Functional Materials Based Direct Ionization Mass Spectrometry For Rapid Analysis Of Complex Biological Samples

Mass spectrometry?MS?is widely applied in various fields of science due to its excellent sensitivity and specificity for chemical detection.Since mass spectrometry uses electric and magnetic fields to accelerate,deflect,separate and detect charged gas-phase ions,the preparation of charged molecules is the premise of mass spectrometry analysis,i.e.,the ionization of samples.Conventionally,for the analysis of complex matrice samples,a series of tedious sample purification processes?such as grinding,digestion,centrifugation,desalting,dilution,fractionation,extraction,drying,reconstitution,chromatography separation,etc.?are needed to remove the sample matrice before ionization process due to the limitation of classic ionization technology.Thus,high-throughput analysis of complex samples is an important topic in mass spectrometry research.The analysis of complex matrice samples has become considerably simpler and faster with the development of ambient mass spectrometry?AMS?,which realized the direct analysis of complex samples without cumbersome sample pretreatment.Through more than ten years of rapid development,AMS has made remarkable progress and wide application in forensic analysis,food safety analysis,drug screening,clinical diagnosis,and life science.For AMS,the key technology is ambient ionization method.Currently,various ambient ionization technologies have been introduced for the analysis of complex gaseous,liquid,and solid samples.However,in practical study,the ionization suppression effect and matrice interference from the sample matrice would greatly influence the sensitivity,repeatability and quantitative performance of AMS,especially in the determination of trace target analytes in complex samples.Therefore,it is necessary to further improve the analytical performance of ambient ionization technology in the rapid analysis of complex samples.Recently,the detection sensitivity and chemical selectivity of AMS?or direct MS?analysis have been significantly improved under the strategy that introduce functional materials for rapid matrice clean-up and target analytes enrichment prior to direct ionization.On the one hand,functional materials can be used to rapidly adsorb and enrich trace target components from the complex matrice.On the other hand,it is necessary to design appropriate and facile ionization source devices for rapidly ionizing the target analytes,so as to realize rapid detection of trace components in complex matrice samples.The rapid capture of trace components from complex matrice by functional materials combined with the rapid direct ionization source will improve the detection sensitivity and quantitative performance of ambient mass spectrometry.The purpose of this doctoral research is to develop methods combining functional materials and ambient ionization technology to rapid analysis of complex matrice samples.In principle,specific functional materials are used for the selective adsorbtion of target components which quickly remove the matrice components and capture the target components for post ionization process.Technically,facile and easy-to-use ionization devices were designed to rapid ionize the target analytes to be tested.In terms of application,the methods proposed in this study could rapid detect trace analytes in complex matrix samples,showing potential application in food safety analysis and clinical research.The main contents of this thesis are as follows:1)Internal extractive electrospray ionization mass spectrometry for quantitative determination of fluoroquinolones captured by magnetic molecularly imprinted polymers from raw milk.In this study,solid-phase extraction based on magnetic molecularly imprinted polymers coupled with internal extractive electrospray ionization mass spectrometry?MMIP-SPE-iEESI-MS?was designed for the quantitative analysis of trace fluoroquinolones?FQs?in raw milk samples.FQs in the raw milk sample?2 mL?were selectively captured by the easily-lab-made magnetic molecularly imprinted polymers?MMIPs?,and then directly eluted by100?L electrospraying solvent?2%ammonia in methanol,w/w?biased with+3.0 kV to produce protonated FQs ions for mass spectrometric characterization.For all the samples tested,the established method showed a low limit-of-detection(LOD?0.03?g L^-1)and a high analysis speed??4 min per sample?.Under optimized experimental condition,satisfactory quantitative performances for norfloxacin,enoxacin,and fleroxacin were achieved by MMIP-SPE-iEESI-MS.The analytic performance for real sample analysis was validated by a nationally standardized protocol using LC-MS,resulting in acceptable relative error values from–3.8%to+4.7%for 6 tested samples.The experimental results demonstrate that MMIP-SPE-iEESI-MS is a new strategy for the quantitative analysis of FQs in complex biological mixtures such as raw milk,showing promising applications in food safety control and biofluid sample analysis.2)Selective enrichment of phosphopeptides and phospholipids from biological matrices on TiO₂ nanowire arrays for direct molecular characterization by internal extractive electrospray ionization mass spectrometry.In this study,trace phospholipids in human plasma,whole blood and undiluted human urine,as well as phosphopeptides in protein digest were selectively captured on a homemade array of TiO₂ nanowires for sensitive characterization by internal extractive electrospray ionization mass spectrometry?TiO₂-iEESI-MS?.Sequential release of captured chemicals from TiO₂ array was achieved by tuning pH of the extraction solvent.A single sample analysis,including sample loading,chemical extraction and MS detection,was accomplished within 3 min.Under optimized experimental condition,satisfactory quantitative performances for phospholipids and phosphopeptides in complex matrice were achieved by TiO₂-iEESI-MS.Based on the orthogonal partial least squares discriminant analysis?OPLS-DA?,TiO₂-iEESI-MS patterns from the blood of 46 patients with ovarian cancer were confidently discriminated from the MS patterns of 46 healthy volunteers.The experimental results indicate the strong potential of TiO₂-iEESI-MS approach for the selective detection of trace phosphopeptides and phospholipids in various biological matrices with high sensitivity,high specificity,low sample consumption,and high throughput.3)Determination of phospholipids from biofluid samples by magnetic TiO₂ nanocomposites based on dispersed solid phase microextraction coupled with nano-electrospray ionization mass spectrometry.In this study,phospholipids in biological samples including human plasma and single cells were selectively captured on magnetic Fe₃O₄@TiO₂ nanocomposites for sensitive molecular characterization by dispersed solid phase microextraction coupled with nano-electrospray ionization mass spectrometry?d-SPME-nanoESI?.In-capillary extraction of phospholipids from small volume of biological sample??1?L?was performed with 50?g Fe₃O₄@TiO₂ nanocomposites which then collected inside a glass capillary tube via an external magnet.The extracted phospholipids were desorbed using 1?L ammonia methanol?1.5%,w/w?within the capillary tube and directly pushed to the connected nanoESI tip for mass spectrometry interrogation.A single sample analysis was accomplished within 3 min.Phospholipids in plasma samples from ovarian cancer patients,pancreatic cancer patients,and healthy volunteers were studied using d-SPME-nanoESI-MS.The phospholipids information from cancer specimens were used to in comparison with the phospholipids molecular patterns from healthy volunteers with the assistance of orthogonal partial least squares discriminant analysis?OPLS-DA?.Furthermore,phospholipids in two types of single cell were also profiled using d-SPME-nanoESI-MS.The experimental results indicate the strong potential of d-SPME-nanoESI-MS for the selective detection of trace phospholipids in various biological matrices with high sensitivity,high specificity,low sample consumption,and high throughput.