台灣株微小亞歷山大藻（ Alexandrium minutum Halim) 毒性成份之純化分析及養殖貝類的麻痺性貝毒調查

Abstract

本研究為首次自台灣株微小亞歷山大藻（Alexandrium minutum Halim）中純化出二種麻痺性貝毒成份。這兩種毒素是取自單一藻株經大量培養之後的水溶性萃取物，先後經二種不同製備層析方法獲得。樣品先經膠濾層析分離後，毒性分液再經離子交換層析，分別以0-0.025N、0.027-0.03N兩種醋酸梯度沖提可得較佳的分離效果。兩毒素經核磁共振氫譜分析進一步肯定為Gonyautoxin-1、Gonyautoxin-4。藻萃取物中尚有其他已被証明但尚未分離之微量毒性成份Gonyautoxin-3、Gonyautoxin-2。測試毒素穩定性的實驗中，觀察到純質Gonyautoxin-1、Gonyautoxin-4在溶於pH=3.3醋酸溶液、溫度-20℃之下，十二天內樣品有逐漸喪失之趨勢。純毒溶液處淤pH值稍高、溫度提升狀態或經冷凍乾燥處理會促進此二種毒素的分解或相互轉換。另一部份的實驗則藉由經年例行性的抽樣分析，檢測出臺灣西南沿海養殖貝類有含麻痺性貝毒現象。受檢測的三種貝類，文蛤、牡蠣以及西施貝，唯有西施貝被檢測出含毒。毒貝幾乎完全來自於屏東地區。高效逆相層析分析的結果顯示，毒貝與該水域微小亞歷山大藻的毒性成份相同，皆為Gonyautoxin-1、Gonyautoxin-2、Gonyautoxin-3,Gonyautoxin-4。有毒西施貝含毒量曾六次超過美日兩國訂定的食用安全標準，足以妨害消費者健康。

This is the first research ever attempting to isolate two components of paralytic shellfish toxins from Alexandrium minutum Halim in Taiwan. The two toxins were obtained by means of two different yet successive preparative chromatography on the water soluble extract that taken from a mass culture of Alexandrium minutum Halim AM-1. The sample was first seperated by a gel filtration chromatography, then the toxin fraction obtained was then puried by an ion exchange chromatography. A better ion exchange seperation was achieved by a linear gradient elution with 0-0.025N acetic acid and a subsequent 0.027-0.03N acetic acid elution on the toxin fraction. Gonyautoxin-1 and Gonyautoxin-4 were thus purified and confirmed by 1H-NMR spectroscopic analysis. There exist two other minor toxins which have been confirmed yet not purified in A. minutum are Gonyautoxin-3 and Gonyautoxin-2. Solution of pure Gonyautoxin1 and Gonyautoxin-4 were found loosing their nature gradually within 12 days even in pH=3.3 acetic acid solution and a temperature below -20℃. Pure form of toxins was also found to degrade or to convert to its epimer in a solution of higher pH or higher temperature. A drying procedure such as lyophilization may cause degradation and interconversion. The second portion of this research is the routine monitoring result of paralytic shellfish toxins in the culturing shellfishes along the south western coast of Taiwan during the year of 1993-1995. Among the three man-raised shellfishes, clam, oyster and purple clam, only purple clam was found to be toxic occasionally. It was also found that almost all the toxic purple clams were from Pintung area. The toxic purple clams were found to contain Gonyautoxin-1, Gonyautoxin-2, Gonyautoxin-3 and Gonyautoxin-4 which were the same composition of toxins found in A. minutum collected in the same area. Sometimes, the toxicity of the purple clam may exceeded the quarantine allowance of U.S. or Japan by the times of six which was thought to cause illness in human.