Study On Methodologies And Applications Of Capillary Electrophoresis Coupled With Amperometric Detection

Capillary electrophoresis (CE) is a powerful liquid phase separation technique.The separation principle is based on the phenomena that ionized molecules ofdifferent mass and charge will travel with different velocities through a fused silicacapillary under the influence of a high-voltage electric field.Owing to the highseparation efficiency,rapid analysis,minimum consume of samples andenvironmental friendliness,it has been flourishing in wide areas such asenvironmental analysis,pharmaceutical analysis,biochemical analysis,and foodanalysis etc.There are many detection methodologies coupled with CE,such as UV-Visibledetection (UV-Vis),Laser Induced Fluorescence detection (LIF),Electrochemicaldetection (ED) and Mass Spectrum (MS).The major barriers for CE systemintegration with LIF and MS etc.are fluidic interface,sample pretreatment(fluorescence derivatization or labeling),etc.Since ED,especially amperometricdetection (AD) has many advantages over other widely used techniques in that it ismuch more economical,without complicated pretreatment procedure,and it providesgood selectivity as well as high sensitivity,so it has flourished to various areas.Fromthe standpoint of broadening its application areas,we mainly focus on thedevelopment of new chemically modified electrode (CME) and new method ofCE-AD.This paper begins with a brief introduction of CE,its separation models as well asvarious detection methodologies.Chapter 2 is the determination of three kinds of carbohydrates together withascorbic acid by capillary zone electrophoresis with amperometric detection(CZE-AD) at a carbon paste modified electrode.In this paper,a kind of novel carbonpaste electrode modified with double modifiers-polyethylene glycol (PEG) and Cu₂O(PEG-Cu₂O CPME) in CZE-AD was applied to simultaneously determine glucose,sucrose,fructose and ascorbic acid (AA).The catalytic electrochemical properties ofPEG-Cu₂O CPME could enhance sensitivity obviously compared with carbon pasteelectrode modified with only PEG or Cu₂O at a relatively lower detection potential(+0.3 V vs.SCE).The four analytes could be perfectly separated within 22 min,the linear ranges were from 1.0×10^-6 to 5.0×10^-5 mol L^-1 and the detection limits were at10^-7 mol L^-1 magnitude (S/N=3).The present working electrode was successfullyemployed to analyze beverage samples with recoveries in the range 93～107% andRSDs less than 4%.The results demonstrated that CZE coupled with the PEG-Cu₂Ocarbon paste modified electrode was of convenient preparation,high sensitivity,goodrepeatability and could be used in the rapid determination of practical samples.Chapter 3 states the development of a novel double modifier carbon pasteelectrode-a MWCNT-Cu₂O CPME and its application on several amino acids.Aminoacids are essential building blocks of biological molecules.Though they are soimportant to human beings,it is to some extent very difficult to separate anddetermine amino acids because of their adsorbability to silica capillary andnon-optical and non-electrochemical activity for detection.In this paper,a carbonpaste electrode modified with multiwall carbon nanotubes and copper (I) oxide(MWCNT-Cu₂O CPME) was fabricated and the electrochemical behaviors ofnineteen kinds of natural amino acids at this modified electrode were studied.TheMWCNT-Cu₂O CPME was also successfully applied to the CZE-AD of six kinds ofamino acids.Under the optimal conditions,these amino acids could be perfectlyseparated within 20 min and their detection limits were as low as 10^-7 or 10^-8 mol L^-1magnitude (S/N=3),which demonstrated that MWCNT-Cu₂O CPME could besuccessfully employed as an electrochemical sensor for amino acids with someadvantages of convenient preparation,high sensitivity and good repeatability.Chapter 4 develops a new kind of vitamin B₁₂ (acquo-cobalamine) chemicallymodified electrode and applies in capillary zone electrophoresis coupled withamperometric detection (CZE-AD) for simultaneous determination of someantioxidants in fruits and vegetables.The catalytic electrochemical properties of thechemically modified electrode could obviously enhance oxidation peak currentsresponses by about five times to glutathione,ascorbic acid,vanillic acid,chlorogenicacid,salicylic acid,and caffeic acid compared with common carbon disk electrode.Furthermore,the effects of working electrode potential,pH and concentration ofrunning buffer,separation voltage and injection time on CZE-AD were investigated.Under the optimum conditions,the six analytes could be completely separated and detected in a borate-phosphate buffer (pH 8.4) within 15 min.Their linear rangeswere from 2.5×10^-7 to 1.0×10^-4 mol L^-1 and the detection limits were as low as 10^-7 or10^-8 mol L^-1 magnitude (S/N=3).The proposed method has been successfullyemployed to monitor the six analytes in practical samples with recoveries in the range96.0-106.0% and RSDs less than 5.0%.Above results demonstrate that capillary zoneelectrophoresis coupled with electrochemical detection using vitamin B₁₂ modifiedelectrode as detector is of convenient preparation,high sensitivity,good repeatability,and could be used in the rapid determination of practical samples.Chapter 5 states simultaneous determination of three isomers ofphenylenediamines (o-,m-p-phenylenediamine) and two isomers ofdihydroxybenzenes (catechol and resorcinol) in hair dyes was performed by capillaryzone eletrophoresis coupled with amperometric detection (CZE-AD).The effects ofworking electrode potential,pH and concentration of running buffer,separationvoltage and injection time on CZE-AD were investigated.Under the optimumconditions,the five analytes could be perfectly separated in a 0.04 mol L^-1borate-phosphate buffer (pH 5.8) within 15 min.A 300μm diameter platinumelectrode had good responses at +0.85 V (versus SCE) for the five analytes.Theirlinear ranges were from 1.0×10^-6 to 1.0×10^-4 mol L^-1 and the detection limits were aslow as 10^-7 mol L^-1 magnitude (S/N=3).The present working electrode wassuccessfully employed to analyse eight kinds of hair dyes samples with recoveries inthe range 91.0-108.0% and RSDs less than 5.0%.Above results demonstrated thatcapillary zone electrophoresis coupled with electrochemical detection using platinumworking electrode as detector was of convenient preparation,high sensitivity,goodrepeatability and could be used in the rapid determination of practical samples.Chapter 6 is about the optimization of CZE for a complicated system.Chrysanthemum is a herbaceous perennial plant possessing antimicrobial,antibacterial,antifungal,antiviral and anti-inflammatory activities,which mostlyowing to it ingredients of flavonoids.Here in this research we focus on the CZEseparation of six kinds of flavonoids in chrysanthemum.With the help of differentadditives into buffer and sample,they could get baseline separation within 20 min andthe LOD could reach 10^-8 g mL^-1 magnitude.The successful practical application on the determination of chrysanthemum samples confirmed the validity and practicabilityof this method.The last chapter is focus on determination of the o-,m-,p-phenylenediamine ando-,m-,p-dihydroxybenzene.Because of the great pKa differences between twogroups of isomers but the similar pKa in each group,it is very difficult to find asuitable pH buffer for the simultaneous separation of the two groups of isomers bycapillary zone electrophoresis (CZE).In the paper,a novel method based on runningbuffer pH gradients was designed for capillary zone electrophoresis withamperometry detection (CZE-AD) to simultaneously separate two groups ofinteresting positional isomers including o-,m-,p-phenylenediamine and o-,m-,p-dihydroxybenzene.It is found that when the pH gradients running was employedappropriately,the six analytes could be separated in less than 20 min and the detectionlimits were as low as 10^-7 mol L^-1.Furthermore,other factors affecting the CZEseparation,such as working potential,ionic strength of running buffer,separationvoltage and sample injection time were extensively investigated.The feasible of themethod was determinated by using it in the chapter 6 chrysanthemum experiment,theresults showed that the novel method can replace adding modifier in running buffer.Experimental results demonstrated that the method could be successfully employedfor the analytes which could not be separated simultaneously under the immobile plugwith defined pH with some advantages of high sensitivity,good repeatability andsmall sample requirement.