Extraction and Purification of TTX and STX

From the ovaries of T. rubripes, TTX with a purity greater than 60% was prepared, and from the xanthid crab Zosimus aeneus, STX with a purity greater than 80% was prepared by solvent partitioning, activated charcoal treatment, and Bio-Gel P-2 (Bio-Rad Laboratories, Inc., Hercules, CA, USA) and Bio-Rex 70 (Bio-Rad) column chromatography according to the previously reported method [37 (link)], and used for the intrarectal TTX/STX administration experiment. Crystalline TTX (Wako Pure Chemical Industries Ltd., Tokyo, Japan) and an aqueous solution of STX purified from the xanthid crab Z. aeneus were used as external standards for the toxin quantification analyses described below.