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Acute Oral Toxicity - OECD Fixed Dose Method
This bulletin provides information regarding the use of data from a new OECD Test Guideline 420 [Fixed Dose Method (FDM)] which measures acute oral toxicity in animals.
A major factor in the development of this new test method was the desire to minimize the use of laboratory animals for testing chemical products. The more traditional test for acute oral toxicity is the oral LD 50 , which uses more animals than the FDM, and one test protocol for this is found in OECD Test Guideline 401.
Oral LD 50 test data are used to evaluate and classify the toxicity hazards of chemical products under the Controlled Products Regulations(CPR) for WHMIS and also under the Canadian Transportation of Dangerous Goods (TDG) Regulations.
Since FDM data are significantly different from LD 50 data, this bulletin provides guidance on how to use FDM data to classify products under WHMIS Class D1, when LD 50 data are not available, and also covers appropriate disclosure of test data on a Material Safety Data Sheet (MSDS). This guidance does not address classification of products under TDG Class 6.1.
It should be noted that no equivalent Fixed Dose methods have been developed so far for the dermal and inhalation routes of exposure.
Currently, sections 46 and 49 of the CPR specify acute oral lethality test criteria for classifying a product in Class D1, based on results from OECD Test Guideline 401 - Acute Oral Toxicity. This Guideline uses lethality as the basis for acute oral toxicity assessment.
The new OECD FDM (Test Guideline 420) departs from the conventional method (Test Guideline 401) in two principal ways:
- FDM considers non-lethal endpoints; and
- it does not develop a quantitative relationship between dose and lethality.
The method relies upon clear signs of toxicity ("evident toxicity"), a non-lethal endpoint as well as lethality, as the basis for acute oral toxicity assessment.
Use of FDM Data in Classifying Products under Class D1
In the OECD FDM, the initial dose is selected from one of four levels: 5, 50, 500 or 2000 mg/kg body weight (b.w.). The initial dose is selected on the basis of a preliminary "sighting study" performed with single animals of one sex. The main study is conducted with 5 animals/sex/dose. Where no evidence of toxicity is seen at the initial dose, or where severe toxicity or mortality are observed, an additional dose level would be added to the study. When deaths have occurred over the same dose range for the two sexes, the data can be pooled to provide data based on a total of 10 animals.
Two of the suggested doses are 50 and 500 mg/kg b.w., which correspond to the cutoff levels for acute oral toxicity for WHMIS classifications D1A (very toxic material) and D1B (toxic material). It should be stressed that the FDM is NOT intended to result in the generation of data for the estimation of an oral LD 50 for a test substance.
In order to apply this protocol to classification under WHMIS, the following interpretations should be applied:
- Where mortality in the FDM is ³ 50% at one of the fixed dose levels, it can be assumed that the LD 50 for the controlled product, when tested in accordance with OECD 401, would likely fall below that fixed dose level. Consequently, it follows that:
- at a dose of 5 or 50 mg/kg b.w. one would classify the controlled product in D1A.
- at a dose of 500 mg/kg b.w. one would classify the controlled product in D1B, provided there was no data available to show that lethality at 50 mg/kg b.w. exceeded 50%.
- at a dose of 2000 mg/kg b.w., one would need to review test data at the next lower dose level (500 mg/kg) to determine if lethality at that level is 50%.
- Where there is 100% survival with or without evident toxicity:
- at a dose of 5 or 50 mg/kg b.w. one would need to review test data at the next highest dose level (50 or 500 mg/kg b.w.) to determine if lethality at that level is ³ 50%.
- at a dose of 500 or 2000 mg/kg b.w. the LD 5o is not very likely to be £ 500 mg/kg b.w., therefore the product would not be classified for acute oral toxicity under WHMIS Class D1.
- Where mortality is < 50% but > 0% at one of the fixed dose levels, it can be assumed that in most cases, the LD 50 value for the substance, when tested in accordance with OECD 401, would fall above that fixed dose level. Consequently, it follows that:
- at a dose of 5 mg/kg b.w. - one would need to review the lethality data for the next highest fixed dose (50 mg/kg) to determine the % lethality at that level. If mortality at the next fixed dose level is ³ 50% the controlled product would be classified D1A, as described in 1(a), above.
- at a dose of 50 mg/kg b.w. - one would need to review the lethality data for the next highest fixed dose (500 mg/kg b.w.) to determine the % lethality at that level. If mortality at the next fixed dose level is ³ 50% the controlled product would be classified in D1B, as described in 1(b), above. However, if lethality at 50 mg/kg b.w. is 40% (i.e. if the data are pooled and 4/10 animals died, especially if the majority of them were of one sex) and no data are available at 500 mg/kg b.w., the conservative approach would be to assume that the LD 50 value for the substance might be below 50 mg/kg b.w., and consequently it would be prudent to classify in D1A.
- at a dose of 500 mg/kg b.w. - if lethality at 500 mg/kg b.w. is 40% (i.e. 4/10 animals died) and no data are available at the next highest fixed dose (2000 mg/kg b.w.), the conservative approach would be to assume that the LD 50 value for the substance might be below 500 mg/kg b.w., and consequently it would be prudent to classify in D1B. If lethality at 500 mg/kg b.w. was lower than 40%, it would be reasonable to assume that the substance is not classified under WHMIS class D1.
- at a dose of 2000 mg/kg b.w. - not classified for acute oral toxicity under WHMIS Class D1.
Disclosure of FDM Data on a MSDS
The implication of the FDM for hazard disclosure under WHMIS is that an oral LD 50 is not generated. Consequently, the LD 50 subitem is not available for disclosure on the MSDS as required by CPR Schedule I, Item 1. However, mortality and evident toxicity data would be available, and both should be disclosed on the MSDS under "effects of acute exposure".
Impact of FDM data
It is expected that the use of this Guideline will have an overall neutral impact on the classification of substances under class D1. Since products need to be classified under both WHMIS and TDG regulations, and this is typically carried out at the same time, it should be recognized that guidance from Transport Canada will be required respecting the application of FDM test data to TDG Class 6.1.
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