臺灣博碩士論文加值系統

本研究探討以高效能液相層析儀（HPLC）同時檢測食品中對羥苯甲酸（P-Hydroxy benzoic acid）、水楊酸（Salicylic Acid）、苯甲酸（Benzoic Acid）、己二烯酸（Sorbic Acid）、及去水醋酸（Dehydrate Acetic Acid）等五種防腐劑及糖精（Saccharin）、甜精（Dulcin）等兩種人工甜味劑之方法。固體樣品以水震盪萃取後過濾，液體樣品則直接以水稀釋過濾後注入HPLC，使用Intersil-ODS2-6mmx15mm 5μm層析管，用甲醇：氰甲烷：5mM檸檬酸緩衝溶液（15：15：70）為移動相，於流速1ml/min，UV225nm波長條件下，歷時20分鐘可同時檢出糖精、對羥苯甲酸、水楊酸、甜精、苯甲酸、己二烯酸、及去水醋酸。線性迴歸係數達r2=0.9994~0.9998，偵測極限（LOD）在訊號對雜訊比（S/N）=3為1~2ng。檢出限量（LOQ）在訊號對雜訊比（S/N）=10為1~2 mg/kg。添加適當混合標準溶液於固體及液體樣品中平均回收率達97.57%。十六件市售樣品，檢測結果有十一件固體樣品含防腐劑或人工甜味劑超過限量，液體樣品有三件檢出含防腐劑，但添加量皆在野i範圍內，其餘二件則未檢出含防腐劑或人工甜味劑。

This program probes into the method of simultaneously examining five preservatives:p-Hydroxy Benzoic Acid, Salicylic Acid, Benzoic Acid, Sorbic Acid, and Dehydrate Acetic Acid, and two artificial sweeteners: Saccharin and Dulcin by HPLC. The solid samples were shacked and extracted by water, and then filtered. The liquid samples were diluted with water and then filtered. After these procedures, they were injected into HPLC. The consistencies of p-Hydroxy Benzoic Acid, Salicylic Acid, Benzoic Acid, Sorbic Acid, Dehydrate Acetic Acid, Saccharin and Dulcin will be presented in twenty minutes by using a Intersil-ODS2-6mmx15mm 5um column, and the buffer solution of Methanol : Acetonitrile : 5mM citric Acid buffer (15:15:70) as mobile phase; under the condition of 1ml/min flow velocity, and 225nm wavelength for detection. The linear correlation factor reached r2=0.9994~0.9998. LOD is 1~2ng, under the condition of signal to noise ratio (S/N)=3. When S/N=10，LOQ is 1~2 mg/kg. The average recovery of adding appropriate standard solution into solid and liquid samples reaches 97.57%.
In total sixteen products tested by this method, there were eleven solid samples containing preservatives or artificial sweeteners, which were over the limit, and three liquid samples containing preservatives, but the additive quantities were within the allowable range, and the others contained no preservatives or artificial sweeteners.

目錄
摘要……………………………………………………………… i
英文摘要∙……………………………………………………… ii
目錄……………………………………………………………… iii
表目錄…………………………………………………………… v
圖目錄 ………………………………………………………… vii
第一章、緒論
一、 前言……………………………………………………… 1
二、 薄層層析法對防腐劑檢測……………………………… 4
三、 薄層層析法對人工甘味劑檢測………………………… 5
四、 氣相薄層層析法對防腐劑檢測………………………… 6
五、 液相層析儀對人工甘味劑及防腐劑檢測……………… 8
六、層析法及液相層析法 ……………………………………11
七、相關食品添加物性質 …………………………………19
第二章、實驗步驟及方法
一、材料及設備………………………………………………26
二、鑑別及定量分析…………………………………………30

iii
第三章、結果與討論
一、 各化合物紫外線吸收波長探討………………………… 33
二、 HPLC系統確立 ………………………………………… 36
三、標準曲線製作……………………………………………… 43
四、回收率試驗 ………………………………………………46
五、儀器偵測極限及檢出限量……………………………… 58
六、市購樣品檢測 ……………………………………………60
七、HPLC法與TLC、GC法比較……………………………61
八、應用HPLC-PDA鑑定 ……………………………………61
第四章、結論 …………………………………………………64
第五章、參考資料……………………………………………66
附件、食品添加物使用範圍及用量標準………………… 72

iv

表目錄
表一、各種防腐劑適用之展開溶媒、發色液及其斑點顏色 … 4
表二、氣相層析儀測定參考條件 ……………………………… 7
表三、層析法的分類 …………………………………………… 12
表四、正相層析與逆相層析之特性比較 ……………………… 15
表五、不同尺寸需求之管住尺寸 ……………………………… 16
表六、各種型態層析管及其官能基 …………………………… 17
表七、高效能液相層析在食品分析之應用 …………………… 18
表八、各種防腐劑及人工甜味劑之uv吸收波峰波長………… 33
表九、不同波長與波峰積分面積 ……………………………… 36
表十、甲醇/氰甲烷比例變化對滯留時間影響 ………………… 39
表十一、甲醇/氰甲烷比例變化對解析度（Rs）影響………… 39
表十二、檸檬酸緩衝溶液濃度對滯留時間影 …………………… 40
表十三、檸檬酸緩衝溶液濃度對解度（Rs）影響 …………… 40
表十四、檸檬酸緩衝溶液配比對滯留時間影響 ……………… 41
表十五、檸檬酸緩衝溶液配比對解析度（Rs）影響 ………… 41
表十六、緩衝溶液pH對滯留時間影響 ………………………… 42
表十七、緩衝溶液pH對解析度（Rs）影響……………………… 43
表十八、濃度（6.25ppm）與積分面積之關係 ………………… 43
v

表十九、濃度（12.5ppm）與積分面積之關係………………… 44
表二十、濃度（25ppm）與積分面積之關係…………………… 44
表二十一、濃度（50ppm）與積分面積之關係………………… 45
表二十二、濃度（100ppm）與積分面積之關係 ………………∙ 45
表二十三、濃度與積分面積之關係……………………………… 47
表二十四、重要導出量…………………………………………… 55
表二十五、固體樣品回收率試驗………………………………… 56
表二十六、液體樣品回收率試驗………………………………… 56
表二十七、市購樣品測試結果…………………………………… 60
表二十八、HPLC法與TLC、GC法比較 ……………………… 61

vi
圖目錄
圖一、糖精之UV吸收圖………………………………………… 34
圖二、對羥苯甲酸之UV吸收圖………………………………… 34
圖三、水楊酸之UV吸收圖……………………………………… 34
圖四、甜精之UV吸收圖………………………………………… 34圖五、苯甲酸之UV吸收圖……………………………………… 35
圖六、己二烯酸之UV吸收圖…………………………………… 35
圖七、去水醋酸之UV吸收圖…………………………………… 35
圖八、分離管以Inertsil 5 ODS 4.0mmX300mm之LC圖譜…… 47
圖九、分離管以Nucleosil 5 ODS 4.6mmX250mm之LC圖譜……∙ 47
圖十、分離管以Inertsil 5 ODS-3 4.0mmX300mm之LC圖譜…… 48
圖十一、移動相以甲醇：氰甲烷：1%醋酸之LC圖譜……………48
圖十二、移動相以甲醇：氰甲烷：1%磷酸之LC圖譜……………49
圖十三、分離管以Inertsil 5 ODS-2 6.0mmX150移動相以甲醇：
氰甲烷：5mM檸檬酸之LC圖譜圖……………………… 49
圖十四、滯留時間對甲醇與氰甲烷比例關係圖………………… 50
圖十五、滯留時間對緩衝溶液濃度關係圖……………………… 50
圖十六、滯留時間對緩衝溶液比例關係圖……………………… 51
圖十七、滯留時間對緩衝溶液pH關係圖………………………… 51
vii
圖十八、甜精之標準曲線圖……………………………………… 52
圖十九、對羥苯甲酸之標準曲線圖 ……………………………… 52
圖二十、水楊酸之標準曲線圖 ……………………………………53
圖二十一、甜精蜜去水醋酸之標準曲線圖……………………… 53
圖二十二、苯甲酸之標準曲線圖………………………………… 54
圖二十三、己二烯酸之標準曲線圖……………………………… 54
圖二十四、去水醋酸之標準曲線圖……………………………… 55
圖二十五、空白樣品之LC圖譜 ………………………………… 57
圖二十六、含己二烯酸樣品之LC圖譜 ………………………… 57
圖二十七、儀器偵測極限(LOD) 之LC圖譜 …………………… 59
圖二十八、方法檢出限量(LOQ) 之LC圖譜 …………………… 59圖二十九、以PDA(225nm)偵測之LC圖譜及紫外線掃瞄圖…… 62
圖三十、PDA之三度空間立體圖………………………………… 63

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