本文選題：柑橘精油 + 代謝圖譜　； 參考：《西南大學(xué)》2014年博士論文

【摘要】：柑橘(Citrus L.)屬于蕓香科(Rutaceae)柑橘亞科(Aurantioideae)植物。中國(guó)是柑橘屬的重要起源中心之一,具有豐富的柑橘野生資源。自林奈1753年確立柑橘屬以來(lái),國(guó)內(nèi)外學(xué)者從形態(tài)到分子對(duì)柑橘屬植物的分類和進(jìn)化問(wèn)題開展了一系列研究。然而,柑橘屬植物的分類、進(jìn)化和重要栽培(雜)種的起源問(wèn)題仍待解決。柑橘精油是廣泛分布于柑橘的果實(shí)、葉片及花中的一類具芳香氣味且常溫?fù)]發(fā)的油狀液體的總稱。柑橘精油的主要成分d-檸檬烯具有抗氧化、抗炎、抗癌等多種生物活性。特別是最新研究發(fā)現(xiàn),柑橘精油的主要成分d-檸檬烯具有緩解由高脂膳食及N硝基左旋精氨酸甲酯(Nw-nitro-L-argininemethylester, L-NAME)誘導(dǎo)的非酒精性脂肪肝大鼠胰島素抵抗的功效。然而,有關(guān)d-檸檬烯對(duì)由高脂膳食誘導(dǎo)的肥胖小鼠的脂代謝紊亂和高血糖方面的研究仍尚未見報(bào)道。 本論文采用氣相色譜與質(zhì)譜技術(shù)對(duì)來(lái)自20種不同柑橘資源的果皮精油的揮發(fā)性成分進(jìn)行了分析,其中10種為中國(guó)特有野生柑橘類型。在柑橘精油代謝圖譜的基礎(chǔ)上,建立了以代謝組學(xué)為基礎(chǔ)的柑橘化學(xué)分類模型,并采用該分類模型對(duì)柑橘屬三個(gè)基本種以及六個(gè)重要栽培(雜)種的分類鑒定和起源問(wèn)題進(jìn)行了分析。在此基礎(chǔ)上,結(jié)合體外細(xì)胞模型以及體內(nèi)小鼠模型,對(duì)柑橘精油的主要成分d-檸檬烯在由高脂膳食誘導(dǎo)的肥胖小鼠脂代謝紊亂及高血糖等代謝綜合征中的防治作用進(jìn)行了系統(tǒng)研究。最后,采用雙熒光素酶報(bào)告基因和實(shí)時(shí)熒光定量PCR技術(shù)對(duì)d-檸檬烯在過(guò)氧化物酶增殖物激活受體(peroxisome proliferator-activated receptor, PPAR)以及肝X受體(liver X receptor, LXR)等核受體信號(hào)通路中的調(diào)節(jié)作用進(jìn)行了探討。 有關(guān)重要研究結(jié)果如下： 1.采用氣相色譜與質(zhì)譜技術(shù)對(duì)20份不同柑橘資源精油代謝圖譜進(jìn)行分析的結(jié)果表明：柑橘精油中的主要揮發(fā)性成分為單萜類物質(zhì),其中d-檸檬烯(55.05-91.06%)、(S)-(-)-苧烯(0.04-28.04%)、月桂烯(3.62-11.32%)、α-蒎烯(0.49-5.26%)以及香檜烯(0.07-7.54%)為柑橘精油的主要化學(xué)成分。對(duì)柑橘屬三個(gè)基本種果皮精油的代謝圖譜的分析結(jié)果表明：枸櫞(C.medica L.)精油的代謝圖譜以含有相對(duì)較低濃度的d-檸檬烯(55.05%)和較高濃度的(S)-(-)-苧烯(19。18%)、月桂烯(7.82%)以及α-蒎烯(3.67%)為主要特征；柚類(C. grandisOsb.)精油的代謝圖譜特征則為d-檸檬烯(85.67%)以及月桂烯(11.32%)的相對(duì)含量較高；寬皮柑橘(C. reticulata Blanco)的代謝圖譜特征為(S)-(-)-苧烯(11.39-21。83%)、α-芋烯崖柏烯(0.45-1.23%)以及α-蒎烯(2.04-4.07%)含量較高。 2.對(duì)柑橘精油代謝圖譜的聚類分析和偏最小二乘法辨別分析的結(jié)果顯示：枸櫞(C. medica L.)、柚(C. grandis Osb.)以及寬皮柑橘(C. reticulata Blanco)等柑橘屬三個(gè)基本種精油的代謝圖譜首先各自聚在一起,然后與其他基本種的代謝圖譜清楚地分離開來(lái)。此外,甜橙(C. sinensis Osb.)、酸橙(C.aurantium L.)、檸檬(C. limon Burm.f.)、粗檸檬(C. jambhiri Lush.)、蘭卜萊檬(C.limonia Osb)以及葡萄柚(C.paradisi Macf.)等重要栽培(雜)種的代謝圖譜首先各自聚在一起,然后分別與其親本或親本之一聚在一起,且聚類結(jié)果與已有形態(tài)學(xué)、DNA分子標(biāo)記和已有化學(xué)分類結(jié)果一致。D-檸檬烯、α-蒎烯、香檜烯與α-萜品烯為柑橘精油代謝圖譜中的特征性成分,是本論文中基于代謝組學(xué)的柑橘化學(xué)分類模型的主要生物標(biāo)記性行物質(zhì)或特征性化合物。 3.利用3T3-L1前脂肪細(xì)胞模型,通過(guò)前脂肪細(xì)胞誘導(dǎo)分化和油紅染色,我們發(fā)現(xiàn),柑橘精油的主要成分d-檸檬烯具有抑制前脂肪細(xì)胞分化的功效。采用C57BL/6小鼠進(jìn)行預(yù)防實(shí)驗(yàn)的結(jié)果表明,d-檸檬烯能夠明顯影響實(shí)驗(yàn)小鼠白色脂肪組織和褐色脂肪組織中脂肪細(xì)胞的大小,并能夠有效預(yù)防由高脂膳食誘導(dǎo)的實(shí)驗(yàn)小鼠血清脂質(zhì)中甘油三酯(p0.01)的上升以及空腹血糖水平(p0.01)的提高,d-檸檬烯還具有預(yù)防小鼠肝臟組織中脂肪積累的功效；治療實(shí)驗(yàn)中,通過(guò)建立高脂膳食誘導(dǎo)的肥胖小鼠模型,d-檸檬烯能夠顯著降低肥胖小鼠血清脂質(zhì)中甘油三酯(p0.01)、低密度脂蛋白膽固醇(p0.05)的含量以及空腹血糖水平(p0.05),并能有效提高肥胖小鼠血清中高密度脂蛋白膽固醇(p0.05)的含量以及小鼠對(duì)葡萄糖的利用率。 4.通過(guò)雙熒光素酶報(bào)告基因研究,我們發(fā)現(xiàn),柑橘d-檸檬烯可以提高PPARα的轉(zhuǎn)錄活性,并能降低LXRβ的表達(dá)水平。實(shí)時(shí)熒光定量PCR檢測(cè)結(jié)果表明,d-檸檬烯能夠顯著提高解偶聯(lián)蛋白(uncoupling protein-2, UCP-2)、乙酰輔酶A羧化酶(acetyl-CoAcarboxylase, ACC)以及過(guò)氧化物酶體增殖物激活受體γ輔激活子la (peroxisome proliferator-activated receptor y coactivator-la, PGC-1α)等PPARa靶基因的表達(dá)水平。此外,d-檸檬烯還能夠抑制膽固醇調(diào)節(jié)元件結(jié)合蛋白(sterol regulatory element-binding protein-1, SREBP-1),載脂蛋白E (apolipo-protein E, ApoE),以及3-羥基-3-甲基戊二酰輔酶A還原酶(3-hydroxy-3-methyl-glutaryl-CoAreductase, HMGR)等LXRP靶基因的mRNA的表達(dá)水平。
[Abstract]:Citrus L . belongs to the genus Aurantioideae of Rutaceae . China is one of the most important centers of citrus , and has rich citrus wild resources . The citrus essential oil is widely distributed in citrus fruits , leaves and flowers . The citrus essential oil is widely distributed in citrus fruits , leaves and flowers .

In this paper , the volatile components of citrus essential oil from 20 different citrus resources were analyzed by gas chromatography and mass spectrometry .

The key findings are as follows :

1 . The main volatile components of citrus essential oil were analyzed by gas chromatography and mass spectrometry . The results showed that the main volatile components of citrus essential oil were monoterpene , d - monene ( 55.05 - 91.06 % ) , myrcene ( 3.62 - 11.32 % ) , 偽 - pinene ( 0.49 - 5.26 % ) and sabinene ( 0.07 - 7.54 % ) as the main chemical constituents of citrus essential oil .
Grapefruit ( C . grandisOsb . ) The metabolic profiles of essential oils were d - monene ( 85.67 % ) and myrcene ( 11.32 % ) .
The metabolic profiles of C . reticulata Blanco were ( S ) - ( - ) - Limonene ( 11.39 - 21.83 % ) , 偽 - morphene ( 0.45 - 1.23 % ) and 偽 - pinene ( 2.04 - 4.07 % ) .

2 . The metabolic profiles of three essential oils , such as C . medica L . , C . grandis Osb . and C . reticulata Blanco , were isolated from the metabolic profiles of three essential oils , such as Citrus medica L . , C . grandis Osb . and C . reticulata Blanco . The metabolic profiles of the important cultivated ( hetero ) species are firstly brought together and then together with one of their parents or parents respectively , and the clustering results are consistent with the existing morphological , DNA molecular markers and the existing chemical classification results . D - Limonene , 偽 - pinene , sabinene and 偽 - terpineene are the characteristic components in the metabolic map of the citrus essential oil , and are the main biomarkers or characteristic compounds of the metabolic group - based citrus chemical classification model in this paper .

3 . Using 3T3 - L1 preadipocytes to induce differentiation and oil - red staining , we found that d - monene of citrus essential oil has the effect of inhibiting the differentiation of preadipocytes . The results showed that d - lemonene can significantly influence the size of fat cells in white adipose tissue and brown adipose tissue of experimental mice , and can effectively prevent the increase of triglyceride ( p0.01 ) and the increase of fasting blood glucose level ( p0.01 ) in the serum lipid of experimental mice induced by high fat diet .
In the treatment experiment , by establishing a model of obesity mice induced by high - fat diet , the content of triglyceride ( p0.01 ) , low - density lipoprotein cholesterol ( p . 05 ) and fasting blood glucose level ( p . 05 ) in the serum lipids of obese mice were significantly reduced by the establishment of a high - fat diet - induced obese mouse model , and the content of high - density lipoprotein cholesterol ( p . 05 ) and the utilization rate of glucose in the serum of obese mice can be effectively improved .

4 . Through the study of luciferase reporter gene , we found that d - lemonene can improve the transcriptional activity of PPAR偽 and decrease the level of expression of LXR 尾 . The results of real - time fluorescence quantitative PCR show that d - monene can significantly improve the expression level of PPARa target genes such as uncoupling protein - 1 , acetyl - CoAscent , ACC , and peroxidase - activated receptor y coactivator - la , PGCs - 1.alpha . , and the expression level of mRNA of LXRP target gene , such as 3 - hydroxy - 3 - methyl - yl - CoAreductase , HMGR , etc .
【學(xué)位授予單位】：西南大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2014
【分類號(hào)】：R151

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