Simultaneous Determination of 5 Phytohormone Residues in Vegetables and Fruits by High Performance Liquid Chromatography with Fluoresence Detector

A method of high performance liquid chromatography with fluorescence detector was applied to the simultaneous determination of five phytohormones residues, including 3-indolyl-acetic acid, 3-indolepropionic acid, 3-indolebutyric acid, 2-naphthoxyacetic acid and 1-naphthylacetic acid, in vegetables and fruits. After the edible part of the vegetable or fruit sample was chopped and mixed well, 10.000 g was taken and added into 20 mL of acetonitrile containing 0.2% (volume fraction) formic acid. The mixture was homogenated for 2 min, then 4 g of anhydrous magnesium sulfate and 1 g of sodium chloride were added and the mixture was vortexed for 5 min. After centrifuging at 8 000 r·min^-1 for 5 min, 10.0 mL of the supernatant was taken and evaporated to near dryness. The residue was dissolved with 5 mL of dichloromethane-acetonitrile (9+1) mixture containing 0.2% (volume fraction) formic acid, and the solution was passed through an amino solid phase extraction cartridge at a flow rate of 1 mL·min^-1. The effluent was collected, concentrated to near dryness on a solvent evaporation workstation, and the residue was dissolved with methanol and made up to 1.0 mL. The solution was filtered through a 0.45 μm filter membrane and analyzed by high performance liquid chromatography. An Agilent Eclipse XDB-C₁₈ column (4.6 mm×250 mm, 5 μm) was used as stationary phase, and a mixture of 0.1% (volume fraction) acetic acid solution and methanol was used as mobile phase for gradient elution. Fluorescence detection was performed at the excitation wavelength of 287 nm and the emission wavelength of 337 nm. The results showed that, linear relationships were found between peak area and mass concentration of the 5 phytohormones in the same range of 0.050-5.0 mg·L^-1, with detection limits (3S/N) in the range of 0.93-2.0 μg·kg^-1. Tests for recovery was made by standard addition method on the base of blank samples, giving results in the range of 72.4%-114%, and relative standard deviations (n=6) less than 4.6%.