發(fā)布時間：2018-06-20 17:34

  本文選題：紅細胞 + 凍干保存　； 參考：《安徽醫(yī)科大學(xué)》2017年碩士論文

【摘要】：目的本實驗研究基于課題組先前的基礎(chǔ),探索凍干紅細胞殘余水分的最佳測定方法,探索凍干條件如溫度、干燥時間對紅細胞殘余水含量的影響,同時研究了殘余水的分布和其紅細胞代謝的影響,為進一步深入探索殘余水在紅細胞冷凍干燥保存方面的影響提供實驗和理論基礎(chǔ)。方法對同一程序凍干的紅細胞分別使用Karl-Fisher萃取法結(jié)合滴定法(Karl-Fisher法)、熱重分析法、核磁共振法測定凍干紅細胞殘余水,分析各組間結(jié)果差異;使用同一程序同時凍干等量的含保護劑的血影細胞、含保護劑的紅細胞、無保護劑的血影細胞、無保護劑的紅細胞,分別測定其殘余水含量并進行對比分析;控制凍干條件(時間、溫度、真空度)制備不同殘余水含量的凍干紅細胞,用Karl-Fisher法測定其殘余水含量,室溫放置保存,分別在凍干之前,凍干完成時、完成1個月后、完成3個月后、完成6個月后測定其ATP活性、SOD活力,分析不同殘余水含量組之間代謝數(shù)據(jù)的變化和關(guān)聯(lián)情況。結(jié)果本研究使用含12%甘油濃度(w/v)的保護劑,使用三種不同方法測定凍干紅細胞、凍干血影細胞、及水分標準品的殘余水含量。Karl-Fisher法測得凍干紅細胞水分含量為(3.37±0.05)%,無保護劑凍干紅細胞水分含量為(1.22±0.09)%,水分標準品水分含量為(5.14±0.13)%;熱重分析法測得凍干品水分含量為(12.40±0.56)%,無保護劑凍干品水分含量為(4.60±0.78)%,水分標準品水分含量為(5.28±0.16)%;核磁共振法測得凍干品水分含量為(0.78±0.09)%,無保護劑凍干品水分含量為(0.89±0.12)%,水分標準品水分含量為(4.99±0.18)%。除水分標準品外,三組間比較結(jié)果差異具統(tǒng)計學(xué)意義(P0.05)。使用方差分析多樣本間LSD檢驗,凍干血影細胞組和凍干紅細胞組比較均為P0.05,差異沒有統(tǒng)計學(xué)意義。血影細胞和紅細胞的殘余水分量無差異。Karl-Fisher法能夠很好地反應(yīng)含保護劑及不含保護劑凍干紅細胞的殘余水分含量,熱重分析法適用于無甘油保護劑的凍干紅細胞水分測定,核磁共振法測定值過低。通過控制凍干條件制備殘余水含量不同的六組凍干紅細胞,由低到高分別為2.80%,5.12%,9.10%,10.77%,11.73%,12.45%。凍干后完成一周、1個月、3個月、6個月后復(fù)水、洗滌,測得ATP含量和SOD活性,同時與凍干前紅細胞的測定值進行比較。各組間ATP值和SOD活力均隨時間有所下降,殘余水含量在2.80%和5.12%的兩組樣本功能隨著時間下降趨勢開始逐漸平緩。結(jié)論對于含甘油保護劑的凍干紅細胞,Karl-Fisher法、熱重法和核磁共振法均可以測定其殘余水含量,測定絕對值之間存在顯著差異。熱重分析法測得結(jié)果偏高,核磁共振法測得結(jié)果偏低;凍干紅細胞殘余水與凍干血影細胞殘余水含量差異無統(tǒng)計學(xué)意義,提示凍干紅細胞殘余水分布于紅細胞膜上;不同殘余水含量組凍干紅細胞的ATP含量和SOD活性水平隨著保存時間的延長均產(chǎn)生不同程度的下降趨勢,殘余水含量2.80%和5.12%組的下降趨勢較其他組別在保存后期趨于平穩(wěn),提示將殘余水含量控制在此區(qū)間將會有效提高紅細胞凍干保存的時間。
[Abstract]:Objective to explore the optimal determination method of residual water in freeze-dried red cells based on the previous basis of the project group, explore the effect of freeze drying conditions such as temperature and drying time on the residual water content of red cells, and study the distribution of residual water and the effect of its red cell metabolism, in order to further explore the residual water freezing in red blood cells. The effects of drying preservation provide experimental and theoretical basis. Methods the residual water of freeze-dried red cells was determined by Karl-Fisher extraction combined with titration (Karl-Fisher), thermogravimetric analysis and nuclear magnetic resonance (NMR), respectively. The blood shadow cells, the red blood cells containing the protectant, the unprotected blood shadow cells and the unprotected red cells, the residual water content was measured and compared. The frozen dry red cells with different residual water content were prepared by controlling the freeze drying conditions (time, temperature and vacuum), and the residual water content was determined by Karl-Fisher method, and stored at room temperature. Do not have frozen dry before the completion of freeze dry, completed 1 months after completion of the completion of 3 months, completed 6 months after the completion of the determination of its ATP activity, SOD activity, analysis of different residual water content of the metabolic data between the changes and association. Results this study used 12% glycerol concentration (w/v) protection agent, the use of three different methods to determine freeze-dried red blood cells, freeze-dried blood The water content of the frozen dry red cells was (3.37 + 0.05)%, the water content of the freeze-dried red cells was (1.22 + 0.09)%, and the water content of the water standard was (5.14 + 0.13)%, and the moisture content of the freeze-dried freeze-dried products was (12.40 + 0.56)% (12.40 + 0.56)%, and the moisture content of the freeze-dried freeze-dried products without protectant was measured by.Karl-Fisher method. The content of the water content was (4.60 + 0.78)%, the water content of the water standard was (5.28 + 0.16)%, the water content of the freeze-dried products was (0.78 + 0.09)%, the water content of the freeze-dried freeze-dried products was (0.89 + 0.12)%, and the water content of the water standard was (4.99 + 0.18)%. The difference of the results between the three groups except the water standard was statistically significant (P0.05). The variance analysis varied the LSD test. The difference between the freeze-dried blood shadow cell group and the freeze-dried red cell group was P0.05, the difference was not statistically significant. There was no difference between the residual water components of the blood shadow cell and the red cell, and the.Karl-Fisher method could react well to the residual water content of the protective agent and the freeze-free red blood cell, and the thermogravimetric analysis was suitable. Six groups of frozen dry red cells with different residual water content were prepared by controlling freeze-drying conditions, from low to high to 2.80%, 5.12%, 9.10%, 10.77%, 11.73%, and 12.45%. were completed for one week, 1 months, 3 months and 6 months after the freeze drying, and the ATP content was measured. The activity of SOD was compared with the determination of red blood cells before freeze-drying. The ATP value and SOD activity of each group decreased with time. The two groups of samples with residual water content in 2.80% and 5.12% began to gradually decrease with time. Conclusion for lyophilized red cells with glycerol protectants, Karl-Fisher, thermogravimetry, and nuclear magnetic Co The residual water in the lyophilized red cell and the lyophilized blood shadow cells were low, and the residual water of the freeze-dried red cells and the lyophilized blood shadow cells had no statistical significance, suggesting that the residual water of the lyophilized red cells was distributed on the red cell membrane; The ATP content and SOD activity level of the frozen dry red cells in the residual water content decreased with the prolongation of the preservation time. The decrease trend of the residual water content 2.80% and 5.12% groups was more stable than the other groups in the later period of preservation, suggesting that the residual water content control in this interval would effectively improve the preservation of red blood cells. Time.
【學(xué)位授予單位】：安徽醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：R318.52

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