chloromethane; methyl chloride

Repeated dose toxicity: inhalation

Administrative data

Endpoint:

chronic toxicity: inhalation

Remarks:

combined repeated dose and carcinogenicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

June 1978 - June 1980

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Comparable to guideline study with acceptable restrictions

Cross-reference

Reason / purpose for cross-reference:

reference to same study

Data source

Reference

Reference 1

Reference Type:

study report

Title:

Unnamed

Year:

1981

Report date:

1981

Reference 2

Reference Type:

other: summary

Title:

Unnamed

Year:

1983

Reference 3

Reference Type:

secondary source

Title:

Methyl Chloride

Author:

Löf, A. et al.

Year:

2000

Bibliographic source:

Concise International Chemical Assessment Document 28

Reference 4

Reference Type:

secondary source

Title:

Chloromethane CAS: 74-87-3

Author:

OECD SIDS

Year:

2004

Bibliographic source:

SIDS Initial Assessment Report for SIAM 15 ; www.iccahpv.com

Reference 5

Reference Type:

secondary source

Title:

Toxicological review of methyl chloride (CAS No. 74-87-3)

Author:

Greenberg, M. and Riddle, J.

Year:

2001

Bibliographic source:

U.S. Environmental Protection Agency, EPA/635/R01/003

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 453 (Combined Chronic Toxicity / Carcinogenicity Studies)

Deviations:

yes

Remarks:

no measurements on food consumption; more animals per sex and dose

GLP compliance:

no

Remarks:

test was performed prior to GLP-requirement

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Chloromethane

EC Number:

200-817-4

EC Name:

Chloromethane

Cas Number:

74-87-3

Molecular formula:

CH3Cl

IUPAC Name:

chloromethane

Details on test material:

- Name of test material (as cited in study report): methyl chloride
- Physical state: gaseous
- Analytical purity: 99.97%
- Purity test date: 1978-04-17, 1978-07-14; 1978-11-14, 1980-01-16
- Lot/batch No.: A-250
- Storage condition of test material: in 24 size IJ cylinders containing 100 pounds

Test animals

Species:

rat

Strain:

Fischer 344

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Breeding Laboratories, Portage, Michigan, USA
- Age at study initiation: 7 weeks
- Weight at study initiation (mean values): females: 68.7 g; males: 78.2 g
- Housing: Rats were placed into stainless steel wire mesh cages manufactured by Allentown Caging Company of Allentown, New Jersey (dimensions: 58 inches long, 12 inches wide, 7 inches high). There were 12 compartments (dimensions: 7 inches high, 4-5 inches wide and 12 inches long) for animal holding in each cage. Rats were caged in individual compartments. Control and test animals were housed in separate rooms during non-exposure periods.
- Diet: Purina Rodent Chow 5001. During the early phases of the studies, the animals were fed a handful of feed blocks. Later on, just 5 or 6 blocks were placed into each feed cup after exposure.
- Water: ad libitum (automatic watering system)
- Acclimation period: 14 days


ENVIRONMENTAL CONDITIONS (holding rooms):
- Temperature: 21.1 °C
- Relative humidity: 45%
- Air changes: 15 per hr
- Photoperiod: 12 hrs dark / 6 hrs light


IN-LIFE DATES: From: 15th June 1978 To: 23th June - 1st July 1980

Administration / exposure

Route of administration:

inhalation

Type of inhalation exposure:

whole body

Vehicle:

other: unchanged (no vehicle)

Remarks on MMAD:

MMAD / GSD: not applicable

Details on inhalation exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: Hinners' type exposure chambers. These chambers were constructed of stainless steel and were five feet square having a pyramidal top and a bottom with a nominal volume of 5 m3. The chambers accommodated five tiers of animals. Each tier of animals was supported on open mesh flooring which was supported by three one and one-quarter inch stainless steel pipes.
- Source and rate of air: Room air that had been filtered with an absolute filter mounted on the intake to each chamber was drawn through the chamber by a large blower mounted on the roof of the facility. The room air had previously been cleaned by passing it through an electrostatic precipitator and a bacteriostatic LiCl solution.
- Method of conditioning air: Chloromethane was introduced into the chamber at the metering orifice so that it was well mixed with the incoming air by turbulence. The desired test concentration in each chamber (50, 225 and 1000 ppm) was regulated passing the chloromethane gas through the precision rotameters mounted on each chamber.
- Temperature, humidity, pressure in air chamber: average temperature: 22.7 °C, average relative humidity 51.9%, slightly negative pressure (approximately 1 inch of water)
- Air flow rate: 1 m3/min
- Air change rate: 12 air changes per hour



TEST ATMOSPHERE
- Brief description of analytical method used: The concentration of the chloromethane in each chamber was monitored every 20 to 24 minutes of each exposure period using GC-FID-analyses. After an initial evaluation, the calibration of the GC was checked approximately weekly by sampling a reference span gas.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

The 24 months overall mean values are:
0.3 +/- 4 ppm
51 +/- 9 ppm
224 +/- 16 ppm
997 +/- 65 ppm

Duration of treatment / exposure:

24 months (interim sacrifices at 6, 12, and 18 months)

Frequency of treatment:

6 hours/day, 5 days/week (excluding holidays)

Doses / concentrations

Remarks:

Doses / Concentrations:
50, 225 or 1000 ppm (103, 465 and 2065 mg/m³)
Basis:
nominal conc.

No. of animals per sex per dose:

120

Control animals:

yes, sham-exposed

Details on study design:

- Dose selection rationale: the dose selection was based on the effects in a 90 day subchronic study
- Post-exposure period: none

Positive control:

no

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily

DETAILED CLINICAL OBSERVATIONS: No

BODY WEIGHT: Yes
- Time schedule for examinations: at the beginning of the study, weekly for the first 6 months, and biweekly thereafter (mice were weighed by compartment groups of four or less)

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

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: No

WATER CONSUMPTION: No

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: prior to the first exposure and within one week of the scheduled sacrifice
- Dose groups that were examined: each animal of all dose groups and controls

HAEMATOLOGY: Yes
- Time schedule for collection of blood: at 6, 12, 18 (interim sacrifices) and 24 months of exposure
- Anaesthetic used for blood collection: Yes (pentobarbital)
- Animals fasted: Yes (22 hours)
- How many animals: typically 10 animals per sex and dose; exception at the 18 month interim sacrifice: 5 males and 10 females in the control, 50 ppm and 225 ppm group; 7 males and 8 females in the highest dose group.
- The following hematology parameters were analysed: hemoglobin (HGB), hematocrit (HCT), white blood count (WBC), red blood count (RBC), mean cell volume (MCV), mean cell hemoglobin (MCH), mean cell hemoglobin concenttration (MCHC), reticulocytes, and different white cell count.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: at 6, 12, 18 (interim sacrifices) and 24 months of exposure
- Animals fasted: Yes (22 hours)
- How many animals: typically 10 animals per sex and dose; exception at the 18 month interim sacrifice: 5 males and 10 females in the control, 50 ppm and 225 ppm group; 7 males and 8 females in the highest dose group
- Sera were analysed for the following clinical chemistry parameters: glucose, blood-urea-nitrogen (BUN), alkaline phosphatase (AP), serum glutamic-oxaloacetic transaminase (SGOT), and serum glutamic-pyruvic transaminase (SGPT). Creatine phosphokinase was analysed at the 6-months interim necropsy only.

URINALYSIS: Yes
- Time schedule for collection of urine: at 6, 12, 18 (interim sacrifices) and 24 months of exposure
- Metabolism cages used for collection of urine: Yes (for a 16-hour period)
- Animals fasted: Yes (but water ad libitum)
- The following determinations were performed: total volume (Vol), secific gravity (Sp. Gr.), pH, glucose (Glu), ketones (Ket), protein (Prot), occult blood (Occ), and urine sediment

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: prior to the scheduled 18 month interim necropsy and at the final scheduled necropsy
- Dose groups that were examined: each animal of all dose groups and control
- Battery of functions tested: posture and gait, facial tone, pupillary reflex, palpebral reflex, extensor thrust/scratch reflex, crossed extensor reflex, clutch response

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes

Interim sacrifices at 6, 12 and 18 months (10 rats per sex and dose at the 6 and 12 month interim sacrifices, 20 rats per sex and dose at the 18 month interim sacrifice); the remaining animale were sacrificed at 24 months.

Tissues examined from each animal: adrenals, aorta, bone marrow, brain, cecum, colon, duodenum, ear canals, epididymides, esophagus, eyes, fallopian tube, femur, heart, ileum, jejunum, kidneys, liver, lungs, mammary gland, mandibular lymph node, mesenteric lymph node, muscle, nasal turbinate, nerve (sciatic), ovaries, pancreas, parathyroid, pituitary, prostate, rectum, salivary gland, seminal vesicles, skin, spinal cord, spleen, stomach, testes, thymus, thyroid, tissue masses, trachea, urinary bladder, uterus

Other examinations:

Fresh liver impression smears were made at necropsy. After drying the smears were examined for porphyrin flourescence.

Statistics:

Sample variance for each exposure group were tested for homogeneity by the method of Bartlett (1937). Depending on the significance of the result further tests were used: ANOVA analysis or Kruskal-Wallis test (Kruskal and Wallis, 1952). Where results of the ANOVA tests were found statistically significant, individual treatment versus control group mean comparison were made using the least significant difference test (LSD) (Steel and Torrie, 1960). Nonparametric equivalents to the LSD test as described by Dunn (1964) and Miller (1966) were used to make treatment versus control group comparisons when the K-W test results were statistically significant.
Chi-square tests (Agresti and Wackerly, 1977; March, 1972) for homogeneity were performed on clinical, opthalmological, and neurobehavioral observations for male rats and female rats.
Mortality and lesion incidence data were prepared using the the acturial life table method developed by Berkson and Gage (1950) in conjunction with the National Cancer Institute’s bioassay data analysis program (Thomas et al., 1977; Gart et al., 1979). Survival curves were calculated from these data by the method of Kaplan and Meier (1958).
The level of significance used in all test procedures with unadjusted data was α = 0.05 and for adjusted data was the unadjusted α divided by the number of treatment groups.

Results and discussion

Results of examinations

Details on results:

CLINICAL SIGNS AND MORTALITY
Clinical observations were unaffected in rats exposed to all concentrations. Rat survival was unaffected by exposure to any concentration.

BODY WEIGHT AND WEIGHT GAIN
The growth rate for male and female rats at 1000 ppm and for female rats at 225 ppm of chloromethane was significantly reduced during the first 24 weeks. From 24 weeks onward, the body weights of these groups generelly remained lower than the control values, but only the 1000 ppm groups were consistently below control values.

OPHTHALMOSCOPIC EXAMINATION
Ophthalmologic examinations revealed changes which were apparently due to a virus and which were also
found in control animals, although at a lower incidence. Corneal cloudiness and opacity at 18 months was significant in female rats (control 2/20, 50 ppm 4/20, 225 ppm 12/20 and 1000 ppm 12/20), but not males, and neither were different from controls at 24 months. Lenticular changes, which appeared in rats only at 18 months, may have been related to exposure. However, the apparent increase in the incidence of anterior lens sutures in male and female rats at 24 months was not statistically significant (P > 0.05).

HAEMATOLOGY
Hematology was unaffected in rats exposed to all concentrations.

CLINICAL CHEMISTRY
Clinical chemistry was unaffected in rats exposed to all concentrations.

URINALYSIS
Urinalysis was unaffected in rats exposed to all concentrations.

NEUROBEHAVIOUR
No neurofunctional impairments were observed that were attributable to chloromethane exposure.

ORGAN WEIGHTS
Organ weights showed significant changes only in rats exposed to 1000 ppm. Increased relative heart weights were found in male rats exposed to 1000 ppm at 12, 18 and 24 months and in female rats at 12 and 24 months. Relative kidney weights were increased in male rats exposed to 1000 ppm at all sacrifice periods but female rats were unaffected. Male rats exposed to 1000 ppm had increased relative liver weights and female rats had decreased absolute liver weights at all times. Decreased absolute brain weights were generally observed at all time periods for male and female rats exposed to 1000 ppm. Testicular weights of male rats exposed to 1000 ppm were decreased at 18 and 24 months when compared to the controls on both an absolute and relative basis. Relative lung weights of male rats were increased at all concentrations but only at the 6-month sacrifices.
No biologically relevant changes in organ or body weights occured in male or female rats at concentration less than 1000 ppm.

GROSS PATHOLOGY/HISTOPATHOLOGY:
The testes were the only organ of the rats considered to have significant chloromethane induced lesions. Bilateral and diffuse degeneration and atrophy of the seminiferous tubules of the testes were first noted in four of ten males exposed to 1000 ppm for 6 months. The effect increased in degree and in number of animals affected until the 18-month sacrifice. By 24 months, the effects of normal ageing prevented interpretation. However, with increased exposure to chloromethane, the resultant significant decrease in bilateral compressive degeneration and atrophy and the significant increase in unilateral compressive degeneration and atrophy (caused by testicular tumors) correlated with a decrease in interstitial cell tumor size. This observation was supported by decreased testicular weights and testes/body weight ratios in rats exposed to 1000 ppm chloromethane. No changes in the testes were detectable at either 50 or 225 ppm.
Sperm granulomas were noted in three male rats at 1000 ppm: two at 6 months and one at 24 months. Their presence could not be directly attributed to chloromethane exposure.
Other lesions noted in rats, such as C-cell carcinoma, pituitary adenomas, and mandibular lymph node hyperplasias, were not related to chloromethane exposure.

Effect levels

Dose descriptor:

NOAEC

Effect level:

465 mg/m³ air (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: reduced body weight

Target system / organ toxicity

Critical effects observed:

not specified