Spectroscopy of Fast Green Solution and Chlorophyll A and B

Authors Avatar

Spectroscopy of Fast Green Solution and Chlorophyll A and B


The purpose of this lab is to be introduced to measure the concentration of an unknown solution using spectroscopy and to isolate the individual pigments of a chloroplast extract to measure the light absorption of chlorophyll A and B. Fast green is Light is a photon, a particle without mass however is a wave. (Petrucci, 2010)

 In these experiments the visible light spectrum having a wavelength between 400 nm to 700 nm was used. A spectrophotometer separates light into distinct bands of energy, allowing on to focus a particular band of energy to measure its absorption from 0 to 100%. (Jones et al., 2007) The spectrophotometer tells the observer the absorbance based on whatever wavelength one wants to find. (Jones et al., 2007) An essential part of observing a substance under a spectrophotometer is that of using a blank. The role of a blank and the spectrophotometer is to set the absorbance of the spectrophotometer to zero allowing the absorbance of one substance to be shown. (Jones et al., 2007)

 In experiment one; water was used as the blank. Based on the results from the spectrophotometer, on is able to make a concentration curve to find the concentration of the unknown solution. (Jones et al., 2007) A spectrophotometer is an instrument that measures the intensity of the light entering a sample and the light exiting a sample and compares the two intensities. Information about the two intensities can be expressed as transmittance. (Vogelman and Evans, 2002) The transmittance is the ratio of the intensity of the exiting light to the entering light. (Jones et al., 2007) This value is expressed as a percentage. Different materials absorb different wavelengths of light. (Karp, 2010) Therefore, the wavelength of maximum absorption by a material is one of the characteristic properties of that material. (Karp, 2010) The transmittance can be related to the absorbance (A) by the formula A is equal to 2 – (log(%T)). For example if the transmittance is 60 percent because 40 percent of the intensity of the light is absorbed passing through the sample.(Jones et al., 2007) The absorption would be 2 – log 60 which is 0.22184 A. Beer’s Law states that the absorbance is directly proportional to the concentration of a solution. (Vogelman and Evans, 2002)

Join now!

 If you plot absorbance versus concentration, the resulting graph yields a straight line. This is called an absorptions spectrum which illustrates the intensity of light absorbed to its relative wavelength. (Karp, 2010) The equation for the straight line can be used to determine the concentration of an unknown solution. (Karp, 2010) This is how the unknown concentration fast green is found in experiment number one. (Karp, 2010)

Materials and Methods

The protocol and materials used for the experiment were completed as written in the Department of Biology of the University of Waterloo Fall term 2010 Biology 130L Lab Manual, pages ...

This is a preview of the whole essay