122(P-80) 4-L-Cysteinetetrodotoxin from the liver of puffer fish and the reaction of tetrodotoxins with thiols
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Metabolic pathway of tetrodotoxin (TTX, 1), a potent neurotoxin in puffer fish, has not been clarified yet, for puffer fish and also for human. We found a new natural analog of TTX, 4-L-cycteineTTX (4-CysTTX, 2) from the liver of the puffer fish, Fugu pardalis, collected in spring in 2000. In this paper, structural confirmation of 2, and the reaction of TTXs with thiols are described. 2 (ca. 0.4mg) was isolated from 200g of liver of the puffer fish by sequential liquid chromatography. Based on the spectroscopic data, 2 was determined as an analog of 1 of which equatorial hydroxyl group at C4 was substituted by sulfur atom of cysteine. Configuration of cysteine was confirmed to be L by using Marfey reagent for cystine obtained from 2 by I_2 oxidation. 2 was produced by the reaction of 4,9-anhydroTTX (4) and large excess (100-500 equiv.) of cysteine in phosphate buffer (pH 7.0-8.0) at 40℃ for 90min. It was pH dependent reaction. Other thiols, glutathione (GSH), mercaptoethanol (ME), and cysteinyl glycine were similarly reacted with 4 to produce sulfur substituted compounds at C4 of 1. On the condition with much more excess of thiols (more than 20000 equiv.) and for longer reaction time (longer than 8hr), ME and GSH were also reacted with 1 to produce the same derivatives from 4.
