Study On The Pharmacognosy Of The Picris Japonica Thunb And Herba Asari

Picris japonica Thunb belongs to family Compositae. In China, it mainly grows in Hebei, Shanxi, Jilin and Heilongjiang provinces. It is often used to herba and could clear heat, detume scence, cure flu and mammary abscess. At present, pharmacognosy study of P. japonica Thunb is very few, and only the chemical composition is covered in several reports. In this paper, its microscopic identification was studied based on the identification and scanning electron microscopic technique. The main contents are as follows:1. The microstructure of P. japonica Thunb.①Root cross-section has an epidermal cell. The old root lateral layers have several cork layers. The laticifer has scattered in the phloem, and medullary ray is obvious.②Stem cross-section has an epidermal cell. Its inner cortex is obviously. There are glandular hairs and non-glandular hairs on the epidermis of stem.③Leaf cross-section has an epidermal cell, has 1-2 palisade tissue cells and don't through the main vein. There are 1-5 main vein vascular bundles, which are collateral bundles, slightly semi-circular array.④Leaf epidermis-chip:The glandular hairs, non-glandular hairs, and stomas distributed on the upper and lower epidermis. The anticlinal walls of the upper epidermis cells are slightly straight, the anticlinal walls of the lower epidermis cells are wavy bending. Andersen matal for inequality or infinitive, the Deputy Guardian cells 4-6.⑤The aboveground part are coated with non-glandular hairs which like bifurcated hooks, the two top cells bifurcated into special hook, thickened obviously, the basal part constituted by a number of elongated cells; There are two types of glandular hairs.⑥Pollen exine has spinulose ornamentation. There are three apertures.⑦Powders observation, two top bifurcation of the non-glandular hairs and debris of the multicellular glandular hairs are seen. Pollen grains common, oval, outer barbed-like protuberances, there are three apertures; Vessels are mostly spiral vessel and bordered pit vessel. There are broken pappus and laticiferous vessels.2. The surface micromorphology for pollen and fruit of P. japonica Thunb. Pollen exine has spinulose ornamentation, whose base is thick. The Thorn-like protuberances are ring-shaped arrangement or dense clusters; The mesh is small and deep, both size and shape are similar; Three germination holes with three large tumor-like enations which has brain striate ornamentation. Different numbers of tumor-like protuberances on the different observation surfaces. Exocarp fruits were arranged around the tile sheet, "tile" tip with finger-like protrusions. The fruit has about five to ten longitudinal ribs and five deep cannelures that across the fruit both sides. There are more finger-like protrusions in the deep cannelures. Exocarp cells are long and narrow, the tip of the finger-like protrusions are dissociative, finger-like protrusions arrangement were stacked around the fruit. Overview on the other side overlooking the pentacle was the middle hole, the surface of many cells tightly arranged in long strips, point hole, the point of view of the fruit surface, such as layer upon layer of mushrooms.The micro structure and micro-morphological characteristics using scanning electron microscopy observation provide an important fundamental basis for the identification of P. japonica Thunb.Herba Asari is a commonly used traditional Chinese medicine, derived from dried roots and rhizomes of the Aristolochiaceae Asarum. heterotropoides Fr. Schmidt var.mandshuricum (Maxim.) Kitag, A. sieboldii Miq. var. seoulense Nakai and A. sieboldii Miq.. It has many kinds of effects, mainly treats cold, headache, toothache and other diseases. Herba Asari (A. sieboldii Miq.) widely distributes in the Yellow River and the Yangtze River basin, which derived from Shaanxi Huayin were traditionally regarded as the best quality. However, it is lack of the effective basis actually. In this paper, through analysis of the volatile oil content and chemical composition, the regional differences in volatile components of Herba Asari were compared. Many records said that the Herba Asari was poisonous since ancient times. However, until now, the toxicity manifestations, toxic components and the toxicity mechanism of Herba Asari are still a lack of understanding. In this paper, a systematic comparative analysis of the commonly used formulations of water decoction of Herba Asari, powder and compound preparations (Mahuangfuzixixin Tang) on the SD rat kidney, liver, lung, stomach, intestines and other important organs of the toxic effects. The results are as follows:1. Volatile oil content and chemical composition analysis. Measured volatile oil contents in leaves, roots and rhizome of Herba Asari at 25 sampling points such as Zhejiang, Jiangxi, Anhui, Shandong, Hunan, Hubei, Sichuan, Chongqing, Shaanxi Provinces, compared its absolute content and composition of volatile components.①The average of volatile oil content in root and rhizome of Herba Asari is 2.87%, but the average of volatile oil content in leaf is 0.23%. Obviously, the volatile oil content in the leaf is far lower than the volatile oil content in the root and the rhizome of Herba Asari.②Some volatile components of Herba Asari between the different sampling points show significance difference (P<0.05), such asα-pinene, sabinene,β—pinene,α-terpene, eucalyptol,γ-pinene,4-terpine,α-terpineol, 3,5-dimethoxytoluene, safrole, methyleugenol, myristicin, n-pentadecane, elemicin and so on. The other volatile components show non-significance differences (P>0.05), such as camphene, myrcene,α-phellandrene, p-cymine and so on.2. The toxicity of Herba Asari was closely related to its preparation and dose. At the dose of 0.94 g·kg-1·d-1(9g/person) Herba Asari powders have caused obvious injury in rats, including considerably high CREA figures and pathological changes in tissues of kidney, liver and lung; and at the dose of 2.81 g·kg-1·d-1 (27g/person) the rats were dead one after another and their kidney, liver and lung were seriously injured. In contrast, Herba Asari decoction and Mahuang-Fuzi-Xixin Tang made no harm to the rats after 60 days drug treament at both doses of 0.625 g·kg-1·d-1(6g/person) and 3.125 g·kg-1·d-1 (30g/person), in other words, there was no meaningful difference in the biochemical indices TPU, CREA and BUN from those of the blank group, and no pathological alterations were recognized in the tissue sections of kidney. However, in Mahuang-Fuzi-Xixin Tang group occured slight injury in the rats'kidney when the dose of Herba Asari was added to 6.25 g·kg-1·d-1 (60g/person). It's showed that in rats the toxic effects of Herba Asari were manifested in the injury of kidney, liver and lung, but not in stomach and intestines. The toxicity of Herba Asari powders was remarkably stronger than those of Herba Asari decoction and Mahuang-Fuzi-Xixin Tang.3. The establishment of AFLP reaction system for Asarum sieholdii Miq. Adopting improved CTAB method to establish the method for extracting Asarum sieholdii Miq's genome DNA which is suitable for AFLP analysis. It is shown that Asarum sieholdii Miq's genome DNA extracted with 2%CTAB,2mol/LNaCl, 1uLβ-Mercaptoethanol is purity and no degradation basically and is suitable for AFLP analysis. AFLP reaction system of Asarum sieholdii Miq was studied initially and the analysis system which is more suitable for AFLP analysis was obtained. Genomic DNA (500 ng) was digested with 10 U EcoR I and 5 U Mse I restriction enzymes at 37℃for 3 h in a volume of 25μl. In ligation reaction (12.5uL) containing 2.5 pmol EcoR I ligation adapters,25 pmol Mse I ligation adapters and 175 units of T4 DNA ligase, the mixture was incubated overnight at 16℃(12h～16h). Pre-amplification products were diluted 150 folds. The selective PCR reaction mixture (20μl) also included EcoR I selective primer (8 pmol), Mse I selective primer (8 pmol), dNTPs (4 nmol), and Taq DNA polymerase (1 U) in 1×PCR buffer. Prior to electrophoresis, the reaction products (8μl) was mixed with formamide loading buffer (3μl) of 98%(w/v) formamide/10 mM EDTA, pH 8.0/0.1% bromophenol blue/0.1% xylene cyanol (w/v). The loading mixture was denatured at 95℃for 5 min, then cooled on ice immediately, and resolved on 6% denaturing polyacrylamide gels. AFLP gels were silver stained and photographed.