Methyl (S)-(-)-lactate

Repeated dose toxicity: inhalation

Administrative data

Endpoint:

short-term repeated dose toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Data source

Reference

Reference Type:

publication

Title:

Subchronic inhalation toxicity of methanol

Author:

Andrews L.S., Clary J.J., Terrill J.B. and Bolte H.F.

Year:

1987

Bibliographic source:

Journal of Toxicology and Environmental Health 20, 117-124

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 412 (Subacute Inhalation Toxicity: 28-Day Study)

Principles of method if other than guideline:

No guideline was explicity followed, but the study design is equivalent/similar to OECD TG 412.

GLP compliance:

not specified

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Methanol

EC Number:

200-659-6

EC Name:

Methanol

Cas Number:

67-56-1

Molecular formula:

CH4O

IUPAC Name:

methanol

Specific details on test material used for the study:

- Name of test materail used: Methanol
- Source: Celanese Corporation, New York, N.Y
- Purity: 99.85%

Test animals

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, Kingston, N.Y., USA
- Age at study initiation: 35 days
- Weight at study initiation: not specified
- Housing:n rack-mounted, wire-mesh cages
- Diet: ad libitum, Purina Laboratory Chow 5001
- Water: ad libitum, tap water
- Acclimation period: 15 days

Administration / exposure

Route of administration:

inhalation: vapour

Type of inhalation exposure:

whole body

Vehicle:

air

Details on inhalation exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: Four-cubic-meter exposure chambers constructed of glass and stainless steel with conical tops and bottoms.
- Method of holding animals in test chamber: The positions of individual animals were rotated on the rack during the study.
- Source and rate of air: Chambers were operated dynamically at an average airflow rate of 2000 L/min. This airflow rate was calculated to provide 1 air change every 3.0 minutes and a T99 equilibrium time of 13.8 min.
- System of generating particulates/aerosols: Test atmospheres were generated by ambient flash evaporation. Liquid methanol was placed in a reservoir and fed with a fluid metering pump via Teflon tubing to a Spraying Systems Atomizer. The atomizer was mounted in the inlet portal of the exposure chamber, and the methanol aerosol was delivered as a fine aerosol into the incoming stream of house-supply chamber air. The aerosol droplets evaporated in the inlet air stream to yield a vapor in the exposure chamber.

TEST ATMOSPHERE
- Brief description of analytical method used: Methanol exposure levels were monitored with a Wilkes MIRAN model IA-CVF infrared analyzer at a wavelength of 3.4 µM. The initial calibration was performed using a 14-point check that was conducted on 2 d and in replicate. After the initial calibration, a three-point check was done each exposure day. The 500-ppm exposure chamber was monitored by the basic MIRAN IA unit. For monitoring of the 2000- and 5000-ppm exposure chambers, a 1:1 (room air: chamber air) dynamic dilution system was employed with the MIRAN. This modification was required for on-scale MIRAN readings. The resultant (diluted) values were multiplied by 2 to obtain the chamber concentration of methanol.
- Samples taken from breathing zone: not stated

Analytical verification of doses or concentrations:

yes

Duration of treatment / exposure:

20 days

Frequency of treatment:

6 hours a day, 5 days a week for a period of 4 weeks

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 ppm (nominal)

Remarks:

Control group

Doses / concentrations 2

Dose / conc.:

500 ppm (nominal)

Remarks:

Low concentration

Doses / concentrations 3

Dose / conc.:

2 000 ppm (nominal)

Remarks:

Mid concentration

Doses / concentrations 4

Dose / conc.:

5 000 ppm (nominal)

Remarks:

High concentration

No. of animals per sex per dose:

5

Control animals:

yes, concurrent no treatment

Positive control:

-

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice a day

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: once a week

BODY WEIGHT: Yes
- Time schedule for examinations: individual body weights were recorded prior to study initiation, on the first day of exposure and weekly thereafter

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: at pretest and at termination. Ophthalmoscopic examination included the following procedures: Lids, lacrimal apparatus, and conjunctiva were examined grossly. The cornea, anterior chamber, lens, vitreous humor, retina, and optic disc were examined by indirect ophthalmoscopy. Mydriasis was induced by tropicamide and atropine.
- Dose groups that were examined: all

HAEMATOLOGY: No
CLINICAL CHEMISTRY: No
URINALYSIS: No
NEUROBEHAVIOURAL EXAMINATION: No

Sacrifice and pathology:

Rats were sacrificed by exsanguination under ethyl ether anesthesia.
The external body surfaces, all orifices, body cavities and their visceral organs, cervical tissues and organs, and carcass were examined. Tissues and organs were examined in situ and after removal. Organ weights were taken for the adrenal glands (paired), heart, kidneys (paired), liver, lungs, spleen, testes (with epididymides paired), and ovaries (paired).
Approximately 35 different tissues per animal were preserved for microscopic examination. The eyes, testes, and epididymides were preserved in Bouin's solution for 48-72 h, washed in 3 changes of 70% ethanol for 72 h (ethanol changed every 24 h), and stored in 10% neutral buffered formalin. The lungs were infused with 10% neutral buffered formalin via the trachea with an amount of fluid approximately equal to 75% of the lung capacity and were placed into 10% neutral buffered formalin. Bone marrow smears were fixed in absolute methanol and stored unstained, for future reference. All of the other tissue samples were routinely preserved in 10% neutral buffered formalin. Slides from the nasal turbinâtes, trachea, lungs, kidneys, esophagus, liver, and eye (with optic nerve) from all animals from control and high-exposure groups were prepared and examined microscopically.

Statistics:

Body weights, organ weights, and organ/body weight ratios were compared by a statistical evaluation of equality of means. This was done using the appropriate one-way analysis of variance technique, followed by a multiple-comparison procedure if needed.
The parametric procedures were the standard one-way ANOVA used to determine which means were significantly different from the control. If a nonparametric procedure for testing equality of means was needed, the Kruskal-Wallis test was used, and if differences were indicated, a summed-rank test was used to determine which treatments differed from the control (Dunnett, 1955, 1964).
A statistical test for trend in the dose levels was also performed. In the parametric case (i.e., equal variance), standard regression techniques with a test for trend and lack of fit were used. In the nonparametric case, Jonckheere's test for monotonic trend was used. The test for equal variance (Bartlett's) was conducted at the 1%, two-sided risk level. All other statistical tests were conducted at the 5% and 1% two-sided risk levels.
Statistical evaluations were not performed when the standard error for the control group or more than one group was 0.0 due to lack of variance. If a standard error for one treated group was 0.0 or when N (number of animals) was less than or equal to 2 animals for any treated group, it was not included in the statistical evaluation.

Results and discussion

Results of examinations

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

Increased instances of discharges about the eyes and nose-lacrimation, mucoid nasal discharge, red nasal discharge, and dried red nasal discharge - were observed. The frequencies of these findings were increased in the treated groups, but only the incidence of mucoid nasal discharge appeared to be dose-related.

Mortality:

no mortality observed

Description (incidence):

All animals survived the duration of the study.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

Body weights for treated rats were comparable to their respective controls.

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:

no effects observed

Description (incidence and severity):

Results of terminal ophthalmoscopic examinations on high-dose rats revealed no ocular abnormalities that could be attributed to methanol exposure.

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Endocrine findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Description (incidence and severity):

Comparisons of organ weights for male rats revealed no statistically significant intergroup differences. Female rats of the mid-dose group had significantly (p≤ 0.05) higher relative spleen weights. These differences by themselves are not of any apparent biological significance.

Gross pathological findings:

no effects observed

Description (incidence and severity):

Gross pathological examination of control and high-dose rats revealed no effects from exposure to methanol.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

no effects observed

Description (incidence and severity):

Histopathological examination of control and high-dose rats revealed no effects from exposure to methanol.

Other effects:

not specified

Effect levels

Key result

Dose descriptor:

NOAEC

Effect level:

5 000 ppm (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: no adverse effects observed

Target system / organ toxicity

Critical effects observed:

no

Any other information on results incl. tables

Chamber concentrations:

Based on infrared analysis of chamber atmospheres, animals in groups I, II, III, and IV received exposure to mean concentrations of 0, 520, 1980, and 5010 ppm, respectively. Mean nominal concentrations were 550 (group II), 2330 (group III), and 5330 (group IV) ppm. Occasional discrepancies between nominal and analytical values were within the limits of expected experimental error (± 10%)

Applicant's summary and conclusion

Conclusions:

In conclusion, in a sub-chronic repeated-dose toxicity study, methanol was administered to rats for 13 weeks via inhalation. The repeated daily exposure to methanol vapors (up to 5000 ppm) does not result in adverse effects on body / organ weights, gross and histopathologic or ophthalmoscopic examinations in rats. Based on the results, the NOAEC was established to be 5000 ppm for methanol.

Executive summary:

In a sub-acute repeated dose toxicity study, methanol (purity 99.85%) was administered to 5 male/female Wistar rats/dose by inhalation in whole body exposure chambers at concentrations of 0, 500, 2000 and 5000 ppm as vapour, for 6 hours per day, 5 days/week for a total of 20 days.

No treatment-related effects on mortality, clinical signs, body / organ weights, gross/histopathologic or ophthalmoscopic examinations were noted. The only treatment and dose-related effect noted was that of mucoid nasal discharge in rats, which was considered reflective of upper respiratory tract irritation.

Based on the results, the NOAEC can be considered to be 5000 ppm, which equals 6.552 mg/L. Thus, in accordance with CLP Regualtion 1272/2008 no classifcation for STOT RE is warranted.