Ultraperformance Ion-Pair Liquid Chromatography Coupled to Electrospray Time-of-Flight Mass Spectrometry for Compositional Profiling and Quantification of Heparin and Heparan Sulfate

2008 ◽  
Vol 80 (4) ◽  
pp. 1297-1306 ◽  
Author(s):  
Albert K. Korir ◽  
John F. K. Limtiaco ◽  
Sarah M. Gutierrez ◽  
Cynthia K. Larive
Keyword(s):  
Mass Spectrometry ◽  
Liquid Chromatography ◽  
Heparan Sulfate ◽  
Time Of Flight ◽  
Ion Pair ◽  
Flight Mass Spectrometry

scholarly journals Studying in vitro metabolism of the first and second generation of antisense oligonucleotides with the use of ultra-high-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry

2020 ◽  
Vol 412 (27) ◽  
pp. 7453-7467
Author(s):  
Anna Kilanowska ◽  
Łukasz Nuckowski ◽  
Sylwia Studzińska
Keyword(s):  
Mass Spectrometry ◽  
Liquid Chromatography ◽  
Liver Microsomes ◽  
Time Of Flight ◽  
Ion Pair ◽  
In Vitro Metabolism ◽  
Detection Methods ◽  
Ion Pair Chromatography ◽  
Flight Mass Spectrometry

Abstract The aim of the present investigation was the analysis and identification of antisense oligonucleotide metabolism products after incubation with human liver microsomes regarding four different oligonucleotide modifications. Separation and detection methods based on the use of liquid chromatography coupled with quadrupole time-of-flight mass spectrometry were developed for this purpose. Firstly, the optimization of mass spectrometer parameters was done to select those which ensure the highest possible sensitivity of oligonucleotide analysis. This step was conducted for two chromatographic modes—ion pair chromatography and hydrophilic interaction liquid chromatography—due to their common application in oligonucleotide analysis. Based on sensitivity results, ion pair chromatography coupled with mass spectrometry was selected for the separation of model oligonucleotide mixtures in order to verify its selectivity for N-deleted metabolite separation. Next, the developed method was applied in the examination of oligonucleotides in vitro metabolism. First, wide optimization of incubation parameters was conducted including the concentration of the reaction buffer components. Obtained results indicated that both 3′-exonucleases and 5′-exonucleases contributed to the biotransformation of oligonucleotides. Moreover, it may be concluded that the number of metabolites depends on oligonucleotide modification and consequently its resistance to enzymatic attack. Thus, the number of the oligonucleotide metabolites decreased with the decrease of the resultant polarity of oligonucleotide caused by chemical modification.

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