Analysis of DSP Toxins and other Lipophilic Toxins in Japanese Bivalves – Performance and Correlation of LC-MS versus Mouse Bioassay

Toshiyuki Suzuki1*,Ph.D.

Taketo Jin2, Yuri Shirota3, Yuki Kosaka4, Junko Kimura2, Naotsugu Hata5, Hajime Uchida1,6, Ryuichi Watanabe1, Ryoji Matsushima1, Hiroshi Nagai 6, Takeshi Yasumoto1, Yutaka Okumura7, Takashi Kamiyama7

1 National Research Institute of Fisheries Science, 2-12-4 Fukuura, Kanazawa-ku, Yokohama, Kanagawa 236-8648, Japan

2 Aomori Prefectural Institute of Public Health and Environmen,. 1-1-1 Higashi-tsukurimichi Aomori, Aomori 030-8566, Japan

3 Japan Frozen Foods Inspection Corporation, 1-24-18 Takasago, Miyagino-ku, Sendai, Miyagi 983-0014, Japan

4 Aomori Prefectural Industrial Technology Research Center Fisheries Research Institute, 10 Moura Tsukidomari Hiranai-machi, Aomori 039-3381, Japan

5Mie Prefecture Fisheries Research Institute, Shima, Mie 517-0404, Japan

6Tokyo University of Marine Science and Technology, 4-5-7, Konan, Minato-ku, Tokyo, 108-8477, Japan

7 Tohoku National Fisheries Research Institute, 3-27-5 Shinhama Shiogama, Miyagi 985-0001, Japan

* corresponding author: tsuzuki@affrc.go.jp


Diarrhetic shellfish poisoning (DSP) is a severe gastrointestinal illness caused by the consumption of shellfish contaminated with DSP toxins. Based on their structures, DSP toxins were initially classified into three groups, okadaic acid (OA) analogues, pectenotoxins (PTXs), and yessotoxins (YTXs). Because PTXs and YTXs have been subsequently shown to have no diarrhetic activities, PTXs and YTXs have recently been eliminated from the definition of DSP toxins. Mouse bioassay (MBA), which is the official testing method of DSP in Japan, has been protecting human health against DSP for more than 30 years. When the toxicity of the bivalves exceeds the quarantine levels (0.05 MU/g wet weight for DSP), harvesting ceases in Japan. No human poisonings due to contaminated bivalves with DSP toxins distributed on commercial markets have been reported in Japan since the establishment of the monitoring system for shellfish toxins in 1979. On the other hand, the technique of electrospray ionization (ESI) liquid chromatography-mass spectrometry (LC-MS) has proven to be one of the most powerful tools for the detection, identification and quantification of DSP and other lipophlic toxins, however LC-MS requires expensive and complicated hardware.

In the present study, performance and correlation of LC-MS versus MBA were investigated. More than 800 bivalve samples collected in Japan were analyzed by both LC-MS and MBA. Comparison between the LC-MS and the MBA indicates that false negative results in OA testing by the MBA in comparison with the LC-MS were less than 3 %. YTXs and PTXs were poorly detected by the MBA due to low recoveries of these lipophilic toxins in the partition procedure in the MBA. The result demonstrates that the MBA, at least with our local profiles of lipophilic toxins, is still useful in Japan for routine monitoring of OA analogues in bivalves. A convenient and economical HPLC method for okadaic acid analogues as 9-anthrylmethyl esters, now refined with an automatic column switching cleanup method, will also be presented.

 

Dr. Toshiyuki Suzuki received his Ph.D. degree in the Chemistry of Marine Products from Hokkaido University in 1993. Dr. Suzuki joined Tohoku National Fisheries Research Institute in 1993, Sendai, Japan. He has been an invited visiting scientist at the Cawthron Institute, New Zealand, and Canada’s National Research Council, Marine Biosciences. Now Director of Biochemistry and Food Technology at the National Research Institute of Fisheries Science at Yokohama, Japan, his research is focused on the development of analytical methods for marine biotoxins and metabolism of marine biotoxins in shellfish. In 2007, Dr. Suzuki was honored with the Scientific Achievement Award in Fisheries Science (The Japanese Society of Fisheries Science Award) for development of analytical methods for shellfish toxins and their application for the elucidation of toxification mechanism in bivalves.