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METHOD DEVELOPMENT FOR THE ANALYSIS OF PROTEINS AND SMALL

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abstract
Analytical separations in conjunction with powerful detection methods make possible the quantitation of large and small molecules important in environmental science, food science, pharmaceutical, and medical matrices. Many of these separations utilize instrumental methods such as capillary electrophoresis (CE) and high performance liquid chromatography (HPLC) to alleviate the complexity of these various matrices. Fluorescence, laser-induced fluorescence (LIF), evaporative light scattering detection (ELSD), and mass spectrometry (MS) are detection methods commonly employed in conjunction with these separation strategies. Three distinct projects to develop and exploit such analytical technologies are described. First, few biological molecules are natively fluorescent, so they often must be labeled with a fluorescent tag in order to be detected via LIF. Furthermore, protein separations using CE-LIF may be complicated by protein interactions with the negatively charged surface of the inner capillary wall, arising from the ionization of surface silanol groups. Presented here is a method for quantitation of protein-dye complexes using a self assembled, surfactant-coated capillary and on-column protein labeling with a non-covalent squarylium dye, Red-1c. Second, seasonal sugar levels are relevant to the relationship between winter anthocyanin production and drought stress in angiosperm evergreen species, and so the development of a method for rapid analysis of sugars in seasonal tree leaf tissue is needed. An HPLC-ELSD method was developed for the quantitation of three sugars, glucose, fructose, and sucrose. The method was subjected to standard addition validation and was shown to provide precise and accurate measurement of all three analytes in real samples.
subject
Capillary Electrophoresis
CE-LIF
CE-MS
Proteins
separation
contributor
Gerardi, Anthony Richard (author)
Colyer, Christa L (committee chair)
Guthold, Martin (committee member)
Dos Santos, Patricia C. (committee member)
Hinze, Willie L. (committee member)
Jones, Bradley T. (committee member)
date
2011-07-14T20:36:16Z (accessioned)
2011-07-14T20:36:16Z (available)
2011 (issued)
degree
Chemistry (discipline)
identifier
http://hdl.handle.net/10339/33489 (uri)
language
en (iso)
publisher
Wake Forest University
title
METHOD DEVELOPMENT FOR THE ANALYSIS OF PROTEINS AND SMALL
type
Dissertation

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