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Description of key information

In repeated dose studies, the principle effects of xylenes were adaptive changes in the liver, organ weight and body weight changes. Inhalation studies in rodents have demonstrated a potential to cause ototoxicity.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Referenceopen allclose all

Endpoint:
sub-chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: GLP status not known, near guideline study, published in peer reviewed literature, limitations in design and/or reporting but otherwise adequate for assessment.
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)
Qualifier:
equivalent or similar to guideline
Guideline:
EPA OPP 82-1 (90-Day Oral Toxicity)
Principles of method if other than guideline:
Observations included assessment of clinical chemistry, haematology, organ weights and microscopic examination of liver and kidney.
GLP compliance:
not specified
Limit test:
no
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Breeding Laboratories (Wilmington, MA) or Simonsen Laboratories (Gilroy, CA, USA)
- Age at study initiation: 6-8 weeks
- Diet: Commercial rodent diet (Ralston Purina Company, St. Louis, MO, USA) ad libitum
- Water: ad libitum
- Housing: 5 per cage
- Acclimation period: quarantined for 1 week

ENVIRONMENTAL CONDITIONS
- Temperature: 21-24°C
- Humidity: 30-70%
- Photoperiod: 12hrs dark / 12hrs light

IN-LIFE DATES: Not reported
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS: Prepared freshly each day in corn oil and dosed at a constant volume of 5 mL/kg bw
Analytical verification of doses or concentrations:
not specified
Duration of treatment / exposure:
90 days
Frequency of treatment:
Daily: once/day
Remarks:
Doses / Concentrations:
0, 150, 750 or 1500 mg/kg/day
Basis:

No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes (normal behavioural response and mortality)
- Time schedule: daily

BODY WEIGHT: Yes
- Time schedule for examinations: weekly

FOOD CONSUMPTION AND COMPOUND INTAKE: No

WATER CONSUMPTION AND COMPOUND INTAKE: No
- Time schedule for examinations:
OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: day of sacrifice
- Method of blood collection: cardiac puncture
- Animals fasted: No data
- How many animals: 10/sex/group
- Parameters examined: RBC, WBC, Hgb, Hct, MCV, MCH, MCHC, platelets, differential count of PMN, Lymph, mono, eosino

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: day of sacrifice
- Animals fasted: No data
- How many animals: 10/sex/group
- Parameters examined: BUN, creatinine, AST, ALT, ALP (females only), LDH, Calcium, Total protein, Albumin, Sodium, Potassium

URINALYSIS: Yes
- Time schedule for collection of urine: once during final week
- Metabolism cages used for collection of urine: Yes
- Animals fasted: No data
- Parameters examined: pH, protein, glucose, bilirubin, urobilinogen, occult blood

NEUROBEHAVIOURAL EXAMINATION: No

Sacrifice and pathology:
GROSS PATHOLOGY: Yes (major organs preserved for future possible examination)

ORGAN WEIGHTS: brain, liver, spleen, lungs, thymus, kidneys, heart and gonads of all animals

HISTOPATHOLOGY: Yes (liver and kidney)
Statistics:
For each treatment group, the mean weekly body weights, clinical chemistry, haematology and organ weights were compared by analysis of variance. Comparisons of each treatment group with their respective vehicle control groups were made by Student's t-test. Results of the urinalysis and gross necropsy findings were not statistically analyzed.
Details on results:
CLINICAL SIGNS AND MORTALITY: No mortalities. Only clinical sign was increase in aggressiveness during handling at 1500 mg/kg, especially males.

BODY WEIGHT AND WEIGHT GAIN: Reduced body weight gain at 1500 mg/kg in males. Overall weight gain was 104, 100 and 86% of control for 150, 750 and 1500 mg/kg respectively.

HAEMATOLOGY: No toxicologically significant effects. Statistically significant changes were present which indicated the presence of mild polycythaemia and leukocytosis in some of the dose groups. These changes were considered not to be toxicologically significant since the clinical health of the rats did not appear to be affected.

CLINICAL CHEMISTRY: Several instances of statistically significant variations were detected between control and treated groups but none are considered to be toxicologically significant. The modest increases in AST values in the 1500 mg/kg females and in ALT values in 1500 mg/kg males and 750 and 1500 mg/kg females, were not of sufficient magnitude to indicate liver damage but are consistent with hepatomegally.

ORGAN WEIGHTS:  Relative liver weights were statistically significantly increased in males (5.7, 17.1 and 37.1 % difference from controls for 150, 750 and 1500 mg/kg/day respectively) and females (13.9 and 27.8% difference from controls for 750 and 1500 mg/kg/day respectively). Relative kidney weights were statistically significantly increased in males (17.5 and 27.0% difference from controls for 750 and 1500 mg/kg/day respectively) and in females (16.4% difference from controls for 1500 mg/kg/day). Relative spleen and heart weights increased slightly in females at 1500 mg/kg (12.2 and 11.5% difference from controls respectively).

HISTOPATHOLOGY: NON-NEOPLASTIC: In male kidneys - dose-related increase (all dose levels 0/9, 3/9, 5/10, 8/10, in 0, 150, 750 and 1500 mg/kg bw/day groups respectively) in the incidence of slight to mild hyaline droplet formation in tubules which the authors concluded to be related to male rat-specific alpha-2u-globulin accumulation and not to be relevant to humans. In females - minimal nephropathy (incidence 1/10, 3/10, 6/10, 7/10 in 0, 150, 750 and 1500 mg/kg bw/day groups respectively). This was described as scattered tubular dilation and atrophy, with occasional tubular regeneration. The kidney changes in female rats are typical of chronic progressive nephropathy seen in ageing rats. There were no treatment-related liver changes.
Dose descriptor:
LOAEL
Effect level:
150 mg/kg bw/day (nominal)
Sex:
male
Basis for effect level:
other: increased relative liver weight, no adverse liver histopathology, no alteration in clinical chemistry or haematological parameters. 750 mg/kg bw/day is a no effect level for decreases in body weight gain.
Dose descriptor:
NOAEL
Effect level:
150 mg/kg bw/day (nominal)
Sex:
female
Basis for effect level:
other: increased liver and kidney weights and increased incidence of minimal kidney nephropathy at 750 and 1500 mg/kg bw/day
Critical effects observed:
not specified
Conclusions:
Treatment-related alterations following subchronic oral treatment with mixed xylenes were mild and limited to decreased body weight gain and increased relative organ weights (affecting primarily the liver and kidney; no histopathological involvement).
Executive summary:

The repeated dose toxicity of technical grade mixed xylenes was investigated in male and female Sprague-Dawley rats following oral gavage administration for 90 days at 0, 150, 750 or 1500 mg/kg/day. Treatment-related alterations were mild and limited primarily to increased relative liver weight (LOAEL 150 mg/kg bw/d for males; NOAEL 150 mg/kg bw/d for females; no histopathological alterations), relative kidney weight (higher LOAELs and NOAELs) and a reduction in body weight gain (males, NOAEL 750 mg/kg/d).

Endpoint:
repeated dose toxicity: oral, other
Remarks:
combined repeated dose and carcinogenicity
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: GLP status not known, near guideline study, published in peer reviewed literature, limitations in design and/or reporting but otherwise adequate for assessment.
Qualifier:
equivalent or similar to guideline
Guideline:
other: EU Method B.32 (Carcinogenicity Test)
Principles of method if other than guideline:
Mixed xylene was administered by oral gavage to groups of 50 male and 50 female F344/N rats at doses of 0, 250 or 500 mg/kg bw/day for 103 weeks. Animals were observed for survival, clinical signs and body weight gain and subject to a full necropsy with tissue histopathology at termination. However, only two dose levels were tested and ophthalmoscopy, food consumption, haematology, blood clinical chemistry and urinalysis were not assessed.
GLP compliance:
not specified
Limit test:
no
Species:
rat
Strain:
other: F344/N
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Breeding Laboratories, Kingston, NY, USA
- Age at study initiation: 7 weeks
- Housing: 5 per sex /cage in Polycarbonate cages
- Diet: NIH 07 Rat and Mouse Ration (Zeigler Bros., Inc., Gardners, PA, USA); available ad libitum
- Water: ad libitum
- Acclimation period: 19 days

ENVIRONMENTAL CONDITIONS
- Temperature: 23° ± 1°C
- Humidity: 40 - 60%
- Air changes: 15 air changes/hr
- Photoperiod: 12 hr/d light; 12 hr/d dark

IN-LIFE DATES: From: 30 June 1980 To: 2 July 1982
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on oral exposure:
Oral (gavage): 0, 250 or 500 mg/kg xylenes (mixed) in corn oil; 4 mL/kg

Preparation: Weighed portions of xylenes (mixed) were placed in a graduated cylinder and mixed with corn oil to achieve the proper volume. The mixtures were shaken vigorously for 10 seconds.

Maximum Storage Time: 2 wks

Storage Conditions: Approximately 24ºC, 46% humidity under fluorescent light.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The concentrations of xylenes in corn oil was analysed by gas chromatography with flame ionization detection following extraction with methanol.
During the 2-year studies, the dose preparations were analyzed once every 2 months, with concentrations varying from 94.6% to 106.9% (within 10% target concentrations).
Duration of treatment / exposure:
5 days per week for 103 weeks.
Frequency of treatment:
Once daily (5 days / week).

Remarks:
Doses / Concentrations:
0, 250 or 500 mg/kg
Basis:
other: nominal concentration
No. of animals per sex per dose:
50 male / 50 female per group

Control animals:
yes, concurrent vehicle
Details on study design:
Dose selection rationale: Based on weight gain depression at 1,000 mg/kg in both sexes in the 14-day studies and in males in the 13-week studies and on the clinical signs in the 14-day studies, doses selected for rats for the 2-year studies were 0, 250, and 500 mg/kg xylenes (mixed) in corn oil by gavage, administered 5 days per week.
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- All animals were observed twice daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Clinical signs were recorded once per day for 16 months and then once per month.

BODY WEIGHT: Yes
- Body weights were recorded weekly for 12 weeks and monthly thereafter

OPHTHALMOSCOPIC EXAMINATION: No data

HAEMATOLOGY: No data

CLINICAL CHEMISTRY: No data

URINALYSIS: No data

NEUROBEHAVIOURAL EXAMINATION: No data

Data were recorded in the NTP Carcinogenesis Bioassay Data System. The data elements included descriptive information on the chemicals, animals, experimental design, survival, body weight, and individual pathologic results.
Sacrifice and pathology:
Necropsy and histopathological examination performed on all animals, where possible. During necropsy, all organs and tissues were examined for grossly visible lesions. Tissues were preserved in 10% neutral buffered formalin, embedded in paraffin, sectioned, and stained with haematoxylin and eosin. The following tissues were examined: gross lesions and tissue masses, mandibular lymph nodes, salivary gland, femur, including marrow, thyroid gland, parathyroids, small intestine, colon, liver, prostate / testis or ovaries / uterus, heart, oesophagus, stomach, brain, thymus, trachea, pancreas, spleen, skin, lungs and mainstem bronchi, kidneys, adrenal glands, urinary bladder, pituitary gland, eyes (if grossly abnormal), and mammary gland.
Statistics:
Survival Analyses: Kaplan and Meier (1958); Cox (1972) and Tarone (1975). All reported P values for the survival analysis are two-sided. Calculation of Incidence for neoplastic and non-neoplastic lesions. Analysis of Tumour Incidence: Mantel and Haenszel (1959). Continuity-corrected tests were used in the analysis of tumour incidence, and reported P values are one-sided. Life Table Analyses-- Mantel-Haenszel (1959) method used to obtain an overall P value. Life table method of Cox (1972) and of Tarone (1975). The underlying variable considered by this analysis is time to death due to tumour. Incidental Tumour Analyses-- (Haseman, 1984) Unadjusted Analyses--Primarily, survival-adjusted methods are used to evaluate tumour incidence. The Fisher exact test for pairwise comparisons and the Cochran-Armitage linear trend test (Armitage, 1971; Gart et al., 1979).
Clinical signs:
effects observed, treatment-related
Mortality:
mortality observed, treatment-related
Body weight and weight changes:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
no effects observed
Details on results:
Mortality - Although the mortality was dose related in male rats (final survival: vehicle control 36/50, low dose 26/50, high dose 20/50), many of the early deaths in the dosed males were gavage related. Survival of the high dose males was significantly lower than that of the vehicle control after week 103.

Bodyweight - Bodyweights of high dose male rats were 5%-8% lower than those of the vehicle controls after week 59.

Tumour findings - There were no significant changes in the incidences of neoplastic or non-neoplastic lesions which were considered to be related to the administration of xylenes (mixed).

Testis findings - Although the overall incidences of interstitial cell tumours were comparable in male rat groups (vehicle control, 43/50; low dose,38/50; high dose, 41/49), survival-adjusted analyses indicated an increased incidence in the high dose group relative to vehicle controls. This apparent effect was due primarily to animals dying between weeks 62 and 92, for which the incidence of interstitial cell tumours was 13/13 for the high dose group compared with 4/9 for vehicle controls. Tumour incidences were comparable during the other time intervals. It is doubtful that this marginal effect is compound related.

Haematopoietic System and Pituitary Gland - Dose-related decreases in the incidences of mononuclear cell leukaemia (vehicle control,22/50; low dose, 18/50; high dose, 11/50) and pituitary gland adenoma or carcinoma (combined) (vehicle control, 24/49; low dose, 22/50; high dose, 12/45) were observed in male rats. However, these differences were due primarily to decreased survival of the high dose group relative to that of the vehicle controls.
Dose descriptor:
NOAEL
Effect level:
250 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: decreased body weights in males from week 59 at 500 mg/kg bw/day and no evidence of systemic toxicity of xylenes (mixed) for male or female F344/N rats given 250 or 500 mg/kg
Critical effects observed:
not specified
Conclusions:
There was no evidence of treatment-related systemic toxicity or carcinogenicity following gavage administration of mixed xylenes to male and female F344/N rats at doses of 0, 250 or 500 mg/kg body weight/day for up to 103 weeks.
Executive summary:

The toxicity of mixed xylene was investigated in male and female F344/N rats following oral (gavage) administration at doses of 0, 250 or 500 mg/kg bw/day for 103 weeks. Animals were observed for survival, clinical signs and body weight gain and subject to a full necropsy with tissue histopathology at sacrifice. There was no evidence of treatment-related systemic toxicity in either sex under these conditions.

Endpoint:
sub-chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: GLP status not known, near guideline study, published in peer reviewed literature, limitations in design and/or reporting but otherwise adequate for assessment.
Qualifier:
no guideline followed
Principles of method if other than guideline:
The purpose of the study was to evaluate the subchronic toxicity of xylenes when administered to mice daily by oral gavage for 13 weeks.
GLP compliance:
no
Limit test:
no
Species:
mouse
Strain:
B6C3F1
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Breeding Laboratories, Kingston, NY, USA
- Age at study initiation: 7 weeks
- Housing: 5 per sex /cage in polycarbonate cages
- Diet: Purina Lab Chow available ad libitum
- Water: ad libitum
- Acclimation period: 15 days

ENVIRONMENTAL CONDITIONS
- Temperature: 22° ± 1°C
- Humidity: 40 - 60%
- Air changes: 15 air changes/hr
- Photoperiod: 12 hr/d; light 12 hr/d

IN-LIFE DATES: From: 6 August 1979 To: 6 November 1979
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on oral exposure:
Oral (gavage): 0, 125, 250, 500, 1000 or 2000 mg/kg xylenes (mixed) in corn oil; dose volume 8 mL/kg.

Preparation: Weighed portions of xylenes (mixed) were placed in a graduated cylinder and mixed with corn oil to achieve the proper volume. The mixtures were shaken vigorously for 10 seconds.

Maximum Storage Time: 2 wks

Storage Conditions: Approximately 23ºC, 46% humidity under fluorescent light.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The dose preparations were analysed once, with concentrations varying from 84.8% to 107.5% of target concentrations.
Duration of treatment / exposure:
5 days per week for 13 weeks
Frequency of treatment:
Once daily (5 days / week)
Remarks:
Doses / Concentrations:
0, 125, 250, 500, 1000 or 2000 mg/kg
Basis:
other: nominal in corn oil
No. of animals per sex per dose:
10 male / 10 female per group
Control animals:
yes, concurrent no treatment
Details on study design:
A thirteen-week study was conducted to evaluate the cumulative toxic effects of repeated administration of mixed xylenes and to determine the doses to be used in the 2-year study.
Observations and examinations performed and frequency:
Animals were checked twice daily; moribund animals were killed, Individual animal weights were recorded weekly.
Sacrifice and pathology:
Necropsy was performed on all animals.
The following tissues examined histopathologically for vehicle control and high dose groups: gross lesions and tissue masses, mandibular lymph node, salivary gland, sternebrae, femur, or vertebrae including marrow, thyroid gland, parathyroids, small intestine, colon, heart, oesophagus, stomach, brain, thymus, trachea, pancreas, spleen, kidneys, liver, gall bladder, prostate/testis or ovaries/uterus, lungs and mainstem bronchi, adrenal glands, urinary bladder, pituitary gland, spinal cord (if neurologic signs present), eyes (if grossly abnormal), and mammary gland.
Statistics:
none
Details on results:
Two female mice that received 2,000 mg/kg died before the end of the study (possibly mis-dosed).

Weakness, lethargy, short and shallow breathing, unsteadiness, tremors, and paresis were observed in the 2,000 mg/kg group 5-10 minutes after dosing and lasted 15-60 minutes.

Mean body weight gain of mice that received 2,000 mg/kg was 7% lower than that of the vehicle controls for males and 17% lower for females.

No compound-related gross or microscopic pathologic lesions were observed.
Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day (nominal)
Sex:
male/female
Basis for effect level:
other: decreased body weight, no gross or microscopic tissue lesions present
Critical effects observed:
not specified
Conclusions:
Treatment-related alterations in this mouse subchronic oral study were mild and limited to transient, reversible CNS depression and decreased body weight gain.
Executive summary:

In a 13-week study, groups of B6C3F1 of each sex received mixed xylenes by oral gavage at 0, 125, 250, 500, 1,000, or 2,000 mg/kg bw/day. Clinical signs (indicative of transient, reversible CNS depression) were present in high dose animals immediately after dosing and lasted for 15-60 minutes. Treatment-related findings were mild and limited to a 7-17% reduction in overall body weight gain with a NOAEL of 1000 mg/kg bw/day. No treatment-related gross or microscopic pathological lesions were observed.


Endpoint:
sub-chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: GLP status not known, near guideline study, published in peer reviewed literature, limitations in design and/or reporting but otherwise adequate for assessment.
Qualifier:
no guideline followed
Principles of method if other than guideline:
The purpose of the study was to evaluate the subchronic toxicity of xylenes when administered to rats daily by oral gavage for 13 weeks.
GLP compliance:
no
Limit test:
no
Species:
rat
Strain:
other: F344/N
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Breeding Laboratories, Kingston, NY, USA
- Age at study initiation: 6 weeks
- Housing: 5 per sex /cage in polycarbonate cages
- Diet: Purina Lab Chow available ad libitum
- Water: ad libitum
- Acclimation period: 15 days

ENVIRONMENTAL CONDITIONS
- Temperature: 22° ± 1°C
- Humidity: 40 - 60%
- Air changes: 15 air changes/hr
- Photoperiod: 12 hr/d; light 12 hr/d

IN-LIFE DATES: From: 6 August 1979 To: 6 November 1979
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on oral exposure:
Oral (gavage): 0, 62.5, 125, 250, 500 or 1000 mg/kg xylenes (mixed) in corn oil; dose volume 4 mL/kg

Preparation: Weighed portions of xylenes (mixed) were placed in a graduated cylinder and mixed with corn oil to achieve the proper volume. The mixtures were shaken vigorously for 10 seconds.

Maximum Storage Time: 2 wks

Storage Conditions: Approximately 23ºC, 46% humidity under fluorescent light
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The dose preparations were analysed once, with concentrations varying from 97.2% to 106% of target concentrations.
Duration of treatment / exposure:
5 days per week for 13 weeks
Frequency of treatment:
Once daily (5 days / week)
Remarks:
Doses / Concentrations:
0, 62.5, 125, 250, 500 or 1000 mg/kg
Basis:
other: nominal in corn oil
No. of animals per sex per dose:
10 male / 10 female per group
Control animals:
yes, concurrent vehicle
Details on study design:
A thirteen-week study was conducted to evaluate the cumulative toxic effects of repeated administration of mixed xylenes and to determine the dose levels to be used in the 2-year study.
Observations and examinations performed and frequency:
Animals were checked twice daily; moribund animals were killed. Individual animal weights were recorded weekly.
Sacrifice and pathology:
Necropsy was performed on all animals. The following tissues examined histopathologically for vehicle control and high dose groups: gross lesions and tissue masses, mandibular lymph node, salivary gland, sternebrae, femur, or vertebrae including marrow, thyroid gland, parathyroids, small intestine, colon, heart, oesophagus, stomach, brain, thymus, trachea, pancreas, spleen, kidneys, liver, prostate/testis or ovaries/uterus, lungs and mainstem bronchi, adrenal glands, urinary bladder, pituitary gland, spinal cord (if neurologic signs present), eyes (if grossly abnormal), and mammary gland
Statistics:
None
Details on results:
All the rats survived to the end of the study.
The change in mean body weight of male and female rats that received 1,000 mg/kg was 15% and 8% lower than that of the vehicle controls after 13 weeks of exposure.
No signs of toxicity were observed, and no compound-related gross or microscopic pathologic lesions were present.
Dose descriptor:
NOAEL
Effect level:
500 mg/kg bw/day (nominal)
Sex:
male/female
Basis for effect level:
other: decreased body weight, no gross or microscopic tissue lesions present at highest dose tested
Critical effects observed:
not specified
Conclusions:
Treatment-related alterations in this rat subchronic oral study were mild and limited to decreased body weight gain.
Executive summary:

The oral toxicity of mixed xylenes was investigated in male and female Fisher F344/N rats following repeated dosing by gavage for 90 days with 0, 62.5, 125, 250, 500 or 1000 mg/kg/day. Treatment-related findings were mild and limited to an 8-15% reduction in overall body weight gain with a NOAEL of 500 mg/kg bw /day. No treatment-related gross or microscopic lesions were observed.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEL
250 mg/kg bw/day
Study duration:
chronic
Species:
rat
Quality of whole database:
Results are available from chronic and sub-chronic studies that have investigated the repeated dose toxicity of mixed xylene in rodents.

Repeated dose toxicity: inhalation - systemic effects

Link to relevant study records
Reference
Endpoint:
sub-chronic toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: GLP status not known, non-guideline study, published in peer reviewed literature, limitations in design and/or reporting but otherwise adequate for assessment.
Qualifier:
no guideline followed
GLP compliance:
not specified
Limit test:
no
Species:
rat
Strain:
Wistar
Sex:
male
Details on test animals or test system and environmental conditions:
No details reported
Route of administration:
inhalation: vapour
Type of inhalation exposure:
whole body
Vehicle:
other: air
Details on inhalation exposure:
no details
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Gas chromatography.
Duration of treatment / exposure:
13 weeks
Frequency of treatment:
6 hours/day, 5 days/week
Remarks:
Doses / Concentrations:
0, 180, 460 or 810 ppm
Basis:
analytical conc.
Remarks:
Doses / Concentrations:
0, 781, 1996 and 3515 mg/m3
Basis:
analytical conc.
No. of animals per sex per dose:
25 males
Control animals:
yes
Details on study design:
Groups of 25 male rats were assigned randomly to each of three graded levels of mixed xylenes or to the solvent- free air-control. Three rats from each group were killed for histological examination after 3 and 7 week intervals. Ten rats per group that survived 13 weeks were used for a challenge exposure to determine whether they had become more or less sensitive as a result of the repeated inhalation of vapour. The remaining rats were terminated after 13 weeks (65 days of exposure).
Positive control:
No
Observations and examinations performed and frequency:
Measurements included body weight change and urine and blood analysis. Evaluations on blood included: haematocrit, total erythrocyte count, reticulocyte count, total and differential leucocyte counts, serum alkaline phosphatase (SAP), serum glutamic pyruvic transaminase (SGPT), serum glutamic oxalacetic transaminase (SGOT) and blood urea nitrogen (BUN).
Sacrifice and pathology:
At termination, the following tissues were taken for microscopic examination: adrenal, brain, pituitary, trachea, thyroid, parathyroid, lung, heart, liver, kidney, spleen, stomach, duodenum, pancreas, ileum, jejunum, colon, skeletal muscle, sciatic nerve, and bone marrow impression smear. All tissues were examined at each time point for the high dose and control groups, but for the low and intermediate dose only lung, liver, kidney, heart, spleen, adrenal, thyroid, parathyroid, trachea, oesophagus, and bone marrow impression smears were examined.
Other examinations:
Groups of ten rats per group that survived exposure over 13 weeks, were subjected to a 4-hr challenge period during which time they inhaled 29.0 mg/L (6700 ppm) of mixed xylenes. Their response was compared to the similarly handled control group and to 20 naive rats that were randomized from the same production lot as those under test but not subjected to daily handling.
Statistics:
No details
Details on results:
Two rats (one control and one low dose) died of extraneous pneumonic infections. All other rats gained weight normally. Minor differences in blood clinical chemistry and haematology were considered not to be adverse effects. Urinalyses performed before rats were sacrificed at 3, 7, and 13 wk revealed no abnormalities. No lesions ascribed to inhalation of mixed xylenes were found at any time point.

No statistically significant differences in median time to death were apparent following the challenge at the end of the study. No protective adaptive mechanism could be claimed as a result of exposure to concentrations as high as 3.5 mg/L (810 ppm) for 6 hr/day x 65 days (5 days /week for 13 weeks.

Dose descriptor:
NOAEC
Effect level:
810 ppm
Sex:
male
Basis for effect level:
other: 3515 mg/m3. No effects at the highest dose tested.
Critical effects observed:
not specified
Conclusions:
The NOAEC of mixed xylenes for male rats exposed 6h/day for 5 days in each of 13 weeks was 3515 mg/m3 (3.5 mg/L).
Executive summary:

Male rats were exposed 6h/day for 5 days in each of 13 weeks to 0, 0.77, 2.0 or 3.5 mg/L (0, 180, 460 or 810 ppm) mixed xylenes. The NOAEC was 3515 mg/m3 (3.5 mg/L).

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEC
3 515 mg/m³
Study duration:
subchronic
Species:
rat
Quality of whole database:
A NOAEC of 3515 mg/m3 was reported by Carpenter et al. (1975) for generalised systemic effects in male rats and male dogs for xylene. Other studies have shown that some xylene isomers adversely affect hearing in the rat, with a sub-chronic NOAEC of 1950 mg/m3 reported for p-xylene; the NOAEC for ototoxicity of m-xylene and o-xylene was greater than 7810 mg/m3 (Gagnaire et al., 2001). The ototoxicity of mixed xylenes appears to be dependent upon composition (Gagnaire et al., 2007), with a sub-chronic LOAEC of 1080 mg/m3 reported for one sample while another had a NOAEC of 2170 mg/m3.

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

The multi-constituent substances covered by this registration comprise individual xylene isomers (m-xylene, o-xylene, p-xylene) and ethylbenzene (<10%). The following information is available to characterise their repeated dose toxicity.

 

Repeated dose toxicity: oral

 

Data are available for laboratory animals exposed to high doses of mixed xylene, adverse effects have been observed in the kidney and liver (IARC, 1989).

 

A near-guideline (equivalent or similar to OECD 408) subchronic oral gavage study with mixed xylene (comprising 18% o-xylene, 62% m- and p-xylene, 20% ethyl benzene) was conducted by Condie et al. (1988). It includes the range of toxicological endpoints routinely investigated in regulatory subchronic studies and is a key study for identifying the effects of mixed xylene. In the study, groups of 10 male and 10 female rats were given 0, 150, 750, or 1500 mg/kg bw/day of mixed xylene in corn oil for 90 consecutive days. A decrease in body weight was observed in males only at 1500 mg/kg bw/day. Although increased relative liver weights were seen at all dose levels in males and in females at 750 and 1500 mg/kg bw/day there were no adverse histopathology findings in the liver. An increased relative kidney weight was observed in high dose males and females and in intermediate dose males. In males there was a dose-related increase in the incidence of slight to mild hyaline droplet formation in tubules at all dose levels. This finding is indicative of alpha-2u-globulin which is considered to be male rat-specific and is not relevant for humans. In females the incidence of minimal nephropathy in females was statistically significantly increased in the 750 and 1500 mg/kg bw/day groups. This finding described as scattered tubular dilation and atrophy, with occasional regeneration and is the chronic progressive nephropathy typically seen in ageing rats.

 

No NOAEL was established in this study for males based on liver weight increases. However increases in liver weight with no adverse histopathological findings are considered to be an adaptive response to administration of mixed xylene rather than an adverse toxicological effect. A dose of 750 mg/kg bw/day is the NOAEL based on effects on male body weight. In females a NOAEL of 150 mg/kg bw/day is based on liver weight increases and the kidney effects observed at dose levels of 750 mg/kg bw/day and higher.

 

Treatment levels between 150 and 750 mg/kg bw/day are covered in a carcinogenicity study in rats (NTP, 1986). The test sample comprised 60% m-xylene, 14% p-xylene, 9% o-xylene, and 17% ethyl benzene. Although this study did not include all of the end points included in chronic studies to current guidelines, the key parameters affected in the sub chronic study, i.e. body weights and detailed pathology and histopathology are included. Rats were dosed with mixed xylene at concentrations of 0, 250 or 500 mg/kg bw/day 5 days per week for 103 weeks. The main finding was a decrease in body weights in males receiving 500 mg/kg bw/day in the second year of the study. There was no other evidence of systemic toxicity including no treatment-related pathology findings. A dose of 250 mg/kg/day was a NOAEL for both sexes and this is considered to be the key study for determining the NOAEL for repeated dose exposure to mixed xylenes via the oral route.

 

A supporting subchronic oral gavage study was conducted by NTP (1986) using groups of 10 male and 10 female rats treated via gavage 5 days/week for 13 weeks with 0, 62.5, 125, 250, 500 or 1000 mg/kg/day. In the same study, groups of 10 male and 10 female mice were similarly dosed with 0, 125, 250, 500, 1000 or 2000 mg/kg/day of mixed xylene in corn oil. The composition of the sample was as described above.

 

Limited toxicological endpoints were evaluated.

 

Treatment-related findings in rats were limited to a reduction in overall body weight gain (15% for males and 8% for females) with a NOAEL of 500 mg/kg/day. High dose mice exhibited transient CNS effects 5-10 minutes after dosing that lasted 15-60 minutes. Other treatment-related findings were limited to a reduction in overall body weight gain (7% for males and 17% for females) with a NOAEL of 1000 mg/kg/day. Neither blood clinical chemistry nor organ weight data were collected for either species. No treatment-related macroscopic or microscopic lesions were observed in the tissues examined including the liver and kidney.

 

 

o-xylene

A guideline OECD 408 90-day oral rat study with o-xylene was conducted (Britton LD, 2020).

 

The test material, o-xylene (CAS95-47 -6), was administered to male and female rats by oral gavage in order to evaluate its sub chronic toxicity. Dose levels were 0, 100, 300, and 1000 mg/kg/day (Groups 1 through 4, respectively), with 10 animals/sex/group dosed daily for at least 90 days.The following were investigated: clinical observations, ophthalmoscopy, body weight, food intake, functional observation battery, haematology, blood chemistry, thyroid hormone analysis, organ weights, macroscopic and microscopic pathology.

 

There were no deaths during the study. Males and females given 1000 mg/kg/day showed decreased activity and unsteady gait from Days 19/18 of dosing onwards (persisting for up to 4 hours after dosing), respectively. The incidence of these clinical signs decreased with time and by Day 72 of dosing, only one male was showing decreased activity and abnormal gait. Over Days 84 to 92 of dosing at most eight out of ten females showed one or both of these signs after dosing. No effects on sensory reaction or grip strength were elicited by o-xylene. In open arena testing, males given 1000 mg/kg/day exhibited effects on arousal, unusual posture and abnormal gait. Twitches or extended rearing were also observed in a small number of individual males given 1000 mg/kg/day. Increased episodes of arousal in the arena and abnormal gait were observed between Days 21 to 70 and Days 21 to 56 of dosing, respectively. Excessive salivation in the arena was also noted between Days 14 and 70 of dosing for males given 300 or 1000 mg/kg/day, with incidence increasing with time. Females given 1000 mg/kg/day also exhibited effects on arousal in the arena, abnormal gait and salivation. Although incidences increased with time similarly to the males, these observations were not seen in either sex after Day 77 of dosing. Movement counts and distance travelled were reduced in males given 100, 300 or 1000 mg/kg/day and this was occasionally accompanied by an increased time at rest. Males given 300 or 1000 mg/kg/day gained less weight over the study than Controls with terminal body weights that were 6 % and 11 % respectively, lower than those of the Controls. There was no effect on body weight for females and no effect on food intake in either sex. There were slight increases in Thyroxine (T4) and no effect on thyroid stimulating hormone (TSH) or triiodothyronine (T3) levels at the end of the treatment period and no adverse effects on haematology, coagulation or blood chemistry parameters. Both sexes given 300 or 1000 mg/kg/day and females given 100 mg/kg/day had statistically significantly higher liver and kidney weights when compared with Controls. In addition, thyroid weights for females given 1000 mg/kg/day were also higher than Controls.

 

Histopathology findings considered to be related to o-xylene were seen in the liver (hepatocellular hypertrophy) and thyroid (follicular cell hypertrophy) in both sexes and, in the kidneys, eosinophilic inclusions (hyaline droplets) in the tubular epithelium (males only). Following α2μ-globulin staining of the kidneys, although positive staining was observed in 3/10 high dose males, minimal staining was also observed in 2/10 Controls. The precise content of the hyaline droplets observed at an increased incidence/severity of within the cortical tubules of male kidneys therefore remained unestablished. Given the physiological differences between the male rat kidney and human, these findings are considered unlikely to represent a hazard to man.

 

In conclusion, when administered by gavage to the Han Wistar rat, once daily for at least 90 days, o-xylene was generally well tolerated in-life at 100 and 300 mg/kg/day with, at the latter dose, only a mild retardation of body weight gain in males and clinical signs being limited to some transient excessive salivation after dosing. However, more profound (decreased activity, unsteady gait, effects on arousal, unusual posture), but transient, clinical signs were apparent at 1000 mg/kg/day and a more marked retardation of body weight gains in males. Target organ pathology was restricted to rat specific adaptive responses consisting of hepatic centrilobular hypertrophy and secondary thyroid follicular hypertrophy consistent with enzyme induction and adverse renal tubular changes in males only likely reflecting proteinuria and inclusion of non α2μ-globulin proteins. Therefore, the No Observed Adverse Effect Level (NOAEL) was defined as 300 mg/kg/day o-xylene in the female rat because of the severity of clinical signs at 1000 mg/kg/day and as 100 mg/kg/day in the male rat, because of the incidence and severity of the renal changes. However, given the physiological differences between the male rat kidney and human, the observed tubular changes are unlikely to represent a hazard to man. Therefore, if the findings in the kidney are disregarded as being human relevant, the NOAEL for the male would be 300 mg/kg/day.

 

m-xylene

A GLP 90-day oral rat study with m-xylene was conducted (Wolfe GW, 1988).

 

The test material, m-xylene (CAS 108-38-3), was administered to male and female rats by oral gavage in order to evaluate its sub chronic toxicity. Dose levels were 0, 100, 200, and 800 mg/kg/day (Groups 1 through 4, respectively), with 20 animals/sex/group dosed daily for at least 90 days. Criteria evaluated included mortality, clinical signs, body weights, food consumption, ophthalmologic examinations, haematology, clinical chemistry, organ weights, organ-to-body weight ratios, gross pathology, and histopathology.

 

Mortality was significantly increased in male Group 3 and female Groups 3 and 4; however, based on histopathology findings, these deaths were most likely the result of vehicle and/or compound aspiration. Salivation was frequently observed in both sexes of Group 4 and only rarely in the other treated groups. Total body weight gain was significantly decreased in male Groups 3 and 4, and female Group 4. Food consumption was significantly decreased from Week 1 to 5 in Group 4 males and from Week 6 to 9 in Groups 3 and 4 males. A few clinical pathology parameters were significantly increased or decreased and with one exception (female Group 3), all were in Group 4. A few significant changes were seen in absolute or relative organ weights, all of which were in male Group 4. The most commonly observed lesions at necropsy were in the lungs (mottled and failure to collapse). These lesions were only seen in the animals found dead. No apparent compound-related effects were present in the ophthalmology or histopathology findings.

 

Based on the results of this study, with the assumption that the decrease in survival was the result of vehicle and/or compound aspiration, the no-observed-adverse-effect level (NOAEL) of m-Xylene was 200 mg/kg/day in the females. However, due to significantly decreased body weight gain in Groups 3 and 4, the NOAEL for the males was 100 mg/kg/day.

 

Data are available for laboratory animals exposed to high doses of mixed xylenes, adverse effects have been observed in the kidney and liver (IARC, 1989).

 

A near-guideline (equivalent or similar to OECD 408) subchronic oral gavage study with mixed xylenes (20% ethylbenzene) was conducted by Condie et al. (1988). It includes the range of toxicological endpoints routinely investigated in regulatory subchronic studies and is a key study for identifying the effects of mixed xylenes. In the study, groups of 10 male and 10 female rats were given 0, 150, 750, or 1500 mg/kg bw/day of mixed xylenes in corn oil for 90 consecutive days. A decrease in body weight was observed in males only at 1500 mg/kg bw/day. Although increased relative liver weights were seen at all dose levels in males and in females at 750 and 1500 mg/kg bw/day there were no adverse histopathology findings in the liver. An increased relative kidney weight was observed in high dose males and females and in intermediate dose males. In males there was a dose-related increase in the incidence of slight to mild hyaline droplet formation in tubules at all dose levels. This finding is indicative of alpha-2u-globulin which is considered to be male rat-specific and is not relevant for humans. In females the incidence of minimal nephropathy in females was statistically significantly increased in the 750 and 1500 mg/kg bw/day groups. This finding described as scattered tubular dilation and atrophy, with occasional regeneration and is the chronic progressive nephropathy typically seen in ageing rats.

 

No NOAEL was established in this study for males based on liver weight increases. However increases in liver weight with no adverse histopathological findings are considered to be an adaptive response to administration of mixed xylenes rather than an adverse toxicological effect. A dose of 750 mg/kg bw/day is the NOAEL based on effects on male body weight. In females a NOAEL of 150 mg/kg bw/day is based on liver weight increases and the kidney effects observed at dose levels of 750 mg/kg bw/day and higher.

 

Treatment levels between 150 and 750 mg/kg bw/day are covered in a carcinogenicity study in rats (NTP, 1986). Although this study did not include all of the end points included in chronic studies to current guidelines, the key parameters affected in the sub chronic study, i.e. body weights and detailed pathology and histopathology are included. Rats were dosed with mixed xylenes at concentrations of 0, 250 or 500 mg/kg bw /day 5 days per week for 103 weeks. The main finding was a decrease in body weights in males receiving 500 mg/kg bw/day in the second year of the study. There was no other evidence of systemic toxicity including no treatment-related pathology findings. A dose of 250 mg/kg/day was a NOAEL for both sexes and this is considered to be the key study for determining the NOAEL for repeated dose exposure to mixed xylenes via the oral route.

 

p-xylene

A GLP 90-day oral rat study with m-xylene was conducted (Wolfe GW, 1988).

 

The test material, p-Xylene (CAS 106-42-3), 99% pure, was administered to male and female Sprague-Dawley rats by oral gavage in order to evaluate its sub-chronic toxicity. Dose levels were 0, 100, 200, and 800 mg/kg/day (Groups 1 through 4, respectively), with 20 animals/sex/group dosed daily for at least 90 days. Criteria evaluated included mortality, clinical signs, body weights. food consumption, ophthalmologic examinations, haematology, clinical chemistry, organ weights, organ-to-body weight ratios, gross pathology, and histopathology.

 

At least one animal in each treated group died before termination of the study and there was a significant increase in mortality in Group 4 males. However, based on histopathology findings, nearly all of the unscheduled deaths were related to aspiration of the vehicle/test material. Salivation was frequently observed and exclusively in both sexes of Group 4. Mean body weights were slightly decreased in Group 4 males and females while total food consumption was slightly increased in these groups. There was one incidental finding in the haematology results. In the clinical chemistries, a few significant differences were present in the Group 4 females at Week 5; however, none were present at Week 13. No apparent compound-related effects were present in the ophthalmology, organ weight, gross pathology, or histopathology data.

 

Based on the results of this study, with the consideration of vehicle/test material aspiration producing the deaths in the treated groups, the no-observed-effect level (NOEL) of p-xylene was 200 mg/kg/day.  

 

Repeated dose toxicity: dermal

 

No studies are available for xylenes (including mixed xylene and the individual xylene isomers).

 

 

Repeated dose toxicity: inhalation

 

The available subchronic inhalation studies are designed primarily to address neurological endpoints (including ototoxicity) in male rats and dogs. These endpoints are discussed in section 5.10.1.1.

 

In a recent study (Gagnaire et al., 2007a) the potential ototoxicity of two samples of mixed xylene was investigated in groups of male rats exposed to 250, 500, 1000 and 2000 ppm for 6 h/day, 6 d/wk over 13 weeks, with a recovery period of 8 weeks. One sample contained 10% ethyl benzene, the other 20% ethyl benzene. There was no adverse effect on body weight at any of the dose levels.

 

In another investigation (Gagnaire et al., 2001), the potential ototoxicity of individual xylene isomers was evaluated using electrophysiological methods in male rats exposed by inhalation to three different concentrations 6 hours/day, 5 days/week for 13 weeks was evaluated. The highest exposure concentration of 1800 ppm had no significant effect on body weight or body weight gain.

 

In an older study by Carpenter (1975), male rats and male dogs were exposed 6h/day for 5 days in each of 13 weeks to 0, 180, 460 or 810 ppm mixed xylene. The highest exposure level was a NOAEC for both species.

 


Justification for selection of repeated dose toxicity via oral route - systemic effects endpoint:
Results from oral repeated dose studies conducted on mixed xylene in the rat indicate a sub-chronic LOAEL of 750 mg/kg bw/d (Condie et al., 1988) and a chronic NOAEL of 250 mg/kg bw/d (NTP, 1986). A sub-chronic NOAEL of 75 mg/kg bw/d has been reported for ethyl benzene (Mellert et al., 2007).

Justification for selection of repeated dose toxicity inhalation - systemic effects endpoint:
A NOAEC of 3515 mg/m3 was reported by Carpenter et al. (1975) for generalised systemic effects in male rats and male dogs. Other studies have shown that some xylene isomers adversely affect hearing in the rat, with a sub-chronic NOAEC of 1950 mg/m3 reported for p-xylene; the NOAEC for ototoxicity of m-xylene and o-xylene was greater than 7810 mg/m3 (Gagnaire et al., 2001). The ototoxicity of mixed xylene appears to be dependent upon composition (Gagnaire et al., 2007), with a sub-chronic LOAEC of 1080 mg/m3 reported for one sample while another had a NOAEC of 2170 mg/m3. A LOAEC of 868 mg/m3 was reported for the ototoxicity of ethyl benzene in the rat (Gagnaire et al, 2007), equivalent to an extrapolated NOAEC of 500 mg/m3.

Repeated dose toxicity: via oral route - systemic effects (target organ) other: all gross lesions and masses

Repeated dose toxicity: inhalation - systemic effects (target organ) other: all gross lesions and masses

Justification for classification or non-classification

No classification for Xylene is warranted as ethylbenzene content is <10%.