N-hexane

Repeated dose toxicity: inhalation

Administrative data

Endpoint:

sub-chronic toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1991

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: This study is classified as reliable with restrictions because no GLP statement was included, but the study was otherwise well-documented and closely followed OECD guideline 413.

Data source

Reference

Reference Type:

publication

Title:

NTP Report on the Toxicity Studies of n-Hexane in B6C3F1 Mice

Author:

Dunnick, JK

Year:

1991

Bibliographic source:

National Toxicology Program, NIH Publication No. 91-3121

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 413 (Subchronic Inhalation Toxicity: 90-Day Study)

GLP compliance:

not specified

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

N-hexane

EC Number:

203-777-6

EC Name:

N-hexane

Cas Number:

110-54-3

Molecular formula:

C6H14

IUPAC Name:

hexane

Details on test material:

- Name of test material (as cited in study report): n-hexane
- Analytical purity: 99%
- Storage condition of test material: stable when stored in the dark at temps. below 60 degree Celsius.

Test animals

Species:

mouse

Strain:

B6C3F1

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Frederick Cancer Research Facility
- Age at study initiation: 7 weeks
- Weight at study initiation: mean weights: 23.3-23.8 g male, 18.3-20.0 female
- Housing: individually
- Diet (e.g. ad libitum): NIH 07 Rat and Mouse Ration, ad libitum overnight during exposure
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 14 days


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21.0-24.8
- Humidity (%): 40-84
- Photoperiod (hrs dark / hrs light): 12 hrs light/12 hrs dark

Administration / exposure

Route of administration:

inhalation: vapour

Type of inhalation exposure:

not specified

Vehicle:

not specified

Details on inhalation exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: 0.5 m3 Hinners-type chambers
- Source and rate of air: Air was passed through a 2 l reserve of n-hexane at 50 degree C with a fritted bubbler. Separate generators were used for each chamber.
- Temperature, humidity, pressure in air chamber: 21.0-24.8 degree C, and 40-84% humidity

TEST ATMOSPHERE
- Brief description of analytical method used: The 10,000 ppm chamber was monitored with a Miran 1A IR. The other chambers were monitored with a Miran 980 IR. Chambers were sampled once per hour. Biweekly samples were also analyzed using GC with flame ionization detection.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

The 10,000 ppm chamber was monitored with a Miran 1A IR. The other chambers were monitored with a Miran 980 IR. Chambers were sampled once per hour. Biweekly samples were also analyzed using GC with flame ionization detection.

Duration of treatment / exposure:

13 weeks

Frequency of treatment:

6 hrs/day, 5 days per week except for one 1000 ppm group that was exposed 22 hrs/day for 5 days/week

Doses / concentrations

Doses / concentrations 1

Remarks:

Doses / Concentrations:
0, 500, 1000, 4000, 10,000 ppm
Basis:
nominal conc.

Doses / concentrations 2

Remarks:

Doses / Concentrations:
1000 ppm
Basis:
nominal conc.

No. of animals per sex per dose:

18 mice per sex

Control animals:

yes, sham-exposed

Details on study design:

- Dose selection rationale: not provided

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily

BODY WEIGHT: Yes
- Time schedule for examinations: weekly

HAEMATOLOGY: Yes
- Parameters checked: leukocytes, lymphocytes, monocytes, eosinophils, segmented neutrophils, hematocrit, hemoglobin, mean corpuscular hemoglobin concentration, mean cell volume, erythrocytes, reticulocytes, platelets

NEUROBEHAVIOURAL EXAMINATION: Yes
- Dose groups that were examined: 0, 1000 (22 hr exposure), 10,000 ppm
- Battery of functions tested: startle response, foot splay, analgesia response, grip strength, locomotor activity, exploratory behaviour

Sacrifice and pathology:

GROSS PATHOLOGY: Yes, brain, heart, right kidney, liver, lung, spleen, right testis, thymus
HISTOPATHOLOGY: Yes, adrenal glands, brain, bronchial lymph nodes, cecum, colon, duodenum, esophagus, gallbladder, gross lesions, tissue masses, heart, ileum, jejunum, kidneys, larynx, liver, lungs, mainstream bronchi, mammary glands, mandibular lymph nodes, mesenteric lymph nodes, mediastinal lymph nodes, nasal cavity, turbinates, pancreas, parathyroid glands, pituitary gland, rectum, salivary glands, sciatic nerves, spinal cord, spleen, sternum including marrow, stomach, testes, ovaries, uterus, thymus, thyroid gland, trachea, urinary bladder

Statistics:

Multiple comparison methods of Dunnett.

Results and discussion

Results of examinations

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):

effects observed, treatment-related

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Gross pathological findings:

no effects observed

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Histopathological findings: neoplastic:

not examined

Details on results:

CLINICAL SIGNS AND MORTALITY
All animals survived to the end of the study. Clinical signs were limited to sneezing in the 10,000 ppm group.

BODY WEIGHT AND WEIGHT GAIN
Mean body weights of males in the 1000 ppm, 22 hr, exposure group and 10,000 ppm group were significantly reduced. The mean body weight of females in the 10,000 ppm group were also signficantly reduced.

HAEMATOLOGY
Segmented neutrophils were significantly increased in male mice exposed to 10,000 ppm.


NEUROBEHAVIOUR
Female mice in the 1000 ppm, 22 hr, exposure group and 10,000 ppm showed decreased locomotor activity.

ORGAN WEIGHTS
Liver, kidney, and heart weights were increased in exposed female mice.


HISTOPATHOLOGY: NON-NEOPLASTIC
Paranodal swellings in the tibial nerve were observed in 10,000 ppm exposed males and females, and the 1000 ppm, 22 hr exposure females. Inflammation and regeneration of the respiratory epithelium and olfactory epithelium, and metaplasia of olfactory epithelium to olfactory epithelium was in mice exposed to 10,000 ppm. Similar lesions, but of less severity were also seen in females in the 4,000 ppm group and 1000, 22 hr exposure, group females. Females in the 1000 and 500 ppm group showed minimal olfactory epithelium changes. Males in the 1000 ppm, 22 hr exposure group, and 1000 ppm group, had minimal lesions. Males in the 4000 ppm group and 500 ppm group did not show nasal lesions.

Effect levels

1

Key result

Dose descriptor:

NOAEC

Effect level:

500 ppm

Sex:

male

Basis for effect level:

other: nasal lesions

2

Key result

Dose descriptor:

LOAEC

Effect level:

1 000 ppm

Sex:

male

Basis for effect level:

other: nasal lesions

3

Key result

Dose descriptor:

LOAEC

Effect level:

500 ppm

Sex:

female

Basis for effect level:

other: nasal lesions

Target system / organ toxicity

Critical effects observed:

not specified

Any other information on results incl. tables

Significant results of mouse repeated dose inhalation study

Concentration	Control	500 ppm	1000 ppm	4000 ppm	10,000 ppm	1000 ppm (22 hr)
Male body weight (g)	33.4 ± 0.55	32.3 ± 0.97	31.2 ± 0.57	31.2 ± 0.75	27.8 ± 0.50	30.0 ± 0.60
Female body weight (g)	25.3 ± 0.49	25.1 ± 0.62	26.4 ± 0.66	25.6 ± 0.75	23.8 ± 0.57	25.4 ± 0.88
Male segmented neutrophils (10^3/microliter)	0.31 ± 0.054	0.40 ± 0.068	0.46 ± 0.108	0.25 ± 0.036	1.16 ± 0.363	0.55 ± 0.079
Locomotor activity in female mice - week 13	170 ± 9.1	168 ± 8.7	166 ± 9.2	152 ± 6.6	141 ± 10.6	126 ± 10.5

Applicant's summary and conclusion

Conclusions:

The LOAEC for female mice was 500 ppm based on nasal lesions. No NOAEC was found for female mice. The LOAEC for male mice was 1000 ppm based on nasal lesions, and the NOAEC was 500 ppm.

Executive summary:

This study examined the effect of 13 weeks inhalation exposure of n-hexane to mice. Groups of 18 female and 18 male mice were exposed to concentrations of 0, 500, 1000, 4000, or 10,000 ppm of test substance for 6 hrs/day, 5 days/week, for 13 weeks. An additional group was exposed to 1000 ppm of test substance for 22 hrs/day, 5 days/week, for 13 weeks. Clinical examinations were done twice daily, and body weights taken weekly. After sacrifice, animals were examined for histopathological parameters, and organ weights. 8 mice of each sex in each exposure group were examined for neurobehaviour. No animals died during the study, and the only neurological effects were decreased locomotion in females exposed to 10000 ppm. Nasal lesions were seen in females in all exposure groups, and in males exposed to 1000 ppm of test substance. The LOAEC for females was therefore 500 ppm (1760 mg/m3), with no NOAEC found. The NOAEC for males was 500 ppm (1760 mg/m3), with an LOAEC of 1000 ppm (3520 mg/m3) based on nasal lesions.