Column chromatography constituent identification

Thin-layer chromatography (TLC) [5] is routinely used in dendrimer synthesis for purity checks and identification of individual constituents of a dendrimer sample and is suitable for monitoring column chromatographic separations. Since chemistry students are introduced to this analytical technique during their first semesters, we shall not go into further details here. 

This limitation has been largely overcome by linking chromatographic columns directly with ultraviolet, infrared, and mass spectrometers. The resulting hyphenated instruments are powerful tools for identifying the components of complex mixtures (see Section 31A-4). An example of the use of mass spectroscopy combined with gas chromatography for the identification of constituents in blood is given in Feature 31-1. 

The separation of nucleotides and deoxynucleotides, previously a formidable task involving the fractional crystallization of heavy metal and alkaloid salts 102) has been made much easier by developments in analytical techniques. Ion-exchange methods may be used for the purification, isolation, and identification of both classes of nucleotides from hydrolysis mixtures 103), Countercurrent distribution 104) and starch 106) and cellulose-column 106) as well as paper-strip chromatography 107) have also proved to be useful in separating nucleotides from natural sources. Spectro-photometric procedures based on the characteristic ultraviolet absorption spectra of the purines and pyrimidines have been the most convenient method to locate, estimate, and identify the fractions obtained in the previous separations. Since the nucleotides are acid in nature, they are often named as acids, e.g., adenylic acid, cytidylic acid. The general constitution of the purine nucleotides (and by analogy the pyrimidine nucleotides) is demonstrated by their hydrolysis by acids to a purine and ribose (or 2-deoxyribose) monophosphate and by alkalies to the nucleosides and phosphoric acid. The order of the constituents in a purine nucleotide must, therefore, be ... 

In this type of chromatc iaphy the position of a compound in the effluent from the chromatogram is less likely to be influenced by the presence of other constituents of the mixture than in the case of adsorption chromatography. Hence, when the procedure is standardized it should be possible to make tentative identification of compounds by their position in the chromatogram, and to predict the position of new compounds if their partition coefficients and the constants of the column are known. 

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