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Toxicological information

Acute Toxicity: inhalation

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Administrative data

Endpoint:
acute toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2019
Report Date:
2019

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 403 (Acute Inhalation Toxicity)
Deviations:
yes
Remarks:
Mean relative humidity in the exposure chamber (28.3%) was slightly below the target range. No impact on the quality of the study
GLP compliance:
yes (incl. certificate)
Test type:
traditional method
Limit test:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Type:
Constituent
Test material form:
solid: particulate/powder
Remarks:
powder
Details on test material:
white or almost white powder

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
- Age: 10 weeks
- Animals per cage: 3 of the same sex
- Change of cages: At least once a week and when deemed necessary throughout the study period
- Inclusion criteria on arrival: Veterinary inspection
- Acclimatisation period: Animals were acclimatised to Vivotecnia housing facilities 20 days prior exposure. Acclimatisation to the nose-only restraining tubes was performed for at least 30 minutes the day of exposure.
- Animal identification: Digit ink marks
- Health status: Specific Pathogen Free (SPF). Females will be nulliparous and non-pregnant
- Light cycle: 12h light : 12h dark
- Temperature: 19.3 – 23.9 ºC
- Relative humidity: 30 – 66 %
- Food: Global diet
- Reference: 2914C
- Supplier: Envigo
- Nutritional / contaminant contents: Certificate of analysis for the batch used in the study is included in Annex III
- Food availability: Ad libitum (animals were deprived of food during exposure)
- Drinking water: Tap water
- Watering: Bottles
- Water availability: Ad libitum (animals were deprived of water during exposure)

Administration / exposure

Route of administration:
inhalation: aerosol
Type of inhalation exposure:
nose only
Vehicle:
air
Remarks:
filtered air
Mass median aerodynamic diameter (MMAD):
>= 3.25 - <= 3.35 µm
Geometric standard deviation (GSD):
>= 2.11 - <= 2.7
Remark on MMAD/GSD:
A respirable aerosol (MMAD in the range of 1-4 μm) could be achieved at the technical limit.
Analytical verification of test atmosphere concentrations:
yes
Duration of exposure:
4 h
Concentrations:
4.42 mg/L (technical limit)
No. of animals per sex per dose:
3 males, 3 females
Control animals:
no
Details on study design:
Please see "Any other information on materials and methods incl. tables"
Statistics:
No statistical analysis was required.

Results and discussion

Effect levels
Key result
Sex:
male/female
Dose descriptor:
LC50
Effect level:
> 4.42 mg/L air (analytical)
Based on:
test mat.
Exp. duration:
4 h
Remarks on result:
not determinable due to absence of adverse toxic effects
Mortality:
One female animal (ID 6) was found dead the day of exposure (study day 1). No relevant findings were observed upon gross necropsy.
Clinical signs:
other: All other animals, regardless of sex, presented wet and dirty fur, chromorrhinorrhea, chromodacryorrhea as well as piloerection immediately after finishing the exposure period. Dirty fur and piloerection lasted until up to day 5 and day 8 of study for mal
Body weight:
A decrease in mean body weight of approximately 6% in males and 7% in females was observed between study day 1 (exposure) and study day 2.
From study day 2 to the end of the observation period (day 15), body weight increased gradually in all animals with the exception of the male ID 2 in which a small decrease (2.7 g, 0.8% ) was observed between study days 2 and 4.
Total mean body weight gains of approximately 6% and 13% were recorded for males and females, respectively, during the 14 day observation period (from study day 1 to study day 15).
Gross pathology:
No relevant macroscopic findings were observed in any of the animals.

Applicant's summary and conclusion

Interpretation of results:
Category 4 based on GHS criteria
Conclusions:
It can be concluded that, under the present experimental conditions:
- The exposure of male and female Sprague Dawley rats to the test item NICOTINE BITARTRATE DIHYDRATE for 4 hours resulted in the premature death of one female animal.
- The marginal body weight loss between study days 1 and 2 in all animals was considered to be mainly due to the stress related to the nose-only exposure procedure, although a contribution of the treatment with the test item cannot be excluded.
- The LC50 of NICOTINE BITARTRATE DIHYDRATE was greater than 4.42 mg/L air (gravimetric aerosol concentration). The test was performed at the technical limit concentration.
- Based on the GHS classification criteria, NICOTINE BITARTRATE DIHYDRATE can be considered to be under human health hazard Category 4 (“harmful if inhaled”) or higher.
Executive summary:

The present study has been designed to evaluate the acute inhalation toxicity of the test item NICOTINE BITARTRATE DIHYDRATE in male and female Sprague Dawley rats by the traditional protocol described in the OECD Guideline Nº 403: Acute Inhalation Toxicity. This method provides lethality data of the test item and allows its classification according to the Globally Harmonized System (GHS) of Classification and Labelling of Chemicals.

For that purpose, a group of three male and three female Sprague Dawley rats was exposed by nose-only, flow-past inhalation to NICOTINE BITARTRATE DIHYDRATE at a mean concentration of 4.42 mg/L air during 4 hours as a limit test.

All animals were observed for clinical signs and mortality during the exposure and the subsequent 14-day observation period. Body weight was recorded just before starting exposure, 24 h, 72 h and one week thereafter and also before sacrifice two weeks after exposure.

At the end of the observation period, all surviving animals were subjected to a gross necropsy and all macroscopic abnormalities were recorded.

The ranges of aerosol concentration, temperature, relative humidity and air flow rate were considered satisfactory for a study of this type. In addition, the test item was considered to be respirable to rats.

One female animal (ID 6) was found dead during exposure (study day 1). Upon gross necropsy, no relevant findings were observed.

All other animals, regardless of sex, presented wet and dirty fur, chromorrhinorrhea, chromodacryorrhea as well as piloerection immediately after finishing the exposure period. Dirty fur and piloerection lasted until up to day 5 and day 8 of study for male and females, respectively. Additionally, immediately after exposure, one male and the two surviving female animals presented shaking. Prostration, and ptosis was also observed in one male (ID 1) and one female (ID 4) animals.

From study day 8 until the end of the 14 day-observation period, no clinical signs were observed in any of the five surviving animals and all of them exhibited a normal behavior.

A decrease in mean body weight of approximately 6% in males and 7% in females was observed between study day 1 (exposure) and study day 2. Thereafter body weight tended to increase gradually in most animals and mean body weight gains over the 14 day observation period of 6% and 13% were recorded for males and females, respectively.

It can be concluded that, under the present experimental conditions:

- The exposure of male and female Sprague Dawley rats to the test item NICOTINE BITARTRATE DIHYDRATE for 4 hours resulted in the premature death of one female animal.

- The marginal body weight loss between study days 1 and 2 as well as the common clinical sings in all animals were considered to be mainly due to the stress related to the nose-only exposure procedure, although a contribution of the treatment with the test item cannot be excluded.

- No macroscopical findings were recorded upon gross necropsy in any of the animals.

- The LC50 of NICOTINE BITARTRATE DIHYDRATE was greater than 4.42 mg/L air (gravimetric aerosol concentration). The test was performed at the technical limit concentration.

- Based on the GHS classification criteria, NICOTINE BITARTRATE DIHYDRATE can be considered to be under human health hazard Category 4 (“harmful if inhaled”) or higher.