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Toxicological information

Repeated dose toxicity: inhalation

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Administrative data

Endpoint:
chronic toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1993-1995
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
comparable to guideline study
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.
Cross-reference
Reason / purpose for cross-reference:
reference to same study
Reference
Endpoint:
carcinogenicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2000
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Fully documented NTP guideline study. NTP studies are subject to critical peer review process.
Reason / purpose for cross-reference:
reference to same study
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 451 (Carcinogenicity Studies)
Deviations:
no
Remarks:
, no significant deviations noted.
GLP compliance:
not specified
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: 7-8 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: 18 days in quarantine.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 24C approx
- Humidity (%): 57-58
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 23 Aug 1993 To: 18 Aug 1995
Route of administration:
inhalation: vapour
Type of inhalation exposure (if applicable):
whole body
Vehicle:
unchanged (no vehicle)
Details on 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: 2-butoxyethanol under nitrogen micrometer pumped onto a heated glass column filled with glass beads. Output of generator quantified by nitrogen flow and 2-butoxyethanol vapour pressure at exit temperature. Transfer lines heated to prevent condensation. Particle detector used to ensure vapour only entered chamber (particle counts never greater than LOD of 200/cm3).

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:
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.
Duration of treatment / exposure:
2 years
Frequency of treatment:
6 hours/day plus chamber equilibration time (12 mins), 5 days per week
Post exposure period:
none
Remarks:
Doses / Concentrations:
0, 31, 62.5, 125ppm
Basis:
nominal conc.
Remarks:
Doses / Concentrations:
0, 29.0-32.6, 57.6-67.0, 117-133ppm
Basis:
analytical conc.
No. of animals per sex per dose:
50
Control animals:
yes, sham-exposed
Details on study design:
- Dose selection rationale: Based on 90 day range finder study described in chapter 7.5.3. Animal distribution among groups designed to equalise initial mean body weight.
- Rationale for selecting satellite groups: Satellite groups for haematology and bone marrow analysis (27 animals per sex in control, mid and high dose groups, 9 animals per sex for low dose group.) One group each for time point sacrifices of 3, 6 and 12 months.
Positive control:
No
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Twice daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: monthly (fortnightly during last 6 months) and at end of study

BODY WEIGHT: Yes
- Time schedule for examinations: start, monthly (fortnightly during last 6 months) 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 and from satellite groups at 3, 6 and 12 months (not 31.2ppm) - 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, reticulocyte, haemoglobin, leucocyte differential count and nucleated erythrocyte count. Morphological analysis of erythrocytes, leucocytes and platelets. Measurements made using an Ortho ELT-8/ds 9000 analyser. Bone marrow cellularity measurements using a Coulter model Zh counter while cytological evaluations of bone marrow cell morphology and myeloid/erythroid ratios were made microscopically.

CLINICAL CHEMISTRY: No

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No

ORGAN WEIGHTS: not measured.
Sacrifice and pathology:
Complete histopathology on all core animals (not satellites). Apart from gross lesions, the following were examined for gross lesions and tissues fixed and processed for examination. : 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, ovary, 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.
Other examinations:
Haematology at 3 and 6 months using satellite groups.
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.
Clinical signs:
no effects observed
Mortality:
no mortality observed
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:
not examined
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:
BODY WEIGHT AND WEIGHT GAIN: Body weights of male and female rats in the 31.2 and 62.5 ppm groups and male rats of the 125 ppm group were generally similar to the controls throughout the experiment. Body weights of female rats of the 125 ppm group were generally lower than those of the chamber control from week 17 until the end of the experiment.

All non-neoplastic findings are reported in the entry for this study in chapter 7.5.3. Neoplastic findings are reported in the 'remarks on results' field.
Relevance of carcinogenic effects / potential:
No clear evidence of carcinogenic effect
Dose descriptor:
NOAEC
Sex:
male/female
Basis for effect level:
other: The NOAEC for other effects is reported in chapter 7.5.3
Remarks on result:
not determinable
Remarks:
no NOAEC identified. Effect type:carcinogenicity (migrated information)

The incidence of benign or malignant phaeochromocytoma (combined) of the adrenal medulla in females exposed to 125 ppm EGBE was not significantly increased compared to the chamber controls, but it did exceed the historical control range. There was only one malignant phaeochromocytoma, which occurred in the 125 ppm group. The data are summarised in the table below:

Neoplastic and non-neoplastic lesions of the adrenal medulla in female F344/N rats

 

Chamber

control


31.2 ppm


62.5 ppm

125 ppm

No. examined

50

50

49

49

Hyperplasia (severity)

11 (1.9)

11 (2.3)

8 (2.1)

17 (2.5)

Benign phaeochromocytoma

3/50 (6 %)

4/50 (8 %)

1/49 (2 %)

7/49 (14 %)

Benign or malignant phaeochromocytoma

3/50 (6 %)

4/50 (8 %)

1/49 (2 %)

8/49 (16 %)

Two nasal tumours were discovered: a chondroma in a 31.2 ppm male and an adenoma in a 62.5 ppm male. In the absence of any preneoplastic changes, these are considered incidental findings. Although the incidence of hyaline degeneration was significantly increased in all exposed groups of male rats, the severity of the lesion was minimal and was not affected by exposure.

Conclusions:
There was no clear evidence of a carcinogenic effect. The marginal effects that were statistically significantly different to historic but not concurrent controls could not be atttributed with confidence to exposure.
Executive summary:

In a 2 year NTP cancer bioassay in rats exposed to 2 -butoxyethanol by the whole body inhalation route to concentrations up to 125ppm, there was no evidence for carcinogenicity in males. The incidence of benign or malignant phaeochromocytoma (combined) of the adrenal medulla in females exposed to the top dose of 125 ppm was not significantly increased compared to the chamber controls, but it did exceed the historical control range. Within this dose group, there was only a single incidence of malignant phaeochromocytoma. The study report recorded that this finding provided equivocal evidence of carcinogenicity in females, this is a very conservative interpretation and the findings cannot be attributed with confidence to exposure

Synopsis

NOAEL (2yr cancer study, rats)>125ppm

Data source

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

Materials and methods

Test guideline
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 453 (Combined Chronic Toxicity / Carcinogenicity Studies)
Deviations:
no
Remarks:
, no significant deviations noted
Principles of method if other than guideline:
NTP carcinogenicity study
GLP compliance:
yes
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
2-butoxyethanol
EC Number:
203-905-0
EC Name:
2-butoxyethanol
Cas Number:
111-76-2
Molecular formula:
C6H14O2
IUPAC Name:
2-butoxyethanol
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 0.0254%, 0.003% acetic acid, <1000ppm peroxide.
- Lot/batch No.: QP-921215-26D2
- 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 or test system and environmental conditions:
TEST ANIMALS
- Source: Taconic Laboratory Animals and Services, Germantown, NY
- Age at study initiation: 7-8 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: 18 days in quarantine.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 24C approx
- Humidity (%): 57-58
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 23 Aug 1993 To: 18 Aug 1995

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: 2-butoxyethanol under nitrogen micrometer pumped onto a heated glass column filled with glass beads. Output of generator quantified by nitrogen flow and 2-butoxyethanol vapour pressure at exit temperature. Transfer lines heated to prevent condensation. Particle detector used to ensure vapour only entered chamber (particle counts never greater than LOD of 200/cm3).

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:
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.
Duration of treatment / exposure:
2 years
Frequency of treatment:
6 hours/day plus chamber equilibration time (12 mins), 5 days per week
Doses / concentrationsopen allclose all
Dose / conc.:
31 ppm (nominal)
Remarks:
Actual dose 29.0 to 32.6ppm over course of study
Dose / conc.:
62.5 ppm (nominal)
Remarks:
Actual dose 57.6 to 67.0ppm over course of study
Dose / conc.:
125 ppm (nominal)
Remarks:
Actual dose 117 to 133ppm over course of study
No. of animals per sex per dose:
50
Control animals:
yes, sham-exposed
Details on study design:
- Dose selection rationale: Based on range finder study described in this chapter. Animal distribution among groups designed to equalise initial mean body weight.
- Rationale for selecting satellite groups: Satellite groups for haematology and bone marrow analysis (27 animals per sex in control, mid and high dose groups, 9 animals per sex for low dose group.) One group each for time point sacrifices of 3, 6 and 12 months.
Positive control:
No

Examinations

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

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: monthly (fortnightly during last 6 months) and at end of study

BODY WEIGHT: Yes
- Time schedule for examinations: start, monthly (fortnightly during last 6 months) 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 and from satellite groups at 3, 6 and 12 months (not 31.2ppm) - 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, reticulocyte, haemoglobin, leucocyte differential count and nucleated erythrocyte count. Morphological analysis of erythrocytes, leucocytes and platelets.

CLINICAL CHEMISTRY: No

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No
Sacrifice and pathology:
HISTOPATHOLOGY: Yes. Complete histopathology on all core animals (not satellites). 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, ovary, 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.
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:
no effects observed
Description (incidence and severity):
No clinical signs were attributed to 2-butoxyethanol exposure.
Mortality:
no mortality observed
Description (incidence):
Survival of exposed male and female rats was similar to the chamber control groups, the mean numbers of survivors at the end of the experiment in each of the 0, 31.2, 62.5 and 125 ppm groups were 19, 11, 21 and 24 males and 29, 27, 23 and 21 females, respectively.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Body weights of male and female rats in the 31.2 and 62.5 ppm groups and male rats of the 125 ppm group were generally similar to the controls throughout the experiment.
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
Description (incidence and severity):
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. These changes occurred at 3, 6 and 12 months in 62.5 ppm group females and 125 ppm group males. Some anaemia also occurred at 3 and 6 months in the 31.2 ppm group females and at 12 months in the 62.5 ppm group males. In females this was characterised by t adose related and significant fall in hematocrit, haemoglobin and erythrocyte count and an increase in MCV. The changes at 31ppm were however small (<5%). Increases in circulating reticulocyte and nucleated erythrocyte counts are consistent with an erythropoietic response to the anaemia. In bone marrow there were approximately 15 % - 35 % decreases in the myeloid/erythroid ratio in the 125 ppm rats of both sexes, but particularly in females. Increases in bone marrow cellularity occured at all time points in females at 125ppm along with a 15-35% decrease in myeloid/erythroid ratio. Significant changes were also seen in males at 125ppm and females at 62.5ppm but only at one time point. The severity of the response was dose related.
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
not specified
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
In the nose there were significant increases in the incidences of hyaline degeneration of the olfactory epithelium in all groups of males (chamber control, 13/48; 31.2 ppm, 21/49; 62.5 ppm, 23/49; 125 ppm, 40/50) and in females exposed to the two higher concentrations of EGBE (chamber control, 13/50; 31.2 ppm, 18/48; 62.5 ppm, 28/50; 125 ppm, 40/49). The severity of the lesion was minimal and did not change with increasing exposure concentration. It is the most common age-related change of the nasal passages of rats (St. Clair & Morgan, 1992) and it has been proposed to have an adaptive or protective role (Buckley et al., 1985).

Incidences of Kupffer cell pigmentation of the liver increased significantly in all exposed groups of male rats (chamber control, 23/50; 31.2 ppm, 30/50; 62.5 ppm, 34/50; 125 ppm, 42/50) and in the two higher exposure groups of female rats (chamber control, 15/50; 31.2 ppm, 19/50; 62.5 ppm, 36/50; 125 ppm, 47/50). The severity of the lesion increased in the 125 ppm group of both sexes.

In the spleen, incidences of fibrosis were significantly increased in the two higher exposure groups of male rats (chamber control, 11/50; 31.2 ppm, 14/50; 62.5 ppm, 19/50; 125 ppm, 20/50), but not in females.
Histopathological findings: neoplastic:
effects observed, treatment-related
Description (incidence and severity):
See results in chapter 7.7 of IUCLID dossier

Effect levels

open allclose all
Dose descriptor:
NOAEC
Effect level:
< 31 ppm
Based on:
test mat.
Sex:
female
Basis for effect level:
haematology
histopathology: non-neoplastic
Remarks on result:
other: adverse effects seen at lowest tested dose
Dose descriptor:
NOAEC
Effect level:
< 31 ppm
Based on:
test mat.
Sex:
male
Basis for effect level:
other: Kupffer cell pigmentation.

Target system / organ toxicity

Critical effects observed:
yes
Lowest effective dose / conc.:
31 ppm
System:
hepatobiliary
Organ:
liver
Treatment related:
yes
Dose response relationship:
yes
Relevant for humans:
no

Any other information on results incl. tables

Neoplastic effects are reported in chapter 7.7.

Applicant's summary and conclusion

Conclusions:
No NOAEC was found. The LOAEL of 31 ppm based on haematological effects seen at all doses tested. in females and Kupffer cell pigmentation in both sexes. Females appear to be more senstive to the haematological effects of 2-butoxyethanol than males.
Executive summary:

In a two year carcinogenicity study, rats were exposed by inhalation to butoxyethanol at doses up to 125ppm for a period of 104 weeks. The study approximated to guideline but parameters known to be insensitive to this substance were not examined. The predominant non-neoplastic effects of note were adverse changes to the haematology, particularly haematocrit, hemoglobin, erythrocytes reductions. Females were particularly sensitive showing dose related and statistically significant reductions in these parameters at 31ppm, although it should be noted that the reduction in these parameters was only around 5%. These effects were only consistently seen at all interim sacrifice points at 125ppm in males. However no NOAEL could be established due to the presence in both sexes of increased Kupffer cell pigmentation at 31ppm.

Synopsis:

NOAEL (rats, 104 weeks, inhalation): <31ppm