Capillary electrophoretic separation of nucleotide isomers via complexation with cyclodextrin and borate.
The electrophoretic behaviour of monophosphorylated nucleotide isomers can be manipulated using complex-forming reactions with beta-cyclodextrin (beta-CD) and borate. Resolution of the 2'- and 3'-isomers of nucleotides is possible when the electrophoresis buffer contains 10 mM CD. The effect of beta-CD concentration on electrophoretic mobility is used to calculate the formation constant, K, of beta-CD-nucleotide complexes. The 3'-isomer of adenosine monophosphate (AMP) forms the strongest complex with beta-CD probably as a result of hydrogen bonding between the phosphate group of AMP and hydroxyls of beta-CD. In addition, complexation of 5'-nucleotides with borate increases the migration time window and leads to better separation. Complex-forming reactions of guanosine monophosphate and uridine monophosphate are shown to be strongly dependent on buffer pH. A mixture of 12 monophosphorylated nucleotides can be separated in less than 15 min using a buffer of 20 mM borate-10 mM beta-CD.
Department of Chemistry, McGill University, Montreal, Quebec, Canada.
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D001881:Borates / Q000737:chemistry*; D003505:Cyclodextrins / Q000737:chemistry*; D004586:Electrophoresis / Q000379:methods*; D006863:Hydrogen-Ion Concentration; D007536:Isomerism; D009711:Nucleotides / Q000737:chemistry / Q000302:isolation & purification*; D047392:beta-Cyclodextrins
D001881:Borates; D003505:Cyclodextrins; D009711:Nucleotides; D047392:beta-Cyclodextrins; C031215:betadex