title: Preparative isolation of six anti-tumour biflavonoids from Selaginella doederleinii Hieron by high-speed counter-current chromatography.
abstract: BACKGROUND
  Biflavonoids are the primary constituents of Selaginella doederleinii Hieron, to which different bioactivities have been attributed, including anti-cancer, anti-inflammatory, anti-oxidant, anti-fungal and anti-virus activity. However, effective methods for separation of these compounds are not currently available.
  
  
  OBJECTIVE
  To develop a high performance and bioassay-guided method for preparative isolation of biflavonoids from S. doederleini via high-speed counter-current chromatography (HSCCC).
  
  
  METHODS
  The anti-proliferation effects of four fractions (70% ethanol, petroleum ether, dichloromethane and acetic ether extracts) of S. doederleinii on five human cancer cells were monitored. The dichloromethane and acetic ether extracts showed good cytotoxicities to the studied cancer cell lines, guiding the subsequent separation. Two solvent systems composed of n-hexane:ethyl acetate:methanol:water (1:2:1.5:1.5, v/v) and n-hexane:ethyl acetate:methanol:water (3:2:3:2, v/v) were developed for separation of the active fractions, respectively. Identification of the biflavonoids was performed by EI-MS(n) , (1) H- and (13) C-NMR.'
  
  
  RESULTS
  Under the optimised conditions, 12.6 mg amentoflavone (91.4%), 6.6 mg robustaflavone (90.4%), 7.5 mg 2'', 3''-dihydro-3', 3'''-biapigenin (98.2%) and 7.3 mg 3', 3'''-binaringenin (90.3%) from acetic ether extract (500 mg) and 6.3 mg heveaflavone (93.5%) and 5.3 mg 7, 4', 7'', 4'''-tetra-O-methyl-amentoflavone (94.5%) from dichloromethane extract (200 mg) were obtained, respectively. The anti-proliferation effects of the six biflavonoids on the five human cancer cells were further verified.
  
  
  CONCLUSIONS
  The study provides methodological references for simultaneously preparative isolation of several bioactive biflavones from the herbal family of Selaginella. It is the first report discovering 2'', 3''-dihydro-3', 3'''-biapigenin and 3', 3'''-binaringenin from this herb and describing their cytotoxicities to human cancer cell lines.
authors.affiliation: Department of Pharmaceutical Analysis, Faculty of Pharmacy, Fujian Medical University, Fuzhou, 350004, China.
authors.forename: Shaoguang
authors.initials: S
authors.lastname: Li
authors.forename: Meifeng
authors.initials: M
authors.lastname: Zhao
authors.forename: Yuxiang
authors.initials: Y
authors.lastname: Li
authors.forename: Yuxia
authors.initials: Y
authors.lastname: Sui
authors.forename: Hong
authors.initials: H
authors.lastname: Yao
authors.forename: Liying
authors.initials: L
authors.lastname: Huang
authors.forename: Xinhua
authors.initials: X
authors.lastname: Lin
mesh_terms: D000970:Antineoplastic Agents / Q000737:chemistry / Q000302:isolation & purification* / Q000494:pharmacology; D044946:Biflavonoids / Q000737:chemistry / Q000302:isolation & purification* / Q000494:pharmacology; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D003377:Countercurrent Distribution / Q000379:methods*; D006801:Humans; D015394:Molecular Structure; D035261:Plant Components, Aerial / Q000737:chemistry; D010936:Plant Extracts / Q000737:chemistry / Q000302:isolation & purification* / Q000494:pharmacology; D010946:Plants, Medicinal; D032503:Selaginellaceae / Q000737:chemistry*; D021241:Spectrometry, Mass, Electrospray Ionization
chemical_list: D000970:Antineoplastic Agents; D044946:Biflavonoids; D010936:Plant Extracts
keywords: Activity-guided preparative isolation; HSCCC; Selaginella doederleinii Hieron; biflavonoids; cytotoxicity