QUANTITATIVE-ANALYSIS OF MODIFIED ANTISENSE OLIGONUCLEOTIDES IN BIOLOGICAL-FLUIDS USING CATIONIC NANOPARTICLES FOR SOLID-PHASE EXTRACTION

Based on a novel method for solid-phase extraction using cationic poly styrene nanoparticles, the suitability of the extraction procedure for quantitation of terminally and backbone-modified antisense oligonucle otides was investigated, Extractions were carried out from both human plasma and urine. Quantitative analysis of the extracted samples was p erformed with capillary gel electrophoresis. In accordance with previo us results obtained with phosphorothioate oligonucleotides in human pl asma, high linearity and accuracy of the assay was demonstrated for an oligodeoxyribonucleotide-palmityl conjugate as well as for a modified oligoribonucleotide. Optimized extraction conditions allow the isolat ion of oligonucleotides in high yields and purity even for concentrati ons in the low nanomolar range, down to 5 nM. Comparing the results ob tained from human plasma and urine, no significant differences in the absolute recovery rates which reach values up to 95% were observed. Ho wever, when the loading capacity of the nanoparticles was exceeded, se lective recovery was observed for the coisolation of phosphodiester an d phosphorothioate oligonucleotides. This effect can be explained by d ifferences in the attractive forces between PO- and PS-oligonucleotide s and the particle surface and appears to be valuable for a modificati on-dependent enrichment of oligonucleotides out of complex mixtures. ( C) 1998 Academic Press.