Application Of Carbon Nanotubes In Separation Of Peptides By Capillary Electrophoresis And Extraction And Separation Of Saikosaponin

Carbon nanotubes（CNTs）have been widely used in various fields due to their unique structure and excellent properties.For example,excellent electrical conductivity,excellent mechanical properties,easy modification,unique hollow pipe structure,and large specific surface area have attracted a lot of attention in recent years.In this research,carbon nanotubes are applied in the separation of peptides by capillary electrophoresis.At the same time,carbon nanotubes are used in the extraction and separation of traditional Chinese medicine saikosaponin.This thesis is divided into six chapters:The first chapter firstly introduces the structure of CNTs and methods of dispersion CNTs.Secondly it introduces the research progress of carbon nanotubes in the field of separation and adsorption.Thirdly it introduces the pharmacological effects of traditional Chinese medicine Bupleurum and the commonly used extraction methods of its active ingredients.The second chapter explores the method of MWCNTs.The dispersive sonication time,the comparison of different surfactants and three kinds of carbon nanotubes.Ultraviolet-visible photometer was used to detect the absorbance and scanning electron microscope（SEM）to characterize the morphology of the material dispersion.After exploring various conditions,it was found that 1 mg of carboxyl multi-walled carbon nanotubes（MWCNTs-COOH）were dispersed into solution including 17.5 m M surfactant SDS and 10%（v/v）N-butanol and sonicated 20 min,the carbon nanotubes were well dispersed.In the third chapter,the dispersed carbon nanotubes were used for peptide separation in capillary electrophoresis.The dispersed MWCNTs-COOH served as a quasi-stationary phase that can reduce peptides adsorption on the capillary wall and can provide more sites to improve the peptides separation.By adjusting the concentration of phosphate,the concentration of sodium dodecyl sulfonate（SDS）,the concentration of carbon nanotubes,pH and voltage conditions were optimized.Under the optimal conditions,10 m M phosphate,8 mg/L MWCNTs-COOH,pH 8.85,and the voltage of 12 k V,six peptides DE-11;DE-10;Met-Enkephalin;Bombesin;Bradykinin and Oxytocin were effectively separated.In the fourth chapter,we used multi-walled carbon nanotubes（MWCNTs）,carboxyl multi-walled carbon nanotubes（MWCNTs-COOH）and hydroxyl multi-walled carbon nanotubes（MWCNTs-OH）to study the extraction and separation of saikosaponin from the Chinese medicine Beichahu.At first,the chromatography and mass spectrometry conditions of five saikosaponins were optimized on the LC-MS platform,and the LC-MS quantitative analysis method of five saikosaponins was established.The ability of absorption saikosaponin was investigated,and it was found that the concentration of saikosaponin had no significant effect on the adsorption;there was no large difference in the adsorption capacity of the three CNTs on saikosaponin A,B₁,and D,but for saikosaponin B₂ and C,the efficiency of MWCNTs-OH was significantly higher than the other two types of carbon nanotubes.Finally,a water-saturated n-butanol solution was selected as the eluent,and it was found that it had a certain elution capacity for the five saikosaponins adsorbed by MWCNTs and MWCNTs-OH.The fifth chapter studies the enrichment ability of MWCNTs,MWCNTs-COOH and MWCNTs-OH on saikosaponin in Bupleurum extract.Because saikosaponin and sugar coexist in the Bupleurum extract,their polarity are similar,but the sugar content is much higher than that of saikosaponin.Therefore,the use of a suitable adsorbent can eliminate sugar interference and specifically adsorb saikosaponin.Firstly,we configured a mixed standard solution of saikosaponin and sugar.Based on the LC-MS platform,whether the three carbon nanotubes of MWCNTs,MWCNTs-COOH and MWCNTs-OH can specifically adsorb saikosaponin in the presence of sugar was performed.It was found that the adsorption rate of sugar by three kinds of carbon nanotubes is very low,below 10%,and the adsorption performance of saikosaponin is high,which shows the specific adsorption ability of carbon nanotubes to saikosaponin.We explore Bupleurum extract,which proves that carbon nanotubes can be used for the enrichment of saikosaponin in traditional Chinese medicine.In chapter 6,we summarized the whole thesis and looked into the future prospect.