臺灣博碩士論文加值系統

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gabapentin的分析方法已被開發完成，利用6-carboxyfluorescein succinimidyl ester (CFSE) 將非螢光性質的gabapentin進行管柱前衍生，在水溶液中，gabapentin的線性範圍可達3個級數，偵測極限為3.7 nM，其遷移時間和訊號面積精密度分別為0.9%、4.5%。結合簡單的樣品前處理，此分析方法成功的應用於血漿樣品中gabapentin分析，在血漿樣品中gabapentin的線性範圍0.03mM-30mM，r = 0.9996，偵測極限為4.8 nM。在確效試驗中，添加gabapentin於血漿樣品中所得的重現性相對標準誤差皆小於10.8%。
另外還進行毛細管電泳搭配雷射激發螢光偵測分析Ibuprofen的方法已被開發出來。將非螢光性質的Ibuprofen以5-AMF進行管柱前衍生，使之帶有螢光。這步驟中Ibuprofen先以hydroxybenzotriazol (HOBT) 和 diisopropylcarbodiimide (DIC)活化後，再添加溶於dimethylformamide (DMF)的5-AMF於室溫下進行衍生。所形成的衍生物利用MEKC分離，再藉由Ar+雷射激發螢光來偵測。其濃度線性範圍可達3個級數，偵測極限在10-9 M範圍，衍生極限可達4×10-7 M，遷移時間和訊號面積的精密度分別為0.23%、7.3%。此方法成功應用於市售藥品中的Ibuprofen分析。

A capillary electrophoresis method with air-cooled argon-ion laser-induced fluorescence detection for gabapentin has been developed. 6-Carboxyfluorescein succinimidyl ester was used for precolumn derivatization of the non-fluorescent drug. Linearity over three orders of magnitudes was generally obtained. The limit of detection for gabapentin in aqueous solution was calculated to be 3.7 nM. The relative standard deviation values on peak size and migration time were 4.5% and 0.9%, respectively. Combined with a simple cleanup procedure, this method can be applied to the determination of gabapentin in human plasma. The ratio of peak area of gabapentin / PPA versus gabapentin，s molar concentration plot showed a good linearity with a correlation coefficient (r) of 0.9996 and the limit of detection of gabapentin in plasma was about 4.8 nM. The Within-Day and Day to Day reproducibilities of gabapentin in plasma were less than 10.8%。
A capillary electrophoresis laser-induced fluorescence detection method is described for the determination of ibuprofen. 5-(Aminomethyl) fluorescein was used for precolumn derivatization of the non-fluorescent drug.
In the derivatization procedure, the ibuprofen was activated by hydroxybenzotriazol (HOBT) and diisopropylcarbodiimide (DIC), then reacted with 5-(aminomethyl) fluorescein in dimethylformamide at ambient temperature. The conjugate was separated by MEKC and detected by a Ar ion laser-induced fluorescence detector. Linearity of three orders of magnitudes was obtained. The concentration limit of detection was in 1.5×10-9 M range. A derivatization limit of detection of 4×10-7 M was achieved for ibuprofen. Precisions of migration times and peak areas were 0.23% and 7.3% respectively. The method was successfully applied to the analysis of ibuprofen in pharmaceuticals.

目錄

中文摘要 I
英文摘要 II
誌謝 IV
目錄 V
圖目錄 IX
表目錄 XI


第一章 緒論 1
1.1前言 1
1.2毛細管電泳的發展歷程 2
1.3毛細管電泳簡介 5
1.3.1基本構造 5
1.3.2分離原理 6
1.3.3進樣方式 9
1.3.4偵測方式 11
1.3.5樣品堆積 15
1.4 Gabapentin簡介 17
1.4.1藥理性質 17
1.4.2分析方法 18
1.4.3研究動機 20
1.5 Ibuprofen簡介 21
1.5.1藥理性質 21
1.5.2分析方法 22
1.5.3研究方向 23


第二章 儀器設備與實驗方法 25
2.1藥品 25
2.2儀器設備 27
2.3 Gabapentin實驗方法 29
2.3.1電泳緩衝溶液的配製 29
2.3.2藥品的配製 29
2.3.3血漿樣品配製 29
2.3.4衍生反應 30
2.3.5毛細管處理 30
2.3.6注射方式 30
2.3.7毛細管電泳(Capillary Electrophoresis, CE)裝置 31
2.3.8雷射激發螢光（Laser Induced Fluorescence, LIF）偵測裝置 31
2.4 Ibuprofen實驗方法 35
2.4.1電泳緩衝溶液的配製 35
2.4.2藥品的配製 35
2.4.3市售樣品的配製 36
2.4.4衍生反應 36
2.4.5注射方式 36


第三章 Gabapentin的結果與討論 37
3.1標準溶液分析 37
3.1.1衍生反應 37
3.1.2最佳分析、衍生條件 38
3.1.3檢量線、衍生極限及再現性試驗 42
3.2血漿樣品的分析 46
3.3靈敏度的提升 50
3.4確效試驗 59


第四章 Ibuprofen的結果與討論 68
4.1衍生條件最佳化 68
4.1.1衍生劑的影響 68
4.1.2 DIC的影響 68
4.1.3 HOBT的影響 71
4.1.4 DIPEA的角色 71
4.1.5衍生時間探討 71
4.1.6 5-AMF-Ibuprofen的分離 74
4.1.7檢量線、再現性測試及偵測極限探討 76
4.1.8衍生極限探討 78
4.2市售藥物的分析 81


第五章 結論與未來展望 84
5.1結論 84
5.1.1 Gabapentin結論 84
5.1.2 Ibuprofen結論 84
5.2未來展望 86
參考文獻 87

圖目錄

圖(一)毛細管電泳儀器裝置圖 5
圖(二)Gabapentin與GABA結構圖 18
圖(三)Ibuprofen的結構 21
圖(四)雷射激發螢光偵測器細部元件構造 33
圖(五)毛細管電泳/雷射激發螢光偵測器裝置圖 34
圖(六)CFSE與Gabapentin的衍生反應圖 39
圖(七)CFSE與水的衍生反應圖 40
圖(八)衍生時間對於Gabapentin訊號面積的影響 41
圖(九)CFSE-Gabapentin衍生產物電泳圖 43
圖(十)CFSE-Gabapentin衍生產物檢量線 44
圖(十一)不同比例ACN去除血漿樣品中蛋白質的電泳圖 47
圖(十二)Gabapentin的血漿樣品電泳圖 49
圖(十三)血漿樣品中不同注射秒數的電泳圖(1) 51
圖(十四)血漿樣品中不同注射秒數的電泳圖(2) 52
圖(十五)注射時間與訊號高度的關係圖 53
圖(十六)注射時間與訊號面積的關係圖 54
圖(十七)注射時間與理論板數的關係圖 55
圖(十八)注射60秒的Gabapentin血漿樣品電泳圖 58
圖(十九)Gabapentin的血漿樣品電泳圖(1) 62
圖(二十)Gabapentin的血漿樣品電泳圖(2) 63
圖(二十一)Gabapentin的血漿樣品電泳圖(3) 64
圖(二十二)Gabapentin的血漿樣品電泳圖(4) 65
圖(二十三)Gabapentin的血漿樣品電泳圖(5) 66
圖(二十四)Gabapentin的血漿樣品電泳圖(6) 67
圖(二十六)5-AMF濃度對於分析物訊號面積的影響 70
圖(二十七)DIC濃度對於分析物訊號面積的影響 72
圖(二十八)HOBT濃度對於分析物訊號面積的影響 73
圖(二十九)衍生時間對於分析物訊號面積的影響 75
圖(三十)Ibuprofen之電泳圖 77
圖(三十一)Ibuprofen的濃度檢量線 79
圖(三十二)市售樣品分析 82


表目錄

表(一)CFSE-Gabapentin衍生產物訊號重現性 45
表(二)血漿樣品檢量線與偵測極限的數據 56
表(三)血漿中Gabapentin的Within-Day與Day to Day確效試驗的數據 60
表(四)血漿樣品前處理回收率試驗 61
表(五)市售藥品之含量 83

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