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Toxicological information

Repeated dose toxicity: inhalation

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Administrative data

Endpoint:
sub-chronic toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
circa 1982
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Essentially a GLP/Guideline study
Cross-reference
Reason / purpose:
reference to same study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1982
Report Date:
1982

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 413 (Subchronic Inhalation Toxicity: 90-Day Study)
Deviations:
yes
Remarks:
feed consumption was not measured.
GLP compliance:
yes
Remarks:
Study was initiated prior to GLP and completed with GLP.
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
Commercial NE (Lot No. 8020-513) was obtained from International Minerals Company (IMC, Chemical Group, Inc., Terre Haute, Indiana). A sample was analyzed for impurities and identified by gas chromatography-mass spectroscopy on an LKB9000 using a 10 ft, 10% 20M Carbowax column. Paraldehyde, 2-nitropropane, nitromethane, and nitroethane were found. To quantitate the compounds present in the test material, an infrared spectrophotometer (IR) analysis was performed which specified the following: Paraldehyde no IR features shown, suggests presence in very low quantities (ppm range)2-Nitropropane 1.5%, maximumNitromethane 1%, maximumNitroethane 97+%, by difference

Test animals

Species:
rat
Strain:
Fischer 344
Sex:
male/female
Details on test animals and environmental conditions:
Fischer 344 rats (15/sex), 9 weeks old, from Charles River Laboratory, Wilmington, MA were used in this study. Each rat was uniquely identified with a numbered metal ear tag. All animals were housed in stainless steel cages with wire bottoms (2 rats/cage) and were acclimated for 14 days prior to exposure in an animal holding room designed to maintain a temperature of 70°F, relative humidity of 50%, and a light cycle of 12 hours light and dark. A standard diet (Purina Laboratory Chow, Ralston Purina Co., St. Louis, MO) and water were removed during the exposure period, but were available ad libitum at all other times. Rats were assigned to control or treatment groups by a statistical randomization procedure using a computer program.

Administration / exposure

Route of administration:
inhalation: vapour
Type of inhalation exposure:
whole body
Vehicle:
air
Remarks on MMAD:
MMAD / GSD: Not applicable.
Details on inhalation exposure:
Exposures were conducted in 1 cubic meter stainless steel and glass Rochester-type chambers under dynamic airflow conditions. Airflow was approximately 175 i/min. Temperature and relative humidity in the animal holding room and in the chambers were controlled by a system designed to maintain temperature at approximately 70°F and relative humidity at approximately 50%.The NE vapor was generated by metering the test material at a controlled rate with a precision pump into a warmed vaporization flask (100-130°C). The vapor was swept with compressed air into the air inlet of the exposure chamber and then diluted to the desired concentration.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Analysis for NE in the exposure chamber atmosphere was performed by an infrared spectrophotometer equipped with a variable path1ength gas cell (Miran I, Foxboro, Norwalk, CT). The wavelength used for the analYSis was 11.5 microns. The analysis was performed 1-2 times per hour for each exposure concentration. Standards for the analysis were prepared by injecting a measured volume of NE into aU-tube connected to a SARAN gas sampling bag. The U-tube was gently heated with an air gun to improve evaporation of NE. The analytical concentration of NE in the exposure chamber was determined by interpolation from a standard curve derived from vapor standards of known concentrations. Complete standard curves were run prior to the first exposure and weekly thereafter. One standard bag was run before every exposure day to check the analytical system. The nominal concentration of NE in the exposure chamber was the ratio of the rate at which the test material was dispensed to the rate of total airflow through the chamber.
Duration of treatment / exposure:
6 hours/day
Frequency of treatment:
5 days/week (excluding holidays) for a 92 day period. (an interim group of 5 rats/sex/concentration were exposed for a 30 day period).
Doses / concentrations
Remarks:
Doses / Concentrations:0, 100, 350 or 1000 ppmBasis:nominal conc.
No. of animals per sex per dose:
15 rats/sex/concentration (an interim group of 5 rats/sex/concentration were exposed for a 30 day period).
Control animals:
yes, sham-exposed
Details on study design:
Four groups of Fischer 344 rats (15/sex), 9 weeks old were exposed to 0, 100, 350 or 1000 ppm (0, 0.3, 1.0, and 3.0 mg/L) of NE. Five rats/sex/exposure concentration were exposed for 6 hours/day, 5 days/week for a 30-day period, respectively, at which times they were necropsied, i.e. the interim kill. The remaining rats (10/sex) were exposed for 6 hours/day, 5 days/week (excluding holidays) for a 92-day period until they were necropsied, i.e. the terminal kill of the l3-week study.
Positive control:
No data

Examinations

Observations and examinations performed and frequency:
Body weights were obtained prior to the first exposure, twice weekly for the first two weeks and once weekly thereafter.
Sacrifice and pathology:
Clinical Observations: Blood was collected for hematology analysis and clinical chemistry studies from the tail vein and from the cervical vessels of rats, respectively. Urine was collected for urinalysis from rats only. The hematologic parameters included red blood cell (RBC) counts, total and differential white blood cell (WBC) counts, hemoglobin (Hgb) concentration and packed cell volume (PCV)[a]. Reticulocytes were determined as described by Wintrobe (1961); and Heinz bodies were determined by a method developed in this laboratory.[b] Urinalysis included pH, glucose, protein, ketones, bilirubin, occult blood, and urobilinogen[c] as well as specific gravity.[d] The clinical chemistry studies included the determination of blood urea nitrogen (BUN), serum glutamic pyruvic transaminase (SGPT), serum glutamic oxaloacetic transaminase (SGOT), serum alkaline phosphatase (AP), glucose, bilirubin, serum creatinine, phosphorus (P), and calcium (Ca)[e]. Blood for methemoglobin determination was collected by orbital sinus puncture from rats and mice and was analyzed by the method described by Hainline (1965) .Pathology: Gross necropsy was performed on all animals. All rats were fasted overnight. All animals were weighed prior to necropsy to determine organ to body weight ratios for the liver, kidney, brain, heart, thymus, and testes (male) which were obtained at necropsy. All animals were anesthetized with methoxyflurane and decapitated after the trachea was clamped. Blood samples were co11ected from the severed ,cervical vessels for clinical chemistry determinations as previously mentioned. A complete gross pathologic examination was performed, including examination of the eyes by pressing a wet microscope slide against the cornea and examining it under bright fluorescent illumination. The eyes from 5 rats/sex/concentration for both the interim and terminal kills were preserved in Zenker's solution. The eyes of the 'remaining rats of the terminal kill were preserved in 10% neutral phosphate-buffered formalin. The lungs were distended to approximately their normal inspiratory volume with 10% neutral phosphate-buffered formalin.Representative specimens of approximately 40 tissues were retained and preserved in 10% neutral phosphate-buffered formalin. A complete set of tissues (except sternum) was prepared and histologically evaluated from rats that died spontaneously and that were selected for the interim and terminal kills. Of the terminal kill, only 5/sex/species of the 0 and 1000 ppm exposure groups were evaluated. As a minimum, sections of liver, lungs, kidneys, heart, thymus, spleen, and nasal turbinates were prepared and evaluated from all spontaneous dead, all interim kill, and 5/sex of the terminal kill rats from the 100 and 350 ppm exposure groups. These tissues were processed for microscopic examination by conventional histologic methods, stained with hematoxylin and eosin and evaluated by light microscopy. In a few cases, a tissue specimen was not obtained at necropsy or was lost during the processing, and tissues such as the parathyroid glands were evaluated only to the extent that they were included in theroutine sections of the thyroid glands.[a] RBC , WBC-Coulter Counter Model ZBI; Hgb-Hemoglobinometer, Coulter Electronics, Hialeah, FL; PCV-Microhematocrit Centrifuge, Clay-Adams Co., New York, NY.[b] Standard Operating Procedures of Pathology Group, Toxicology Research, Dow Chemical USA.[c] Multistix, Ames Co., Elkhart, IN.[d] T. S. Meter American Optical Co., Buffalo, NY[e] CentrifiChem System 400, Methods File, Union Carbide Corp., Rye, NY.ReferencesHainline Jr., A. (1965). in Standard Methods of Clinical Chemistry, Academic Press, New York and London, Vol. 5, pp 143-157.
Other examinations:
No additional information available.
Statistics:
Hematology, methemoglobin, specific gravity of urine, clinical chemistry, body weights, organ weights, and organ to body weight values were evaluated by analysis of variance and Dunnett's test using a level of significance of p <0.05.

Results and discussion

Results of examinations

Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Cyanotic skin color, dull and dark red eyes in males and females and unkept appearance in females
Mortality:
mortality observed, treatment-related
Description (incidence):
Cyanotic skin color, dull and dark red eyes in males and females and unkept appearance in females
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
decreased body weight
Food consumption and compound intake (if feeding study):
not specified
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
not specified
Haematological findings:
effects observed, treatment-related
Description (incidence and severity):
Increased reticulocytes and Heinz bodies. Methemoglobin levels elevated at all concentrations
Clinical biochemistry findings:
effects observed, treatment-related
Description (incidence and severity):
Increased bilirubin levels in 1000 ppm females. Decreased glucose levels of males and females exposed to 350 or 1000 ppm.
Urinalysis findings:
no effects observed
Behaviour (functional findings):
not specified
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
Due to decreased body weights a number of absolute and/or relative organ weights were affected.
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
Increased spleen size
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Nasal turbinates exhibited chronic active inflammation of the olfactory epithelium, salivary gland changes, spleen exhibited extramedullary hematopoiesis, liver had fatty change and kidneys had decreased cytoplasmic granularity of proximal tubular cells.
Histopathological findings: neoplastic:
not specified
Details on results:
For each exposure chamber the mean daily time-weighted average (TWA) analytical concentration was very close to the intended target concentrationand the mean daily calculated nominal concentration. The close agreement between the analytical and nominal'concentrations for each exposure chamber indicates that the test material losses were minimal in the vapor generation and exposure system. Also, the temperature and relative humidity values were compatible between the chambers. Clinical observations: Two clinical findings consistent with the grossly observable treatment-induced methemoglobinemia were (1) dull, dark red eyes and (2) grayish or bluish colored skin of the extremities (cyanosis). The dull, dark red eye color was not very distinctive in the 350 ppm group and was not noticed until after 4 weeks of exposure. However, this eye color was very pronounced in the 1000 ppm group and was noted after the firstexposure and thereafter. The gray color of the hairless skin of the extremities was noted in the 350 ppm rats after 9 weeks of exposure, whereas a more perceptible bluish-gray color was seen in the 1000 ppm group after 4 exposures and continued to the end of the study. These color changes were visible after exposure and were not visible in the morning prior to the 'next exposure, approximately 19 hours later. Swelling in the salivary gland region was found in some rats of the 100 and 350 ppm groups after the 5th and 6th exposures and in the 1000 ppm group after exposures 2 through 6. This observation is consistent with a mild transient viral infection (sialodacryoadenitis) which commonly occurs in this laboratory, since there were no histopathological changes in the salivary glands or vicinity thereof in the interim kill rats. Increased amounts of porphyrin pigment around the nares were observed in the rats of the 1000 ppm exposure group during the 1st and 3rd exposure weeks. This transient clinical sign was possibly another indication of the sialodacryoadenitis infection suggested by the swelling in thesalivary gland region, stated previously, and was not judged to be a treatment-related effect.During this study female rats of the 100, 350, and 1000 ppm exposure groups had an unkempt appearance which was an expression of their general weakened condition secondary to the toxicity of the test material.Body Weights: The animal randomization procedure used in this study did not assure equal group mean body weight at the beginning of the study.Despite the fact that the 1000 ppm female rats weighed statistically significantly less than their controls prior to the start of the study, there was obvious growth retardation of this group along with the males of the 1000 ppm group and the male and female rats of the 350 ppm group. All of these treatment groups had statistically significant body weight decreases when compared to controls during the last month of the study, clearly a treatment-related effect. Group mean body weights for both sexes of the 100 ppm group were comparable to their controls. Organ Weights: No toxicologically significant changes were found in any of these mean weights. However, at both kills the fasted mean body weights of the females of the 350 ppm exposure group and of the males and females of the 1000 ppm exposure group were statistically significantly decreased. Their decreased body weights were responsible for the secondary decreases and increases in the absolute and/or relative weights, respectively, of the liver, kidney, heart, brain, thymus, and testes. No toxicological importance was attributed to the statistically significant increases in the absolute heart weight of the 1000 ppm females of the terminal kill or in the absolute testes weight of the 100 ppm males of the terminal kill.Hematology: Hematology parameters of male and female rats were obtained prior to the interim kill and twice before the terminal kill to examine the reproducibility of the packed cell volume (PCV), red blood cell (RBC), hemoglobin (Hgb), and white blood cell (WBC) results obtained at the first analysis before the terminal kill. Prior to the interim kill the statistically significant increases of the WBC counts of the 1000 ppm exposure group may have been a treatment-related effect, possibly a result of the chronic active inflammation seen histologically in the nasal turbinates. However, due to the variability in the mean WBC counts obtained prior to the terminal kill, this was considered a sporadic finding and of no toxicologic significance.Of the two hematological analyses prior to the terminal kill, the following are the only results considered to have any possible toxicological significance. In the second analysis prior to the terminal kill, the 1000 ppm female rats had a statistically significantly increased PCV with a decreased (though not statistically significant) RBC count. This data may have been indicative of the increased number of reticulocytes present, approximately 4 times greater than the controls. At the interim and second terminal analyses the statistically significantly lowered hemoglobin values of the 1000 ppm male rats was a possible exposure-related effect.In addition, the appearance of reticulocytes and Heinz bodies found prior to the interim and terminal kills showed: the 100 ppm exposure group was comparable to the controls, whereas the 350 ppm and 1000 ppm exposure groups showed increases in these parameters over control. The increased emergence of these reticu10cytes and Heinz bodies was probably associated with the methemoglobinemia observed in these two treatment groups.Urinalysis: All parameters of both time periods were interpreted to be unaffected by exposures to NE.Clinical Chemistry Studies: Statistically significant changes in alkaline phosphatase activity (AP) of the 1000 ppm females and in the serum phosphorus levels of the male and/or female NE exposed rats which occurred at the interim and/or terminal kills were not associated with gross or histopathologic exposure-related changes in livers or kidneys and thus were not diagnostic of adverse treatment-related effects on those tissues.At the terminal kill, the 1000 ppm female rats had statistically significantly increased bilirubin levels which could have been a result of exposure to the test material. At the terminal kill, glucose levels of the male and female rats of the 350 and/or 1000 ppm exposure groups that were statistically significantly decreased were indicative of the poor nutritional state of the rats of these groups, a general response to the toxicity of NE. There was a statistically significant increase in the glucose level of the 350 ppm females of the interim kill. However this was considered to be of no toxicological importance because of its isolated occurrence and lack of dose-response.Methemoglobin: Prior to the interim kill on exposure day 20, experiment day 29, methemoglobin (MetHb) analysis was performed approximately 15 hours after exposure; the MetHb levels of all NE exposed rats were comparable to their controls. However, further MetHb analysis was prompted by the clinical observation of dull dark red eyes and bluish skin of the 1000 ppm rats' extremities which were found only after exposure and would disappear by morning (~l9 hrs later). Because female rats exposed to 1000 ppm NE visually appeared to be affected more than the males, the analysis was performed on the 1000 ppm females and the control group right after exposure. The statistically significantly increased amount of MetHb found in the 1000 ppm females clearly demonstrated that time was the essential factor for the determination of MetHb. Subsequent evaluation of the other exposure groups (both sexes) showed a concentration-related increased amount of MetHb in the male and female rats of all treatment groups. To more completely characterize the time-dependent MetHb effect, a timed sequence of analyses was performed the day before the terminal kill. The determinations were performed. right after exposure (<1/2 hr), 4 and 19 hours later, on rats and mice (only the 1000 ppm NE exposure groups were sampled at approximately 4 hours after exposure). By morning (19 hrs after exposure) the MetHb levels of the 100 and 350 ppm exposure groups of ratswere comparable to the controls, and only the 1000 ppm exposure groups of rats were statistically significantly increased. The data indicate initially higher concentrations of MetHb in rats than in mice, but a faster rate of MetHb disappearance in mice than in rats.Gross Pathology and HistopathologyMany gross pathology and histopathology observations were typical of naturally-occurring spontaneous lesions, conmonly seen in this age and strain of rats and mice used in this study and will not be discussed. Interim Kill-Rats: Treatment-related effects specifically attributed to the test material occurred at all 3 exposure groups which involved either the spleen, nasal turbinates, liver or kidneys. Upon gross examination, the spleen appeared enlarged in a few of the 350 ppm males and 1000 ppm females and in all of the 1000 ppm males. It was also dark colored in all 350 and 1000 ppm males and in most 1000 ppm females. At histopathologic examination, the spleen showed increased congestion of the red pulp in all 350 ppm males and in all 1000 ppm rats as well as an increase in extramedullary hematopoieSis in a few 350 ppm male rats, inmost 1000 ppm female rats and in all 1000 ppm male rats. These alterations in the spleen were considered to be consistent with the methemoglobinemia induced by exposure to NE.Nasal turbinate lesions found only by histopathological inspection were (1) chronic active inflammation in the region of the olfactory epithelium found in one female of the 100 ppm group, all males and a single female of the 350 ppm group, and all rats of the 1000 ppm group, and (2) degeneration in the olfactory epithelium observed in all males and most females of the 350 ppm exposure group and in all of the 1000 ppm group. The degeneration consisted of a flattening of the olfactory mucosal epithelium and intercellular mucosal edema of the olfactory epithelium in the dorsal medial portion of the nasal turbinates. These responses of inflammation and degeneration were the result of exposure to NE.Only histopathological changes were found in the liver and kidneys. The liver effects observed were described as vacuolization, consistent with fatty change, seen in most male rats exposed to 1000 ppm HE. Decreased cytoplasmic granularity of predominantly the proximal tubular cells of the kidneys was observed in a few male rats of the 1000 ppm group. Three findings that were secondary to the toxicity of the test material and occurred only in the 1000 ppm treatment group were (1) a gross observation in two female rats of thickened stomach walls histopathologically described as submucosal gastric edema and was probably the result of not eating and/or stress, (2) a decrease in adipose tissue reserves of both sexes that represented a decreased food intake, and (3) perineal soiling of most females that was viewed as a nonspecific expression of the sickened condition of the animals. There was no effect noted in the salivary gland of female rats exposed to 1000 ppm. In males, the slight effect noted (salivary gland acini vacuolization) was observed in both control and 1000 ppm exposed rats.Terminal Kill-Rats: Exposure associated changes caused directly by NE were found in the spleen, nasal turbinates, salivary glands, liver, or kidneys of all 3 exposure groups. The spleen was grossly observed to be dark in most of the 350 ppm males and dark and enlarged in all male and female rats of the 1000 ppm group. Microscopic changes included (1) an increase in congestion of all male rats exposed to NE and in the majority of the females of the 350 and 1000 ppm groups, and (2) an increase in extramedullary hematopoiesis in all treated males and in a single or a few female rats of each NE exposure group. All of these changes were considered the result of NE induced methemoglobinemia.Treatment-related effects in the nasal turbinates were not observed on gross inspection but were evident upon histopathologic examination. Chronic active inflammation and degeneration were observed in the region of the olfactory epithelium of a few males and females exposed to 350 ppm NE and of most exposed to 1000 ppm NE. Degeneration was characterized by a flattening of the epithelium and intercellular mucosal edema of the olfactory region in the dorsal medial portion of the nasal turbinates and was moderate in severity in rats exposed to 1000 ppm but slight in those at 350 ppm.Salivary gland alterations associated with treatment were limited to histopathologic observations. A decreased cytoplasmic granularity and decreased eosinophilic staining of the ductal epithelial cells was seen in all treated male and female rats. The severity varied from very slight in rats exposed to 100 ppm to slight in those exposed to 350 and 1000 ppm. In addition. vacuolization of the salivary gland acini was seen in most females exposed to 1000 ppm of NE.Histopathologic changes in the liver were characterized by vacuolization consistent with fatty change and were present in rats of all exposure levels excluding 100 ppm males and control females. The severity of the vacuolization varied from very slight in the control males to slight in the others. This observation was believed to be related to exposure to NE. Several alterations specific to the rats of the 1000 ppm exposure group occurred in the kidney, thymus, adipose tissue, and haircoat. Renal changes consisted of a dark appearance upon gross inspection of a few males. Histologically the kidney lesions were observed as a decreased cytoplasmic granularity of the renal cortical (predominantly proximal) tubular epithelial cells of all males in the 1000 ppm exposure group. Such changes were the result of treatment. Decreased thymic size was noted at necropsy in a few males of the 1000 ppm group but no microscopic alterations were recognized. thus this was judged to be a secondary (nonspecific) treatment-related effect. The decrease in adipose tissue in a few of. the 1000 ppm males and the perineal soiling of a few 1000 ppm females were also considered to be nonspecific changes secondary to the toxicity of the test material.

Effect levels

open allclose all
Dose descriptor:
NOAEC
Effect level:
100 ppm
Based on:
test mat.
Sex:
male
Basis for effect level:
other: Local effects; Histopathologic changes noted in nasal turbinates following 6 hr/day, 5 days/week for 13 weeks exposure to 350 ppm
Dose descriptor:
NOAEC
Effect level:
ca. 100 ppm
Based on:
test mat.
Sex:
female
Basis for effect level:
other: Local effects; Histopathologic changes noted in one of 10 nasal turbinates following 6 hr/day, 5 days/week for 4 weeks but not 13 weeks exposure to 100 ppm
Dose descriptor:
LOAEC
Effect level:
100 ppm
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: Systemic effects; Histopathologic changes noted in salivary gland following 6 hr/day, 5 days/week for 13 weeks exposure to 100 ppm.

Target system / organ toxicity

Critical effects observed:
not specified

Any other information on results incl. tables

Table 1 Mean Methemoglobin (MetHb) Values for Rats Prior to the 93-Day Terminal Kill 13-Week Study

 Exposure          % MetHb (Mean + S.D.)
 Level, ppm  N  Immediately After Last (64th) Exposure  4 hrs After Last Exposure  19 hrs After Last Exposure
 Males        
 0  5  0.4 + 0.4  N.D.  0.5 + 0.3
 100  5  2.4 + 0.5  N.D.  0.4 + 0.3
 350  5  12.9 + 1.5*  N.D.  0.6 + 0.2
 1000  5  50.7 + 5.4*  58.6 + 6.1 a  1.5 + 0.8*
 Females        
 0  5  0.5 + 0.3  N.D.  0.5 + 0.3
 100  5  5.3 + 1.7  N.D.  0.8 + 0.8
 350  5  30.7 + 3.9*  N.D.  0.8 + 0.5
 1000  5  61.8 + 6.0*  64.1 + 4.6a  1.9 + 0.3

*Statistically significant deviation from control mean using Dunnett's test, p <0.05.

aNo statistical analysis was performed on this data since no control data was determined at this time period.

Applicant's summary and conclusion

Conclusions:
Exposure of rats to 1000 ppm NE resulted in decreased body weight gain, elevated MetHb levels with cyanosis, increased reticulocytes and Heinz bodies in peripheral blood and associated splenic congestion and extramedullary hematopoiesis. Other major target organ effects in these animals included degenerative and inflammatory changes in the olfactory nasal epithelium, hepatocellular vacuolization, decreased cytoplasmic granularity of renal cortical tubular epithelium and ductal epithelial cells of the salivary glands. Rats exposed to 350 ppm NE showed similar but less severe changes in MetHb, spleen, nasal turbinates and salivary glands. Minimal changes in MetHb, spleen and salivary glands were observed in rats :xposed to 100 ppm.
Executive summary:

The subchronic toxicity of nitroethane was examined in rats. Groups of rats were exposed to 0, 100, 350, or 1000 ppm (0, 0.3, 1.0 or 3.0 mg/l) of NE for 6 hr/day, 5 days/wk for a total of 64-65 exposures with an interim sacrifice of rats after 20-21 exposures. Parameters monitored were clinical observations, body weights, organ weights, hematologic characteristics including methemoglobin (MetHb) determination, clinical chemistries, urinalysis, gross pathology and histopathology.

Exposure of rats to 1000 ppm NE resulted in decreased body weight gain, elevated MetHb levels with cyanosis, increased reticulocytes and Heinz bodies in peripheral blood and associated splenic congestion and extramedullary hematopoiesis. Other major target organ effects in these animals included degenerative and inflammatory changes in the olfactory nasal epithelium, hepatocellular vacuolization, decreased cytoplasmic granularity of renal cortical tubular epithelium and ductal epithelial cells of the salivary glands. Rats exposed to 350 ppm NE showed similar but less severe changes in MetHb, spleen, nasal turbinates and salivary glands. Minimal changes in MetHb, spleen and salivary glands were observed in rats exposed to 100 ppm.