以毛細管電泳結合線上樣品前濃縮步驟進行兒茶酚胺類化合物之偵測

Determination of Catecholamines by On-line Sample Concentration Steps Coupled to Capillary Electrophoresis

劉鈺偉(Yuh-Woei Liu)

指導教授 ： 黃悉雅

中原大學/理學院/化學研究所/碩士(2011年)

https://doi.org/10.6840/cycu201100608

摘要

摘要
　　本論文利用毛細管電泳技術結合線上樣品濃縮步驟，偵測兒茶酚胺類（多巴胺、正腎上腺素與腎上腺素）化合物。
　　第一部分為利用紫外光偵測方式結合陽離子選擇性徹底注射（Cation-Selective Exhaustive Injection and Sweeping, CSEI-sweeping）的線上濃縮步驟。針對CE分離條件（微胞與磷酸鹽濃度）與線上前濃縮條件（樣品基質、酸段濃度、水段及酸段注射時間）做最佳化條件測試，最佳偵測極限為47.48 ~ 87.20 pg/mL，檢量線線性範圍為0.9994 ~ 0.9995（濃度範圍：1 ~ 100 ng/mL），濃縮倍率可達5 700 ~ 6 900倍。相較於文獻中只檢測多巴胺，此方法可同時偵測三種兒茶酚胺類化合物，且有較低的偵測極限。
　　第二部分應用在螢光偵測的系統上，使用fluorescein isothiocyanate（FITC）做為兒茶酚胺類化合物之螢光衍生劑，針對衍生條件（衍生劑濃度、衍生反應時間、衍生溶液組成與pH值）做最佳化測試。最佳偵測極限可達2.61 ~ 4.50 pg/mL（ppt）。為了達到更好的偵測感度，將螢光衍生後的兒茶酚胺類化合物進行線上樣品前濃縮，使用場放大樣品注射法（Field-Amplified Sample Stacking, FASS），針對線上前濃縮條件（樣品及水段注射時間、樣品基質濃度變化）做最佳化測試，最佳偵測極限為21.87 ~ 34.16 pg/L（ppq），線性範圍在0.9997 ~ 0.9999（濃度範圍：0.005 ~ 5 ng/mL），濃縮倍率可達114 ~ 132倍，相較於文獻，此方法有較低的偵測極限。將使用上述兩種方法運用在實際樣品的測試（生物樣品與人體尿液）。

關鍵字

毛細管電泳 ； 螢光 ； 兒茶酚胺

並列摘要

Abstract
In this study, on-line concentration-capillary electrophoresis was used to determine catecholamines (dopamine, norepinephrine, epinephrine). Different methods were employed namely, CE-UV and CE-LIF combined with cation-selective exhaustive injection and sweeping（CSEI-sweeping) and field-amplified sample stacking (FASS), respectively.
In the CE-UV coupled with CSEI-sweeping method, separation conditions such as micelle and phosphate concentrations and, on-line concentration conditions such as sample matrix and acid plug concentrations, water and acid plug length were optimized to find the suitable conditions for catecholamines determination. The obtained linearity of the calibration curve ranged from 0.9994 to 0.9995, and the detection limits（S/N=3) ranged from 47.48 to 87.20 pg/mL. All analytes were concentrated to about 5 700 to 6 900 fold by the optimal CSEI-CE/UV method.
　　On the other hand, in the fluorescence detection of catecholamines, derivatization conditions such as fluorescein isothiocyanate (FITC, derivatizing agent) concentration, reaction time, buffer and pH value were optimized to find the best conditions prior to analysis. The detection limits (S/N=3) obtained in the normal injection method ranged from 13.07 to 22.50 pg/mL. In order to achieve higher sensitivities, on-line sample concentration by FASS method for the determination of catecholamines after derivatization was employed. On-line concentration steps such as sample injection time, water plug length and sample matrix concentration were optimized to find the best conditions. The linearity of the analytes obtained were from 0.9997 to 0.9999, and the detection limits（S/N=3） ranged from 0.11 to 0.17 pg/mL. This method allowed all analytes to be concentrated to about 114- to 132-fold by the optimal FASS-CE/LIF method.
　　The above-mentioned two methods were used to test real samples such as biological samples and human urine.

並列關鍵字

capillary electrophoresis ； fluorescence ； catecholamine

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