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標題: | 二角多甲藻水層萃取物之毒性研究 The study of toxicity of water-soluble extract from Peridinium bipes |
作者: | 塗珮琪 |
出版年 : | 2003 |
學位: | 碩士 |
摘要: | 德基水庫因過度優養化而發生由二角多甲藻(Peridinium bipes)—後文簡稱二甲藻形成之淡水紅潮,本研究以水混合海砂對所採集之二甲藻進行研磨萃取,發現所得之水層粗萃取物具有多樣生物活性。13種測試藻株中有7種對水層粗萃取物較具敏感性,其生長速率皆受到不同程度的抑制,本研究進一步以銅綠微囊藻(Microcystis aeruginosa)進行生理活性之抑制測試。發現在二甲藻水層粗萃取物處理下,可觀測到微囊藻產氧速率隨著劑量與測試時間增加而遞減,耗氧速率則反之。光合作用之第二光系統之螢光發散量(685nm)亦與所添加之劑量成正相關,故推論可能藉阻斷電子傳遞鏈來抑制光合作用。二甲藻水層粗萃取物亦造成微囊藻無法維持正常細胞質之酸鹼度,同時發生大量鉀離子滲漏至胞外的現象。初步純化後之水層萃取物分樣,在10分鐘內甚至可造成胞內鉀離子之完全滲漏,此現象推論係細胞膜破裂之效應,而不只是膜上特定離子運輸酵素受到抑制。另以二甲藻水層粗萃取物對11種消化?進行敏感度測試,其中以凝血?最為敏感,不過,酵素受抑制作用之機制則未知。進一步純化水層粗萃取物後發現,活性物質之分子量均小於3kD,再進行陽離子交換層析分離後得到三個毒性分樣,藉由其210nm與254nm之吸收值與蛋白質含量測定等性質,推測活性物質可能為具有環狀構造之胺基酸之勝?(peptides)。 Water bloom resulted from Peridinium bipes, a freshwater dinoflagellate, has commonly occurred in Dergie dam of the mid of Taiwan. Allelopathic activity was found in the water-soluble extracts of this alga. This study is aimed to study the toxic effects and to characterize the allelochemicals. Seven of thirteen strains of phytoplankton tested were susceptible to the water-soluble extracts of P bipes. A further study done with Microcystis aeruginosa showed that the photosynthetic O2-evolution and electron transfer between photosystems I and II, but not the respiration, were inhibited in the presence of extracts. This agreed with the phenomenon of the enhancement in the emission of PSII fluorescence when the extract was added to M. aeruginosa cells. The treatment with P. bipes extract has caused the concentration-and time-dependent reductions in uranine accumulation in M. aeruginosa cells, associated with a significant and rapid leakage of K+ from intra- to extra-cellular medium. This was considered a phenomenon of damage in membrane integrity of the treated cells. As a result, a change in the membrane permeability along with a loss in the pH gradient across the plasma membranes might occur. Further study was done to test the inhibition of proteases in vitro by the presence of P. bipes extracts. Among 11 proteases tested, thrombin was the most sensitive one. When the extracts were further purified by centricon and centriprep, the fractions with toxic activity were found at those <3 kD. From them, three fractions were obtained when purified with ion chromatography. These fractions showed positive reaction with protein staining, suggesting that they were peptides. Although P. bipes seems to be cable of producing various kinds of inhibitors, present study suggests that peptide is one of such compounds accounting for allelopathic effects. The primary effects of such allelochemicals seem to act on the plasma membranes of the target algal cells and to inhibit the proteases of the predators. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/75423 |
全文授權: | 未授權 |
顯示於系所單位: | 植物科學研究所 |
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