建立金針菇免疫調節蛋白 FIP-fve 品管指標及評估 FIP-fve 之生物可及性

Establishment of Quality Control Indicator and Bioaccessibility Evaluation for Fungal Immunomodulatory Protein FIP-fve

陳偉齊(Wei-Chi Chen)

指導教授 ： 許輔

國立臺灣大學/生物資源暨農學院/園藝學研究所/碩士(2015年)

https://doi.org/10.6342/NTU.2015.00233

摘要

金針菇含有免疫調節蛋白 FIP-fve，具有抗過敏、抗腫瘤、免疫調節等生物活性，因此金針菇具有開發為健康食品素材的潛力。健康食品的開發必須要提出功效成分的科學證據，並能管理品質，使其在食用上須安全無虞。目前 FIP-fve 沒有一套完善的分析方法，從產品開發到品管皆有困難。因此，本研究的目的為建立分析 FIP-fve 的品管指標，測定金針菇中 FIP-fve 的含量，作為產品開發和品管依據，同時評估金針菇產品在食用上的安全性，並研究 FIP-fve 的生物可及性。
第一部分，建立 HPLC 分析 FIP-fve 的方法，並予以確效，並評估專一性、線性、準確度、精密度和偵測極限。結果顯示此分析方法穩定且值得信賴，並應用於後續的分析。第二部分，研究乾燥金針菇中 FIP-fve 的含量變化，找出最適當的乾燥條件用於往後的大量生產，結果發現金針菇分別經真空冷凍乾燥、真空微波乾燥和熱風乾燥後，三者 FIP-fve 含量並無顯著差異，且在熱風乾燥時 FIP-fve 含量會隨著溫度上升而顯著的下降。第三部分，探討食用安全性，對滅菌處理及金針菇的火菇毒素 (flammutoxin, FTX) 進行研究。首先將金針菇產品以加馬輻射滅菌，發現並不影響 FIP-fve 的含量。另一方面，火菇毒素為金針菇具溶血活性的毒蛋白，實驗發現火菇毒素於冷凍乾燥的金針菇中含量最多，其次為熱風和微波乾燥，而當熱風乾燥溫度提高時火菇毒素含量亦會顯著的下降。利用小鼠紅血球測其溶血活性，發現在熱風乾燥 70℃ 後，仍具有溶血活性。為解決火菇毒素的問題，初步以 60℃ 熱水預處理 5 分鐘後再進行50℃ 熱風乾燥，可使火菇毒素含量顯著下降並保留大部分的 FIP-fve。第四部分，在個別模擬胃及腸道消化的模式，在胃蛋白酶和胰酶的作用下，FIP-fve 僅少部分被消化。使用連續式胃腸模擬消化 FIP-fve，其在胃及腸道各消化兩小時後，FIP-fve 並不會降解。綜合以上結果，本實驗對金針菇健康食品開發從品管、安全性到消化已做一全面性的研究，證明金針菇具有作為健康食品的可行性。

關鍵字

FIP-fve ； 分析方法確效 ； 火菇毒素 ； 生物可及性

並列摘要

Enoki mushroom was known as containing FIP-fve, a fungal immunomodulatory protein with several biological activities including anti-allergy, anti-tumor, and immunomodulation, and it showed a high potential as a good functional ingredient. The development of a successful health food product demanded the necessary requirements: (1) its ingredients should provide healthy and functional benefits with significant science evidences, (2) its functional compartments should be determined, and (3) the levels of the bio-activating compartments should be assured. Nevertheless, it lacked an appropriate analytical method available for FIP-fve analysis currently, which limited the utilization and application of enoki mushroom as a functional ingredient. Therefore, the aims of this study were to establish and to validate an analytical method as a quality control indicator of FIP-fve determination. In addition, the processing technologies to dehydrate the mushroom, to retain most of FIP-fve contents, and to eliminate the activity of hemolytic protein flammutoxin (FTX) were researched. Moreover, the bioaccessibility of FIP-fve within enoki juice and products were performed in this study.
First, we established and validated a HPLC method for FIP-fve determination. Through considering its specificity, linearity, accuracy, precision and quantitation limit, the results showed that this method was reliable and could be applied for subsequent analysis. Second, we studied the variation of FIP-fve contents in dried enoki mushroom, to figure out the best dehydration conditions for industrial production. The contents of FIP-fve showed no significant difference among vacuum freeze drying, vacuum microwave drying and hot air dehydration methods. Moreover, content of FIP-fve would be markedly dropped while the dehydration temperature was risen. Third, we investigated the processing stability of FIP-fve and FTX. Mushroom products exposed to gamma radiation presented no discernible difference in FIP-fve content. In addition, freeze dried mushroom yielded the highest level of FTX, which was followed by hot air dehydration and microwave drying, respectively. As the same as FIP-fve, content of FTX was dropped significantly while temperature rising and it was found that the hot air dehydration at 70℃ could not completely destroy the hemolytic activity of FTX. In order to provide a safe mushroom preparation, the mushrooms were preheated and treated using hot water at 60℃ before hot air dehydration at 50℃. Under this hot-water treatment, content of FTX dropped significantly and retained most of the FIP-fve levels. Forth, FIP-fve was found to be slightly degraded when using the model of mimicked stages of digestion in stomach and intestine. FIP-fve was not degraded when using the mimicked continuous gastrointestinal digestion model. In conclusion, this study made comprehensive researches of FIP-fve preparation and provided more evidences for enoki mushroom as a functional food ingredient for health food utilization.

並列關鍵字

FIP-fve ； analytical method validation ； flammutoxin ； bioaccessibility

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