Understanding Absorbance
Absorbance (A) in spectroscopy is a quantitative measure of the amount of light absorbed by a sample when a light beam passes through it. It quantifies how much incident light is diminished by the sample, providing a direct relationship to the concentration of the absorbing substance.
How it is Measured
Absorbance is typically measured using a spectrophotometer. This instrument shines a specific wavelength of light through a sample and measures the amount of light that passes through (transmittance). Absorbance is calculated from the ratio of the initial light intensity (I₀) to the transmitted light intensity (I) using the formula A = log₁₀(I₀/I).
Relationship to Concentration (Beer-Lambert Law)
The Beer-Lambert Law establishes that absorbance is directly proportional to the concentration (c) of the absorbing species and the path length (b) of the light through the sample. This fundamental relationship, expressed as A = εbc (where ε is the molar absorptivity coefficient), is vital for determining unknown concentrations in solutions.
Importance and Applications
Absorbance measurements are critical in diverse scientific fields, including chemistry, biochemistry, and environmental science. They are utilized for identifying substances, quantifying concentrations (e.g., DNA, proteins, chemical contaminants), monitoring reaction kinetics, and investigating molecular structures.