Aqueous normal phase retention of nucleotides on silica hydride-based columns: Method development strategies for analytes revelant in clinical analysis
✍ Scribed by Maria T. Matyska; Joseph J. Pesek; John Duley; Mazin Zamzami; Steven M. Fischer
- Book ID
- 102443857
- Publisher
- John Wiley and Sons
- Year
- 2010
- Tongue
- English
- Weight
- 304 KB
- Volume
- 33
- Category
- Article
- ISSN
- 1615-9306
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✦ Synopsis
Abstract
An aqueous normal phase HPLC method coupled with UV or ESI/MS detection was used for the determination of a wide variety of nucleotides, essential in metabolomics studies. Fifteen nucleotides were tested in clinically relevant mixtures at levels of 100 μg/mL for UV detection and 1 μg/mL for ESI‐MS detection. Analysis times for all protocols developed were less than 20 min. The chromatographic conditions were changed to achieve optimized retention and separation of the nucleotides studied. The aqueous normal phase‐HPLC methods were developed utilizing two columns, one having a minimally modified hydride surface another having an undecanoic acid moiety on a hydride surface. Volatile, low ionic strength mobile phases were used. Negative ion mode ESI‐MS at near neutral pH mobile phase, combined with a TOF detector provided a highly sensitive and specific method, which is equally suitable for quadrupole and ion trap instruments.