Two-dimensional liquid chromatography-mass spectrometry for the characterization of modified oligonucleotide impurities

Oligonucleotide analysis by hydrophilic interaction liquid chromatography-mass spectrometry in the absence of ion-pair reagents

“…Alternatives to the traditional IP-RP-LC of oligonucleotides have been explored over the years, including capillary gel electrophoresis [29,30], size exclusion [31,32], anion exchange [30,33,34], mixed-mode [35], monolithic columns [36,37], non-porous polymeric particles [38][39][40], phenyl, fluorophenyl and cyano stationary phases [13,41,42], and various combinations of these techniques in multi-dimensional approaches [34,43]. A promising alternative to IP-RP for LC-MS analysis is hydrophilic interaction liquid chromatography (HILIC) [44].…”

Recent Developments in Two-Dimensional Liquid Chromatography: Fundamental Improvements for Practical Applications

“…Finally, Roussis et al have demonstrated the use of mLC-LC and LC × LC separations involving different combinations of AEX, RP, and IP-RP separation modes to identify impurities present in synthetic oligonucleotides as well as degradants observed in stability studies. 47 This work provides a compelling example of the ability of 2D-LC to achieve separations “...not currently possible by mass spectrometry alone or 1D-LC”.…”

Two‐dimensional liquid chromatography analysis of all‐trans‐, 9‐cis‐, and 13‐cis‐astaxanthin in raw extracts from Phaffia rhodozyma

“…Hypersil Gold C18 column (100 mm × 2.1 mm, 1.9 µm) from Thermo Fisher Scientific (San Jose, CA, USA) was used as the first column in the 2-D-LC system. The mobile phase was composed of methanol (A) and ultrapure water (B) as described in previous studies [22,29]. The elution program had the following parameters: 0 min 20%B, 4 min 0%B, 5-7 min 90%B, 9-12 min 20%B.…”

“…In recent years, 2-D-LC has been developed to analyze target compounds from complex solution/extracts. This chromatographic method employs two different columns that have different separation characteristics, and whereby components are subjected to two distinct separative displacements that dramatically increase the separation effectiveness in comparison to 1-D HPLC [22,23]. Generally, two kinds of online patterns, including comprehensive 2-D-LC and heart-cutting 2-D-LC, are used for an efficient metabolite analysis.…”

“…In heart cutting 2-D-LC pattern, only parts of the eluents from the first column are chosen to transfer to the second column for further separation. Both comprehensive and heart cutting 2-D-LC approaches are very powerful for the determination of target compounds in complex matrices [22][23][24]. For instance, a comprehensive 2-D-LC method has been successfully used to separate 23 different carotenoids in the fruit of Red Mamey (Pouteria sapote) [25]; similarly, a heart cutting 2-D-LC was successfully exploited to determine tropane alkaloids in herbal medicines [26].…”