Atractylodis Rhizoma Volatile Oil Fingerprint Study And 2 Strains Of Fungi On The Tissue Culture Plantlets Of Atractylodes Lancea (Thunb.) DC

In recent years,the international demand for ATRACTYLODIS RHIZOMA is increasing.Atractylodes lancea(Thunb.)DC is the genuine herb of ATRACTYLODIS RHIZOMA.While the plant resources and the ecological environment of the ATRACTYLODIS RHIZOMA authentic region has been seriously destroyed.In addition,the wild Atractylodes lancea(Thunb.)DC has the low reproductive ability.As a result,the resource of authentic Atractylodes lancea(Thunb.)DC is at the edge of extinction and a growing number of the no authentic ATRACTYLODIS RHIZOMA circulates on the market.Therefore,it is necessary to assess the quality of ATRACTYLODIS RHIZOMA in different regions,study differences and relationships between the suthentic and the no authentic ATRACTYLODIS RHIZOMA,and establish a quality evaluation system of ATRACTYLODIS RHIZOMA.Thus,clinical curative effects of ATRACTYLODIS RHIZOMA can be further determined.In this study,we firstly use silica gel chromatography separating four kinds of active ingredients in ATRACTYLODIS RHIZOMA:atractylon,hinesol,?-eudesmol,and atractylodin.In petroleum ether-ethyl acetate(95:5)layer,under the Chromogenic condition of diaminobenzene formaldehyde,atractylon is purplish red,its Rf value is 0.82,and its molecular weight is 216.33.After heating,atractylodin is blue blob,its Rf value is 0.67,and its molecular weight is 182.23.?-eudesmol is blue after heating,its Rf value is 0.24,and its molecular weight is 222.37.Hinesol is also blue after heating,its Rf value is 0.23,and its molecular weight is 222.37.According to chromatography,mass spectrometry,and nuclear magnetic data,structures of these compounds have been determined.Via purity test with area normalization method,we find the purity of these four compounds is all above 98%.Recent years,the chromatographic fingerprint similarity analysis(SA)combined with multi-ingredients quantitative determination(MD)has been confirmed and developed for the Chinese Herbal Medicine(CHM)quality control.In Chinese medicinal system,identification of CHM is not only depending on botanical classification standards,but producing areas are obtained great attentions.However,in modern researches,the information of producing areas of CHM is lack.Rhizome Atractylodes is a typical CHM for dampness-removing.We use the GC-FID establishing a method combined SA and MD for the quality control of Rhizoma Atractylodis.20 batches of Rhizoma Atractylodis from different areas are studied.Then based on the information contained in the fingerprint,we draw a dendrogram by Hierarchical Clustering Analysis(HCA).According to the results of HCA and MD,we establish a producing areas discrimination model via the Canonical Discriminant Analysis(CDA).Finally,our experiments have been carried out to verify the model.Results of methodology study show that the method we established has higher precision,stability,repeatability,and accuracy.Based on the result of experiments testing,the producing area model established by CDA is proved reliable.Our work on the quality control and producing area discriminant of Rhizoma Atractylodis provides an effective method.CDA is a useful tool for the identification of CHM.The strategy of establishing the method proposed in this study can be applied in other CHM.Finally,we examine effects of two fungal strains(Fusarium oxysporum and Fusarium spp.)on the volatile oils of tissue culture plantlets of Atractylodes lancea(Thunb.)DC.After a serious of observation of leaves' enzyme activity and volatile oil components of roots,we find that the atractylon,hinesol,and ?-eudesmol are not main components in tissue culture plantlets.In the Treatment group that inoculated with two fungal strains at the same time,we find that the enzyme activity and volatile oil component have remarkable differences.phenylalanineammonialyase(PAL)and polyphenol oxidase(PPO)reached peaks two days after inoculation,cata-lase(CAT)and superoxide dismutase(SOD)reached peaks four days after inoculation,and the accumulation of volatile oil peaks in the second day.Then the enzyme activity and volatile component rapidly reduce.The existence of antagonistic microbes stimulates the increase of enzyme activity and the accumulation of volatile components appeared at early stage.However,in this experiment,the increase of enzyme activity and the accumulation of volatile oil are not sustainable.Based on the objective and scientific evaluation of traditional medicine quality,we separate active ingredient of ATRACTYLODIS RHIZOMA,analyze chromatographic fingerprint of ATRACTYLODIS RHIZOMA in different regions,and study effects of pathogen infection on the tissue culture plantlets of Atractylodes lancea(Thunb.)DC.Our researches on ATRACTYLODIS RHIZOMA active substances has good application value,which provides a standardized reference for objectively evaluating the quality of ATRACTYLODIS RHIZOMA a certain guidance for the improvement of quality,and the prevention of pathogen in the cultivation of ATRACTYLODIS RHIZOMA.