血红蛋白液态和固态制剂稳定性研究
文献类型：学位论文
| 作者 | 李小爽 |
| 学位类别 | 硕士 |
| 答辩日期 | 2012-05-22 |
| 授予单位 | 中国科学院研究生院 |
| 导师 | 徐霞 |
| 关键词 | 血红蛋白 液态制剂 冷冻干燥 高铁血红蛋白 稳定性 |
| 其他题名 | Study on Aqueous and Solid Formulations for preservation of Hemoglobin |
| 学位专业 | 化学工程 |
| 中文摘要 | 血红蛋白是制备血液代用品或血红蛋白类氧载体的重要原料，被广泛地用于医药行业，随着血液代用品的进一步深入研究，血红蛋白在长期保存过程中的稳定性成为了研究的关键环节。血红蛋白在保存过程中容易发生构象改变，其四级结构是由4个亚基通过疏水相互作用结合而成，在液相中可解聚为二聚体或单聚体；同时由于其亚基上血红素中心的亚铁离子易氧化，容易由氧合血红蛋白变为高铁血红蛋白而失去载氧能力。另外，由冷冻干燥方法制备血红蛋白的固态制剂也可能引起高铁血红蛋白含量的增加，从而导致血红蛋白而丧失载氧能力。本文研究了在利用氨基酸或糖类做保护剂的条件下血红蛋白液态及固态制剂的稳定性问题，利用数学模型研究了制剂冷冻干燥过程物理参数（温度、pH、崩塌温度等）变化，主要结论如下：（1）在25°C和40°C下，Gly、Cys和Glu能够显著降低在血红蛋白保存过程中的氧化程度，而Vc、Met并未对血红蛋白的氧化产生明显抑制作用，还原作用不是主要抑制血红蛋白维持氧合态的主要因素。（2）建立数学模型，获得了不同降温速率下10 mM磷酸盐缓冲液冷冻时温度及冰晶半径变化曲线，并计算得到了磷酸钠盐及磷酸钾盐缓冲液在溶液结晶后的pH变化曲线，磷酸钠盐缓冲液由初始7.2在冷冻至-40 °C后降低至5.4；磷酸钾盐缓冲液的pH值由初始7.2在冷冻至-40°C后升高至7.5。（3）冻干过程中pH及崩塌温度测定：a)开发可视化方法测定冻结状态下的pH；b)在常用浓度条件下，磷酸钠盐缓冲液不会对体系的崩塌温度造成显著影响，表面活性剂对体系的崩塌温度有一定影响，Pluronic F68能够使剂型的Tc降低，但对于不同的体系，降低程度有所不同。（4）不同比例的冻干保护剂会对血红蛋白冻干过程的二级结构造成影响，葡萄糖醛酸与血红蛋白的最适摩尔比为128:1，山梨醇在摩尔比为64:1及128:1时，血红蛋白α螺旋破坏程度最低，为最适宜的摩尔比，葡萄糖醛酸及山梨醇对血红蛋白在冻干过程中的自氧化并没有明显的抑制作用。 |
| 英文摘要 | Hemoglobin, which has been widely used in pharmaceutical industry, is an important source of preparing blood substitute or hemoglobin based oxygen carrier. The stability for hemoglobin during long-time preservation is critical for further research on blood substitute. The conformation of hemoglobin changes easily during preservation. Hemoglobin easily degrades into dimer or mono when preserved in aqueous fomulation because it is comprised of four subunits which are combined by hydrophobic interaction. Meanwhile, hemoglobin can become methemoglobin which losses the ability to carry oxygen due to the easy oxidation of ferrous ion in the center of the heme. In addition, the content of methemoglobin may increase when preparing solid hemoglobin formulation using freeze-drying. In the present study, the stability of hemoglobin in aqueous and solid formulation using amino acid or sugar as protectants was investigated. We simulated changes of physical parameters (temperature, pH, collapse temperature et al) during freezing using mathematical models. The main results are as follows: (1) the oxidization of hemoglobin during the preservation process remarkbly was reduced in the presence of Gly, Cys and, Glu when keeping at 25°C and 40°C. Reduction was not the main factor which preventing hemoglobin from changing into methemoglobin. (2) Changes in temperature and ice crystal radius for phosphate system (10 mM) were obtained based on the mathematical modelling when different cooling rates were applied. Meanwhile, the pH shift in the system of sodium and potassium phosphate after freezing was simulated. When freezed to -40°C pH decreased from 7.2 to 5.4 for sodium phosphatesystem while pH increased from 7.2 to 7.5 when freezed to -40°C for potassium phosphate buffer. (3) Measurement of pH and collapse temperature during freeze-drying: a. A visual method was developed for measuring pH in the frozen state. b. Sodium phosphate buffer had no significant effect on collapse temperature at common concentration while surfactant had some effect on it. collapse temperature was decreased in the presence of Pluronic F68. (4) The ratio of lyoprotectant to hemoglobin had an significant effects on the secondary structure of hemoglobin. The secondary structure of hemoglobin was maintained at the molar ratio of 128 with glucuronic acid, and at 64 and 128 with sobitol.However, both glucuronic acid and sorbitol had no obvious effect on autooxidation of hemoglobin. |
| 语种 | 中文 |
| 公开日期 | 2013-09-25 |
| 源URL | [http://ir.ipe.ac.cn/handle/122111/1848]  |
| 专题 | 过程工程研究所_研究所（批量导入） |
| 推荐引用方式 GB/T 7714 | 李小爽. 血红蛋白液态和固态制剂稳定性研究[D]. 中国科学院研究生院. 2012. |
入库方式： OAI收割
来源：过程工程研究所
浏览0
下载0
收藏0
其他版本
除非特别说明，本系统中所有内容都受版权保护，并保留所有权利。