| Isatidis Radix,the root of Isatis indigotica,called Banlangen in Chinese is usually used Traditional Chinese Medicine(TCM)being valued to have antiviral,antibacterial,antioxidant,immune enhancing activities and traditionally used for treatmenting seasonal febrile diseases,mumps,pestilence,eruptive diseases,inflammatory diseases with redness of skin,and sore throat.At present,the study of chemical composition and the function mechanism of antiviral research in Isatidis Radix is not deep enough.This dissertation consists of four parts,which belong to the chemical constituents of Isatidis Radix.The studies are as follows:In Chapter 1,a detailed review about latest research progress of chemical components,pharmacological activities and quality control of Isatidis Radix is summarized.Besides,the research approach was carried out by analyzing present state of Isatidis Radix research.In Chapter 2,chemical constituents from the effective extraction of Isatidis Radix were studied.Many chromatography means were used in separation and purification,and the structures of all compounds were identified by the means of spectroscopic analysis and physicochemical properties.As a result,20 compounds were obtained from the bioactive extract of Isatidis Radix,and their structures were elucidated as:salicylic acid(1),p-hydroxybenzoic acid(2),uracil(3),adenosine(4),guanosine(5),uracil riboside(6),sucrose(7),syringin(8),Indole-3-acetonitrile-6-O-β-D-glucopyranoside(9),indirubin(10),isaindigodione(11),2,4,(1H,3H)-quinazolinedione(12),Myo-inositol(13),7S,8R,8’R-(+)-lariciresinol-4,4’-bis-O-β-D-glucopyranoside(14),(9E,2S,3R)-3-hydroxymethyl-2-N-(2’-hrdroxynonacosanoyl)-9-tridecasphingenine(15),cyclo-(Tyr-Leu)(16),(1’R,2’R,3’S,4’R)-1,2,4-triazole nucleoside(17),2-amine-4-quinlinecarboxylic acid(18)and 3-thiophenecarboxylic acid(19),2-pyrrolidone-N-methylene-β-D-arabinoside(20).Among them,compounds 16-19 were obtained from this plant for the first time,compound 20 was new compound.In Chapter 3,the composition analysis study on the Isatidis Radix of different regions were performed,and the results were summarized in four sections.Section 1:the composition analysis study of the alkaloids.The contents of epigoitrin,indole-3-acetonitrile-6-O-β-D-glucopyranoside,2,4,(1H,3H)-quinazolinedione in Isatidis Radix were simultaneously determined with HPLC spectrophotom-etry method.The result showed that almost all of these Isatidis Radix were rich in this three alkaloids,and the contents of epigoitrin were higher than other alkaloids.Section 2:the composition analysis study of the nucleosides and nucleobases.To explore the profiles of the nucleosides and nucleobases in Isatidis Radix,an HILIC-UPLC-MS/MS method was established and validated.The method was successfully applied for the simultaneous characterization and quantitation of 16 nucleosides and nucleobases in ten samples which contained from 8 cultivation regions.The result showed that almost all of these Isatidis Radix were rich in nucleosides and nucleobases,but their contents were obviously various.The highest concentration was found to be from Shanxi,whereas the lowest was from Heilongjiang.The average contents of xanthine are relative higher than the other tested compounds,while the thymidine are relative lower than the other tested compounds.Section 3:the composition analysis study of the free amino acids.With the application of principal component analysis,a rapid,sensitive and non-derivative method for the determination of 22 free amino acids in Isatidis Radix by HILIC-UPLC-MS/MS was established and validated.The results showed that almost all of these samples were rich in free amino acids,but their contents were obviously various.As for the individual compounds determined in the experiments,proline was found to be the most abundant free amino acid in all samples,and its average content in these investigated samples was 7.28 mg/g,which accounted for more than 33%of the total amino acids tested in this study.However,the hydroxyproline was the lowest except for not detected.8 kinds of essential amino acids were widely distributed in all the samples.In addition,without the protein amino acids,three nonprotein amino acids,including y-amino-butyric acid,hydroxyproline,and citrulline,were found in the Isatidis Radix samples.The results of the principal component analysis showed that the chemical composition of Shanxi,Anhui 1,Sichuang were different from other origins.Section 4:the composition analysis study of flavonoids.A comparative study on the total flavonoids distribution in Isatidis Radix in different origins was carried out using UV-Vis spectrophotometry.The results showed that content of total flavonoids in Isatidis Radix was relative lower,and their contents were obviously various.The average content of total flavonoids as follow:Neimengu(0.180%)>Henan(0.124%)>Gansu2(0.115%)>Heilongjiang(0.109%)>Hebei(0.084%)>Sichuang(0.064%)>Shanxi(0.048%)>Gansul(0.023%)>Anhui 1(0.012%)>Anhui2(0.006%).In Chapter 4,a simple,rapid and sensitive method was developed for the analysis of indole-3-acetonitrile-6-O-β-D-glucopyranoside in rat plasma by liquid chromatography with tandem mass spectrometric(LC-MS).The results showed the specificity,precision,accuracy and recovery of this method was suitable for determination indole-3-acetonitrile-6-O-β-D-glucopy-ranoside in rat plasma. |
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