1-Methoxycarbonylindolizine-3,5-dicarbaldehyde as a derivatization reagent for amino compounds in high-performance capillary electrophoresis

Abstract

The indolizine derivative 1-methoxycarbonylindolizine-3,5-dicarbaldehyde (IDA) was synthesized from 2-(1,3-dioxolan-2-yl)pyridine in four steps. The reactivity of the reagent towards primary amines was investigated by using alanine (Ala) as a model compound. The reagent easily reacted with Ala in 20 mmol l^–1 phosphate–borate buffer at pH 10 containing 25–50% v/v of ethanol in the dark at room temperature, and the reaction was completed within 15 min. The IDA derivative of alanine (IDA-Ala) showed a strong absorption at 280 nm (ε= 3.31 × 10⁴ l mol^–1 cm^–1) and 409 nm (ε= 2.18 × 10⁴ l mol^–1 cm^–1). The derivative also showed fluorescence at 482 nm when irradiated at 282 or 414 nm. These wavelengths did not overlap with those of IDA. IDA-Ala showed a 1.5 times stronger absorption than that produced by the ninhydrin method, and the fluorescence intensity of IDA-Ala was about three times that produced by o-phthalaldehyde-Ala. On analysis of IDA-Ala by high-performance capillary electrophoresis (HPCE) in the capillary zone electrophoresis (CZE) mode with detection at 280 nm, the calibration graph showed good linearity over the range 0.0172–21.5 µmol ml^–1 of alanine. The detection limit for alanine was about 5 nmol ml^–1. The relative standard deviations in the determination of IDA-Ala at 10.8 and 21.5 µmol ml^–1 were less than 2%(n= 5). Application to the analysis of a mixture of amino acids by HPCE using the micellar electrokinetic chromatography mode is also described.