高渗、低温和氧化条件下酵母细胞应激产生活性物质的研究
发布时间：2018-10-15 11:10
【摘要】：本论文对酵母细胞在高渗、低温和氧化条件下的应激反应进行了研究,筛选了耐高渗酵母突变株,通过低剂量刺激酵母细胞产生活性衍生物—LYCD,开辟了制备LYCD的新途径,考察了LYCD对氧化损伤的酵母细胞的保护作用。 采用对数期正常酵母细胞为研究对象,对其进行低剂量高渗(KCl)和氧化(H_2O_2)预处理后发现细胞对更高剂量产生了抗性,并且这种抗性具有自我适应性和交叉适应性。此外,在低温和氧化应激反应之间也存在交叉适应性。这种交叉适应性说明,高渗和氧化应激、低温和氧化应激对细胞的损伤在一定程度上具有相似性,预处理后可能诱导出相似的保护物质。 测定了酵母经高渗、低温和氧化预处理前后细胞内几种重要抗氧化剂(GSH、SOD、CAT)和总抗氧化能力(T-AOC)的变化,结果表明:预处理都可以使正常酵母细胞中抗氧化剂含量提高,总抗氧化能力增强。并分析了正常酵母预处理前后细胞内脂质过氧化物—丙二醛(MDA)的含量,发现预处理后,再置于氧化致死条件下,MDA的含量降低,细胞的存活率也升高。这不仅说明预处理使细胞抗氧化水平提高,减少了MDA的积累,细胞增加了对这几种不良环境的抗性,更直接说明高渗、低温与氧化关系密切。 将经过高渗和低温预处理后的酵母进行破壁,分别得到LYCD_(KCl)和LYCD_(LT),发现两者都对细胞抗氧化具有保护活性。分别对其处理条件进行优化,确定了LYCDK_(KCl)的最佳处理条件为:1%KCl预处理90min,LYCD_(LT)的最佳处理条件为:4℃预处理60min。 通过紫外诱变筛选了耐高渗酵母突变株(CYK),与正常酵母相比,发现耐高渗突变株对氧化具有很强的抗性,而且耐高渗酵母提取物(LYCDK)也具有保护细胞抵抗氧化的能力。分析了正常酵母和耐高渗酵母以及预处理对耐高渗酵母细胞内GSH、SOD、CAT、T-AOC和MDA的差异,发现耐高渗酵母胞内抗氧化水平高于正常酵母菌株,且预处理增加了LYCDK中抗氧化剂的含量,降低了MDA水平。 对正常酵母细胞预处理前后蛋白的变化进行了电泳分析,结果表明,一些蛋白是H_2O_2和KCl都可诱导合成的,另一些蛋白是由H_2O_2或KCl特异性诱导合成的。 探讨了H_2O_2和紫外线条件下,LYCD对酵母细胞氧化损伤的保护作用。在H_2O_2(10mmol/L)和紫外线(20W紫外灯下30cm处照射60s)氧化损伤酵母细胞模型中添加LYCD后,细胞存活率显著上升,且有剂量依赖性。LYCD提高了紫外线氧化损伤细胞上清液中GSH、SOD和CAT含量,降低了MDA水平。
[Abstract]:In this paper, the stress reaction of yeast cells under hyperosmotic, low temperature and oxidizing conditions was studied. The mutant strain of hyperosmotic yeast was screened, and a new way to prepare LYCD was opened up by low dose stimulation of yeast cells to produce active derivative-LYCD,. The protective effect of LYCD on oxidative damaged yeast cells was investigated. Normal yeast cells in logarithmic phase were pretreated with low dose and high osmotic (KCl) and oxidation (H_2O_2). It was found that the cells developed resistance to higher doses and the resistance was self-adaptive and cross-adaptive. In addition, there is cross adaptation between low temperature and oxidative stress reaction. This cross-adaptation indicated that the damage to cells caused by hyperosmotic and oxidative stress, hypothermia and oxidative stress was similar to a certain degree, and similar protective substances might be induced after pretreatment. The changes of several important antioxidants (GSH,SOD,CAT) and total antioxidant capacity (T-AOC) in yeast before and after hypertonic, low temperature and oxidation pretreatment were measured. The results showed that pretreatment could increase the content of antioxidants in normal yeast cells. Total antioxidant capacity increased. The content of lipid peroxide-malondialdehyde (MDA) in normal yeast cells was analyzed before and after pretreatment. It was found that after pretreatment, the content of MDA decreased and the survival rate of cells increased. This indicated that pretreatment not only increased the antioxidant level of cells and reduced the accumulation of MDA, but also increased the resistance of cells to these adverse environments, which directly indicated that high permeability and low temperature were closely related to oxidation. The yeast treated with hypertonic and cryogenic pretreatment was broken into the wall, and LYCD_ (KCl) and LYCD_ (LT), were obtained, respectively, which showed that both of them had protective effect on the antioxidant activity of the cells. The optimum treatment conditions of LYCDK_ (KCl) were determined as follows: 1%KCl pretreatment 90 min LYCD _ (LT) treatment conditions were: 4 鈩，
本文编号：2272359
[Abstract]:In this paper, the stress reaction of yeast cells under hyperosmotic, low temperature and oxidizing conditions was studied. The mutant strain of hyperosmotic yeast was screened, and a new way to prepare LYCD was opened up by low dose stimulation of yeast cells to produce active derivative-LYCD,. The protective effect of LYCD on oxidative damaged yeast cells was investigated. Normal yeast cells in logarithmic phase were pretreated with low dose and high osmotic (KCl) and oxidation (H_2O_2). It was found that the cells developed resistance to higher doses and the resistance was self-adaptive and cross-adaptive. In addition, there is cross adaptation between low temperature and oxidative stress reaction. This cross-adaptation indicated that the damage to cells caused by hyperosmotic and oxidative stress, hypothermia and oxidative stress was similar to a certain degree, and similar protective substances might be induced after pretreatment. The changes of several important antioxidants (GSH,SOD,CAT) and total antioxidant capacity (T-AOC) in yeast before and after hypertonic, low temperature and oxidation pretreatment were measured. The results showed that pretreatment could increase the content of antioxidants in normal yeast cells. Total antioxidant capacity increased. The content of lipid peroxide-malondialdehyde (MDA) in normal yeast cells was analyzed before and after pretreatment. It was found that after pretreatment, the content of MDA decreased and the survival rate of cells increased. This indicated that pretreatment not only increased the antioxidant level of cells and reduced the accumulation of MDA, but also increased the resistance of cells to these adverse environments, which directly indicated that high permeability and low temperature were closely related to oxidation. The yeast treated with hypertonic and cryogenic pretreatment was broken into the wall, and LYCD_ (KCl) and LYCD_ (LT), were obtained, respectively, which showed that both of them had protective effect on the antioxidant activity of the cells. The optimum treatment conditions of LYCDK_ (KCl) were determined as follows: 1%KCl pretreatment 90 min LYCD _ (LT) treatment conditions were: 4 鈩，
本文编号：2272359
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