title: Fluorescence detection by intensity changes for high-performance thin-layer chromatography separation of lipids using automated multiple development.
abstract: Changes in emission of berberine cation, induced by non-covalent interactions with lipids on silica gel plates, can be used for detecting and quantifying lipids using fluorescence scanning densitometry in HPTLC analysis. This procedure, referred to as fluorescence detection by intensity changes (FDIC) has been used here in combination with automated multiple development (HPTLC/AMD), a gradient-based separation HPTLC technique, for separating, detecting and quantifying lipids from different families. Three different HPTLC/AMD gradient schemes have been developed for separating: neutral lipid families and steryl glycosides; different sphingolipids; and sphingosine-sphinganine mixtures. Fluorescent molar responses of studied lipids, and differences in response among different lipid families have been rationalized in the light of a previously proposed model of FDIC response, which is based on ion-induced dipole interactions between the fluorophore and the analyte. Likewise, computational calculations using molecular mechanics have also been a complementary useful tool to explain high FDIC responses of cholesteryl and steryl-derivatives, and moderate responses of sphingolipids. An explanation for the high FDIC response of cholesterol, whose limit of detection (LOD) is 5 ng, has been proposed. Advantages and limitations of FDIC application have also been discussed.
authors.affiliation: Instituto de Carboquímica, Consejo Superior de Investigaciones Cientificas, C/Miguel Luesma, 4, 50018 Zaragoza, Spain. vcebolla@icb.csic.es
authors.forename: Vicente L
authors.initials: VL
authors.lastname: Cebolla
authors.forename: Carmen
authors.initials: C
authors.lastname: Jarne
authors.forename: Pilar
authors.initials: P
authors.lastname: Domingo
authors.forename: Andrés
authors.initials: A
authors.lastname: Domínguez
authors.forename: Aránzazu
authors.initials: A
authors.lastname: Delgado-Camón
authors.forename: Rosa
authors.initials: R
authors.lastname: Garriga
authors.forename: Javier
authors.initials: J
authors.lastname: Galbán
authors.forename: Luis
authors.initials: L
authors.lastname: Membrado
authors.forename: Eva M
authors.initials: EM
authors.lastname: Gálvez
authors.forename: Fernando P
authors.initials: FP
authors.lastname: Cossío
mesh_terms: D001599:Berberine; D002784:Cholesterol; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D002855:Chromatography, Thin Layer / Q000379:methods*; D003720:Densitometry / Q000379:methods; D008055:Lipids / Q000737:chemistry / Q000302:isolation & purification*; D012680:Sensitivity and Specificity; D013050:Spectrometry, Fluorescence / Q000379:methods*
chemical_list: D008055:Lipids; D001599:Berberine; D002784:Cholesterol