Fingerprints Of Drugs And Ananasis Of Active Constituents Of Gushudan

Gushudan prescription is formed according to "The Basic Concept of Establishing Platforms for Operational Techniques in Systems of Elaborately Selecting Small Prescriptions of Traditional Chinese Medicine" and "The Database Management System of PUJIFANG". The prescription is composed of Herba Epimedii, Fructus Cnidii, Rhizoma Drynariae and Radix Salviae Miltiorrhizae.HPLC-Fingerprints that can indicate substances existing in Herba Epimedii, Fructus Cnidii and Rhizoma Drynariae were established with ultraviolet detection respectively. The analysis was performed on a Diamonsil^TM C₁₈ column. A gradient elution of acetonitrile and water (with 0.4% acetic acid) was used in the analysis of Herba Epimedii and Rhizoma Drynariae, and the fingerprint analysis of Fructus Cnidii used acetonitrile and water as mobile phase. The column temperature was maintained at 25℃. Detection wavelength was 270nm and the flow rate was 1.0mL·min^-1. Peaks existing in all sample chromatograms were assigned as "common peaks". The HPLC-FPS similarities of Fructus Cnidii or Rhizoma Drynariae samples were evaluated by Similarity Evaluation System for Chromatographic Fingerprint of TCM of the Chinese Pharmacopoeia Committee. There are 8 "common peaks" in the fingerprint of Rhizoma Drynariae and 7 "common peaks" in the fingerprint of Fructus Cnidii. Good similarities with cosine coefficient above 0.90 were found in fingerprints of Fructus Cnidii or Rhizoma Drynariae from different sources. When the fingerprint data of Herba Epimedii were analyzed by using Cluster Analysis as the measure, Herba Epimedii can be departed to 2 kinds. There are 23 "common peaks" in the fingerprint of Epimedium koreanum and similarities with cosine coefficient above 0.80 were found in fingerprints. Herba Epimedii, Fructus Cnidii and Rhizoma Drynariae can be differentiated from their fakes by the fingerprint.Osteoblast-like UMR106 cells were employed as an in vitro model. The cell proliferation was measured as active indices of stimulating osteoblastic bone formation. The effect of rat serum after administration of Gushudan prescription on osteoblastic proliferation were examined with MTT method. An increase of 45.1% in osteoblastic proliferation was found.An LC-MS/MS method was used for simultaneous determination of icariin, icarisideâ…¡, naringin and osthole in various tissues of rats. With carbamazepine as internal standard, tissues samples were prepared by protein precipitation. The LC-MS/MS analysis was performed on an ACQUITY UPLC^TM BEH C₁₈ column using a linear gradient elution of 0.1% aqueous acetic acid and acetonitrile. The column temperature was maintained at 40℃. The flow rate was set at 0.25mL·min^-1. The mass detection was performed by means of electrospray ionization (ESI) mass spectrometry in positive ion mode with multiple reaction monitoring (MRM). Linear calibration curves of icariin, icarisideâ…¡, naringin and osthole were obtained over the concentration range of 20-400ng·g^-1, 20-400 ng·g⁽-1_, 100-2000ng·g^-1 and 10-1000ng·g^-1, respectively. Tissue distribution results reveal preferential distribution to the stomach and intestine, and display substantial penetration into liver and kidney. No constituent was detected at heart, spleen, lung, adrenal gland or cerebra. The results of this study are helpful for the systematic assessment of the disposition of Gushudan in vivo and ultimately for rational drug therapy.In conclusion, under the direction of theory and clinical practice of TCM, This research is a fundamental work for the research and development of Gushudan prescription.