Development and application of electroanalytical techniques for biological matrices
Degree GrantorUniversity of Canterbury
Degree NameDoctor of Philosophy
This thesis presents a study on the development and application of electrochemical techniques for analysis in biological fluids. Two main problems which occur when measurements are made directly in biological matrices were addressed. Firstly, interferences from other electro active components and secondly, adsorption of high molecular weight organic species (such as proteins) onto the electrode surface. Much of the work described utilised glassy carbon electrodes covalently modified by a diazonium salt coupling procedure. This method of modification produced stable electrode surfaces suitable for use in flowing streams and with organic solvents (such as those used in reversed phase-HPLC). The properties of the chemically modified electrodes (CMEs) could be varied by changing the para substituent of the aryl species grafted onto the electrode surface. Modification with p-phenylacetate and p-benzoate moieties generated CMEs which were negatively charged at physiological pH (7.4) and were selective for cationic analytes relative to anionic species. The suitability of these CMEs as probes for the measurement of the catecholamine neurotransmitter dopamine (DA) was investigated in detail. The CMEs exhibited fast response times to DA and good selectivity to DA over ascorbic acid making them suitable for DA measurements with fast timescale electrochemical techniques. Uncharged CMEs were also prepared using the diazonium salt procedure. The selectivity of p-alkylbenzene modified electrodes was investigated in batch and flow injection analysis conditions. The selective measurement of biologically important species (acetaminophen and chlorpromazine) over interferents (ascorbic acid and uric acid), was investigated. Chlorpromazine was able to be measured in the presence of uric acid using a retention-time based technique coupled with a p-phenylacetate/p-methylbenzene modified detector. The simultaneous determination of acetaminophen, ascorbic acid and uric acid was achieved at an array of modified detectors. Covalently modified electrodes were used in protein adsorption studies. The extent of protein adsorption was evaluated by monitoring the electrochemical response of suitable probe analytes. Protein adsorption was influenced by monolayer modification. Anionic groups close to the electrode surface did not affect protein adsorption whilst hydrophobic groups increased protein adsorption. Charged groups which extended further into solution decreased protein adsorption. Protein adsorption was also examined at different carbon materials, electrochemically pretreated glassy carbon and CMEs prepared by methods other than diazonium salt modification. Glassy carbon was less prone to adsorption of high molecular weight foul ants compared to other unmodified carbon electrodes. Some assembly of anionic surfactant on basal plane graphite and incorporation of anionic surfactant in graphite epoxy decreased protein adsorption relative to unmodified electrodes. Electrodes coated with the polymer, cellulose acetate, decreased protein adsorption but gave poor reproducibility and suffered from time-dependent response.