Determination of multi-index components and quality evaluation of Dilong formula granules / 药学实践杂志

Objective To establish the method for the simultaneous determination of hypoxanthine, inosine, guanosine and adenosine in Dilong formula granules by HPLC and compare the fingerprints of Dilong formula granules from different manufacturers by HPLC chromatogram. Methods The contents of hypoxanthine, inosine, guanosine and adenosine were determined by Thermo AcclaimTM120C18 column (4.6 mm×250 mm 5 μm). The mobile phase was acetonitrile-water. Gradient elution with flow rate of 0.6 ml/min was used. Column temperature was 25 ℃. Detection wavelength was 254 nm. 10 batches of samples were tested. The HPLC chromatogram were compared and analyzed by using the similarity Evaluation system of chromatographic fingerprint of traditional Chinese Medicine (version 2012.130723). Results The linear ranges for the detection of hypoxanthine, inosine, guanosine and adenosine showed good linear relationships within their own ranges (r≥0.999 9). The average recoveries were 99.20%~102.98% with RSD of 0.26 %~0.71%. The contents of 4 components in 10 batches of samples were 0.740 0~4.457 4 mg/g, 2.132 3~7.805 0 mg/g, 0.325 4~1.596 1 mg/g, 0.537 2~2.222 9 mg/g respectively. The similarity between HPLC chromatogram and control fingerprints of Dilong formula granules from different manufacturers was greater than 0.91. Conclusion The method could be used to determine the contents of hypoxanthine, inosine, guanosine and adenosine in Dilong formula granule. HPLC fingerprints could be used to evaluation the quality in Dilong formula granule. The similarity of HPLC fingerprints from different manufacturer production of Dilong formula granule is high, but 4 contents in composition are difference.

Study on the good agricultural practice for Shanghai Pheretima / 药学实践杂志

Objective To establish the Good Agricultural Practice（GAP）for standard operation procedure of Shanghai Pheretima. Methods According to the comprehensive technical requirements of GAP standardized production of Chinese herbal medicine, the key technologies of standardized cultivation and processing were studied in Sangxin Town, Chongming District, Shanghai. Results The production area environment, variety characteristics, breeding technology, pest control, harvesting and processing, packaging, storage and transportation, quality monitoring and other technical links of Shanghai Pheretima were clearly defined and standardized. Conclusion This study provides practical basis for the standardization and modernization quality management of Shanghai Pheretima.

Research progress on Pheretima / 药学实践杂志

Pheretima is one kind of Traditional Chinese Medicine (TCM) with multiple pharmacological effects .Resear-ches on chemical constituents ,pharmacological effects ,identification ,artificial breeding ,machining and quality evaluation of Pheretima were reviewed in this paper .Study gaps and key points were pointed out to provide the basis on follow-up work .

From Earthworm Research to See How to Make Progress in Dilemma of Traditional Chinese Medicine Resources and Quality / 世界科学技术-中医药现代化

Resource of traditional Chinese medicine (TCM) is one of the greatest treasures of China. It made magnif-icent contribution in Chinese lives and breeds. Quality is the foundation of safety, effective and quality control of TCM. With the social development, problem with raising need and falling resource and quality had became more and more serious. It slowed down the speed of TCM modernization and internationalization. This study used Shanghai earthworm, the authentic TCM of Shanghai, as an example. Focus on resource and quality problems, such as species confusion, resource shortage, rough machining, low quality and no standards, this study tried to work out the reason-able and feasible solution. This effort funded a way out of a stalemate of TCM resources and quality. It is important to TCM modernization and internationalization.

Tissue culture and rapid regeneration of Hypericum perforatum / 第二军医大学学报

Objective: To study the proper medium for tissue culture of the explants from Hypericum perforatum . Methods: The root, stem with node and leaf of the Hypericum perforatum seedling were used as explants, and cultured in basic Murashige and Skoog (MS) medium supplemented with different kinds and different concentrations of hormones. The callus, buds and roots were induced. The cultivation of the tube seedling were carried out. Results: Different hormones and explants had different effects on the induction of callus and the formation of buds. 2,4 D could promote the formation of the callus of Hypericum perforatum . Among cytokinins, 6 BA had great effect on the formation of buds, but the effects of KT and ZT were insignificant. The combination of NAA 1 mg/L and 6 BA 0.2 mg/L was found most effective in promoting the rooting of Hypericum perforatum . Conclusion: The results indicate that the callus is easy to be induced when MS medium supplemented with 2,4 D. The root of explants is most suitable for the rosette bud rapid induction.

Establishing better lines and regenerating system of hairy roots of Isatis indigotica with Ri T-DNA / 第二军医大学学报

Objective: To establish the culture system of hairy root of Isatis indigotica and to induce the regeneration plant of hairy root. Methods: Hairy root of Isatis indigotica was obtained from infected cotyledon explants after infection with Agrobacterium rhizogenes strains A4, R1601 and ATCC15834. The better lines were selected. The growth curve was surveyed and extrinsic factors affecting the growth of hairy roots were investigated. The regeneration plant was induced on MS media with different hormones. The transformation of Ri T DNA was examined through high voltage paper electrophoresis. Results: The hairy root was originally obtained from Isatis indigotica . The regeneration plant was induced on MS media with BA. The result of high voltage paper electrophoresis confirmed the transformation of T DNA from Ri plasmid to the hairy root and regeneration plant. Conclusion: The acquisition of hairy root of Isatis indigotica and regeneration plant of it lay a foundation for the production of active component and introduction of foreign gene. [

Enhanced resistance of transgenic tetraploid Isatis indigotica by Bt and CpTI to moths / 中草药

Objective Two insect resistance genes Bacillus thuringiensis(Bt) crystal protein gene CryIA(c) and cowpea trypsin inhibitor gene CpTI were introduced into tetraploid Isatis indigotica to enhance the resistance to moths.Methods I.indigotica was transformed with a plasmid,pGBI121S4ABC,containing CryIA(c) Bt and CpTI and the selectable gene(Npt-Ⅱ) driven by the CaMV35S promoter via Agrobacterium tumefaciens LBA4404 mediated transformation.Then PCR and Southern blotting assay were conducted followed by moth bioassay test.Results Co-transgenic rate of the two genes in tetraploid I.indigotica was 16.67%.The integration and expression of introduced genes in T0 regenerated transgenic plants were confirmed by Southern blotting assays.Insect bioassay test demonstrated transgenic lines had significant inhibition to moths,compared with wild type control.Conclusion Co-transformation of Bt and CpTI genes is an effective strategy to enhance the resistance to moths for tetraploid I.indigotica.

Optimization of cDNA-AFLP reaction system for Carthamus tinctorius / 中草药

Objective To construct for the heredity linkage map and to study the functional gene research of Carthamus tinctorius,the factors affecting cDNA amplified fragment length polymorphism(cDNA-AFLP) of C.tinctorius were investigated with developing and optimizing the cDNA-AFLP reaction system.Methods Improved Trizol method was used to extract RNA from compounds in new petals specific to safflower.With the help of M-MLV RTase without RNase activity combined with replacement synthesis method,double-stranded cDNA was synthesized from total RNA;cDNA was digested by restriction enzyme MseI/EcoRI and ligated by two steps.Then the products were provided for pre-amplification and selected amplification of different concentration gradients.After tiny modifications of system concentration,finally PAGE electrophoresis and silver-staining were performed.Results High purity and integrated total RNA for later cDNA synthesis were obtained and high quality cDNA was synthesized with the help of M-MLV RTase without RNase activity combined with replacement synthesis method.The cDNA-AFLP reaction system in C.tinctorius was as follows: 250 ng integrity cDNA was digested thoroughly at 37 ℃ for 6 h,and ligated 12 h at 16 ℃.Furthermore,the sample dilution multiplication was 10 fold for pre-amplification and 150 fold for selected amplification under the proper system concentration.According to the above reaction system,the polymorphous strips with high resolution power in PAGE electrophoresis were clear and stable.Conclusion The cDNA-AFLP reaction system established and optimized in this experiment is suitable for the functional gene analysis of C.tinctorius.

Identification of Yangti (crispate rhubarb) by near-infrared diffuse reflectance spectroscopy / 中草药

Object To establish a new method for the identification of Yangti (Chinese herb from Rumex L., Polygonaceae) by near infrared diffuse reflectance spectrometry. Methods Cluster analysis and discriminative analysis were adopted for their identification. Results The method can identify crude Yangti to a certain degree with results coincident with that of the traditional phytotoxnomy. Conclusion This method can be used for the rapid and accurate differentiation of crude drug of Rumex L..

Genetic transformation of Eclipta prostrata by Ri T-DNA / 中草药

Object To establish the culture system of hairy root of Eclipta prostrata (L.) L.. Methods Hairy root of E. prostrata was obtained from infected cotyledon explants after infection with Agrobacterium rhizogenes strains A4, R1601 and ATCC15834, elite strains were screened and growth curves determined. The transformation of Ri T DNA was examined through high voltage paper electrophoresis. Results The hairy root was originally obtained from E.prostrata. The result of high voltage paper elctrophoresis confirmed the transformation of T DNA from Ri plasmid to the hairy root. Conclusion The acquisition of hairy root of E. prostrata provided further a foundation for the industrial production of active drug component.

Study on high-performance liquid chromatographic fingerprints of different germplasms of Flos Carthami / 第二军医大学学报

Objective:To study the high-performance liquid chromatographic(HPLC) fingerprints of different germplasms of Flos Carthami. Methods: HPLC was applied in chromatographic separation of 36 different germplasms of Flos Carthami collected from 11 provinces of China.The chromatography condition was: Agilent Zorbax C_(18) column(4.6 mm?250 mm,5 mm);mobile phase: CAN,0.5% H_3PO_4(from 1090 to 8020);running time: 60 min;flow speed: 1.0 ml/min;detection wavelength: 265 nm;and temperature: 20℃.The data were processed by Chromatographic Fingerprint Station designed by Department of Pharmaceutical Analysis,School of Pharmacy,Second Military Medical University.Different germplasms of Flos Carthami were subjected to cluster analysis and their similarity was also evaluated.Results: The different germplasms of Flos Carthami in this study were grouped into 2 chemical types.Samples from northwest of China were of high similarity;samples from the middle China were of high similarity to those from South China.The similarity of some germplasms of Flos Carthami was significantly different.Conclusion: The chemical components of different germplasms of Flos Carthami are obviously different from each other;selection of germplasm is very important in quality control of Flos Carthami.

STUDIES ON VARIATION OF LANATA C CONTENTS IN LEAVES OF DIGITALIS LANATA WITH RADIOIMMUNOASSAY / 第二军医大学学报

The contents of lanata C in leaves of biennial Digitalis lanata were determined by means of radioimmunoassay.The results show that the contents of lanata C in leaves of Digitalis rapidly increased half a month before blossom and reached maximum between a week before and a week after blossom, and then decreased gradually. The contents of lanata C in a day reached maximum between .1 and 6 0'clock in the afternoon.This study will provide useful information for selecting an appropriate time to collect.