2000 Fiscal Year Final Research Report Summary
Studies on paralytic toxins in a ribbon worm adherent to cultured oysters in Hiroshima Bay, Hiroshima Prefecture, Japan
| Project/Area Number |
11660205
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Fisheries chemistry
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| Research Institution | Hiroshima University |
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Principal Investigator |
ASAKAWA Manabu Hiroshima University, Faculty of Applied Biological Science, Associate Professor, 生物生産学部, 助教授 (60243606)
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| Project Period (FY) |
1999 – 2000
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| Keywords | Hiroshima Bay / ribbon worm / tetrodotoxin / oyster / テトロドトキシン / 付着生物 |
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| Research Abstract |
In 1998, in Hiroshima Bay, Hiroshima Prefecture, Japan, specimens of the ribbon worm, "himomushi" adherent to the shells of cultured oysters were found to contain toxins which showed strong paralytic action in mice. The specimens were identified as an undescribed species of the genus Procephalothrix (formerly Cephalothrix). The maximum toxicity (as tetrodotoxin, TTX) was 14,734 MU/g whole body. The "himomushi" toxin (HMT) was extracted from the worm with 80% methanol acidified with acetic acid and the extract defatted with dichloromethane. The aqueous layer was chromatographed on activated charcoal and the unbound and bound toxic fractions were analyzed by high-performance liquid chromatography and gas chromatography-mass spectrometry. It was rather unexpectedly revealed from these results that HMT was comprised of TTX, 4-epiTTX, anhydro TTX and unidentified toxins. The main toxic component of HMT was isolated from a pooled specimen by a method which consisted of treatment with activat
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ed charcoal, chromatography on Bio-Gel P-2 and Bio-Rex 70 (H^+ form), and finally crystallization from an acidified methanolic solution. The recrystallized toxin showed a specific toxicity of 3,520 MU/mg. This toxin showed (M+H)^+ and (M+H-H_2O)^+ ion peaks at m/z 320 and 302, respectively by electrospray ionization-mass spectrometry(ESI-MS). The molecular weight thus determined was 319 for TTX in accordance with its reported molecular weight. The absorption band at 3353, 3235, 1666, 1612 and 1076cm^<-1> were observed in infrared spectrum of this toxin. This spectrum was indistinguishable from that of TTX.The ^1H-NMR spectrum for the recrystallized toxin was the same as that for TTX.The pair of doublets centered at 2.33 (J=10.0 Hz) and 5.48ppm(J=10.0 Hz), which are characteristic of tetrodotoxin, were shown to be coupled by double irradiation. By GC-MS analysis, the alkali-hydrolyzate of this toxin, like TTX, showed the quinazoline skeleton (C_9-base).
These findings are the first direct evidence of the existence of TTX in the ribbon worms found on the surface of the shells of oysters hanging onto floating oyster culture rafts in Hiroshima Bay, Hiroshima Prefecture. Less
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Research Products
(2 results)
