本文選題：沙蜇　切入點(diǎn)：溶血活性　出處：《青島大學(xué)》2017年碩士論文

【摘要】：研究目的研究沙蜇毒素致紅細(xì)胞損傷(溶血活性)的機(jī)制,評(píng)估不同濃度的維生素C對(duì)沙蜇毒素溶血活性的影響。研究方法采用沙蜇自溶法,從沙蜇中分離刺絲囊細(xì)胞,利用超聲波細(xì)胞破碎法提取刺絲囊細(xì)胞中的毒素,以牛血清白蛋白為標(biāo)準(zhǔn),測(cè)得毒素蛋白的濃度。選取健康自愿者,男女各8名,各抽取新鮮血3~4ml,重復(fù)洗滌紅細(xì)胞2~3次后,配制成20%(V/V)人血紅細(xì)胞懸浮液和0.45%(V/V)人血紅細(xì)胞懸浮液。取不同體積的沙蜇毒素加入到1ml的20%人血紅細(xì)胞懸浮液中,加0.9%的氯化鈉溶液稀釋至5ml,使得沙蜇毒素終濃度為0、2、4、8、16μg/ml,應(yīng)用硫代巴比妥(TBA)法,檢測(cè)不同濃度的沙蜇毒素作用后,溶血反應(yīng)體系當(dāng)中的脂質(zhì)過(guò)氧化產(chǎn)物丙二醛(MDA)的水平。取一定體積的沙蜇毒素,加入到1ml的0.45%人血紅細(xì)胞懸浮液中,后加入不同體積的維生素C,再加入0.9%的氯化鈉溶液稀釋至5ml,使維生素C終濃度分別0、0.5、1、1.5、2、2.5mmol/L,測(cè)定不同濃度的維生素C對(duì)沙蜇毒素溶血活性的影響。結(jié)果實(shí)驗(yàn)數(shù)據(jù)所得,本研究中沙蜇毒素蛋白濃度為395.45μg/ml。不同濃度的沙蜇毒素作用后,紅細(xì)胞溶血體系中的MDA水平隨毒素劑量增大呈依賴性升高,實(shí)驗(yàn)中沙蜇毒素濃度為16μg/ml時(shí),MDA濃度由基礎(chǔ)濃度(0.79±0.07μmol/L)升高至(19.90±1.22μmol/L),采用單因素方差分析及LSD-t檢驗(yàn),實(shí)驗(yàn)各組與對(duì)照組比較,差異有統(tǒng)計(jì)學(xué)意義(p0.05);實(shí)驗(yàn)各組互相比較,差異有統(tǒng)計(jì)學(xué)意義(p0.05)。所研究濃度范圍內(nèi)的維生素C可以拮抗沙蜇毒素的溶血活性,隨著維生素C濃度的升高,溶血活性不斷下降。當(dāng)維生素C濃度在0~1mmol/L范圍內(nèi),毒素的溶血活性下降明顯,由75.79%(未添加維生素C)顯著下降至31.95%;當(dāng)維生素C濃度在1~2.5mmol/L范圍內(nèi),溶血活性由31.95%下降至26.66%,下降趨勢(shì)減慢。采用單因素方差分析及LSD-t檢驗(yàn),實(shí)驗(yàn)各組與對(duì)照組相比,差異有統(tǒng)計(jì)學(xué)意義(p0.05);實(shí)驗(yàn)各組互相比較,當(dāng)實(shí)驗(yàn)中維生素C濃度為1.5、2、2.5mmol/L時(shí),差異無(wú)統(tǒng)計(jì)學(xué)意義(p0.05),其余各組差異有統(tǒng)計(jì)學(xué)意義(p0.05)。結(jié)論脂質(zhì)過(guò)氧化損傷是沙蜇毒素溶血活性的機(jī)制之一,有其他機(jī)制參與了溶血過(guò)程。維生素C能夠一定程度上抑制沙蜇毒素的溶血活性,可以作為預(yù)防和治療沙蜇蜇傷的候選藥物。沙蜇毒素中存在對(duì)紅細(xì)胞產(chǎn)生脂質(zhì)過(guò)氧化損傷的成分,為沙蜇毒素中溶血蛋白的提純提供了實(shí)驗(yàn)幫助。意義了解沙蜇毒素致紅細(xì)胞損傷的機(jī)理,為沙蜇蜇傷的預(yù)防和治療提供實(shí)驗(yàn)依據(jù),為深入研究沙蜇毒素中各類生物活性物質(zhì)提供基礎(chǔ)資料。
[Abstract]:Objective to study the mechanism of erythrocyte damage (hemolytic activity) induced by jellyfish toxin and to evaluate the effect of different concentrations of vitamin C on the hemolytic activity of jellyfish toxin. The toxin was extracted by ultrasonic cell fragmentation method. The concentration of toxin protein was determined by using bovine serum albumin as the standard. Healthy volunteers, eight men and eight men, were selected to extract 3ml of fresh blood and washed red blood cells 23 times. We prepared 20 V / V human erythrocyte suspensions and 0.45 V / V human erythrocyte suspensions. Different volumes of jellyfish toxin were added to the 20% human erythrocyte suspensions of 1ml. When 0.9% sodium chloride solution was added to 5 ml, the final concentration of jellyfish toxin was 816 渭 g / ml, the final concentration of jellyfish toxin was 816 渭 g / ml. The method of thiobarbital TBA was used to detect the effect of different concentrations of jellyfish toxin. The level of malondialdehyde (MDA), a product of lipid peroxidation in hemolytic reaction system. A certain volume of jellyfish toxin was added to 0.45% human erythrocyte suspension of 1ml. Different volumes of vitamin C were added, then 0.9% sodium chloride solution was added to dilute to 5 ml. The final concentration of vitamin C was 0 ~ (0.5) ~ (1) ~ (-1) ~ (1.5) ~ (2.5) mmol / L, respectively. The effects of different concentrations of vitamin C on the hemolytic activity of jellyfish stings were determined. In this study, the concentration of jellyfish toxin protein was 395.45 渭 g / ml. The level of MDA in erythrocyte hemolysis system increased in a dependent manner with the increase of toxin dose after different concentrations of jellyfish toxin. When the concentration of jellyfish toxin was 16 渭 g/ml, the concentration of malondialdehyde increased from 0.79 鹵0.07 渭 mol / L to 19.90 鹵1.22 渭 mol 路L ~ (-1) 路L ~ (-1) 路L ~ (-1). The results of single factor variance analysis and LSD-t test showed that there was a significant difference between the experimental groups and the control group (P 0.05). The difference was statistically significant (P 0.05). Vitamin C in the studied concentration range could antagonize the hemolytic activity of jellyfish toxin. With the increase of vitamin C concentration, the hemolytic activity decreased. When the concentration of vitamin C was in the range of 0~1mmol/L, The hemolytic activity of the toxin decreased significantly from 75.79 (without vitamin C) to 31.95. When the concentration of vitamin C was in the range of 1~2.5mmol/L, the hemolytic activity decreased from 31.95% to 26.66%, and the downward trend slowed down. Univariate analysis of variance and LSD-t test were used. Compared with the control group, there was a significant difference between the experimental groups and the control group (p 0.05), and when the concentration of vitamin C in the experiment was 2.5 mmol / L, the concentration of vitamin C in the experiment was 2.5 mmol / L. There was no significant difference between the two groups (P 0.05). Conclusion the damage of lipid peroxidation is one of the mechanisms of the hemolytic activity of jellyfish toxin. There are other mechanisms involved in the hemolysis process. Vitamin C can inhibit the hemolytic activity of jellyfish toxin to some extent. It can be used as a candidate drug for the prevention and treatment of jellyfish stings. It provides experimental help for the purification of hemolytic protein from jellyfish toxin. The significance of this study is to understand the mechanism of erythrocyte damage induced by jellyfish toxin, and to provide experimental basis for the prevention and treatment of jellyfish stings. To provide basic data for further study of various bioactive substances in jellyfish toxin.
【學(xué)位授予單位】：青島大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：R751

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