Beer's law states that for a parallel beam of monochromatic radiation passing through homogeneous solutions of equal pathlength the absorbance is proportional to the concentration.
i.e. For solutions 1 and 2
(Absorbance 1)/(Absorbance 2) = (Concentration 1)/(Concentration 2)

Lambert's law states that for a parallel beam of monochromatic radiation passing through homogeneous solutions of equal concentration the absorbance is proportional to the pathlength.
i.e. For pathlengths A and B
(Absorbance A)/(Absorbance B) = (Pathlength A)/(Pathlength B)

The absorbance of a 2-cm layer of a 0.0006 %^w/v solution is 0.48
What would be the absobance of a 1-cm layer of a 0.0009 %^w/v solution of the same substance under the same conditions?

0.64      0.16      0.36      1.44      0.48     

The Beer-Lambert law states that for a parallel beam of monochromatic radiation passing through a homogeneous solution the absorbance is proportional to the product of the concentration and pathlength.

Absorbance = constant × concentration × pathlength

The value of the constant depends on the substance, the solvent, the wavelength and the units used for concentration and pathlength. Two common methods of expressing the constant are A(1%,1cm) and molar absorptivity.

A(1%, 1cm) is the absorbance of a 1-cm layer of a 1%^w/v solution.

A = A(1%, 1cm) × C × L

Where C is the concentration as %^w/v and L is the pathlength in cm.

A 2-cm layer of a 0.003 %^w/v solution of Ethamivan exhibits a maximum at 280 nm with an absorbance 0.92.
What is the A(1%, 1cm) of Ethamivan under these conditions?

304      3420      153      307      46700     

A 4-cm layer of a 0.00003 mol litre^-1 solution of Ethamivan has an absorbance of 0.411 at the maximum at 280 nm

What is the molar absorpivity of Ethamivan under these conditions?

153      1120      2725      3425      13700     

A(1%, 1cm) values are generally found in the pharmaceutical literature while molar absorptivities are found in the chemical literature. You may not always be able to find the information in the most convenient form.

A 1-cm layer of a 0.0008 %^w/v solution of Thiabendazole in 0.1M hydrochloric acid has an absorbance of 0.47 at the maximum at 243 nm.

Calculate the molar absorptivity (litre mol^-1 cm^-1) under these conditions.

1.18 × 10³      1.18 × 10⁴      1.43 × 10⁴      588      517     

When preparing solutions for absorbance measurements you will need to know what concentrations to prepare in order to obtain suitable absorbances. For routine analytical work absorbances greater than 1 should be avoided.

Sulphadiazine in ethanol has a molar absorptivity of 2.11×10⁴ mol^-1 litre cm^-1 at the maximum at 270 nm.

From the list below, select the concentrations (mol litre^-1 and %^w/v) which would give an absobance of 1.0 in a 2-cm cell at 270 nm

4.74×10^-5 mol litre^-1      2.37×10^-5 mol litre^-1      6.40×10^-3 mol litre^-1      1.18×10^-3 %^w/v      6.40×10^-4 %^w/v      5.92×10^-4 %^w/v      3.20×10^-4 %^w/v     

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