利用1-Naphthylacetic acid之可移除性衍生試劑分析膽固醇

Using 1-Naphthylacetic acid as a removable derivative reagent for cholesterol analysis

翁芝綾(Chih-Ling Weng)

指導教授 ： 吳信昇

高雄醫學大學/藥學院/藥學系碩士班/碩士(2015年)

摘要

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

關鍵字

膽固醇 ； 衍生試劑 ； HPLC-UV

並列摘要

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

並列關鍵字

Cholesterol ； Derived agent ； HPLC-UV

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