Spectrophotometry

Spectrophotometry: 

Spectrophotometry Measures the amount of light that is absorbed or transmitted by a chemical Different substances will absorb/transmit different frequencies of light Amount of absorbance is proportional to the concentration

Using a Spectrophotometer: 

Using a Spectrophotometer Samples are placed in a cuvette Small square vial or small test tube Always run a blank A blank is like a control Standards Solutions of known concentration are run to create a standard curve Unknowns can be compared to the standard curve to determine their concentration

Beer’s Law: 

Beer’s Law Relates absorbance to solution concentration A = a b c A = Absorbance a = Beer’s Law Constant b = path length (width of cuvette) c = solution concentration

Using Beer’s Law to Create a Calibration Curve: 

Using Beer’s Law to Create a Calibration Curve By plotting absorbance vs. concentration for a series of solutions of known concentrations, a calibration curve (generally, a linear plot) can be created. The concentration of unknown solutions (falling within the range of concentrations used to create the calibration curve) can be determined by plotting the absorbance value.