| 题名 | 在线生物气溶胶质谱仪的研制及应用研究 |
| 作者 | 梁苗 |
| 学位类别 | 博士 |
| 答辩日期 | 2014-11 |
| 授予单位 | 中国科学院研究生院 |
| 授予地点 | 北京 |
| 导师 | 束继年 |
| 关键词 | 生物气溶胶，基质辅助激光解吸/电离，在线生物气溶胶质谱仪，在线 混合进样，在线分析，脂类生物标记物，bioaerosol, MALDI, online bioaerosl mass spectrometry, online analysis, online mixing technique, lipid biomarker |
| 其他题名 | Development of the an online bioaerosol mass spectrometer and its initial application for bioaerosol detection |
| 学位专业 | 环境工程 |
| 中文摘要 | 生物气溶胶（bioaerosol）是大气气溶胶的重要组成部分，会对大气环境和人类健康产生不利影响，同时由于生物气溶胶在环境中的物理和化学特性易变，所以研究和发展生物气溶胶的在线检测技术具有非常重要的意义。基质辅助激光解吸/电离技术（Matrix-Assisted Laser Desorption/Ionization，MALDI）是一种软电离技术，适用于生物大分子的分析和检测。MALDI 与飞行时间质谱技术（Time-of-Flight Mass Spectrometry, TOF MS）结合，可以快速、连续地检测宽质量范围的生物大分子物质。传统的MALDI-TOF MS 包含复杂的样品准备过程，所以，发展在线的MALDI-TOF MS 检测设备并将其应用于生物气溶胶领域，对于保护大气环境和保障公众安全具有非常重要的意义。 本论文介绍了一台自主研发的在线生物气溶胶质谱仪，它由在线混合进样装置和MALDI TOF 气溶胶质谱仪（MALDI TOF AMS）两部分构成，可以实现基质的在线添加，生物气溶胶的连续进样和在线MALDI 质谱分析。本研究在对在线生物气溶胶质谱仪进行调试和参数优化的基础上，完成了将此仪器应用于典型生物气溶胶的初步研究。主要研究成果及创新性如下： （1）在线生物气溶胶质谱仪的在线混合进样装置主要由两个雾化器和一个混合三通构成，它可以实现干燥样品颗粒和基质液滴颗粒在输送过程中的在线混合，并产生样品/基质混合气溶胶颗粒。混合颗粒直接通入MALDI TOF AMS 进行质谱解析。MALDI TOF AMS 主要由真空腔体、激光系统、反射式质谱装置和时序控制系统四部分构成。其中真空腔体由束源室、差分室、电离室和离子飞行时间管四部分构成；激光系统由波长为266 nm 的激光器、光学反射透镜和聚焦透镜构成；反射式质谱装置由V 型反射离子飞行元件和微通导板离子检测器组成；时序控制系统主要是一台DG535 脉冲发生器。此仪器采用垂直引出和二次反射技术消除基质离子噪声和脉冲电场对信号的干扰。经过调试后，此仪器可以准确检测PEG1000，分辨率达到2000。 （2）本研究利用在线生物气溶胶质谱仪通过对蜡酯（十六酸十六酯）气溶胶进行在线检测，得到了较高准确度和信噪比的十六酸十六酯离子信号，显示了此仪器的在线基质添加能力和在线分析能力。实验还研究了基质种类、基质溶剂配比、样品浓度、激光能量等实验因素对检测结果的影响，并优化了实验条件。最终得到的蜡酯气溶胶在线检测的最优实验条件如下：基质为LiDHB，样品浓度为0.8 mg/mL，基质与样品的摩尔比为8:1，基质溶剂为异丙醇:氯仿:正己醇=1:1:2 的混合溶剂，激光强度为1.2 mJ/pulse。仪器检测十六酸十六酯的分辨率为~1400，检出限为~0.8 pmol。 （3）本研究利用MALDI TOF 气溶胶质谱仪对四种花粉的脂类标记物进行检测，显示了此仪器对生物气溶胶进行快速种类鉴别的能力。实验首先采用改进的脂提方法对花粉脂质进行提取，然后利用MALDI TOF 气溶胶质谱仪检测到五种常见的花粉脂质，包括膜系统磷脂和花粉外膜脂质，并对质谱图中特征脂质信号峰进行了初步物质归属。结果表明，通过对不同花粉的脂质标记物进行检测，并对其特征脂质指纹峰进行对比，可以对花粉进行初步地物种鉴别和分类。 （4）本研究对三种不同的脂提方法进行了优化和对比，结果显示不同脂提方法对不同花粉脂质的提取效率有所差异。其中微波辅助甲酸消解法提取出的花粉脂质种类最多，同时产生的信号峰较多，信号较强，更适用于花粉脂质生物标记物的检测。 以上在线生物气溶胶的初步研究工作，表明这台仪器可以连续在线地对生物气溶胶进行检测，同时具备对生物气溶胶进行实时采样同时在线分析的潜力。在公共卫生健康、公共安全等领域具有重要应用价值。 |
| 英文摘要 | Bioaerosols are suspended sediments including allergens and microbial products which contribute to poor air quality and infectious diseases. Bioaerosol possesses widely variable physical and chemical properties in the atmpspheric environment. For these reasons, the development of rapid and on-line detection of bioaerosol has very important significance. Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) is a soft ionization analytical technique. The major advantages of MALDI-MS technique is the little or no fragmentations of mass spectra. Consequently, MALDI-MS is quite suitable for the analysis and detection of intact high-molecular- weight biomolecules. The technique of MALDI associating with TOF-MS represents the potential for rapid detection of bioaerosols. In this study, an online bioaerosl mass spectrometry has been developed. This instrument includes two parts: an in situ matrix/analyte mixing device and a MALDI TOF aerosol mass spectrometer (MALDI TOF AMS). The former part is fitted at the entrance of the MALDI TOF AMS. The main results were as follows: (1) The in-situ matrix/analyte mixing device mainly consisted of two droplet s generators and a mixing tee. The dry analyte particles and matrix droplets can continuously mix together in the mixing tee and the resulted analyte/matrix particles are then directly sampled into the following MALDI TOF AMS. The MALDI TOF AMS mainly consists of four parts: the vacuum chamber, laser system, reflectron-MS and timing sequence control system. The vacuum chamber is composed of four parts: the source chamber, the differential chamber, the deflection chamber and the detection chamber. The laser system is mainly composed of a 266 nm laser generator and the optical lens. The reflectron-MS is mainly composed of a set of ion reflectron-flight component and a multi-microchannel plates. The timing sequence control system is mainly consisted of a DG535 pulser. This instrument was debugged using PEG1000. The mass resolution is ~1600. (2) The performance of the online MALDI-TOF MS was evaluated with palmityl palmitate and lithium 2,4-dihydroxybenoic acid. Strong and stable MALDI signals of palmityl palmitate were obtained. Two factors, namely, the matrix solvent mixture and the analyte concentration, were optimized. Initial experimental results demonstrate that the technique provides an alternative method for online MALDI analysis. The mass resolution of the instrument reached 1400 m/Δm. The detection limit of the instrument for palmityl palmitate was ~0.8 pmol. (3) The lipidic extracts of pollen grains were analyzed using the MALDI TOF AMS. The lipidic extracts were premixed with the matrix. The sample/matrix aerosol was sampled constiously and analyzed rapidly. Membrane-associated phospholipids (phosphatidylserine and phosphatidylcholine) and diacylglycerol in pollen coats were observed. Results show that the characteristic lipid profiling can been used to characterize pollen. (4) The lipids in pollen grains were extracted by the methods of microwave-assisted formic acid digestion, microwave-assisted 2-aminoethanol solvolysis, and ultrasonic wave-assisted 2-aminoethanol solvolysis. The method of microwave-assisted formic acid digestion presented better lipid profiles. Initial experimental results demonstrate that online bioaerosl mass spectrometry provides an alternative method for online analysis of the bioaerosol. In addition, this development has the potential for online bioaerosol sampling and analysis. |
| 内容类型 | 学位论文 |
| 源URL | [http://ir.rcees.ac.cn/handle/311016/34501]  |
| 专题 | 生态环境研究中心_大气环境科学实验室 |
| 推荐引用方式 GB/T 7714 | 梁苗. 在线生物气溶胶质谱仪的研制及应用研究[D]. 北京. 中国科学院研究生院. 2014. |
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