Chromatography partition, nucleoside separation

Many schemes for fractionating nucleotides, nucleosides and bases on sulphonated polystyrene resins have been published. The main difficulty with these methods is variation between resin batches (e.g. Anderson et al. 1963). Nucleotide separations can be achieved on DEAE-cellulose (Whatman Data Sheet 13, 1967) and DEAE-Sephadex (Piers et al. 1965b) but these media do not seem to be widely used. Gel filtration columns will separate some nucleotide components. Ligand exchange chromatography and partition chromatography of nucleosides are useful for minor components. 

A reversed-phase chromatography technique for the preparative separation of polar compounds has been demonstrated with the separation of a simple nucleoside mixture. A group separation of the 2 -deoxyribonucleoside, nucleotide and nucleobase constituents of normal and modified nucleic acids was achieved by gel permeation chromatography, and further separation was achieved on a hydrophilic acrylate polymer operating in the partition mode. Mononucleotides are selectively bound at low pH to Fe(ni) immobilized on agarose gel due to their free phosphate ester group, and can be recovered with a neutral eluant. Nucleosides and molecules with phosphate diester groups are not retained. ... 

Sephadex has been used to a lesser extent in the nucleic acid field. - The G-50 type will separate RNA of high molecular weight from nucleosides or bases. However, Sephadex does not separate the complex mixture of mono- to hexanucleotides obtained by enzymic hydrolysis of RNA. Sephadex-25 has been used as the supporting medium for partition chromatography of yeast-soluble RNA. ... 

Purine metabolites were measured via fluorimetric detection of the rate of H2O2 production during sequential catabolism of uric acid. H 02 oxidizes nonfluorescent dichlorofluorescin to fluorescent dichlorofluorescein under conditions that allow for detection of lOpmoles of purine substrate, Deoxy- and ribo-nucleosides may be partitioned by boronate affinity chromatography thus allowing their separate determination. 

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