Abstract
Detection of target compounds, particularly trace amounts of micropollutants or biomarkers in complex samples, is a key step of chromatographic analysis. In principle, any qualitative and/or quantitative analysis demands a suitable signal to noise ratio allowing chromatographic detection with an appropriate contrast. In planar chromatography and related techniques (e.g., microfluidic-paper-based devices) this can be achieved by online or pre/postseparation reactions of analytes with a variety of visualization reagents. This process is commonly described as staining and/or derivatization and allows effective detection of components of interest, which in their native form are transparent to a given detector at specific data acquisition conditions. Typically, the analytical signal results from physicochemical interactions, chemical reactions, or UV/thermal decomposition. Visualized zones on a thin-layer chromatography (TLC) plate can be acquired by detectors usually operated within the UV–Visible–IR range in the absorbance or fluorescence mode recorded in reflectance or transmission by digital cameras, scanners, or video densitometers. This chapter provides a practical overview of commonly applied derivatization protocols, including application techniques and reagents types, which are frequently used for fast, sensitive, and nonexpensive detection of a wide range of target compounds on thin-layer chromatography plates, bars, or strips coated with different stationary phases. Derivatization techniques in terms of both quantitative and qualitative analysis, detection of specific physicochemical or biological properties of target compounds, as well as selected problems associated with signal optimization are highlighted. Moreover, visualization protocols for detection of target components on TLC plates included in the General Monographs of the European Pharmacopoeia are summarized as an example of the extensive use of staining and derivatization reagents for qualitative and quantitative analysis.
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