If Caffeine Contains the Properties of a Mutagen

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Many common products, like preserved foods, cleaning agents, and vehicle airbags, contain known mutagens that are hazardous to humans. Caffeine is present in coffee and energy drinks, beverages consumed on an everyday basis by many people. It is important to recognize how interactions with these substances can affect the body, specifically an organism’s DNA. The purpose of this experiment is to determine if caffeine contains the properties of a mutagen necessary to revert his ̄ S. typhimurium bacteria from an auxotroph into a prototroph.

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Terms central to understanding this lab include: mutagen, auxotroph, prototroph, Ames test, and revertant. A mutagen is a property of a substance that possesses the ability to permanently alter the makeup of DNA and change the message of gene expression. The original sample of his ̄ S. typhimurium is an auxotroph, meaning that it requires a growing medium with histidine, as it lacks the ability to produce its own. 

The revertants yielded by this experiment are bacterial cells that mutated from auxotrophs to prototrophs. These revertants are said to be prototrophs because their mutations allow them to produce their own histidine, along with all other molecules necessary for self-sufficiency. The Ames test is an assay on S. typhimurium bacteria to test for mutagenicity of compounds added to the bacterial sample. It is important to state that the process of testing caffeine’s genotoxicity on bacteria is suitable for extrapolation onto its effect on humans as the possible mutagen acts on the DNA of both organisms. Also, caffeine can only be determined to be a mutagen, rather than a carcinogen, because carcinogenicity determination falls outside the scope of the Ames testing results.

The strain of bacteria used, TA1538 his ̄ S. typhimurium, has vital auxotrophic mutations that allow it to be a proper subject for this experiment. The TA1538 bacterial strain is valuable here as it has a single base pair deletion of C in its HisD gene that results in a frameshift mutation yielding a premature introduction of a stop codon in frame 133 following the exchange of two amino acids. This is what prevented the bacteria from producing its own histidine. TA1538 also had a mutation of the rfa gene that caused the lipopolysaccharide cell wall to be defectively weak, which allowed for increased susceptibility of the cell to external mutagens. The defective excision repair mechanisms interfered with the cell’s ability to remove the emerging mutations during DNA proofreading and replication.

In accordance with the popularity of caffeinated beverages and prior research, caffeine is likely to have a low mutagenicity rate. The null hypothesis states that caffeine would not act as a mutagen on the his ̄ S. typhimurium, while the alternative hypothesis states that caffeine would act as a mutagen in the tested bacteria as shown by the production of revertant colonies.  

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