Rape oil, reaction products with diethanolamine

Endpoint summary

Description of key information

A 90-d oral study conducted on structurally similar 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 (Sharrat et al., 1961). Furthermore, a 28-day oral repeated dose toxicity study conducted with structurally similar, 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 (Potokar, 1983). Therefore it is scientifically justified to use the subacute oral rat NOAEL of 750 mg/kg bw/d to derive the oral DNEL.
Regarding the dermal route, the following dose descriptors resulted from 90-d and 2-yr NTP studies on structural analogue ODEA: 
• Sub-chronic dermal rat: NOAEL (systemic effects): 100 mg/kg bw/d based on body weight, organ weight and clinical chemistry alterations at the ≥200 mg/kg bw/d; NOAEL (local effects): 25 mg/kg bw/d based on non-neoplastic lesions of the skin at ≥50 mg/kg bw/d. Sub-chronic dermal mouse LOAEL for systemic effects: 50 mg/kg bw/d based organ weight changes at ≥50 mg/kg bw/d; LOAEL for local effects: 50 mg/kg bw/d based on non-neoplastic lesion of the skin
• Chronic dermal rat: NOAEL (systemic effects): 50 mg/kg bw/d based on BW changes at the LOAEL; LOAEL (local effects): 50 mg/kg bw/d based on non-neoplastic lesion of the skin at all tested doses. Chronic dermal mouse: NOAEL (systemic effects): 15 mg/kg bw/d based on body weight changes; LOAEL (local effects): 15 mg/kg bw/d based on non-neoplastic lesions of the skin. 
Giving preference to rat species as well as considering the highest NOAEL below the lowest LOAEL, the NOAEL for systemic effects of 50 mg/kg bw/d from the 2-yr study in rats will be used. For the local effects, since the 90-d and 2-yr rat studies result in LOAELs of 50 mg/kg bw/d this will be further used.

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: see 'Remark'

Remarks:

Read across study conducted on structural analogue amides, C12-18(even-numbered) and C18(unsatd.), N,N-bis(hydroxyethyl) hence maximum reliability rating of 2 assigned according to ECHA guidance, although study is conducted equivalent or similar to OECD guideline 407.

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 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

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: inhalation - local effects

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

Study period:

From 06 May 1993 to 16 May 1995

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Study well documented, meets generally accepted scientific principles, acceptable for assessment and compliant with GLP. Conducted on the structural analogue oleic acid diethanolamine condensate.

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 2-year toxicity study in F344/N rats was conducted by NTP to evaluate the carcinogenic potential by the repeated dermal exposure to the test substance.

GLP compliance:

yes

Species:

rat

Strain:

Fischer 344

Sex:

male/female

Details on test animals or test system and environmental conditions:

- Source: Taconic Laboratory Animals and Services (Germantown, NY)
- Age at study initiation: 7 wk
- Housing: Housed individually in Polycarbonate cages
-Method of distribution: Animals were distributed randomly into groups of approximately equal initial mean body weights.
- Bedding: Sani-Chip® heat-treated hardwood chips (P.J. Murphy Forest Products Corp., Montville, NJ), changed weekly
- Diet: NIH-07 open formula pelleted diet (Zeigler Brothers, Inc., Gardners, PA), available ad libitum
- Water: Tap water (Columbus municipal supply) via automatic watering system (Edstrom Industries, Inc., Waterford, WI), available ad libitum
- Acclimation period:Time held before studies: Males: 13 d and Females: 14 d
- Cages: Polycarbonate (Lab Products, Inc., Maywood, NJ), changed weekly and rotated every 2 wks
- Animal number per cage: 1
- Cage Filters: Spun-bonded polyester Du Pont 2024 (Snow Filtration, Co., Cincinnati, OH), changed every 2 wks
- Racks: Stainless steel drawer-type (Lab Products, Inc., Maywood, NJ), changed and rotated every 2 wks

ENVIRONMENTAL CONDITIONS
- Temperature: 21.1-23.3°C
- Relative humidity: 31-73 %
- Air changes: 10/h
- Photoperiod: 12 h dark/12 h light

IN-LIFE DATES: From: 1993-05-06 To: 1995-05-16

Type of coverage:

open

Vehicle:

ethanol

Details on exposure:

PREPARATION OF DOSING SOLUTIONS: The dose formulations were prepared every 3 weeks by mixing the test substance by stirring or sonicating with 95% ethanol to give the required concentrations. The test substance formulations were applied on shaved skin of the test animals.
-The dose formulations were stored at room temperature, protected from light, in amber glass bottles for up to 28 days.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Periodic analyses of the dose formulations of the test substance from the beginning, middle, and end of the studies were analyzed at the study laboratory using HPLC. All the samples from the formulations were analysed every 9 weeks during the 2-year study and were within 10% of the target concentration.

-Stability of dose formulations: Stability was confirmed for at least 28 days when stored in sealed containers, protected from ultraviolet light, at up to room temperature or for 3 hours when stored open to air and light.

Duration of treatment / exposure:

2-yr

Frequency of treatment:

5 exposures/wk

Remarks:

Doses / Concentrations:
0, 50, 100 mg/kg bw/day (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: The dose selection was based primarily on the incidences and severities of skin lesions observed at the site of application during the 13-week dermal studies. The doses of 200 and 400 mg/kg bw/day exhibited reduced mean body weight and body weight gain along with high incidences of skin lesions at the site of application; thus were considered inappropriate for a 2-year study. Further, lesions of the skin were also present at the site of application in groups administered 100 mg/kg bw/day; however, the incidence were less than those observed in the 200 and 400 mg/kg bw/day groups. Moreover, it was considered unlikely that these lesions would progress and become life threatening over the period of a 2-year study. Therefore, 100 mg/kg bw/day was selected as the high dose for rats in the 2-year study. In groups treated with 50 mg/kg bw/day, the incidences of skin lesions diminished considerably and lesion severities were minimal. Therefore, 50 mg/kg bw/day was selected as the low dose.

Observations and examinations performed and frequency:

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Clinical findings were recorded monthly and at the end of the studies.
BODY WEIGHT: Yes
- Time schedule for examinations: Weighed initially, weekly for 13 weeks, approximately monthly thereafter and again at the end of the studies
HAEMATOLOGY: No
CLINICAL CHEMISTRY: No

Sacrifice and pathology:

SACRIFICE: At the end of the 2-year study animals were sacrificed by carbon dioxide asphyxiation.

GROSS PATHOLOGY: Yes

HISTOPATHOLOGY: Yes

- Complete histopathology was performed on all the rats at the end of the study. In addition to gross lesions and tissue masses, the tissues examined were: adrenal gland, bone with marrow, brain, clitoral gland, esophagus, gallbladder (mice), heart with aorta, large intestine (cecum, colon and rectum), small intestine (duodenum, jejunum and 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 (site of application), spleen, stomach (forestomach and glandular), testis with epididymis and seminal vesicle, thymus, thyroid gland, trachea, urinary bladder, and uterus.

- All major tissues were fixed and preserved in 10% neutral buffered formalin processed and trimmed, embedded in paraffin, sectioned to a thickness of 5 to 6 μm, and stained with hematoxylin and eosin for microscopic examination.

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 non-neoplastic 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 pair wise 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. Values of P greater than 0.5 are presented as 1-P with the letter N added to indicate a lower incidence or negative trend in neoplasm occurrence relative to the control group (e.g., P=0.99 is presented as P=0.01N).

Analysis of Continuous Variables: Organ and body weight data were analysed using the parametric multiple comparison procedures of Dunnett (1955) and Williams (1971, 1972).

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

(no effects on survival, but there was a treatment-related mild to moderate irritation of the skin at the site of application in dosed males and females; vehicle control, 0/50; 50 mg/kg bw/day, 17/50; 100 mg/kg bw/day, 32/50; females: 3/50, 46/50, 50/50).

Dermal irritation:

effects observed, treatment-related

Description (incidence and severity):

(reduced mean body weights of males (i.e., slightly) and females at 100 mg/kg bw/day ).

Mortality:

mortality observed, treatment-related

Description (incidence):

(no effects on survival, but there was a treatment-related mild to moderate irritation of the skin at the site of application in dosed males and females; vehicle control, 0/50; 50 mg/kg bw/day, 17/50; 100 mg/kg bw/day, 32/50; females: 3/50, 46/50, 50/50).

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

(reduced mean body weights of males (i.e., slightly) and females at 100 mg/kg bw/day ).

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:

no effects observed

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

(see below under the 'details on results' section)

Histopathological findings: neoplastic:

no effects observed

Description (incidence and severity):

(see below under the 'details on results' section)

Details on results:

Histopathology:
- Non-neoplastic lesions:
- Skin lesions: Minimal to moderate non-neoplastic lesions of the skin at the site of application were observed. The major alterations included (thickening of the epidermis, sebaceous gland and epidermal hyperplasia, hyperkeratosis, parakeratosis, chronic active dermal inflammation and ulcer) which were significantly increased in dosed males and females relative to the vehicle control.
- Forestomach lesions: No significant treatment related effects were observed. The increased incidence of hyperkeratosis and ulceration in 50 mg/kg bw/day males were not considered to be treatment related as these effects were not observed in females.
- Testis: No significant treatment related effects were observed.
- Thyroid gland lesions: No significant treatment related effects were observed.

- Neoplastic lesions:
- Skin lesions: No significant treatment related effects were observed. Few observed skin neoplasms (one subcutaneous fibroma in one vehicle control male and one subcutaneous fibrosarcoma in each of the 50 and 100 mg/kg bw/day male groups) were not considered to be significant as the incidences did not follow a pattern indicative of an association with the test substance.
- Testis: No significant treatment related effects were observed. Increased interstitial cell adenoma in males at 100 mg/kg bw/day was not considered significant as similar incidences were reported in vehicle controls in historical NTP dermal studies.
- Thyroid gland: No significant treatment related effects were observed. Marginal increase in the incidence of follicular cell adenoma or carcinoma observed in males at 50 mg/kg bw/day was not considered dose related and no follicular cell hyperplasias were observed .

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: body weight changes at the LOAEL

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: irritation and non-neoplastic lesion of the skin at both the tested doses

Critical effects observed:

not specified

Conclusions:

Under the test conditions, the NOAEL for systemic effects can be considered to be at 50 mg/kg bw/d and for local effects the LOAEL can be considered at the lowest dose of 50 mg/kg bw/d.

Executive summary:

A study was conducted by NTP to evaluate the carcinogenic potential following repeated dermal exposure to oleic acid diethanolamine condensate (ODEA) in F344/N rats.

Groups of 50 male and 50 female rats were dermally exposed to 0, 50 or 100 mg test substance/kg bw/day in ethanol at a frequency of 5 d/wk for a period of 104 weeks. Survival, clinical findings, body weight and histopathology of different organs were evaluated at specific time intervals.

Survival of the dosed male and female rats was similar to that of the vehicle control groups. The mean body weights of males and females (week 24 onwards) were reduced than those of the vehicle control group at 100 mg/kg bw/day. Dose dependent increase in irritation (i.e., mild to moderate) and non-neoplastic lesions (i.e., minimal to moderate) of the skin was observed at the site of application in males and females. The non-neoplastic lesions included epidermal hyperplasia, sebaceous gland hyperplasia, hyperkeratosis, parakeratosis, chronic active dermal inflammation, and ulcer. No significant neoplastic lesions or evidence of carcinogenic activity was observed at any tested dose levels in skin, testis and thyroid gland.

As a result, for systemic effects, the NOAEL can be considered to be at 50 mg/kg bw/d and for local effects the LOAEL can be considered at the lowest dose of 50 mg/kg bw/d.

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 requirement for this tonnage band is sufficiently met with the available data.

Repeated dose toxicity: dermal - local effects

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

LOAEL

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

In a 90-day oral study conducted in rats (Gaunt IF et al., 1965), the no observed effect level (NOEL) was determined to be 50 mg/kg bw/d for structurally similar lauric acid diethanolamine condensate (LDEA) based on growth retardation, biochemical changes and an increase in kidney weights seen in the 250 mg/kg bw/d dose group and growth retardation, haematological effects (anaemia) and increased kidney and liver weights seen in the 500 and 1000 mg/kg bw/d dose groups. The growth retardation observed at 250 mg/kg bw/d was considered to be associated with diminished food intake which was attributed to the palatability of the test substance. This was further considered to have influenced the other effects noted in the higher dose groups as noted by the investigator.

A 28-day oral study conducted in rats (Potokar, 1983) with the structurally similar amides, 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 biologically relevant parameters investigated at any of the dose levels tested (i.e. 70 , 250 , 750 mg/kg bw/day. There were changes in the fore-stomach at some doses including controls however the authors attributed this to the use of olive oil and found the fore-stomach changes to be reversible at the end of the exposure period.

Dermal

Two sub-chronic NTP studies (NTP report 481, 1999) were conducted to evaluate the cumulative toxic effects following repeated dermal exposure to the structural analogue oleic acid diethanolamine condensate (ODEA) in F344/N rats as well as inB6C3F1 mice.

In the rat study, groups of 10 male and 10 female rats were dermally exposed to 0, 25, 50, 100, 200, or 400 mg ODEA/kg bw/day in ethanol at a frequency of 5 d/wk for a period of 13 weeks. All animals survived until study termination. The final mean body weights and body weight gains of 200 and 400 mg/kg bw/day males and the mean body weight gain of 400 mg/kg bw/day females were significantly less than those of the vehicle controls. The only chemical-related clinical finding was irritation of the skin at the site of application in most males administered 100 mg/kg bw/day or greater and in all females administered 50 mg/kg bw/day or greater. Segmented neutrophil counts and alkaline phosphatase concentrations were increased significantly on specific days in the 200 and/or 400 mg/kg bw/day male and female groups. Kidney weights of 200 and 400 mg/kg females were significantly greater than those of the vehicle controls. There was a dose dependent increase in the incidence of non-neoplastic lesions of the skin at the site of application, including epidermal hyperplasia, parakeratosis, chronic active dermal inflammation, suppurative epidermal inflammation, and sebaceous gland hypertrophy in dosed rats. However, these effects were minimal in the 50 mg/kg bw/day dose group.

Under the conditions of the 13-week study in rats, the NOAEL for systemic and local effects can be considered to be at 100 and 25 mg/kg bw/day respectively.

In the mice study, groups of 10 male and 10 female mice were dermally exposed to 0, 50, 100, 200, 400 or 800 mg ODEA/kg bw/day in ethanol at a frequency of 5 d/wk for a period of 13 weeks. All male and female mice except one 800 mg/kg bw/day male survived until the end of the study. Final mean body weights and body weight gains of 800 mg/kg bw/day males and females and 400 mg/kg bw/day females were significantly less than those of the vehicle controls. Clinical findings in dosed mice included irritation of the skin at the site of application. Irritation occurred in all surviving dosed males and in most females administered ≥100 mg/kg bw/day. The heart weights of 400 and 800 mg/kg bw/day males and females and 200 mg/kg bw/day females and the kidney weights of 50, 100, and 400 mg/kg bw/day males were significantly greater than those of the vehicle controls. Relative to the vehicle controls, the liver weights were increased in all dosed groups. Dose dependent increase in the incidences of non-neoplastic skin lesions included epidermal hyperplasia, parakeratosis, suppurative epidermal inflammation, chronic active dermal inflammation, sebaceous gland hypertrophy, and ulcer.

Under the conditions of the 13-week study in mice, the LOAEL for both systemic and local effects can be considered to be at 50 mg/kg bw/day.

Two year chronic dermal studies (NTP report 481, 1999) performed to assess the carcinogenic affects of the structural analogue oleic acid diethanolamine condensate (ODEA) 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 ODEA/kg bw/d of the test substance to 50 male and female test animals in each group. Five exposures per week were given for 104 weeks.The mean body weights of males and females (week 24 onwards) were reduced than those of the vehicle control group at 100 mg/kg bw/day. Dose dependent increase in irritation (i.e., mild to moderate) and non-neoplastic lesions (i.e., minimal to moderate) of the skin was observed at the site of application in males and females. The non-neoplastic lesions included epidermal hyperplasia, sebaceous gland hyperplasia, hyperkeratosis, parakeratosis, chronic active dermal inflammation, and ulcer. No significant neoplastic lesions or evidence of carcinogenic activity was observed at any tested dose levels in skin, testis and thyroid gland. Under the test conditions, no evidence of carcinogenic activity was observed with the test substance at any tested dose levels in rats. As a result, for systemic effects, the NOAEL can be considered to be at 50 mg/kg bw/d and for local effects the LOAEL can be considered at the lowest dose of 50 mg/kg bw/d.

B6C3F1 micewere administered doses of 0, 15 or 30 mg ODEA/kg bw/d to 50 male/female test animals in each group. 5 exposures per week were given for 105 weeks.5 males and 5 females were considered for the 3 -month interim evaluation mice. The mean body weights of females (week 76 onwards) were reduced than those of the vehicle control group at 30 mg/kg bw/day. The only significant treatment related clinical finding was irritation of the skin at the site of application in 30 mg/kg bw/day males. The incidences of epidermal hyperplasia, sebaceous gland hyperplasia, and chronic active inflammation of the dermis in all dosed groups were significantly increased relative to the vehicle controls at 3 months and at 2 years. The increased incidences of hyperkeratosis in dosed males at 3 months and in dosed males and females at 2 years, of parakeratosis in 30 mg/kg bw/day males at 3 months and 2 years, and of ulcer in 30 mg/kg bw/day males and exudate in 30 mg/kg bw/day males and females at 2 years were also attributed to the test substance administration. No significant neoplastic lesions or evidence of carcinogenic activity was observed at any tested dose levels in skin and lymph nodes. Under the test conditions, no evidence of carcinogenic activity was observed with the test substance at any tested dose levels in mice. As a result, for systemic effects, the NOAEL can be considered to be at 15 mg/kg bw/d and for local effects, the LOAEL can be considered to be at 15 mg/kg bw/d (NTP 481report, 1999).

Inhalation

Due to the low vapour pressure and high viscosity of HE Rape Oil, reaction product with diethanolamine, inhalation exposure is unlikely to occur under the conditions of normal and foreseeable use and therefore health effects from inhalatory exposure to the substance are considered not to be of concern.

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

A 90-d oral study conducted on a structurally similar LDEA 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 indicating the diet was unacceptable to the animals (Sharrat et al., 1961). Further, 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/d based on absence of treatment-related effects at any of the tested dose levels (Potokar, 1983). Based on this, it is scientifically justified to use the sub-acute oral rat NOAEL of 750 mg/kg bw/d for this chemical safety assessment as the point of departure (POD).

Regarding the dermal route, 90-d and 2-yr chronic studies on the structural analogue ODEA have been conducted in both rats and mice. The following dose descriptors have been derived based on the studies:

• Sub-chronic dermal rat: NOAEL for systemic effects: 100 mg/kg bw/d based on body weight and organ weight changes, clinical chemistry alterations at the ≥200 mg/kg bw/d; NOAEL for local effects: 25 mg/kg bw/d based on irritation at ≥100 mg/kg bw/d and non-neoplastic lesions of the skin at ≥50 mg/kg bw/d.

• Sub-chronic dermal mouse LOAEL for systemic effects: 50 mg/kg bw/d based organ weight changes at ≥50 mg/kg bw/d; LOAEL for local effects: 50 mg/kg bw/d based on non-neoplastic lesion of the skin at all doses.

• Chronic dermal rat: NOAEL for systemic effects: 50 mg/kg bw/d based on body weight changes at the LOAEL; LOAEL for local effects: 50 mg/kg bw/d based on irritation and non-neoplastic lesion of the skin and hyperkeratosis and chronic active inflammation of the forestomach at both the tested doses.

• Chronic dermal mouse: NOAEL for systemic effects: 15 mg/kg bw/d based on body weight changes at 30 mg/kgt bw/d; LOAEL for local effects: 15 mg/kg bw/d based on incidences of non-neoplastic lesions of the skin.

 

Giving preference to rat species and longer duration study as well as considering the highest NOAEL below the lowest LOAEL (i.e., 100 mg/kg bw/d) approach, the NOAEL for systemic effects of 50 mg/kg bw/d from the chronic dermal study in rats will be used as the POD to derive the DNEL dermal systemic value as a conservative approach.

 

Since both the sub-chronic and chronic dermal rat studies available for ODEA results in LOAELs of 50 mg/kg bw/d for the local effects, this dose level will be further used in this chemical safety assessment as POD for dermal DNEL for long term local effects.

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. Both studies were conducted on structural analogues.

Justification for selection of repeated dose toxicity inhalation - systemic effects endpoint:
The substance is a liquid with a low vapour pressure. Due to its physical chemical properties, it is unlikely that it will form inhalable dust, mist or fumes when handled and used in solid form. In case inhalable forms of the substance (either pure or in aqueous solutions) are created under particular conditions (e. g. spraying, elevated temperature/pressure), appropriate risk management measures such as closed systems, exhaust ventilation or wearing of respirators are implemented to control exposure. Under such conditions, the risk to humans following inhalation exposure can be considered minimal and further testing involving vertebrate animals may be omitted, in accordance with Annex XI (1.2) of the REACH regulation.

Justification for selection of repeated dose toxicity inhalation - local effects endpoint:
The substance is a liquid with a low vapour pressure. Due to its physical chemical properties, it is unlikely that it will form inhalable dust, mist or fumes when handled and used in solid form. In case inhalable forms of the substance (either pure or in aqueous solutions) are created under particular conditions (e. g. spraying, elevated temperature/pressure), appropriate risk management measures such as closed systems, exhaust ventilation or wearing of respirators are implemented to control exposure. Under such conditions, the risk to humans following inhalation exposure can be considered minimal and further testing involving vertebrate animals may be omitted, in accordance with Annex XI (1.2) of the REACH regulation.

Justification for selection of repeated dose toxicity dermal - systemic effects endpoint:
Rat long term study and provides the highest NOAEL below the lowest LOAEL (i.e., 100 mg/kg bw/d) approach, the NOAEL for systemic effects of 50 mg/kg bw/d from the chronic dermal study in rats will be used to derive the DNEL dermal systemic value as a conservative approach.

Justification for selection of repeated dose toxicity dermal - local effects endpoint:
No key study selected as both the sub-chronic and chronic dermal rat studies available for the structural analogue ODEA result in LOAELs of 50 mg/kg bw/d for the local effects; this dose level will be further used in this chemical safety assessment to derive the dermal DNEL for long term local effects.

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

Justification for classification or non-classification

Based on the observed effects in the above repeated dose studies and the available NOAELs and LOAELs, it can be concluded that HE Rape Oil, reaction product with diethanolamine does not require classification according to EC criteria (67/548/EEC) and CLP criteria (EC 1272/2008) based on the effects observed in all the repeated dose toxicity studies conducted on structural analogues.