Amides, C12-18 and C18-unsatd., N,N-bis(hydroxyethyl)

Repeated dose toxicity: dermal

Administrative data

Endpoint:

chronic toxicity: dermal

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Not available

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

test procedure in accordance with generally accepted scientific standards and described in sufficient detail

Cross-reference

Cross-reference 1

Reason / purpose for cross-reference:

reference to same study

Cross-reference 2

Reason / purpose for cross-reference:

reference to other study

Data source

Reference

Reference Type:

publication

Title:

Unnamed

Year:

2001

Materials and methods

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

Test material

Test material information

Constituent 1

Reference substance name:

Amides, C8-18 (even numbered) and C18-unsatd., N,N-bis(hydroxyethyl)

EC Number:

931-329-6

Molecular formula:

The alkyl chain length of the amide ranges between 8 and 18 carbon atoms

IUPAC Name:

Amides, C8-18 (even numbered) and C18-unsatd., N,N-bis(hydroxyethyl)

Test material form:

liquid: viscous

Test animals

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 (PJ 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

Administration / exposure

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

Duration of treatment / exposure:

104 wk

Frequency of treatment:

Five exposures per week

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 mg/kg bw/day (nominal)

Doses / concentrations 2

Dose / conc.:

50 mg/kg bw/day (nominal)

Remarks:

Corresponding to 85 mg/mL in ethanol

Doses / concentrations 3

Dose / conc.:

100 mg/kg bw/day (nominal)

Remarks:

Corresponding to 170 mg/mL in ethanol

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.

Examinations

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.

Results and discussion

Results of examinations

Clinical signs:

effects observed, treatment-related

Dermal irritation:

effects observed, treatment-related

Description (incidence and severity):

The only clinical finding attributed to dosing was irritation of the skin at the site of application in 100 mg/kg bw/d females.

Mortality:

mortality observed, treatment-related

Description (incidence):

Survival rates of dosed male and female rats were similar to those of the vehicle controls.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

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

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

Description (incidence and severity):

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.

Histopathological findings: neoplastic:

effects observed, treatment-related

Description (incidence and severity):

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 bw 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 bw male rats was significantly greater than that in the vehicle controls.

Effect levels

1

Key result

Dose descriptor:

NOAEL

Remarks:

(systemic effects)

Effect level:

ca. 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

2

Key result

Dose descriptor:

LOAEL

Remarks:

(local effects)

Effect level:

ca. 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

Target system / organ toxicity

Critical effects observed:

not specified

Any other information on results incl. tables

For detailed tables kindly refer to the attached background materials section of the IUCLID.

Applicant's summary and conclusion

Conclusions:

Under the study conditions, the 2-yr NOAEL was considered to be 50 mg/kg bw/day in rat.

Executive summary:

A study was conducted to evaluate the long-term repeated dose dermal toxicity of the test substance, C8-18 and C18-unsatd. DEA, in compliance with GLP. The experiment was conducted in compliance with GLP. Groups of 50 male and 50 female rats were exposed to 0, 50 or 100 mg/kg bw/day of the substance in ethanol by dermal application, 5 times per week, for 104 weeks. Mortality, clinical signs and bodyweight were recorded throughout the study. At necropsy, a gross macroscopic examination and complete histopathology were carried out. The survival rates of treated male and female rats were similar to those of controls. There were no significant differences in bodyweight throughout the groups. The only treatment-related clinical finding was irritation of the skin at the site of application in 100 mg/kg bw/day females. 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. There were marginal increases in the incidences of renal tubule adenoma or carcinoma (combined) in 50 mg/kg bw/day females. The severity of nephropathy increased with increasing dose in female rats. 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. Under the study conditions, there was no evidence of carcinogenic activity of the test substance in male rats at any dose. 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 treatment uncertain. Under the study conditions, the 2-yr NOAEL was considered to be 50 mg/kg bw/day in rat (Irwin, 2001).