Studies On The HPLC Fingerprint Chromatograph Of Toad Venom From Different Origins

Toad venom, in the form of dried toad secretions derived from either Bufo bufo gargarizans Cantor or B. melanostictus Schneider, has been widely used as a cardio tonic, diuretic, anodyne and antineoplastic agent in Asia. The medicinal resource of toad venom is abundant in China. Besides the above 2 species, other Bufo sp. such as B. b. andrewsi Schimidt and B. raddei STRAUCH were also used to provide secretions. Further more, toad venom is an expensive medicine, the adulterant can be found in commercial products frequently. The confusion of the allied species and the adulteration took great troubles in the quality control of the drug and its preparations.In this paper, a standard and scientific strategy for quality control of toad venom was established, based on a series of studies, including the HPLC fingerprint chromatographic analysis, the chemical constituent isolation and identification, and the animal collection and variety certification studies. It would be a great help for the quality standardization of toad venom crude drug and its preparations.Main contents and results as follows:1. In this paper the chemical constituents, the pharmaceutical activities and the quality control method for toad venom were reviewed and compiled.2. A HPLC method for simultaneous determination of the major bioactive bufogenins and bufotenines in toad venom was elaborated. The optimized HPLC conditions insured the simultaneous determination of 10 bufogenins and 4 bufotenines in toad venom. The method had good linearity, reproducibility, precision and accuracy, and had been applied to quality evaluation of TCM toad venom. The HPLC analytical method for bufotenines was established for the first time. The developed method could be used to analyze the bufotenines in toad venom and its preparations basis of water-soluble substances.3. Chemical studies on toad venom from different species were performed. 19 pure compounds were isolated from the venom of B. b. gargarizans, 5 pure compounds were isolated from the venom of B. melanosticus and 5 pure compounds were isolated from B. raddei. The structures were identified through the studies of physico-chemical properties and spectroscopic analysis. These compounds were further purified to the...