Abstract
Dynamic metabolic flux analysis requires efficient and effective methods for extraction, purification and analysis of a plethora of naturally-occurring compounds. One area of metabolism that would be highly informative to study using metabolic flux analysis is the tricarboxylic acid (TCA) cycle, which consists of short-chain carboxylic acids. Here, we describe a newly-developed method for extraction, purification, derivatization and analysis of short-chain carboxylic acids involved in the TCA cycle. The method consists of snap-freezing the plant material, followed by maceration and a 12–15 h extraction at −80 °C. The extracts are then subject to reduction (to stabilize β-keto acids), purified by strong anion exchange solid phase extraction and methylated with methanolic HCl. This method could also be readily adapted to quantify many other short-chain carboxylic acids.
Original language | English (US) |
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Pages (from-to) | 165-174 |
Number of pages | 10 |
Journal | Journal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences |
Volume | 1028 |
DOIs | |
State | Published - Aug 15 2016 |
Bibliographical note
Publisher Copyright:© 2016 Elsevier B.V.
Keywords
- Carboxylic acid
- Gas chromatography-mass spectrometry
- Metabolic flux
- Methylation
- Polar metabolites
- Tricarboxylic acid cycle