Dual-cloud point extraction and tertiary amine labeling for selective and sensitive capillary electrophoresis-electrochemiluminescent detection of auxins

The low concentrations of the auxins in samples of plant tissue necessitate the use of selective and sensitive techniques for their quantification. Herein a selective and sensitive method based on dual-cloud point extraction (dCPE) and tertiary amine labeling for the quantification of indole-3-acetic acid (IAA) and indole-3-butyric acid (IBA) by capillary electrophoresis-electrochemiluminescence (CE-ECL) is proposed. The procedure for dCPE included two cloud point processes with Triton X-114 as the extractant. The two auxins became hydrophobic in an acidic solution and were extracted into surfactant-rich phase after the first cloud point procedure. They were then back-extracted into the alkaline aqueous phase during the second cloud point step. The extracted auxins were reacted with 2-(2-aminoethyl)-1-methylpyrrolidine (AEMP) in acetonitrile that contained N,N′-dicyclohexylcarbodiimide and 3,4-dihydro-3-hydroxy-4-oxo-1,2,3-benzotriazine to produce their AEMP-derivatives. The two auxin-AEMP-derivatives were subjected into CE and detected by Ru(bpy)32+-based ECL. The preconcentration factors for IAA and IBA with dCPE were 40.5 and 43.4, respectively. The on-capillary detection limits (S/N = 3) were 2.5 and 2.8 nM for IAA and IBA. This protocol presents a clear advantage in that it reduces the interference from the matrixes extensively and gives a high sensitivity for the detection of auxins. The proposed method was applied successfully to the detection of the two auxins in acacia tender leaves, buds, and bean sprout.