University:
The Pennsylvania State University
Course:
BMB 442 | Biochemistry and Molecular Biology
Academic year:

2024
Views:

132

Pages:

1
Author:

Alexis H.

Spectrophotometry and Beers Law Introduction An absorbance spectrophotometer is an instrument that measures the fraction of the incident light transmitted through a solution. In other words, it is used to measure the amount of light that passes through a sample material and, by comparison to the initial intensity of light reaching the sample, they indirectly measure the amount of light absorbed by that sample Because different compounds absorb light at different wavelengths, a spectrophotometer can be used to distinguish compounds by analyzing the pattern of wavelengths absorbed by a given sample Additionally, the amount of light absorbed is directly proportional to the concentration of absorbing compounds in that sample, so a spectrophotometer can also be used to determine concentrations of compounds in solution The transmittance is defined as the ratio of the light energy transmitted through the sample to the energy transmitted through the reference blank When you adjust the wavelength on a spectrophotometer, you are changing the position of the prism or diffraction grating so that different wavelengths of light are directed at the slit Beers Law Beers law States that the absorbency of a light absorbing material is proportional to its concentration in solution A = "lc A is absorbance Epsilon is the molar absorptivity or extinction coefficient of the substance l is the sample path length measured in cm (width of cuvette usually 1 cm c is the molar concentration of the solution