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cis/trans Carotenoid Extraction, Purification, Detection, Quantification, and Profiling in Plant Tissues

  • Yagiz Alagoz
  • Namraj Dhami
  • Chris Mitchell
  • Christopher I. CazzonelliEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 2083)

Abstract

Reverse phase high-performance liquid chromatography (HPLC) is the method of choice used in biological, health, and food research to identify, quantify, and profile carotenoid species. The identification and quantification of cis- and/or trans-carotene and xanthophyll isomers in plant tissues can be affected by the method of sample preparation and extraction, as well as the HPLC column chemistry and the solvent gradient. There is a high degree of heterogeneity in existing methods in terms of their ease, efficiency, and accuracy. We describe a simple carotenoid extraction method and two different optimised HPLC methods utilizing C18 or C30 reverse-phase columns. We outline applications, advantages, and disadvantages for using these reverse phase columns to detect xanthophylls and cis-carotenes in wild-type photosynthetic leaves and mutant dark-grown etiolated seedlings, respectively. Resources are provided to profile individual species based upon their spectral properties and retention time, as well as quantify carotenoids by their composition and absolute levels in different plant tissues.

Key words

Carotenoid cis-carotene Molar coefficient Composition Plants HPLC C30 C18 

Notes

Acknowledgments

This work is supported by Australian Research Council Discovery Grant DP130102593 (to C.I.C). We acknowledge the Western Sydney University and Hawkesbury Institute for the Environment for providing a Ph.D. scholarship award to Y.A. and N.D. and financial support to purchase chemicals required for this research.

Author contributions: Y.A. wrote the methods chapter, prepared figures and tables. Y.A., N.D. and C.M. optimised methods for carotenoid extraction and HPLC separation. C.I.C outlined and edited the methods chapter. C.I.C supervised Y.A. and N.D. All authors contributed to editing this chapter.

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2020

Authors and Affiliations

  • Yagiz Alagoz
    • 1
  • Namraj Dhami
    • 1
  • Chris Mitchell
    • 1
  • Christopher I. Cazzonelli
    • 1
    Email author
  1. 1.Hawkesbury Institute for the EnvironmentWestern Sydney UniversityPenrithAustralia

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