| Prunella vulgaris L. (Labiatae), also known as the "self-heal," was a perennial herb commonly found in North Asia, Europe and North Africa. The dried spica of P. vulgaris, Prunellae Spica, a standard medicinal material in the Chinese Pharmacopoeia and occasionally as a folk medicine to treat sore throat, fever, goiter, hypertension, tuberculous lymphadenitis and mammary gland hyperplasia. Several studies on the pharmacological activities of P. vulgaris have been previously reported, including hepatoprotection, antihyperglucemia, antifugal, anti-tumor and anti-inflammatory activities. As the key raw material, there were dozens of Chinese proprietary medicine manufactured with P. vulgaris. In recent years, the demand for P. vulgaris has steadily increased in the world market. The wild population of P. vulgaris cannot meet this growing need. Long-term since, P. vulgaris was often cultured by traditional planting technology. The many problems associated with actual production process have emerged. All this shows that the cultivation techniques of P. vulgaris could not to promote the development of traditional Chinese medicine. Therefore, the good agricultural practice (GAP) and quality control of P. vulgaris were investigated, and improved the yield and quality of P. vulgaris. Specific results were as follows:1. Textual research on medicinal works of past dynasties and field work were adopted. The results showed, the natural distributions of P. vulgaris were concentrated distribution in Sichuan Province, Huaihe River Basin, and Middle-Lower Yangtze River Valley in Chinese history; the traditional efficacy of P. vulgaris between ancient and modern was basically identical; The whole grass of P. vulgaris as medicinal parts (from late Ming dynasty to late Qing dynasty and early stage Republic of China), both whole grass and spicas as medicinal parts (from a mid-term Republic of China to 1963), and the semi-maturity or maturity of spicas as medicinal parts (from 1963 to today). The processing method for medicinal parts of P. vulgaris adopted sun drying and shady drying in ancient China, but the sun drying was only used in modern China. The original plants were originated from two species (P. vulgaris L. and Prunella asiatica Nakai.); Prunella hispida Benth.was used as a certified medicinal materials in Yunnan province from Ming Dynasty to modern time. The dietotherapy history of P. vulgaris in China could be traced back to Ming Dynasty or before.2. The various germplasm resources of P. vulgaris came from all over the country were investigated by analysis of correlation analysis, path analysis and principle component analysis in randomized block experiment. The results showed that the 6 morphological characteristics were greatly different from each other in various germplasm. The spicas yield per plant had a very significantly positive correlation with the number of spicas per plant and fresh leaves weight per plant, meanwhile the correlation between the spicas yield per plant and spicas length was very significantly. Three principal components which accounted for 87.533%of total variance were extracted from the principal component analysis. The strong growth ability, the number of spicas per plant and spicas length were focused on the high yield breeding and planting of P. vulgaris germplasm.3. Through the dynamic sampling of P. vulgaris in different growing stage, and the dry weight of plant, the amount of dry matter and the content of N, P and K within each organ of the plant were measured. The results showed that the dry weight of P. vulgaris linearly increased with increase of nitrogen, phosphorus and potassium accumulation with significant correlation coefficients at whole growth period. The balance of nitrogen, phosphorus and potassium accumulation was beneficial for dry matter accumulation, which linearly increased with increase of nutrient balance index. The maximum stage accumulation rate of dry matter from mid-April to beginning of May, reached 63.71%. The amount of dry matter was mainly distribution in leaves from late March to mid-April. The distribution ratio in stem was highest from beginning of May to late May. Until mid-June, the distribution ratio of dry matter in spicas was higher than other organs. The absorption amounts of N, P and K were lower before mid-April, and the nutrient element was concentrated in plant leaves. From late April to beginning of May, the N, P and K accumulation reached maximum of total absorptive capacity, which accounted for 72.35%、59.82% and 87.41%, and the mineral elements were concentrated distribution of plant stems. When transferred to reproductive stage, both accumulation amount and distribution rate of nutrient elements in spicas was growth rapidly, which reached maximum until mid-June. From elongation stages to quaring period was the period of maximal accumulation of dry matter and showed the maximal efficiency of N, P and K nutrition. From elongation stages to quaring period is the most crucial period in the growth development stage of P. vulgaris, and should be pay more attention to the supply of mineral nutrition and moisture of plant to promote spicas formation and dry matter accumulation in this stage. The average N, P205and K2O absorption amount per 100 kg dry matter of spicas were 1.62 kg,0.36 kg,2.88 kg respectively, and the ratio of N:P2O5 : K2O was 1: 0.22: 1.77.4. The experiment was a randomized complete block design arranged in split plot. The growth periods of P. vulgaris were investigated. The morphological characteristics, biomass of each organ, spicas yield and content of active components at harvest time were determined. The results showed, with the delay of sowing date, the growing period of P. vulgaris was shortened; the morphological characteristics, biomass of each organ and spicas yield were decreased significantly in P. vulgaris. With postpone of sowing date, the content of active components in P. vulgaris were significantly increased. Considering the two factors between spicas yield and quality in P. vulgaris, the suitable sowing period for P. vulgaris was from 5th September to 5th October in Lujiang County, Anhui Province.5. The six planting densities were designed in the plot experiment. Morphological index per plant, dry matter accumulation in each organ, spicas yield and content of active components in P. vulgaris were determined and analyzed. The results showed that plant height of P. vulgaris significantly improved with increasing density, but crown width, leaf area per plant, number of spicas per plant, spica length of P. vulgaris significantly declined with increasing density. Dry weight of P. vulgaris and biomass of each organ significantly decreased with density increasing. Distribution ratio of dry matter in root has less effect by planting density, distribution rate of dry matter in leaf and stem decrease with density increasing, the distribution ratio of dry matter in spicas increase with increasing density. The treatment D has a highest spicas yield. The content of water-soluble extracts and total flavonoids in P. vulgaris reached the highest in treatment C, the maximum content of polysaccharides in P. vulgaris was treatment D, the highest amount of ursolic acid, oleanolic acid and rosmarinic acid in P. vulgaris were found in treatment A. Considering the two factors between spicas yield and quality in P. vulgaris, the planting density was set as 25 cm X 25 cm in Lujiang county, Anhui Province.6. Field plot experiment with a 4-variable quadratic orthogonal rotation combination design was adopted, and a function model of spicas yield in P. vulgaris was established. Under different fertilizer treatments, the content of water-soluble extracts, total flavonoids, polysaccharides, ursolic acid, oleanolic acid and rosmarinic acid were determined. The results showed, the model analysis showed that spicas yields of P. vulgaris were significantly influenced under the N, P2O5 K2O and OF applications, among there factors, N has the greatest effects on spicas yield of P. vulgaris, secondly for P2O5, thirdly for K2O, and OF has the smallest effects on spicas yield. At the lower fertility levels, the spicas yields of P. vulgaris were improved along with the increasing of N, P2O5 K2O and OF applications, but the spicas yields decreased when N, P2O5 K2O and OF were applied too much. The main active components level in P. vulgaris spicas were significantly increased by optimized application of N, P2O5, K2O and OF. In this experiment condition, the spicas yields of P. vulgaris could reach to 722 kg·hm-2, and quality of spicas could effectively improve when the fertilizer rates were N 303.9～335.1 kg·hm-2, P2O5 432.5～500.6 kg·hm-2, K2O 206.6～240.2 kg·hm-2, OF 2312.5～2687.5 kg·hm-2.7. The factors studied included two water conditions and three fertilizer regimes. The results showed that moderate drought stress dramatically decreases spica biomass production but increases the content of RA, UA and OA in spicas of P. vulgaris. Total RA, UA and OA yields were found to be significantly higher in well-watered plants than in drought-stressed plants. An appropriate amount of fertilizer could alleviate the negative effect of drought stress on the growth of P. vulgaris and its production of RA, UA, and OA. The interaction of water and fertilizer treatments significantly influences vegetative dry weight, reproductive dry weight and total RA, UA and OA yields in P. vulgaris. The results suggest that the application of the proper amount of fertilizer aids P. vulgaris production in arid and semi-arid regions and that the combined use of fertilizer and properly timed exposure to drought stress can enhance total RA, UA and OA yields in P. vulgaris.8. The active compounds of P. vulgaris were measured, and the main chemical components in P. vulgaris were used for comprehensive evaluation. The results showed that the accumulation of the active constituents, including water-soluble extracts, ethanol-soluble extracts, total flavonoids, polysaccharides, ursolic acid, oleanolic acid and rosmarinic acid levels in P. vulgaris were found to highest in the squaring stage (on 5 May); The contents of water-soluble extracts and ethanol-soluble extracts in P. vulgaris whole herbs were remarkable higher than in spicas. The levels of ursolic acid and oleanolic acid in P. vulgaris spicas were significantly higher than in whole herbs. From squaring stage(on 5 May) to the flowering stage (on 20 May), the contents of total flavonoids, polysaccharides and rosmarinic acid in P. vulgaris spicas were significantly higher than in whole herbs, but from maturity stage (on 15 June) to the wilting stage (on 25 June), our results showed the reverse pattern:the total flavonoids, polysaccharides and rosmarinic acid levels of P. vulgaris whole herbs were remarkable higher than of spicas. Both sun and shade drying were found to exhibit no remarkable influence on active constituents of P. vulgaris, including water-soluble extracts, ethanol-soluble extracts, total flavonoids, polysaccharides, ursolic acid and oleanolic acid. The content of rosmarinic acid in P. vulgaris was significantly reduced by the sun drying. The highest sums of comprehensive evaluation were spicas of P. vulgaris in the squaring stage under open-air sun drying condition. The yield and quality of P. vulgaris were considered, harvest period should selecte in the squaring stage (beginning of May), whole herbs of P. vulgaris should be harvested, and sun drying method need to be adopted.9. The spicas of P. vulgaris.in different storage period (including current year、first-year and second-year) were collected from Lujiang base in Anhui province. And the medicinal material could be dealed with srandom sampling, with the methods of morphologic characteristics and commercial size on P. vulgaris, the samples were graded into three grades; according to the methods from the Pharmacopoeia of the People’s Republic of China (2005), the moisture content, water-soluble extracts, ethanol-soluble extracts, total ash and acid insoluble ash were measured, respectively, utilizing the UV spectrophotometry to detection the contents of total flavonoids and polysaccharides, the levels of ursolic acid, oleanolic acid and rosmarinic acid were adopted by HPLC. The results showed that moisture content of P. vulgaris in different storage period was showed on significant differences; the total ash and acid insoluble ash of P. vulgaris showed significantly increase during prolonged storage; however, the content of water-soluble extracts, ethanol-soluble, total flavonoids, polysaccharides, ursolic acid, oleanolic acid, rosmarinic acid showed significantly decrease during prolonged storage. The spicas of P. vulgaris exhibited no rules differences between the quality and grading standards, The moisture content of spicas in different degree showed significant differences; the second-degree belongs to high content of total flavonoids; the high content of polysaccharides was first-degree; the total ash and acid insoluble ash of spicas exhibited gradually decreased from first-degree to third-grade; and the content of water-soluble extracts, ethanol-soluble extracts, ursolic acid, oleanolic acid, rosmarinic acid showed gradually increased from first-degree to third-grade. The current year was a suitable storage time of spicas from P. vulgaris could be selected; and the traditional grading standards failed to reflect the quality of herbal medicine, therefore, we suggested that the traditional standards should be abolished. 10. The GC method was applied to determine residuals of organochlorine pesticides, the contents of Cu、Pb、Cr、As、Cd and Hg were measured by ICP. The results showed, the contents of residuals of organochlorine pesticides and heavy metals were significant differences in two planting bases, but all of the residuals between organochlorine pesticides and heavy metals were below the GB standards. Residuals of organochlorine pesticides and heavy metals in soil and P. vulgaris from two planting bases were conformed to GAP.11. The essential oils from the fresh flowers, dried flowers leaves and stems of P. vulgaris were isolated by hydrodistillation. The collected oils were analyzed by capillary GC and GC-MS. The results showed that a total of 24,28,19 and 21 compounds were identified, accounting for 95.22%,83.47%,80.03%and 79.48%of fresh flowers, dried flowers, leaves and stems oil, respectively. The main constituent of the essential oil of fresh flowers, dried flowers and stem oils, respectively, was hexadecanoic acid (58.12%,69.99% and 38.44%). Aromadendrene (55.35%) was identified as the highest main constituent for leaves. The composition of essential oil from fresh flowers, dried flowers, leaves and stems of P. vulgaris were discrepant. |
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