CAPILLARY ELECTROPHORETIC SEPARATION OF NUCLEOTIDE ISOMERS VIA COMPLEXATION WITH CYCLODEXTRIN AND BERATE

The electrophoretic behaviour of monophosphorylated nucleotide isomers can be manipulated using complex-forming reactions with beta-cyclodex trin (beta-CD) and berate. Resolution of the 2'- and 3'-isomers of nuc leotides is possible when the electrophoresis buffer contains 10 mM CD . The effect of beta-CD concentration on electrophoretic mobility is u sed to calculate the formation constant, K, of beta-CD-nucleotide comp lexes. The 3'-isomer of adenosine monophosphate (AMP) forms the strong est complex with beta-CD probably as a result of hydrogen bonding betw een the phosphate group of AMP and hydroxyls of beta-CD. In addition, complexation of 5'-nucleotides with berate increases the migration tim e window and leads to better separation. Complex-forming reactions of guanosine monophosphate and uridine monophosphate are shown to be stro ngly dependent on buffer pH. A mixture of 12 monophosphorylated nucleo tides can be separated in less than 15 min using a buffer of 20 mM bor ate-10 mM beta-CD.