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Toxicological information

Repeated dose toxicity: inhalation

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Administrative data

Endpoint:
sub-chronic toxicity: inhalation
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
1992
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
Fully documented study to a recognised protocol and performed by an organisation known to operate to high standards and that uses a peer review process for its work. Read-across based on supporting substance (analogue approach). The data in this record is for the metabolite 2-butoxyethanol (CAS; 111-76-2, EC no 203-905-0) for which the systemic toxicity will be the same. Full details of the justification for the use of an analogue for this end point are included in the document appended to chapter 13 of this dossier.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2000

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 413 (Subchronic Inhalation Toxicity: 90-Day Study)
Deviations:
yes
Remarks:
, no clinical chemistry, urine analysis, opthalmology
Principles of method if other than guideline:
Range finder study for a carcinogenicity assay. Animals exposed up to concentrations causing clear toxicity and subject to most of the assays normally used in a guideline subchronic study. Those parameters known to be insensitive to 2-butoxyethanol were not examined
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): 2-butoxyethanol
- Physical state: liquid
- Analytical purity: >99%
- Impurities (identity and concentrations): Water 0.02%, 0.00% acetic acid, 105ppm peroxide.
- Lot/batch No.: QP-911021-26D1
- Source: Dow Chemical, Plaquemine, LA
- Stability under test conditions: No degradation detected (monitored for acid, peroxide and by GC)
- Storage condition of test material: Room temperature in the dark.

Test animals

Species:
rat
Strain:
Fischer 344
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Taconic Laboratory Animals and Services, Germantown, NY
- Age at study initiation: 6 weeks
- Housing: individual stainless steel, wire bottomed cages.
- Diet: NIH-07 open formulate, pelleted, ad libitum except during exposure period
- Water: tap, ad libitum, automated watering system
- Acclimation period: 11-12 days in quarantine.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 24C approx
- Humidity (%): 55 approx
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 23/24 March 1992 To: 22-25 June 1992

Administration / exposure

Route of administration:
inhalation: vapour
Type of inhalation exposure:
whole body
Vehicle:
other: unchanged (no vehicle)
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: Stainless steel 1.7m3
- Method of conditioning air: charcoal filtration
- Temperature, humidity, pressure in air chamber: 24C approx, 55% approx.
- Air change rate: 15/hr
- Vapour generation: The test material was held in a stainless-steel reservoir under a nitrogen blanket. The material was pumped into a glass column filled with glass beads and heated by a flexible electric heat tape encircling the column. Vapor temperature was monitored at the top of the condenser column by a temperature sensor. The vapor-laden air was transferred through a heated Teflon distribution line and diluted with HEPA- and charcoal-filtered air. Three-way valves in the chamber inlet ducts allowed vapors to be diverted to the exhaust until a stable concentration of test material was built up in the distribution line. At each chamber, vapor moving through the inlet duct was further diluted with filtered air to the appropriate concentration of test material. The total active mixing volume of each chamber was 1.7 m3. A small particle detector was placed in the chambers to measure concentrations of aerosol. No particle counts above the minimum resolvable level (200 particles/cm3) were detected

TEST ATMOSPHERE
- Brief description of analytical method used: On line gas chromatography, sampling every 15 mins. Calibration with off-line gas chromatography, which were themselves calibrated with gravimetrically prepared samples of 2-butoxyethanol.
- Samples taken from breathing zone: no
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Chamber concentrations were monitored with an on-line gas chromatograph (GC). The monitor was coupled with the inhalation chambers by a computer-controlled 12-port stream select valve. The GC was calibrated by comparing chamber concentration data to data from grab samples analyzed by an off-line GC. The grab samples were collected in bubblers containing water. The off-line GC was calibrated with gravimetrically prepared standards. Chamber concentration uniformity was maintained throughout the study. Buildup and decay rates for chamber concentrations were determined with and without animals in the chambers. The time to achieve 90% of the target concentration and the time for decay to 10% of the target concentration was 12.5 minutes. Studies of 2-butoxyethanol degradation and monitoring for impurities were conducted throughout the studies by comparing bubbler samples to a reference sample. No significant degradation was observed during the studies.
Duration of treatment / exposure:
14 weeks average
Frequency of treatment:
6 hours/day plus chamber equilibration time (12 mins), 5 days per week
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
0, 31, 62.5, 125, 250, 500ppm
Basis:
nominal conc.
Remarks:
Doses / Concentrations:
0, 29.8-32.2, 60.3-63.9, 119-131, 238-260, 478-516 ppm
Basis:
analytical conc.
No. of animals per sex per dose:
10
Control animals:
yes, sham-exposed
Details on study design:
- Dose selection rationale: Maximum dose set as maximum that could be acheived without generating aerosol particles.
Positive control:
No

Examinations

Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Twice daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: weekly and at end of study

BODY WEIGHT: Yes
- Time schedule for examinations: start, weeekly and at end of study

FOOD CONSUMPTION:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: No

WATER CONSUMPTION: No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: at end of study prior to sacrifice - collected from retroorbital sinus
- Anaesthetic used for blood collection: Yes (identity) CO2/O2
- Animals fasted: No data
- How many animals: all survivors
- Parameters checked: erythrocyte, leucocyte, platelet counts; MCV, MCHg, MCHg concentration; hematocrit, leucocyte differential count and nucleated erythrocyte count.

CLINICAL CHEMISTRY: No

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No
Sacrifice and pathology:
HISTOPATHOLOGY: Yes. Complete histopathology performed on top and bottom dose animals plus 250ppm females. Apart from gross lesions, the following were examined: adrenal gland, bone with marrow, brain, clitoral gland, esophagus, heart, large intestine (cecum, colon, rectum), small intestine (duodenum, jejunum, ileum), kidney, larynx, liver, lung, lymph nodes, (mandibular, mesenteric, brochial, mediastinal), mammary gland, nose, overy, pancreas, parathyroid gland, pituitary gland, preputial gland, prostate gland, salivary gland, spleen, stomach (fore and glandular), testes (with epididymis and seminal vesicle), thymus, thyroid gland, trachea, urinary bladder, uterus. Histopathology on remaining animals: bone with marrow, forestomach, kidney, spleen of male rats and nose, salivary gland, tail and thymus of female rats.
Other examinations:
none
Statistics:
Probability of survival estimated by product limit procedure of Kaplan and Meier (1958). Dose related effects tested with Cox's method (1972) for group equality and Tarone's 1975 life table test to identify dose related trends. Reported p values 2 sided. The ply-k test was used to assess for lesion prevalence (modified version of Cochran-Armitage test). Organ and body weight assess using Dunett's parametric multicomparison procedure. Haematology data analysed using method of Shirley and Dunn for nonparametric data along with Jonckheere's method to test for trend. Dixon and Masssey (1951) method used to eliminate outlier values.

Results and discussion

Results of examinations

Clinical signs:
effects observed, treatment-related
Mortality:
mortality observed, treatment-related
Body weight and weight changes:
effects observed, treatment-related
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:
effects observed, treatment-related
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:
CLINICAL SIGNS AND MORTALITY: Six female rats were found moribund and killed during the study: five in the 500 ppm group (four in week 1, one in week 5) and one in the 250 ppm group (in week 8). Clinical findings were most prevalent in rats of both sexes exposed to 125, 250 or 500 ppm and included abnormal breathing, pallor, red urine stains, nasal and eye discharge, lethargy and either increased salivation or lacrimation. All females of the 500 ppm group, particularly during the first two weeks, developed alternating blue and white bands on their tails that caused them to self-mutilate and loose the distal portion of their tails.

BODY WEIGHT AND WEIGHT GAIN: By the end of the study, body weight gains were significantly reduced in females of the 500 ppm group, but were unaffected in all other groups. The mean final body weights (197 +/- 4 g) and body weight gains (89 +/- 3 g) of females exposed to 500 ppm were significantly less than controls (217 +/- 5 and 105 +/- 4 g, respectively).

HAEMATOLOGY: Haematological examination showed that inhalation of 2-butoxyethanol resulted in the development of a persistent and exposure-related macrocytic, normochromic, responsive anaemia, as indicated by decreased haematocrit values, haemoglobin concentrations and erythrocyte counts in the 125 ppm or greater group males and in all groups of exposed females. The effects were dose related and statistically significant, albeit small at the lower doses (eg females at 31ppm: reduction ~5%). This evidence of a sex difference in the severity of the anaemia was also seen in the 500 ppm group, in which the indicators were slightly more severe in the females than in the males. Evidence of an erythropoietic response was shown by increases in reticulocyte and nucleated erythrocyte counts in males of the 125 ppm or greater groups and females of the 62.5 ppm or greater groups. Other haematological changes were decreases in lymphocyte and monocyte counts in males of the 125 ppm or greater groups and increased platelet counts in females of the 125 or 500 ppm groups.

ORGAN WEIGHTS: Some organ weight changes were observed. These were: increases of the kidney of males in the 500 ppm group and females in the 125 ppm or greater groups; increases of the liver of males in the 250 or 500 ppm and females in the 125 ppm or greater groups; and decreases of the thymus of females in the 500 ppm group.

HISTOPATHOLOGY: NON-NEOPLASTIC: Female rats that were killed moribund exhibited a number of histopathologic changes. Thrombosis occurred in a number of tissues in high dose females. Thromboses were associated with areas of infarction in the tail and necrosis in the incisors and liver. Thrombosis was present in the atrium of the heart, in the nasal septum, in central veins of the liver associated with large foci of necrosis, in the lung, the femur, tail, and dental pulp. There were areas of necrosis within bone marrow. Affected marrow was infiltrated by macrophages. In the most severely affected vertebrae in the tail, there was growth plate degeneration with no evidence of renewed longitudinal growth. Atrophy of the spleen and thymus; inflammation, necrosis, ulceration and hyperplasia of the forestomach; centrilobular degeneration of the liver; and renal tubule degeneration were also observed in rats killed moribund. Similar effects were seen in animals that survived to study termination. Bone marrow necrosis and infarcts were found in the tails of all surviving females exposed to 500 ppm. Minimal hematopoetic cell proliferation of the spleen was noted in females exposed to > = 62.5 ppm (N = 1, 10, 8 and all 5 survivors in 62.5, 125, 250 and 500 ppm groups) and all males exposed to > = 250 ppm. Bone marrow hyperplasia was increased in all males exposed to > = 250 ppm and females exposed to > = 62.5 ppm (N = 8, 10, all 9 survivors and all 5 survivors in 62.5, 125, 250 and 500 ppm groups). Increased pigmentation of Kupffer cells in the liver was also noted in males exposed to > = 125 ppm (N = 7, 10 and 10 at 125, 250 and 500 ppm) and all surviving females exposed to > = 62.5 ppm. Renal tubule pigmentation was noted in 8/10 males exposed to 250 ppm, all males exposed to 500 ppm, and all surviving females exposed to > = 125 ppm. Minimal forestomach inflammation and hyperplasia were noted in 2 or 3 males exposed to > = 250 ppm. Epithelial hyperplasia of the forestomach were noted in 1 female each in the 250 and 500 ppm groups.

Effect levels

open allclose all
Dose descriptor:
NOAEC
Effect level:
< 31 ppm
Sex:
female
Basis for effect level:
other: haematology (haematocrit, hemoglobin, erythrocytes all reduced)
Dose descriptor:
NOAEC
Effect level:
62.5 ppm
Sex:
male
Basis for effect level:
other: haematology (as females plus reticulocytes, MCV, leucocytes, monocytes count reduced)

Target system / organ toxicity

Critical effects observed:
not specified

Applicant's summary and conclusion

Conclusions:
No NOAEC was found for female rats. LOAEL was 31 ppm based on haematological effects seen at all doses tested. A NOAEL of 62.5 ppm was found for male rats, based on haematotoxic effects seen at 125 ppm. Females appear to be more senstive to the haematological effects of 2-butoxyethanol than males.
Executive summary:

In a range finder study, rats were exposed by inhalation to butoxyethanol at doses up to 500ppm for a period of 14 weeks. The study approximated to guideline but parameters known to be insensitive to this substance were not examined. Substantial toxicity was noted at the higher doses but the predominant effects at lower doses were adverse changes to the haematology, particularly haematocrit, hemoglobin, erythrocytes reductions. No NOAEC was found for female rats; the LOAEL was 31 ppm based on haematological effects seen at all doses tested. It should be noted that although the effects were dose related and statistically significant, at 31ppm the reduction in these parameters was only around 5%. A NOAEL of 62.5 ppm was found for male rats, based on haematotoxic effects seen at 125 ppm.

Synopsis:

NOAEL (rats, 14 weeks, inhalation): males 62.5ppm, females <31ppm. Similar values would apply for butoxyethyl acetate.