Determination of abscisic acid content in a normal environment by ultra-performance liquid chromatography–mass spectrometry (UPLC-MS/MS)

Approximately 50 mg of powder from the same tissues used for sequencing was obtained per tube at low temperature and immediately weighed with a 1/10,000 balance. Approximately 0.5 mL of the extract solution (isopropanol:H₂O:HCl = 2:1:0.002, v/v/v) was mixed with the powder in a centrifuge tube, and 10 μL of 1 ng μL⁻¹ D6 abscisic acid (ABA) was added as an internal standard. The subsequent UPLC-MS/MS extraction procedure followed the method with slight modifications²³. Approximately 50 µL of the extracted sample solution was injected into a reverse-phase C18 Gemini HPLC column for analysis. The ABA content was determined using ultra-performance liquid chromatography–electrospray ionization triple quadrupole mass spectrometry (UPLC-ESI-MS/MS) (Agilent 5500, Santa Clara, CA, USA). The areas of the peaks in the chromatogram were quantified using MassHunter software (Agilent, Santa Clara, CA, USA).