Studies On The Correlation Between The Structural Development Of Achyranthes Bidentata BL. And The Accumulation Of Major Medicinal Components Together With Its Forming Of Genuineness

The plant Achyranthus bidentata Blume belongs to Amaranthaceae family, its dried roots, has been used in traditional Chinese medicine. In China this specie is mainly distributed in the area of Guhuaiqingfu such as Wuzhi, Wenxian, Qinyang, Mengxian, huixian and Boai in Henan Province. Therefore, name of locally produced drug has been prefixed with the word "Huai", which has been passed down through history. The mainly medicinal ingredients in its dried roots were saponin, ecdysterone, and polysaccharide. This dissertation systematically investigated the development and structural features and the histochemistry orientation of triterpenoid saponin of various vegetative organs, the accumulation of triterpenoid saponin in different sowing-season A. bidentata and the accumulation of ecdysterone in conventional sowing-season A. bidentata, determined the contents of mainly medicinal ingredient, inorganic elements in cultivated soil and roots in A. bidentata from different producing areas by using anatomical, phytochemistry and histochemistry methord. The correlation between the content of chemical constituents in roots from different areas and environmental factors were analysted by SPSS13.0. Finally the genuineness of A. bidentata were approached. The main experimental results are as follows.There were tertiary structures which were originated from a supernumerary cambium in the axes of A bidentata. The development of A. bidentata root and stem was divided into five developmental stages, which were promeristem, primary meristem, primary structure, secondary structure and tertiary growth. The primary and initial secondary growth of the root and stem was similar to those of ordinary dicotyledonous plants. Generally, the primary xylem was diarch. The secondary structure consisted of periderm and secondary vascular tissue from the outer to the inner. The cork cambium formed from pericycle cells that resumed their dividing capabilities. The enlargement of its roots, however, was primarily resulted from the differentiation and development of the tertiary structure formed by the extra cambium. The first ring of extra cambium originates from the parenchymatous and vascular ray cells outside the secondary phloem and each following rings initiated in the outmost foundamental parenchyma cells which were derived from its previous ring of extra cambium. In the supernumerary cambiums, there were not distinction between the fusiform initial and the ray initial. Its cells present storied arrange from a longitudinal section through root. The tertiary vascular bundles were neatly arranged in a concentric ring, separated by parenchyma cells. The numbers of ring were the same as that of extra cambium and continually increased to 4-5 cycles with the development of the root in the annual root of A bidentata. However, there were two types of anomalous structures in the stem of A bidentata. In the development of stem, the fascicular cambium was initiated, the interfascicular cambium was not formed in the secondary growth of stem. A supernumerary cambium ring originated from the one or two layers cells of procabium which remained the outside of secondary vascular bundle when the differentiation of secondary vascular bundle was finished. And the activities of the supernumerary cambium produced a round tertiary vascular bundle. In addition, there was no correlation between the two medullary bundles that also originated from procambium and normal vascular bundles in the position. But there were multiformity on the types of the structure of medullary bundle. And sometimes incomplete amphivasal bundles were formed.The leaves of A. bidentata were typically bifacial, and the structure, origin and development of which was similar to those of ordinary dicotyledonous plants, consisting of epidermis, mesophyll and veins. There were two types non-glandular hairs whose cell numbers were obviously different in both the upper and lower epidermal layers, while stomata were distributed only in the lower epidermis. There were three to four layers of palisade tissue cells in mesophyll, with several chloroplasts. There were a few spongy tissue cells with an irregular cell shape, containing a few chloroplasts, and intercellular space. The major vein consisted of several collateral bundles. There were 7-8 collateral bundles ranged in parenchyma tissue but no medullary bundle in the semicircular petiole. The flower of A bidentata was disexual and cross pollination, including five stamens and one pestil. The anther contained four pollen sacs which were divided into two sections. There were granular protuberances on the surface of elliptical pollen in anther. The length of pistil was more than stamen. The pistil consisted of one carpel. There was one ovule in the ovary which was epistasis with one locular. The single stigma with a lot of digitations on its surface was wet type. The intraepidermal cells of the hollow style formed entad processes constituting particular passage cells. The utricle was composed of pericarp and a seed. The seed type belonged to a dicotyledonous albuminous seed.Histochemistry, scanning electron microscope and energy-dispersive X-ray microanalysis analysis of A. bidentata vegetative structures indicated that, in the root primary structures, saponins were mainly distributed in the pericycle, primary phloem and parenchyma cells between primary phloem and primary xylem. In the secondary root structures, saponins were mainly distributed in secondary phloem and parenchyma cells of the phelloderm, and as well as in supernumerary cambium and phloem of tertiary vascular bundle after the tertiary structure maturated gradually in the roots. Saponins were mainly found in the phloem cells of normal vascular bundles and medullary vascular bundles of stems, and in palisade tissue and phloem cells of the main vein vascular bundles in leaves. The tertiary structure was not only main parts in the roots of A. bidentata, but also important storage region of triterpenoid saponin in its growth and development. The stem may be the essential transport organ of triterpenoid saponin, while palisade tissue cells may be the primary synthesis location.Phytochemistry results showed that there were dynamic change trend of oleanolic acid contents in different sowing-season A. bidentata roots, which revealed a "high-low-high" pattern. Reproductive growth was not beneficial to the accumulation of saponins in roots. There was no significant difference between and summer-sowing roots in yield except for the long growth periods of spring-sowing A. bidentata. However, the contents of oleanolic acid in sprin-sowing A. bidentata roots are lower than smmer-sowing. Therefore, early July would be the optimal seeding time of A. bidentata. That not only enhances the efficiency of land usage, but also improves the yield and quality of A. bidentata.Study on dynamic change of ecdysterone during the growth and development of A. bidentata indicated that the optimal period of ecdysterone accumulation was vegetative growth phase. Whereas reproductive growth was not beneficial to accumulation of ecdysterone in roots. In November, the relative levels of ecdysterone in various organ of A. bidentata were fruits>leaves >roots>stems. So the leaves may be the synthesis location of ecdysterone, the fruits may be the main storage organ, while roots may be the secondary storage organ, the stem may be the essential transport organ. The contents of ecdysterone in leaves remained at a higher level during the whole growth of A. bidentata, which might be related to the biological behavior and activity of pollination insects. It is chemical ecology significance that there were ecdysterone in A. bidentata.Comparative research on A. bidentata from different producing areas showed thatThere were different on the outward appearances. Roots of A. bidentata from geo-authentic producing areas were longer and more bulky than that of non-authentic while no obvious differences on internal structures. The contents of oleanolic acid, ecdysterone, polysaccharide and Cr were higher, but the Zn were lower in the geo-authentic A. bidentata roots. The geo-authentic crude drugs had a strong tendency to accumulate Fe, Cu, Cr and P. The contents of Zn, B and Al were higher, while the Cu were lower in the cultivated soil of geo-authentic areas.There were obvious relationship between the concentrations of inorganic elements in A. bidentata and those in their corresponding soil. There were good relationship between the absorption and accumulation of Na and Al, Ti; Ca and Mg, B, Cd; Mg and B; Al and Cd; Cd and Ti, while were the antagonism relationship between Pn and Cu, K; Cr and B, Zn. There were positive relationship obviously between oleanolic acid and polysaccharide and Cr, whereas Ca, Mg, B and Zn was not beneficial to accumulation of that. The annual average evaporation of geo-authentic were lower than non-authentic areas, which was beneficial to accumulation of ecdysterone and polysaccharide in A. bidentata roots. But the concentrations of available phosphorus and potassium were lower in cultivated soil of the geo-authentic, which was beneficial to accumulation of oleanolic acid in in A. bidentata roots.