The purpose of this experiment is to determine the maximum absorbance of fast green, and the chlorophylls, also in the case of fast green create a concentration curve to determine an unknown substance. Each test will use the spectrophotometer.
Spectroscopy is the study of light. A spectrophotometer is a machine used to determine the absorbance of light at any given wavelength. It does this by using a source of white light through a prism, which gives multiple wavelengths that can be individually focused (Ayyagari and Nigam, 2007). Substances are put into cuvettes that are glass or quartz containers that light can easily travel through. The light that is being focused travels through the substance gets absorbed by the substance and is reflected back and read by galvanometer which had the ability to detect electric currents (Verma, R). The absorbance reading is then given, absorbance is usually between 0.0 – 2.0, any higher than 2.0 may mean not enough light is getting through to the galvanometer (Bhowmik and Bose, 2011). When using the spectrophotometer it is necessary to use a control or blank to zero or tare the machine in between every new wavelength or concentration, this control is water (Ayyagari and Nigam, 2007). The correlation between the numbers acquired through spectroscopy can be seen using the Beer- Lambert Law. The Beer- Lambert law states that the amount of light absorbed at a certain wavelength is proportional to the concentration of the absorbing substance (Fankhauser, 2007).
The first experiment involves fast green, a stain for protein that is used to demonstrate histones, as well as a marker dye (Lundbland and Macdonald, 2010). The purpose of this experiment is to obtain a concentration curve. A concentration curve also known, as a calibration curve is a plot of absorbance versus concentration with this it is possible to determine any unknown concentration of fast green if you have either the absorbance of it
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