Determination of four kinds of endogenous hormones in poplar dialyzate by HPLC with microdialysis

A dialyzate was obtained from the tender stems of living poplar (Populus wutunensis) by microdialysis (MD). Four kinds of endogenous hormones, namely, benzylamino purine, gibberellic acid, indole acetic acid, and abscisic acid, were then simultaneously determined by HPLC. The molecular weight cutoff (MWCO) of the MD probe was 30 kDa. The perfusion rate was 1 μL/min, and the perfusion liquid was Millipore-Q water. HPLC was performed using an Inertsil ODS-SP column. The mobile phase comprised 60:40 (v/v) methanol and water containing 0.2% phosphoric acid. The elution rate was 0.6 mL/min rate at 30°C. The average recoveries of the four kinds of hormones ranged from 93% to 109%, with correlation coefficients greater than 0.998. The determinations of endogenous hormones in plants as real time, living, and online were realized by MD-HPLC. The cells and macromolecules were excluded by the MD probe so centrifugation or protein precipitation was not required prior to analysis. MD-HPLC is a highly sensitive and selective method for the simultaneous profiling and quantification of a wide variety of plant hormone groups. This method is convenient for studying the mechanism of plant hormones.