-C18 alkylamide

Link to relevant study records

Reference

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:

REPORTING FORMAT FOR THE ANALOGUE APPROACH

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
This read-across is based on the hypothesis that source and target substances have similar toxicological properties because
• they are manufactured from similar or identical precursors under similar conditions
• they share structural similarities with common functional groups: tertiary amines, amides, and fatty acid chains with comparable length and degree of saturation (corresponding to scenario 2 of the read-across assessment framework)

The read-across hypothesis is based on structural similarity of target and source substances. The target and source chemicals have a very similar structure in that they are comprised of a hydrophobic (alkyl) and hydrophilic (amine headgroup) end. Due to this motif they form micelles (colloidal dispersions) and have surfactant properties.
Based on available experimental data, including key physicochemical properties and data from genotoxicity studies, the read-across strategy is supported by a similar toxicological profile of all substances.

Therefore, read-across from the existing toxicity studies conducted with the source substances is considered as an appropriate adaptation to the standard information requirements of the REACH Regulation for the target substance, in accordance with the provisions of Annex XI, 1.5 of the REACH Regulation.

A justification for read-across is attached to IUCLID section 13.

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
See justification for read-across attached to IUCLID section 13.

3. ANALOGUE APPROACH JUSTIFICATION
See justification for read-across attached to IUCLID section 13.

4. DATA MATRIX
See justification for read-across attached to IUCLID section 13.

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across: supporting information

Species:

rat

Strain:

other: Crl:WI (Han)

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Deutschland, Sulzfeld, Germany
- Age at study initiation: 10-14 weeks.
- Weight at study initiation: 174-231 g
- Fasting period before study: none
- Housing: individually in Macrolon plastic cages (MIII type, height 18 cm). Sterilized sawdust as bedding material and paper as cage enrichment/nesting material were supplied.
- Diet: Free access to pelleted rodent diet (SM R/M-Z from SSNIFF® Spezialdiäten GmbH, Soest, Germany).
- Water: Free access to tap-water
- Acclimation period: At least 5 days prior to treatment.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20.7 - 21.2
- Humidity (%): 50.1 – 74.8
- Air changes (per hr): at least 10
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 06 September 2016 (first delivery of mated females) To: 29 September 2016

Route of administration:

oral: gavage

Vehicle:

corn oil

Details on mating procedure:

Untreated females were mated at the Supplier, and were at Day 0 or 1 post-coitum on arrival at the Test Facility.
- Proof of pregnancy: vaginal plug referred to as day 0 of pregnancy

Duration of treatment / exposure:

From Days 6 to 20 post-coitum, inclusive

Frequency of treatment:

Once daily, 7 days/week

Duration of test:

14 days

Dose / conc.:

15 mg/kg bw/day

Dose / conc.:

40 mg/kg bw/day

Dose / conc.:

100 mg/kg bw/day

No. of animals per sex per dose:

22 females/dose

Control animals:

yes, concurrent vehicle

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

Rales and piloerection were noted at 100 mg/kg bw/day for five and one female(s), respectively. Although these signs recovered after 1-5 days, they might be related to treatment with test item.
Salivation was noted for all females at 100 mg/kg bw/day, compared to none in the other groups. This finding was considered to be a physiological response rather than a sign of systemic toxicity considering the nature and minor severity of the effect and its time of occurrence (i.e. after dosing).
Moreover, alopecia was noted for one single female at 15 mg/kg bwd/day, which was not considered to be treatment related.

Mortality:

mortality observed, non-treatment-related

Description (incidence):

Treatment at 100 mg/kg bw/day resulted in two unscheduled deaths.
One female at 100 mg/kg bw/day was sacrificed in extremis on Day 7 post-coitum (before dosing), the day after having received the first dose. Observations preceding early sacrifice included squeaking, rales, labored (severe) and shallow (moderate) respiration, gasping and hypothermia. At necropsy, dark red foci were noted on the thymus. This female was pregnant and had 8 normal implantations in development.
Another female of the 100 mg/kg bw/day group was found dead on Day 21 post-coitum, the day of planned necropsy. No toxicologically relevant clinical signs were noted for this female. Body weight and food consumption was slightly lower for this female, when compared to the other females of the 100 mg/kg bw/day group. This was considered to be (partly) caused by the number of fetuses, as she had only 3 (dead) fetuses and 4 early resorptions. No macroscopic alterations were noted at necropsy.
Although these mortality findings were incidental, it could not be excluded that these unscheduled deaths were related to treatment.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

There was a trend towards lower body weights at 100 mg/kg bw/day from Day 12 post-coitum onwards. Body weight gain was statistically significantly lower at 100 mg/kg bw/day than controls on Days 15 and 18 post-coitum. For uterus corrected body weight gain was considered to be unaffected by treatment up to 100 mg/kg bw/day.
Mean body weight and body weight gain at 15 and 40 mg/kg bw/day remained in the same range as controls.
One female from the control group and three females from the 100 mg/kg bw/day group had significantly lower body weights and weight gains when compared to the other females of the same group. This was considered to be (partly) due to the relatively low number of fetuses, as two females had 1 fetus, and two other females had 3 fetuses. In addition, for two females (one with one fetus and one with two fetuses) no body weight gain was observed over Days 12 to 21 post-coitum, which was considered to be a direct treatment-related effect rather than only caused by the low number of fetuses.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

Food consumption before and after correction for body weight was statistically significantly reduced at 100 mg/kg bw/day on Days 9 to 12 post-coitum. This recovered during the remainder of the treatment period.
At 15 and 40 mg/kg bw/day, food consumption before or after correction for body weight were similar as controls.

Food efficiency:

not examined

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

no effects observed

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

effects observed, non-treatment-related

Description (incidence and severity):

Alopecia was noted for one single female at 15 mg/kg bw/day, confirming the clinical sign observed during the in-life phase.
Dark red foci on the thymus were noted for one female at 100 mg/kg bw/day, which was early sacrificed.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

not examined

Histopathological findings: neoplastic:

not examined

Other effects:

no effects observed

Number of abortions:

no effects observed

Pre- and post-implantation loss:

no effects observed

Description (incidence and severity):

Pre- or post-implantation loss were unaffected by treatment up to and including 100 mg/kg bw/day.

Total litter losses by resorption:

no effects observed

Early or late resorptions:

no effects observed

Dead fetuses:

no effects observed

Changes in pregnancy duration:

not examined

Changes in number of pregnant:

not examined

Other effects:

effects observed, non-treatment-related

Description (incidence and severity):

Statistically significantly lower numbers of corpora lutea (10.7 per female) were noted in the 100 mg/kg bw/day group when compared to the concurrent control group (12.2 per female). As treatment started from implantation onwards, i.e. Day 6 post-coitum, this was not considered to be treatment related. Consequently, the number of implantation sites at 100 mg/kg bw/day were slightly lower than controls as well (9.6 versus 11.4 per female). This was not statistically significant and within the range of available historical control data and not considered to be treatment related.

Key result

Dose descriptor:

NOAEL

Effect level:

40 mg/kg bw/day

Based on:

test mat.

Basis for effect level:

body weight and weight gain

clinical signs

Key result

Abnormalities:

no effects observed

Fetal body weight changes:

no effects observed

Description (incidence and severity):

There were no effects on fetal body weights (both sexes) noted by treatment up to 100 mg/kg bw/day.
Mean combined (male and female) fetal body weights were 5.3, 5.3, 5.2 and 5.4 gram for the control, 15, 40 and 100 mg/kg groups, respectively
Migrated Data from removed field(s)
Field "Fetal/pup body weight changes" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsFetuses.FetalPupBodyWeightChanges): not examined

Reduction in number of live offspring:

effects observed, non-treatment-related

Description (incidence and severity):

Mean litter sizes were 10.7, 10.7, 11.0 and 9.1 viable fetuses/litter for the control, 15, 40 and 100 mg/kg bw/day groups, respectively.
The slightly lower number of fetuses/litter at 100 mg/kg bw/day when compared to controls was not statistically significant and within the range of available historical control data. As it was related to the lower numbers of corpora lutea and implantation sites at 100 mg/kg bw/day, it was not considered to be treatment related.
There were two females, one in the control group and one in the 100 mg/kg bw/day group, which only had one viable fetus. This was considered to be chance findings.

Changes in sex ratio:

no effects observed

Description (incidence and severity):

The male:female ratio was unaffected by treatment up to 100 mg/kg bw/day.
Mean sex ratios (males:females) were 55:45, 49:51, 43:57 and 50:50 for the control, 15, 40 and 100 mg/kg groups, respectively.

Changes in litter size and weights:

effects observed, non-treatment-related

Description (incidence and severity):

There were no treatment-related effects on litter size of any group.
Mean litter sizes were 10.7, 10.7, 11.0 and 9.1 viable fetuses/litter for the control, 15, 40 and 100 mg/kg groups, respectively.
The slightly lower number of fetuses/litter at 100 mg/kg bw/day when compared to controls was not statistically significant and within the range of available historical control data. As it was related to the lower numbers of corpora lutea and implantation sites at 100 mg/kg bw/day, it was not considered to be treatment related.

Changes in postnatal survival:

not examined

External malformations:

effects observed, non-treatment-related

Description (incidence and severity):

There were no treatment related effects on external morphology following treatment up to 100 mg/kg bw/day.
External malformations occurred in all groups. A small lower jaw was observed in one fetus of 100 mg/kg bw/day group and was skeletally confirmed. An absent tail combined with anal atresia were found in one fetus of 40 mg/kg bw/day group, and in 15 mg/kg bw/day group an omphalocele in one fetus and anasarca in another fetus were noticed. Omphalocele and anasarca were observed in single control fetuses as well. The single occurrence and/or group distribution of the above malformations did not indicate a treatment relationship and therefore all were considered to be chance findings.
External variations were not seen in any group.

Skeletal malformations:

effects observed, non-treatment-related

Description (incidence and severity):

There were no treatment related effects on skeletal morphology following treatment up to 100 mg/kg bw/day.
Malformations were observed in two fetuses of 100 mg/kg bw/day group and single fetuses of control, 15 and 40 mg/kg bw/day groups. A fetus from the 100 mg/kg bw/day group (which had a small lower jaw and a variety of visceral malformations) appeared to have severely malaligned sternebrae and a bent scapula as well. Another fetus of the 100 mg/kg bw/day group had malpositioned metatarsals.
The other affected fetuses (control, 15 and 40 mg/kg bw/day groups, respectively) all had a vertebral anomaly with or without associated rib anomaly. In addition, the fetus from the 15 mg/kg bw/day group (that also had an omphalocele and malpositioned kidneys) had a sternal anomaly and bent limb bones as well. The group distribution and/or single occurrence of these malformations did not suggest any treatment relationship and therefore all were considered to be chance findings.
Skeletal variations occurred at an incidence of 86.7%, 89.3%, 78.5% and 76.0% per litter in controls, 15, 40 and 100 mg/kg bw/day, respectively. All the ones noted, were not considered treatment-related, as they occurred in the absence of a dose-dependent relationship, infrequently and/or at frequencies that were within the range of available historical control data.

Visceral malformations:

effects observed, non-treatment-related

Description (incidence and severity):

There were no treatment related effects on visceral morphology following treatment up to 100 mg/kg bw/day.
In total five fetuses were viscerally malformed in this study. The two fetuses of 15 mg/kg bw/day group, one fetus of the 100 mg/kg bw/day group and one control fetus that were affected externally, appeared to have one or more visceral malformations as well. The other affected fetus was a 40 mg/kg bw/day group fetus. Due to the group distribution and variety of malformation observed, all were considered not to be toxicologically relevant.
Visceral variations were observed in 10.3%, 9.1%, 7.3% and 7.5% of fetuses per litter in controls, 15, 40 and 100 mg/kg bw/day groups, respectively. These all occurred in the absence of a dose-related incidence trend, infrequently and/or at frequencies that were within the range of available historical control data.

Other effects:

no effects observed

Key result

Dose descriptor:

NOAEL

Effect level:

100 mg/kg bw/day

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: no treatment-related effects observed at the highest tested dose.

Key result

Abnormalities:

no effects observed

Key result

Developmental effects observed:

no

Conclusions:

In a GLP-compliant guideline study with rats, the administration of the source substance by oral gavage to pregnant rats during gestation days 6-21 did not result in developmental effects on the offspring at the highest tested dose of 100 mg/kg bw/day. This level was considered to be a NOAEL for developmental effects. Based on the clinical signs and reduced body weight gain at the highest dose level in maternal animals, the NOAEL for maternal toxicity was set at 40 mg/kg bw/day. This result is applicable also to the target substance DIMAPDO.

Effect on developmental toxicity: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

100 mg/kg bw/day

Study duration:

subacute

Species:

rat

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

Experimental data on developmental toxicity is not available for the target substance DIMAPDO. For the assessment of effects of DIMAPDO to reproduction results from the following study is taken into consideration. A justification for read-across is attached to IUCLID section 13.

 

In a GLP-compliant OECD guideline 414 study, the source substance N-[3-(dimethylamino)propyl] C 12-C18 alkylamide was administered to groups of 22 pregnant rats at dose levels of 0 (concurrent vehicle controls), 15, 40 and 100 mg/kg bw/day. Two mortalities occurred at the highest dose level (day 7 and day 21); however, in the absence of corroborative gross pathological and histopathological findings these deaths are likely to have been incidental. Rales and piloerection were observed in 5 and 1 female of the high-dose group, respectively, but disappeared after 1 -5 days. Lower body weight gains were observed on days 15 and 18 of gestation. Based on these effects the maternal NOAEL was set at 40 mg/kg bw/day.

Statistically significantly lower numbers of corpora lutea were noted in the high-dose group in comparison to the controls, resulting in a lower number of implantation sites; however, as the treatment started after the implantation, i.e. from day 6 post-coitum, this finding was considered to be not related to the treatment. Furthermore, the number of implantation sites was within the range of historical control data. There were no effects on the number of early and late resorption, post-implantation losses or the number of viable fetuses. Body weights and sex ratio of fetuses were unaffected by the treatment. There were no treatment-related external, visceral or skeletal variations or malformations at any dose level. Based on this, the NOAEL for developmental toxicity was set at the highest tested dose of 100 mg/kg bw/day.

Justification for classification or non-classification

Results of existing studies with structural very similar substances indicate that DIMAPDO does not need to be classified for toxicity to reproduction, developmental toxicity and teratogenicity according to the criteria given in regulation (EC) 1272/2008. Therefore labelling is not necessary.