Capillary electrophoresis with wavelength-resolved laser-induced fluorescence detection

Zhang, Xin; Stuart, Jeffrey N.; Sweedler, Jonathan V.

doi:10.1007/s00216-002-1288-9

Cited by 52 publications

(39 citation statements)

References 74 publications

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“…The laboratory-assembled capillary electrophoresis (CE) system with 257 nm excitation and a charge-coupled device/spectrograph for wavelength-resolved fluorescence emission detection was employed for ultra sensitive detection of 5-HT and its metabolites (Fuller et al 1998;Park et al 1999;Zhang et al 2001;Zhang et al 2002;Stuart et al 2003). Briefly, approximately 3 nL of sample was injected onto a 50-lm inner diameter/150-lm outer diameter capillary and separated by an applied voltage of 21-25 kV.…”

Section: Capillary Electrophoresismentioning

confidence: 99%

Systemic serotonin sulfate in opisthobranch mollusks

Stuart¹,

Ebaugh²,

Copes³

et al. 2004

Journal of Neurochemistry

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Serotonin (5-hydroxytryptamine, 5-HT) is a ubiquitous modulatory neurotransmitter with roles as a neurohormone and neurotransmitter. However, few studies have been performed characterizing this molecule and its related metabolites in circulating fluids. Here, we demonstrate native 5-HT sulfate, but much lower levels of 5-HT, in hemolymph of the marine mollusk Pleurobranchaea californica. The metabolite 5-HT sulfate forms from 5-HT uptake and metabolism in central ganglia of Aplysia californica and in the visceral nerve and eye of Pleurobranchaea, but not in hemolymph itself. In addition, 5-hydroxyindole acetic acid (5-HIAA), while not detected in hemolymph, forms in higher quantities than does 5-HT sulfate in the eye and visceral nerve, and c-glu-5-HT is also observed in this area but never in hemolymph. As systemic 5-HT sulfate appears not to originate from the optic region or from systemic 5-HT, 5-HT sulfate likely derives from the nervous system. Circulating 5-HT sulfate is at least 10-fold higher during the light portion of a 12 : 12-h light/dark cycle than during the dark portion (p < 0.0007), but there is no obvious trend for free systemic tryptophan (Trp) (p > 0.3) in Pleurobranchaea. 5-HT in mollusks is associated with general arousal state; thus, diurnal systemic changes in a 5-HT catabolite may reflect a regulatory role for indole catabolism in behavioral rhythms. Keywords: capillary electrophoresis, hemolymph, marine mollusk, serotonin, sulfation. Serotonin (5-hydroxytryptamine, 5-HT) is a biogenic amine responsible for a variety of tasks in the nervous system. For instance, 5-HT is implicated in learning and memory in the central nervous system (CNS) and emesis and peristaltic reflex in the enteric nervous system (ENS) (Bartsch et al. 2000;Buhot et al. 2000;Endo et al. 2000). Several organs serve as targets for 5-HT, but the fate of 5-HT in target tissues, let alone the physiological effects of the resulting metabolites, has not been fully ascertained. Knowledge of the products of transformed 5-HT, as well as their distribution patterns, will assist in illuminating the processes leading to impaired neurotransmission and eventually aid in designing compounds that could potentially inhibit disease in neural and surrounding tissues.Marine mollusks present an excellent model for the study of the nervous system because of their large cell somata and well-characterized neural networks. In gastropod mollusks, 5-HT is involved in many neurophysiological processes, including sensitization and facilitation of withdrawal reflexes (Hawkins et al. 1993), feeding behavior (Gillette and Davis 1977) and locomotion (McPherson and Blankenship 1991). However, 5-HT-related metabolites have received considerably less attention.Our laboratory recently demonstrated that 5-HT metabolism in Aplysia californica involves sulfation and c-glutamylation with trace amounts of monoamine oxidase activity present in the form of 5-hydroxyindole acetic acid (5-HIAA) (Stuart et al. 2003). In particular, 5-HT sulfate is present ...

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Section: Capillary Electrophoresismentioning

confidence: 99%

Systemic serotonin sulfate in opisthobranch mollusks

Stuart¹,

Ebaugh²,

Copes³

et al. 2004

Journal of Neurochemistry

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“…The method combines the high separation efficiencies and small sample volumes of CE with the low detection limits and high resolution fluorescence spectra obtained from the LIF detection system, which uses a sheath-flow cuvette, spectrograph, and charge-coupled device array. 13 CE with wavelengthresolved fluorescence detection has been used for a wide variety of applications including DNA sequencing, cellular and neurochemical analyses, and rapid clinical assays of biological fluids. 14 The CE-LIF instrumentation was configured for optimal separation of negatively charged species, 13,15 as we anticipate for the carboxylate modified Si-np-COO − at basic pH.…”

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confidence: 99%

“…Calibrations of the system using a range of organic fluorophores indicate that the 8.9 min band corresponds to neutral or near-neutral species, while the remaining bands at longer elution times fall in the ranges typical of small organics with one or two negative charges. 13 For example, the fluorescein dianion elutes at ϳ22 min under the same conditions. Fluorescence spectra for the major bands are shown in Fig.…”

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confidence: 99%

Capillary electrophoresis of ultrasmall carboxylate functionalized silicon nanoparticles

Eckhoff

Stuart

Sutin

et al. 2006

The Journal of Chemical Physics

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Capillary electrophoresis is used to separate ultrasmall (∼1nm) carboxylate functionalized Si nanoparticles (Si-np-COO−) prepared via hydrosilylation with an ω-ester 1-alkene. The electropherograms show a monodisperse Si core size with one or two carboxylate groups added to the surface. On-column detection of their laser-induced fluorescence demonstrates that the individual Si-np-COO− have narrow emissions (full width at half maximum =30–40nm) with a nearly symmetric lineshape. Preparative scale electrophoresis should be a viable route for purification of the Si-np-COO− for further study and future applications.

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“…The use of a spectrometer adds a second dimension of information to the analysis. A tandem measurement of the fluorescence emission spectrum allows resolution of analytes with similar migration times and more reliable identification of analytes than is available by migration time alone (26) . The amount of analyte, determined by the integral of the peak, can be calibrated by spiking cellular lysates with known amounts of analyte.…”

Section: Coupling Ce-lif To a Microbeammentioning

confidence: 99%