Development and application of a new in-line coupling of a miniaturized boronate affinity monolithic column with capillary zone electrophoresis for the selective enrichment and analysis of cis-diol-containing compounds.
An integrated, miniaturized and fully automated system was developed for the analysis (preconcentration/purification, separation and detection) of cis-diol containing molecules in complex matrices. This innovative in-line coupling system was achieved via the in-situ and localized synthesis of a short segment of silica-based monolith at the inlet of a 75-μm inner diameter fused silica capillary. The monolithic segment was locally functionalized with an acrylamide derivative of phenylboronic acid by free radical photopolymerization within 10min of irradiation time. Efficiency of the photopolymerization reaction was followed by frontal affinity chromatography of 1,2-dihydroxybenzene (catechol) as cis-diol model solute. An active-site amount of 0.43nmolcm-1(9.8nmolμL-1) of phenylboronic acid moieties was obtained, with a Kdvalue of about 290μM close to reported value for the phenyl boronate-catechol complex. The optimal conditions of use of the miniaturized boronate affinity monolithic column (μBAMC) were determined and adapted to the in-line coupling with capillary electrophoresis. Catechol was specifically preconcentrated in a pH 8.5 phosphate buffer/MeOH (80/20, v/v) mixture. A volume up to 20 times the monolith volume can be percolated with a quantitative recovery yield. Three catecholamines were purified, preconcentrated and in-line separated. Elution from the μBAMC was performed with a small plug of acidic solution, allowing field amplified sample stacking of solutes within the plug before their in-line electrophoretic separation at pH 8.75. This unique in-line coupling was successfully used for the fully automated analysis of catecholamines neurotransmitters in urine samples, highlighting the purification efficiency of the μBAMC and the potential of such a fully integrated approach. In addition to the low sample volume required (less than 2μL), the limits of detection (LOD) accomplished with this coupling were estimated at 9.0, 9.5 and 4.8ngmL-1for dopamine, adrenaline and noradrenaline respectively, which improves the LOD of theses solutes compared to other CE methods.
Université de Lyon, CNRS, Université Claude Bernard Lyon 1, ENS de Lyon, Institut des Sciences Analytiques, UMR 5280, 5 rue de la Doua, F-69100 VILLEURBANNE, France. Electronic address: maria.espina-benitez@isa-lyon.fr.
Maria Betzabeth
MB
Espina-Benitez
Université de Lyon, CNRS, Université Claude Bernard Lyon 1, ENS de Lyon, Institut des Sciences Analytiques, UMR 5280, 5 rue de la Doua, F-69100 VILLEURBANNE, France.
Jérôme
J
Randon
Université de Lyon, CNRS, Université Claude Bernard Lyon 1, ENS de Lyon, Institut des Sciences Analytiques, UMR 5280, 5 rue de la Doua, F-69100 VILLEURBANNE, France.
Claire
C
Demesmay
Université de Lyon, CNRS, Université Claude Bernard Lyon 1, ENS de Lyon, Institut des Sciences Analytiques, UMR 5280, 5 rue de la Doua, F-69100 VILLEURBANNE, France.
Vincent
V
Dugas
D001897:Boronic Acids / Q000737:chemistry*; D002395:Catecholamines / Q000652:urine; D002396:Catechols / Q000737:chemistry; D002846:Chromatography, Affinity / Q000295:instrumentation; D019075:Electrophoresis, Capillary / Q000295:instrumentation*; D006801:Humans; D057230:Limit of Detection; D018377:Neurotransmitter Agents / Q000652:urine; D012822:Silicon Dioxide / Q000737:chemistry; D016482:Urinalysis / Q000295:instrumentation / Q000379:methods*
D001897:Boronic Acids; D002395:Catecholamines; D002396:Catechols; D018377:Neurotransmitter Agents; D012822:Silicon Dioxide; C034221:catechol
Boronate affinity chromatography; Capillary electrophoresis; Catecholamines; In-line coupling; Silica monolith; Urine