N-methylformamide

Repeated dose toxicity: inhalation

Administrative data

Endpoint:

short-term repeated dose toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Comparable to Guideline study with minor restrictions (only males used [however, LC50 study suggested that males are more susceptible]).

Data source

Reference

Reference 1

Reference Type:

study report

Title:

Unnamed

Year:

1983

Report date:

1983

Reference 2

Reference Type:

publication

Title:

2-Week inhalation study of N-monomethylformamide in rats

Author:

Kennedy GL, Ferenz RL, Burgess BA, Stula EF

Year:

1990

Bibliographic source:

Fundam Appl Toxicol 14: 810-816

Reference 3

Reference Type:

publication

Title:

Inhalation toxicity of N-methylformamide in the rat

Author:

Ferenz RL, Burgess BA, Stula EF, and Kennedy GL

Year:

1984

Bibliographic source:

Toxicologist 4: 65

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

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

Deviations:

no

GLP compliance:

not specified

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

N-methylformamide (NMF)

IUPAC Name:

N-methylformamide (NMF)

Details on test material:

purity 99.5% (0.2% dimethylformamide, 0.2% water, 0.1% unknown impurities)
Source: Dupont, Chemicals and Pigments Department
stable throughout the exposure phase of the study (GC method)

Test animals

Species:

rat

Strain:

other: Crl:CD BR

Sex:

male

Details on test animals or test system and environmental conditions:

Initial body weight: 207-242 g, age: 7-8 weeks
2 rats per cage
acclimatisation period 1 week
certified diet and water ad libitum
no data about housing conditions

Administration / exposure

Route of administration:

inhalation

Type of inhalation exposure:

nose only

Vehicle:

other: air

Remarks on MMAD:

MMAD / GSD: vapour

Details on inhalation exposure:

- Exposure apparatus: liquid test substance syringe-driven into a heated (180ºC) 3-neck flask, where it flash evaporated; unheated dilution air passed through the flask and carried vapours to the exposure chamber.
- Method of holding animals in test chamber: placed in cylindrical. perforated. stainless-steel restrainers equipped with conical nose pieces; restrainers were inserted Into face plates on the exposure chamber; nose of each rat protruded into chamber
- Method of conditioning air: no data

- Temperature near breathing zone: 24-27°C, one reading 34°C at exposure day 10
- humidity, pressure in air chamber: no data
- Air flow rate: 25 l/minute

TEST ATMOSPHERE
- Brief description of analytical method used: see below
- Samples taken from breathing zone: yes

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Atmospheric concentrations analyzed via gas chromatography (GC with a flame ionization detector; calibrated daily); sampling every 30 minutes
mean concentrations +- standard deviation in ppm: 50+-8, 130+-25, 398+-91.

Duration of treatment / exposure:

2 weeks
5 sacrificed immediately after the 10th exposure, 5 rats used solely for urine sampling, 5 rats sacrified after a post exposure observation period of 14 days

Frequency of treatment:

6h/day, 5 days/week

Doses / concentrations

Remarks:

Doses / Concentrations:
50, 130, 400 ppm (ca. 0.12, 0.32, 0.98 mg/l)
Basis:
analytical conc.

No. of animals per sex per dose:

15 (5 sacrificed immediately after the 10th exposure, 5 rats used solely for urine sampling, 5 rats sacrified after a post exposure observation period of 14 days)

Control animals:

yes, sham-exposed

Details on study design:

Post-exposure observation period: 14 days in a subgroup of 5 animals

Positive control:

no

Examinations

Observations and examinations performed and frequency:

All rats weighed and observed daily (excluding weekends) throughout the exposure and recovery periods.
Urine samples for analysis of NMF (GC method) were collected overnight from 5 rats on exposure days 1, 4, and 9 and on recovery days 3, 6, and 13.

Urine analysis clinical parameters: 10 rats, overnight urine sampling following the 9th exposures; 5 rats, overnight urine following the 13th day of recovery; measured parameters: urine volume, osmolality, ph, occult blood, protein, sugar, bilirubin, acetone, urobilinogen.

Hematology: blood sampling (tail vein) after the last exposure and day 14 of recovery period; erythrocyte count, hemoglobin concentration, hematocrit, platelet count, and total and differential leukocyte counts.

Clinical chemistry: Serum samples analyzed activities of alkaline phosphatase. alanine aminotransferase (ALT), aspartate aminotransferase( AST), urea nitrogen, creatinine, total protein, and cholesterol.

Sacrifice and pathology:

Necropsy of rats sacrificed after the 10th exposure and after the recovery period.
Absolute and relative organs weights measured: heart, liver, lungs, kidneys, spleen, testes, and thymus.

Histopathology of the following organs: adrenal glands, thyroid gland, esophagus, stomach, duodenum, parcreas, jejunum, ileum, cecum, colon. liver, spleen, thymus, mediastinal lymph nodes, eye, brain, trachea, heart, nose, urinary bladder, lungs, sternum, kidneys, testes, and epididymides.

Statistics:

signilicance at the 0.05 probability level
ANOVA, Bartlett's test and Dunnett's test

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:

no effects observed

Clinical biochemistry findings:

effects observed, treatment-related

Urinalysis findings:

no effects observed

Behaviour (functional findings):

not examined

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

Details on results:

Body weight
Rats exposed to 130 ppm had significantly lower body weights during the first week of exposure (slight but significant effect; mean bw 229 g versus 237 g in control); no effects on body weight were detected at later time points. However, rats exposed to 400 ppm had significantly lower body weights throughout the study (also in recovery period) with severe weight depression during the exposure phase and weight gain at a rate parallel to that of controls during the recovery phase. No effects at 50 ppm.

Clinical chemistry
50 ppm: no compound-related effects in rats
>=130 ppm: dose dependent increase in serum cholesterol concentrations.
400 ppm: decreased serum alkaline phosphatase activities, and increased serum ALT and AST activities; changes related to hepatotoxicity. All compound-related effects observed at the end of the exposure period were absent after the recovery period.

Pathologic examination revealed no compound-related macroscopic lesions in any rats.
Organ weights
50 ppm: no effects
>= 130 ppm: increased relative liver weight (no effects after recovery).
400 ppm: relative testes weights were increased following 10 exposures, no microscopic findings were observed in the testes, and no differences in testes weight were found fourteen days following exposure.

Histopathology
>=130 ppm: compound-related effects following exposure in the livers; lesions included pale cytoplasm, increase in the number of mitotic figures, and cytoplasmic lipid vacuolation; changes interpreted as being degenerative and regenerative in nature; after 14 days non-exposure period partial recovery at 400 ppm and complete recovery at 130 ppm occurred. See also following table.

NMF concentration in urine (limit of detection was 10 ppm)
NMF excreted in a dose-dependent fashion in the urine of exposed rats; it was detected in the urine of rats exposed to >=130 ppm on day 1 and in rats exposed to 50 ppm on day 4. In all test groups, urinary levels of NMF generally increased throughout the exposure period and then decreased throughout the recovery period. At the end of the recovery period, NMF was still detectable in rats of mid and high dose level, but not in rats exposed to 50 ppm.

Effect levels

1

Dose descriptor:

NOAEC

Effect level:

50 ppm (analytical)

Sex:

male

2

Dose descriptor:

LOAEC

Effect level:

130 ppm (analytical)

Sex:

male

Basis for effect level:

other: reduced body weight; hepatotoxicity

Target system / organ toxicity

Critical effects observed:

not specified

Any other information on results incl. tables

Histopathological effects in the liver of rats after sub-acute inhalation exposure to N-methylformamide

Incidences in male rats immediately after 10 exposures (a) and after the 14 day recovery period (b)

5 rats in each group evaluated for effects

Investigated parameter	Dose level in ppm
	0	50	130	400
Centrilobular cytoplasmic vacuoles	0(a) 1(b)	0(a) 4(b)	0(a) 1(b)	0(a) 0(b)
Focal haematopoiesis	0(a) 1(b)	1(a) 0(b)	1(a) 0(b)	0(a) 0(b)
Hepatocellular cytoplasmic lipid vacuoles	0(a) 0(b)	0(a) 0(b)	1(a) 0(b)	5(a) 0(b)
Increased centrilobular mitotic figures	0(a) 0(b)	0(a) 0(b)	3(a) 0(b)	2(a) 0(b)
Pale cytoplasm, hepatocellular	0(a) 0(b)	0(a) 0(b)	3(a) 1(b)	0(a) 5(b)
Pale cytoplasm, hepatocellular, centrilobular	0(a) 0(b)	0(a) 0(b)	2(a) 0(b)	5(a) 0(b)

Applicant's summary and conclusion

Conclusions:

In a sub-acute inhalation study in male rats the target organ was the liver, the NOAEC was 50 ppm (0.12 mg/l) and the LOAEC 130 ppm (0.32 mg/l).

Executive summary:

Comparable to Guideline study with minor restrictions (only males used [however, LC50 study suggested that males are more susceptible]).

Groups of 15 male Crl:CD BR rats each were exposed by nose-only inhalation 6 hr/day, 5 days/week, for 2 weeks to either 0 (control), 50, 130, or 400 ppm N-methylformamide (NMF). Five rats per group were killed following the 10th exposure, five were sacrificed after a 14-day post exposure recovery period and five rats were used to determine urinary NMF excretion. Parameters investigated were clinical observations and body weights, urinalysis, hematology, clinical chemistry and gross and microscopic pathology including organ weights. Liver damage occurred in rats exposed to >= 130 ppm. This was detected by increased relative liver weights and histopathological alterations. At 400 ppm also effects on serum enzyme activity (ALT, AST, alkaline phosphatase) were found. After the 14 days non-exposure period partial recovery was seen at 400 ppm and complete recovery at 130 ppm. The amount of NMF excreted in the urine was dependent on the exposure concentration and NMF was present 14 days post exposure at the higher exposure levels.

Conclusion: In a sub-acute inhalation study in male rats the target organ was the liver, the NOAEC was 50 ppm (0.12 mg/l) and the LOAEC 130 ppm (0.32 mg/l).