Study On The Application Of Pretreatment And High Resolution Mass Spectrum Technologies On Biological Samples

High resolution mass spectrum(HRMS)has extensive applications and shows broad prospects in the modern analytical chemistry filed,with many advantages including high sensitivity and great accuracy.Due to the complexity of the matrix of biological samples,sample clean-up is crucial before bioanalysis since it prolongs the lifespan of MS instruments,as well as contributes to the generation of reliable and reproducible data.Based on online solid phase(SPE)pretreatment and HRMS technologies,we are able to optimize the experiment scheme and establish a trustworthy,specific,sensitive and efficient biological sample analysis method which provides favorable technical support for the drug concentration monitoring in the body.Firstly,U3000 was coupled with Q Orbitrap for the first time to establish a fully automated system named online-SPE-LC-HRMS.In addition,the application in the biological sample analysis was also dicussed for the first time.We performed a systematic comparison and study on the quantitative ability of MS/MS targeted ion fragmentation scan(t-MS~2),targeted-selected ion monitoring(t-SIM)and full MS-SIM(F-SIM)mode for sofosbuvir(SF)in human plasma.Our results showed that t-MS~2 and t-SIM mode exhibited higher sensitivity for sofosbuvir in human plasma compared to F-SIM mode,and the results also indicated that HRMS was a substitute for triple quadrupole spectrometry(QqQ)analysis tools on biological samples.The lower limit of quantitation(LLOQ)was 0.5 ng/mL.The total running time was 10 min.Secondly,we developed a sensitive and automated method for simultaneous quantification of multi-components including lamotrigine(LTG,internal standard)phenobarbital(PB),phenytoin(PHT),carbamazepine(CBZ),and its one of its active metabolites named carbamazepine-10,11-epoxide(CBZE)in human plasma.t-MS~2mode was selected as the quantification mode in this study.Scan points were evaluated based on good separation,the order was LTG,PB,CBZE,PHT and CBZ,they are positive,negative,positive,negative and positive ions separately,the inclusion list and scan groups were evaluated.The total running time of this method was 10 min.This method has been applied successfully in clinical application.Thirdly,two-dimensional(2D)separation technique was firstly incorporated into the design of online-SPE.Basing on the design optimization,an online-dual-SPE system was established,it was developed by Oasis~?HLB and Hypercarb Hypersep to online pretreatment of amphotericin B(AMB),fluconazole(FZ)and flucytosine(FC)simultaneously in human plasma and cerebrospinal fluid,which show rather broad physicochemical properties.The majority of online-dual-SPE stop-flow heart-cutting2D-LC approaches utilized a combination of two different SPE cartridges retention and elution mechanisms:(1)to develop a sample cleanup procedure followed by separate process by SPE cartridges;(2)to transfer selected cuts from one cartridge to another with different properties so that the selectivity of the separation;(3)to apply double internal standards to obtain better quantitative results.The total running time was 7 min.The method is simple,rapid and with broad linearity range,therefore it is used clinically.