title: An UPLC-MS/MS method for highly sensitive high-throughput analysis of phytohormones in plant tissues.
abstract: BACKGROUND
  Phytohormones are the key metabolites participating in the regulation of multiple functions of plant organism. Among them, jasmonates, as well as abscisic and salicylic acids are responsible for triggering and modulating plant reactions targeted against pathogens and herbivores, as well as resistance to abiotic stress (drought, UV-irradiation and mechanical wounding). These factors induce dramatic changes in phytohormone biosynthesis and transport leading to rapid local and systemic stress responses. Understanding of underlying mechanisms is of principle interest for scientists working in various areas of plant biology. However, highly sensitive, precise and high-throughput methods for quantification of these phytohormones in small samples of plant tissues are still missing.
  
  
  RESULTS
  Here we present an LC-MS/MS method for fast and highly sensitive determination of jasmonates, abscisic and salicylic acids. A single-step sample preparation procedure based on mixed-mode solid phase extraction was efficiently combined with essential improvements in mobile phase composition yielding higher efficiency of chromatographic separation and MS-sensitivity. This strategy resulted in dramatic increase in overall sensitivity, allowing successful determination of phytohormones in small (less than 50 mg of fresh weight) tissue samples. The method was completely validated in terms of analyte recovery, sensitivity, linearity and precision. Additionally, it was cross-validated with a well-established GC-MS-based procedure and its applicability to a variety of plant species and organs was verified.
  
  
  CONCLUSIONS
  The method can be applied for the analyses of target phytohormones in small tissue samples obtained from any plant species and/or plant part relying on any commercially available (even less sensitive) tandem mass spectrometry instrumentation.
authors.affiliation: Department of Cell and Metabolic Biology, Leibniz Institute of Plant Biochemistry, Weinberg 3, Halle (Saale), 06120, Germany.
authors.forename: Gerd Ulrich
authors.initials: GU
authors.lastname: Balcke
authors.affiliation: Department of Cell and Metabolic Biology, Leibniz Institute of Plant Biochemistry, Weinberg 3, Halle (Saale), 06120, Germany.|Present address: Max Planck Institute for Chemical Ecology, Department of Biochemistry, Hans-Knoell-Str. 8, Jena, 07745, Germany.
authors.forename: Vinzenz
authors.initials: V
authors.lastname: Handrick
authors.affiliation: Department of Cell and Metabolic Biology, Leibniz Institute of Plant Biochemistry, Weinberg 3, Halle (Saale), 06120, Germany.
authors.forename: Nick
authors.initials: N
authors.lastname: Bergau
authors.affiliation: Department of Cell and Metabolic Biology, Leibniz Institute of Plant Biochemistry, Weinberg 3, Halle (Saale), 06120, Germany.
authors.forename: Mandy
authors.initials: M
authors.lastname: Fichtner
authors.affiliation: Department of Cell and Metabolic Biology, Leibniz Institute of Plant Biochemistry, Weinberg 3, Halle (Saale), 06120, Germany.
authors.forename: Anja
authors.initials: A
authors.lastname: Henning
authors.affiliation: Department of Cell and Metabolic Biology, Leibniz Institute of Plant Biochemistry, Weinberg 3, Halle (Saale), 06120, Germany.
authors.forename: Hagen
authors.initials: H
authors.lastname: Stellmach
authors.affiliation: Department of Cell and Metabolic Biology, Leibniz Institute of Plant Biochemistry, Weinberg 3, Halle (Saale), 06120, Germany.
authors.forename: Alain
authors.initials: A
authors.lastname: Tissier
authors.affiliation: Department of Cell and Metabolic Biology, Leibniz Institute of Plant Biochemistry, Weinberg 3, Halle (Saale), 06120, Germany.
authors.forename: Bettina
authors.initials: B
authors.lastname: Hause
authors.affiliation: Department of Cell and Metabolic Biology, Leibniz Institute of Plant Biochemistry, Weinberg 3, Halle (Saale), 06120, Germany.|Faculty of Chemistry and Mineralogy, Institute of Bioanalytical Chemistry, Centre for Biotechnology and Biomedicine, Leipzig University, Deutscher Platz 5, Leipzig, 04103, Germany.
authors.forename: Andrej
authors.initials: A
authors.lastname: Frolov