本文關(guān)鍵詞： Pulicaria iuloides 精油 抗氧化 抗菌 抗糖尿病 抗癌　出處：《江南大學(xué)》2017年博士論文　論文類型：學(xué)位論文

【摘要】：Pulicaria inuloides是Asteraceae科Pulicaria屬植物,它被廣泛用作抗癌藥物,并用于治療糖尿病、風(fēng)濕、痛風(fēng)、膀胱炎、腎炎、痛經(jīng)、牙痛、頭痛等,并具有抗菌,抗凝血等功效。P.inuloides含有生物堿、揮發(fā)油、萜類、酚酸、類黃酮、單寧酸等成分。第一階段,從藥用芳香植物Pulicaria inuloides-Astereceae和Ocimum forskoleiLamaiceae葉子中得到精油,從精油中分離并鑒定生物活性物質(zhì)。然后測(cè)定分離出的物質(zhì)的抗氧化活性、總酚含量和抗微生物活性。結(jié)果表明與O.forskolei相比,P.inuloides有較高的總酚量(O.forskolei為54.6±30 mg GAE/g提取物,P.inuloides為144±5.32 mg GAE/g提取物);P.inuloides具有較高的抗氧化活性,DPPH清除率分別為26.76±0.11%(O.forskolei)和92.92±0.10%(P.inuloides),IC50分別為73.03±0.05(O.forskolei)和4.5±0.05(P.inuloides);P.inuloides有更高的β胡蘿卜素漂白值:分別為41.03±6.35%(O.forskolei)和90.77±0.21%(P.inuloides)。P.inuloides的精油能夠抑制除鼠傷寒沙門氏菌和志賀氏痢疾桿菌以外的所有測(cè)試微生物,并且對(duì)大腸桿菌的最小抑制濃度(MIC)為3.0μg/m L。O.forskolei精油僅抑制白色念珠菌。第二階段,通過(guò)用氣相色譜-質(zhì)譜聯(lián)用法鑒定P.inuloides和O.forskolei根中的揮發(fā)性化合物結(jié)果表明,與O.forskolei相比,P.inuloides根中含有較高的總酚含量(P.inuloides為55.4±0.1,O.forskolei為35.3±0.2mg GAE/g提取物);P.inuloides具有較低的抗氧化活性,DPPH清除率分別為23.07±0.06%(P.inuloides)43.97±0.18%(O.forskolei),IC50值分別為80.01±0.03(P.inuloides)和31.18±0.01(O.forskolei);P.inuloides有較低的β胡蘿卜素漂白值:分別25.3±0.3%(P.inuloides)和39.9±0.7%(O.forskolei)。P.inuloides精油能抑制除鼠傷寒沙門氏菌,痢疾桿菌和大腸桿菌以外的所有測(cè)試微生物。在第三階段,研究溶劑類型對(duì)Pulicaria inuloides(Arecaceae)中多酚、單寧、花青素和類黃酮提取的影響,以及對(duì)其抗氧化和抗菌活性影響的研究。提取中分別采用超聲處理30分鐘,頻率為50 Hz,速度為70 rpm。結(jié)果表明,總多酚含量(TPC)、總黃酮(TFC)和總的丹寧含量(TTC)以及抗氧化能力在使用甲醇提取時(shí)達(dá)到最大。P.inuloides的甲醇和乙醇提取物對(duì)于所有選定的微生物都具有較低的抗菌活性。二乙醚提取物對(duì)選定的所有被測(cè)試細(xì)菌沒(méi)有抗菌活性。第四階段,蚤草屬inuloides葉的乙醇提取物的GC-MS分析顯示10峰指示存在10個(gè)化合物。第五階段,研究從P.inuloides中提取分離的酚類化合物的抗氧化和抗菌活性,并對(duì)其脂肪酸和生育酚含量進(jìn)行評(píng)價(jià)。通過(guò)大孔樹(shù)脂柱層析分離得到酚類化合物,然后利用高效液相色譜法進(jìn)行分析。提取物中γ-生育酚具有最高含量,大約占總生育酚的52.76%,其次是α-生育酚(29.83%)和δ-生育酚(17.41%)。另外,P.inuloides中含有大量的鉀,鎂,鈉,鐵和鈣,含量分別為159.5,29.5,14.2,13.875和5.225mg/100g。利用掃描電鏡、透射電鏡(TEM,SEM and EDX)揭示了P.inuloides的內(nèi)部結(jié)構(gòu)。第六階段,將P.inouloides葉子中的精油(PIEO)應(yīng)用在海產(chǎn)品juvenile kachlan(Trachinotus ovatus,Linnaeus)上。結(jié)果表明,P.inuloides的精油抑制所有測(cè)試的微生物,對(duì)大腸桿菌的最小抑制濃度(MIC)為2.0μg/m L。本研究以純水為對(duì)照組,研究了Pi EO應(yīng)用在魚片上對(duì)單核細(xì)胞增生李斯特氏菌、大腸桿菌和金黃色葡萄球菌的影響。結(jié)果表明,以純水做參照,應(yīng)用了Pi EO后(25℃),立即消除了魚片上大部分單核細(xì)胞增生李斯特氏菌、大腸桿菌和金黃色葡萄球菌,降低了固相細(xì)胞計(jì)數(shù)值(SPC)。在10℃下12天內(nèi)也可減少以上細(xì)菌的含量,在10℃保存條件下,延長(zhǎng)了食品的貨架期。在貯藏結(jié)束時(shí),SPC計(jì)數(shù)7 log cfu/g。感官評(píng)價(jià)證明P.inouloides精油處理后的魚片是可以被消費(fèi)者接受的。結(jié)果還表明,從超市(江蘇,無(wú)錫)購(gòu)買的魚產(chǎn)品擁有良好的衛(wèi)生及健康條件,而從當(dāng)?shù)厥袌?chǎng)購(gòu)買的魚產(chǎn)品則不達(dá)標(biāo)準(zhǔn)。在下一階段,驗(yàn)證P.inouloides葉中的精油對(duì)于糖尿病的效果。精油的化學(xué)分析結(jié)果表明它的成分中含有生物堿、類固醇、皂甙、糖苷、還原糖和萜類化合物。在250μg/ml的濃度下,P.inuloides葉子精油對(duì)α-淀粉酶和α-葡糖苷的抑制活性分別為85.50%和86.52%,IC50值分別為140.49μg/ml和128.33μg/ml。P.inuloides葉子精油在實(shí)驗(yàn)劑量水平(400 mg/kg體重)下可以降低糖尿病大鼠的空腹血糖水平。到了實(shí)驗(yàn)的第21天,在P.inuloides葉子精油處理組,大鼠血糖值下降到與糖尿病藥物處理組大鼠沒(méi)有明顯差異(P0.05)。在鏈脲佐菌素(STZ)誘導(dǎo)糖尿病模型大鼠中,P.inuloides葉子精油可以降低大鼠的總膽固醇,甘油三酯和低密度脂蛋白(LDL)水平,同時(shí)會(huì)增加高密度脂蛋(HDL)水平。在實(shí)驗(yàn)的21天內(nèi),大鼠體重保持穩(wěn)定。在鏈脲佐菌素(STZ)誘導(dǎo)糖尿病模型大鼠中,反應(yīng)肝功能的標(biāo)志物谷丙轉(zhuǎn)氨酶(ALT)、谷草轉(zhuǎn)氨酶(AST)和堿性磷酸酶(ALP)含量的顯著升高。而P.inuloides葉子精油的處理可以顯著降低大鼠體內(nèi)這些酶的水平(P0.01)。在鏈脲佐菌素(STZ)誘導(dǎo)糖尿病模型大鼠中,肌酐、尿素氮、總蛋白等腎功能標(biāo)記物含量顯著升高。而P.inuloides葉子精油的處理可以顯著降低大鼠體內(nèi)以上指標(biāo)的水平(P0.01)。這些結(jié)果表明,糖尿病大鼠都患有腎功能障礙。肝臟組織病理學(xué)研究表明,與對(duì)照組相比,P.inuloides葉子精油能廣泛改善胰島細(xì)胞的生理狀況。P.inuloides的毒理試驗(yàn)結(jié)果顯示,P.inuloides沒(méi)有任何導(dǎo)致急性毒性的跡象,如呼吸窘迫,昏迷或死亡等。在最后一部分細(xì)胞實(shí)驗(yàn)中,結(jié)果顯示對(duì)P.inuloides葉子精油MCF-7細(xì)胞,具有很強(qiáng)的細(xì)胞毒活性的作用,IC50分別為12.9和3.00μl/ml。Pi EO的精油抑制所有測(cè)試的微生物(Listeria)。并運(yùn)用光學(xué)顯微鏡對(duì)MCF-7細(xì)胞形態(tài)進(jìn)行了分析。
[Abstract]:Pulicaria inuloides Asteraceae is a plant of the genus Pulicaria, which is widely used as anticancer drugs, and for the treatment of diabetes, rheumatism, gout, cystitis, nephritis, dysmenorrhea, toothache, headache, and has antibacterial, anti coagulation effect of.P.inuloides containing alkaloids, terpenes, volatile oil, flavonoids, phenolic acid, tannic acid and other ingredients. Stage, get the essential oil from Pulicaria inuloides-Astereceae and Ocimum of medicinal and aromatic plants leaves forskoleiLamaiceae, isolation and identification of bioactive substances from oil. Then the determination of antioxidant activity of the isolated material, and the anti microbial activity of total phenolic content. The results showed that compared with O.forskolei, P.inuloides had higher total phenolic content (O.forskolei 54.6 + 30 mg GAE/g extract, P.inuloides 144 + 5.32 mg GAE/g extract); P.inuloides has the higher antioxidant activity, DPPH clearance rates were 26.76 + 0.11% (O.forskolei) and 92.92 + 0.10% (P.inuloides), IC50 were 73.03 + 0.05 and 4.5 + 0.05 (O.forskolei) (P.inuloides); P.inuloides has a higher beta carotene bleaching values were 41.03 + 6.35% and 90.77 + 0.21% (O.forskolei) (P.inuloides).P.inuloides essential oil can inhibit all the tested microorganisms except Salmonella and Shigella dysenteriae, and the minimum inhibitory concentration of Escherichia coli (MIC) 3 g/m L.O.forskolei oil only inhibit Candida albicans. The second stage, by using gas chromatography - mass spectrometry method for identification of P.inuloides and volatile compounds in the roots of O.forskolei. The results showed that compared with O.forskolei, total phenol the content of P.inuloides in root contains higher (P.inuloides = 55.4 + 0.1, 35.3 + O.forskolei 0.2mg GAE/g extract); P.inuloides has lower antioxidant activity, DPPH clearance rates were 23. 07 + 0.06% (P.inuloides) 43.97 + 0.18% (O.forskolei), IC50 = 80.01 + 0.03 (P.inuloides + 0.01) and 31.18 (O.forskolei); P.inuloides had lower beta carotene bleaching value: 25.3 + 0.3% (P.inuloides + 0.7%) and 39.9 (O.forskolei).P.inuloides essential oil can inhibit the removal of Salmonella typhimurium all the tested microorganisms except bacteria, Bacillus dysenteriae and Escherichia coli. In the third stage, Pulicaria inuloides of the type of solvents (Arecaceae) in polyphenols, tannins, anthocyanins and flavonoids extraction effect, and the effect of antioxidant and antibacterial activity research. Extraction were used in ultrasonic treatment for 30 minutes, frequency is 50 Hz. The speed of 70 rpm.. The results showed that the total polyphenol content (TPC), total flavonoids (TFC) and total tannins (TTC) and antioxidant capacity reached the maximum.P.inuloides of methanol and ethanol extracts of methanol extraction in use For all selected microorganisms with antimicrobial activity is low. Two selected ether extracts of all tested bacteria have antibacterial activity. The fourth stage, the ethanol extract of leaves of the pulicaria inuloides GC-MS analysis showed 10 peaks indicating the presence of 10 compounds. The fifth stage extraction of phenolic compounds isolated from antioxidant P.inuloides and the antibacterial activity, and to evaluate the fatty acid and tocopherol content. Phenolic compounds by macroporous resin column chromatography, high performance liquid chromatography and then use analysis. In the extract of gamma tocopherol has the highest content of total tocopherol 52.76%, followed by alpha tocopherol (29.83%) and delta tocopherol (17.41%). In addition, contains a lot of potassium, P.inuloides sodium, magnesium, iron and calcium, the content of 159.5,29.5,14.2,13.875 and 5.225mg/100g. respectively by scanning electron microscopy, transmission electron microscopy (TEM SEM, and EDX) to reveal the internal structure of P.inuloides. In the sixth stage, the essential oils will the leaves of P.inouloides (PIEO) used in juvenile kachlan (Trachinotus ovatus, seafood Linnaeus). The results show that the P.inuloides essential oils on all tested microorganisms, the minimum inhibitory concentration of Escherichia coli (MIC) 2 g/m L. in this study, using pure water as the control group, the Pi EO application in the study of fish Listeria monocytogenes List S effect of bacteria, Escherichia coli and Staphylococcus aureus. The results showed that using pure water as the reference, the application of Pi EO (25 C), immediately eliminates most fish Listeria monocytogenes bacteria List S, Escherichia coli and Staphylococcus aureus, reduce solid cell count (SPC). The content of 10 C within 12 days can also reduce bacteria, 10 degrees in the preservation conditions, prolong the shelf life of food in the storage node. Beam, SPC count 7 log cfu/g. P.inouloides after the treatment of the sensory evaluation of fish oil can be accepted by consumers. The results also show that from the supermarket (Jiangsu, Wuxi) has a good health and health conditions to buy fish products, and purchased from the local market of fish products is not up to the standard in the next. Stage, verification of P.inouloides in the leaf essential oil for diabetes. The results showed that the chemical analysis of essential oil composition containing alkaloids, steroids, saponins, glycosides, reducing sugar and terpenoids. At a concentration of 250 g/ml, inhibited P.inuloides activity of leaf essential oils of alpha amylase and alpha glucoside respectively. 85.50% and 86.52%, IC50 values were 140.49 g/ml and 128.33 g/ml.P.inuloides in leaf oil level of experimental dose (400 mg/kg body weight) can reduce fasting blood glucose levels in diabetic rats. On the twenty-first day of the experiment, In the P.inuloides leaf oil treatment group, blood glucose of rats and decreased to the diabetes drug treatment group (P0.05). There was no significant difference in streptozotocin (STZ) induced diabetic rats, P.inuloides leaf essential oil can reduce the total cholesterol of rats, three glycerol ester and low density lipoprotein (LDL at the same time will increase the level), high density lipoprotein (HDL) levels. On the 21 day of the experiment, the body weight of rats remained stable. In streptozotocin (STZ) induced diabetic rats in response, liver function markers alanine aminotransferase (ALT), aspartate aminotransferase (AST) and alkaline phosphatase (ALP) significantly increased the content of P.inuloides. And the processing of leaf essential oils can significantly reduce the levels of these enzymes in rats (P0.01). In streptozotocin (STZ) induced diabetic model rats, serum creatinine, urea nitrogen, total protein and other markers of renal function were significantly increased and P.inu. Loides leaf oil can significantly reduce the rats above index level (P0.01). These results showed that diabetic rats had renal dysfunction. The histopathological study showed that, compared with the control group, the toxicological test results of P.inuloides leaf oil can widely improve islet cell physiological status of.P.inuloides showed no P.inuloides any signs of acute toxicity of lead, such as respiratory distress, coma or death. In the last part of the cells in the experiment, the results showed that for P.inuloides leaves oil MCF-7 cells, has strong cytotoxic activity and IC50 were 12.9 and 3 l/ml.Pi EO oils inhibit all the tested microorganisms (Listeria). The use of optical microscope to analyze MCF-7 cell morphology.

【學(xué)位授予單位】：江南大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2017
【分類號(hào)】：TQ460.1

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本文編號(hào)：1490103