臺灣博碩士論文加值系統

帕金森氏症是一種常見的神經退化性疾病，主要原因是因為黑質紋狀體的多巴胺神經元死亡導致多巴胺的缺少與氧化壓力及α-突觸核蛋白功能障礙。本論文擬從傳統中草藥山楂中尋找可以預防帕金森氏症的可能性，研究顯示山楂具有清除自由基與抗發炎活性。本論文的目的是探討山楂萃取物對於保護大鼠腎上腺髓質嗜鉻細胞瘤細胞對抗巴拉刈之毒性。首先測定山楂萃取物總多酚、總類黃酮、總單寧與粗三萜的含量及抗氧化能力。進一步將山楂95%乙醇萃取物進行分離與純化，並利用高效能液相層析儀比對萃取物與標準品。使用巴拉刈誘導大鼠腎上腺髓質嗜鉻細胞瘤細胞產生毒性，測定山楂萃取物對於巴拉刈誘導細胞毒性之保護效果與山楂萃取物減少乳酸脫氫酶濃度、活性氧產生和細胞內鈣離子增加，以及分析α-突觸核蛋白、Bcl-2、Bax與caspase 3蛋白表現量。研究結果提供實驗依據，做為評估山楂萃取物是否具有預防帕金森氏症的參考依據。

Parkinson's disease is one common neurodegenerative disease. Parkinson disease is loss of dopaminergic neurons in the substantia nigra leading to a dopamine deficit in the striatum as well as oxidative stress and α-synuclein dysfunction. In this study, we used a traditional Chinese medicine Crataegus pinnatifida as a possible remedy for Parkinson's disease. Many studies suggest that C. pinnatifida possesses free radical scavenging and anti-inflammatory activities. The purpose of this thesis is to explore whether the extracts from C. pinnatifida protect rat adrenal medulla pheochromocytoma cells against the paraquat toxicity. In the beginning, we measured the contents of total polyphenol, flavonoid, tannin, and crude triterpenoid and antioxidant capacities for the extracts from C. pinnatifida. The 95% ethanolic extract of the C. pinnatifida was further isolated and purified, and the extracts are then compared with the standards by high performance liquid chromatography. Using paraquat-induced rat adrenal medulla pheochromocytoma cells cytotoxicity to examine whether these extracts can protect rat adrenal medulla pheochromocytoma cells against the paraquat-induced cytotoxicity and reduce the lactate dehydrogenase concentration, reactive oxygen species production, and intracellular calcium increment. We also analyses the expression of protein α-synuclein, Bax, Bcl-2 and caspase 3. The results from this study provide an experimental basis in the prevention of Parkinson's disease.

中文摘要…………………………I
英文摘要…………………………II
目錄…………………………………IV
前言…………………………………1
一、帕金森氏症(Parkinson’s disease，PD) ………1
二、PD治療方法與藥物使用現況………………………3
三、巴拉刈(Paraquat，PQ) ………………………………5
四、中草藥-山楂(Crataegus pinnatifida) ……………6
五、PC12細胞株………………………………………………7
研究目的…………………………………9
實驗材料………………………………………10
實驗方法……………………………………14
一、山楂的萃取與分離純化方法……………………………14
二、萃取物總酚、總類黃酮、總單寧與粗三萜含量測定………16
三、抗氧化活性測定…………………………17
四、PC12 cells培養與保存……………………………………18
五、樣品保護作用機制之評估………………………………20
實驗結果……………………28
一、山楂95%乙醇萃取物與分層萃取物的產率及成分含量測定…………………………………………………28
二、山楂95%乙醇萃取物與分層萃取物的抗氧化能力…………29
三、山楂95%乙醇萃取物之分層萃取物與標準品比較………30
四、保護劑與傷害劑對於PC12細胞之毒性與保護作用………35
五、山楂95%乙醇萃取物與分層萃取物對於PQ誘導PC12細胞傷害之保護作用機制評估………36
實驗討論…………42
一、山楂95%乙醇萃取物與分層萃取物的產率及成分含量測定………………………………………42
二、山楂95%乙醇萃取物與分層萃取物的抗氧化能力………42
三、山楂95%乙醇萃取物之分層萃取物與標準品比較………42
四、山楂95%乙醇萃取物與分層萃取物對抗PQ誘導PC12細胞傷害之保護作用與對於PC12細胞存活率之影響………43
五、成分標準品對抗PQ誘導PC12細胞傷害之保護作用與對於PC12細胞存活率之影響…………………………………….. 43
六、山楂95%乙醇萃取物、分層萃取物與crataegolic acid對於PQ誘導PC12細胞傷害之保護作用機制……………… 44
七、山楂95%乙醇萃取物、分層萃取物與crataegolic acid對於PQ誘導PC12細胞內蛋白表現變化之影響…………45
八、山楂95%乙醇萃取物、分層萃取物與crataegolic acid對於PQ誘導PC12細胞傷害之細胞核染色實驗(Hoechst 33258染色)………46
九、綜合討論………46
結論………48
參考文獻………49

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