Analysis of phosphorylated carbohydrates by high-performance liquid chromatography–electrospray ionization tandem mass spectrometry utilising a β-cyclodextrin bonded stationary phase

• Published in: Journal of Chromatography A Vol. 803; no. 1; pp. 111 - 119
• Main Authors: Feurle, J., Jomaa, H., Wilhelm, M., Gutsche, B., Herderich, M.
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier B.V 17.04.1998; Elsevier
• Subjects: Analytical chemistry; Carbohydrates; Chemistry; Chromatographic methods and physical methods associated with chromatography; D-1-Deoxyxylulose-5-phosphate; Exact sciences and technology; Other chromatographic methods; Sugar phosphates; D-1-Deoxyxylulose-5-phosphate; Carbohydrates; Sugar phosphates; Hexose; Chemical analysis; Aldose; Enzyme; Coupled method; Transferases; Ketose; Reaction product; Transketolase; HPLC chromatography; Pyruvate; Fragmentation pattern; Glycosyl phosphate; Phosphorus Organic compounds; Electrospray; Cyclodextrin; Pentose; Mass spectrometry MS/MS; Organic phosphate; Qualitative analysis; Bonded stationary phase; Enzymatic reaction
• ISSN: 0021-9673
• DOI: 10.1016/S0021-9673(97)01214-4

Chromatographic separation of phosphorylated carbohydrates was achieved on a β-cyclodextrin (CD) bonded HPLC column. Applying acetonitrile–aqueous ammonium acetate gradient elution, the separation of sugar phosphates relied on the combination of anion-exchange and hydrophilic interaction properties of CD bonded phases in the presence of ammonium ions. Furthermore, this solvent system allowed on-line coupling to electrospray ionization mass spectrometry (ESI-MS) and direct structural characterisation of carbohydrate phosphates by tandem mass spectrometry (MS–MS). In addition to the analysis of various pentose and hexose phosphates, the HPLC–ESI-MS–MS method was successfully applied to demonstrate the enzymatic formation of D-1-deoxyxylulose 5-phosphate from pyruvate and glyceraldehyde phosphate catalysed by yeast transketolase.