Study On Novel Liquid Chromatography-Electrochemical Detection And Its Application In Neurotoxicity Of Endogenous Toxins To Parkinsonian Neurons

With the fast development of life science,accurate analysis of biological samples is the most crucial.However,many conventional analytical methods can't match the requirements because of the complex component and low content of biological samples.High performance liquid chromatography coupled with electrochemical detection(HPLC-ED)has become one of the most important approaches in the fields of chemical,biological,medical and environmental analysis since its appearance in the 1970's for its high separation effectiveness,sensitivity and selectivity and its easiness of control.Parkinson's disease(PD)is one of the most common neurodegenerative diseases in the aged and its pathological mechanism still remains unknown.In this dissertation, we laid our emphasis on the research of novel electrochemical detectors(ECD),and established some new methods of sample separation and analysis in life science by HPLC-ED coupled with microdialysis sampling technique.Some kinds of chemically modified electrodes(CME)that could enhance the sensitivity and selectivity in vivo analysis were developed and used as HPLC-ECD.These new analytical methods were successfully applied to study on neurotoxicity of endogenous toxins to Parkinsonian neurons.The dissertation includes three chapters:Part 1.Study the Concentration Changes of Monoamine Neurotransmitters in Rat Brain after the Administration of(R)-salsolinol and(R)-N-methylsalsolinol by Liquid Chromatography with Acetylcholine Modified ElectrodeThe purpose of this paper was to study the concentration changes of monoamine neurotransmitters in rat brain after the administration of(R)-salsolinol and(R)-N-methylsalsolinol. A novel electrochemical detector with the acetylcholine film for high performance liquid chromatography was described.It was found that the chemically modified electrode exhibited efficiently electrocatalytic effect on the current responses of(R)-Sal and(R)-NMSal and monoamine neurotransmitters with high sensitivity,stability,and long-life of activity.Combined with microdialysis sampling,the method was successfully applied to study the concentration changes of the monoamine neurotransmitters in rat brain after the injection of(R)-salsolinol and (R)-N-methylsalsolinol.Part 2.Study of the nicotine prevention effect on striatal dopamine loss caused by(R)-N-methylsalsolinol in rat brainThe purpose of this paper is to study the protective effects of nicotine on striatal DA loss caused by(R)-NMSal in rat striatum.In this study,we assayed the changes of nitric oxide synthase activity,and also detected the concentration of DA and its metabolites by high performance liquid chromatography with Pt-polyvinyl alcohol (Pt/PVA)chemically modified electrode in rat striatum.The results showed that nicotine could significantly increase striatal monoamine neurotransmitters release to the basal level.Part 3.Effect of(R)-Salsolinol and N-Methyl-(R)-salsolinol on the Balance Impairment Between Dopamine and Acetylcholine in Rat BrainIn this paper,after injection of(R)-Sal or(R)-NMSal into the rat brain straitum,the levels of DA and its metabolites were detected by high performance liquid chromatography with electrochemical detection.We assessed the influence of neurotoxins on acetylcholinesterase (ACHE)activity and developed a microdialysis-electrochemical device to measure ACh levels with enzyme modified electrodes.The results show that(R)-Sal and(R)-NMSal exerted a considerable effect on the balance between DA and ACh by impairing cholinergic system as well as dopaminergic system.This result is helpful for better understanding the potential role of endogenous neurotoxins in the pathogenesis of PD.