利用質譜儀建立阿斯匹靈之快速分析方法

Rapid determination of aspirin by mass spectrometer

劉璟樺(Jing-Hua Liu)

指導教授 ： 呂濟宇

高雄醫學大學/醫學院/生物化學研究所/碩士(2014年)

摘要

Aspirin有抗血小板及抗發炎之特性，其中主要是透過抑制COX-1活性而造成的，已有研究證實每天服用低劑量75-150 mg能預防心血管疾病及癌症，但仍有部分人會對aspirin產生抵抗性。 因此，本篇主要目的是利用基質輔助雷射脫附游離飛行時間質譜儀(MALDI-TOF-MS)之技術建立aspirin之快速分析方法，應用在藥品及血漿檢測。此技術與傳統液相層析儀器相比，具有靈敏度高、樣品需求量少、操作簡單及偵測時間極短之優勢。在藥劑分析上，添加鹼金屬化合物NaOH及基質2,5-dihydroxybenzoic acid (25DHB)，偵測下限最低可至5 μg/mL﹔而在血漿分析上則是利用化學衍生法，以8-(bromomethyl) quinoline (8-BrMQ)衍生試劑與aspirin進行反應，同時添加18-crown-6及K2CO3之催化劑，偵測濃度最低可至0.1 μg/mL，此兩種方法之相對標準偏差(relative standard deviation；R.S.D.) 和相對誤差(relative error；R.E.)皆小於10%。已知aspirin可誘發氧化壓力而造成癌細胞凋亡，本篇以aspirin培養大鼠腎小管上皮細胞(NRK-52E)，利用liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS)進行細胞分析，探討是否同樣會造成細胞凋亡。此外，以 aspirin合併有抗氧化作用的薑黃素(curcumin)，以及aspirin合併治療糖尿病之藥物格林齊特(gliclazide)，分別探討對細胞凋亡的影響。實驗結果發現aspirin會讓細胞產生氧化壓力以及細胞凋亡相關之蛋白，合併curcumin有減輕細胞凋亡的現象，而合併gliclazide則對細胞凋亡的影響並不明顯。

關鍵字

阿斯匹靈 ； 質譜儀

並列摘要

Aspirin has anti-platelet and anti-inflammatory characteristics. Primarily through inhibition of cyclooxygenase-1(COX-1) activity noted studies has demonstrated low-dose aspirin 70~150 mg/day can prevent from cardiovascular disease and certain cancer, but it still has aspirin resistance for some people. Therefore, the main purpose of this study is to use the matrix-assisted laser desorption ionization time of flight (MALDI-TOF-MS) technology to build a rapid method for analysis of aspirin in drug and plasma. Compared with conventional liquid chromatography instruments, the advantages of the MALDI-TOF-MS are high sensitivity, less sample required, easy to operate and detect in short time. In drug analysis, the addition of alkali metal compound such as NaOH and matrix 2,5-dihydroxybenzoic acid (25DHB), the detection limit of aspirin could be attained 5 μg/ml. In plasma analysis, 8-(bromomethyl) quinoline (8-BrMQ) was used to react with aspirin, and the reaction was catalyzed with 18-crown-6 and K2CO3. The detection limit of aspirin could be attained 0.1 μg/ml, relative standard deviation (R.S.D.) and relative error (R.E.) of the two methods are less than 10%.
Aspirin is known to induce oxidative stress caused apoptosis in cancer cell. Hence, this study used aspirin to treat rat renal tubular epithelial cells (NRK-52E) and screen the apoptosis-related protein by liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS). Moreover, combination of antioxidant curcumin and gliclazide, the effects on apoptosis also examine separately. It was found that aspirin could cause oxidative stress and express apoptosis-related proteins. Aspirin combind with curcumin could reduce the apoptosis of cells, combined with gliclazide is not obvious.

並列關鍵字

aspirin ； mass spectrometer

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