Abstract
Ultrahigh-performance liquid chromatography has been used for the separation and analysis of unmodified and modified antisense oligonucleotides. For this reason, we tested various columns of low particle sizes in our analysis of unmodified and phosphorothioate oligonucleotides. The influence of both the type and concentration of ion-pair reagent on the retention of the studied biomolecules was tested. The developed methods were used for separation of unmodified oligonucleotides and to determine antisense oligonucleotides in human serum samples. The results proved that octadecyl and phenyl columns are the most selective in the resolution of oligonucleotides which differ in the position of single nucleotides in the sequence. The phenyl column was selected and applied for the analysis of phosphorothioate oligonucleotides in serum samples. The calibration plots showed good linearity within the test concentration ranges. The intra-day CV of the calibration curve slopes was in the range of 1.6 to 4.2 %. The limits of detection (LODs) were in the range of 0.11–0.16 μg mL−1, while the limit of quantification (LOQ) values were between 0.35 and 0.51 μg mL−1. FigureDetermination of antisense phosphorothioate oligonucleotides in serum
Highlights
Oligonucleotides are components of nucleic acids, which are built of nucleotide monomers [1, 2]
They may be divided into unmodified and modified oligonucleotides [1, 2]. The latter are synthetic compounds with a modification of either the nitrogen base or phosphate group. These alterations may be connected by introduction of the methyl group at each phosphorus in the oligonucleotide chain or at the bases; methylphosphonates are synthesized [1]
Two groups of oligonucleotides were chosen for the present investigation: unmodified and modified ones (Table 1)
Summary
Oligonucleotides are components of nucleic acids, which are built of nucleotide monomers [1, 2] They play an important role in most living organisms. They may be divided into unmodified and modified oligonucleotides [1, 2] The latter are synthetic compounds with a modification of either the nitrogen base or phosphate group. Phosphorothioates are the most popular [1] Their structure is modified by the replacement of one of the nonbridging oxygens by sulfur at each phosphorus in the chain. Such modifications improve their enzymatic stability and cell membrane permeability, as phosphorothioate oligonucleotides are used in antisense therapy [1, 2]. They are applied for the synthesis of deoxyribonucleic and ribonucleic acids or for polymerase chain reactions [1, 2]
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
