Dibasic amines as competitive ions improve the resolution between polyanionic nucleotides.
Aliphatic diamines when used as single ion pairing reagents were capable of resolving 3'-,5'- and 2'-,5'- nucleotidyl diphosphates from one another while conventional ion pairing reagents did not separate these positional isomers. The use of 1,2-diamines resulted in the greatest resolution while increasing spacing between the amino groups progressively reduced the resolution while increasing the retention volume. A competitive ion pairing system was also developed using triethylamine as an additional ion pairing reagent. Using this system ethylenediamine, 1,2- and 1,3-diaminopropane were nearly equivalent in their ability to resolve adenosine 3'-phosphate 5'-phosphate, from adenosine 2'-phosphate 5'-phosphate, and adenosine 3'-phosphate 5'-beta-methylenephosphosulfate (3'-mePAPS) from adenosine 2'-phosphate 5'-beta-methylenephosphosulfate (2'-mePAPS), respectively. The ability to easily resolve these positional isomers allows the use of a more simplified synthetic procedure that does not involve the use selective protecting groups to specifically phosphorylate the 2' or 3' hydroxyl group. We have used this procedure on a semipreparative scale to obtain small quantities of both mePAPS and 2'-mePAPS for use in enzymatic studies.
Callahan
L
L
Division of Cytokine Biology, Food and Drug Administration, Bethesda, Maryland 20892.
Schwartz
N
N
Ng
K
K
D000227:Adenine Nucleotides / Q000032:analysis*; D000244:Adenosine Diphosphate / Q000032:analysis; D001667:Binding, Competitive; D002852:Chromatography, Ion Exchange; D003959:Diamines; D010724:Phosphoadenosine Phosphosulfate / Q000031:analogs & derivatives / Q000032:analysis
D000227:Adenine Nucleotides; D003959:Diamines; C072513:adenosine 3'-phosphate 5'-methylenephosphosulfate; C072514:adenosine 2'-phosphate 5'-methylenephosphosulfate; C045716:adenosine 2',5'-diphosphate; D010724:Phosphoadenosine Phosphosulfate; D000244:Adenosine Diphosphate; C010840:adenosine 3'-phosphate-5'-phosphate