In Situ Ionization Mass Spectrometry Analysis Of Carbohydrates

Carbohydrates are the most abundant organic compound in nature,and acting as the energy materials and structural materials of living organisms.They participate in various physiological and pathological activities,playing vital roles in human beings' life.In-depth study of carbohydrates is important for the understanding of their biological functions.However,the identification and structural elucidation of carbohydrates remain a great challenge to scientific community,due to the large diversity,structural complexity and the existence of enormous isomers of carbohydrates.Mass spectrometry is a powerful tool for the detection and structural analysis of chemical substances,and ambient ionization mass spectrometry(AIMS)allows for the direct analysis of raw samples in an open environment,where the analyte ionization process can be performed at atmospheric pressure.Direct analysis real-time mass spectrometry(DART-MS)is one of the rapidly developing AIMS techniques in recent years,with less pre-treatment and fast analysis speed(?10 s/per sample).It has shown various advantages in environmental,pharmaceutical and biomedical analysis,but the studies reported on the carbohydrate analysis are very rare.Therefore,efficient analytical methods for the analysis of carbohydrates based on mass spectrometry,mainly involved AIMS,are established in this thesis including the following aspects:(1)Evaluation of saccharides' structures by the pyrolysis process in DART-MS.In this chapter,a comprehensive study of the analytical patterns of DART-MS was carried out on different types of saccharides standards with a wide range of analytes covering from monosaccharides and disaccharides to oligosaccharides and polysaccharides,with molecular weights ranging from a few hundred Da to several hundred k Da.DART-MS provides an analytical protocol for the efficient detection of all saccharide standards.The systematical study of DART working gases revealed that the use of argon kept relatively higher structural integrity of oligosaccharides which contributes to the analysis of their structures,while polysaccharides with larger molecular weights required helium with higher ionization energy as working gas.At the same time,the analysis of the saccharide standards with larger molecular weights required the higher optimal temperature of the DART working gas.The study in this chapter lays the foundation for the potentially wide application of DART-MS for the analysis of carbohydrates to a great extent.(2)Rapid differentiation of disaccharide isomers by combining TMAH methylation and DART-MS.In this chapter,the identification of disaccharide isomers was achieved by the rapid methylation reaction of tetramethylammonium hydroxide(TMAH)with disaccharide isomers,combined with the real-time detection by DART-MS.The results indicate that the methylation of disaccharides by TMAH occurs as a transient gas-phase reaction in the DART ion source.Eleven disaccharide isomers with different linkages,compositions and conformations yielded different methylation products,which were effectively distinguished.The method was also applied to the analysis of disaccharide mixtures in honey and a variety of disaccharide compounds were detected and identified.Semi-quantitative analysis of sucrose and isomaltose fractions in honey was also performed in this work.In this chapter,a new simple and efficient method for the differentiation of disaccharide isomers with good stability and feasibility is established and proven to be promising in the analysis of real samples.(3)Rapid fingerprint analysis for herbal polysaccharides using direct analysis in real time mass spectrometry.In this chapter,a new method for the rapid fingerprint analysis of herbal polysaccharides was established by combining mechanochemical extraction(MCE)and DART-MS.It was found that different herbal polysaccharides produced characteristic fingerprint profiles on DART-MS,and they could be effectively distinguished by combining multivariate data analysis.Meanwhile,polysaccharides were also differentiated in complex mixtures.In addition,the polysaccharide contents of Dendrobium officinale from different origins were accurately determined by the standard addition method.The direct DART-MS analysis of MCE extracts of herbal polysaccharides allowed the whole process to be completed within minutes.This study makes the fingerprint analysis of herbal polysaccharides much shorter,reproducible and of high application value.(4)Combining online size exclusion chromatography and electrospray ionization mass spectrometry to characterize plant polysaccharides.In this chapter,an analytical protocol of combining size exclusion chromatography and source-induced-dissociation-electrospray ionization mass spectrometry(SEC-SID-ESI-MS)was developed for the on-line separation and structural elucidation of intact polysaccharide macromolecules.The method was optimized and successfully applied to the analysis of plant polysaccharides using dextran(MW 80 k Da)as the optimization sample.The plant polysaccharides were first separated by size exclusion chromatography and the eluted fractions were subsequently transported online to electrospray ionization mass spectrometry for detection.By introducing source-induced-dissociation(SID)in electrospray ionization,plant polysaccharide macromolecules can be cleaved into smaller molecular weight fragments(m/z < 3000),thus realizing the detection of polysaccharide macromolecules with mass spectrometry.This work is a viable option for obtaining valuable structural information from intact polysaccharides.