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CE-LIF ANALYSIS OF INTACT MARINE MICROBES ALONG WITH THEIR CONSTITUENT PHYCOBILIPROTEINS AND PHYCOBILIN PIGMENTS

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abstract
Cyanobacteria contribute significantly to the marine biosphere as primary producers and as participants in global carbon and nitrogen cycles, and so it is vital to develop efficient methods for cyanobacteria quantitation and characterization. Capillary electrophoresis (CE) methods with dual-wavelength laser induced fluorescence (LIF) detection schemes were developed to analyze cyanobacteria according to 1) the separation and quantitation of their natively fluorescent phycobiliproteins, and 2) their electrophoretic profiles as intact organisms. CE-LIF results were compared to those obtained by way of a newly developed, high efficiency separation method known as polymer enhanced capillary transient isotachophoresis (PectI). Four phycobiliproteins (R-PE, B-PE, C-PC, and APC) were determined by CE and PectI, with detection limits below 10 ppb achieved for each protein. Additionally, the phycobilin pigments giving rise to the native fluorescence of these proteins were themselves studied by fluorimetry after liberation from the proteins by way of chemical cleavage with DTT or TCEP. Whereas conventional CE was unable to distinguish between two Synechoccocus sp. bacteria (CCMP 1333 and CCMP 833) in a mixture, the new PectI method was shown to improve the electrophoretic profiles of these intact bacteria samples and to differentiate between them based on the method’s spectral and temporal resolution capabilities.
subject
Capillary Electrophoresis
Cyanobacteria
PectI
Phycobiliproteins
Separations
contributor
Vaughan, Benjamin Ambler (author)
Colyer, Christa L (committee chair)
Jones, Bradley T (committee member)
Alexander, Rebecca W (committee member)
date
2015-06-23T08:35:47Z (accessioned)
2016-06-22T08:30:09Z (available)
2015 (issued)
degree
Chemistry (discipline)
embargo
2016-06-22 (terms)
identifier
http://hdl.handle.net/10339/57130 (uri)
language
en (iso)
publisher
Wake Forest University
title
CE-LIF ANALYSIS OF INTACT MARINE MICROBES ALONG WITH THEIR CONSTITUENT PHYCOBILIPROTEINS AND PHYCOBILIN PIGMENTS
type
Thesis

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