Derivatization at Ultratrace Levels on Solid Phase Sorbents

Defense Date


Graduation Date

Spring 1-1-2005


Campus Only

Submission Type


Degree Name



Chemistry and Biochemistry


Bayer School of Natural and Environmental Sciences

Committee Chair

Mitchell E. Johnson

Committee Member

H. M. Skip Kingston

Committee Member

Jeffry D. Madura


analyte isolation, cannabinoid, fluorescence


The use of derivatization to improve analyte response is widespread in analytical techniques. It helps improve detection limits when analytes are present at low concentrations or their chemistry does not permit detection by traditional methods such as UV detection. Another way to enhance analytical signal is through preconcentration techniques. Preconcentration allows a sample of low concentration to be analyzed and it also lowers mass detection limits for a particular sample. In this work, solid phase sorbents are used to combine preconcentration with derivatization in a single step. The use of a chromatographic stationary phase allows for lower detection limits and better reaction efficiency. The technique has been applied to a 2.1 × 10.0 mm guard column and on a microchip format in conjunction with analysis by HPLC. Both techniques provide an improvement in the reaction efficiency and mass detection limit. The analytes of interest are primary amines and they are derivatized with 5-fluorescein isothiocyanate (FITC). Work with phenylalkylamines on the guard column is related to biologically significant neurotransmitters and displays the usefulness of online derivatization with possible automation to eliminate multiple steps that normally accompany analysis. Mircochips packed with solid phase were used to derivatize a series of fatty amines. These amines are related to a series of biologically significant compounds known collectively as primary fatty acid amides (PFAM's). This class of compounds, particularly oleamide, has distinct biological function and the detection of these compounds at nanomolar levels through an online derivatization technique will advance the progress in screening for and understanding these unique molecules. Work surrounding the separation of a series of PFAM's using HPLC is also discussed. The work shows a need for a more sensitive technique, but illustrates the use of HPLC as a powerful separation technique. The combination of online sample clean-up, preconcentration and derivatization will eventually lead to a method of determining primary fatty acid amides as a class of lipids. By incorporating derivatization on a microchip with techniques such as microchip capillary electrophoresis, a micro total analysis system for clinical diagnosis can eventually be realized.





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