Date of Award

Spring 2015

Project Type

Dissertation

Program or Major

Chemistry

Degree Name

Doctor of Philosophy

First Advisor

Vernon N Reinhold

Second Advisor

W Rudolf Seitz

Third Advisor

Erik Berda

Abstract

Heparin/heparin sulfates are carbohydrate polyionic polymers that participate in a host of critically important biological processes such as blood anticoagulation, pathogen infection, cell differentiation, growth, migration and inflammation, to mention a few. A century has passed since heparin’s initial discovery with a fair understanding of its overall composition. Unfortunately, there has been no structural work at the detailed chemical level that might support a synthetic effort. In this study, I utilize a chemical derivatization strategy (dual permethylation) that imparts isotopic structural specificity, which can be followed by step-wise disassembly in an ion trap mass spectrometer, (MSn). A set of analytical protocols is described that introduces a probable sequencing strategy for heparin analogs. Following derivatization, the O-sulfo groups are converted to deutero methyl esters and the N-sulfo groups are converted to N-deutero acetyl groups. Mass shifts among derivatized products with differential isotopic labeling verified the numbers of N-sulfo groups and O-sulfo groups, while sulfo group positions were characterized by MSn analysis on selected precursor ions. Documentation has been studied with simple heparin/heparin sulfates disaccharides and a synthetic octasulfated pentasaccharide (Arixtra®). Determination of this analogous composition, sequence and sulfonation patterns were accomplished by MSn. The protocols provide unique insights into monomer sequence with monomer structural detail observed by cross-ring fragments often resolved by deuterated isotopic labeling.

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