What factors contribute to deviations from the Beer-Lambert’s Absorbance law?

Beer–Lambert’s Absorbance law is a universally accepted relationship which helps calculation of concentration of an absorbing species from measured absorbance values. Under ideal conditions absorbance versus concentration plot is a straight line passing through the origin. However, under real situations a curvature in the plot is observed beyond a particular concentration. The curvature can be both positive or negative depending on the nature of absorbing species and experimental conditions.在早期的文章中讨论了与法律的偏差。本文将讨论负责此类偏差的原因。

Positive deviations are observed when the measured absorbance values are more than the expected theoretical values and negative deviations result when the measured values are lower than the theoretical values.

Limitations of the Law

啤酒 - 兰伯特的法律通常表现出低于关键限额的浓度。对于大多数吸收分子，在10mm以上的浓度下观察非线性行为。然而，一些吸收分子如亚甲基蓝色倾向于表现出与浓度的线性度低至10μm的偏差。溶质 - 较高浓度甚至氢键的溶剂相互作用负责这种偏差。

化学因素

改变博士

Some absorbing molecules undergo change in colour with change in pH of the solvent medium. An example is Phenol red which changes colour from yellow in acidic media to red in basic media due to internal migration of proton accompanied by a switch between a single and an adjacent double bond.

另一个例子是二聚角的水溶液，从黄色变为橙色，随着铬酸盐 - 二甲酸酯转化而增加氢离子的浓度

\（2Cro_7 ^ 2 ^ - + 2 h ^ + \）\（\ rectionleftrarrows \）\（cr_2o_7 ^ 2 ^ - + h_2o \）
重铬酸铬酸
（黄色）（橙色）

Complexation , Dissociation or Association

由于关联引起的复杂形成可能导致浓度变化的颜色变化。一个例子是氯化钴，其由于关联而从绿色变为蓝色的颜色

\(2CoCl_2\) \(\rightleftarrows\) \(Co(CoCl_4)\)
（粉红色）（蓝色）

The degree of association increases with increase in concentration

Instrumental factors

波长选择

Beer- Lambert’s law holds true strictly for single wavelength light. However, due to inefficiencies of monochromator a range of wavelengths gets isolated. In addition stray light also results from internal reflections from optical components such as gratings, lenses and optical windows. Absorption resulting from stray light contributions will show deviations from the Beer- Lambert’s law

Mismatch of solution and cells

Sample and reference blank solutions composition variations or optical mismatch of windows of cuvettes will result in deviations from Beer- Lambert’s law.

To minimize the deviations the following precautions should be taken while reading absorbance of solutions to provide high accuracy of results:

• Always use optically matched pair of measuring cells
• 分析物的浓度应低于啤酒的线性范围。
• pH should be maintained at specified value for both blank and sample solutions
• 空白和样品溶液的组成应尽可能地匹配