title: Comparative Analysis of the Bufonis Venenum by Using TLC, HPLC, and LC-MS for Different Extraction Methods.
abstract: OBJECTIVE
  Toad venom, called Chan-Su, is a traditional Oriental medicine secreted from the auricular and the skin glands of the Bufo bufo gargarizanz Cantor or B. melanosticus Schneider and has been widely used in China, Korea and other parts of Asia for the treatment of pain, heart conditions, and cancer. We examined the concentrations of the main chemical constituents within a commerciallyavailable toad venom product and compared the levels for different extraction methods.
  
  
  METHODS
  Toad venom was extracted using either cold or hot water, ethanol (EtOH), methanol (MeOH), or ethyl acetate (EtOAc), was fractionated using precipitation or reflux, and was then analyzed using thin layer chromatography (TLC), high-performance liquid chromatography (HTLC), and liquid chroma-tography - mass spectrometry (LC-MS). Individual components were identified by comparisons of the retention times, the ultraviolet spectra, and mass spectras and differences in chemical constituents for different solvents and extraction methods are presented.
  
  
  RESULTS
  Components with authentic standards, including serotonin and bufodienolides (cinobufagen, bufalin, cinobufalin, and resibufogenin), were detected. The water extract of toad venom contained the greatest amount of serotonin (75.7 ± 0.1 mg/g), but very small amounts of bufodienolides (3.8 ± 0.0 mg/g). In contrast, the use of MeOH or EtOH extraction solutions resulted in 5-26 times higher concentrations of bufodienolides, with only trace amounts of serotonin. The relative and the absolute concentrations of the component also varied based on the extraction method; i.e., EtOH extracts yielded the greatest total amounts of bufodienolides, and EtOAc precipitation had the lowest amounts of bufodienolides.
  
  
  CONCLUSIONS
  Toad venom consists of serotonin and several bufodienolides, and the choice of solvent to extract chemical the constituents is important as a way to enrich the purported active components for treating different conditions.
authors.affiliation: East-West Cancer Center, Dunsan Oriental Hospital of Daejeon University, Daejeon, Korea.
authors.forename: Hyo-Jae
authors.initials: HJ
authors.lastname: Lee
authors.affiliation: Integrative Medicine Program, Department of General Oncology, The University of Texas MD Anderson Cancer Center, Houston, USA.
authors.forename: Fan-Pei
authors.initials: FP
authors.lastname: Koung
authors.affiliation: Korean Pharmacopuncture Institute, Seoul, Korea.
authors.forename: Ki-Rok
authors.initials: KR
authors.lastname: Kwon
authors.affiliation: Korean Pharmacopuncture Institute, Seoul, Korea.
authors.forename: Dae-In
authors.initials: DI
authors.lastname: Kang
authors.affiliation: Integrative Medicine Program, Department of General Oncology, The University of Texas MD Anderson Cancer Center, Houston, USA.
authors.forename: Lorenzo
authors.initials: L
authors.lastname: Cohen
authors.affiliation: Integrative Medicine Program, Department of General Oncology, The University of Texas MD Anderson Cancer Center, Houston, USA.
authors.forename: Pei-Ying
authors.initials: PY
authors.lastname: Yang
authors.affiliation: East-West Cancer Center, Dunsan Oriental Hospital of Daejeon University, Daejeon, Korea ; Integrative Medicine Program, Department of General Oncology, The University of Texas MD Anderson Cancer Center, Houston, USA ; Korean Pharmacopuncture Institute, Seoul, Korea.
authors.forename: Hwa-Seung
authors.initials: HS
authors.lastname: Yoo
keywords: Bufonis venenum; HPLC; LC/MS; TLC; bufadienolides; bufotoxin