Integrated on‐chip derivatization and electrophoresis for the rapid analysis of biogenic amines

Abstract

We demonstrate the monolithic integration of a chemical reactor with a capillary electrophoresis device for the rapid and sensitive analysis of biogenic amines. Fluorescein isothiocyanate (FITC) is widely employed for the analysis of amino-group containing analytes. However, the slow reaction kinetics hinders the use of this dye for on-chip labeling applications. Other alternatives are available such as o-phthaldehyde (OPA), however, the inferior photophysical properties and the UV lambdamax present difficulties when using common excitation sources leading to a disparity in sensitivity. Consequently, we present for the first time the use of dichlorotriazine fluorescein (DTAF) as a superior in situ derivatizing agent for biogenic amines in microfluidic devices. The developed microdevice employs both hydrodynamic and electroosmotic flow, facilitating the creation of a polymeric microchip to perform both precolumn derivatization and electrophoretic analysis. The favorable photophysical properties of the DTAF and its fast reaction kinetics provide detection limits down to 1 nM and total analysis times (including on-chip mixing and reaction) of <60 s. The detection limits are two orders of magnitude lower than current limits obtained with both FITC and OPA. The optimized microdevice is also employed to probe biogenic amines in real samples.

An integrated microfluidic device in polyester for electrophoretic analysis of amino acids

“…Both precolumn and postcolumn reaction chambers have been successfully integrated into microfluidic systems with laser-induced fluorescence (LIF) detection [3,4,6,7]. However, on-column labeling with a low yield may be unsuitable for a miniaturized detection system with lower sensitivity than LIF.…”

Determination of biogenic amines in HeLa cell lysate by 6‐oxy‐(N‐succinimidyl acetate)‐9‐(2'–methoxycarbonyl) fluorescein and micellar electrokinetic capillary chromatography with laser‐induced fluorescence detection

“…Compared to the poor sensitivity of UV absorption detection, a remarkable improvement in detection limits in fluorescence detection has been reported to at least 2-3 orders of magnitude [10][11][12]26]. LIF is often the method of interest for low-concentration analytes because of its high sensitivity.…”

“…The argon ion laser emitting at 488 nm is the most frequent excitation source employed for detecting biogenic amines. However, only a small quantity of derivatizing reagents such as FITC [24,27], 5-(4,6-dichloro-s-triazin-2-ylamino)fluorescein (DTAF) [10,[28][29][30][31], and 5(6) carboxyfluorescein N-succinimidyl ester (CFSE) [32], which are suitable for this ion laser, has been developed. Nevertheless, these reagents have their drawbacks.…”

Precolumn diastereomerization and micellar electrokinetic chromatography on a plastic microchip: Rapid chiral analysis of amino acids

“…In both studies [38,39], amino acids had to be derivatized with FITC for 5-12 h in an offline condition for the fluorescence detection prior to injection into the microchips. In order to maintain the advantages of the microchip analysis, such as high speed assay, low reagent consumption, and ease of integration, a chip based precolumn derivatization/separation protocol [43][44][45][46] is also preferred for the chiral separation and fluorescence detection of amino acids.…”