Quantitative Evaluation of Thin-Layer Chromatograms

The first part of the book consists of a detailed treatment of the fundamentals of thin-layer chromatography, and of measurement techniques and apparatus for the qualitative and quantitative evaluation of thin-layer chromatograms. In situ prechromatographic derivatization techniques used to improve the selectivity of the separation, to increase the sensitivity of detection, and to enhance the precision of the subsequent quantitative analysis are summarized in numerous tables. 

Quantitative evaluation of thin-layer chromatograms can be performed by direct, in situ visual, and indirect elution techniques. Visual evaluation involves comparison of the sizes and intensities of color or fluorescence between sample and standard zones spotted, developed, and detected on the same layer. The series of standards is chosen to have concentrations or weights that bracket those of the sample zones. After matching a sample with its closest standard, accuracy and precision are improved by respotting a more restricted series of bracketing standards with a separate sample spot between each of two standard zones. Accuracy no greater than 5-10% is possible for trained personnel using visual evaluation. The determination of myco-toxins in food samples is an example of a practical application of visual comparison of fluorescent zones. 

Quantitative evaluation of thin-layer chromatograms can be performed by direct, in situ visual and indirect elution techniques. Visual evaluation involves comparison of the sizes and intensities of color or fluorescence between sample and standard zones spotted, developed, and detected on the same layer. The series of standards is chosen to have... 

Falk, H. and Krummen, K. 1975. A new method for the quantitative evaluation of thin-layer chromatograms, J. Chromatogr., 103 279-288. 

H. Ganshibt Quantitative Evaluation of Thin-Layer Chromatograms... 

Methods for the quantitative evaluation of thin-layer chromatograms can be divided into two categories. In the first, solutes are assayed directly on the layer, either by visual comparison, area measurement, or densitometry. In the second, solutes are eluted from the sorbent before being examined further. Quantification of radioactive zones is not considered here but is discussed in Chapter 13. 

Phenylalanine (1), rerf-leucine (2), 5,5-dimethylthiazolidine-4-carboxylic acid (3), and a-hydroxy-phenylalanine (4) have been chosen as models for the direct quantitative evaluation of thin-layer chromatograms. Emphasis has been placed on the evaluation of detection limits for the TLC-separated enantiomers, because exact determination of trace levels of a D- or L-enantiomer in an excess of the other is increasingly important (171,201,203-205). 

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