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Updated: Feb 3, 2026

A Novel Saturation Mutagenesis Approach: Single Step Characterization of Regulatory Protein Binding Sites in RNA Using Phosphorothioates
Published on: August 21, 2018
Development of SPE method for the extraction of phosphorothioate oligonucleotides from serum samples
Łukasz Nuckowski1, Anna Kaczmarkiewicz1, Sylwia Studzińska1
1Chair of Environmental Chemistry & Bioanalytics, Faculty of Chemistry, Nicolaus Copernicus University in Toruń, 7 Gagarin Str, PL-87-100 Toruń, Poland.
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.
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