AMES mutagenicity test, base oils

The Ames test employs several strains of Salmonella typhimurium which have been selected based on their sensitivity to mutation.  The test substance (liquid) or an extract of the test substance and the test organism are mixed together in a soft agar solution.   Reverse mutation can then be observed.

Study Outline

The Ames test employs several strains of Salmonella typhimurium which have been selected based on their sensitivity to mutation because of: 1) an increased cell wall permeability, due to mutation which causes partial loss of the lipopolysaccharide barrier that coats the bacterial surface and results in increased permeability to large molecules (rfa mutation); 2) a mutation in the bacterial cell system to excise and repair defects in the DNA, resulting in the inability to repair damaged or mutated sections (uvrB mutation); and 3) R-factor plasmids (some strains) and a multicopy plasmid (some strains) which contain error-prone DNA repair systems.  The tester strains also require histidine for growth, due to a mutation in the gene which controls production of histidine.
The Salmonella reverse mutation (Ames) tests are performed by mixing the test substance (liquid) or an extract of the test substance and the test organism together in a soft agar solution, which contains only small amounts of histidine.   The histidine permits the inoculated test organism to undergo a limited number of divisions, but is insufficient to permit normal growth.  If, however, the strain undergoes a reverse mutation, (spontaneous or induced by the test substance or a positive control material), the organism no longer requires histidine to grow and can produce a visible colony or revertant.  Only mutations to the test organism in the region of the histidine gene will cause the test organism to undergo a reverse mutation to an organism which then no longer requires histidine.  The tester strains are selected to detect various types of mutagens.  The tester strains employed are TA97A, TA98, TA100, TA102 and TA1535.

A spot test is also performed where the test substance or an extract of the test substance is spotted on a plate containing the test organism with a small amount of histidine.  As the test substance diffuses through the agar, it becomes more dilute.  This allows for detection of mutagens which cause some inhibition of the tester strains.  The tests are performed both with and without S-9 activation.  The S-9 activation system is designed to simulate mammalian liver enzyme systems and is used to detect substances which undergo metabolic activation from non-mutagenic forms.

Sample Requirements

Solids with uniform surface area and thickness <0.5 mm: 120 cm2
Solids with uniform surface area and thickness >0.5 mm: 60 cm2
Solids without uniform surface area: 4 grams
Liquids: 10 mL

Nelson Experts:

• Chad Summers 801-290-7892, 800-826-2088

Sample Requirements

Solids with uniform surface area and thickness <0.5 mm: 120 cm2
Solids with uniform surface area and thickness >0.5 mm: 60 cm2
Solids without uniform surface area: 4 grams
Liquids: 10 mL