臺灣博碩士論文加值系統

本研究之目的為應用波長範圍介於400至2498 nm之近紅外光譜（near infrared spectroscopy, NIR），開發快速檢測白米中黃麴毒素（aflatoxin）之方法。國際癌症研究機構（International Agency for Research on Cancer, IARC ）已於2002年將黃麴毒素列為一級致癌物，確定對人體具有致癌性。臺灣位於亞熱帶，高溫高濕的環境有利於黴菌之生長，所以如何快速且準確的檢驗出白米中的黃麴毒素含量，為本研究之重點。由人工汙染的白米樣品利用FOSS NIRS 6500型分光光度計所測得的光譜值和化學分析方法──液相層析串聯式質譜儀(LC-MS/MS)所得的數據，透過支援向量機（support vector machine, SVM）和多重線性迴歸（multiple linear regression, MLR）分別來建立定性和定量的校正模型，定性分析部分，分別對有無遭受汙染和不同汙染濃度進行判別，預測正確率分別為88.89 %和87.5 %。定量分析部分，黃麴毒素濃度35 ppb以上之樣品數為36個，檢量線之判定係數（RSQ）為0.74，標準預測誤差（standard error of prediction, SEP ）為13.552 ppb；黃麴毒素濃度35 ppb以下之樣品數為35個，檢量線之判定係數為0.92，標準預測誤差（SEP）為3.533 ppb，由分析結果推論黃麴毒素以35 ppb為界，呈現非線性關係。本研究證實近紅外光譜法結合化學計量學可以快速且準確檢測白米中的黃麴毒素。

The aim of this study is to develop a rapid and accurate method for detecting aflatoxin B1 (AFB1) content in rice by using near infrared (NIR) spectroscopy. AFB1 has been classified by the International Agency for Research on Cancer (IARC) as Group 1: The agent (mixture) is carcinogenic to humans. In Taiwan, environment with high temperature and humidity provides favorable conditions for fungal propagation and aflatoxin production in stored rice. In the present study, the AFB1 contents of the artificially contaminated rice were evaluated by NIR spectroscopy associated with chemometric methods. The NIR spectra and AFB1 contents of the rice samples were measured by FOSS NIRS 6500 spectrophotometer and liquid chromatography tandem-mass spectrometry (LC-MS/MS). The prediction accuracies of the qualitative models based on support vector machine (SVM) algorithm for all samples and the contaminated samples were 88.89 % and 87.5 %, respectively. The coefficient of determination (R2) of the calibration models built by multiple linear regression (MLR) for the samples with AFB1 concentrations higher and lower than 35 ppb were 0.74 and 0.92, respectively. In summary, integration of NIR spectroscopy and chemometrics provides a useful tool for monitoring AFB1 contents in rice.

中文摘要 I
英文摘要 II
誌謝 III
目錄 V
表目次 VIII
圖目次 X
第一章 緒論 12
第二章 文獻回顧 14
第一節 米 14
第一項 米之需求量 14
第二項 米之分類 14
第二節 黃麴毒素之介紹 15
第一項 黃麴毒素之來源 15
第二項 黃麴毒素之特性 16
第三項 黃麴毒素之毒性與中毒症狀 17
第四項 各國對黃麴毒素的限量標準 18
第五項 黃麴毒素之檢測方法 20
第三節 近紅外光譜法 22
第一項 紅外光譜法之簡介 22
第二項 近紅外光譜技術之原理 23
第三項 原子分子振動情形 23
第四項 近紅外光譜之檢測 24
第四節 近紅外光分析技術 26
第一項 近紅外光譜儀之介紹 26
第二項 光譜前處理 28
第三項 定性分析 29
第四項 定量分析 31
第五項 檢量線之建立 32
第六項 近紅外光譜技術之應用及特點 33
第五節 近紅外線光譜法於受黃麴菌及黃麴毒素汙染農產品之相關研究 36
第三章 研究動機與目的 43
第四章 材料與方法 44
第一節 實驗材料 44
第一項 標準品製備 44
第二項 樣品製備 44
第三項 藥品與試劑 47
第四項 器材與儀器 47
第二節 實驗流程 48
第三節 分析方法 49
第一項 近紅外光譜法（NIR） 49
第二項 高效液相層析串聯式質譜儀 51
第三項 分析軟體 54
第四項 數據分析 55
第五項 性能評估的相關統計 56
第五章 結果與討論 58
第一節 近紅外光譜分析 58
第二節 黃麴毒素之LC-MS/MS分析 63
第三節 黃麴毒素之SVM光譜定性分析 69
第四節 黃麴毒素之光譜定量分析 72
第一項 部分最小平方迴歸（partial least squares regression, PLSR）分析 72
第二項 多重線性迴歸（multiple linear regression, MLR）分析 72
第六章 結論 85
參考文獻 86

表目次
表一、臺灣對食品中黃麴毒素限量之標準 19
表二、美國食品中黃麴毒素之限量標準 19
表三、歐盟食品中黃麴毒素之限量標準 20
表四、近紅外光譜技術的一些代表性應用 34
表五、近期有關於使用NIR定性/定量檢測不同農產品中黃麴毒素汙染的文獻 37
表六、最近發表關於使用NIR檢測不同農產品中感染到黃麴菌的文獻 39
表七、農產品研究中有關黃麴毒素研究之波長整理 40
表八、農產品研究中有關受黃麴菌汙染研究之波長整理 42
表九、LC-MS/MS分析黃麴毒素B1之沖提（gradient）條件 53
表十、白米樣品中的黃麴毒素 67
表十一、所有白米樣品之實際黃麴毒素濃度（ppb） 68
表十二、對原始光譜之SVM預測結果 70
表十三、SVM預測所有含有黃麴毒素樣品之結果 71
表十六、樣品濃度在60 ppb以上MLR分析之結果（18個樣品） 74
表十七、樣品濃度在50 ppb以上MLR分析之結果（24個樣品） 76
表十八、樣品濃度在40 ppb以上MLR分析之結果（29個樣品） 79
表十九、樣品濃度在30 ppb以上MLR分析之結果（42個樣品） 79
表二十、樣品濃度在35 ppb以上MLR分析之結果（36個樣品） 80
表二十一、樣品濃度在35 ppb以下MLR分析之結果（35個樣品） 80
表二十二、黃麴毒素B1含量35 ppb以上白米樣品之分組情形 81
表二十三、黃麴毒素B1含量35 ppb以下白米樣品之分組情形 81

圖目次
圖一、105年每人每日糧食供給量 14
圖二、米之分類 15
圖三、產生黃麴毒素之菌種：(A)黃麴菌，(B)寄生麴菌 16
圖四、黃麴毒素B1、B2、G1、G2的化學結構式 18
圖五、近紅外光的吸收帶 24
圖六、單波分光器示意圖 26
圖七、二極體陣列光譜儀示意圖 27
圖八、濾波光譜儀示意圖 27
圖九、近紅外線光譜儀的採樣配置簡圖 28
圖十、稻米中黃麴毒素B1光譜圖 29
圖十一、人工類神經網絡示意圖 30
圖十二、用於光譜數據建模的多變量數據分析方法 32
圖十三、實驗步驟流程圖 49
圖十四、（A）FOSS NIRS 6500實驗室型分光光度計搭配樣本自動傳送配件（B）量測標準品使用之cuvette（C）量測白米使用之small ring cup 50
圖十五、光譜流程圖 51
圖十六、高效液相層析質譜測定黃麴毒素流程圖 54
圖十七、黃麴毒素B1標準品溶液之原始光譜 59
圖十八、不同濃度黃麴毒素B1標準品溶液之一次微分光譜 59
圖十九、不同濃度黃麴毒素B1標準品溶液之二次微分光譜 60
圖二十、黃麴毒素B1標準品之線性迴歸圖 60
圖二十一、所有白米樣品之原始光譜圖 61
圖二十二、所有白米樣品之一次微分光譜 61
圖二十三、受黃麴毒素汙染白米樣品之二次微分光譜 62
圖二十四、不同黃麴毒素B1標準品與白米樣品之層析圖 65
圖二十五、黃麴毒素B1標準品曲線 66
圖二十六、白米樣品中黃麴毒素濃度分布 66
圖二十七、測定白米中黃麴毒素B1時，理論值與實際值間之關係 67
圖二十八、黃麴毒素35 ppb以上校正組白米樣品之檢量線預測結果 82
圖二十九、黃麴毒素35 ppb以上驗證組白米樣品之檢量線預測結果 82
圖三十、黃麴毒素35 ppb以下校正組白米樣品之檢量線預測結果 83
圖三十一、黃麴毒素35 ppb以下驗證組白米樣品之檢量線預測結果 83

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