Registration Dossier

Diss Factsheets

Administrative data

Description of key information

A 90-d oral study conducted on a read across substance, C12 DEA from which a NOAEL of 50 mg/kg bw/d was established, showed effects at higher dose levels. However it is unclear whether the effects noted were related to the test substance itself or was a result of the nutritional deficiencies due to the un-palatability of the diet as evidenced by scattering of food (Sharrat et al., 1961). Further, a 28-d oral repeated dose toxicity study conducted with read across substance, C12-18 and C18-unsatd. DEA in rats revealed a NOAEL of >750 mg/kg bw/d based on absence of treatment-related effects at any of the tested dose levels. Based on this, it is scientifically justified to use the 28-d oral rat NOAEL of 750 mg/kg bw/d for this CSA as the POD. Regarding dermal route, 90-d and 2-yr studies on structurally similar substance, C8-18 and C18-unsatd. DEA have been conducted in both rats and mice. The following dose descriptors have been derived from these studies:

- Sub-chronic dermal rat: NOAEL for systemic effects: 50 mg/kg bw/d based on renal tubule regeneration; LOAEL for local effects: 25 mg/kg bw/d based on non-neoplastic lesions on the skin; Sub-chronic dermal mouse: NOAEL for systemic effects: 200 mg/kg bw/d based on organ-weight changes; LOAEL for local effects: 50 mg/kg bw/d based non-neoplastic lesions on the skin.

- Chronic dermal rat: NOAEL for systemic effects: 50 mg/kg bw/d based on pancreatic acinar atrophy and nephropathy; NOAEL for local effects: 50 mg/kg bw/d based on non-neoplastic lesions on the skin; Chronic dermal mouse LOAEL: 100 mg/kg bw/d based on thyroid gland follicular cell hyperplasia, hepatic and renal neoplasms and non-neoplastic lesions on the skin.

As a conservative approach and giving preference to a longer duration study, the lower NOAEL of 50 mg/kg bw/d for the systemic effects and the NOAEL of 50 mg/kg bw/d for local effects will be used as the POD for deriving DNEL dermal value of long term systemic and local effects respectively.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records
Reference
Endpoint:
short-term repeated dose toxicity: oral
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
1983
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Read across study hence maximum reliability rating of 2 assigned according to ECHA guidance, although study is conducted equivalent or similar to OECD guideline 407.
Justification for data waiving:
other:
Reason / purpose for cross-reference:
reference to same study
Reason / purpose for cross-reference:
reference to other study
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity Study in Rodents)
Deviations:
no
Principles of method if other than guideline:
Not applicable
GLP compliance:
no
Limit test:
no
Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Mus-Rattus, Brunntal, Germany
- Strain: Wistar rat, MuRa Han 67 SPF
- Age at study initiation: between 6-7 weeks
- Weight at study initiation: 109 (f) - 114 (m) g
- Housing: plastic cages, 3 males and 5 females/cage
- Diet (e.g. ad libitum): ad libitum (Altromin Ratdiet No. 1424 DK, Altromin GmbH, Lage, Germany)
- Water (e.g. ad libitum): ad libitum (tap water)
- Acclimation period: 6 d

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19-22
- Humidity (%): 60-80
- Air changes (per hr): 11
- Photoperiod (hrs dark / hrs light): 12/12
Route of administration:
oral: gavage
Vehicle:
olive oil
Details on oral exposure:
Doses were adpated weekly to the body weight; application volume - 5 mL/kg bw
Analytical verification of doses or concentrations:
no
Details on analytical verification of doses or concentrations:
No information
Duration of treatment / exposure:
28 d
Frequency of treatment:
Daily once, 5 times/wk
Remarks:
Doses / Concentrations:
70 mg/kg bw/d (Days 1-28)
Basis:
actual ingested
Remarks:
Doses / Concentrations:
250 mg/kg bw/d (Days 1-28)
Basis:
actual ingested
Remarks:
Doses / Concentrations:
750 mg/kg bw/d (Days 1-14)
Basis:
actual ingested
Remarks:
Doses / Concentrations:
1500 mg/kg bw/d (Days 15-28)
Basis:
actual ingested
No. of animals per sex per dose:
10/sex/dose for main study; 5/sex/dose for 4 month recovery period


Control animals:
yes, concurrent no treatment
Details on study design:
According to standard procedure
Positive control:
Not necessary
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily


DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: end of study


BODY WEIGHT: Yes
- Time schedule for examinations: weekly


HAEMATOLOGY: Yes
- Time schedule for collection of blood: end of study
- Anaesthetic used for blood collection: Yes (ether)
- How many animals: 10 per dose and sex
- Parameters checked: Hematocrit, erythrocytes, leukocytes, hemoglobin, thrombocytes, mean corpuscular volume, white blood cell differential


CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: end of study
- How many animals: 10 per sex and dose
- Parameters checked: GPT, GOT, AP, glucose, urea, total protein, calcium, phosphate, cholesterol


URINALYSIS: Yes
- Time schedule for collection of urine: end of study
- Metabolism cages used for collection of urine: No
- Parameters checked: urea, creatinine, sodium, potassium, glucose, calcium, AP


NEUROBEHAVIOURAL EXAMINATION: No


Other: Groups of 5 male and 5 female rats kept for an additional 4 month recovery period.
Sacrifice and pathology:
GROSS PATHOLOGY: Yes
the following tissues/organs were examined:
adrenal gland (2)
aorta thoracica
brain (cornu ammonis)
coagulating gland with seminal vesicle
epididymis (2)
eye with optic nerve (2)
heart
intestine, large
intestine, small
kidney (2)
liver
lungs
lymph node (cervical) (1)
lymph node (mesenteric) (1)
mucles
oesophagus
ovary (2)
pancreas
prostate
salivary glands (mandibular,
parotid and sublingual gland)
skin
spleen
stomach
testicle (2)
thymus
thyroid (2) (incl. parathyroids)
tongue
trachea (incl. larynx)
urinary bladder
uterus (incl. cervix and oviducts)


HISTOPATHOLOGY: Yes
see gross pathology
Other examinations:
None
Statistics:
't' test used for statistical analysis of all parameters except organ weight (U-test)


Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
not specified
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Clinical biochemistry findings:
effects observed, treatment-related
Urinalysis findings:
no effects observed
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
no effects observed
Details on results:
The doses up to 1500 mg/kg body weight/day were tolerated by all animals without lethality. The body weight gain and total increase of body weights did not differ from the control group. Absolute and relative organ weights showed no significant changes in the substance groups compared to the control group, except the organ weight of the liver which is slightly increased for the males of group 4 (750/1500 mg/kg bw) and increased adrenal glands weight in high dose females. This result is considered of no relevance.

The pathological and histological evaluation did not show significant compound related gross or microscocopic organ injury of liver, kidneys, adrenals, heart, lung, spleen and gonads; dose dependent reversible local findings were restricted to the fore stomach mucosa (important hyperplasia and ulcerations, in some cases also hyper- and parakeratosis of the forestomach of males and females at the high dose. Similar effects but less intense at the medium and low doses). The biochemical parameters calcium, blood sugar, urea, creatinine, cholesterine, GGT, GOT, GPT and LDH did not show any critical signs. Only slight shifts which were not dose-dependent could be observed. These signs were considered as not substance depending.
Dose descriptor:
NOAEL
Effect level:
> 750 mg/kg bw/day (nominal)
Sex:
male/female
Basis for effect level:
other: No biologically relevant treatment-related effects observed on any of the parameters recorded at any dose, also test animals treated with 1500 mg/kg bw (Days 15-28) showed no adverse effect
Critical effects observed:
not specified

None

Conclusions:
Under the test conditions, the 28d NOAEL of structurally similar 'amides, C12-18(even-numbered) and C18(unsatd.), N,N-bis(hydroxyethyl)' to rats can therefore be considered to be greater than 750 mg/kg bw/d.
Executive summary:

A study was conducted to determine the oral toxicity of structurally similar 'amides, C12-18(even-numbered) and C18(unsatd.), N,N-bis(hydroxyethyl)' to rats after 28 d of exposure.

Groups of 10 male and 10 female rats were orally gavaged with the test substance diluted in olive oil 5 d/wk for 28 d. Clinical signs, bodyweight, hematology, clinical chemistry, urinalysis, gross and microscopic pathology were recorded. Additional groups of 5 male and 5 female rats were kept for a 4 month recovery period.

No treatment-related adverse effects were observed at any of the doses. Changes in the forestomach at some doses including controls were attributed to the use of olive oil and found to be reversible after end of exposure.

Under the test conditions, the 28d NOAEL of 'amides, C12-18(even-numbered) and C18(unsatd.), N,N-bis(hydroxyethyl)' to rats can therefore be considered to be greater than 750 mg/kg bw/d.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
750 mg/kg bw/day
Study duration:
subacute
Species:
rat
Quality of whole database:
The information requirements for this tonnage band is sufficiently met with the available data.

Repeated dose toxicity: inhalation - systemic effects

Link to relevant study records
Reference
Endpoint:
sub-chronic toxicity: inhalation
Data waiving:
other justification
Justification for data waiving:
other:
Critical effects observed:
not specified
Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: inhalation - local effects

Link to relevant study records
Reference
Endpoint:
sub-chronic toxicity: inhalation
Data waiving:
other justification
Justification for data waiving:
other:
Critical effects observed:
not specified
Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - systemic effects

Link to relevant study records
Reference
Endpoint:
chronic toxicity: dermal
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Read across study hence maximum reliability rating of 2 assigned according to ECHA guidance, although study is well documented, meets generally accepted scientific principles, acceptable for assessment
Reason / purpose for cross-reference:
reference to same study
Reason / purpose for cross-reference:
reference to other study
Principles of method if other than guideline:
A two-year dermal study was conducted in rat to evaluate the carcinogenic potential of the test substance.

GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
Fischer 344
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Taconic Laboratory Animals and Services, (Germantown, NY)
- Age at study initiation: 6 wk
- Housing: Housed individually in Polycarbonate cages (Lab Products, Inc., Maywood, NJ)
- Bedding: Sani-Chip heat-treated hardwood chips (P.J. Murphy Forest Products Corp., Montville, NJ)
- Diet : NIH-07 open formula pelleted diet (Zeigler Brothers, Inc., Gardners, PA), ad libitum
- Water : Tap water (Columbus municipal supply), ad libitum
- Acclimation period: 11 to 12 d

ENVIRONMENTAL CONDITIONS
- Temperature : 20.0 -23.9 °C
- Humidity : 33-70 %
- Air changes : 10/hr
- Photoperiod : 12 h dark/12 h light

IN-LIFE DATES: From: Feb. 1, 1993 To: Jan. 31, 1995
Type of coverage:
open
Vehicle:
ethanol
Details on exposure:
The test substance formulation was applied to the shaved skin of test animals. No further details provided.


Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Dose formulations were analysed approximately every 2 months using HPLC. In addition to dose formulation analysis prior to dosing, samples collected after dosing (animal room samples) were analysed periodically.
Duration of treatment / exposure:
104 wk
Frequency of treatment:
Five exposures per week
Remarks:
Doses / Concentrations:
0, 50 or 100 mg/kg bw/d (0, 85, or 170 mg/mL in ethanol)
Basis:
nominal per unit body weight
No. of animals per sex per dose:
50/sex/dose
Control animals:
yes, concurrent vehicle
Details on study design:
Dose selection rationale: Doses of 200 or 400 mg/kg bw/d in the 14 wk study were associated with reduced mean body weights, mild anemia, and significantly increased incidences and severities of lesions of the skin at the site of application. Therefore, these doses were considered inappropriate for a 2-year study. At 100 mg/kg bw/d, the incidences of skin lesions, especially ulceration, were less than at 200 mg/kg bw/d, and in general, the severities were minimal to mild. Therefore, 100 mg/kg bw/d was selected as the high dose for this 2-yr study.
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Observed twice daily


DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Clinical findings were recorded initially, at 4 wk intervals during the study, and at the end of the study


DERMAL IRRITATION (if dermal study): Yes


BODY WEIGHT: Yes
- Time schedule for examinations: Animals were weighed initially, weekly during Week 1 through 13, at 4 wk intervals thereafter, and at the end of the studies


Sacrifice and pathology:
SACRIFICE: Carbon dioxide asphyxiation
GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes, Complete histopathology was performed on all rats. In addition to gross lesions and tissue masses, the following tissues
were examined: adrenal gland, bone and marrow, brain, clitoral gland, esophagus, heart, large intestine (cecum, colon, rectum), small intestine (duodenum, jejunum, ileum), kidney, liver, lung, lymph nodes (mandibular and mesenteric), mammary gland, nose, ovary, pancreas, parathyroid gland, pituitary gland, preputial gland, prostate gland, salivary gland, skin, spleen, stomach (forestomach and glandular), testis (and
epididymis and seminal vesicle), thymus, thyroid gland, trachea, urinary bladder, and uterus
Statistics:
Survival Analyses: The probability of survival was estimated by the product-limit procedure of Kaplan and Meier (1958). Possible dose-related effects on survival were analysed by Cox’s (1972) method for testing two groups for equality and Tarone’s (1975) life table test to identify dose-related trends. All reported P values for the survival analyses were two-sided.
Analysis of Neoplasm and Nonneoplastic Lesion Incidences: The Poly-k test (Bailer and Portier, 1988; Portier and Bailer, 1989; Piegorsch and Bailer, 1997) was used to assess neoplasm and nonneoplastic lesion prevalence. Tests of significance included pairwise comparisons of each dosed group with controls and a test for an overall dose-related trend. Continuity-corrected tests were used in the analysis of lesion incidence, and reported P values are one sided.
Analysis of Continuous Variables: Organ and body weight data, which historically have approximately normal distributions, were analyzed with the parametric multiple comparison procedures of Dunnett (1955) and Williams (1971, 1972). Jonckheere's test (Jonckheere, 1954) was used to assess the significance of the dose-related trends. Average severity values were analyzed for significance with the Mann-Whitney U test. Treatment effects were investigated by applying a multivariate analysis of variance (Morrison, 1976) to the transformed data to test for simultaneous equality of measurements across dose levels.
Clinical signs:
effects observed, treatment-related
Dermal irritation:
effects observed, treatment-related
Mortality:
mortality observed, treatment-related
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
not examined
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Histopathological findings: neoplastic:
effects observed, treatment-related
Details on results:
CLINICAL SIGNS AND MORTALITY: Survival rates of dosed male and female rats were similar to those of the vehicle controls. The only clinical finding attributed to dosing was irritation of the skin at the site of application in 100 mg/kg bw/d females.


BODY WEIGHT AND WEIGHT GAIN: The mean body weights of dosed male and female rats were similar to those of the vehicle controls throughout the study.


HISTOPATHOLOGY:
Skin: No neoplasms of the skin were attributed to treatment with test material. Incidences of squamous cell papilloma, keratoacanthoma, trichoepithelioma, basal cell adenoma, or carcinoma (combined) were significantly decreased in 100 mg/kg bw/d male rats. Incidences of epidermal hyperplasia, sebaceous gland hyperplasia, parakeratosis, and hyperkeratosis in all dosed groups were significantly greater than those in the vehicle control groups. The severities of these lesions generally increased with increasing dose and ranged from minimal to mild. Females in the 100 mg/kg bw/d group had a significantly greater incidence of ulceration at the site of application than did the vehicle controls.

Kidney: Incidences of renal tubule hyperplasia in dosed females were significantly greater than those of the vehicle controls, and the incidence of renal tubule adenoma in 50 mg/kg bw/d males was marginally increased. Incidences of chronic nephropathy were similar between vehicle control and dosed groups of male and female rats; however, the severity of nephropathy increased with increasing dose in female rats. The incidences of renal tubule adenoma in all groups of males and of renal tubule carcinoma in 50 mg/kg bw/d females exceeded the historical control ranges. An extended evaluation of the kidney revealed additional renal tubule adenomas in vehicle control and dosed males, and renal tubule adenomas and/or carcinomas in dosed females. When the single and step sections were combined, the incidences of renal tubule hyperplasia in dosed females and of renal tubule adenoma or carcinoma (combined) in 50 mg/kg bw/d females were significantly greater than those of the controls. In female rats, the combined single and step section evaluations of the kidney revealed a significant dose- related increase in the incidence of renal tubule hyperplasia and two adenomas and two carcinomas in the 50 mg/kg bw/d group but only one neoplasm (an adenoma), in the 100 mg/kg group. Renal tubule neoplasms are uncommon in female F344/N rats, and the presence of four neoplasms in the 50 mg/kg bw/d group, combined with the increased incidence of hyperplasia, is suggestive of an association with chemical exposure. However, the absence of an increase in neoplasms in the 100 mg/kg bw/d group in the presence of increased hyperplasia makes the association with chemical exposure uncertain.

Forestomach: The incidences of chronic active inflammation (vehicle control, 1/50; 50 mg/kg bw, 3/50; 100 mg/kg bw, 10/50), epithelial hyperplasia (2/50, 5/50, 13/50), and epithelial ulcer (1/50, 3/50, 11/50) were significantly increased in the forestomach of 100 mg/kg bw females. The severities of these lesions were similar among all groups.

Pancreas: The incidence of pancreatic acinar atrophy in 100 mg/kg male rats was significantly greater than that in the vehicle controls.

Dose descriptor:
NOAEL
Remarks:
(systemic effects)
Effect level:
50 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: pancreatic acinar atrophy and nephropathy at the higher dose
Dose descriptor:
LOAEL
Remarks:
(local effects)
Effect level:
50 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: based on non-neoplastic lesions of the skin at all doses
Critical effects observed:
not specified

All dose formulations analysed during the 2 year studies were within 10% of the target concentration.

Conclusions:
Under the test conditions, there was no evidence of carcinogenic activity of the test substance in male rats in any of the tested dose levels. There was an equivocal evidence of carcinogenic activity in female rats based on a marginal increase in the incidences of renal tubule neoplasms. However, the absence of an increase in neoplasms in the 100 mg/kg bw/d group in the presence of increased hyperplasia makes the association with chemical exposure uncertain.
Executive summary:

A two-year dermal study was conducted in F344/N rats to evaluate the carcinogenic potential of 'amides, C8-18 (even-numbered) and C18-unsatd., N,N-bis(hydroxyethyl)' (in the form ofcoconut oil acid diethanolamine condensate).

 

 Doses studied include 0, 50, or 100 mg/kg bw/d test substance (0, 85, or 170 mg/mL in ethanol). 50 male/female test animals were used in each group. 5 exposures per week were given for 104 wk. The animals were observed twice daily, body weights and clinical findings were recorded periodically. Necropsy was performed on all animals and complete histopathology was performed.

 

The survival rates of treated male and female rats were similar to those of the vehicle controls. The mean body weights of dosed males and females were similar to those of the vehicle controls throughout the study. The only chemical-related clinical finding was irritation of the skin at the site of application in 100 mg/kg bw/d females. There were marginal increases in the incidences of renal tubule adenoma or carcinoma (combined) in 50 mg/kg bw/d females. The severity of nephropathy increased with increasing dose in female rats.

 

 Non neoplastic lesions of the skin at the site of application included epidermal hyperplasia, sebaceous gland hyperplasia, parakeratosis, and hyperkeratosis, and the incidences and severities of these lesions increased with increasing dose. The incidences of chronic active inflammation, epithelial hyperplasia, and epithelial ulcer of the forestomach increased with dose in female rats, and the increases were significant in the 100 mg/kg bw/d group.

 

 Under the test conditions, there was no evidence of carcinogenic activity of the test substance in male rats administered 50 or 100 mg/kg bw/d. There was an equivocal evidence of carcinogenic activity in female rats based on a marginal increase in the incidences of renal tubule neoplasms. However, the absence of an increase in neoplasms in the 100 mg/kg bw/d group in the presence of increased hyperplasia makes the association with chemical exposure uncertain.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEL
50 mg/kg bw/day
Study duration:
chronic
Species:
rat
Quality of whole database:
The information requirements for this tonnage band is sufficiently met with the available data.

Repeated dose toxicity: dermal - local effects

Link to relevant study records
Reference
Endpoint:
chronic toxicity: dermal
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Read across study hence maximum reliability rating of 2 assigned according to ECHA guidance, although study is well documented, meets generally accepted scientific principles, acceptable for assessment
Reason / purpose for cross-reference:
reference to same study
Reason / purpose for cross-reference:
reference to other study
Principles of method if other than guideline:
A two-year dermal study was conducted in rat to evaluate the carcinogenic potential of the test substance.

GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
Fischer 344
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Taconic Laboratory Animals and Services, (Germantown, NY)
- Age at study initiation: 6 wk
- Housing: Housed individually in Polycarbonate cages (Lab Products, Inc., Maywood, NJ)
- Bedding: Sani-Chip heat-treated hardwood chips (P.J. Murphy Forest Products Corp., Montville, NJ)
- Diet : NIH-07 open formula pelleted diet (Zeigler Brothers, Inc., Gardners, PA), ad libitum
- Water : Tap water (Columbus municipal supply), ad libitum
- Acclimation period: 11 to 12 d

ENVIRONMENTAL CONDITIONS
- Temperature : 20.0 -23.9 °C
- Humidity : 33-70 %
- Air changes : 10/hr
- Photoperiod : 12 h dark/12 h light

IN-LIFE DATES: From: Feb. 1, 1993 To: Jan. 31, 1995
Type of coverage:
open
Vehicle:
ethanol
Details on exposure:
The test substance formulation was applied to the shaved skin of test animals. No further details provided.


Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Dose formulations were analysed approximately every 2 months using HPLC. In addition to dose formulation analysis prior to dosing, samples collected after dosing (animal room samples) were analysed periodically.
Duration of treatment / exposure:
104 wk
Frequency of treatment:
Five exposures per week
Remarks:
Doses / Concentrations:
0, 50 or 100 mg/kg bw/d (0, 85, or 170 mg/mL in ethanol)
Basis:
nominal per unit body weight
No. of animals per sex per dose:
50/sex/dose
Control animals:
yes, concurrent vehicle
Details on study design:
Dose selection rationale: Doses of 200 or 400 mg/kg bw/d in the 14 wk study were associated with reduced mean body weights, mild anemia, and significantly increased incidences and severities of lesions of the skin at the site of application. Therefore, these doses were considered inappropriate for a 2-year study. At 100 mg/kg bw/d, the incidences of skin lesions, especially ulceration, were less than at 200 mg/kg bw/d, and in general, the severities were minimal to mild. Therefore, 100 mg/kg bw/d was selected as the high dose for this 2-yr study.
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Observed twice daily


DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Clinical findings were recorded initially, at 4 wk intervals during the study, and at the end of the study


DERMAL IRRITATION (if dermal study): Yes


BODY WEIGHT: Yes
- Time schedule for examinations: Animals were weighed initially, weekly during Week 1 through 13, at 4 wk intervals thereafter, and at the end of the studies


Sacrifice and pathology:
SACRIFICE: Carbon dioxide asphyxiation
GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes, Complete histopathology was performed on all rats. In addition to gross lesions and tissue masses, the following tissues
were examined: adrenal gland, bone and marrow, brain, clitoral gland, esophagus, heart, large intestine (cecum, colon, rectum), small intestine (duodenum, jejunum, ileum), kidney, liver, lung, lymph nodes (mandibular and mesenteric), mammary gland, nose, ovary, pancreas, parathyroid gland, pituitary gland, preputial gland, prostate gland, salivary gland, skin, spleen, stomach (forestomach and glandular), testis (and
epididymis and seminal vesicle), thymus, thyroid gland, trachea, urinary bladder, and uterus
Statistics:
Survival Analyses: The probability of survival was estimated by the product-limit procedure of Kaplan and Meier (1958). Possible dose-related effects on survival were analysed by Cox’s (1972) method for testing two groups for equality and Tarone’s (1975) life table test to identify dose-related trends. All reported P values for the survival analyses were two-sided.
Analysis of Neoplasm and Nonneoplastic Lesion Incidences: The Poly-k test (Bailer and Portier, 1988; Portier and Bailer, 1989; Piegorsch and Bailer, 1997) was used to assess neoplasm and nonneoplastic lesion prevalence. Tests of significance included pairwise comparisons of each dosed group with controls and a test for an overall dose-related trend. Continuity-corrected tests were used in the analysis of lesion incidence, and reported P values are one sided.
Analysis of Continuous Variables: Organ and body weight data, which historically have approximately normal distributions, were analyzed with the parametric multiple comparison procedures of Dunnett (1955) and Williams (1971, 1972). Jonckheere's test (Jonckheere, 1954) was used to assess the significance of the dose-related trends. Average severity values were analyzed for significance with the Mann-Whitney U test. Treatment effects were investigated by applying a multivariate analysis of variance (Morrison, 1976) to the transformed data to test for simultaneous equality of measurements across dose levels.
Clinical signs:
effects observed, treatment-related
Dermal irritation:
effects observed, treatment-related
Mortality:
mortality observed, treatment-related
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
not examined
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Histopathological findings: neoplastic:
effects observed, treatment-related
Details on results:
CLINICAL SIGNS AND MORTALITY: Survival rates of dosed male and female rats were similar to those of the vehicle controls. The only clinical finding attributed to dosing was irritation of the skin at the site of application in 100 mg/kg bw/d females.


BODY WEIGHT AND WEIGHT GAIN: The mean body weights of dosed male and female rats were similar to those of the vehicle controls throughout the study.


HISTOPATHOLOGY:
Skin: No neoplasms of the skin were attributed to treatment with test material. Incidences of squamous cell papilloma, keratoacanthoma, trichoepithelioma, basal cell adenoma, or carcinoma (combined) were significantly decreased in 100 mg/kg bw/d male rats. Incidences of epidermal hyperplasia, sebaceous gland hyperplasia, parakeratosis, and hyperkeratosis in all dosed groups were significantly greater than those in the vehicle control groups. The severities of these lesions generally increased with increasing dose and ranged from minimal to mild. Females in the 100 mg/kg bw/d group had a significantly greater incidence of ulceration at the site of application than did the vehicle controls.

Kidney: Incidences of renal tubule hyperplasia in dosed females were significantly greater than those of the vehicle controls, and the incidence of renal tubule adenoma in 50 mg/kg bw/d males was marginally increased. Incidences of chronic nephropathy were similar between vehicle control and dosed groups of male and female rats; however, the severity of nephropathy increased with increasing dose in female rats. The incidences of renal tubule adenoma in all groups of males and of renal tubule carcinoma in 50 mg/kg bw/d females exceeded the historical control ranges. An extended evaluation of the kidney revealed additional renal tubule adenomas in vehicle control and dosed males, and renal tubule adenomas and/or carcinomas in dosed females. When the single and step sections were combined, the incidences of renal tubule hyperplasia in dosed females and of renal tubule adenoma or carcinoma (combined) in 50 mg/kg bw/d females were significantly greater than those of the controls. In female rats, the combined single and step section evaluations of the kidney revealed a significant dose- related increase in the incidence of renal tubule hyperplasia and two adenomas and two carcinomas in the 50 mg/kg bw/d group but only one neoplasm (an adenoma), in the 100 mg/kg group. Renal tubule neoplasms are uncommon in female F344/N rats, and the presence of four neoplasms in the 50 mg/kg bw/d group, combined with the increased incidence of hyperplasia, is suggestive of an association with chemical exposure. However, the absence of an increase in neoplasms in the 100 mg/kg bw/d group in the presence of increased hyperplasia makes the association with chemical exposure uncertain.

Forestomach: The incidences of chronic active inflammation (vehicle control, 1/50; 50 mg/kg bw, 3/50; 100 mg/kg bw, 10/50), epithelial hyperplasia (2/50, 5/50, 13/50), and epithelial ulcer (1/50, 3/50, 11/50) were significantly increased in the forestomach of 100 mg/kg bw females. The severities of these lesions were similar among all groups.

Pancreas: The incidence of pancreatic acinar atrophy in 100 mg/kg male rats was significantly greater than that in the vehicle controls.

Dose descriptor:
NOAEL
Remarks:
(systemic effects)
Effect level:
50 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: pancreatic acinar atrophy and nephropathy at the higher dose
Dose descriptor:
LOAEL
Remarks:
(local effects)
Effect level:
50 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: based on non-neoplastic lesions of the skin at all doses
Critical effects observed:
not specified

All dose formulations analysed during the 2 year studies were within 10% of the target concentration.

Conclusions:
Under the test conditions, there was no evidence of carcinogenic activity of the test substance in male rats in any of the tested dose levels. There was an equivocal evidence of carcinogenic activity in female rats based on a marginal increase in the incidences of renal tubule neoplasms. However, the absence of an increase in neoplasms in the 100 mg/kg bw/d group in the presence of increased hyperplasia makes the association with chemical exposure uncertain.
Executive summary:

A two-year dermal study was conducted in F344/N rats to evaluate the carcinogenic potential of 'amides, C8-18 (even-numbered) and C18-unsatd., N,N-bis(hydroxyethyl)' (in the form ofcoconut oil acid diethanolamine condensate).

 

 Doses studied include 0, 50, or 100 mg/kg bw/d test substance (0, 85, or 170 mg/mL in ethanol). 50 male/female test animals were used in each group. 5 exposures per week were given for 104 wk. The animals were observed twice daily, body weights and clinical findings were recorded periodically. Necropsy was performed on all animals and complete histopathology was performed.

 

The survival rates of treated male and female rats were similar to those of the vehicle controls. The mean body weights of dosed males and females were similar to those of the vehicle controls throughout the study. The only chemical-related clinical finding was irritation of the skin at the site of application in 100 mg/kg bw/d females. There were marginal increases in the incidences of renal tubule adenoma or carcinoma (combined) in 50 mg/kg bw/d females. The severity of nephropathy increased with increasing dose in female rats.

 

 Non neoplastic lesions of the skin at the site of application included epidermal hyperplasia, sebaceous gland hyperplasia, parakeratosis, and hyperkeratosis, and the incidences and severities of these lesions increased with increasing dose. The incidences of chronic active inflammation, epithelial hyperplasia, and epithelial ulcer of the forestomach increased with dose in female rats, and the increases were significant in the 100 mg/kg bw/d group.

 

 Under the test conditions, there was no evidence of carcinogenic activity of the test substance in male rats administered 50 or 100 mg/kg bw/d. There was an equivocal evidence of carcinogenic activity in female rats based on a marginal increase in the incidences of renal tubule neoplasms. However, the absence of an increase in neoplasms in the 100 mg/kg bw/d group in the presence of increased hyperplasia makes the association with chemical exposure uncertain.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEL
281 µg/cm²
Study duration:
chronic
Species:
rat
Quality of whole database:
The information requirements for this tonnage band is sufficiently met with the available data.

Additional information

Oral

A 28 day oral study conducted in rats (Potokar, 1983) with structurally similaramides, C12-18(even-numbered) and C18-unsatd., N,N-bis(hydroxyethyl)resulted in a no observed adverse effect level (NOAEL) of >750 mg/kg bw/day (male/females) since no treatment-related effects were noted in any of the parameters investigated at any of the dose levels tested (i.e., 70, 250 and 750 mg/kg bw/day). There were changes in the forestomach at some doses including controls, however the authors attributed this to the use of olive oil and found the forestomach changes to be reversible at the end of the exposure period.

In a 90 day oral study conducted in rats (Gaunt IFet al., 1965), the no observed effect level (NOEL) was determined to be 50 mg/kg bw/day for LDEA based on growth retardation, biochemical changes and an increase in kidney weights seen in the 250 mg/kg bw/day dose group and growth retardation, haematological effects (anaemia) and increased kidney and liver weights seen in the 500 and 1000 mg/kg bw/day dose groups. The growth retardation at 250 mg/kg bw/day and above was considered to be caused by the decrease in food intake due to the palatability of the diet which may have also influenced the other effects noted in the higher dose groups.

Dermal

In a subchronic dermal study (NTP report 479, 2001) , 0, 25, 50, 100, 200 or 400 mg/kg bw/day of structurally similar amides, C8 -18 and C18-unsatd., N,N-bis(hydroxyethyl) administered in ethanol for 14 weeks to groups of 10 male and 10 female F344/N rats resulted in a NOAEL of 50 mg/kg bw/day. The NOAEL was established based on the significant decrease in mean body weight and body weight gain in the 200 and 400 mg/kg bw/day dose groups, skin irritation at the site of application in 100, 200 and 400 mg/kg bw/day (males and females) dose groups, as well as haematological changes (minimal microcytic anaemia) and decreased cholesterol and triglyceride concentrations. Histopathologic lesions of the skin at the site of application included epidermal hyperplasia, sebaceous gland hyperplasia, chronic active inflammation, parakeratosis and ulcer. The incidence and severity of these skin lesions generally increased with dose in males and females. Greater incidences of renal tubule regeneration in 100, 200 and 400 mg/kg bw/day females were also observed. Hence, the NOAEL for systemic effects of the test substance can be considered to be 50 mg/kg bw/day and the LOAEL for local effects 25 mg/kg bw/day.

In a subchronic dermal study (NTP report 479, 2001), 0, 50, 100, 200, 400 or 800 mg/kg bw/day of structurally similar amides, C8-18 (even-numbered) and C18 -unsatd., N,N-bis(hydroxyethyl) administered for 14 weeks to groups of 10 male and 10 female B6C3F1 mice resulted in a NOAEL of 100 mg/kg bw/day. The NOAEL was established based on the incidence of chronic active inflammation in all dose groups at 200 mg/kg bw/day and above. The incidences and severities of these skin lesions generally increased with increasing dose. At 400 mg/kg bw/day significant increase in relative liver weights was noted in females and at 800 mg/kg bw/day significant increase in relative liver, kidney (male and females) and lung weights (females) in males and females. Hence, the NOAEL of the test substance for systemic effects can be considered to be 200 mg/kg bw/day and the LOAEL for the local effects 50 mg/kg bw/day.

 Two year chronic dermal studies (NTP report 479, 2001) performed to assess the carcinogenic affects of structurally similar amides, C8-18 (even-numbered) and C18-unsatd., N,N-bis(hydroxyethyl) conducted in rats and mice allow derivations of NOAEL (rat) and LOAEL (mouse) as described in the following:

F344/N rats were administered doses of 0, 50, or 100 mg/kg bw/day of test substance (0, 85, or 170 mg/mL in ethanol). Five exposures per week were given for 104 weeks. The animals were observed twice daily, body weights and clinical findings were recorded periodically. Necropsy and histopathology was performed on all animals. The survival rates of treated male and female rats were similar to those of the vehicle controls. The mean body weights of dosed males and females were similar to those of the vehicle controls throughout the study. The only chemical-related clinical finding was skin irritation at the site of application in the 100 mg/kg bw/day dose group (females). There were marginal increases in the incidences of renal tubule adenoma or carcinoma (combined) in 50 mg/kg bw/day dose group (females). The severity of nephropathy increased with dose in female rats. Non-neoplastic lesions of the skin at the site of application included epidermal hyperplasia, sebaceous gland hyperplasia, parakeratosis and hyperkeratosis, and the incidences and severities of these lesions increased with dose and ranged from minimal to mild. The incidences of chronic active inflammation, epithelial hyperplasia and epithelial ulcer of the forestomach increased with dose in female rats, and the increases were significant in the 100 mg/kg bw/day group. There was no evidence of carcinogenic activity of the test substance in male rats administered 50 or 100 mg/kg bw/day. There was an equivocal evidence of carcinogenic activity in female rats based on a marginal increase in the incidences of renal tubule neoplasms. However, the absence of an increase in neoplasms in the 100 mg/kg bw/day group in the presence of increased hyperplasia makes the association with chemical exposure uncertain. Hence, the NOAEL for systemic effects and local effects was 50 mg/kg bw/day.

B6C3F1 mice were administered doses of 0, 100 or 200 mg/kg bw/day of test substance (0, 50 or 100 mg/mL in ethanol). Five exposures per week were given for 104 to 105 weeks. The animals were observed twice daily, body weights and clinical findings were recorded periodically. Necropsy was performed on all animals and complete histopathology was performed. Survival of dosed male and female mice was generally similar to that of the vehicle controls. Mean bodyweights of 100 mg/kg bw/day females from week 93 and 200 mg/kg bw/day females from week 77 were less than those of the vehicle controls. The only clinical finding attributed to treatment was skin irritation at the site of application in males administered 200 mg/kg bw/day. The incidences of hepatic neoplasms (hepatocellularadenoma, hepatocellular carcinoma and hepatoblastoma) were significantly increased in male and/or female mice. The incidences of eosinophilic foci in dosed groups of male mice were increased relative to that in the vehicle controls. The incidences of renal tubule adenoma and renal tubule adenoma or carcinoma (combined) were significantly increased in 200 mg/kg bw/day males. Several non-neoplastic lesions of the skin at the site of application were considered treatment-related. Incidences of epidermal hyperplasia, sebaceous glandhyperplasia and hyperkeratosis were greater in all dosed groups of males and females than in the vehicle controls. The incidences of thyroid gland follicular cell hyperplasia in all test groups of males and females were significantly greater than those in the vehicle control group. Based on the above results, there was a clear evidence of carcinogenic activity in male B6C3F1 mice however this has been attributed to the presence of free DEA. Therefore, the LOAEL for systemic and local effects was 100 mg/kg bw/day.

Inhalation:

Due to the low vapour pressure inhalation exposure is unlikely to occur under the conditions of normal and foreseeable use and therefore health effects from inhalatory exposure to the substance is considered not to be of concern.

The following information is taken into account for any hazard / risk assessment:

A 28 day oral repeated dose toxicity study conducted with structurally similar, amides, C12-18(even-numbered) and C18 -unsatd., N,N-bis(hydroxyethyl) in rats revealed a NOAEL of >750 mg/kg bw/day based on absence of treatment-related effects at any of the dose levels (Potokar, 1983). A 90 day oral study conducted on LDEA from which a NOAEL of 50 mg/kg bw/day was established, showed effects at higher dose levels. However, it is unclear whether the effects noted were related to the test substance itself or were a result of the nutritional deficiencies due to the unpalatability of the diet as evidenced by scattering of food (Sharratet al., 1961). Thus it is scientifically justified to use the 28 day oral rat NOAEL of 750 mg/kg bw/day for this chemical safety assessment as the point of departure (POD).

Regarding the dermal route, 90 day and 2 year chronic studies on structurally similar substance, C8-18 and C18-unsatd., N,N-bis(hydroxyethyl) was conducted in both rats and mice. The following dose descriptors have been derived based on those studies:

·        Sub-chronic dermal rat: NOAEL for systemic effects: 50 mg/kg bw/day based on renal tubule regeneration, alteration in body weights, haematological and reduced cholesterol and triglyceride levels at the higher doses; LOAEL for local effects: 25 mg/kg bw/day based on minimal severity neoplastic (epidermal and sebaceous gland hyperplasia in males) lesions of the skin at the lowest dose.

·        Sub-chronic dermal mouse: NOAEL for systemic effects: 200 mg/kg bw/day based on organ-weight changes and epididymal spermatozoal concentration at higher doses; LOAEL for local effects: 50 mg/kg bw/day based on minimal severity epidermal and sebaceous gland hyperplasia at the lowest dose.

·        Chronic dermal rat: NOAEL for systemic effects: 50 mg/kg bw/day based on pancreatic acinar atrophy and nephropathy at the higher dose; NOAEL for local effects: 50 mg/kg bw/day based on non-neoplastic lesions at higher doses

·        Chronic dermal mouse LOAEL: 100 mg/kg bw/day based on body weight changes, thyroid gland follicular cell hyperplasia, hepatic neoplasms, renal tubule adenoma, and several non-neoplastic lesions of the skin at all dose levels.

The NOAEL for systemic effects of 50 mg/kg bw/day based on the chronic dermal study in rats will be used to derive the DNEL dermal systemic value since as it is also the lowest NOAEL (50 mg/kg bw/day) below the consistent highest LOAEL (i.e., 100 mg/kg bw/day) obtained across all the sub-chronic and chronic studies in rats and mice.

To derive dermal DNEL for local effects the lowest NOAEL of 50 mg/kg bw/day from the chronic dermal rat study will be used.

Justification for selection of repeated dose toxicity via oral route - systemic effects endpoint:

The oral 28-day study is the more robust since the effects noted in the 90 day study was considered to be caused by the decrease in food intake due to the palatability of the diet which may have also influenced the other effects noted in the higher dose groups as noted by the investigator.

Justification for selection of repeated dose toxicity inhalation - systemic effects endpoint:

Due to the low vapour pressure amides, C16-18 (even-numbered) and C18-unsatd., N,N-bis(hydroxyethyl) inhalation exposure is unlikely to occur under the conditions of normal and foreseeable use and therefore health effects from inhalatory exposure to the substance is considered not to be of concern.

Justification for selection of repeated dose toxicity inhalation - local effects endpoint:

Due to the low vapour pressure amides, C16-18 (even-numbered) and C18-unsatd., N,N-bis(hydroxyethyl) inhalation exposure is unlikely to occur under the conditions of normal and foreseeable use and therefore health effects from inhalatory exposure to the substance is considered not to be of concern.

Justification for selection of repeated dose toxicity dermal - systemic effects endpoint:

As a conservative approach and giving preference to a longer duration study, the NOAEL for systemic effects of 50 mg/kg bw/d based on the chronic dermal study in rats will be used as the POD to derive the DNEL dermal systemic value. This is also the lowest NOAEL (50 mg/kg bw/d) below the consistent highest LOAEL obtained across all the sub-chronic and chronic studies (i.e., 100 mg/kg bw/d).

Justification for selection of repeated dose toxicity dermal - local effects endpoint:

The lowest NOAEL of 50 mg/kg bw/d, this dose level will be further used in this chemical safety assessment as POD for dermal DNEL for long term local effects.

Repeated dose toxicity: dermal - systemic effects (target organ) digestive: stomach; other: skin

Justification for classification or non-classification

Based on the observed effects in all the above repeated dose studies and the available NOAELs and LOAELs, it can be concluded that amides, C16-18 (even-numbered) and C18-unsatd., N,N-bis(hydroxyethyl) does not require classification according to EC criteria (67/548/EEC) and CLP criteria (EC 1272/2008).

 

Categories Display