Question

In an Ames test using his- Salmonella bacteria a researcher determines that adding a test compound plus the S9 extract produces a large number of his+ revertants, but mixing the his- strain plus the test compound without adding S9 does not produce an elevated number of his+ revertants. Explain this result. Also state whether or not you think this compound should be considered a potential mutagen.

Answer 1

Ames test: This test is used to determine whether the given test compound is a mutagenic compound or non mutagenic compound. When the test compound added to the media, allows the Salmonella bacteria (His^-) to grow that is, it is reverting the Salmonella His^- mutants to its wild type Salmonella His⁺, there by causing changes in Histidine producing gene and acting as a mutagenic agent.

S9 extract which is the rat liver extract, is usually added to the above media (Salmonella His^- mutants + growth media minus Histidine compund+ test compound+ S9 extract) in order to stimulate the effect of metabolism on the test compound, as most of the compounds like benzopyrene, are by themselves not mutagenic but their metabolic products that are formed once they are metabolised in the liver do act as mutagenic agents. Thus adding the S9 extract to the medium in Ames test truly confirm the actual mutagenic nature of the compound.

In the above case mentioned the test compound is giving smaller number of Salmonella His⁺ revertents without the addition of S9 extract, but by the addition of S9 extract the test compound is producing elevated levels of Salmonella His⁺ revertents. This suggests that the test compound which is basically a mutagen is becoming more potential mutagen in its metabolized form when it is being processed by the liver extract enzymes (S9 extract). In actual metabolism process in an organism, the test compound should go through the metabolism in liver and its metabolised compound should be actually tested for mutagenic nature.

In the above experiment it is clear that addition of S9 extract to the test compound is producing elevated levels of Salmonella His⁺ revertents. Thus its metabolised form is considered as potential mutagen which in turn indicates that the test compund itself is a potential mutagen.