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研究生:楊詠平
研究生(外文):Yung-Ping Yang
論文名稱:多點式葉片葉綠素螢光感測器之研發
論文名稱(外文):Development of a multipoint chlorophyll fluorescence sensor for plant leaves
指導教授:尤瓊琦
指導教授(外文):Chung-Chyi Yu
口試委員:黃振文洪滉祐
口試委員(外文):Jenn-Wen HuangHuaang-Youh Hurng
口試日期:2016-07-26
學位類別:碩士
校院名稱:國立中興大學
系所名稱:生物產業機電工程學系所
學門:工程學門
學類:機械工程學類
論文種類:學術論文
論文出版年:2016
畢業學年度:104
語文別:中文
論文頁數:53
中文關鍵詞:葉綠素螢光葉綠素濃度逆境
外文關鍵詞:chlorophyll fluorescencechlorophyll contentstress
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本研究旨在建立一具備多點葉綠素螢光量測與葉綠素濃度量測之低成本感測器,藉以降低使用葉綠素螢光量測之價格門檻及改善傳統單點葉綠素螢光無法代表葉片整體之缺點。試驗首先將建立一單點量測系統,並進行葉綠素螢光量測可行性試驗,量測昭和草(Crassocephalum crepidioides)經除草劑DCMU(3-(3′,4′-dichlorophenyl)-1,1-dimethylurea)處理前後與不同溫度下之螢光反應,藉以測試此系統量測之能力。試驗結果獲致葉片經除草劑DCMU處理後,其所量測之螢光淬滅量,將會大幅下降,並於處理後15分達到平衡;而葉片處於低溫環境與高溫環境下,其所量測之螢光淬滅量皆低於室溫之環境,且高溫亦對其造成不可逆之破壞;上述試驗結果皆與前人之研究有相符之處,同時亦顯示本試驗自製葉綠素螢光量測系統具葉綠素螢光量測之能力。
最適量測點數之模擬試驗,首先係利用自製之葉綠素螢光成像系統對各成長階段之小白菜(Brassica rapa chinensis)進行葉綠素螢光成像擷取,並利用影像處理程式對其進行適合實際感測器使用條件之模擬,模擬點數範圍為1~4點,以各點隨機量測所得之數據間離散程度與實際使用限制作為評估。試驗結果獲致增加量測點數可有效降低不同量測位置所造成量測間之差異,其中三點量測於實際應用上較為靈活,其限制遠小於四點量測,故本研究將以三點作為系統之量測點數,並以此結果製作一具備三點葉綠素螢光量測與葉綠素濃度之感測器。
試驗進一步將研究建置之多點式葉片葉綠素螢光量測系統與市售之儀器進行比較,其包含葉綠素濃度量測與手持葉綠素計(SPAD-502Plus, KONICA MINOLATA, Japan)及葉綠素螢光量測與市售葉綠素螢光儀(FMS1, Hansarech, USA)比較試驗,試驗結果顯示自製之葉綠素濃度量測與手持葉綠素計所量測之數值具有良好之相關性,其決定係數R2為0.997;而葉綠素螢光量測所得之螢光淬滅量與市售儀器量測參數ϕII於小白菜葉片經DCMU處理前後之反應皆有相同之趨勢,結果顯示自製之葉綠素螢光量測與濃度量測皆具有一定之可靠性。
試驗最終以研究所建置之多點式葉片葉綠素螢光感測器實際應用於量測小白菜葉片於缺水環境下之螢光反應,試驗結果獲致於缺水環境下之小白菜葉片其所量測之葉綠素螢光淬滅量將低於無水缺之正常葉片,而重新給予水分後,其螢光淬滅量則有一增加之現象,顯示本研究所建置之多點式葉片葉綠素螢光量測系統具備逆境造成不可逆傷害前即測得其螢光變化之能力,未來可提供相關業者進行葉菜類植物生長過程現象之應用評估。


The purpose of this study is to develop a low-cost, multipoint chlorophyll fluorescence sensor and relative chlorophyll content measuring system due to the prohibitive cost of using a chlorophyll fluorescence technique and the unreliability of traditional single-point chlorophyll fluorescence measurement. The experiment first established a single-point chlorophyll fluorescence measuring system, the feasibility of which was evaluated by measuring the chlorophyll fluorescence of the leaves (Crassocephalum crepidioides), which were treated by DCMU(3-(3′,4′-dichlorophenyl)-1,1-dimethylurea) and temperature stress. The results showed that (1) there was a significant decline of photochemical chlorophyll fluorescence quenching after treatment by DCMU and stable levels were reached after 15 minutes of decline. (2) Both high and low temperature stress of leaves can cause a drop in chlorophyll fluorescence quenching. According to the results above, our proposed chlorophyll fluorescence measuring system showed a superior performance.
Second, the simulation of the number of points in the multipoint chlorophyll fluorescence measuring system was established by using the image-processing techniques on chlorophyll fluorescence images of leaf (Brassica rapa chinensis) obtained by the proposed system we made. The coefficient of variation of random measuring data and working area were analyzed to determine the optimal number of measuring points. The simulation results showed that the number of optimal measuring points was three.
Third, the reliability of the proposed chlorophyll fluorescence and relative chlorophyll content measuring system we made was evaluated by comparing the obtained data with a commercial instrument. The result showed that there was a good correlation between the data of relative chlorophyll content or the proposed and commercial systems(SPAD-502Plus, KONICA MINOLATA, Japan); the coefficient of determination, R2, was 0.997. There was a similar trend between the chlorophyll fluorescence quenching measured by the proposed sensor and chlorophyll fluorescence parameter ϕII measured by a commercial instrument (FMS1, Hansarech, USA) after the leaves(Brassica rapa chinensis) have been treated by DCMU. The result displayed that the proposed sensor has good performance in terms of reliability.
Final, the proposed multipoint chlorophyll fluorescence sensor was actually applied to measure the chlorophyll fluorescence of leaves (Brassica rapa chinensis) under water deficit conditions. The result showed that the chlorophyll fluorescence quenching of leaves under water deficit conditions was less than that of the control leaves, and, after rewatering, it returned to the control level. The result demonstrated that the proposed sensor can detect the stress conditions in plants before irreversible damage is caused.



摘要 i
Abstract iii
目錄 v
圖目錄 viii
第一章 緒論 1
1-1 前言 1
1-2 研究目的 2
第二章 文獻探討 3
2-1 葉綠素螢光概述 3
2-2 常見葉綠素螢光研究之方法 6
2-2-1飽和脈衝法(saturation pulse method) 6
2-2-2快速葉綠素螢光誘導動力學分析OJIP-test 7
2-3葉綠素螢光研究之應用 8
2-4感測器製作相關條件 11
2-4-1激發光源與葉綠素螢光接收波長之選取 11
2-4-2感測元量測件位置之差異 12
2-4-3葉綠素濃度與葉綠素螢光 13
第三章 實驗材料與方法 14
3-1實驗流程設計 14
3-2試驗材料 16
3-3實驗設備 17
3-3-1自製葉綠素螢光量測系統 17
3-3-2自製之簡易葉綠素螢光成像系統 19
3-3-3植物生長箱 20
3-3-4葉綠素螢光儀 21
3-3-5手持葉綠素計 21
3-4實驗方法 22
3-4-1葉綠素螢光量測可行性實驗 22
3-4-2最適量測點數模擬實驗 23
3-4-3多點式葉片葉綠素螢光感測器之可靠性實驗 26
3-4-3-1葉綠素濃度量測與手持葉綠素計之比較 26
3-4-3-2葉綠素螢光量測與葉綠素螢光儀器之比較 26
3-4-4小白菜缺水環境實驗方法 27
第四章 結果與討論 28
4-1葉綠素螢光量測可行性試驗 28
4-1-1昭和草經除草劑DCMU處理前後之葉綠素螢光差異 28
4-1-2昭和草葉片於不同溫度下之葉綠素螢光差異 31
4-2最適量測點數模擬試驗結果 34
4-3多點式葉片葉綠素螢光感測器可靠性實驗 40
4-3-1葉綠素濃度量測與手持葉綠素計 40
4-3-2葉綠素螢光量測與市售儀器之比較 41
4-4小白菜缺水環境試驗結果 45
第五章 結論 47
第六章 參考文獻 50



圖目錄
圖2- 1葉綠體類囊體膜上之光合電子傳遞鏈 4
圖2- 2飽和脈衝法(Büchel & Wilhelm, 1993) 7
圖2- 3 JIP-TEST(Strasserf & Srivastava, 1995) 8
圖2- 4葉綠素螢光再吸收之現象(Buschmann, 2007) 12
圖3- 1實驗流程圖 15
圖3- 2小白菜 16
圖3- 3放大電路工作原理圖 18
圖3- 4 LCD螢幕顯示圖 19
圖3- 5自製之簡易葉綠素螢光成像系統 20
圖3- 6最適點數模擬條件示意圖 24
圖3-7單點與雙點可量測面積模擬成像 25
圖4- 1葉綠素螢光反應隨塗抹除草劑DCMU時間之變化 29
圖4- 2葉綠素螢光淬滅量隨塗抹除草劑DCMU時間之變化 30
圖4- 3最大葉綠素螢光值(Fm) 隨塗抹除草劑DCMU時間之變化 30
圖4- 4葉片於不同溫度之葉綠素螢光反應 32
圖4- 5葉綠素螢光淬滅量與溫度之關係 33
圖4- 6各成長階段之小白菜葉綠素螢光成像之R圖層 34
圖4- 7量測點數與變異係數之關係 36
圖4- 8量測點數與無法量測面積之關係 36
圖4- 9三點量測於葉片上量測之示意圖 37
圖4- 10小白菜發芽後14天之葉片螢光成像與三點模擬結果 38
圖4- 11四點量測於葉片上量測之示意圖 38
圖4- 12三點量測感測夾具之成品側視與上視圖 39
圖4- 13自製葉素濃度與市售儀器量測之關係 40
圖4- 14自製葉綠素螢光量測與市售儀器量測之比較(一) 41
圖4- 15自製葉綠素螢光量測與市售儀器量測之比較(二) 42
圖4- 16經DCMU處理後不同時間之螢光淬滅過程 44
圖4- 17缺水環境與控制組之螢光淬滅量與時間之關係 46



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