AAT Bioquest

What factors will affect absorbance?

Posted October 30, 2020


Absorbance measures the amount of light with a specific wavelength that a given substance prevents from passing through it.

The two main factors that affect absorbance are concentration of the substance and path length.

Relation between concentration and absorbance: Absorbance is directly proportional to the concentration of the substance. The higher the concentration, the higher its absorbance. This is because the proportion of light that gets absorbed is affected by the number of molecules that it interacts with. Solutions that are more concentrated have a larger number of molecules that interact with the light that enters, thus increasing its absorbance. In a diluted solution the absorbance is low because fewer molecules are available to interact with the light.

Relation between concentration and path length: Absorbance is also directly proportional to the path length, where path length refers to the distance the light travels through the substance. With a longer path length, the light interacts with a larger number of molecules as it travels the longer distance through the solution. This increases the absorbance.

Additional resources

Theoretical and experimental investigation of factors affecting precision in molecular absorption spectrophotometry

Absorption Spectrum Viewer

DNA Concentration Calculator

Beer Lambert Law Calculator