Characterization of Uncoupling Effects in Bamboo Shoot Mitochondria

Abstract

本研究以兩種竹筍分別是Phyllostachys edulis (冬筍) 與Bambusa oldhamii (綠竹筍) 分離出粒線體為材料，研究脂肪酸的去偶合作用。 本實驗使用呼吸控制比值 (RC ratio) 評估粒線體的呼吸作用效率。對於P. edulis粒線體而言，未加任何脂肪酸的控制組的RC ratio為1.38，加了亞麻油酸 (LA) 或棕櫚酸 (PA) 後，RC ratio分別成為1.12 與 1.03。加了抑制劑oligomycin後，state 4 rate由18.71 nmol O2 mg min-1 protein-1減少為15.02 nmol O2 mg min-1 protein-1。加入去偶合劑FCCP後，state 4 rate由18.01 nmol O2 mg min-1 protein-1急速升高為31.68 nmol O2 mg min-1 protein-1。然而，當我們加入抑制alternative oxidase (AOX) 作用的抑制劑SHAM後，state 4 rate並沒有明顯的變化。 對於B. oldhamii粒線體而言，未加任何脂肪酸的控制組的RC ratio為1.08，加了LA或PA後，RC ratio分別成為1.18 與 1.12。加入相關的抑制劑後，state 4 rate並沒有明顯的變化。 對於P. edulis與B. oldhamii這兩種竹筍粒線體來說，在分離粒線體的過程中在備製溶液中加入BSA，所得到的粒線體呼吸速率皆會比分離過程中沒有加BSA得到的粒線體的呼吸速率高。在反應溶液中加BSA，LA或PA對於P. edulis粒線體所產生的去偶合作用會被抑制，不過這種現象並沒有在B. oldhamii粒線體中被發現。Bamboo shoot mitochondria were isolated from Phyllostachys edulis and Bambusa oldhamii to study the uncoupling effect of fatty acids and respiratory inhibitors on mitochondrial. This study characterized the phosphorylation and non-phosphorylation respirations. The respiratory control ratio (RC ratio) was determined to be an index of how coupled the mitochondria are. In P. edulis mitochondria, the RC ratios were 1.12 and 1.03 in the presence of linoleic acid (LA) and palmitic acid (PA) as compared to the control of 1.38. With addition of oligomycin, the non-phosphorylation respiration rate was decreased from 18.71 to 15.02 nmol O2 mg min-1 protein-1. With addition of FCCP, the non-phosphorylation respiration rate increased from 18.01 to 31.68 nmol O2 mg min-1 protein-1. However, the non-phosphorylation respiration rate did not significantly change when SHAM, inhibiting alternative oxidase (AOX), was added. In B. oldhamii mitochondria, the RC ratios were 1.18 and 1.12 in the presence of LA and PA as compared to the control of 1.08. There was no significant change in non-phosphorylation respiration with addition of respiratory inhibitors. In P. edulis and B. oldhamii mitochondria, isolated with BSA, the respiration rates were both higher than without BSA. With addition of BSA in the reaction buffer, the uncoupling effect of LA and PA on non-phosphorylation respiration rate of P. edulis mitochondria can be inhibited. However, there was no significant change in B. oldhamii mitochondria.中文摘要……………………………………………I 英文摘要……………………………………………II 目錄…………………………………………………III 圖表目錄……………………………………………IV 前言…………………………………………………1 材料與方法…………………………………………13 結果與討論…………………………………………16 結論…………………………………………………56 參考文獻……………………………………………57 附錄…………………………………………………6