RCEES OpenIR  > 环境化学与生态毒理学国家重点实验室
单颗粒电感耦合等离子体质谱进样系统的改进和方法应用
孙玉贞
Subtype博士后
Thesis Advisor江桂斌 ; 胡立刚
2019-12
Degree Grantor中国科学院生态环境研究中心
Place of Conferral北京
Degree Name理学博士
Degree Discipline生物学
Keyword单颗粒电感耦合等离子体质谱,进样系统,纳米金,二氧化硅包覆纳米金,体内分布 sIngle Particle Inductively Coupled Plasma Mass Spectrometry, Sample Introduction, Aunps, Au@sio2, Distribution In Vivo
Abstract

         纳米材料因自身尺寸而具有独特的性质,使其在工业、日用品和医疗保健等领域有着广泛应用。因此,不可避免的导致其释放在环境中,引起了人们对其的环境危害、生态系统影响以及人体健康效应评价的热切关注。纳米技术的持续发展需要纳米剂量学为其提供可靠而准确的分析技术来定量和表征纳米材料。近几十年,已经发展了商业化的检测仪器和标准方法来表征纳米材料。其中单颗粒电感耦合等离子体质谱(single particle inductively coupled plasma-mass spectrometry, spICP-MS)是一种新兴的稳定而准确的测定纳米颗粒的方法,可以用来表征和定量环境浓度下的金属基纳米颗粒的粒径大小、组成成分、颗粒的质量浓度和数量浓度等。spICP-MS还可以直接检测溶液中的纳米颗粒和溶解态的离子信号而不需要进行分离预处理,可以精确检测环境浓度样品中的颗粒粒径和粒径分布。虽然spICP-MS对于纳米材料的分析显示出巨大潜力,但是依然存在着一些挑战需要解决。样品的基质干扰和传输效率是影响spICP-MS检测纳米颗粒的灵敏度和应用范围的重要影响因素,为了提高传输效率,减少基质影响,扩大应用范围,本项目通过改进spICP-MS的进样系统,提高了传输效率,减少了基质影响,并采用改进的spICP-MS方法检测了血液样品中纳米颗粒随时间的动态变化,为纳米材料在医药领域的应用提供了分析方法,同时还分析了类似大气颗粒物的硅包覆金纳米颗粒在小鼠体内的分布,为研究大气细颗粒物的毒性提供研究依据。
       论文的研究主要包括以下四个方面:
       (1) spICP-MS进样系统的改进
       spICP-MS的进样系统会直接影响纳米材料的测定,因为包括将纳米颗粒样品传输到等离子体、将样品气化成气溶胶以及样品进入等离子体的比例等都直接由进样系统决定。为了提高传输效率,减少样品基质对检测影响,扩大样品检测范围,本项目采用microFAST自动进样系统取代蠕动泵进样,原有蠕动泵进样,样品流速只能选择0.3 mL/min和0.5 mL/min,而采用microFAST自动进样系统,样品流速可从0.005 mL/min调节到5 mL/min,最小进样量可低至20 μL。同时采用无挡流板的旋流雾化室取代传统的双通道雾化室以及带挡流板的雾化室,可以大大的提高传输效率,一般带挡流板的雾化室在样品流速小于等于10 μL/min时,传输效率大约在30 %左右,而本项目采用无挡流板雾化室的传输效率可以达到92.9 %。同时,本项目改进的进样系统减少了样品基质对检测的影响,减化了样品前处理过程,扩大了spICP-MS的样品分析范围。
       (2) spICP-MS方法检测不同形状的纳米金颗粒
       spICP-MS作为一种新型的颗粒物分析技术,简化了复杂的样品前处理过程,同时还可以分析颗粒物的粒径大小、质量浓度、颗粒浓度以及对应的化学成分,获得多维信息。但是目前spICP-MS对纳米颗粒的分析,主要基于球形纳米颗粒获得粒径大小和分布,对于其他形状的纳米颗粒研究还鲜少报道。而纳米线和纳米棒等其他形状的纳米材料因为其特殊的性能,已经越来越广泛的应用在医药和工业等多种领域。为了评价非球形纳米材料的环境行为和毒性效应,需要研究spICP-MS方法对不同形状纳米颗粒的表征和定量分析。本项目采用spICP-MS方法对不同粒径的纳米金球颗粒和不同长径比的纳米金棒进行分析,结合UV-Vis和TEM方法对纳米金球和纳米金棒进行表征,来探究纳米颗粒形状对spICP-MS分析方法的影响。
      ( 3) spICP-MS方法检测不同时间血液样品中纳米金颗粒
       spICP-MS是一种灵敏度高、分析时间短且针对金属基纳米颗粒进行表征的新型分析方法。目前已经广泛应用在环境、食品以及制造业等诸多领域。研究纳米颗粒在血液中的动态变化对于理解其在体内的吸收和代谢至关重要,血液中纳米颗粒的浓度随时间的变化可以揭示出纳米颗粒是否可以穿越生物学屏障以及是否会沉积到脏器中等信息。然而,因为血液样品一般其体积较小,浓度较低并且需要用生物酶或者生物碱来对生物样品进行硝解。样品硝解的过程可能会影响纳米颗粒的离子化程度和传输效率。自动进样系统以及改进的旋流雾化室可以直接分析稀释后的血液样品而不用硝解,有效地减少了硝解给样品带来的误差,提高了工作效率。本研究采用改进的spICP-MS方法,对暴露了三种不同浓度5 μg/L、250 μg/L和500 μg/mL AuNPs的小鼠血液进行测定,结果表明在30 s后血液中的大部分AuNPs已经被代谢,只有暴露浓度为500 μg/mL的小鼠血液中检测到浓度为32 μg/mL的AuNPs;当对暴露后血液中的AuNPs颗粒随时间的变化进行分析时,研究发现在暴露后1小时左右,血液中的纳米颗粒浓度达到峰值,此后随着时间的增加,血液中纳米颗粒的浓度逐渐减少。该方法的回收率为111.74%,方法可信。
       (4) spICP-MS方法检测二氧化硅包覆纳米金颗粒在小鼠体内的分布特征
       颗粒物在人体内的吸收、分布、蓄积和代谢对于了解颗粒物的毒性机制至关重要,为了了解颗粒物在人体内的吸收和分布,本文采用二氧化硅包覆纳米金颗粒对小鼠进行呼吸暴露,解剖后采用spICP-MS分析其在体内的分布。结果表明Au@SiO2颗粒会部分沉积在肺部,其余会穿过肺泡-毛细血管屏障进入血液循环,并迁移到心脏和肝脏中,最终可能引起这些脏器的病变。纳米颗粒在动物体内的迁移和分布除了受传统因素包括颗粒分布到组织器官的速率等影响之外,还会由暴露途径和纳米特征等因素决定。研究Au@SiO2纳米颗粒在动物体内的分布有助于了解其在体内的靶向作用和行为,同时因为其表面为二氧化硅,与大气环境中的二氧化硅颗粒表面性质一致,由此亦可为大气中类似粒径的二氧化硅颗粒进入动物体内后可能的迁移和分布规律提供研究依据。
        本项目通过改进单颗粒电感耦合等离子体质谱的进样系统,提高了雾化效率,扩大了样品检测范围,为spICP-MS的广泛应用提供了研究基础。同时,本研究应用改进的spICP-MS方法检测了不同时间血液样品中纳米金颗粒的浓度,为研究纳米颗粒作为药物或者药物载体在血液中的释放情况提供了研究方法;进一步采用该方法研究了类大气颗粒物的二氧化硅包覆纳米金颗粒在小鼠体内的分布情况,为研究大气细颗粒物进入人体后的分布规律提供了研究基础。
 

Other Abstract

       Nanomaterials have unique properties due to their size, which makes nanomaterials widely use in industries, commodities, medical and other fields. The widespread use of nanomaterials inevitably leads to their release in the environment, which has caused people's eager attention to its environmental hazards, ecosystem impacts and health effects. The development of nanotechnology requires nanodosimetry to provide reliable and accurate analytical techniques to quantify and characterize nanomaterials. In recent decades, commercial instrumentation and standard methods have been developed to characterize nanomaterials. Single particle inductively coupled plasma-mass spectrometry (spICP-MS) is a promising method for determination of nanoparticles. It can directly detect nanoparticle and dissolved ion signals in solution without separation pretreatment, which can also accurately detect particle size, distribution, composition, mass concentration and number concentration in environmental concentration samples. Although spICP-MS shows great potential for the analysis of nanomaterials, there are still some challenges. The sample matrix interference and transmission efficiency are important factors influencing the sensitivity and application range of spICP-MS. In order to improve the transmission efficiency, reduce the matrix influence and expand the application range, this research improves the spICP-MS sample introduction system. The improved method increases the transmission efficiency, reduce the influence of matrix. The improved spICP-MS method was employed to detect the dynamic changes of nanoparticles in blood samples with time, which provided an analytical method for the application of nanomaterials in the field of medicine. Moreover, spICP-MS also was used to analyzed the distribution of AuNPs@SiO2 in mice. The AuNPs@SiO2 is similar to fine atmosphere particles. Its distribution in mice provides useful information for studying the toxicity of fine particulate matter in the atmosphere.
The research mainly includes the following four aspects:
       1. Improvement of spICP-MS sample introduction
       The sample introduction of spICP-MS directly affects the determination of nanomaterials, including the transfer of nanoparticles to the plasma, the gasification of the sample into an aerosol, and the proportion of sample entering the plasma. In order to improve the transmission efficiency and reduce the influence of sample matrix, this research uses microFAST automatic sample introduction system to replace peristaltic pump. The sample flow rate is 0.3 mL/min or 0.5 mL/min with peristaltic pump. While using the microFAST autosampler system, the sample flow rate can be adjusted from 0.005 mL/min to 5 mL/min, and the minimum injection volume can be as low as 20 μL. At the same time, the cyclonic spray chamber without baffle is used to replace the traditional two-channel spray chamber and the spray chamber with baffle, which can greatly improve the transmission efficiency. Generally, the transmission efficiency of the spray chamber with the baffle is about 30% when the sample flow rate less than 10 μL/min. However, the transmission efficiency of the baffle spray chamber without baffle can reach 92.9 % at the similar flow rate. At the same time, the improved sample introduction of this project also reduces the influence of the sample matrix on the detection, reduces the sample pretreatment process, and expands the sample analysis range of spICP-MS.
      2. Detection of AuNPs with different shapes by spICP-MS
       As a powerful nanoparticle analysis technology, spICP-MS reduces the complexity of sample preparation process, and also analyzes particle size, mass concentration, particle concentration and corresponding chemical composition to obtain multidimensional information. However, the current analysis of nanoparticles by spICP-MS is mainly based on the size and distribution of spherical nanoparticles, which is rarely reported for other shapes of nanoparticles. Nanomaterials such as nanowires and nanorods have been widely used in various fields such as medicine and industry because of their special properties. In order to evaluate the environmental behavior and toxic effects of non-spherical nanomaterials, it is necessary to study the characterization and quantitative analysis of different shapes of nanoparticles by spICP-MS method. In this project, the nano-gold spheres with different particle sizes and gold nanorods with different aspect ratios were synthesized firstly, and then UV-Vis and TEM methods were employed to characterize the shape and size. Finally, the concentration and size were characterized and quantified by spICP-MS, which provides useful information for the detection of nanoparticles with different shapes.
       3. Dynamic changes in nanoparticles in blood samples detected by spICP-MS
       spICP-MS is a new analytical method with high sensitivity, short analysis time and characterization of metal nanoparticles. It has been widely used in many fields such as environment, food and manufacturing. The dynamic changes of nanoparticles in the blood are essential for understanding their absorption and metabolism in the body. The concentration of nanoparticles in the blood changes over time to reveal whether the nanoparticles can cross the biological barrier and whether they will deposit into the organ. However, biological samples are generally small in size, low in concentration, and require biological enzymes or alkaloids to nitrate the biological samples. The process of nitration may affect the ionization and transport of nanoparticles. The introduction system and the improved cyclone chamber allow direct analysis of the diluted blood sample without digestion, effectively reducing the error caused by digestion. The swirling spray chamber without the baffle can greatly increase the transmission efficiency while allowing a large biological cell to enter, reducing the pretreatment steps of the blood sample. In this work, the blood of mice exposed to three different concentrations of 5 μg/L, 250 μg/L, and 500 μg/mL AuNPs was measured by the spICP-MS method of the improved injection system. The results showed that the blood was 30 s later. Most of the AuNPs in the blood have been metabolized, and only 32 μg/mL were detected in the blood of mice exposed to 500 μg/mL. The study shown that the concentration of nanoparticles in the blood peaked about 1 hour after exposure, and then the concentration of nanoparticles in the blood gradually decreased with time. The recovery rate of this method is 111.74%, and the method is credible.
       4. Distribution characteristics of silica-coated gold nanoparticles in mice detected by spICP-MS
       The absorption, distribution, accumulation and metabolism of particulate matter in vivo are essential for understanding the toxicity mechanism of particulate matter. In order to understand the absorption and distribution of particulate matter in vivo, the mice were exposed to breath by Au@SiO2 particles. After dissection, the distribution in vivo was analyzed by spICP-MS. The results shown that the Au@SiO2 particles partially deposited in the lungs, and some passed through the alveolar-capillary barrier to enter the blood circulation and migrated to the heart and liver, which may eventually cause lesions in these organs. The distribution of nanoparticles in animals is influenced by factors such as exposure pathways and nanofeatures, in addition to traditional factors including the rate at which particles are distributed to tissues and organs. Studying the distribution of Au@SiO2 nanoparticles in vivo helps to understand their targeting and behavior in vivo, and because the surface is silica, it is consistent with the surface properties of silica particles in the atmosphere. It reveals the possible migration and distribution of silica particles with similar particle size in the atmosphere, and provides a basis for studying the migration and biodistribution of silica particles in animal fine particles.
       By improving the sample introduction system of single particle inductively coupled plasma mass spectrometry, this project has improved the transmission efficiency and expands the detected range of sample species. At the same time, spICP-MS method has been used to detect the concentration of gold nanoparticles in blood samples at different times, providing a research method for studying the release of nanoparticles in blood as drugs or drug carriers. The method was also used to study the distribution of silica coated gold nanoparticles like as atmospheric particulate matter in mice, which provided a basis for studying the distribution of atmospheric fine particulate matter into human body and the changes in blood concentration.

Pages87
Language中文
Document Type学位论文
Identifierhttp://ir.rcees.ac.cn/handle/311016/42321
Collection环境化学与生态毒理学国家重点实验室
Recommended Citation
GB/T 7714
孙玉贞. 单颗粒电感耦合等离子体质谱进样系统的改进和方法应用[D]. 北京. 中国科学院生态环境研究中心,2019.
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