# A better understanding of carotenoid metabolism in tomato fruit by using the metabolomics capabilities of full-scan Orbitrap MS

Lieven Van Meulebroek (UGent) , Julie Vanden Bussche (UGent) , Kathy Steppe (UGent) and Lynn Vanhaecke (UGent)
Author
Organization
Project
IWT 101188
Abstract
Carotenoids are important secondary metabolites, which are mainly synthesized by plants, algae and certain types of bacteria and fungi. In epidemiological and clinical studies, associations were found between the intake of the concerned dietary compounds and the prevalence of chronic-degenerative diseases. Because of these beneficial health effects, carotenoids are intensively studied to better understand amongst others their partitioning and metabolism in plants. In this context, phytohormones might fulfil an important role since these compounds have a regulatory function in various plant processes. This study aimed for a better understanding of carotenoids’ metabolism by investigating the regulating role of phytohormones in the occurrence of tomato fruit carotenoids. The realization of the outlined objective completely fits into the foodomics concept, which is intended to be a global discipline that includes all of the emerging working areas in which food, analytical techniques and bioinformatics are combined. Completing this aim required the development of analytical methods for quantification of both carotenoids and phytohormones, present in tomato plant tissue. For the extraction of phytohormones (with a representative for each hormonal class), the developed protocol included solid liquid extraction with Bieleski solvent (methanol/water/formic acid), a purification step using a 30 kDa Amicon centrifugal filter unit, and an enrichment phase. Extracts were analyzed using UHPLC-Orbitrap ExactiveTM MS. The high resolution (up to 100,000 FWHM) and the full-scan principle of the applied mass spectrometer were particularly suited to realize a metabolomic approach. The protocol for extraction of the carotenoids (lutein, zeaxanthin, α-carotene, β-carotene, lycopene) was developed by means of a factorial d-optimal design and consisted of liquid-liquid extraction, using methyl-tert-butyl ether and methanol. Extracts were also analyzed by Orbitrap ExactiveTM MS. For chromatographic separation, a C30 HPLC-column was used. Both methods were successfully validated since linearity (R2 > 0.99), repeatability (< 15%), within-laboratory reproducibility (< 20%), specificity and recovery (> 85%) were satisfactory. Additionally, the obtained LOD (S/N ≥ 3) and LOQ (S/N ≥ 10) values were good for all targeted analytes. Subsequently, an experiment was set up in which tomato plants were subjected to either a control or drought treatment. The purpose was to trigger a stress response in the tomato plant, resulting in altered phytohormone and increased carotenoid concentration levels. Statistical interpretation of the analytical concentrations, obtained from these fruits, allows to investigate the relation between the concerned components. To this end, suited software programs, including SieveTM (fingerprinting) and SimcaTM (data analysis), have been applied. The different aspects of the described foodomic approach will be discussed.
Keywords
Phytohormones, Metabolomics, Orbitrap-MS, Carotenoids, Bio-informatics

## Citation

Chicago
Van Meulebroek, Lieven, Julie Vanden Bussche, Kathy Steppe, and Lynn Vanhaecke. 2013. “A Better Understanding of Carotenoid Metabolism in Tomato Fruit by Using the Metabolomics Capabilities of Full-scan Orbitrap MS.” In Recent Advances in Food Analysis, 6th International Symposium, Abstracts. Prague, Czech Republic: Institute of Chemical Technology.
APA
Van Meulebroek, L., Vanden Bussche, J., Steppe, K., & Vanhaecke, L. (2013). A better understanding of carotenoid metabolism in tomato fruit by using the metabolomics capabilities of full-scan Orbitrap MS. Recent Advances in Food Analysis, 6th International symposium, Abstracts. Presented at the 6th International symposium on Recent Advances in Food Analysis (RAFA 2013), Prague, Czech Republic: Institute of Chemical Technology.
Vancouver
1.
Van Meulebroek L, Vanden Bussche J, Steppe K, Vanhaecke L. A better understanding of carotenoid metabolism in tomato fruit by using the metabolomics capabilities of full-scan Orbitrap MS. Recent Advances in Food Analysis, 6th International symposium, Abstracts. Prague, Czech Republic: Institute of Chemical Technology; 2013.
MLA
Van Meulebroek, Lieven, Julie Vanden Bussche, Kathy Steppe, et al. “A Better Understanding of Carotenoid Metabolism in Tomato Fruit by Using the Metabolomics Capabilities of Full-scan Orbitrap MS.” Recent Advances in Food Analysis, 6th International Symposium, Abstracts. Prague, Czech Republic: Institute of Chemical Technology, 2013. Print.
@inproceedings{4252116,
abstract     = {Carotenoids are important secondary metabolites, which are mainly synthesized by plants, algae and certain types of bacteria and fungi. In epidemiological and clinical studies, associations were found between the intake of the concerned dietary compounds and the prevalence of chronic-degenerative diseases. Because of these beneficial health effects, carotenoids are intensively studied to better understand amongst others their partitioning and metabolism in plants. In this context, phytohormones might fulfil an important role since these compounds have a regulatory function in various plant processes.
This study aimed for a better understanding of carotenoids{\textquoteright} metabolism by investigating the regulating role of phytohormones in the occurrence of tomato fruit carotenoids. The realization of the outlined objective completely fits into the foodomics concept, which is intended to be a global discipline that includes all of the emerging working areas in which food, analytical techniques and bioinformatics are combined. Completing this aim required the development of analytical methods for quantification of both carotenoids and phytohormones, present in tomato plant tissue. For the extraction of phytohormones (with a representative for each hormonal class), the developed protocol included solid liquid extraction with Bieleski solvent (methanol/water/formic acid), a purification step using a 30 kDa Amicon centrifugal filter unit, and an enrichment phase. Extracts were analyzed using UHPLC-Orbitrap ExactiveTM MS. The high resolution (up to 100,000 FWHM) and the full-scan principle of the applied mass spectrometer were particularly suited to realize a metabolomic approach. The protocol for extraction of the carotenoids (lutein, zeaxanthin, \ensuremath{\alpha}-carotene, \ensuremath{\beta}-carotene, lycopene) was developed by means of a factorial d-optimal design and consisted of liquid-liquid extraction, using methyl-tert-butyl ether and methanol. Extracts were also analyzed by Orbitrap ExactiveTM MS. For chromatographic separation, a C30 HPLC-column was used. Both methods were successfully validated since linearity (R2 {\textrangle} 0.99), repeatability ({\textlangle} 15\%), within-laboratory reproducibility ({\textlangle} 20\%), specificity and recovery ({\textrangle} 85\%) were satisfactory. Additionally, the obtained LOD (S/N \ensuremath{\geq} 3) and LOQ (S/N \ensuremath{\geq} 10) values were good for all targeted analytes. Subsequently, an experiment was set up in which tomato plants were subjected to either a control or drought treatment. The purpose was to trigger a stress response in the tomato plant, resulting in altered phytohormone and increased carotenoid concentration levels. Statistical interpretation of the analytical concentrations, obtained from these fruits, allows to investigate the relation between the concerned components. To this end, suited software programs, including SieveTM (fingerprinting) and SimcaTM (data analysis), have been applied. The different aspects of the described foodomic approach will be discussed.},
author       = {Van Meulebroek, Lieven and Vanden Bussche, Julie and Steppe, Kathy and Vanhaecke, Lynn},
booktitle    = {Recent Advances in Food Analysis, 6th International symposium, Abstracts},
isbn         = {9788070808610},
language     = {eng},
location     = {Prague, Czech Republic},
publisher    = {Institute of Chemical Technology},
title        = {A better understanding of carotenoid metabolism in tomato fruit by using the metabolomics capabilities of full-scan Orbitrap MS},
year         = {2013},
}