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ARS Home » Midwest Area » Columbia, Missouri » Plant Genetics Research » Research » Publications at this Location » Publication #414033
Research Project: Adaptation of Grain Crops to Varying Environments Including Climates, Stressors, and Human Uses

Location: Plant Genetics Research

Title: Sterol and brassinosteroid hormone quantification by LC/MS of picolinyl ester derivatives

Author
item DILKES, BRIAN - Purdue University
item Best, Norman

Submitted to: Cold Spring Harbor Protocols
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 2/21/2025
Publication Date: 4/17/2025
Citation: Dilkes, B.P., Best, N.B. 2025. Sterol and brassinosteroid hormone quantification by LC/MS of picolinyl ester derivatives. Cold Spring Harbor Protocols. https://doi.org/10.1101/pdb.prot108646.
DOI: https://doi.org/10.1101/pdb.prot108646

Interpretive Summary: Brassinosteroids are small plant hormones that regulate all aspects of plant growth and development. They are in very small abundance in maize which makes them difficult to measure in plant tissue. They are neutrally charged compounds that makes it difficult to identify them with mass spectrometry techniques. We have developed a mass spectrometry method that allows for these plant hormones to be accurately measured at low concentrations. This manuscript lays out a step-by-step method to extract, derivatize, and measure brassinosteroids in a small amount of maize tissue. We provide the masses and retention times for a number of sterols and brassinosteroids that were identified by us. This provides a novel method for the research community to measure brassinosteroids in plant tissue to conduct genetic and biochemical experiments.

Technical Abstract: Brassinosteroids are small steroidal hormones that regulate plant growth, differentiation, and defense. They are low abundance in plant tissues and are difficult to assess via mass spectrometry due to poor ionization. In this protocol, we provide a method for the extraction, detection, and quantification of a subset of sterol and brassinosteroid metabolites using a derivatization method to improve ionization during liquid chromatography coupled with mass spectrometry. Multiple reaction monitoring, which is the utilization of metabolite fragments made in the instrument, is used to distinguish the sterols from other metabolites in complex mixtures to allow the simultaneous detection of a wide variety of steroids, including brassinosteroids. In maize, genetic resources have permitted multiple insights into the role of brassinosteroids in growth and development, and the addition of this convenient protocol to quantify their levels in plant tissue will enable a deeper physiological and biochemical understanding.