The Researches On Preparation And Activities Of Paralytic Shellfish Poisons

As a type of bioactive compounds, paralytic shellfish poisons (PSP) were mainly produced by dinoflagellate. PSP toxins have potential application in researches of harmful algal bloom toxicity, neurology and molecular biology, and development of new drug. However, the lackness of pufified PSP hampered the related researches. In this thesis, using the PSP-producing dinoflagellate A/exandrium tarnarense, the technology on seperation and purification of PSP toxins have been studied and developed. Meanwhile, the activities of PSP were studied and discussed. The methods for culturing of Alexandrium tarnarense were established and improved in the lab. The methods on concentrating and harvesting the cells were discussed. Crude PSP solution were obtained by sonicating Alexandrium tamarense cells and then centrifugating. After the impurities were excluded by large size gel filtration, PSP were then seperated and purified by small size gel filtration and ion-exchange resin. The results showed that the impurities can be effectively excluded by large size gel, gonyautoxin (GTX) and C toxins could be seperated small size gel filtration, GTX2,3 and GTX 1,4 could be seperated by ion-exchange resin. GTX2,3 and GTX1,4 were the final products. Mouse assay and fluoresence detector were used in the whole preparation process to monitor the toxins. The mice used for test of toxicity showed typical symptoms of PSP poisoning. The extracted PSP could reversibly inhibit thevoltage-gated sodium current of mouse motor nerve terminals and the whole NG 108-15 cells, with the identical active mechanism of the standard PSP. The retention time of the purified GTX2,3 and GTX 1,4 were the same as PSP standards by HPLC and the m/z of GTX2,3 and GTXI,4 were the same as PSP standards by MS. The pharmaceutical activities of the purified toxins were studied with animal models. The local anesthetic, analgesia, muscular relaxation of GTX2,3 were studied by rat sciatic anesthesia, mouse hot plate and mouse climbing web tests respectively. The results showed that the local anesthetic of rabbit was significant after percutaneous sciatic nerve injections with GTX2,3 equivalent to O.4tg STX/kg. The analgesia of mouse was significant after muscular injections with GTX2,3 equivalent to 5.99p.g STX/kg. The muscular relaxation of mouse was significant after subpercutaneous injections with GTX2,3 equivalent to 6.49ug S TX/kg.