Quantifying Plant Stress Hormones

Determination of ABA, its glycosylated form (ABA-GE), and catabolites (phaseic and dihydrophaseic acid) in flavedo, pulp, bark and xylem of the different genotypes were performed as described by Diretto et al. (2020) (link) with slight modifications. Briefly, frozen tissues were lyophilized and ground to a fine powder, and replicate samples of 200 mg of each genotype were extracted as previously described (Welsch et al., 2008 (link)). LC-HRMS was carried out using a Ultimate UHPLC-DAD (Dionex) coupled to a Q-Exactive quadrupole Orbitrap mass spectrometry System (Thermo Fisher Scientific), equipped with a C18 Luna column (150 x 2.0mm, 3μm) as described before (Noronha et al., 2022 (link)). Internal standard-based quantification was carried out using the MS data and the quantification software available in the Xcalibur 7.0 software package (Thermo Fisher Scientific, Bremen, Germany). Results are expressed as fold changes with respect to the internal standard.