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研究生:林憶芳
研究生(外文):Yi-Fang Lin
論文名稱:毛細管電泳/雷射激發螢光分析人體血液中AminoglycosideAntibiotics含量之研究
論文名稱(外文):Determination of Aminoglycoside Antibiotics in Human Plasma by Capillary Electrophoresis with Laser-induced Fluorescence Detection
指導教授:張 玉 珍陳 政 男
指導教授(外文):Yu-Chen ChangYu-Chen Chang
學位類別:碩士
校院名稱:朝陽科技大學
系所名稱:應用化學系碩士班
學門:自然科學學門
學類:化學學類
論文種類:學術論文
論文出版年:2006
畢業學年度:94
語文別:中文
論文頁數:112
中文關鍵詞:毛細管電泳胺基配醣體類抗生素
外文關鍵詞:aminoglycoside antibioticscapillary electrophoresis
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Aminoglycoside Antibiotics 為一廣效性之抗生素,主要是用來治療經由革蘭氏陽性菌及革蘭氏陰性菌感染所引起之肺結核。由於Aminoglycoside Antibiotics 在化學療法中,通常伴隨著耳毒性及腎毒性等副作用,對耳朵會造成無法回復的聽力受損,所以該如何快速又有效地監控血液中Aminoglycoside Antibiotics 之濃度是很重要的。
本研究利用毛細管電泳搭配氣冷式氬離子雷射激發螢光偵測系統應用於Aminoglycoside Antibiotics 的分析,利用6-carboxyfluorescein succinimidyl
ester (CFSE) 將不具螢光性質的Aminoglycoside Antibiotics 進行管柱前衍生,胺基配醣體類抗生素的線性範圍少於三個級數( 0.01 ~ 5 μM,r 0.99≧ 94) ,偵測極限在nM 的範圍。結合簡單的血液樣品前處理,此分析方法成功地應用於血漿樣品中Aminoglycoside Antibiotics 之分析,在血漿樣品中Paromomycin、Tobramycin、Bekanamycin 及Kanamycin 的線性範圍為0.15~30 mM,r>0.9912,偵測極限分別為24 nM、17 nM、15 nM及14 nM。在確效試驗中,intra-day 和inter-day 的相對誤差分別小於3.7 及3.1%。回收率88~106%,其相對標準偏差皆落於9.1%內。
Aminoglycoside antibiotics is one kind of antibiotics which has been used
extensively. It could treat the phthisis, which is infected by gram-negative
bacilli and gram-positive bacilli. Aminoglycoside antibiotics usually causes
the side effect of ototoxicity and nephrotoxicity in clinical chemotherapy. It
will damage the hearing, and the damage is very difficult to be recovered.
Therefore, it is very important to control the concentration of aminoglycoside in
human blood.
In this research, capillary electrophoresis with laser-induced fluorescence
detection method was applied for the analysis of aminoglycoside antibiotics in
human plasma. 6-Carboxyfluorescein succinimidyl ester (CFSE) was used for
precolumn derivatization of aminoglycoside antibiotics which is nonfluorescent.
The linear ranges of aminoglycosides were less than 3 orders (0.01~5 μM,
r 0.9994≧ 94), and the limits of detection were 3.9~8.2 nM. Combining with the
simple pretreatment, the developed method was successfully applied to
determine the aminoglycosides in human plasma. The linear range of
Paromomycin、Tobramycin、Bekanamycin and Kanamycin in human plasma is
0.15~30 mM (r>0.9912), and the limits of detection were be 24 nM、17
nM、15 nM and 14 nM, respectively. The relative error of intra-day and
inter-day were less than 3.7 and 3.1%. The recoveries were 88~106%in
human plasma.
摘要 …………………………………………………………………………………………Ⅰ
Abstract………………………………………………………………………………………Ⅱ
誌謝 …………………………………………………………………………………………Ⅲ
目錄………………………………………………………………………………………… Ⅳ
圖目錄……………………………………………………………………………………… Ⅶ
表目錄……………………………………………………………………………………… Ⅸ
壹、緒論…………………………………………………………………………………… 1
一、前言…………………………………………………………………………………… 1
二、毛細管電泳之發展…………………………………………………………………… 2
三、毛細管電泳簡介……………………………………………………………………… 6
1 基本構造………………………………………………………………………………… 6
2 分離原理………………………………………………………………………………… 7
3 進樣方式………………………………………………………………………………… 10
4 偵測方式………………………………………………………………………………… 12
四、 樣品堆積…………………………………………………………………………… 15
1 等速電泳法……………………………………………………………………………… 15
2 電場放大樣品堆積法…………………………………………………………………… 18
V
3 巨大樣品體積堆積……………………………………………………………………… 18
4 清掃堆積法……………………………………………………………………………… 19
5 酸鹼調節堆積…………………………………………………………………………… 19
6 乙�城嚙n法……………………………………………………………………………… 20
五、簡介……………………………………………………………………………………21
1 藥理性質………………………………………………………………………………… 21
2 過去之分析方法………………………………………………………………………… 24
六、研究動機………………………………………………………………………………27
貳、儀器設備與實驗方法…………………………………………………………………28
一、藥品……………………………………………………………………………………28
二、儀器設備………………………………………………………………………………31
三、實驗方法………………………………………………………………………………33
1 毛細管處理……………………………………………………………………………… 33
2 電泳緩衝溶液配製……………………………………………………………………… 33
3 藥品配製………………………………………………………………………………… 33
四、血漿樣品配製…………………………………………………………………………34
五、衍生反應………………………………………………………………………………35
六、注射方式………………………………………………………………………………35
VI
七、毛細管電泳裝置………………………………………………………………………35
1 雷射激發螢光偵測裝置………………………………………………………………… 36
參、結果與討論……………………………………………………………………………39
一、偵測系統測試…………………………………………………………………………39
二、衍生條件之探討………………………………………………………………………41
1 衍生反應………………………………………………………………………………… 41
2 衍生試劑之探討………………………………………………………………………… 49
3 衍生緩衝溶液 pH 之探討……………………………………………………………… 51
4 衍生溫度之探討………………………………………………………………………… 51
5 衍生時間之探討………………………………………………………………………… 51
三、分離條件之探討………………………………………………………………………56
1 電泳緩衝溶液pH 值之探討………………………………………………………………60
2 電泳緩衝溶液濃度之探討……………………………………………………………… 60
3 分離電壓之探討………………………………………………………………………… 67
四、最佳分析條件………………………………………………………………………… 69
1 標準樣品中檢量線、再現性試驗……………………………………………………… 71
五、血漿樣品分析………………………………………………………………………… 74
六、血漿樣品前處理……………………………………………………………………… 77
VII
七、靈敏度之提升………………………………………………………………………… 83
八、確效試驗……………………………………………………………………………… 84
肆、結果與未來展望……………………………………………………………………… 95
一、結論…………………………………………………………………………………… 95
二、未來展望……………………………………………………………………………… 96
參考文獻…………………………………………………………………………………… 97
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