蛋白質磷酸酵素活性抑制分析及高效液相層析法於藻類營養補充食品內微囊藻毒分析之應用   全文下載

Application of Protein Phosphatase 1 Inhibition Assay and Reversed Phase Liquid Chromatography for the Detection of Microcystins in Algal Dietary Supplements

本研究嘗試調整現有的蛋白質磷酸酵素活性抑制分析法與高效液相層析法於市售螺旋藻及綠球藻類營養補充食品中微囊藻毒（microcystins）的定性定量檢測。利用添加定量毒素的藻樣，測試酵素與化學分析法的各種不同樣品前處理過程以改善毒素分析的靈敏度與精確度。結果顯示本研究所設計的矽膠固相萃取可以顯著的減低樣品萃出液中的介質干擾，因而改善上述兩種分析法對微囊藻毒的偵測。藻樣中的各種不同微囊藻毒可藉高效液相層析在有標準品與可容許的解析度下可分別定性定量，而在蛋白質磷酸酵素抑制分析上可獲得不同毒素累加的活性，再依標準品的酵素抑制分析轉換成毒素標準品MCYST-LR 當量，本研究所改善的操作與定量方法，在上述兩種模式下均可達50 ppb的靈敏度，可應用於藻類健康食品可能受到微囊藻毒污染的品管檢測。應用本研究所建立的方法於市售藻類營養食品上的微囊藻毒分析，證實分析到的九樣產品都未受到微囊藻毒的污染。

Protein phosphatase 1 inhibition assay and reversed phase high performance liquid chromatography were adjusted for the detection of microcystins in Chlorella and Spirulina dietary supplements. Various sample preparation procedures for the enzyme and chemical assays were tested by the toxin-spiked samples to improve the sensitivity and accuracy of these methods. A silica gel solid phase extraction coupled with a deactivated elution had remarkably reduced the matrix interference from sample extracts and improved the detection of microcystins in both assays. High performance liquid chromatography could give the composition of different microcystins in the sample if the resolution of separation and toxin standards were available. On the contrary, protein phosphatase 1 inhibition assay gave an accumulated inhibition activity of the sample that could be converted to MCYST-LR equivalent according to the IC50 of this standard. A good sensitivity of 50 ppb of microcystin in dried algal samples, was reached using the methods established in this research. Both enzyme and chemical assays were applied for the detection of possible microcystin contamination in the commercial products of algal dietary supplement. It was found that all nine samples of algal product examined in this study were free from microcystins.