| Anoectochilus roxburghii(A.roxburghii),a kind of Anoectochilus herb belonging to the family of Orchidaceae,is mainly produced in Fujian and Taiwan province.It also has another name as “Gold Thread” and been known as “King of medicine” in the folk with the efficacy of analgesic cough,heat-clearing and detoxicating,dispel wind and wet.It can not only be used to treat cough up blood in tuberculosis,the disease such as diabetic,but also relieve the pain from cystitis,rheumatism and rheumatoid arthritis.With the development and utilization of its medicinal value and health care function,the wild resources of A.roxburghii become increasingly scarce.At present,the industry of A.roxburghii mainly focuses on tissue culture and artificial cultivation.But this kind of current situation also exists a lot of inadequacy,for example,the quality of A.roxburghii in the market is uneven,and there is even a phenomenon that someone passes off fish-eyes as pearls.Meanwhile,whether it is necessary to transplant the seedlings after tissue culture,how long should them be transplanted,and the base selection and management measures for the cultivation of A.roxburghii have not been standardized yet.In this study,we used the tissue culture seedlings which has been called "Fujianxiaoyuanye" as experimental materials.Then,we transplanted the five months old tissue culture plants into the simulated wild environment under the forest to dynamically explore the rules of quality change and residue of the plant growth regulator.After that,we compare the difference of microstructure between tissue cultured clematis and transplanted clematis,and analyze the dynamic changes of soil nutrients and microbial communities in the process of imitation wild cultivation.We hope our research can provide theoretical basis for the establishment of quality standards and standardized cultivation to A.roxburghii industry.The main results are as follows.1.Through the statistical analysis of biomass' s dynamic of transplanted A.roxburghii under forest in different months,we found that plant height,weight and leaf number of A.roxburghii increased with the time passing by after transplanting.Specifically,the plant height and weight reached maximum after transplanted for six months and leaf number reached maximum after transplanted for eight months.Meanwhile,the drying rate of the whole plants showed a trend of "M" type.It has a higher drying rate from six to eight months after transplanting.According to the growth rate of A.roxburghii,it can be concluded that the growth rhythm of A.roxburghii after transplanting is as follows: the first two months after transplanting are slow growth period,three months to seven months are rapid growth period,and the eighth month is slow growth period.Moreover,we found that the numbers of vascular bundles in stems,the area of leaf mesophyll cells and stomata density increased clearly after transplanted by comparing microstructure of stem and leaf between tissue culture seedlings and transplanted seedling.What's more,the calcium oxalate crystal can be observed in stem and leaf after transplanted for several months.2.We dynamically analyzed of the content of active ingredients and plant growth regulator in different months transplanted plants and found that total polysaccharides accumulated as the time goes on.The content reached maximum in the seventh month.Furthermore,we further analyzed the monosaccharide content and components of polysaccharide by using HPLC method and found that polysaccharide of tissue culture seedlings and transplanted seedling are contain mannose,glucose,galactose and arabinose.Particularly,rhamnose appeared in the transplanted plants from sixth months.What's more,the main components of polysaccharide is glucose,and galactose takes the second place.Total flavonoids increased slowly at the first three months after transplanted,and decreased gradually from four to seven months.We also used the HPLC to analyze twelve kinds of free amino acids in A.roxburghii.The result showed that the content of various amino acids in the tissue culture seedlings was higher than those in the transplanted seedling.Moreover,the contents of Zn,Mn,Ca and Mg element were decreased after transplantation,while the contents of Fe and Cr element were fluctuated.In general,there was no significant accumulation of feature elements during transplanting.Besides,the plant growth regulator 2,4-D was detected in the tissue culture seedlings,and its content gradually decreased as the time goes by.Finally,it can't be detected after fourth months.3.The contents of organic matter,available phosphorus and soil pH were gradually decreased with the passage of months,and there was a significant negative correlation between the content of organic matter and planting time.However,the total nitrogen,potassium,phosphorus,available nitrogen and available potassium were positively correlated with the planting time.In addition,both the number of microorganisms and the microbial biodiversity were slightly increased with the transplanting time by analysis the microbial community structure.From the perspective of phylum level,the bacterial community structure of rhizoplane soil of A.roxburghii in the process of transplanting gradually resembles that of the wild type.Furthermore,the main microbial phyla were Proteobacteria and Acidobacteria.But the microbial community structure changed constantly along with the time pass by.Bacteroides and lactobacillus were significantly concentrated in the M2 sample,while Flavobacterium,Pseudomonas and Neosphaingella were highly abundant in the M4 sample.However,the abundance of Burkholderia,Blastomonas and Thermoacidella in M8 samples was significantly higher than that in other samples.The change of soil microbial community in rhizosphere may be closely related to the growth of A.roxburghii.In general,the transplantation of tissue culture seedlings of A.roxburghii is very necessary and the transplantation time should be no less than four months.According to the biomass and effective component content,the optimal harvest time should be seven months after the transplantation.The soil rich in organic matter and microorganism with high vegetation coverage and type of broad-leaved forest and bamboo forest should be selected in the selection of cultivation base of A.roxburghii.In the transplanting process of A.roxburghii,phosphorus should be properly supplemented,soil acidity and alkalinity should be adjusted and organic matter should be appropriately added during long-term planting,or the soil in this area should be restored after harvest for a period of time. |
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