發(fā)布時(shí)間：2018-06-03 19:08

  本文選題：抗氧化 + 乳酸菌�。� 參考：《江南大學(xué)》2009年碩士論文

【摘要】： 目的:從乳酸發(fā)酵食品中分離篩選具有較高抗氧化活性的乳酸菌,研究其抗氧化能力并初步探討其作用機(jī)制,為開(kāi)發(fā)功能性乳制品提供理論依據(jù)。 方法:以體外清除DPPH的能力為指標(biāo),在相同菌量條件下,從32株受試菌株中篩選出具抗氧化活性的乳酸菌,正交實(shí)驗(yàn)優(yōu)化其發(fā)酵條件,以最大程度上提高其抗氧化水平。小鼠結(jié)腸癌細(xì)胞CT-26傳代培養(yǎng)后分為對(duì)照組、模型組和乳酸菌干預(yù)組。模型組用含0.45 mmol/L H2O2(終濃度)的RPMI-1640培養(yǎng)基作用于細(xì)胞2 h建立損傷模型,干預(yù)組將109cfu/mL的具抗氧化活性的乳酸菌與含0.45 mmol/L H2O2的RPMI-1640培養(yǎng)基于37℃孵育3 h,過(guò)濾除菌后取上清作用于細(xì)胞2 h,分別測(cè)定各組細(xì)胞活力(MTT法),測(cè)定上清液LDH、MDA及SOD含量,掃描電鏡觀察細(xì)胞表面形態(tài),Hoechest 33342染色檢測(cè)細(xì)胞核變化,HepG2細(xì)胞輔助驗(yàn)證受試菌對(duì)其保護(hù)作用。 結(jié)果:從受試的32株乳酸菌株中,篩選得到一株具有較強(qiáng)抗氧化能力的嗜酸乳桿菌874,DPPH清除率在35%左右。利用單因素實(shí)驗(yàn),以發(fā)酵得到的菌體的DPPH清除率為指標(biāo),從接種量、培養(yǎng)基初始pH值、培養(yǎng)時(shí)間、培養(yǎng)溫度、吐溫種類(lèi)、生長(zhǎng)因子種類(lèi)及添加量,碳、氮源種類(lèi)及添加量等多個(gè)方面對(duì)篩得菌株的生長(zhǎng)條件進(jìn)行了優(yōu)化,并就碳、氮源及生長(zhǎng)因子(乳糖和豆芽汁)添加量等對(duì)清除DPPH自由基影響較大的因素進(jìn)行了正交實(shí)驗(yàn),確定了篩得菌株的優(yōu)化培養(yǎng)基及最佳培養(yǎng)條件。優(yōu)化培養(yǎng)基成分為乳糖1%,豆芽汁12.5%,葡萄糖2.5%,蛋白胨:牛肉浸膏=1:1,其余同MRS培養(yǎng)基。 最佳培養(yǎng)條件為:接種量為3%,培養(yǎng)基初始pH值為6.4,37℃下發(fā)酵20 h,吐溫-80作為促生長(zhǎng)因子。經(jīng)優(yōu)化后,菌株DPPH清除率由(35.15±0.49)%左右增至(66.70±0.42)%,清除羥自由基的能力由(19.60±0.99)%提高至(33.30±1.84)%,還原能力由(105.75±1.34)%增至(159.50±22.20)%。體外細(xì)胞培養(yǎng)研究結(jié)果表明:干預(yù)組細(xì)胞活力顯著高于模型組(P0.01),培養(yǎng)上清液中LDH及MDA含量顯著低于模型組(P0.01),SOD含量高于模型組(P0.01);掃描電鏡下模型組細(xì)胞表面微絨毛消失,出現(xiàn)大量凋亡小體,干預(yù)組形態(tài)接近正常;模型組經(jīng)Hoechst-33342熒光染色可見(jiàn)明顯的顆粒狀和固縮狀熒光,干預(yù)組呈彌漫均勻熒光。 結(jié)論:篩選得到1株嗜酸乳酸菌,經(jīng)優(yōu)化其DPPH清除率達(dá)(66.70±0.42)%,羥自由基清除率為(33.30±1.84)%,還原能力為(159.50±22.20)%;完整菌體可保護(hù)細(xì)胞免受H2O2造成的氧化損傷,其作用機(jī)制可能與乳酸菌可以清除自由基(活性氧)等有關(guān)。
[Abstract]:Objective: to isolate and screen lactic acid bacteria with high antioxidant activity from lactic acid fermented food, to study its antioxidant ability and to explore its mechanism, and to provide theoretical basis for the development of functional dairy products. Methods: according to the ability of scavenging DPPH in vitro, lactic acid bacteria with antioxidant activity were screened from 32 tested strains under the same amount of bacteria, and the fermentation conditions were optimized by orthogonal experiment in order to improve the antioxidant level to the greatest extent. Mouse colon cancer cells were divided into control group, model group and lactobacillus intervention group after subculture of CT-26. The model group was treated with RPMI-1640 medium containing 0.45 mmol/L H _ 2O _ 2 (final concentration) for 2 h to establish the injury model. In the intervention group, lactic acid bacteria with antioxidant activity of 109cfu/mL and RPMI-1640 containing 0.45 mmol/L H2O2 were incubated for 3 h at 37 鈩，

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