螢光化衍生及高效能液相層析法對血漿及製劑中有機酸藥物之分析

Fluorogenic Derivatization and Liquid Chromatographic Method for the Analysis of Organic Acid Drugs in Plasma and Pharmaceutical Preparations

林明君(Ming-Chun Lin)

指導教授 ： 吳信隆

高雄醫學大學/藥學院/藥學研究所/博士(2005年)

摘要

本研究之目的針對不具呈色基團之有機酸藥物，利用螢光導向衍生反應配合液相層析，建立了高感度及高選擇性分析法，並應用於製劑或血漿中chenodiol、ursodiol、valproic acid、undecylenic acid及zinc undecylenate之微量分析，對各有機酸分析法之重點歸納如下：

1. 膽汁酸藥物Chenodiol及ursodiol二種立體異構物之同時分析。以衍生試劑2-(2-naphthoxy)ethyl 2-(piperidino)ethanesulfonate (NOEPES)對chenodiol及ursodiol進行衍生，所得衍生物以HPLC螢光法進行分析並應用於該二種膽汁酸藥物市售製劑之含量分析。本法感度高又具選擇性，對chenodiol及ursodiol的定量範圍介於1.0–30.0 μM；其偵測極限對chenodiol及ursodiol分別為0.4和0.2 μM (S/N=3，注入體積為10 μL)。

2. 血漿中抗癲癇藥物valproic acid (VA)之分析。本方法是將VA從酸化的血漿中以甲苯萃取，所得甲苯萃取液直接以NOEPES衍生，結果產生之衍生物以高感度螢光偵測器 (激發波長為230 nm及發射波長為350 nm)分析，本法之定量範圍VA在10.0–200.0 μM，又其偵測極限為0.6 μM (S/N=3，注入體積為10 μL)。本法簡單、靈敏度高，應用於服用抗癲癇藥物VA病人血漿中濃度之監測，僅使用少量的檢品體積 (10-50 μL的血漿)，甚具實用性。

3. 抗黴菌藥物Undecylenic acid (UA)及zinc undecylenate (ZnUA)之同時分析。UA及ZnUA在18-crown-6和KHCO3的催化下，與NOEPES進行衍生，所得衍生物以高效能液相層析螢光法分析(激發波長為230 nm及發射波長為350 nm)，二種待測物的定量範圍皆在3.0–50.0 μM，又其偵測極限為0.3 μM (S/N=3，注入體積為10 μL)。本法主要發現在UA及ZnUA並存下，以氰甲烷可定量萃取UA，並應用於UA及ZnUA複方製劑之個別成份之含量分析。

關鍵字

微量分析 ； 衍生反應 ； 螢光法 ； 液相層析

並列摘要

Organic acids are widely found in biological and pharmaceutical samples. Many important organic acids are lack of chromophore in structure; therefore, they can not be directly analyzed at trace levels by absorption spectrophotometry. We have developed several methods for the sensitive analysis of bile acids, valproic acid, undecylenic acid and zinc undecylenate. The analytes were derivatized with 2-(2-naphthoxy)ethyl 2-(piperidino)ethanesulfonate (NOEPES) catalyzed by 18-crown-6 ether and potassium bicarbonate. The resulting derivatives were analyzed by isocratic HPLC with fluorimetric detection (excitation at 235 nm or 230 nm and emission at 350 nm). The main results of our study are summarized as follows:

1. Chenodiol and ursodiol are diastereomeric bile acids and widely used as anticholelithogenic. A sensitive and selective method was established for the simultaneous determination of chenodiol and ursodiol for the quality control of pharmaceutical preparations. The linear range for the analysis of the drugs was 1.0-30.0 μM with the detection limits (S/N=3, 10 μL injection) of 0.4 and 0.2 μM, respectively, for chenodiol and ursodiol. The method was demonstrated to the analysis of chenodiol in capsules and ursodiol in tablets.

2. A simple and sensitive method is described for the analysis of valproic acid in human plasma. The linear range for the analysis of the drugs was 10.0-200.0 μM with the detection limit of 0.6 μM (S/N=3, 10 μL injection). Toluene was used for the extraction of valproic acid from plasma and the toluene extract obtained was subjected to subsequent derivatization without solvent replacement. The simple method was applied to the analysis of valproic acid in plasma of patients using only small amount of sample (10-50 μL plasma).

3. A simple and selective approach is described for the analysis of undecylenic acid and zinc undecylenate in pharmaceutical preparations. The lower quantitation of the method is 3.0 μM of undecylenic acid with a detection limit of about 0.3 μM (S/N=3 with 10 μL injection). We found that acetonitrile is the selective solvent for differentially dissolving undecylenic acid from coexisted zinc undecylenate in compound formulation that makes the analysis of the analytes simple and accurate. Application of the method to the analysis of undecylenic acid and zinc undecylenate in ointment, powder, and solution preparations proved feasible.

並列關鍵字

Undecylenic acid ； Zinc undecylenate ； Ursodiol ； Valproic acid ； Fluorimetry ； HPLC ； Deivatization ； Chenodiol

參考文獻

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