Study On CE Methods For The Simultaneous Separation And Determination Of Three Types Of Biogenic Amines With Different Cheimcal Structures

Biogenic amines?BAs?refer to the class of the low molecular weight compounds containing nitrogen with biological activity.They can also be considered the low molecular weight organic base which formatted by that H+ of the ammonia hydrogen are replaced by alkyl or aryl.BAs widely exist in the human body or food.They play important physiological roles,and are the important symbols of food freshness and safety.According to their chemical structures,BAs can be divided into aliphatic,aromatic and heterocyclic species.Due to the different detection characters of BAs,it is essential to develop a rapid,simple and sensitive analysis method and simultaneously detect three types of BAs without derivatization.The thirteen key BAs containing aliphatic,aromatic and heterocyclic types were selected as the analytes.And,this paper completed the detailed study targeting at the simultaneous determination of BAs owning different chemical structures.The first chapter firstly introduced the relative knowlegde of capillary electrophoresis?CE?technique,including the basic knowledge,the instrument structure,separation mode,injection methods,detection methods,and the characteristic and deficiency of CE.Secondly,the classification,distribution,physiological function,harm and the main analysis methods of BAs were also introduced.Then,the application of CE techniques in the analysis of BAs was summarized in detail.And finally,this chapter pointed out the purpose and significance of this study based on the current analytical methods.The second chapter developed a CE-ultraviolet?UV?method to detect aliphatic,aromatic and heterocyclic BAs at the same time,which combines with the principles of the direct UV detection and indirect UV detection.The UV absorbing compound need be selected as an UV probe that dissolved in the background electrolyte?BGE?as the co-ion.The analyte ions form the sample zone by the physical replacement with co-ions.And then,analytes owning different UV absorption abilities could be detected by the changes of UV signal at the detection window.In this paper,imidazole?Im?was selected as the UV probe in BGE.Under the precondition of the transfer ratio between visualization agent and the analyte at 1:1,the detecting principle was discussed in detail.The a-cyclodextrin?a-CD?was firstly applied for the separation of BAs.In addition,the influences of the p H of BGE,Im concentration,a-CD concentration,hydrodynamic injection?HDI?time and separation voltage were examined in detail.The optimal BGE: 12.0 mmol/L Im+10.0 mmol/L a-CD?p H 4.50 adjusted by 1.0 mol/L acetic acid?.And other CE condition: detection wavelength 200 nm,separation voltage +15 k V and HDI 0.5 psi?5 s.The thirteen BAs selected were separated under the optimal condition.The limits of detection?LODs??S/N=3?of this method was 0.4³.7 ?mol/L,and the interday relative standard deviation?RSD?of peak area and migration time were 2.5⁴.7% and 0.7¹.3%,respectively.The third chapter aimed at enhancing the sensitivity of CE-UV method developed in second chapter by other injection modes such as electrokinetic injection?EKI?and pressure-assisted electrokinetic injection?PAEKI?.The experimental results showed that both EKI and PAEKI can improve the sensitivity by the effective stacking,and the LODs?S/N=3?reached nmol/L level.Especially,the sensitivity of CE-UV method with PAEKI was the best.Under the injection condition +7.8 k V?-0.1 psi?injection voltage?external pressure?,the optimal injection time for the four BAs by indirect UV detection and 9 BAs by direct detection were 0.35 min and 0.70 min,respectively.Through the effective accumulation by PAEKI,the LODs?S/N=3?of the four BAs by inrect UV detection were 8.7²4.3 nmol/L,and the LODs?S/N=3?of the others by direct UV detection were 0.3⁴.5 nmol/L.Compared with HDI methods,the enhancement factors can reach 318¹408 times.In addition,for the optimization of PAEKI conditions,pressure-BGE velocity and voltage-EOF velocity could be made to find the balance points between the injection voltage and external pressure.It was a more rapid,convenient,systematic procedure with high accuracy.The fourth chapter also developed another CE method for the simultaneous determination of the aliphatic,aromatic and heterocyclic BAs combining with UV detection and capacitively-coupled contactless conductivity detection?C⁴D?.Ionic liquid----1-butyl-3-methylimidazolium tetrafluoroborate?[BMIM][BF₄]?was used in the final BGE for the both detection.The BAs owning UV absorption ability can be analyzed with UV detector,while Spd,Put,HA and Cad which own low or none UV absorption ability can be detected by C4 D detector.The influences of BGE and instrument parameters were studied,in the UV detection and C4 D detection,respectively.And then,the optimal BGE was 8.0 mmol/L [BMIM][BF₄]+ 12.0 mmol/L ?-CD?p H 4.50 adjusted by 0.1 mol/L Na OH?and the other CE condition: HDI 0.5 psi?5 s,separation voltage +15 k V and detection wavelength 200 nm.Under the optimal CE condition,Spd,Put,HA and Cad were qualified and quantified by C4 D detector,and the LODs?S/N=3?were 0.8¹.4 ?mol/L.Other BAs could be analyzed by UV detector,and the LODs?S/N=3?were 0.2¹.1 ?mol/L.The BGE solution that can be applied for the two detectors,can rich the analytical information of method.In addition,this study has great guiding significance for the analysis of underivative BAs by CE coupled with double detectors in the future.The fifth chapter determined the thirteen BAs in beers with a mature liquid-liquid extraction procedure and by CE-UV method developed in second chapter,which verified the practicability and reliability of this analytical method.In addition,due to the complex matrix interference in the actual samples such as human urine and food,the solid phase extraction?SPE?method was also studied in detail by thirteen standard BAs.This chapter explored the preliminary study of the adsorption results by using C18 and MCX columns,which lays a good foundation for the practice applications with CE methods developed in this paper.The sixth chapter stated the conclusions and prospects of this thesis.