Journal article
Factors influencing the separation of oligonucleotides using reversed-phase/ion-exchange mixed-mode high performance liquid chromatography columns
Journal of Chromatography A, v 1304, pp 69-77
23 Aug 2013
PMID: 23859796
Featured in Collection : UN Sustainable Development Goals @ Drexel
Abstract
•Mixed-mode stationary phases were evaluated for the analysis of oligonucleotides.•A single column and method can be used for complete oligonucleotide analysis.•Mixed-mode columns with SAX-LC conditions provided the best separation.•SM-C18 column with NaCl salt gradient provided the best peak shape and separation.
New mixed-mode columns consisting of reversed-phase and ion-exchange separation modes were evaluated for the analysis of short RNA oligonucleotides (∼20mers). Conventional analysis for these samples typically involves using two complementary methods: strong anion-exchange liquid chromatography (SAX-LC) for separation based on charge, and ion-pair reversed-phase liquid chromatography (IP-RPLC) for separation based on hydrophobicity. Recently introduced mixed-mode high performance liquid chromatography (HPLC) columns combine both reversed-phase and ion-exchange modes, potentially offering a simpler analysis by combining the benefits of both separation modes into a single method. Analysis of a variety of RNA oligonucleotide samples using three different mixed-mode stationary phases showed some distinct benefits for oligonucleotide separation and analysis. When using these mixed-mode columns with typical IP-RPLC mobile phase conditions, such as ammonium acetate or triethylammonium acetate as the primary ion-pair reagent, the separation was mainly based on the IP-RPLC mode. However, when changing the mobile phase conditions to those more typical for SAX-LC, such as salt gradients with NaCl or NaBr, very different separation patterns were observed due to mixed-mode interactions. In addition, the Scherzo SW-C18 and SM-C18 columns with sodium chloride or sodium bromide salt gradients also showed significant improvements in peak shape.
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Details
- Title
- Factors influencing the separation of oligonucleotides using reversed-phase/ion-exchange mixed-mode high performance liquid chromatography columns
- Creators
- Mirlinda Biba - Merck Research Laboratories, Rahway, NJ 07065, USAEileen Jiang - Merck Research Laboratories, Rahway, NJ 07065, USABing Mao - Merck Research Laboratories, Rahway, NJ 07065, USADaniel Zewge - Merck Research Laboratories, Rahway, NJ 07065, USAJoe P Foley - Drexel University, Philadelphia, PA 19101, USAChristopher J Welch - Merck Research Laboratories, Rahway, NJ 07065, USA
- Publication Details
- Journal of Chromatography A, v 1304, pp 69-77
- Publisher
- Elsevier
- Resource Type
- Journal article
- Language
- English
- Academic Unit
- Chemistry
- Web of Science ID
- WOS:000323017500009
- Scopus ID
- 2-s2.0-84881027279
- Other Identifier
- 991014878442904721
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- Collaboration types
- Industry collaboration
- Domestic collaboration
- Web of Science research areas
- Biochemical Research Methods
- Chemistry, Analytical