Development of an ion-pair reverse-phase liquid chromatographic/tandem mass spectrometry method for the determination of an 18-mer phosphorothioate oligonucleotide in mouse liver tissue.
A quantitative method for the determination of a partially modified, 2'-ribose alkoxy 18-mer phosphorothioate oligonucleotide, in liver tissue has been developed. A liquid:liquid extraction, ion-pair reverse phase chromatographic separation, and tandem mass spectrometry were used to achieve a quantitation range of 125 to 10,000 ng g(-1) mouse liver tissue. A total cycle time of 5 min was obtained while maintaining separation of three potential impurities. Separations were performed using a Discovery RP-Amide C16, 100 x 2 mm column packed with 5 microm particles. The separation was facilitated by the use of triethylamine (TEA) and hexafluoroisopropanol (HFIP) as ion-pair agents. The method has subsequently been used for the determination of other phosphorothioate oligonucleotides in support of discovery research.
Murphy
Anthony T
AT
Lilly Research Laboratories, Lilly Corporate Center, Indianapolis, IN 46285, USA. atm@lilly.com
Brown-Augsburger
Patricia
P
Yu
Rosie Z
RZ
Geary
Richard S
RS
Thibodeaux
Stefan
S
Ackermann
Bradley L
BL
D000818:Animals; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D008099:Liver / Q000737:chemistry / Q000378:metabolism*; D051379:Mice; D016376:Oligonucleotides, Antisense / Q000032:analysis* / Q000737:chemistry; D010710:Phosphates / Q000737:chemistry; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D013873:Thionucleotides / Q000032:analysis* / Q000737:chemistry
D016376:Oligonucleotides, Antisense; D010710:Phosphates; D013873:Thionucleotides; C083120:phosphorodithioic acid