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超臨界流體萃取茵陳色原酮及抑制肝癌細胞生長的探討
No full textThis study examined the effect of Soxhlet solvent and Supercritical CO2 (SC-CO2) extracts of Artemisia capillaris T. (AC) and Artemisia annua L. (AA) on the growth inhibition of rat hepatic stellate cells (HSC) and two human hepatoma cells (Hep-G2 and HUH7). Most of extracts of Artemisia annua L and Artemisia capillaris T. present growth inhibitions on the HSC and on the Hep-G2 and HUH7 by a varied degree. Concentration ranged from 0.01 μg/ml to 70 μg/ml for HSC cells, concentration ranged from 15 μg/ml to 400 μg/ml for hepatoma cells. Further cancer cell treatments using three pure compounds revealed that capillarisin is relatively a significant growth inhibitor for HSC cells and for two human hepatoma cells. A two factor centercomposite scheme of Response Surface Methodology was then designed for supercritical CO2 added with ethylacetate extraction at 50℃. Pressure ranged from 2000 psig to 3000 psig and co-solvent ranged from 5 wt% to 10 wt%. The AC extracted of supercritical carbone dioxide added with 7.5 wt% ethyl acetate at 2500 psig and 50℃results in the hightest purity (39.52 wt%) of capillarisin and the best growth inhibition of two hepatoma cells. AC sample's IC50 values of growth inhibition of Hep-G2 and HUH7 cells are 72 ppm and 40 ppm, respectively, inuestrgated ender concentrations ranged from 15 μg/ml to 120 μg/ml.根據最近發表的文獻得知，青蒿和茵陳蒿內含的幾種指標成分能有效的抑制癌細胞與肝纖維化。本研究發現青蒿與茵陳蒿的萃出物，均能有效地抑制大白鼠肝纖維細胞(HSC)，且對兩株人類肝癌細胞HEP-G2、HUH7，呈現不同程度的生長抑制效果。對HSC肝纖維細胞，青蒿樣品濃度測試範圍0.01 μg/ml至60 μg/ml；茵陳蒿濃度範圍1 μg/ml至70 μg/ml。對兩株肝癌細胞，青蒿與茵陳蒿濃度範圍均由15 μg/ml至400 μg/ml。SPSS統計軟體的數據指出，茵陳蒿的各溶劑萃出物中綠原酸、香豆素、茵陳色原酮純度與對活化型態的兩株肝癌細胞的生長抑制，有直接的關連性。其中以茵陳色原酮的純度對肝癌細胞(Hep-G2 和HUH7)生長抑制的效果，相關性最為顯著。接著，以壓力與乙酸乙酯添加比率為應答曲面的實驗設計(RSM)，進行茵陳蒿超臨界提取物的色原酮純度，對癌細胞生長抑制率的探討(以半數存活有效濃度IC50值為應答值)。超臨界萃取的實驗數據指出，在2500 psig，50℃，7.5 wt%乙酸乙酯的添加，2小時萃出物中茵陳色原酮的純度可達39.52 wt%最大值，72小時萃出物可以有效地抑制兩株人類肝癌細胞的生長(Hep-G2的IC50值最低為72 μg/ml； HUH7的IC50值最低為40 μg/ml)，相當於95%純度的茵陳色原酮的效果。摘要……………………………………………………………..………………………2
ABSTRACT…………...………………………………………………………………. .3
目錄………………………………………………………………………………….…..4表目錄…………………………………………………………………………………...5
圖目錄 ………………………………………………………………………………….6
符號說明… ……………………….…………………………………………………… 8
序論…………………………………………………………………………………….11
第一節、茵陳蒿的介紹……………………………………………………………11
第二節、肝纖維化與肝癌的介紹………………………………………………….14
文獻回顧……………………………………………………………………………….18
第一節、茵陳蒿的分析及萃取方法……………………………………………...18
第二節、超臨界流體萃取的發展與應用………………………………………...20
研究動機……………………………………………………………………………….22
研究目的及方法……………………………………………………………………….23
實驗部分………………………………………………………………………………24
第一節、化學藥品與材料……………………………………………………..…24
第二節、儀器與設備………… ……………………………………………….....27
第三節、蒿類植物的樣品萃取…………………………………………………..34
第四節、樣品分析與數據計算……..……………………………………………39
第五節、細胞培養……………………..…………………………………………40
第六節、細胞抑制數據之計算……………………..……………………………43
第七節、應答曲面的實驗分析(RSM)...…………………………………………44
結果與討論……………………………………………………………………………46
結論……………………………………………………………………………………61
參考文獻………………………………………………………………………………64
表一、溶劑萃提茵陳蒿中綠原酸、香豆素與色原酮的最大回收率及純度數據比
較。…………………………….…..….…………………………………….....72
表二、乾的二氧化碳與添加乙酸乙酯的二氧化碳茵陳蒿超臨界萃出物中茵陳色
原酮的回收率(R)，純度(P)，總產率數據比較。.…….………………...……..73
表三、在50℃與2小時，添加乙酸乙酯超臨界二氧化碳萃取茵陳色原酮之雙變
數中心混層的應答曲面實驗設計數據。……………………………....….…74
表四、100 μg/ml青蒿及茵陳蒿樣品對HUH7 生長抑制率和純度的關係。….….…76
表五、100 μg/ml青蒿及茵陳蒿樣品對Hep-G2 生長抑制率和純度的關係。....….77
表六、100μg/ml青蒿及茵陳蒿樣品對BNL 抑制率和純度的關係。……..…….....78
表七、青蒿及茵陳蒿萃取樣品對大白鼠肝纖維細胞72小時的 IC50、IC90值。 ….79
表八、青蒿樣品處理48小時與72小時後肝癌細胞生長抑制的 IC50值(μg/ml).. 80
表九、100μg/ml茵陳蒿樣品處理48小時肝癌細胞IC50、IC90值(μg/ml)與三種
成份純度的關係。…………………………………………………..……...…81
表十、100μg/ml茵陳蒿樣品處理72小時肝癌細胞IC50、IC90值(μg/ml)與三種
成份純度的關係。…………………………………………………...…..……82
附表一、添加乙酸乙酯的近臨界二氧化碳操作條件與溶劑的臨界點。…………111
附表二、應答曲面模式ANOVA分析，超臨界壓力與共溶劑添加量對茵陳色原
酮純度的影響(設計點2500 psig , 7.5wt% EA added)。…………………..112
附表三、應答曲面ANOVA分析，超臨界壓力與共溶劑添加量對茵陳色原酮回
收率的影響(設計點2500 psig , 7.5wt% EA added) 。.…………….……..113
附表四、應答曲面模式分析，超臨界壓力與共溶劑添加量對總產率的影響. (設計點
2500 psig , 7.5wt% EA added)………………………………………….….114
附表五、應答曲面模式分析，超臨界壓力與共溶劑添加量對人類肝癌細胞
Hep-G2的影響。………………………..…………………………………115
附表六、應答曲面模式分析，超臨界壓力與共溶劑添加量對人類肝癌細胞
HUH7的影響。……………………….……………..……..………………116
附表7、以HPLC偵測茵陳蒿三主要成份於各萃出液中之線性範圍、線性方程
式、線性相關係數及偵測極限。…………….…………… ………………...117
圖 1、索式迴流萃取裝置簡圖。………...…………..…….........…………...………83
圖 2、超臨界萃取裝置圖。….....…………………...………………………………. 84
圖 3、茵陳蒿中主要成分結構式。………………..……………………………..... .85
圖 4、茵陳蒿各部份含量及其生化活性。………………………….………………86
圖 5、SEM下的青蒿和茵陳蒿。…………………………………………..…….…..87
圖 6、24小時，以200毫升(95% Ethyl Acetate+ 5% Ethanol)索式萃取10
克茵陳蒿花業種子中三成份萃出量和時間之關係。……....…………88
圖 7、24小時，以200毫升(95% Ethyl Acetate+ 5% Ethanol)索式萃取10
克茵陳蒿花業種子中三成份純度和時間之關係。……………….……88
圖 8、16小時，以200毫升(95% Ethyl Acetate+ 5% Ethanol)索式萃取10
克茵陳蒿四部份中各成份萃出量之比較。….……………….………….......89
圖 9、16小時，以200毫升(95% Ethyl Acetate+ 5% Ethanol)索式萃取10
克茵陳蒿四部份中各成份純度之比較。……..……………………………….89
圖10、添加5wt% EA超臨界二氧化碳萃取時，靜置時間影響茵陳色原酮純度及
回收率(2500 psigc和40℃, 4000 psigc和40℃ ) 。…………….……….……90
圖11、添加固定比例乙酸乙酯的二氧化碳萃取時，靜置時間影響茵陳色原酮純
度及回收率(在2500pisg和40℃) 。…………………...…………………...…90
圖12、添加5 wt%乙酸乙酯的二氧化碳，萃取時間影響茵陳色原酮純度(時間不
含洩壓時CO2的體積) 。……... .………………………………………...……91
圖13、超臨界二氧化碳萃取，溫度影響茵陳色原酮純度(乙酸乙酯5wt%)。…..….92
圖14、超臨界二氧化碳萃取，溫度影響茵陳色原酮回收率(乙酸乙酯5wt%)。…….92
圖15、乙酸乙酯5wt%的超臨界二氧化碳萃取，壓力影響茵陳色原酮純度。……...93
圖16、乙酸乙酯5wt%的超臨界二氧化碳萃取，壓力影響茵陳色原酮回收率。……93
圖17、2500 psig的超臨界二氧化碳萃取，溫度與乙酸乙酯的比例影響茵陳色原
酮純度。………………….………………………………………..………..…94
圖18、2500 psig的超臨界二氧化碳萃取，溫度與乙酸乙酯的比例比例影響茵陳
色原酮回收率。……….…………………………………………………...….94
圖19、茵陳蒿萃出樣品對HUH7處理48小時的生長抑制。………………………..95
圖20、茵陳蒿萃出樣品對HUH7處理72小時的生長抑制。………………………..95
圖21、茵陳蒿萃出樣品對Hep-G2處理48小時的生長制。……………………….….96
圖22、茵陳蒿萃出樣品對Hep-G2處理72小時的生長抑制……………..………….96
圖22、茵陳蒿萃出樣品對Hep-G2處理72小時的生長抑制。……………..……….97
圖24、茵陳蒿萃出樣品對BNL處理72小時的生長抑制。………………………… 97
圖25、茵陳蒿超臨界萃出樣品對HUH7處理48小時的生長抑制。…………………98
圖26、茵陳蒿超臨界萃出樣品對HUH7處理72小時的生長抑制。…………………98
圖27、茵陳蒿超臨界萃出樣品對BNL處理48小時的生長抑制……………………99
圖28、茵陳蒿超臨界萃出樣品對BNL處理72小時的生長抑制。…………………99
圖29、茵陳蒿超臨界萃出樣品對Hep-G2處理48小時的生長抑制。……………100
圖30、茵陳蒿超臨界萃出樣品對Hep-G2處理72小時的生長抑制。……………100
圖31、七種青蒿樣品對HUH7處理48小時的生長抑制。………………………101
圖32、七種青蒿樣品對HUH7處理72小時的生長抑制。……………………..…101
圖33、七種青蒿樣品對HPE G2處理48小時的生長抑制。…………………..…102
圖34、七種青蒿樣品對Hep-G2處理72小時的生長抑制。………………………102
圖35、七種青蒿樣品對BNL處理48小時的生長抑制。…………………………103
圖36、七種青蒿樣品對BNL處理72小時的生長抑制。…………………………103
圖37、100μg/ml青蒿與茵陳蒿樣品處理48和72小時，HUH7的生長抑制比較.104
圖38、100μg/ml青蒿與茵陳蒿樣品處理48和72小時，Hep-G2生長抑制比較.105
圖39、100μg/ml青蒿與茵陳蒿樣品處理48和72小時，BNL生長抑制比較。…105
圖40、超臨界二氧化碳萃取,茵陳色原酮純度與變數因子之應答曲面。…………106
圖41、超臨界二氧化碳萃取製程,茵陳色原酮回收率與變數因子之應答曲面。…107
圖42、超臨界二氧化碳萃取製程,總產率與變數因子之應答曲面。……………..108
圖43、50℃與2小時的超臨界二氧化碳萃取製程,培養72小時人類肝癌細胞
Hep-G2 生長抑制IC50值與變數因子之應答曲面。………………...……109
圖44、50℃與2小時的超臨界二氧化碳萃取製程,人類肝癌細胞HUH7 生長抑
制IC50值與變數因子之應答曲面。…………………………………….…110
附圖1、(A)HPLC的層析圖譜(λ=280nm)；(B)HPLC定義分析茵陳蒿成分的溶
媒梯度。…………………………...………………………………………118
附圖2、HPLC 層析茵陳蒿三成分溶於甲醇基質溶液之檢量線。…...……..……119
附圖3、茵陳蒿三成分溶於三種基質溶液的檢量線。…………………………….12
Going Beyond Counting First Authors in Author Co-citation Analysis
Full text linkThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
Variations on the Author
Full text link“Variations on the Author” discusses two of Eduardo Coutinho’s recent films (Um Dia na Vida, from 2010, and Últimas Conversas, posthumously released in 2015) and their contribution to the general question of documentary authorship. The director’s filmography is characterized by a consistent yet self-effacing form of authorial self-inscription: Coutinho often features as an interviewer that rather than express opinions propels discourses; an interviewer that is good at listening. This mode of self-inscription characterizes him as an author who is not expressive but who is nonetheless markedly present on the screen. In Um Dia na Vida, however, Coutinho is completely absent form the image, while Últimas Conversas, on the contrary, includes a confessional prologue that moves the director from the margins to the center of his films. This article examines the ways in which these works stand out in the filmography of a director who offers new insights into the notion of cinematic authorship
Appropriate Similarity Measures for Author Cocitation Analysis
Full text linkWe provide a number of new insights into the methodological discussion about author cocitation analysis. We first argue that the use of the Pearson correlation for measuring the similarity between authors’ cocitation profiles is not very satisfactory. We then discuss what kind of similarity measures may be used as an alternative to the Pearson correlation. We consider three similarity measures in particular. One is the well-known cosine. The other two similarity measures have not been used before in the bibliometric literature. Finally, we show by means of an example that our findings have a high practical relevance.information science;Pearson correlation;cosine;similarity measure;author cocitation analysis
Dispelling the Myths Behind First-author Citation Counts
Full text linkWe conducted a full-scale evaluative citation analysis study of scholars in the XML research field to explore just how different from each other author rankings resulting from different citation counting methods actually are, and to demonstrate the capability of emerging data and tools on the Web in supporting more realistic citation counting methods. Our results contest some common arguments for the continued
use of first-author citation counts in the evaluation of scholars, such as high correlations between author rankings by first-author citation counts and other citation
counting methods, and high costs of using more realistic citation counting methods that are not well-supported by the ISI databases. It is argued that increasingly available digital full text research papers make it possible for citation analysis studies to go beyond what the ISI databases have directly supported and to employ more
sophisticated methods
koamabayili/VECTRON-author-checklist: VECTRON author checklist
No full textWe have done our best to complete the author checklist relating to the use of animals in the hut study. Note that the objective for the hut study was to evaluate the IRS treatment applications for residual efficacy against Anopheles mosquitoes, including the local An. coluzzii mosquito population. Cows were only used to attract mosquitoes into the huts and no tests were carried out directly on the cows. The author checklist is intended for use with studies where experiments are carried out on animals, which is why we have had such difficulty in completing this for the hut study, as many of the questions do not relate to how the cows were used
Author-wise bibliometric analysis based on entropy.
No full textAuthor-wise bibliometric analysis based on entropy.</p
Author Under Sail The Imagination of Jack London, 1893-1902
No full textIn Author Under Sail, Jay Williams offers the first complete literary biography of Jack London as a professional writer engaged in the labor of writing. It examines the authorial imagination in London's work, the use of imagination in both his fiction and nonfiction, and the ways he defined imagination in the creative process in his business dealings with his publishers, editors, and agents. In this first volume of a two-volume biography, Williams traverses the years 1893 to 1902, from London's "Story of a Typhoon" to The People of the Abyss. The Jack London who emerges in the pages of Author Under Sail is a writer whose partnership with publishers, most notably his productive alliance with George Brett of Macmillan, was one of the most formative in American literary history. London pioneered many author models during the heyday of realism and naturalism, blurring the boundaries of these popular genres by focusing on absorption and theatricality and the representation of the seen and unseen. London created an impassioned, sincere, and extremely personal realism unlike that of other American writers of the time. Author Under Sail is a literary tour de force that reveals the full range of London as writer, creative citizen, and entrepreneur at the same time it sheds light on the maverick side of machine-age literature.Intro -- Title Page -- Copyright Page -- Dedication -- Contents -- Acknowledgments -- Introduction -- 1. Spirit Truth -- 2. From Absorption to Theatricality and Back Again -- 3. "I Will Build a New Present" -- 4. Sons as Authors -- 5. Fathers as Publishers -- 6. The Daughter as Author -- 7. Lovers as Authors -- 8. At Sea with the Family -- 9. Yellow News, Yellow Stories -- 10. The Return Home -- Notes -- Bibliography -- Index -- About Jay WilliamsIn Author Under Sail, Jay Williams offers the first complete literary biography of Jack London as a professional writer engaged in the labor of writing. It examines the authorial imagination in London's work, the use of imagination in both his fiction and nonfiction, and the ways he defined imagination in the creative process in his business dealings with his publishers, editors, and agents. In this first volume of a two-volume biography, Williams traverses the years 1893 to 1902, from London's "Story of a Typhoon" to The People of the Abyss. The Jack London who emerges in the pages of Author Under Sail is a writer whose partnership with publishers, most notably his productive alliance with George Brett of Macmillan, was one of the most formative in American literary history. London pioneered many author models during the heyday of realism and naturalism, blurring the boundaries of these popular genres by focusing on absorption and theatricality and the representation of the seen and unseen. London created an impassioned, sincere, and extremely personal realism unlike that of other American writers of the time. Author Under Sail is a literary tour de force that reveals the full range of London as writer, creative citizen, and entrepreneur at the same time it sheds light on the maverick side of machine-age literature.Description based on publisher supplied metadata and other sources.Electronic reproduction. Ann Arbor, Michigan : ProQuest Ebook Central, YYYY. Available via World Wide Web. Access may be limited to ProQuest Ebook Central affiliated libraries
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