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Toxicological information

Repeated dose toxicity: oral

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Administrative data

Endpoint:
sub-chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
supporting study
Study period:
1996
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Justification for type of information:
Ethanol/ Ethyl Alcohol is both reagents used in the manufacture of Reaction mass of ethanol and sodium O-ethyl dithiocarbonate and sodium hydroxide. Therefore, Ethanol/ Ethyl Alcohol need to be considered in the assessment of Reaction mass of ethanol and sodium O-ethyl dithiocarbonate and sodium hydroxide

Data source

Reference
Reference Type:
publication
Title:
NTP Technical Report on Toxicity Studies on Urethane in Drinking water and Urethane in 5% Ethanol Administered to F344 Rats and B6C3F1 Mice.
Author:
National Toxicology Program (1996)
Year:
1996
Bibliographic source:
NTP, Research Triangle Park, NC, USA

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
EPA OPPTS 870.3100 (90-Day Oral Toxicity in Rodents)
Deviations:
yes
Remarks:
single dose only used, no opthalmology, clinical chemistry or haematology.
Principles of method if other than guideline:
The study was primarily to assess the toxicity of urethane with ethanol as the vehicle. From the perspective of ethanol, the study involved a test of ethanol and a set of unexposed controls
GLP compliance:
not specified
Limit test:
no

Test material

Reference
Name:
Unnamed
Test material form:
other: liquid
Details on test material:
-Ethyl Alcohol
Ethanol/ Ethyl Alcohol is both reagents used in the manufacture of Reaction mass of ethanol and sodium O-ethyl dithiocarbonate and sodium hydroxide. Therefore, Ethanol/ Ethyl Alcohol need to be considered in the assessment of Reaction mass of ethanol and sodium O-ethyl dithiocarbonate and sodium hydroxide

Test animals

Species:
mouse
Strain:
B6C3F1
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Age at study initiation: 43 - 46 days.
- Source: Taconic Farms, Germantown, NY
- Quarantine: 18-21 days.
- Animals checked for rodent virus antibodies before and after study (and found to be negative.)
- Housing: singley
- Food ad libitum: NIH-07 open formula diet (Zeigler Brothers Inc, Gardeners, PA)
- Water ad libitum

- temperature: 69-75F
- humidity: 35-65%
- Light cycle: 12 hours dark/12 hours light
- air changes: 10 per hour minimum

IN-LIFE DATES: From: Males: 10/12th December 1990 to: 12/14th March 1991. Females: 11/13th December 1990 to: 13/15th March 1991

Administration / exposure

Route of administration:
oral: drinking water
Vehicle:
water
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS: Ethanol was diluted in deionized water. Storage at 4C +/-3 for max of 3 weeks before renewal.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Stability of dosing solutions checked throughout study and found to be within 10% of nominal concentration at all times.
Duration of treatment / exposure:
90 days
Frequency of treatment:
7 days/week ad libitum
Doses / concentrations
Remarks:
Doses / Concentrations:
5% w/v in deionized water
Basis:
nominal in water
No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle
Details on study design:
- Rationale for selecting satellite groups: Satellite animals were included for haematological and clinical chemistry examination at 3 and 23 days.
- Doses were based on literature reports

Examinations

Observations and examinations performed and frequency:
DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Weekly

BODY WEIGHT: Yes
- Time schedule for examinations: Weekly

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): Yes
- Time schedule for examinations: Weekly
Sacrifice and pathology:
Complete necropsies performed on all animals
GROSS PATHOLOGY: Yes (see below)
HISTOPATHOLOGY: Yes (see below)
Organs wieghed: heart, right kidney, liver, lungs, right testis, and thymus.
Histopathologic Examination: adrenal glands, brain (three sections), clitoral glands, esophagus, eyes (if grossly abnormal), femur and marrow, gallbladder (mice only), gross lesions and tissue masses, heart, kidneys, large intestine (cecum, colon, rectum), liver (two sections), lungs, lymph nodes (mandibular and mesenteric), mammary gland, nasal cavity and turbinates (three sections), ovaries, pancreas, parathyroid glands, pituitary gland, preputial glands, prostate gland, salivary gland, seminal vesicle, small intestine (duodenum, jejunum, ileum), spinal cord and sciatic nerve (if neurologic signs were present), spleen, stomach (forestomach and glandular stomach), testes (with epididymis), thigh muscle (if neuromuscular signs were present), thymus, thyroid gland, trachea, urinary bladde,r uterus, and vagina (females in vaginal cytology studies only).
Other examinations:
Sperm motility was assessed at termination. Source used left epididymis. Parameters examined: sperm motility, density and spermatid head count.
Vaginal cytology was assessed for all females of each group by daily (final 12 days) from vaginal smear and cell staining. Relative number of leukocytes, mucleated epithelial cells and large squamous epithelial cells uded to identify estrous stage.
Statistics:
Statistical tests were t-tests and F-tests.
Organ and body weight: parametric multiple comparisons procedures of Williams or Dunnett. Sperm analysis: Non parametric analysis method of Shirley or Dunn.
Jonckheere's test to assess significance of dose response trends and whether a trend sensitive test (William's or Shirley's test was more appropriate for pairwise comparisons than a test that does not assume a monotonic dose response (Dunnett's or Dunn's test).
Significance tested at p<0.05 and p<0.01.

Results and discussion

Results of examinations

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 examined
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
no effects observed
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
not specified
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Histopathological findings: neoplastic:
not examined
Details on results:
ORGAN WEIGHTS
Males: relative and absolute liver weight was increased and there were increases in absoluted heart, liver, kidney and lung weight. No effects in females.

HISTOPATHOLOGY
Males: Minimal nephropathy occurred in 30% of treated animals and in 10% controls which may be biologically but is not statistically significant. Females: Minimal nephropathy occurred in 1 treated animals but not in controls which may be biologically but is not statistically significant. However, this finding is not regarded as conclusive

OTHER: Sperm count in the cauda epididymis was decreased (~30%). No effects on estrous cycle length. The only treatment-related change in female mice was a small change to the time spent in dioestrus and pro-oestrus but it was unclear whether this was significant. Cycle length was not significantly changed.

Effect levels

open allclose all
Dose descriptor:
NOAEL
Effect level:
< 9 700 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male
Basis for effect level:
other: organ weight changes, decreased sperm count, marginal nephropathy
Dose descriptor:
NOAEL
Effect level:
> 9 400 mg/kg bw (total dose)
Based on:
test mat.
Sex:
female
Basis for effect level:
other: no clear significant biological changes seen

Target system / organ toxicity

Critical effects observed:
not specified

Applicant's summary and conclusion

Conclusions:
In a well conducted study that closely followed guidelines, mice were dosed exposed to ethanol in drinking water at a level of 5% for a period of 90 days. Only a single dose level was used as the study was primarily looking at the toxicology of urethane. To establish the effect of ethanol on urethane disposition, two parallel studies were run, one using distilled water as the vehicle for the urethane, the second using 5% ethanol solution as the vehicle. The study allowed a comparison of the two vehicles used. In male mice, relative and absolute liver weight was increased and there were increases in absoluted heart, liver, kidney and lung weight. There was some evidence for a marginal increase in nephropathy in male mice, but the increase was not statistically clear. Sperm count in the cauda epididymis was also decreased (~30%). Female mice showed no effects apart from a small change to the time spent in dioestrus and pro-oestrus but it was unclear whether this was either statisticall or biologically significant. Cycle length was not significantly changed. A marginal NOAEL of >5% (>9400mg/kg) is selected for females and a LOAEL of 9700mg/kg for males.
Ethanol/ Ethyl Alcohol is both reagents used in the manufacture of Reaction mass of ethanol and sodium O-ethyl dithiocarbonate and sodium hydroxide. Therefore, Ethanol/ Ethyl Alcohol need to be considered in the assessment of Reaction mass of ethanol and sodium O-ethyl dithiocarbonate and sodium hydroxide
Executive summary:

In a well conducted study that closely followed guidelines, mice were dosed exposed to ethanol in drinking water at a level of 5% for a period of 90 days. Only a single dose level was used as the study was primarily looking at the toxicology of urethane. To establish the effect of ethanol on urethane disposition, two parallel studies were run, one using distilled water as the vehicle for the urethane, the second using 5% ethanol solution as the vehicle. The study allowed a comparison of the two vehicles used. In male mice, relative and absolute liver weight was increased and there were increases in absoluted heart, liver, kidney and lung weight. There was some evidence for a marginal increase in nephropathy in male mice, but the increase was not statistically clear. Sperm count in the cauda epididymis was also decreased (~30%). Female mice showed no effects apart from a small change to the time spent in dioestrus and pro-oestrus but it was unclear whether this was either statisticall or biologically significant. Cycle length was not significantly changed. A marginal NOAEL of >5% (>9400mg/kg) is selected for females and a LOAEL of 9700mg/kg for males.