海洋地质与环境重点实验室知识库

微塑料作为一种新型的环境污染物，近年来受到日益关注。自2004年沉积物中的微塑料被报道以来，微塑料现已在近海海岸带以及深海环境中均被发现。南极海冰、马里亚纳海沟、海洋生物甚至是人类体内都有微塑料的存在。而对于海洋环境来说，海洋沉积物被认为是微塑料聚集的“汇”，然而海洋沉积物微塑料检测方法缺乏统一标准，且因研究地区不同而存在差异。选择合适的分析手段是研究海洋沉积物样品的关键，既要保证分析效率，又能够获得样品特征信息。

除光学目视鉴定法能够提供颗粒物理信息外，光谱学领域的傅里叶红外光谱法以及拉曼光谱法被视为常用的无损分析手段，能够实现微塑料的分子内部结构表征。傅里叶红外光谱法利用红外吸收光谱表征分子内部信息，拉曼光谱法通过拉曼散射反映分子结构信息。目前，海洋沉积物样品所需要的前处理环节颇为繁杂，需要进行有机质的酸碱消解，样品颗粒染色等步骤来辅助定性分析以及样品计数统计，而傅里叶红外光谱法对于小于20微米的微粒不能发挥良好的检测效果。本研究针对海洋沉积物中小粒径微塑料，基于共聚焦显微拉曼光谱技术，旨在改进微塑料样品处理步骤，通过实验室模拟以及实际海洋沉积物样品来探索快速高效的小粒径微塑料检测方法。

本研究建立了完备的微塑料拉曼光谱库，包含生活及工业用途的十八种塑料种类，并对其拉曼峰位进行详尽归属，用于与样品的拉曼特征峰进行比对，获得微塑料样品的分子结构特征。对于沉积物中微塑料样品前处理环节，通过改进抽滤装置并省略有机质消解步骤，以实现样品富集并保留样品原有形貌特征。通过模拟海洋沉积物环境进行微塑料回收实验，针对微塑料不同特性进行系统探究，得出适于拉曼光谱技术的分析参数。将此完整的检测流程应用于海洋沉积物样品中，以验证方法的有效性，得出研究区小粒径微塑料特征。对此，得出以下结论：

（1）拉曼光谱可以反映微塑料结构信息，并能够对海洋沉积物中获得的不同种微塑料做到特定区分。由于拉曼光谱反映出分子的振动特征，对于傅里叶红外光谱法不明显的非极性分子振动可以清晰反映，特别是对于微塑料中具有碳碳双键的物质，能够明显地表征出分子内部结构特征。其他振动类型如碳氢键振动、苯环呼吸振动、羰基振动等表现出的拉曼峰位，也可作为不同类型微塑料的特征来进行分析。

（2）共聚焦显微拉曼分析手段能够获得高信噪比的未经有机质消解的微塑料拉曼光谱。在实验室模拟海洋沉积物环境回收微塑料实验中，微塑料样品经密度浮选后未经有机质消解步骤，在拉曼光谱中仍显示清晰的拉曼信号，并能够实现微塑料类型的归属。此外，从海滩沉积物中分离出的微塑料样品，也能够未经有机质消解得到有效的拉曼光谱，并得出微塑料的类型、粒径、形貌等特征信息。

（3）激光功率、微塑料的粒径、微塑料表面颜料覆盖均会对分析结果产生不同影响，并表征在拉曼信号中。由于功率改变带来塑料表面温度的改变，同一粒径的微塑料在由低功率至高功率的检测过程中，拉曼光谱会出现形变。反之，不同粒径在同一功率下也会发生拉曼信号的偏移。当微塑料表面有颜料覆盖时，会掩盖原本的拉曼有效信号，对此，将特定颜料颗粒的光谱也纳入微塑料光谱库中，完备分析机制。

（4）基于建立的检测流程与拉曼分析参数，将小粒径微塑料分析流程应用到山东省青岛市汇泉湾海洋沉积物中，获得了汇泉湾海洋沉积物中粒径在500微米以下的微塑料样品信息，并且对粒径小于10微米的聚丙烯（PP）颗粒实现特征分析。结果表明，粒径在50微米以下的微塑料在500微米以下的样品中占有56%的比重，为探究研究区微塑料污染情况与海洋环境中微塑料的存在方式提供数据支持。

Microplastics has attracted increasing attention in recent years as a new type of environmental pollutant. Since the report of microplastics in sediment in 2004, microplastics have been found in offshore coastal zones and in the deep sea, in the Antarctic, the Mariana Trench, within marine organisms, and even in human bodies. As for the marine environment, marine sediments are hypothesized to be major sinks for the accumulation of microplastics. However, the detection methods for microplastics in marine sediments are lack of unified standards and vary according to the study area. For microplastics separated from sediments, the selection of appropriate detection and identification methods is of vital importance to the research, which could ensure the efficiency of detection and identification without damaging the samples.

Currently, in addition to optical visual identification could provide physical information of particles, Fourier transform infrared transform and Raman spectroscopy are regarded as nondestructive spectroscopic identification methods which can realize the characterization of the internal structure of molecules. FTIR characterize the structure of molecules according to infrared absorption spectrum, and Raman spectroscopy reflects the information of molecular structure by Raman scattering. However, at present, for the detection of microplastics in sediments, the pretreatment steps of samples are quite complicated, such as acid-base digestion of organic matter, staining of sample particles and other steps to assist qualitative identification and sample counting statistics. Additionally, FTIR spectroscopic method could hardly detect particles less than 20 μm, which would become the main constraint for the detection of microparticles in the lower size range. Based on confocal micro-Raman spectroscopy, this study aims to simplify the pretreatment procedure of microplastic samples and explore fast and efficient detection methods of smaller microplastics from marine sediment by applying simulation experiment and identification of actual marine sediments samples.

In this study, a complete Raman reference library of microplastics was established, including 18 kinds of plastics for commercial and industry use. Raman peaks were assigned in detail, which could be compared with the characteristic peaks of samples to acquire the qualitative characteristics of microplastics samples. For the pretreatment steps of microplastic samples in sediments, the filtration device was improved and the digestion of organic matter was omitted to achieve rapid detection. The recovery experiment of microplastics was carried out by simulating the sediment environment, in addition, the characteristics of microplastics were systematically summarized, and the detection parameters suitable for Raman technology were obtained. Furthermore, this simplified detection process was conducted to marine sediment samples to verify the effectiveness of the method. In this regard, the conclusions are as follows:

(1) Raman spectra reflect the structural information of microplastics, and different kinds of microplastics from marine sediment could be distinguished. Since the Raman spectra represents the vibration characteristics of molecules, the non-polar molecular vibrations which are not obvious for FTIR could be reflected clearly, especially for microplastics which have carbon-carbon double bonds. The internal structure of the molecule could be obviously characterized. Other Raman characteristic peaks, such as carbon-hydrogen bond vibration, benzene ring respiratory vibration and carbonyl vibration, could be characterized by different types of microplastics.

(2) Raman spectra of microplastics without digestion of organic matter with high signal-to-noise ratio could be obtained by confocal micro-Raman detection. In the environmental recovery of simulated sediments in the laboratory, the microplastic samples were extracted without digestion of organic matter, besides, the Raman signals were still clearly displayed, and the classification of microplastics could be realized. The Raman spectra of microplastic from the marine sediments without digestion of organic matter could still present high quality. Moreover, the characteristic information of microplastic type, particle size, shape could be reached.

(3) The laser power, the particle size of the microplastic and the pigment coating on the surface of the microplastic all have different effects on the detection, and could be characterized in the Raman signal. Due to the change of the plastic surface temperature caused by the change of power, the microplastics with the same particle size appear spectral deformation in the performance from low power to high power, on the contrary, the background level would change with different particle size at the same power. When the microplastic surface is covered with pigment, the original Raman effective signal would be masked. Therefore, the spectra of specific pigment particles should also be included in the reference library to ensure the completeness.

(4) Based on the established detection process and Raman analysis parameters, the detection process of microplastics with small particle size was applied to the marine sediments of Huiquan Bay. The characteristics of microplastics with a particle size of less than 500um in the marine sediment samples of Huiquan Bay were obtained, and the analysis of PP particles less than 10 microns was realized. The results indicate that microplastics with particle size less than 50 microns account for 56% of the samples with particle size less than 500 microns, which provides data support for exploring the pollution of microplastics in the study area and the existing pattern of microplastics in the marine environment.

第1 章 绪论 ......................................................................................... 1
1.1 课题研究背景 .............................................................................................. 1
1.2 微塑料现有的取样及分离处理方法 ............................................................ 2
1.2.1 取样方法 ............................................................................................... 2
1.2.2 分离方法 ............................................................................................... 3
1.2.3 样品前处理方法 .................................................................................... 3
1.3 微塑料现有的分析方法 ............................................................................... 4
1.4 微塑料研究中存在的问题 ........................................................................... 5
1.5 研究内容与意义 .......................................................................................... 5
1.6 研究创新性 .................................................................................................. 6
第2 章 共聚焦激光拉曼光谱技术 ...................................................... 9
2.1 拉曼光谱学发展历程 ................................................................................... 9
2.2 拉曼散射原理 .............................................................................................. 9
2.3 拉曼光谱技术特点及谱图特征 ..................................................................10
2.3.1 技术特点 ..............................................................................................10
2.3.2 拉曼谱图特征 ...................................................................................... 11
2.4 共聚焦显微拉曼光谱仪 ..............................................................................12
2.4.1 共聚焦原理 ..........................................................................................12
2.4.2 共聚焦显微拉曼光谱仪结构及特点 ....................................................13
2.5 拉曼光谱技术在微塑料领域研究应用 .......................................................14
第3 章 材料与实验方法 .................................................................... 17
3.1 海洋沉积物及微塑料样品的准备 ..............................................................17
3.1.1 海洋沉积物样品取样 ...........................................................................17
3.1.2 微塑料样品制备 ...................................................................................18
3.2 流程设置及实验装置 ....................................................................................20
3.2.1 处理流程设置 ......................................................................................20
3.2.2 实验装置 ..............................................................................................21
3.3 共聚焦显微拉曼光谱仪 ..............................................................................22
3.3.1 光谱仪参数 ..........................................................................................22

3.3.2 实验参数分析 ......................................................................................23
第4 章 微塑料拉曼光谱库的建立 .................................................... 25
4.1 拉曼选择定则与化学键振动 ......................................................................25
4.2 微塑料拉曼光谱库建立及峰位归属分析 ...................................................26
4.3 结果与讨论 .................................................................................................45
4.4 小结 .............................................................................................................46
第5 章 沉积物中微塑料颗粒的拉曼光谱模拟检测及干扰因素分析
............................................................................................................... 49
5.1 基于拉曼分析技术的微塑料模拟回收实验验证 .......................................49
5.1.1 类型分析 ..............................................................................................49
5.1.2 粒径分析 ..............................................................................................51
5.1.3 回收率分析 ..........................................................................................52
5.2 有机质消解对拉曼检测的影响 ..................................................................53
5.3 功率影响分析 .............................................................................................55
5.4 微塑料颗粒中颜料参杂与荧光干扰因素分析 ...........................................57
5.4.1 颜料覆盖的影响分析 ...........................................................................57
5.4.2 荧光干扰分析 ......................................................................................57
5.4.3 含颜料微塑料颗粒的拉曼光谱检测参数优化与数据库构建 .............58
5.5 结果与讨论 .................................................................................................59
5.6 小结 .............................................................................................................60
第6 章 海洋沉积物样品应用实例 ...................................................... 63
6.1 海洋沉积物环境微塑料研究意义 ..............................................................63
6.2 沉积物样品共聚焦显微拉曼光谱分析 .......................................................63
6.2.1 海滩实际样品获取与处理 ...................................................................63
6.2.2 微米量级的微塑料样品特征分析 ........................................................64
6.2.3 基于拉曼光谱的微塑料类型分析 ........................................................66
6.3 汇泉湾微塑料污染的粒径分布与对比 .......................................................70
6.4 结果与讨论 .................................................................................................71
6.5 小结 .............................................................................................................72
第7 章 结论和展望 ............................................................................. 73
7.1 主要结论 .....................................................................................................73
7.2 研究展望 .....................................................................................................74

参考文献 ............................................................................................... 75
致 谢 ................................................................................................... 89
作者简历及攻读学位期间发表的学术论文与研究成果 ..................... 91