The OECD guidelines for inhalation toxicity studies detail protocols for assessing the toxicity of chemicals over acute (14 days), sub-acute (28 days), and chronic (90 days) durations using mammalian species. The guidelines emphasize the importance of behavioral, physiological, and biochemical observations, as well as monitoring chamber conditions for effective exposure setups. Comprehensive data reporting and analysis, including individual animal observations and chemical properties, are outlined to ensure the reliability of toxicity assessments.

1. OECD GUIDELINES FOR THE TESTING OF
CHEMICALS INHALATION TOXICITY STUDY
INHALATION TOXICITY STUDY
 This test guidelines is designed to fully characterised test
chemicals toxicity by inhalation route for acute duration (14
days), sub acute (28 days), and chronic duration(90 days).
 This test guidelines also suggests additional investigation,
such as toxicokinectics and systemic toxicity study to better
characterize the toxicity of test chemical.
 This inhalation toxicity study can be used for quantitative risk
assessment.

2.  Acute inhalation studies can be found in the guidelines
Documents No. 39.
 Acute inhalation toxicity guidelines – 403.
 Sub- acute inhalation toxicity guidelines- 412.
 Chronic inhalation toxicity guidelines- 413.

3. ( ACUTE TOXICITY OR SINGLE DOSE STUDIES)
PRELIMINARY
 single dose tested in 2 rodents species.
 Observation for 14 days after dosing
 Sign, and symptoms reported
 microscopic and macroscopic evaluation.

4. DEFINITIVE-
 Group of 20 animal of either sex
 5 animals of each sex are observed for 48 hrs.
 Remaining 5 animals of each sex are 14 days.
 Changes in body wt.
 Pathological changes
 Macroscopic and microscopic evalution.

5. REPEATED DOSE STUDIES/ SUB-ACUTE TOXICITY
 2 mammalian species(one should be non rodents)
 Long duration studies.
 Parameters observed and recorded are..
 Behavioral
 Physiological
 Biochemicals
 Microscopic observation

6. CHRONIC TOXICITY STUDIES
 To evaluate the toxicity of chemicals.
 2 species required- rodents and non- rodents
 Rodents- 6-24 months
 Non- rodents- 12 months or longer
 Parameter monitored and recorded are-
 Mortality
 Pathology and histopathology
 Wt. change

7. Description of the method
Selection of species…
 Rodents species is rat.
 Rodents species is used.
 One male and one female animals may be performed for the
study.
 Healthy young adults animal of laboratory strains should be
used.
 If female are used they should be non- pregnant.
 Should be b/w 8 and 12 weeks old.
 Weight should be 20% of the mean body wt.

8. Housing and feeding condition
 The temperature of the experimental animal room should be
22 degree celcius.
 The relative humidity should be at least 30% and not exceed
70%.
 Lighting should be artificial, being 12 hours dark,light
Preparation of animals
 The animals are acclimatised to the laboratory conditions for
at least 5 days prior to the start of exposure.
 Animals are selected for used in the study and provide
individual identification.

9. Main study
No. of animals and concentration levels..
 5 animals of one sex will be used for each concentration.
 Each group consists of max. of 5 male and 5 female rodents
are exposed to the test chemicals for 6 hrs per day on 5 day
per week for a periods of 4 weeks (14 days for acute, 28 days
for subacute, and 90 days for chronic).

10. Exposure condition
Administration of concentration
 Animal are exposed to test chemicals as a gas, vapours,
aerosol or a mixture.
 Hygroscopic and chemical reactive test chemical should be
tested under dry air condition.
Test chemical preparation in a vehicle-
 Test chemical should be tested without vehicle.
 If it is necessary to use a vehicle to generate an appropriate
test chemical concentration and particle size, water should be
used whenever possible.

11. Monitoring of exposure conditions
Chamber air flow
 The flow of air through exposure chamber should be carefully
controlled.
 Oxygen concentration Should be at least 19% and carbon di
oxide concentration should not exceed 1%.
 If measurement on the 1st day of exposure show that these
gases are at proper levels, no further measurement should be
necessary.

12. Chamber temp. and relative humidity
 The air temp. in the animal breathing room should be
monitored continuously and recorded at least 3 times during
each exposure, chamber temp. should be maintain at 22
degree celcius.
 The relative humidity should be maintain in the range of 30 to
70%.
Particle size distribution
 Particle size distribution of fine aerosols should be determined
by cascade impactor.

13. Observation
 The animal should be clinically observed before, during and after
each exposure periods as well as during the post exposure periods.
 When animals are sacrified, time of death should be recorded.
 Change in the skin , fur, eyes , mucous membrane, change in
respiratory and circulatory system are observed and recorded.
Body wt.
 Individual animal wt. should be recorded before the first
exposure(day 0), twice weekly or at the time of death or
euthansia.
Food and water consumption
 Food and water consumption may also be measured weekly.

14. Data and reporting
Data
 Individual animal data should be provided.
 All data should be summarised in tabular form….
 No. of animal used
 No. of animal displaying sign of toxicity
 No. of animal dead during the test.
 Time of death
 Discription

15. Test report
Test report should include the following information…
 Source
 Lot no.
 Stability of test chemicals
 Solubility of the test chemical in solvent, if known.
 Measurement of PH and osmolality.
 Physical appearance, water solubility and physicochemical
properties.
 Chemical identification such as IUPAC or CAS name , CAS no.
 Purity

16. Vehicle
 Use of vehicle and justification for choice of vehicle(if other
than water).
Test animal
 Species/strain used
 Microbiological status of the animal.
 Acclimatised period.
 No. ,age and sex of animal
 Source housing condition.
 Diet.

17. Test condition
 Details of test chemical preparation.
 Equipment used to monitor chamber temperature humidity
and airflow.
 Equipment used to collect sample for determination of
chamber concentration and particle size distribution.
 Details of food and water quality.
Result
 Tabulation of chamber temperature, humidity and airflow.
 Tabulation of chamber nominal and actual concentration data.
 Tabulation of response data and concentration level for each
animal.

18. THANK
YOU
KANCHAN GUPTA
M.PHARM(Pharmacology)
Advance institute of biotech and paramedical sciences