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A UPLC‐MS/MS method for quantification of metabolites in the ethylene biosynthesis pathway and its biological validation in Arabidopsis

(2024) NEW PHYTOLOGIST. 243(3). p.1262-1275
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Abstract
The plant hormone ethylene is of vital importance in the regulation of plant development and stress responses. Recent studies revealed that 1-aminocyclopropane-1-carboxylic acid (ACC) plays a role beyond its function as an ethylene precursor. However, the absence of reliable methods to quantify ACC and its conjugates malonyl-ACC (MACC), glutamyl-ACC (GACC), and jasmonyl-ACC (JA-ACC) hinders related research. Combining synthetic and analytical chemistry, we present the first, validated methodology to rapidly extract and quantify ACC and its conjugates using ultra-high-performance liquid chromatography coupled to tandem mass spectrometry (UPLC-MS/MS). Its relevance was confirmed by application to Arabidopsis mutants with altered ACC metabolism and wild-type plants under stress. Pharmacological and genetic suppression of ACC synthesis resulted in decreased ACC and MACC content, whereas induction led to elevated levels. Salt, wounding, and submergence stress enhanced ACC and MACC production. GACC and JA-ACC were undetectable in vivo; however, GACC was identified in vitro, underscoring the broad applicability of the method. This method provides an efficient tool to study individual functions of ACC and its conjugates, paving the road toward exploration of novel avenues in ACC and ethylene metabolism, and revisiting ethylene literature in view of the recent discovery of an ethylene-independent role of ACC.
Keywords
1-aminocyclopropane-1-carboxylic acid, ethylene biosynthesis, glutamyl-ACC, jasmonyl-ACC, malonyl-ACC, method validation, synthetic chemistry of standards, UPLC-MS/MS quantification method, 1-AMINOCYCLOPROPANE-1-CARBOXYLIC ACID, LIQUID-CHROMATOGRAPHY, 1-(MALONYLAMINO)CYCLOPROPANE-1-CARBOXYLIC ACID, QUANTITATIVE-ANALYSIS, MAJOR CONJUGATE, IDENTIFICATION, PRECURSOR, PHYTOHORMONES, ISOLEUCINE, SYNTHASE

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MLA
Cao, Da, et al. “A UPLC‐MS/MS Method for Quantification of Metabolites in the Ethylene Biosynthesis Pathway and Its Biological Validation in Arabidopsis.” NEW PHYTOLOGIST, vol. 243, no. 3, 2024, pp. 1262–75, doi:10.1111/nph.19878.
APA
Cao, D., Depaepe, T., Sanchez Muñoz, R., Janssens, H., Lemière, F., Willems, T., … Van Der Straeten, D. (2024). A UPLC‐MS/MS method for quantification of metabolites in the ethylene biosynthesis pathway and its biological validation in Arabidopsis. NEW PHYTOLOGIST, 243(3), 1262–1275. https://doi.org/10.1111/nph.19878
Chicago author-date
Cao, Da, Thomas Depaepe, Raul Sanchez Muñoz, Hilde Janssens, Filip Lemière, Tim Willems, Johan Winne, Els Prinsen, and Dominique Van Der Straeten. 2024. “A UPLC‐MS/MS Method for Quantification of Metabolites in the Ethylene Biosynthesis Pathway and Its Biological Validation in Arabidopsis.” NEW PHYTOLOGIST 243 (3): 1262–75. https://doi.org/10.1111/nph.19878.
Chicago author-date (all authors)
Cao, Da, Thomas Depaepe, Raul Sanchez Muñoz, Hilde Janssens, Filip Lemière, Tim Willems, Johan Winne, Els Prinsen, and Dominique Van Der Straeten. 2024. “A UPLC‐MS/MS Method for Quantification of Metabolites in the Ethylene Biosynthesis Pathway and Its Biological Validation in Arabidopsis.” NEW PHYTOLOGIST 243 (3): 1262–1275. doi:10.1111/nph.19878.
Vancouver
1.
Cao D, Depaepe T, Sanchez Muñoz R, Janssens H, Lemière F, Willems T, et al. A UPLC‐MS/MS method for quantification of metabolites in the ethylene biosynthesis pathway and its biological validation in Arabidopsis. NEW PHYTOLOGIST. 2024;243(3):1262–75.
IEEE
[1]
D. Cao et al., “A UPLC‐MS/MS method for quantification of metabolites in the ethylene biosynthesis pathway and its biological validation in Arabidopsis,” NEW PHYTOLOGIST, vol. 243, no. 3, pp. 1262–1275, 2024.
@article{01HZYN67644EQW3G9SVC8KAR3W,
  abstract     = {{The plant hormone ethylene is of vital importance in the regulation of plant development and stress responses. Recent studies revealed that 1-aminocyclopropane-1-carboxylic acid (ACC) plays a role beyond its function as an ethylene precursor. However, the absence of reliable methods to quantify ACC and its conjugates malonyl-ACC (MACC), glutamyl-ACC (GACC), and jasmonyl-ACC (JA-ACC) hinders related research.
Combining synthetic and analytical chemistry, we present the first, validated methodology to rapidly extract and quantify ACC and its conjugates using ultra-high-performance liquid chromatography coupled to tandem mass spectrometry (UPLC-MS/MS). Its relevance was confirmed by application to Arabidopsis mutants with altered ACC metabolism and wild-type plants under stress.
Pharmacological and genetic suppression of ACC synthesis resulted in decreased ACC and MACC content, whereas induction led to elevated levels. Salt, wounding, and submergence stress enhanced ACC and MACC production. GACC and JA-ACC were undetectable in vivo; however, GACC was identified in vitro, underscoring the broad applicability of the method.
This method provides an efficient tool to study individual functions of ACC and its conjugates, paving the road toward exploration of novel avenues in ACC and ethylene metabolism, and revisiting ethylene literature in view of the recent discovery of an ethylene-independent role of ACC.}},
  author       = {{Cao, Da and Depaepe, Thomas and Sanchez Muñoz, Raul and Janssens, Hilde and Lemière, Filip and Willems, Tim and Winne, Johan and Prinsen, Els and Van Der Straeten, Dominique}},
  issn         = {{0028-646X}},
  journal      = {{NEW PHYTOLOGIST}},
  keywords     = {{1-aminocyclopropane-1-carboxylic acid,ethylene biosynthesis,glutamyl-ACC,jasmonyl-ACC,malonyl-ACC,method validation,synthetic chemistry of standards,UPLC-MS/MS quantification method,1-AMINOCYCLOPROPANE-1-CARBOXYLIC ACID,LIQUID-CHROMATOGRAPHY,1-(MALONYLAMINO)CYCLOPROPANE-1-CARBOXYLIC ACID,QUANTITATIVE-ANALYSIS,MAJOR CONJUGATE,IDENTIFICATION,PRECURSOR,PHYTOHORMONES,ISOLEUCINE,SYNTHASE}},
  language     = {{eng}},
  number       = {{3}},
  pages        = {{1262--1275}},
  title        = {{A UPLC‐MS/MS method for quantification of metabolites in the ethylene biosynthesis pathway and its biological validation in Arabidopsis}},
  url          = {{http://doi.org/10.1111/nph.19878}},
  volume       = {{243}},
  year         = {{2024}},
}

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