Experiment to Determine How Much Caffeine and Nicotine it Takes to Cause Human Cell Death (Introduction and Materials/Method Sections Only)

Experiment to Determine How Much Caffeine and Nicotine it Takes to Cause Human Cell Death (Introduction and Materials/Method Sections Only)
Caffeine for many, and nicotine, for some, is used in their diet almost every day. As many already know, too much nicotine is not found to be safe, but how much nicotine is always the question. This also goes for caffeine. Most of the population has caffeine in their daily diet, but there is always the question of how much caffeine is actually damaging to cells, and ultimately, the body. Since testing on humans is not a valid option, scientist use other resources, such as Saccharomyces cerevisae, to find out just how much nicotine or caffeine can be taken before mutations or even death starts to occur within the cells. As defined by one of the studies done by Institute of Biochemistry and Biophysics, Warsaw, Poland, caffeine is a natural purine analogue that elicits pleiotropic effects leading ultimately to cell?s death by a largely uncharacterized mechanism1. Nicotine is already looked as being very harmful to one?s body, given that it is tied with smoking. A lot of research has been done with nicotine and cell death. One of these was done on whole rat embryo cultures used to study the effects of nicotine at the neural tube stage of development.2 At the highest concentration, abnormalities were present in the majority of cells. Looking at the affects of nicotine and caffeine on Saccharomyces cerevisiae, one can find the outcomes and be able to relate them to humans. It was predicted that the higher levels of nicotine and caffeine used will kill the cells, and that nicotine will show the effect of cell death the most and with the most mutations or reversions.
Materials and Methods
Within this experiment, ten synthetic complete medium Petri dishes (SC), and ten synthetic ?dropout? medium Petri dishes (SD) are used. Five of the SC Petri plates will be used for nicotine, five for caffeine, and vice versa for the SD Petri dishes. The synthetic complete medium Petri dishes are being used to show any cell death that occurs, and the synthetic ?dropout? medium Petri dishes are being used to show the number of reversions or mutations in the cells that occur. Since the nicotine and caffeine that will be used starts off as 1%, serial dilutions have to be made. The serial dilutions are very similar to the biology L113 laboratory manual3. However, the concentrations used for this experiment are 0, .00005, .0001, .00015, and .0002. Each of these solutions were made independently by the techniques used in the L113 laboratory manual. The 0 will be the control for the experiment, and is used to compare with the other Petri dishes that actually do have concentrations of either nicotine or caffeine introduced to them. The yeast used for this experiment is Saccharomyces cerevisae, commonly known as bakers yeast. Eppendorf tubes are also needed, to make the dilutions, and also a flamed glass spreader with 95% ethanol. The spreading of the yeast is then made to the SD and SC Petri dishes with the same techniques used in the L113 laboratory manual. However, when one pipettes the yeast into the Petri dishes, the nicotine or caffeine is also introduced with the appropriate concentrations, being .0005, .0001, .00015, or .0002. After spreading the yeast, they are stored in a room for two days at 30 degrees for the yeast to grow. After the two days, each Petri dish is counted for yeast colonies.

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