Aim: Comprehensive development of a method for SPE extraction of antisense phosphorothioate oligonucleotide and its metabolites and their determination with the use of UHPLC.
Results: Polymer-based adsorbent and high percentage of methanol in elution solvent provided high recoveries compared with silica-based octadecyl cartridge. As to the type and concentration of ion pair reagent and organic solvent, the mixture of 5 mM of N,N-dimethylbutylamine/150 mM of 1,1,1,3,3,3-hexafluoroisopropanol and methanol was selected. Relatively high recoveries in the range of 79.2-81.2% with the SDs of 3.4-6.2% were obtained for the oligonucleotide and its metabolites extracted from human serum.
Conclusion: The developed method may be successfully applied for routine analysis of antisense oligonucleotides in serum since it is relatively easy, quick and reliable.
Keywords: SPE; antisense oligonucleotides; ion pair reagent; polymeric and silica cartridges; serum samples.