| Mass spectrometry technology with many advantages,such as high sensitivity(detectable pg level),fast analysis speed,intuitive information and so on,brings much more support for drug development.The nano-ESI-FTMS technique enables the qualitative and quantitative identification of amides and peptides and is usually applied for the proteomics researching and the identification for clinically endogenous peptide hormone markers.The "mass spectroscopic imaging" is a modern analytical technique that mainly represents "in situ bioinformatics".It is characterized by the use of high-resolution mass spectrometry to achieve spatial distribution and metabolic changes of endogenous biomolecules after drugs and other exogenous substances acting on the body,regardless of the active form in which the drug enters the body or whether there is a barrier from the blood-brain barrier.As a result,an on-line research mode has been explored for the mechanism of drug action,and it has an irreplaceable advantage in clinical use for targeted screening of tumors.Ginseng radix is the dry root of Panax ginseng C.A.Meyer which belongs to the Panax genus in Araliaceae family.Previous studies on ginseng have focused on the isolation and identification of saponin components,and also activity evaluation and mechanism research of them.Studies have shown that ginseng proteins and peptides content is three times as much as saponin’s.However,there are still few studies on ginseng peptides,especially the type,the combination for amino acids.In addition,ginsenoside Rg1 as the main component of ginsenosides,whose activity performance is considered consistent with the efficacy of ginseng.Rg1 has significant activity on cerebral system diseases,but there are still obvious deficiencies in the differential expression and metabolism of endogenous substances in brain tissue regulated by Rg1.One of the reasons is the complex transmitters and hormones composition of brain which makes it difficult to fully characterize the brain function for the detection of one or two indicators.Secondly,due to the influence of analytical techniques,it is difficult to achieve a complete visual representation of endogenous substances;thus far,the research on the mechanism of Rg1 acting on the brain’s central nervous system and the manifestation of endogenous differences has not been clearly clarified.Therefore,for the purpose of constructing a set of methods to detect the peptides clearly and to find the difference in brain tissues after taken saponins,this study takes traditional Chinese medicine ginseng(Panax ginseng C.A.Meyer.)as an object material and uses the nanoLCMS/MS system to identify the endogenous peptide components in this medicine.A set of methods and databases is wished to be established which also could be used to analysis and detect other plant peptides as a successful example.Here,we use mass spectrometry imaging technology to analyze the "in situ information" change in mice brain tissue after taken Gensenoside Rg1,a typical saponin components in Panax ginseng,which also will to be wished to provide a reference for the similar study for evaluate the drug efficacy or mechanism clarification.So far,the results are the following:1.306 ginseng peptides were characterized by data mining and de novo sequencing based on nanoLC-MS/MS analysis combined with different fragmentation methods of mass spectrometry,of which 123 peptides were included in the database and 183 peptide sequences were obtained by de novo sequencing.Out of the 306 sequences,a total of 120 sequences contained post-translational modifications,which exhibiting the richness of ginseng polypeptide sequences.This result enriches the database of ginseng endogenous peptides,and also provides the basis for further biological research and drug development.2.UPLC-TOF/MS/MS technique was used to map the metabolic profiles of hypothalamus,cerebellum,prefrontal cortex,hippocampus,striatum and other regions of mice’s brain tissues after intragastric administration of Rg1.Comprehensive T-test and pattern recognition results can distinguish between the Rg1 administration group and the control group in each brain region.The striatum is the brain region with the most significant differences in metabolic profiles.Through targeted MS/MS analysis of differential ions,a total of 66 compounds were identified,including 23 amino acids and their derivatives,intermediates of the tricarboxylic acid(TCA)cycle,small amounts of lipids and other substances related to the nervous system,involving alanine,aspartate and glutamate metabolic pathways,and the TCA cycle pathway.3.The MALDI-FT/ICR-MS in situ mass spectrometry imaging system was used to characterize the regulation of the brain histochemical spectrum of ginsenoside Rg1.The in situ spatial distribution density maps of 2000 ions were obtained.In combination with bioinformatics tools,a total of 67 differential ions were identified in the brain-derived material database,65 of which were lipids,covering 12 lipids with different physiological functions.The differences involve multiple brain areas such as striatum and corpus callosum which have been shown to be closely related to the occurrence of neurological diseases characterized by motor dysfunction and learning and memory impairment,demonstrating that the potential therapeutic effects of Rg1 in combating and alleviating the associated central nervous system disorders.Above research results not only enrich the research contents of ginseng,but also give an important support for the plant polypeptide database collecting,also provides a detailed mass spectral data support for the characterization and sequencing detection for peptides from plants,including plants from ginseng genus,meanwhile,the results taken by UPLC-TOF/MS/MS and MALDI-FT/ICR-MS techniques to detect the difference by Rg1 caused in brain tissues were helpful to know or reveal the mechanism of cerebral disease for ginseng saponins Rg1 treatment role in the brain. |
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