High-performance anion-exchange chromatography using on-line electrolytic eluent generation for the determination of more than 25 intermediates from energy metabolism of mammalian cells in culture

Ritter, Joachim; Genzel, Yvonne; Reichl, Udo

doi:10.1016/j.jchromb.2006.06.004

Cited by 34 publications

(30 citation statements)

References 44 publications

(79 reference statements)

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“…High performance anion exchange chromatography using a CarboPac PA1 column separated 20 nucleotides and sugar nucleotides, although UDP-GlcNAc and UDP-GalNAc peaks overlapped in that 55 min method [7]. Ion chromatography afforded improved separation of UDP-GlcNAc and UDPGalNAc in a 65 min method [8]. Lectin affinity chromatography showed significant improvements in discrimination between epimers, but has not been widely adopted [9].…”

Section: Introductionmentioning

confidence: 99%

Enzymatic analysis of uridine diphosphate N-acetyl-d -glucosamine

Namboori

Graham

2008

Analytical Biochemistry

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The Methanococcus maripaludis MMP0352 protein belongs to an oxidoreductase family that has been proposed to catalyze the NAD + -dependent oxidation of the 3″-position of uridine diphosphate N-acetyl-D-glucosamine (UDP-GlcNAc), forming a 3-hexulose sugar nucleotide. The heterologously expressed MMP0352 protein was purified and shown to efficiently catalyze UDP-GlcNAc oxidation forming one NADH equivalent. This enzyme was used to develop a fixed endpoint fluorometric method to analyze UDP-GlcNAc. The enzyme is highly specific for this acetamido sugar nucleotide, and the procedure had a detection limit of 0.2 μM UDP-GlcNAc in a 1-ml sample. Using the method of standard addition, UDP-GlcNAc concentrations were measured in deproteinized extracts of Escherichia coli, Saccharomyces cerevisiae and HeLa carcinoma cells. Equivalent concentrations were determined by both enzymatic and chromatographic analyses, validating this method. This procedure can be adapted for the high-throughput analysis of changes in cellular UDP-GlcNAc concentrations during time series experiments or inhibitor screens.

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Section: Introductionmentioning

confidence: 99%

Enzymatic analysis of uridine diphosphate N-acetyl-d -glucosamine

Namboori

Graham

2008

Analytical Biochemistry

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“…Ritter et al [45] described the hyphenation of an electrolytic on-line eluent generation device with high performance anion-exchange chromatography coupled with UV detection for the determination of a wide range of intracellular metabolites from mammalian cells. The detection wavelength of the UV detector was switched from 220 to 260 nm and the detection limits were in the range of mM.…”

Section: Hyphenation In Liquid Chromatographymentioning

confidence: 99%

Hyphenation in sample preparation: Advancement from the micro to the nano world

Ali

Gupta

Aboul‐Enein³

et al. 2008

J of Separation Science

116

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Analysis at trace levels, an ideal area of application for hyphenated techniques, is steadily gaining importance. Many sample pre-concentration and clean-up methods have been hyphenated with core analytical techniques to accomplish the task of low level detection. The present article describes the state of the art of hyphenation of various techniques such as solid phase extraction, micro-solid phase extraction, dialysis, and chromatographic modalities etc. with liquid chromatography, gas chromatography, capillary electrophoresis, and spectroscopic methods. Besides, attempts have been made to address the hyphenation approach in microfluidic devices.

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“…Traditionally, laborious enzymatic assays [5][6][7][8][9] and HPLC-ultraviolet light detection [10] have been used to determine the concentrations of one or several analytes. However, mass spectrometric (MS) detection offers superior selectivity and the simultaneous profiling of tens of metabolites.…”

Section: Introductionmentioning

confidence: 99%