Studies On Identification, Flavonoids Determination And Antitumour Activities Of Sedum Bulbiferum Makino

“Zhuyajingtian”（Sedum bulbiferum Makino）was an ethnic medicine originatedfrom Genus Sedum plants．The crude drug was harvested in late spring or earlysummer．It was recorded that this medicine was used to treat abdominal pain due tofood stagnation，rheumatic paralysis，malaria，hemostasis，and was also employed asa folk medicine in therapies of carbuncles and furuncles，traumatic injury，scalds，as well as snake bite by Tujia Nationality in West Hunan．As systemic methods of identification and quality analysis of “Zhuyajingtian”were not reported before，the author carried out a series of researches includingmacroscopic，microscopic and TLC identification，determination of flavonoidscontents，extraction and purification methods of total flavonoids，and the anti-tumoreffects of several fractions of the drugs．The preliminary methods for quality analysiswere established，which can be used as a reference of the quality control，theestablishment of quality standard，development and application of the resource，etc．．TLC identification was also conducted in order to differentiate the medicinalplant and3other related medicine from Genus Sedum（Herba Sedi sarmentosi，HerbaSedi Linearis，Sedi Emarginati Herba），and the genetic relationships among themwere discussed by comparing their microstructures and TLC results．Macroscopic， microscopic and TLC results showed that identificationcharacteristics of this crude drug were distinct， such as exodermis and steleparenchyma covered with purplish-red cell，pith cells which were smaller with thincell wall (pith cells were thick-walled in some plants from Sedum)，numerousstomata (2，sometimes3were adjacent) on the lower epidermis of leaf； some clearcharacteristic fluorescent spots were shown in the TLC with reference substance（quercetin） shown in the same place．Compared with Herba Sedi sarmentosi（S.sarmentosum Bunge），Herba Sedi Linearis （S. lineare Thunb.）and Sedi EmarginatiHerba（S. emarginatum Migo.），the differences in microscopic characteristics of“Zhuyajingtian” were obvious．For example，the cell walls of endodermis and exodermis of stem cross section were not thicking；the pith of the stem was somewhatsubrounded with thin-walled cells．All these differences help to distinguish the4Sedum plant drugs．Macroscopic，microscopic and TLC researches came to aconclusion that S. sarmentosum Bunge and S. lineare Thunb. possessed a closerrelationship compared with “Zhuyajingtian” or S. emarginatum Migo.．Through the HPLC sample analysis，it was found that “Zhuyajingtian”contained flavonoid glycosides， and their hydrolyzates included quercetin andkaempferol． And the HPLC method was established to determine the contents ofquercetin and kaempferol in “Zhuyajingtian” from various collectedperiods．YMC-Pack ODS-A（250×4.6mm，5μm）column was used． The mobilephase was methanol-0.4%phosphoric acid（50:50）with flow rate of1.0mL·min-1．The detection wavelength was set at360nm．The column temperature was30℃．Andsample injection volume was10μL．The methodology of HPLC was examined bystandard curve preparation，and analysis of precision，stability，repeatability andaverage recovery．The linear ranges of quercetin and kaempferol were0.03～0.36μg（r=0.9999）and0.08～0.96μg（r=0.9999），respectively，and their averagerecoveries were98.90%（RSD=1.15%）and98.27%（RSD=1.70%），respectively．Themethod was accurate，reproducible and can be used for content determination andquality control of this crude drug．The results demonstrated that the contents ofquercetin and kaempferol in the samples collected in late April was higher than thosecollected in May and June．Furthermore，the samples collected in late April andmid-May contained higher content of flavonoids than that in June．Therefore，it waspreliminarily considered that this crude drug could be collected in the period from lateApril to mid-May．The study on total flavonoid purification methods of “Zhuyajingtian” revealedthat macroporous resin method resulted in a low flavonoid content in the extraction，while the coupling technique of macroporous resin and polyamide absorption columnshowed a good concentration effect． Therefore， the flavonoid content of theextractions of“Zhuyajingtian”reached52.1%，and this have also laid the foundationfor further research of pharmacological activity． The inhibition activities of the different extract fractions and total flavonoids ofSedum bulbiferum Makino on cancer cells proliferation of liver，esophageal and colon(HepG2，EC109，SW480) were tested by MTT assay．In this research，the growthand proliferation of the3cancer cells were all significantly inhibited by the totalflavonoids and ethyl acetate fractions of Sedum bulbiferum Makino．Inhibiting actionincreased when the concentration was improved．Because the content of flavonoidswas comparatively high in ethyl acetate，so it was reasonable to deduce thatflavonoids should be the main active compositions of anti-tumor in “Zhuyajingtian”．In summary，the relevant studies of pharmacognosy laid a foundation for furtherresearch，development and application of this species．Simultaneously， it alsoprovided a useful referance for study and development of Sedum plants．...