Studies On The Relationship Between The Structures, Development And Major Medicinal Components Accumulation Of Polygala Tenuifolia Willd And Polygala Sibiric L.

"Yuan-Zhi" has been used in traditional Chinese medicine. Chinese Pharmacopoeia took Polygala tenuifolia Willd and Polygala sibirica L.as two original plants of "Yuan-Zhi". P. tenuifolia Will and P. sibirica L. which belong to Polygalaceae are perennial herbaceous plants, and the main medical parts of them are dry roots. The main medicinal ingredients in its dry roots were saponin, xanthone, oligosaccharide polyesters, polysaccharide and fat oil. Anatomical, histochemical and phytochemial methods were used systematically investigated the structural features and the development of vegetative organs in plant of Polygala tenuifolia and Polygala sibirica, and the localization of saponin xanthone, fat oil and polysaccharide in vegetative organs of the two plants, and localization of saponin by cytochemistry, as well as the accumulation and the dynamic variation rule of saponin content. At the same time, the quality of crude drug of Polygala tenuifolia and Polygala sibirica were compared. The contents of saponin and polysaccharide of cultivated crude drug of Polygala tenuifolia. from different places of major producing areas were also compared and the correlation between the content of main chemical constituents and main environmental factors were analysed. The main experimental results are as follows:The development of roots of P. tenuifolia could be divided into four stages: promeristem, primary meristem, primary structure and secondary growth. In primary structure, primary xylem was diarch; there is no pith in the center. The secondary structure included periderm and secondary vascular tissue. There were three characteristics of secondary structure of Polygala tenuifolia. Fristly, the secondary phloem made up about 3/5-2/3 of the root diameter, and the diameter of the roots increases depend on normal secondary growth not exceptional secondary growth. Secondly, the secondary phloem contained mainly parenchyma with abundant storeage, the sieve vessel and companion cells were few. Thirdly, Polygala tenuifolia had developed periderm with thick cork, and secondary xylem contained developed vessels which distributed frequently. The structure features of cork and secondary xylem were adapted to the xeromorphic characteristics of Polygala tenuiflia.The development of stems could also be divided into four stages: promeristem, primary meristem, primary structure and secondary growth. The stem composed of epidermis, cortex and vascular bundle. The ring of sclerenchymatous cells that developed between cortex and phloem might be the apoplastic protective screen which could protect the stem from drought. The cellls of cortex that close to the epidermis specialized into assimilating tissue. Xylem was developed. These characteristics adapted to arid environment.The leaves of Polygala tenuiflia were typically bifacial, and the structure, origin and development of leaves was similar to those of ordinary dicotyledonous plants, There were epidermis hair and stomatal apparatus on the epidermis. The stoma chamber was big. There were crystal cells among the mesophyll. These characteristics of leaf also adapt to arid environment.The structures of vegetative organs of P. sibirica were similar to that of Polygala tenuiflia. However, the secondary phloem of P. sibirica made up about 1/3 of the root diameter, the ratio was lower than that of Polygala tenuiflia. There were difference between the density of epidermis hair and stomatal apparatus of the two plants. There were not crystal cells among the mesophyll of P. sibirica.The flower of Polygala tenuiflia and P. sibirica was bisexual, zygomorphous one. The shape of sepal differentiated into petal, and the petal in center of flower differentiated into keel, and the upper partial of petal specialized into the cristate shape. The structures of pistil and stamen were special. The filaments of eight stamens combined partially, and the stigma of pistil was split into two parts, which had unequal height. These characteristics were adapted to entomophilous pollination. The pistil consisted of two carpels. There was one anatropous ovule in the ovary which was epistasis with two loculars. The capsule was composed of pericarp and a seed. The type of seed belonged to a dicotyledonous albuminous one.The histochemical localization results of Polygala tenuiflia by 5% vanillin-glacial acetic acid-perchloric acid solution showed that saponins were distributed in epidermis, cortex and the parenchyma of central cylinder in primary structure, while in secondary structure saponins mainly in secondary phloem, phelloderm, rarely in xylem ray and xylem parenchyma, and in stem mainly in phloem, rarely in epidermis and cortex, and mainly in mesophyll of leaf, rarely in epidermis. Histochemical localization results of P. sibirica were similar to the result of Polygala tenuiflia. The Mg-Hcl localization results of xanthones indicated that phelloderm parenchyma cells and secondary phloem were storage locus. The localization by SudanⅢand osmic acid indicated that there were abundant fat oil in secondary phloem, phelloderm . However, in root of P. sibirica L., fat oil could not be found. PAS reaction results showed polysaccharide were also existed in root periderm and secondary phloem of the two plants. It displayed that the secondary phloem was the main storage site of main medicinal components. The ultrastructure research result indicated that the parenchyma cell which accumulate the saponin have plentiful organelle. The cytochemistry method of PbAc—settling showed that the settleable solids of PbAc were found in vacuoles or around the tonoplast in the parenchyma cells which accumulate the saponin, such as parenchyma cells of phloem, cortex cells and sieve tube etc. It proved the location of saponin of histochemical results.The phytochemical results also showed that the saponin accumulated in the vegetative organs of P. tenuifolia and P. sibirica, with higher content in roots and lower content in the aerial parts that included stems and leaves. This result was consistent with the findings obtained through the histochemistry experiment. Based on these results, we advocate the utilization of aerial parts of P. tenuifolia and P. sibirica facing the shortage of Yuan-Zhi" resource today. According to the dynamic variance of saponins content of P. sibirica at the developmental stage, the P. sibirica should be gathered at period from April to May. The content of total saponin and segenenin in different parts of root showed obvious variation. The content in the "skin areas" was much higher than that of xylem. The result is consistent with the histochemical localization. Accordingly, the crude drug that had thick "skin areas" and thin xylems was of top grade.The HPLC result showed that during the growth period the senegenin percent content of roots in four different years had dynamic variant trend, the senegenin percent content had the similar result, that is annual roots'> biennial roots'> triennial roots'>quadrennial roots', it showed that the younger root had higher content. We guessed that this result came because triterpenoid in P. tenuifolia may be chemical defenses. As far as the increasing rate of roots of P. tenuifolia was concerned, the length, diameter, thickness of "skin areas" as well as dry weight content of roots increased most quickly from the second to the third growth year. The increasing rate of roots decreased at fourth growing year. In the post-fruit period, the total senegenin content of the triennial roots was 2.66 times as much as that of the biennial roots, and 3.97 times as that of the annuals. However, the total senegenin content of the quadrennial root was only 1.20 times as much as that of the triennial roots. In fact, it is the total amount of secondary metabolic product in plant that should be considered while the medicinal material is selected. So the middle ten days of August of the third growing year was optimal collecting period for the drug of Polygala tenuifolia.The comparision of roots of two plant indicated that the diameter, thickness of "skin area", dry weight as well as senegenin contents of roots in P. tenuifolia were all higher than that of P. sibirica. So, the crude drug quality of P.tenuifolia was better than that of P. sibirica. We should choose Polygala tenuifolia as cultivated material.The shape of cultivated plants of P. tenuifolia was more hairchested and had more branches and thicker root, comparising with the wild one. The wild plants of P. tenuifolia having less branches and the root were nonuniform, the lateral root were thiner. Judging from the senegenin contents of "skin areas" and root, the quality of wild plant was higher than that of the cultivated plant as a whole. However, analyzing from the dry weight and total senegenin contents, the roots of triennial cultivated P. tenuifolia had the highest level, which was also higher than that of the wild plant. The result indicated that the cultivated P. tenuifolia could instead of the wild, and the cultivated P. tenuifolia shoud be gathered at third growth year because of the highter total senegenin contents.Results on Polygala tenuifolia from different places of major producing areas in Shanxi, Shaanxi, Hebei province showed that there were obvious differences (p<0.05) in the contents of saponins and polysaccharide. This indicated that the quality of major producing areas were very different. Analyzing from contents of saponins and polysaccharide, the quality of crude drug of "Xin-Jiang" and "Wen-Xi" were higher. The variety of "Yuan-Zhi" from "Xin-Jiang" and "Wen-Xi" could be preferred for breeding. The analysis of major environmental factor showed that the climate of different places of major producing areas were similar, having dryclimate and higher accumulated temperature. The soil were not fertile and alkalescent, with lower content of N and highter content of K. Correlation indicated that the content of saponin had negative correlation with the organic matter of soil, but had positive correlation with the element of Mn. The content of saponin had positive correlation with content of polysaccharide.