Objective: To establish the characteristic map method for simultaneous determination of four kinds of active ingredients including arenobufagin in toad skins from different areas

and provide the basis for its quality control and medicinal material identification. Method: A Phenomenex Gemini (4.6 mm×250 mm

5 μm)column was used with the mobile phase consisting of acetonitrile(A)-water(B)(0-30 min

10%-45%A;30-45 min

45%-60%A);flow rate 1.0 mL·min-1;column oven temperature was 35℃;detection wavelength was 296 nm.The characteristic map method was used to determine the contents of four bufogenin constituents such as arenobufagin. Result: The content of arenobufagin in the toad skin from Sichuan province was highest (0.252 4%

0.158 4%

0.181 1%

0.125 8% and 0.108 8% respectively for large flowered skin

middle black skin

super large skin

brown skin and small black skin)

followed by the black skin from Jiangsu province (0.227 5%). The content of cinobufotalin was highest in large flowered skin from Sichuan province (0.214 3%)

followed by black skin from Jiangsu province (0.147 8%). The contents of cinobufagin in the small black skin from Sichuan province (0.051 0%) and small flowered skin from Shandong province (0.136 1%) were higher than those from other areas and varieties. In general

the contents of four kinds of active ingredients in Sichuan province were higher

especially the large flowered skin from Sichuan Province. Conclusion: The characteristic map method was accurate and reliable in determining the contents of these four bufogenin constituents

which can be used for identification and quality control of told skins. In addition

the contents of the four bufogenin constituents in different areas and varieties were significantly different.