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Repeated dose toxicity - oral: A Combined Repeated Dose Toxicity Study With the Reproduction/Developmental Toxicity Screening Test (oral gavage in rats) on potassium nitrate from Product Safety Laboratories, 2002 provide a NOAEL of 1500 mg/kg/day 
Repeated dose toxicity - inhalation: A repeated dose inhalation toxicity study for nitrogen dioxide gas reported a NOAEC of ≥ 2.15 ppm (4.11 mg/m^3).
Repeated dose toxicity - dermal: No data

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records
Reference
Endpoint:
short-term repeated dose toxicity: oral
Remarks:
combined repeated dose and reproduction / developmental screening
Type of information:
other: Experimental study of a supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
From 2002-01-08 to 2002-02-23
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
Well documented GLP study. Study was performed according to guideine OECD 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test) with minor deviations (only 5 animals per sex tested instead of 10). The read-across rationale can be found in the category approach document attached in Section 13.
Reason / purpose for cross-reference:
reference to same study
Qualifier:
according to guideline
Guideline:
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
Deviations:
yes
Remarks:
5 animals of each sex instead of 10 animlas of each sex tested
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Raleigh, NC
- Age at study initiation: males were 64 days of age on day of arrival; females were 61 days of age on day of arrival
- Weight at study initiation: 225 - 325 grams for male rats; 161-219 for female rats
- Fasting period before study: no data
- Housing: Animals were individually housed except for during the cohabitation and lactation period in wire mesh suspended stainless steel cages which conformed with GLP requirements. During cohabitation each pair of rats were housed in the male rat's cage. Beginning no later than Day 20 of gestation, female rats were individually housed in polyethylene shoebox cages containing nesting material with wire mesh lids. Each dam and litter was housed in a common nesting box during the lactation/postnatal period
- Use of restrainers for preventing ingestion (if dermal): not applicable
- Diet (e.g. ad libitum): Purina Certified Rodent Diet #5002; ad libitum
- Water (e.g. ad libitum): automatic dispenser; ad libitum and when females and litters were housed in shoebox cages via water bottle; ad libitum
- Acclimation period: 12 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18 - 22 °C
- Humidity (%): 43-66%
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): 12 hours dark/12 hours light

IN-LIFE DATES: From: 2002-01-08 To: 2002-02-23
Route of administration:
oral: gavage
Vehicle:
other: distilled water
Details on oral exposure:
Dose calculations: Individual doses were calculated based on the most recent weekly body weights and were adjusted each week to maintain targeted dose level for all rats in the General Toxicity groups (i.e., mg/kg/day). All doses were administered by volume of 10 mL/kg after correcting for concentration of the test mixture. Control animals received the vehicle only at the same volume as the test groups.
Dose preparations: The test substance (011101-3D) was ground in a Krups coffee mill (Model 203) prior to use and again upon receipt of additional test substance (020122-1D). A quantity sufficient to cover the grinding blade was added to the coffee mill and ground to a fine powder. Appropriate amounts of ground test material were accurately weighed into a 100 mL volumetric flask and diluted to volume with distilled water for each of the low, mid and high concentrations. Given that there was visual evidence (i.e. settling of test substance to bottom of cup) of a small amount of precipitate, the dosing mixtures were constantly stirred on a magnetic stirr plate while being sampled to dose the test animals during the study.

DIET PREPARATION:
- Rate of preparation of diet (frequency): not applicable, route of administration of test substance is via gavage
- Mixing appropriate amounts with (Type of food): not applicable
- storage temperature of food: no data
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The test substance was assumed to be homogenous and stable at the time drawn by syringe to dose the test animals. Analysis of dosing mixtures, therefore, were limited to concentration verification of representative preparations intended for the control, low, intermediate and high dose levels in the study. Representative dosing mixtures of each concentration during the study were provided to the analytical department at three time points during the study prior to animal exposure, near the middle (test days 24 and 28) and near the end of the study (test day 45). Vehicle control samples were inadvertently not submitted for analysis. Each dose preparation was evaluated by flame atomic absorption spectroscopy for total potassium (SOAC Official Method 975.03) (1988). A reference standard of potassium (999 µg/mL), supplied by EM Science, was used for calibration.
Duration of treatment / exposure:
daily
Dose frequency: each animal was dosed by oral intubation using a stainless steel ball-tipped gavage needle attached to an appropriate syringe. Dose administration was daily (7 days/week) for all adult animals as follows:
Male: general toxicity groups: for at least 28 days of exposure
Female rats: for at least 28 days of exposure
Frequency of treatment:
daily
Remarks:
Doses / Concentrations:
0, 250, 750 and 1500 mg/kg/day
Basis:
other: Doses were selected based on parameters assessed in a range-finding study at concentrations up to 1000 mg/kg/day)
No. of animals per sex per dose:
5 males and 5 females
Control animals:
other: yes, but no data mentioned
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Twice daily for viability and cage-side observations were performed daily during acclimation, premating and mating, gestation, and lactation periods, except when scheduled detailed observations were conducted. All observations were recorded.


DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Observations were performed and recorded at least once during the acclimation period for all male and female rats. Observations were performed and recorded approximately once per week during the premating and mating periods for females of the reproduction groups during the gestional days (GD7, GD14 and GD20) and lactational (LD4 only) periods. Female rats were evaluated for adverse clinical signs during parturition. Maternal behaviour was checked on LD0 and LD4 and recorded. The date and clock time of all observations and/or mortality was recorded.

BODY WEIGHT: Yes
- Time schedule for examinations: Individual body weights were recorded at least twice during the acclimation period (including the day after receipt) before pairing and mating. All male rats were weighed weekly during the premating and mating periods and at the time of sacrifice. Mated females were weighted on GD0, 7, 14 and 20, and on the day of delivery (LD0) and LD4 (prior to terminal sacrifice). Females showing no evidence of mating were assigned a GD0 after cessation of cohabitation and body weights were measured accordingly. Females in the general toxicity groups were weighed weekly and at the time of sacrifice. Body weight gains were calculated for males and females during each appropriate interval.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
Although not a feeding study, food consumption was determined weekly during the premating period (no mating period) for all males and females. Individual food consumption was measured and recorded weekly thereafter for the females in the general toxicity groups and during the gestational period for the females in the reproductive groups. Food consumption was also recorded on LD0 and LD4. The data were then used to calculate food efficiency for the associated intervals.

FOOD EFFICIENCY:
Data from food consumption were used to determine food efficiency for associated intervals.

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No data

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: See detailed clinical observations
- Dose groups that were examined: No data

HAEMATOLOGY: Yes (collected via orbital sinus bleeding)
- Time schedule for collection of blood: Day 28 of treatment
- Anaesthetic used for blood collection: Yes: Isoflourane anesthesia
- Animals fasted: Yes: 18 hours prior to blood collection
- How many animals: 5 males and 5 females/dose level
- Parameters checked: hematocrit, hemoglobin concentration, erythrocyte count, total and differential leukocyte count, platelet count, prothrombin time and activated partial thromboplastin time.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: on Day 28 of treatment
- Animals fasted: Yes: 18 hours prior to blood collection
- How many animals: 5 males and 5 females/dose level
- Parameters checked: calcium, phosphorus, chloride, sodium, potassium, fasting glucose, serum alanine aminotransferase (SGPT), serum aspartate aminotransferase (SGOT), gamma glutamyl transpeptidase, urea nitrogen, albumin, blood creatinine, total bilirubin, total serum protein, globulin, total cholesterol, alkaline phosphatase and magnesium measurements..

URINALYSIS: No data
- Time schedule for collection of urine: No data
- Metabolism cages used for collection of urine: No data
- Animals fasted: No data
- Parameters checked in table: No data

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: during the final days of treatment
- Dose groups that were examined: Five male and females/dose group (including controls)
- Battery of functions tested: sensory activity / grip strength / motor activity: excitability, autonomic function, gait and sensorimotor coordination (open field and manipulative evaluations), reactivity and sensitivity (elicited behaviour) and other abnormal clinical signs including but not limited to convulsions, tremors, unusual or bizarre behaviour, emaciation, dehydration, and general appearance. The rats were observed in random without the observer aware of the dose group. Motor activity was also evaluated. Each animal was evaluated for a single one-hour phase, with photobeam counts accumulated over six, 10-minute intervals. Total movements (consisting of fine and active movements) were considered an appropriate measure for the assessment of potential behaviour effects in this screening level study).
Sacrifice and pathology:
GROSS PATHOLOGY: Yes: Necropsy: gross necropsy of all males and females included an initial examination of external surfaces and orifices, as well as the cranial, thoracic and abdominal cavities and their contents. Rats were examined for gross lesions. Gross lesions were retained in neutral buffered 10% formalin (NBF). At scheduled sacrifice, all survivors were euthanized by exsanguination from the abdominal aorta under isoflourane anesthesia. All animals were subjected to a full necropsy that included examination of the external surface of the body, all orifices and the thoracic, abdominal and cranial cavities and their contents. The liver, kidneys, adrenals, brain, heart, thymus, spleen, ovaries, testes and epididymides (of all animals sacrificed by design) were weiged wet as soon as possible after dissection to avoid drying. The following organs and tissues from all animals were preserved in NBF for possible future histopathological examination: all gross lesions, lungs, brain- including sections of the medulla/pons, cerebellar cortex and cerebral cortex, spinal cord (3 levels: cervical, mid-thoracic, and lumbar), eyes, pituitary, thyroid/parathyroid, thymus, trachea, heart, sternum with bone marrow, salivary glands, liver, spleen, kidneys, adrenals, pancreas, ovaries, testes, uterus (with attached urinary bladder, cervix and vagina), accessory sex organs (epididymides, prostate, and seminal vesicles), female mammary gland, skin, aorta, esophagus, stomach, duodenum, jejunum, ileum, cecum, colon, rectum, representative lymph node, and peripheral nerve (sciatic).

HISTOPATHOLOGY: Yes: histopathological examination was performed on the preserved organs and tissues of the reproductive and general toxicity group animals from the control (groups I and II) and high dose (groups VII and VIII). In addition, gross lesions of potential toxicological significance noted in any test groups were also examined. Microscopic findings were graded.
Statistics:
Mean and standard deviations were calculated for all quantitative data. Except for clinical pathology data were the contract laboratory, Huntingdon Life Sciences, elected to use statistics to aid in the data interpretation; no further statistical treatment of the study was conducted due to small group sizes.
Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
no effects observed
Food efficiency:
no effects observed
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
no effects observed
Haematological findings:
no effects observed
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
not specified
Behaviour (functional findings):
no effects observed
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
no effects observed
Details on results:
CLINICAL SIGNS AND MORTALITY: there were no treatment-related deaths and no signs of overt clinical toxicity

BODY WEIGHT AND WEIGHT GAIN: there were no effects on body weight, food consumption, or food efficiency.

FOOD CONSUMPTION AND COMPOUND INTAKE: There were no effects of test-substance treatment on food consumption in males. There were no effects of food consumption on females during pre-mating; during weeks 3, 4 and 5 for females in the General Toxicity Group; or during gestation and lactation. Food consumption was not measured during the mating period.

FOOD EFFICIENCY: Food efficiency was also unaffected by treatment.

WATER CONSUMPTION AND COMPOUND INTAKE: not applicable

OPHTHALMOSCOPIC EXAMINATION: no effects

HAEMATOLOGY: No test substance related haematological changes were associated with the test substance treatment

CLINICAL CHEMISTRY: A slight increase in blood urea nitrogen was observed in male and female rats at 1500 mg/kg/day and in female rats at 750 mg/kg/day. Although outside the range of the historical control, the absence of other indicators or renal dysfunction (e.g., creatinine) discounted the clinical siginificance of this endpoint. Minimal changes in electrolyte levels in male rats (e.g., 10% decrease in potassium, 4% decrease in calcium, and 22% increase in phosphorus) and female rats (3% decrease in chloride, 4% decrease in magnesium) were observed at 1500 mg/kg/day. These changes were within the range of historical control and were not considered to be of biological significance.

URINALYSIS: no data

NEUROBEHAVIOUR: Functional observational battery (FOB) and motor activity tests identified no treatment-related changes in behaviour, function, or motor activity

ORGAN WEIGHTS: Mean organ weights and organ-to-body weight ratios for both the reproduction and general toxicity test groups, in general, were considered comparable to their respective control groups. Any slight increases or decreases from the control were incidental, not dose-related and judged not to be of toxicological importance.

GROSS PATHOLOGY: There were a number of gross observations correlated to microscopic findings. The dilatation of the uterus (horns) observed in several female rats from the general toxicology group (group II - animal #8133 and 8134, group VIII - animal #8205, 8206, 8207 and 8208) was considered to be a function of the estrus stage (generally proestrus, but sometimes early estrus). These gross observations and others, along with their microscopic correlates, were all considered identical background findings not attributable to administration of the test substance.

HISTOPATHOLOGY: No treatment-related histopathological changes were reported.

HISTORICAL CONTROL DATA (if applicable): no data
Dose descriptor:
NOAEL
Effect level:
1 500 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: No adverse effects were seen on general toxicity endpoints.
Critical effects observed:
not specified

NOAEL: 1500 mg/kg/day (general toxicity)

LOAEL: No adverse effects were seen on general toxicity endpoints.

Conclusions:
A NOAEL of 1500 mg/kg/day for general toxicity was derived after exposure to potassium nitrate. NO adverse effects were seen on general toxicity endpoints.
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
Study duration:
subacute
Species:
rat
Quality of whole database:
The study has a klimisch code 2.

Repeated dose toxicity: inhalation - systemic effects

Link to relevant study records
Reference
Endpoint:
sub-chronic toxicity: inhalation
Type of information:
other: Experimental study of a supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: GLP and guideline study nitrogen dioxide. The read-across rationale can be found in the category approach document attached in Section 13.
Justification for type of information:
As the target substance is a nitrate, read-across can be applied to other substances from the nitrate group. The read-across rationale can be found in the category approach document attached under 'Attached justification', in Section 13 of IUCLID and the CSR.
Qualifier:
according to guideline
Guideline:
OECD Guideline 413 (Subchronic Inhalation Toxicity: 90-Day Study)
Qualifier:
according to guideline
Guideline:
OECD Guideline 412 (Subacute Inhalation Toxicity: 28-Day Study)
GLP compliance:
yes (incl. QA statement)
Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals or test system and environmental conditions:
About 7 weeks old male and female rats were used. The animals were identified individually by tattooing the
respective animal number into the ears. The female animals were nulliparous and non-pregnant. Only animals free
from clinical signs of disease were used for the study.
During the period when the rats were not exposed they were housed singly in wire cages. Underneath the cages,
waste trays were fixed containing bedding material. The animals were kept in fully air-conditioned rooms in which
a temperature in the range of 20 - 24°C and relativ e humidity in the range of 30 - 70% were ensured by means of a
central air-conditioning system. A light/dark rhythm of 12 hours was maintained (12 h dark, 12 h light). The animals
were maintained on milled mouse/rat laboratory diet and tap water ad libitum.
Route of administration:
inhalation: gas
Type of inhalation exposure:
whole body
Duration of treatment / exposure:
90 days
Frequency of treatment:
6 hours a day on 5 days per week for 13 weeks (65 exposures)
Remarks:
Doses / Concentrations:
0.1, 0.5 and 1 ppm NO2 (= 0.19 mg/m3, 0.96 mg/m3, 1.91 mg/m3); infrared photometric measurement resulted in measured concentrations of 0.25, 0.82 and 2.15 ppm NO2
Basis:

No. of animals per sex per dose:
groups of 15 male and 10 female rats
Control animals:
other: yes; exposed to clean air
Observations and examinations performed and frequency:
During the whole study, the general state of health was controlled twice on working days and once on weekends or holidays. On exposure days clinical observation was performed before, during and after exposure. During preflow period and on post exposure day clinical findings were recorded once each working day. Body weight of the animals was determined.
Lung lavage was performed in 5 male animals from each concentration group (groups 01 – 31). Bronchoalveolar Lavage Fluid (BALF) was evaluated biochemically and cytologically. Moreover, in 10 male and 10 female animals of each concentration group (groups 0 – 3) BrdU minipumps were impanted, pulmonary cell proliferation and apoptose rates were determined. In these animals, clinical chemical and hematological examination of the blood,
as well as a complete necropsy including gross pathological evaluation and weighing of selected organs were performed. The respiratory tract and the mediatitinal lymph nodes of these animals were examined histopathologically.
Statistics:
Means and standard deviations of each test group were calculated for several parameters.
Details on results:
During the whole study period the animals showed no clinical signs and findings different from normal. Concerning the mean body weights, body weight changes, food consumption and food efficiency, there were no statistical significant differences between the exposed animals and the air controls. Clinical pathology examinations revealed no treatment-related changes in the hematology, clinical chemistry and bronchoalveolar lavage parameters.
Regarding pathology, a minimal increase of alveolar histiocytosis in only 3 of 10 test animals of the male group 3 animals exposed to the target concentration of 1 ppm was detected. This may be interpreted as a first indication of a compound-related irritation, but further morphological alterations of the lung parenchyma are missing. Especially,
no biological relevant increases of cell proliferation (BrdU-stain) or apoptosis (TUNEL-stain) could be detected or any kind of significant organ weight changes. Therefore, the few single cases of a minimal (grade 1) alveolar histiocytosis are regarded to be an only incidental increase and of spontaneous origin.
Dose descriptor:
NOAEC
Effect level:
2.15 ppm
Sex:
male/female
Basis for effect level:
other: The inhalation of measured concentrations of up to 2.15 ppm NO2 by male and female rats did not cause any treatment-related findings in all examined parameters.
Critical effects observed:
not specified
Conclusions:
The inhalation of measured concentrations of up to 2.15 ppm NO2 by male and female rats did not cause any treatment-related findings in all examined parameters including biochemical and cytological examinations of the broncho alveolar lavage fluide, hematological and clinical chemical examination of the blood, histopathological examination of the whole respiratory tract, cell proliferation and apoptosis rate in the lung.
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEC
4.11 mg/m³
Study duration:
subchronic
Species:
rat
Quality of whole database:
The study has a klimisch code 2.

Repeated dose toxicity: inhalation - local effects

Link to relevant study records
Reference
Endpoint:
sub-chronic toxicity: inhalation
Type of information:
other: Experimental study of a supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: GLP and guideline study nitrogen dioxide. The read-across rationale can be found in the category approach document attached in Section 13.
Justification for type of information:
As the target substance is a nitrate, read-across can be applied to other substances from the nitrate group. The read-across rationale can be found in the category approach document attached under 'Attached justification', in Section 13 of IUCLID and the CSR.
Qualifier:
according to guideline
Guideline:
OECD Guideline 413 (Subchronic Inhalation Toxicity: 90-Day Study)
Qualifier:
according to guideline
Guideline:
OECD Guideline 412 (Subacute Inhalation Toxicity: 28-Day Study)
GLP compliance:
yes (incl. QA statement)
Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals or test system and environmental conditions:
About 7 weeks old male and female rats were used. The animals were identified individually by tattooing the
respective animal number into the ears. The female animals were nulliparous and non-pregnant. Only animals free
from clinical signs of disease were used for the study.
During the period when the rats were not exposed they were housed singly in wire cages. Underneath the cages,
waste trays were fixed containing bedding material. The animals were kept in fully air-conditioned rooms in which
a temperature in the range of 20 - 24°C and relativ e humidity in the range of 30 - 70% were ensured by means of a
central air-conditioning system. A light/dark rhythm of 12 hours was maintained (12 h dark, 12 h light). The animals
were maintained on milled mouse/rat laboratory diet and tap water ad libitum.
Route of administration:
inhalation: gas
Type of inhalation exposure:
whole body
Duration of treatment / exposure:
90 days
Frequency of treatment:
6 hours a day on 5 days per week for 13 weeks (65 exposures)
Remarks:
Doses / Concentrations:
0.1, 0.5 and 1 ppm NO2 (= 0.19 mg/m3, 0.96 mg/m3, 1.91 mg/m3); infrared photometric measurement resulted in measured concentrations of 0.25, 0.82 and 2.15 ppm NO2
Basis:

No. of animals per sex per dose:
groups of 15 male and 10 female rats
Control animals:
other: yes; exposed to clean air
Observations and examinations performed and frequency:
During the whole study, the general state of health was controlled twice on working days and once on weekends or holidays. On exposure days clinical observation was performed before, during and after exposure. During preflow period and on post exposure day clinical findings were recorded once each working day. Body weight of the animals was determined.
Lung lavage was performed in 5 male animals from each concentration group (groups 01 – 31). Bronchoalveolar Lavage Fluid (BALF) was evaluated biochemically and cytologically. Moreover, in 10 male and 10 female animals of each concentration group (groups 0 – 3) BrdU minipumps were impanted, pulmonary cell proliferation and apoptose rates were determined. In these animals, clinical chemical and hematological examination of the blood,
as well as a complete necropsy including gross pathological evaluation and weighing of selected organs were performed. The respiratory tract and the mediatitinal lymph nodes of these animals were examined histopathologically.
Statistics:
Means and standard deviations of each test group were calculated for several parameters.
Details on results:
During the whole study period the animals showed no clinical signs and findings different from normal. Concerning the mean body weights, body weight changes, food consumption and food efficiency, there were no statistical significant differences between the exposed animals and the air controls. Clinical pathology examinations revealed no treatment-related changes in the hematology, clinical chemistry and bronchoalveolar lavage parameters.
Regarding pathology, a minimal increase of alveolar histiocytosis in only 3 of 10 test animals of the male group 3 animals exposed to the target concentration of 1 ppm was detected. This may be interpreted as a first indication of a compound-related irritation, but further morphological alterations of the lung parenchyma are missing. Especially,
no biological relevant increases of cell proliferation (BrdU-stain) or apoptosis (TUNEL-stain) could be detected or any kind of significant organ weight changes. Therefore, the few single cases of a minimal (grade 1) alveolar histiocytosis are regarded to be an only incidental increase and of spontaneous origin.
Dose descriptor:
NOAEC
Effect level:
2.15 ppm
Sex:
male/female
Basis for effect level:
other: The inhalation of measured concentrations of up to 2.15 ppm NO2 by male and female rats did not cause any treatment-related findings in all examined parameters.
Critical effects observed:
not specified
Conclusions:
The inhalation of measured concentrations of up to 2.15 ppm NO2 by male and female rats did not cause any treatment-related findings in all examined parameters including biochemical and cytological examinations of the broncho alveolar lavage fluide, hematological and clinical chemical examination of the blood, histopathological examination of the whole respiratory tract, cell proliferation and apoptosis rate in the lung.
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEC
4.11 mg/m³
Study duration:
subchronic
Species:
rat
Quality of whole database:
The study has a klimisch code 2.

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Repeated dose toxicity - oral: A reliable study on potassium nitrate was identified: Product Safety Laboratories. 2002. Potassium Nitrate: Combined Repeated Dose Toxicity Study With the Reproduction/Developmental Toxicity Screening Test (Oral Gavage Study in Rats). Study Number 11686 – Owned by TFI. A NOAEL of 1500 mg/kg/day was identified from that study. This oral study on potassium nitrate shows that nitrate ions are not causing effects both systemically and locally. Effects caused by nitric acid after oral exposure will be due to the H+ ions, rather than due to the nitrate ions.But with the introductory sections to Annexes VII-X point at a specific adaptation to the standard information requirements as in vivo testing shall be avoided with corrosive substances at concentration/dose levels causing corrosivity. At low concentration, nitric acid as soon as entering the body is distributed widely as nitrate, which is an essential element to organisms and its fate in the human body is regulated. The H+ ions (if present at low concentration) will also be internally buffered/regulated.

Repeated dose toxicity - Inhalation: No repeated dose inhalation study following OECD guidelines is available on nitric acid. A reliable NOAEC for repeated dose inhalation was not identified for nitric acid based on available information. Nitric acid decomposes to nitrogen dioxide (NO2) so exposure to nitric acid includes co-exposure to NO2. Some of the studies reported their exposure doses in NO2 concentrations or conducted studies directly with NO2 (which in turns hydrolyzes to give nitric acid) . Limited animal studies (dogs, rats, rabbit) exist on nitric acid exposed to a wide range of concentrations. These studies primarily focused on non-OECD standard endpoints, and the majority of the studies used one dose level and therefore a definitive and reliable nitric acid NOAEC could not be determined.

One of the few studies that exposed animals to multiple concentrations of nitric acid was a rabbit study (Schlesinger et al, 1994) that exposed them to nominal concentrations of 50, 150 or 450 ug/m3 for 4 h/day, 3 days/week for 4 weeks. The study identified a LOAEC of ≤50 ug/m3 based on a significant reduction of superoxide levels; however at this concentration no changes were observed in the total number of cells recovered by lavage, differential count, cell viability, LDH or total soluble protein in lavage fluid, and no indication of airway hyporesponsivity compared to control.

In a 90-day inhalation study (BASF, 2006) rats were exposed to NO2 concentrations of 0.25, 0.82 or 2.15 ppm for 6-hours a day on 5 days per week for 13 weeks. A NOAEC of ≥ 2.15 ppm (4.11 mg/m3) was identified based on no clinical signs and findings different from normal during the whole study period. Concerning the mean body weights, body weight changes, food consumption and food efficiency, there were no statistical significant differences between the exposed animals and the air controls. Clinical pathology examinations revealed no treatment-related changes in the hematology, clinical chemistry and bronchoalveolar lavage parameters.

Repeated dose toxicity - Dermal: Testing is waived based on the following justification: studies are available for the inhalation route of exposure on nitric acid and for the oral route of exposure on potassium nitrate. According to the REACH Regulation, only one route of exposure should be tested for repeated dose toxicity (column 2 adaptation, annex VIII, section 8.6.1). Therefore, it is not necessary to perform a repeated dose toxicity study via the dermal route of exposure.


Justification for selection of repeated dose toxicity via oral route - systemic effects endpoint:
Reliable study with potassium nitrate for oral repeated dose toxicity is available. The read-across rationale can be found in the CSR document attached in Section 13.

Justification for selection of repeated dose toxicity inhalation - systemic effects endpoint:
Reliable study with nitrogen dioxide for inhalation repeated dose toxicity is available. The read-across rationale can be found in the discussion below.

Justification for selection of repeated dose toxicity inhalation - local effects endpoint:
Reliable study with nitrogen dioxide for inhalation repeated dose toxicity is available. The read-across rationale can be found in the discussion below.

Justification for classification or non-classification

No repeated dose inhalation studies conducted with standard methodology were available for nitric acid. One study of sufficient quality identified a LOAEC of ≤50 ug/m3 for nitric acid in rabbits based on a significant reduction of superoxide levels; however, at this concentration no changes were observed in the total number of cells recovered by lavage, differential count, cell viability, LDH or total soluble protein in lavage fluid, and no indication of airway hyporesponsivity compared to control. In a 90-day inhalation study with NO2 a NOAEC of ≥ 2.15 ppm (4.11 mg/m3) in rats was identified based on no clinical signs and findings different from normal during the whole study period. Clinical pathology examinations revealed no treatment-related changes in the hematology, clinical chemistry and bronchoalveolar lavage parameters. Nitric acid has been identified as corrosive, which could lead to possible future systemic effects. However, no evidence of this was identified in the repeated dose studies. Therefore, no classification can be made due to inconclusive data.