Rapid and sensitive hormonal profiling of complex plant samples by liquid chromatography coupled to electrospray ionization tandem mass spectrometry

The present work reports a rapid, specific and sensitive method using ultrahigh-performance liquid chromatography coupled to electrospray ionization tandem spectrometry (UPLC/ESI-MS/MS) to analyze quantitatively the major hormones found in plant tissues within six minutes, including auxins, cytokinins, gibberellins, abscisic acid, 1-amino-cyclopropane-1-carboxyic acid (the ethylene precursor), jasmonic acid and salicylic acid. Sample preparation, extraction procedures and UPLC-MS/MS conditions were optimized for the determination of all plant hormones and are summarized in a schematic extraction diagram for the analysis of small amounts of plant material without time-consuming additional steps such as purification, sample drying or re-suspension.This new method is applicable to the analysis of dynamic changes in endogenous concentrations of hormones to study plant developmental processes or plant responses to biotic and abiotic stresses in complex tissues. An example is shown in which a hormone profiling is obtained from leaves of plants exposed to salt stress in the aromatic plant, Rosmarinus officinalis.Hormones play a pivotal role in most physiological processes in plants. These structurally diverse compounds that act usually at nanomolar levels include five groups of the so-called "classic" hormones, comprising auxins, cytokinins, gibberellins (GA), abscisic acid (ABA) and ethylene, and several other plant growth regulators, including jasmonates, salicylates, brassinosteroids, polyamines or the very recently discovered strigolactones, which fit several of the criteria to be considered hormones [1-3]. Furthermore, the list of plant hormones is expected to increase due to a better understanding of plant growth and development and stress responses, and the use of technological advances in analytical methods.Recent studies support the contention that hormone actions build a signaling network and mutually regulate several signaling and metabolic systems, such as auxi