| Many drugs and pesticides are chiral compounds,and their enantiomers have great differences in biological activity,pharmacodynamic action,toxic effects and metabolic pathways.Generally,only one enantiomer can match the targeting molecule.It shows high drug activity,and the other enantiomers are often ineffective and even harmful to humans.Therefore,to comprehensively control the quality of chiral drugs and scientifically evaluate the toxicity of chiral pesticides,it is necessary to determine the content of enantiomers,which is of great significance to ensure people's medication safety and food safety.Since the two enantiomeric structures are extremely similar and generally exist in complex systems,the separation of enantiomers becomes very difficult,and the development of separation materials and techniques with independent intellectual property rights,good separation selectivity,high resolution efficiency and moderate price is the key to solving the above-mentioned chiral safety problems.Derivatized cyclodextrin as a stationary phase ligand,which has a unique cavity structure,exhibits excellent chiral recognition characteristics through cavity inclusion and portal hydrogen bonding,and cyclodextrin is more soluble,better chemical structure stability and more flexible use in a variety of chromatographic modes than cellulose.It is an economical and practical chiral separation material and has broad application prospects.This thesis focuses on the preparation of new cholesterol and phenylureido-based mono-derivatized cyclodextrin stationary phases,chromatographic evaluation,mechanism and their practical application,including the following aspects:1.The differences in enantiomeric activities of common chiral drugs and chiral pesticides and their separation methods were reviewed.The performance and characteristics of several types of chiral separation materials commonly used in high performance liquid chromatography were summarized.The research,application and development trend of cyclodextrin stationary phases were prospected,which provides the basis of the thesis for the establishment of new cholesterol derivatized cyclodextrin stationary phases.2.For the first time,a mono-cholesterol-derivatized?-cyclodextrin was synthesized,which is a novel supramolecular compound with both hydrophobicity,hydrophilicity and encapsulation ability.It was bonded to the surface of the ordered mesoporous silica SBA-15 by a coupling agent to obtain a novel cholesterol-?-cyclodextrin bonded phase?CHCDP?.The chemical structures of the ligand and its stationary phase were characterized by infrared spectroscopy,mass spectrometry,nuclear magnetic resonance,elemental analysis,etc.The binding amount of three different batches of the prepared stationary phases was about 0.100.13?mol/m2.The preparation method is simple with good reproducibility.3.The reversed-phase chromatographic performance of the new stationary phase was evaluated using benzene homologs,polycyclic aromatic hydrocarbons,positional isomers,cholesterol and statin hypolipidemic agents as achiral probes.It was found that CHCDP can completely separate nine polycyclic aromatic hydrocarbons and five benzene homologs within 30 min.The result was showed that the introduction of cholesteryl can enhance the hydrophobicity of the cyclodextrin stationary phase and significantly improve its reversed-phase chromatographic performance.The ln k'of the benzene homologue was linear with the number of methylene groups?CH2,n?(ln k'=0.4355n-0.0702,R2=0.9351,the logarithm of selectivity factor ln??CH2?=0.4355),indicating CHCDP has strong reversed-phase chromatographic performance,similar to ODS(ln k'=0.5312n+0.2985,R2=0.9950,ln??CH2?=0.5312),and has a wide range use in reversed-phase chromatography.CHCDP could also separate effectively the positional isomers?o,m,p?of nitroaniline and hydroxybenzoic acid because CHCDP retained the hydrogen bonding in ports and inclusion in cavity of the cyclodextrin in addition to hydrophobicity with strong spatial recognition.The atorvastatin?44.13 min?with carboxyl group retained much stronger on CHCDP than lovastatin?2.93 min?without carboxyl group,which was significantly associated with hydrogen bonding.In summary,the cholesterol cyclodextrin-based stationary phase is a kind of new separation materials with multiple action sites for various analytes.4.A directly use different classes of acid-alkaline chiral drugs such as flavanones,triazoles,amino acids,?-blockers as probes to evaluate the new stationary phase in reversed phase mode and polar organic mode,which was also convenient for the development of analysis methods for related chiral drugs in the next step.In the reversed-phase mode,using simple methanol-water as the mobile phase,CHCDP exhibited higher separation selectivities for flavanones,triazoles and amino acids,and the resolution of 2'-hydroxyl flavanone two enantiomers was 1.94,the hexaconazole was 1.91,the dansyl-serine was 2.15,and the analysis time was shorter?<20 min?.The above solutes contain hydroxyl groups in the vicinity of the chiral carbon,and the steric hindrance is also relatively small,so hydrogen bond,inclusion and cholesterol steric hindrance should play an important role in the chiral separations.In the polar organic mode,CHCDP can resolve seven?-blockers,in which atenolol?Rs,1.57?,metoprolol?1.48?,esmolol?1.43?and arotinolol?1.54?can achieve baseline separation.This is because the linear molecules,such as atenolol easy to enter the cyclodextrin cavity to form inclusions,and hydrogen bonding with the amide group in order to chiral recognition.It is indicated that the hydrophobic cholesteryl group can improve the reversed-phase chromatography of CHCDP for chiral analytes by synergistic chiral separation.The prepared CHCDP is a kind of multifunctional separation materials with good chiral and achiral separation capabilities.5.Based on the optimization of chromatographic separation and mass spectrometry detection conditions,the new stationary phase was applied to the actual sample detection of chiral pesticide enantiomers in fruits and vegetables.The QuEChERS method was used for sample extraction.Combined the sample purification with PSA,GCB and Fe3O4 self-assembled magnetic materials,methanol-0.1%formic acid?35/65,v/v?was used as mobile phase to establish a analysis method for simultaneous rapid resolution and quantification of the enantiomeric residues of six pesticides,such as fluconazole,hexaconazole and triticonazole in cucumbers and tomatos.A new LC-MS/MS method for The multi-ion monitoring?MRM?and positive ion mode of electrospray ionization mass spectrometry were used to simultaneously measure the enantiomers of the above three triazole pesticides within 30 min.All enantiomers showed good linear correlation?R?0.9994?in the concentration range of25-500?g/mL,and low the detection limits?LODs<0.6?g/kg,LOQs<2?g/kg?,high recoveries?89.8697.19%?,good reproducibility and high stability?intra-day and inter-day RSDs 2.85.9%,n=5?.6.Using a variety of sites of cholesterol cyclodextrin stationary phase,the lipids in human blood were screened in groups to expand their new applications in the analysis of achiral compounds.Based on LC-Q-TOF/MS method,the lipids of different blood types in blood were collected rapidly.The statistical analysis of lipid distribution was carried out by using statistical software Markerview and SIMCA-P.Secondly,in order to obtain complete lipid information,the experiment combines positive and negative ion detection mode electrospray ionization?±ESI?and atmospheric pressure chemical ionization?APCI?ion technology to collect complete information on polar and non-polar lipids in blood.Experimental results showed that CHCDP could separate and classify the lipids according to their structure,and the lipid profile map was obtained.The profile could be used to quickly view the difference of lipids.There were210 lipid species in ESI?+?,122 lipid species in ESI?-?,and 1 lipid species?cholesterol?in APCI mode.After subtracting 42 kinds of lipids detected in the three modes,291kinds of lipids were actually found.The proportion of lipids detected was 47.76%of glycerophospholipids,22.33%of glycerides,13.40%of fatty acids,6.20%of sphingolipids,5.84%of glycolipids,and 4.50%of cholesterol esters,which indicated the categories and content of lipids in the blood had large differences.Through the PCA principal component and OPLS-DA model analysis,it was found that the blood type can be clearly classified according to the distribution of lipids,indicating that the distribution of lipids was related to blood type,and further lipids with higher contribution to each blood type were further confirmed.In addition to studying the cholesterol functionalized cyclodextrin stationary phase,this thesis also prepared and evaluated another type of ureido-based functionalized cyclodextrin stationary phases,including trifluoromethylphenylurea,m-chlorophenylurea and 3,5-dimethylphenylurea mono-derivatized?-cyclodextrin stationary phases?FPCDP,CPCDP,MPCDP?.Their chromatographic performance were compared,and the effects of the electron-withdrawing group CF3,Cl and the electron-donating CH3 on the chiral separation performance of the ligand were investigated,namely the?-acid type and the?-basic urea-cyclodextrin stationary phase.The results showed that triazole chiral pesticides and?-blockers with weakly basic drugs were better resolved on?-acid chiral columns with electron-withdrawing groups FPCDP and CPCDP,such as the enantioresolution of hexaconazole is as high as 2.56and 1.78 in MeOH/H2O?35/65,v/v?,and the stronger the electron-withdrawing ability in the ligand,the better the separation effect of the triazole pesticide.The floxacins and ibuprofen acidic chiral drugs were only separated on the?-basic MPCDP column.In addition,flavonoids were better resolved on polar FPCDP and CPCDP,especially on CPCDP.Sulfonylation amino acids showed large differences in the three stationary phases,for example,leucine and isoleucine had a resolution of 2.77 and 3.18 on CPCDP,2.12 and 2.69 on FPCDP,1.88 and 2.68 on MPCDP.The separations of threonine and serine on MPCDP were better with the resolution?1.97 and 1.69?,respectively.The ureido-based functionalized cyclodextrin stationary phases had high stability,rich hydrogen bonding sites,good separation selectivity,and strong separation ability for?-acid or?-alkaline solutes.The chromatographic properties and applications of the urea-based functionalized cyclodextrin stationary phase require further research in the future. |
PDF Full Text Request