The Relationship Between The Micro-ecology Endophytic Fungi And The Accumulation Of Active Constituents Of Cynomorium Songaricum Rupr.

Cynomorium songaricumis a rare parasitic plant that grows in the arid desert area,known to parasitize the plant roots of Nitraria L.and has important pharmacological effects,such as anti-aging,promoting digestion,prevention of cancer and anticancer etc..In this study,at first of all,C.songaricum and its host plant N.tangutorum from Alxa Left Banner were investigated by ITS2 molecular marker analysis.On this basis,dynamic accumulation process of seven active ingredients?polysacharides,flavonoids,tannins,gal--lic acid,protocatechuic acid,catechin and ursolic acid?during the five different periods were studied.At the same time,the study collected and analyzed the data of C.songaricum endophytic fungi which had been studied by the lab staff.The relationship between the ec--ological succession of endophytic fungi and the accumulation of active constituents of C.-songaricum during the five different periods were investigated through Pearson correlation software.The ITS2 sequence characteristics and genetic relationship of C.songaricum and its host N.tangutorum was identified by DNA bar code?DNA barcoding?technology.The results showed that two species were clustered into one clade.Among them,the K₂P genetic distance of the same genus but different species was significantly higher than the intraspecific genetic distance.Moreover,genetic distance between C.songaricum and N.tangutorumis greater than two species intraspecific genetic distance.Using ITS2 sequence structure prediction map can prove the genetic relationship between two species and congeneric more accurately and visually.Sequence structure of the same species C.-songaricum including stem loop number,size and position was consistent.Sequence structure among different species from the same genera N.tangutorum existed a great difference.C.songaricum polysaccharide was determinated by phenol-sulfuric acid method,its content was the highest in March-germination period;total flavone content which was determined by NaNO₂-AI?N0₃?₃ colorimetric method was the highestat the germination stage;determination of tannin content was in accordance with the national pharmacopoeia method,the results showed the highest content was in November-winter period;C.songaricum gallic acid,protocatechuic acid and catechin were extracted by methanol solution and determined by HPLC,the results showed the content of gallic acid was the highest in the June-vigorous period,protocatechuic acid and catechin content was the highest in the germination period;determination of ursolic acid content was by chloroform extraction and HPLC method,the highest content was in the winter period.The extraction and measurement methods of seven compounds in seven areas?Da-Jing,Gu-Lang,Jin-Ta,Alashan-Left-Banner,Gua-Zhou,Nan-Hu and Min-Qin?were the same as before.The results show that the content of polysaccharide was the highest in Da-Jing,followed by the Nan-Hu and Min-Qin;ursolic acid,tannin and gallic acid content was highest in Alashan-Left-Banner;catechin and protocatechuic acid content was highest in Nan-Hu;the content of total flavonoids was highest in Gua-Zhou.But the seven active ingredient content in Gu-Lang,Jin-Ta and Min-Qin region is generally lower,high content area was mainly in Alashan-Left-Banner,Nan-Hu and Da-Jing.Through the analysis of the previous data about separation and identification of C.songaricum endophytic fungi,the results showed 91 strains endophytic fungi including29 representative plant were separated and identified successfully from 15 healthy plants about 400 pieces of tissue,their homology were more than 97%.Through the statistical analysis of the number of endophytic fungi,it was found that the species abundance of endophytic fungi at different developmental stages was significantly different.Five dominant strains were selected for the study.They were Penicillium spp.,Fusarium spp.,Ilyonectria spp.,Aspergillus spp.,and Trichurus spp.respectively.Using the Pearson correlation software to explore the relationship between the ecological succession of endophytic fungi and the accumulation of seven kinds of active constituents during the five developmental phase,Fusarium species and the content of ursolic acid was highly correlated?p<0.05?,Penicilliumhad significantly negative correlation with the polysaccharide?p<0.01?,there is a guided relationship between the endophytic fungi and active compounds,through the interaction between each other,some endophytic fungi may promote or inhibit the accumulation of C.songaricum active ingredients.This research not only analyzed the accumulation of active components but also studied the relationship between the ecological succession of endophytic fungi and the accumulation of active constituents of C.songaricum systematically and integrally,which provideda a data support for the collection of medicinal materials and safety of market medication,also provided a reliable scientific basis for further research on the interaction mechanism between plants and endophytic fungi.