本文選題：分心木 + 體外抗腫瘤��； 參考：《新疆醫(yī)科大學(xué)》2016年碩士論文

【摘要】：目的:1.研究分心木不同提取物的體外抗腫瘤活性,篩選出體外抗腫瘤作用的活性部位。2.對活性部位的化學(xué)成分進(jìn)行系統(tǒng)研究。3.優(yōu)化分心木總皂苷的提取純化工藝。4.探究分心木總皂苷、總黃酮和總多糖的體外抗氧化活性。方法:1.95%乙醇回流提取分心木,收集提取液旋轉(zhuǎn)蒸發(fā)成浸膏后,依次用石油醚、氯仿、乙酸乙酯、水飽和正丁醇對乙醇提取物進(jìn)行萃取;采用MTT法測定分心木各提取物對人肝癌細(xì)胞7404、人非小細(xì)胞肺癌細(xì)胞A549及人結(jié)腸癌細(xì)胞Hct116、Caco-2增殖的影響,初步篩選出體外抗腫瘤的活性部位;流式細(xì)胞儀測定分心木活性部位對4種癌細(xì)胞凋亡的影響,進(jìn)一步確認(rèn)其活性部位。2.采用溶劑萃取、大孔吸附樹脂、硅膠柱層析、反相硅膠柱及重結(jié)晶等方法對活性部位的化學(xué)成分進(jìn)行分離和純化,根據(jù)理化性質(zhì),波譜學(xué)數(shù)據(jù)以及文獻(xiàn)對照確定所得化合物的結(jié)構(gòu)。3.以齊墩果酸為對照品,利用紫外可見分光光度法測定分心木中總皂苷的含量;利用單因素實驗結(jié)果,設(shè)計正交實驗,研究分心木總皂苷最佳提取工藝;利用大孔吸附樹脂對分心木總皂苷進(jìn)行純化工藝的研究。4.以VC作為陽性對照,檢測分心木中總皂苷、總黃酮和總多糖對超氧陰離子(-O2)、羥基自由基(-OH)、DPPH自由基的清除能力及其鐵還原能力測定。結(jié)果:1.分心木不同提取物對4種癌細(xì)胞的增殖均具有一定的抑制作用,但分心木乙酸乙酯和正丁醇提取物對4種癌細(xì)胞增殖的抑制作用最強,初步判斷乙酸乙酯和正丁醇提取物為分心木體外抗腫瘤活性的主要活性部位。通過流式細(xì)胞儀測定乙酸乙酯和正丁醇提取物對4種癌細(xì)胞凋亡的影響,在不同濃度下兩種提取物誘導(dǎo)4種癌細(xì)胞凋亡,其給藥濃度在400μg/mL時,4種癌細(xì)胞的凋亡率都大于50%。2.對體外抗腫瘤活性部位(正丁醇部位)進(jìn)行分離純化,從中分離出3個化合物,分別為:對苯二甲酸二甲酯(I)、槲皮苷(II)、Jugnaphthalenoside A(III)。3.總皂苷最佳提取工藝為25倍量的80%乙醇,80℃回流提取2次,每次2 h;最佳純化工藝為采用D101大孔吸附樹脂,最大上樣量為180 mL,水洗脫量為8BV,洗脫劑為50%乙醇,洗脫體積為7BV,總皂苷純度由27.4%上升到50.8%,總皂苷的洗脫率可達(dá)到82.4%。4.體外抗氧化實驗結(jié)果表明總皂苷、總黃酮和總多糖均具有較好的對DPPH自由基,羥基自由基,超氧陰離子自由基清除能力和鐵還原能力,并在一定的質(zhì)量濃度范圍內(nèi)成量效關(guān)系。結(jié)論:1.乙酸乙酯和正丁醇提取物是分心木發(fā)揮體外抗腫瘤作用的主要活性部位。2.本研究首次從分心木正丁醇提取物中分離得到化合物I和III。3.實驗表明優(yōu)選出的提取純化工藝穩(wěn)定可行,適用分心木總皂苷的提取純化。4.分心木總皂苷、總黃酮及總多糖具有體外抗氧化活性。
[Abstract]:Purpose 1.To study the antitumor activity of different extracts of distraction tree in vitro, and to screen out the active site of anti-tumor effect. 2. 2.The chemical constituents of active parts were systematically studied.Optimization of extraction and purification process of total saponins. 4.To explore the anti-oxidation activity of total saponins, flavonoids and polysaccharides of distraction wood in vitro.Methods 1. 95% ethanol was refluxed to extract distraction wood. The extract was then extracted by petroleum ether, chloroform, ethyl acetate and water saturated n-butanol.The proliferation of human hepatoma cell line (7404), human non-small cell lung cancer cell line A549 and human colon cancer cell line Hct116Caco-2 was determined by MTT assay.Flow cytometry was used to determine the effect of the active site of distraction tree on apoptosis of four kinds of cancer cells, and to confirm the active site. 2. 2.The active parts were separated and purified by solvent extraction, macroporous adsorption resin, silica gel column chromatography, reverse phase silica gel column and recrystallization.The structure of the compound was determined by spectroscopic data and literature comparison.The content of total saponins in distraction wood was determined by ultraviolet visible spectrophotometry with oleanolic acid as reference substance, and the optimum extraction process of total saponins was studied by orthogonal experiment.Study on purification of total saponins by macroporous resin. 4.The scavenging ability of total saponins, flavonoids and polysaccharides in distraction trees against superoxide anion (O _ 2), hydroxyl radical (OH) -OHH _ (2) and their iron reduction ability were determined with VC as a positive control.The result is 1: 1.Different extracts of distraction tree had certain inhibitory effects on the proliferation of four kinds of cancer cells, but ethyl acetate and n-butanol extracts had the strongest inhibitory effect on the proliferation of four kinds of cancer cells.It was preliminarily determined that ethyl acetate and n-butanol extract were the main active sites of antitumor activity in vitro.The effects of ethyl acetate and n-butanol extract on the apoptosis of four kinds of cancer cells were determined by flow cytometry. The apoptotic rates of four kinds of cancer cells were higher than 50. 2 at the concentration of 400 渭 g/mL.The antitumor active site (n-butanol) was isolated and purified in vitro. Three compounds were isolated, I. E. dimethyl terephthalate, Quercetin II, Jugnaphthalenoside Ajii. 3.The optimum extraction process of total saponins was 25 times of 80% ethanol at 80 鈩，

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