title: Amine Analysis Using AlexaFluor 488 Succinimidyl Ester and Capillary Electrophoresis with Laser-Induced Fluorescence.
abstract: Fluorescent probes enable detection of otherwise nonfluorescent species via highly sensitive laser-induced fluorescence. Organic amines are predominantly nonfluorescent and are of analytical interest in agricultural and food science, biomedical applications, and biowarfare detection. Alexa Fluor 488 N-hydroxysuccinimidyl ester (AF488 NHS-ester) is an amine-specific fluorescent probe. Here, we demonstrate low limit of detection of long-chain (C9 to C18) primary amines and optimize AF488 derivatization of long-chain primary amines. The reaction was found to be equally efficient in all solvents studied (dimethylsulfoxide, ethanol, and N,N-dimethylformamide). While an organic base (N,N-diisopropylethylamine) is required to achieve efficient reaction between AF488 NHS-ester and organic amines with longer hydrophobic chains, high concentrations (>5 mM) result in increased levels of ethylamine and propylamine in the blank. Optimal incubation times were found to be >12 hrs at room temperature. We present an initial capillary electrophoresis separation for analysis using a simple micellar electrokinetic chromatography (MEKC) buffer consisting of 12 mM sodium dodecylsulfate (SDS) and 5 mM carbonate, pH 10. Limits of detection using the optimized labeling conditions and these separation conditions were 5-17 nM. The method presented here represents a novel addition to the arsenal of fluorescent probes available for highly sensitive analysis of small organic molecules.
authors.affiliation: Jet Propulsion Laboratory, California Institute of Technology, Pasadena, CA 91109, USA ; Weill Cornell Graduate School of Medical Science, New York, NY 10065, USA.
authors.forename: Christian G
authors.initials: CG
authors.lastname: Kendall
authors.affiliation: Jet Propulsion Laboratory, California Institute of Technology, Pasadena, CA 91109, USA ; Georgia Institute of Technology, Atlanta, GA 30332, USA.
authors.forename: Amanda M
authors.initials: AM
authors.lastname: Stockton
authors.affiliation: Jet Propulsion Laboratory, California Institute of Technology, Pasadena, CA 91109, USA ; University of California, Los Angeles, CA 90095, USA.
authors.forename: Stephen
authors.initials: S
authors.lastname: Leicht
authors.affiliation: Jet Propulsion Laboratory, California Institute of Technology, Pasadena, CA 91109, USA.
authors.forename: Heather
authors.initials: H
authors.lastname: McCaig
authors.affiliation: Jet Propulsion Laboratory, California Institute of Technology, Pasadena, CA 91109, USA.
authors.forename: Shirley
authors.initials: S
authors.lastname: Chung
authors.affiliation: Jet Propulsion Laboratory, California Institute of Technology, Pasadena, CA 91109, USA.
authors.forename: Valerie
authors.initials: V
authors.lastname: Scott
authors.affiliation: Jet Propulsion Laboratory, California Institute of Technology, Pasadena, CA 91109, USA.
authors.forename: Fang
authors.initials: F
authors.lastname: Zhong
authors.affiliation: Jet Propulsion Laboratory, California Institute of Technology, Pasadena, CA 91109, USA.
authors.forename: Ying
authors.initials: Y
authors.lastname: Lin