Extraction, purification, methylation and GC–MS analysis of short-chain carboxylic acids for metabolic flux analysis

Nathan D. Tivendale; Erin M. Jewett; Adrian D. Hegeman; Jerry D. Cohen

doi:10.1016/j.jchromb.2016.05.042

Extraction, purification, methylation and GC–MS analysis of short-chain carboxylic acids for metabolic flux analysis

Nathan D. Tivendale, Erin M. Jewett, Adrian D. Hegeman, Jerry D. Cohen

Horticultural Science

Research output: Contribution to journal › Article › peer-review

1 Scopus citations

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)
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	https://doi.org/10.1016/j.jchromb.2016.05.042
State	Published - Aug 15 2016

Bibliographical note

Funding Information:
This research was funded by the National Science Foundation ( IOS-1238812 ). We further acknowledge supplemental support from the Gordon and Margaret Bailey Endowment for Environmental Horticulture and the Minnesota Agricultural Experiment Station.

Publisher Copyright:
© 2016 Elsevier B.V.

Copyright:
Copyright 2016 Elsevier B.V., All rights reserved.

Keywords

Carboxylic acid
Gas chromatography-mass spectrometry
Metabolic flux
Methylation
Polar metabolites
Tricarboxylic acid cycle

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Extraction, purification, methylation and GC–MS analysis of short-chain carboxylic acids for metabolic flux analysis. / Tivendale, Nathan D.; Jewett, Erin M.; Hegeman, Adrian D.; Cohen, Jerry D.

In: Journal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences, Vol. 1028, 15.08.2016, p. 165-174.

Research output: Contribution to journal › Article › peer-review

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title = "Extraction, purification, methylation and GC–MS analysis of short-chain carboxylic acids for metabolic flux analysis",

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.",

keywords = "Carboxylic acid, Gas chromatography-mass spectrometry, Metabolic flux, Methylation, Polar metabolites, Tricarboxylic acid cycle",

author = "Tivendale, {Nathan D.} and Jewett, {Erin M.} and Hegeman, {Adrian D.} and Cohen, {Jerry D.}",

note = "Funding Information: This research was funded by the National Science Foundation ( IOS-1238812 ). We further acknowledge supplemental support from the Gordon and Margaret Bailey Endowment for Environmental Horticulture and the Minnesota Agricultural Experiment Station. Publisher Copyright: {\textcopyright} 2016 Elsevier B.V. Copyright: Copyright 2016 Elsevier B.V., All rights reserved.",

year = "2016",

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doi = "10.1016/j.jchromb.2016.05.042",

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AU - Jewett, Erin M.

AU - Hegeman, Adrian D.

AU - Cohen, Jerry D.

N1 - Funding Information: This research was funded by the National Science Foundation ( IOS-1238812 ). We further acknowledge supplemental support from the Gordon and Margaret Bailey Endowment for Environmental Horticulture and the Minnesota Agricultural Experiment Station. Publisher Copyright: © 2016 Elsevier B.V. Copyright: Copyright 2016 Elsevier B.V., All rights reserved.

PY - 2016/8/15

Y1 - 2016/8/15

N2 - 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.

AB - 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.

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KW - Metabolic flux

KW - Methylation

KW - Polar metabolites

KW - Tricarboxylic acid cycle

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ER -

Extraction, purification, methylation and GC–MS analysis of short-chain carboxylic acids for metabolic flux analysis

Abstract

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Keywords

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