| Metabolic analysis can detect and interpret the final products in biological processes,which can help studing the actual physiological processes in organisms.The metabolic profile based on biological fluids(mainly including serum,plasma,urine and cerebrospinal fluid)can not only distinguish patients from healthy controls,but also find and identify potential biomarkers of diseases.Therefore,it has become one of the irreplaceable means in disease diagnosis and monitoring.So far,metabolic analysis based on body fluids has been successfully applied to diagnose many diseases,such as cancer,stroke,mental disease,autoimmune disease.And it also has found a series of potential biomarkers for accurate diseases diagnosis or diseases classification.Compared with other technology for metabolic detection,Laser desorption/ionization mass spectrometry(LDI-MS)has shown its great advantages in metabolic diagnosis by achieving high throughput extraction and rapid acquisition of metabolic information for biological samples nearly non-pretreatment.So it has been regarded as an effective tool for metabolic detection with broad commercial application potential.During the LDI-MS process,analytes firstly need to be mixed with matrix,then the metabolites are effectively desorbed and ionized under the irradiation of ultraviolet laser.However,uneven distribution of matrix and analytes due to manual operating procedures can affect the reproducibility of LDI-MS.In addition,low desorption/ionization efficiency has also been a major bottleneck in the clinical application of LDI-MS.Therefore,it is extremely urgent to reasonably design and apply LDI-MS platform to improve the reproducibility of LDI-MS detection from the sample pretreatment procedures and improve the desorption/ionization efficiency of LDI-MS.This thesis constructed and studied a ternary LDI-MS chip with enhanced desorption/ionization efficiency for metabolic fingerprint which improves detection reproducibility through a uniform surface of the chip,improves desorption/ionization efficiency by designing and optimizing the structural parameters of the chip.Then,we studied the mechanism of its high desorption/ionization efficiency,and explored its practical application of this ternary chip.The thesis mainly includes the following three parts:The first part of this thesis mainly reviewed and summarized the metabonomics research based on the LDI-MS.Firstly,we introduced the significance and problems of current metabonomics research;Then,we summarized the matrices and chips have been applied in LDI-MS metabolic detection and emphatically reviewed the performance and application of these platform.Finally,we explained the purpose and major content of this thesis.In the second part of the thesis,we constructed a LDI-MS ternary chip with enhanced desorption/ionization efficiency for metabolic fingerprint identification,and explored the mechanism of its high desorption/ionization efficiency.Firstly,a layer of uniform mesoporous silica membrane was grown on ITO slide through two-phase growth strategy as the substrate of the LDI-MS chip to provide support and heat insulation.Then gold nanoparticles(Au NP)were deposited on the silica layer through sputtering.After that,1H,1H,2H,2H perfluorodextrinol was used to form a highly perfluorinated nanostructure on the gold layer through gold sulfur coupling,Finally,LDI-MS chip with appropriate hydrophobicity and high desorption/ionization efficiency was obtained.In the process of synthesis,we optimized the parameters of the chip,including the synthesis conditions of mesoporous silica films,the sputtering conditions of gold nanoparticles and the loading conditions of initiators to obtain excellent LDI performance.In order to explain the enhanced LDI performance,we used benzylpyridinium salt(BP)as "chemical thermometer" to study the detailed desorption/ionization mechanism of the LDI-MS chips.This work constructed a ternary LDI-MS chip and studied the mechanism of its high desorption/ionization efficiency,which will bring new inspiration for the future of designing nanomaterial platform for high-performance metabolic analysis and large-scale diagnosis.The third part of the thesis mainly discussed the application of metabolic fingerprint identification by the LDI-MS ternary chip with enhanced desorption/ionization efficiency.Firstly,in order to verify the feasibility of the chip in detecting actual biological fluids,we tested its salt tolerance and protein tolerance of the chip.Then,in order to explore the sensitivity of the chip,we tested the limit of detection of the chip in detecting metabolites.Then,the LDI-MS ternary chip was used to directly analyze metabolites(～1 μL)to prove its repeatability and reliability in actual samples detection.Finally,the optimized LDI-MS chip was used to obtain the urine fingerprint,which can combine with machine learning to distinguish between kidney stone patients and healthy controls.This part explored the application of LDIMS ternary chip with enhanced desorption/ionization efficiency in metabolic fingerprint identification,providing a new strategy for developing a new platform for metabolic fingerprint identification.To sum up,this thesis provides a reasonable idea to design new LDI-MS chip through the ternary LDI-MS chip.It verifies the importance of the structural parameters for the improvement of the detection performance,explores the structure-activity relationship between the chip structural parameters and the enhanced desorption efficiency.Futhermore,it also obtains the urine metabolic fingerprints of kidney stone patients and healthy controls based on LDI-MS,realizes the diagnosis of kidney stone,which is helpful to design high-performance metabolic analysis and large-scale diagnosis application platform based on nanomaterials. |
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