The Investigation And Evaluation Of The Ancestral Resources And The Correlation Between Chemical Composition And Environmental Factors

This object firstly investigates and analyzes the current situation of the resources of CORTEX DAPHNES in the main distribution area of China.Based on this,the germplasm identification and the determination of the main chemical components of the investigated materials are performed to improve its medicinal quality evaluation system.And the correlations between major chemical constituents,ecological factors,and soil factors were be analyzed,and the place of suitability of different original plants was evaluated.The research results have important theoretical significance for the rational use and protection of the wild medicinal resources of CORTEX DAPHNES,the improvement of the quality evaluation system,the planning of the best cultivated areas of CORTEX DAPHNES and the standardization of its modern cultivation.Based on the literature survey,this study firstly adopted the survey method combined with the sampling method of the existing site and the method of the line sample method and the method of combining the area of the vegetation and soil type maps to the four provinces of Shaanxi,Gansu,Sichuan,and Qinghai.The wilderness resources of CORTEX DAPHNES of 23 counties were investigated and their ecological communities were analyzed.On the basis of resource investigation,the method of microscopic identification of medicinal powders was used to identify different species of CORTEX DAPHNES,and then the DNA barcodes technology based on ITS sequences,HPLC fingerprinting technology and content determination technology were applied for germplasm identification and quality evaluation.At the same time,through the analysis of specific sites of ITS sequences,NJ cluster tree analysis,similarity analysis of HPLC fingerprints,cluster analysis,principal component analysis,and the difference analysis of the contents of major components to more fully and accurately identify and assess the CORTEX DAPHNES collected from different distribution areas and different species and resolve the similarities and differences of the materials in genetic and chemical aspects.Finally,this project studied the correlation of the main chemical constituents and the ecological factors and soil factors.The correlation analysis and stepwise regression analysis were used to screen the main environmental factors that affected the accumulation of chemical constituents.The topic has achieved the following research results:(1)The total reserves of wild CORTEX DAPHNES in 23 counties in four provinces of Shanxi,Gansu,Qinghai,and Sichuan are approximately 583.62 t,of which the reserve of Daphne giraldii Nitsche is approximately 97.64 t,accounting for 16.7%;the reserve of Daphne retusa Hemsl about 93.32 t,accounting for 16%;Daphne tangutica Maxim reserves of about 392.66 t,accounting for 67.3%.Based on the vegetation and soil type characteristics of the survey sites,it was concluded that areas with the same vegetation and soil type in the non-surveyed sites may also have the distribution of CORTEX DAPHNES.The product of the average amount of CORTEX DAPHNES in unit area of each province and the total area of growth with suitable CORTEX DAPHNES was used to estimate the reserves in the entire province.According to this estimation method,the total reserves of wild CORTEX DAPHNES from the four provinces are approximately 8429.33 t,the reserves of Daphne giraldii Nitsche are approximately 1407.68 t,the reserves of Daphne tangutica Maxim is approximately 5672.87 t,and the reserves of Daphne retusa Hemsl is approximately 1348.68 t.we only found the cultivated resource in a mountain at an altitude of about 2 800 m in Kang county of Gansu province,which cropping scope was about 33 000 m2 and the source comes from wild Daphne tangutica Maxim seedlings.After transplanting,there are no other fine cultivation measures such as watering,fertilizing,etc.The cultivation is about 40 cm in height in two years,and the annual growth height is less than 10 cm,and the cultivated resource can't provide medicinal products at present(2)The microscopic identification of medicinal powders could not effectively identify different species of CORTEX DAPHNES.The similarity analysis of DNA bar codes based on ITS sequences showed that the consistency of Daphne giraldii Nitsche and Daphne tanguticaMaxim was about 93%;the consistency of Daphne giraldii Nitsche and Daphne retusa Hemsl was about 92%;the agreement between Daphne tangutica Maxim and Daphne retusa Hemsl is 99%.Although the sequence identity between Daphne tangutica Maxim and Daphne retusa Hemsl was relatively high,there were differences in sequences of 59bp and 155bp.The Daphne tangutica Maxim genotype was C and the Daphne retusa Hemsl genotype was A at 155bp;The genotype of Daphne tangutica Maxim is G and the gene of Daphne retusa Hemsl is A at 155bp.Therefore,through these two specific sites,it is also possible to discriminate between Daphne tangutica Maxim and Daphne retusa Hemsl.The NJ-cluster tree analysis,HPLC cluster analysis and principal component analysis have high consistency,and can identify and group the CORTEX DAPHNES of different origins and different species.The results of HPLC assay showed that there was a certain difference between daphnetin,syringoside,and 7-hydroxycoumarin in different species of CORTEX DAPHNES,and there were also certain differences in the three chemical components of CORTEX DAPHNES in different distribution areas.However,in terms of the content of one component of the daphnetin,the CORTEX DAPHNES of Ledu,Qinghai,and Tianzhu,Gansu Province have the best quality.(3)The correlation analysis between the main components and the ecological factors of CORTEX DAPHNES showed that the chemical composition of CORTEX DAPHNES was related to most of the ecological factors,and the longitude and average annual relative humidity of its habitat had the greatest impact on its chemical composition.The equations show that the dominant ecological factors affecting the chemical composition of the founder are longitude,annual average rainfall and annual average sunshine intensity.The correlation analysis and stepwise regression analysis between the main components and the soil factors of CORTEX DAPHNES showed that there was no significant correlation between the chemical composition and the soil factor,and there was no significant correlation between soil organic matter,NPK and other nutrients and their chemical composition.However,soil types have a certain correlation with their chemical composition.The reserves of wild CORTEX DAPHNES are continuously decreasing,and government departments should formulate corresponding protection measures.At the same time,we should strengthen the research on the artificial cultivation of CORTEX DAPHNES to achieve the sustainable use of the materials.The DNA barcodes technique,HPLC fingerprinting technology,and determination of the main components in the herbal can be used together to successfully identify different species of CORTEX DAPHNES and to comprehensively and accurately evaluate quality of the herbal from different species and different origins.From the perspective of improving the content of daphnetin and other chemical constituents,the ideal areas for artificial cultivation of CORTEX DAPHNES are Qinghai,Gansu,and the western regions of Sichuan in China.These regions have low longitude,high altitude,relatively dry climate,and strong sunshine intensity.There is a large temperature difference between day and night.The soil types are mostly chernozem and chestnut-calcium,and a small amount of forest soil.The regions that meet these conditions include Tianzhu of Gansu Province,Ledu,Datong and Hualong in Qinghai Province,and Kangding in Sichuan Province,among which Tianzhu County of Gansu Province is the best choice.Of course,the precursor of increasing the chemical composition is to ensure the normal growth of the plant,so in the artificial cultivation medicinal materials should be finely managed to meet the natural factors such as the natural water for the original plant growth,and the balance point that the biomass and chemical composition both can be improved should be found.The CORTEX DAPHNES resources can better serve the pharmaceutical industry.