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研究生:翁芝綾
研究生(外文):Chih-Ling Weng
論文名稱:利用1-Naphthylacetic acid之可移除性衍生試劑分析膽固醇
論文名稱(外文):Using 1-Naphthylacetic acid as a removable derivative reagent for cholesterol analysis
指導教授:吳信昇吳信昇引用關係
指導教授(外文):Shihn-Sheng Wu
學位類別:碩士
校院名稱:高雄醫學大學
系所名稱:藥學系碩士班
學門:醫藥衛生學門
學類:藥學學類
論文種類:學術論文
論文出版年:2015
畢業學年度:103
語文別:中文
論文頁數:51
中文關鍵詞:膽固醇衍生試劑HPLC-UV
外文關鍵詞:CholesterolDerived agentHPLC-UV
相關次數:
  • 被引用被引用:0
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本研究的目的是開發1-Naphthylacetic acid (1-NAA) 為膽固醇衍生分析方法的衍生試劑,以取代原先的衍生試劑4-[N-methyl, N-(1-naphthymethyl)amino]-4-oxobutanoic acid (NAOB)。
膽固醇和1-NAA衍生物的高效能液相層析分析條件,使用XBridgeTM Phenyl (3.5 μm 4.6x150 mm) 管柱,移動相為Acetonitrile (ACN) : H2O (85:15, v/v),流速1.0 mL/min,樣品注入量10 μL,在UV 281 nm檢測。膽固醇和1-NAA之衍生物與2-Naphthyl myristate (internal standard) 的滯留時間依序分別為16.9和5.20分鐘。膽固醇和1-NAA之衍生物相當穩定,在室溫下其ACN水溶液濃度幾乎不變可達48個小時。
探討其最適衍生分析條件,反應時間為30分鐘,膽固醇比衍生試劑、催化劑和耦合試劑的莫耳濃度比依序分別為180,100和360。偵測極限為0.38 μg/mL,最低偵測濃度為1.24 μg/mL,其檢量線的範圍為1.50 ~ 300 μg/mL且相關係數同日內和異日間分別為1.000和0.999。將膽固醇添加入頭髮樣品中回收率大於97.3 %,顯示本分析方法有良好的可靠性。
關鍵字: 膽固醇、衍生試劑、HPLC-UV


The purpose of this study is to develop 1-naphthylacetic acid (1-NAA) as a derivative agent for cholesterol analysis, and hope it can become a substitute for previous derivative agent, (4-[N-methyl, N- (1-naphthymethyl) amino] -4-oxobutanoic acid (NAOB)), of our method.
The HPLC analytic conditions for derivative of cholesterol and 1-NAA as follows; a XBridgeTM Phenyl (3.5 μm 4.6x150 mm) column was used, mobile phase was composed of acetonitrile (ACN): H2O (85:15, v/v), flow rate was 1.0 mL/min, 10 μL sample solution was injected, and detection was set at UV 281 nm. Retention times of the derivative of cholesterol and 1-NAA and 2-naphthyl myristate (internal standard) were 16.9 and 5.20 minutes, respectively. The concentrations contained the derivative of cholesterol in aqueous ACN solution were almost stable at room temperature up to 48 hours.
Optimal derivatization conditions for cholesterol as following; reaction time was 30 minutes, the molar ratios of derivative agent, 1-NAA, catalytic agent, DMAP, and coupling reagent, EDC, to cholesterol were 180, 100 and 360, respectively. Limit of detection was 0.38 μg/mL, limit of quantification was 1.24 μg/mL, determination range was 1.50 ~ 300 μg/mL, and the correlation coefficients for intra-day and inter-day were 1.000 and 0.999 respectively. The recoveries of cholesterol spiked in hair sample were more than 97.3%, which indicating a good reliability of this method.
Keywords: Cholesterol, Derived agent, HPLC-UV

中文摘要 i
英文摘要 ii
目錄 iii
圖目錄 v
表目錄 vi
中英文縮寫對照表 vii
第壹章 研究背景 1
第貳章 實驗部分 4
第一節、實驗試藥與材料 4
第二節、儀器設備 6
第三節、試藥溶液之配置 7
第四節、膽固醇標準品衍生步驟 8
第五節、1-NAA與膽固醇之層析條件探討 8
第六節、實驗最適條件探討流程圖 11
第七節、最適化衍生條件之探討 12
1. 衍生反應中衍生試劑 (1-NAA) 需要量之探討 12
2. 衍生反應中催化試劑 (DMAP) 需要量之探討 13
3. 衍生反應中耦合試劑 (EDC) 需要量之探討 14
4. 反應系中反應時間之探討 15
5. 減壓抽氣時間之探討 16
6. 衍生物之安定性 16
7. 檢量線之建立及其精密度和準確度之測定 17
8. 同日內 (Intra-day) 18
9. 異日間分析 (Inter-day) 19
10. 方法應用及頭髮中膽固醇萃取回收率之探討 20
第參章 結果與討論 21
1. 1-NAA與膽固醇之層析條件探討 21
22
2. 最適化衍生條件之探討 23
(1) 衍生試劑需要量 23
(2) 催化試劑需要量 24
(3) 耦合試劑需要量 27
(4) 最適化反應時間 29
(5) 最適當之衍生條件 31
(6) 衍生物之安定性 33
3. 檢量線之建立及其精密度和準確度之測定 34
(1) 檢量線之建立 34
(2) 同日內與異日間分析之精密度與準確度 35
(3) 本方法應用及頭髮中膽固醇萃取之回收率 36
第肆章 結論 39
參考文獻 40


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