理學院: 化學研究所指導教授: 戴桓青施旻翰Shih, Min-HanMin-HanShih2017-03-022018-07-102017-03-022018-07-102016http://ntur.lib.ntu.edu.tw//handle/246246/272085生物體的胺基酸分析十分重要但同時也需要面對許多挑戰。生物樣品的複雜性與樣品內微量的胺基酸需仰賴高解析能力與高靈敏度的分析方法，在許多的分析方法中，鄰苯二醛（OPA）試劑的衍生化方法是最廣為人用的。該衍生化方法搭配高效能液相層析儀與螢光偵檢器的使用，可以有效地提高解析能力與靈敏度，達到分析生物樣品內胺基酸的目的。在我的研究中，鄰苯二醛衍生化方法更被進一步的優化及透過串聯質譜儀來提高定性的準確度，並且應用在研究思覺失調症（Schizophrenia）大鼠模型中藥物治療對大鼠腦部胺基酸與神經傳導物質的影響。 思覺失調症是一種極為盛行且嚴重的精神疾病，全球超過百分之一的人口皆深受其害。截至目前為止仍未發展出有效的治療方法，而這也是目前的首要議題。麩胺酸神經傳導系統中N-甲基-D-天門冬胺酸（NMDAR）受體功能低下是目前被廣泛研究於緩解思覺失調症之症狀的新興假說。根據此假說，藉由作用於N-甲基-D-天門冬胺酸受體上之甘胺酸調節位（GMS）的藥物，可以提升N-甲基-D-天門冬胺酸受體的功能並達到治療的效果。在臨床試驗上亦證實增加突觸間甘胺酸濃度將可以提高於甘胺酸調節位的作用，進而提升N-甲基-D-天門冬胺酸受體功能，因此甘胺酸被視為治療思覺失調症的潛力藥物。但是只有高劑量的甘胺酸才能通過血腦屏障進入腦中，而過高的劑量將會導致其他相關的副作用。因此，肌胺酸作為一種甘胺酸轉運體抑制劑，可以透過抑制甘胺酸的回收，以提高突觸間甘胺酸濃度至安全的範圍內，取代甘胺酸及其他直接作用於甘胺酸調節位之制效劑。 在此研究中，透過鄰苯二醛的衍生化方法，分析以肌胺酸注射後之大鼠的腦脊髓液中之甘胺酸濃度。在透過單一注射中等劑量的肌胺酸後，我們以抽取腦脊髓液的時間分成兩群不同組別，分別為 20 分鐘與80分鐘，與單一注射生理食鹽水的大鼠為控制組進行研究與分析。我們發現在單一注射肌胺酸80分鐘後，甘胺酸以及絲胺酸相較於控制組皆有顯著的濃度增加。同時，甘胺酸及D型絲胺酸於生物體內皆會作用於甘胺酸調節位，因此我們的研究結果可以提供強而有力的數據支持肌胺酸為有效治療思覺失調症的潛力藥物。Amino acids analysis in biological samples is a challenging task due to complex matrix and low concentrations of amino acids. The standard analytical technique for amino acids relies on derivatization with O-phthalaldehyde (OPA), followed by HPLC and fluorescence detection. In my thesis, this technique was applied to the study of amino acid neurotransmitters in rat model of schizophrenia. Schizophrenia is a pervasive and debilitating mental disorder that severely affects more than 1 % of the population worldwide. More effective therapeutic treatments of schizophrenia are greatly required. A new hypothesis, based on the hypofunction of the N-methyl-d-aspartate receptor (NMDAR) transmission, a subtype of glutamate receptors, has emerged as a promising target on developing alternative medications to ameliorate schizophrenia symptoms. Enhance the activity of NMDAR via glycine modulation site (GMS) can improve the schizophrenia symptoms. Clinical evidence suggests that increasing synaptic glycine levels at the GMS of NMDARs is a possible approach in the treatment of schizophrenia. Unfortunately, due to the poor penetration across the blood-brain barrier, a high dosage of glycine is needed, which leads to concomitant peripheral side effects. An alternative approach to increase GMS agonist availability is to block glycine reuptake by introducing sarcosine, a naturally occurring glycine transport inhibitor, which may increase extra-synaptic glycine levels in a relatively safe manner. In this study, glycine levels in cerebrospinal fluids (CSF) of sarcosine-treated rats were detected using the OPA method. After a single dose of sarcosine, we measured CSF amino acids at two different time points (20, 80 minutes), and compared with the control group that received a single dose of saline. We found that both glycine and serine became significantly elevated after 80 minutes. Since D-serine and glycine both bind to the GMS site on NMDAR in vivo, our data provided very strong evidence to support the therapeutic effects of sarcosine.2126345 bytesapplication/pdf論文公開時間: 2019/8/24論文使用權限: 同意有償授權(權利金給回饋學校)思覺失調症N-甲基-D-天門冬胺酸受體肌胺酸腦脊髓液胺基酸分析鄰苯二醛高效能液相層析儀螢光偵檢法質譜儀schizophreniaNMDARsarcosineCSFamino acids analysisOPAHPLCfluorescence detectionmass spectrometry[SDGs]SDG3thesis10.6342/NTU201602720http://ntur.lib.ntu.edu.tw/bitstream/246246/272085/1/ntu-105-R03223119-1.pdf