Title Page
Contents
ABSTRACT 15
Chapter 1. Introduction 17
1.1. Research background and objectives 17
1.2. Scope and structure of the dissertation 18
Chapter 2. Literature review 20
2.1. Environmental monitoring of micro(nano) plastics (MNPs) 20
2.1.1. MNPs in natural environment 20
2.1.2. Detection and characterization methods for MNPs 23
2.1.3. Modeling approaches for automated interpretation of MNP data 27
2.1.4. Microfluidic system for MNP analyses 33
2.2. Impact assessment of MNPs 34
2.2.1. Ecological impacts on the soil environment 34
2.2.2. Ecological impacts on the aquatic environment 35
2.2.3. Fates in natural environments 36
Chapter 3. Monitoring system for automated quantification and size detection of MNPs 38
3.1. Introduction 38
3.2. Materials and Methods 40
3.2.1. Preparation of spherical and fragmented MNPs 40
3.2.2. Preparation of the microfluidic system 41
3.2.3. Automated analyses of MNPs under fluorescence microscope and particle tracking algorithms 43
3.3. Results and Discussion 45
3.3.1. Morphological analyses based on fluorescence microfluidic system 45
3.3.2. Automated quantification and size detection of MNPs 47
3.3.3. Performance evaluation: Particle concentration versus mass concentration 49
3.3.4. Performance evaluation: Automated quantification versus manual quantification 51
3.3.5. Performance evaluation: Dispersion in natural waters 51
3.4. Summary 54
Chapter 4. Effects of MNPs on soil microbial communities 55
4.1. Introduction 55
4.2. Materials and Methods 56
4.2.1 Characterization of polystyrene (PS) MNPs 56
4.2.2. Soil sampling and incubation 58
4.2.3. Soil microbial biomass analysis 59
4.2.4. Inorganic nitrogen analysis 59
4.2.5 Extracellular enzyme assays 59
4.2.6. Analysis of soil bacterial community composition 60
4.2.7 Statistical analyses 61
4.3. Results and Discussion 62
4.3.1. Effects of PS MNPs on soil microbial biomass 62
4.3.2. Effects of PS MNPs on soil inorganic N and soil enzyme activity 62
4.3.3 Effects of MNPs on the composition of soil bacterial communities 66
4.4. Summary 71
Chapter 5. Effects of MNPs on aquatic microbial communities 72
5.1. Introduction 72
5.2. Materials and Methods 73
5.2.1. UV aging and protein adsorption of PS MNPs 73
5.2.2. Exposure test 74
5.2.3. Microbial responses upon exposure to pristine and aged MNPs 76
5.2.4. Statistical analyses 78
5.3. Results and Discussion 79
5.3.1. Characteristics of pristine and aged MNPs 79
5.3.2. Microbial responses of pure culture 82
5.3.3. Microbial responses of freshwater microbial communities 85
5.3.4. Cellular interaction between MNPs and E. coli 87
5.3.5. Different patterns of microbial responses of pure culture and freshwater microbial communities 89
5.4. Summary 92
Chapter 6. Fates of MNPs in soil environments under climate change 93
6.1. Introduction 93
6.2. Materials and Methods 94
6.2.1. Preparation of pristine and aged MNPs 94
6.2.2. Transport experiments in water-saturated porous media 95
6.2.3. Determination of attachment efficiency 97
6.2.4. Statistical analyses 97
6.3. Results and Discussion 97
6.3.1. Characteristics of pristine and aged MNPs 97
6.3.2. Effects of aging on the transport of MNPs 99
6.3.3. Effects of water type saturated in the porous media on the transport of MNPs 104
6.3.4. Effects of temperature on the transport of MNPs 105
6.4. Summary 108
Chapter 7. Fates of MNPs in aquatic environments under climate change 109
7.1. Introduction 109
7.2. Materials and Methods 110
7.2.1. Preparation of pristine and aged MNPs 110
7.2.2. Sedimentation experiments 111
7.2.3. Determination of aggregation kinetics and interaction energy 112
7.2.4. Statistical analyses 113
7.3. Results and Discussion 113
7.3.1. Characteristics of pristine and aged MNPs 113
7.3.2. Aggregation kinetics of pristine and aged MNPs 116
7.3.3. Stability of MNPs in natural waters 122
7.3.4. Effects of water temperature on the stability of MNPs 127
7.4. Summary 130
Chapter 8. Conclusions 131
References 133
Appendices 158
[Appendix 1] CO₂ uptake monitoring system 158
[Appendix 2] CO₂ release monitoring system 161
Abstract (in Korean) 164
Table 2-1. Characteristics and physicochemical properties of MNPs observed in environmental samples. 21
Table 4-1. Zeta potentials and hydrodynamic diameters of PS MNPs (average ± one standard error; n=3). 57
Table 7-1. Characteristics of natural waters (average ± one standard error; n=2). 126
Figure 1-1. The structure of the dissertation. 19
Figure 2-1. Particle size detection range of analytical equipment adopted for analyzing MNPs (A: Lorenzo-Navarro et al., 2021, 2020, B: Renner et al., 2017, C:... 22
Figure 3-1. Preparation steps for creating a microchannel using PDMS (a). Schematic comparison between the general and current design of a microfluidic device (b and c). 42
Figure 3-2. Microfluidic device (a) and microscopic image of microchannel (b). Red box indicates the observation region for data acquisition (180×110 μm). Acquisition... 42
Figure 3-3. Data processing steps for automated quantification and size detection using particle tracking algorithms. Process includes background subtraction and gray... 44
Figure 3-4. Fluorescence image of (a) 500-nm PS beads, (b) 1000-nm PS beads, and (c) fragmented PS and PVC MNPs. A single frame was extracted from the video record of MNPs passing through the microchannel. 46
Figure 3-5. Fluorescence images of PS beads with size ranging from 100 to 5000 nm. The minimum size detection limit of the microfluidic system was 200 nm. 47
Figure 3-6. Particle concentrations and diameters of MNPs estimated using automated particle tracking algorithms (a-c). Particle size distribution of MNPs (d-f). 48
Figure 3-7. Linear regression analysis showing the relationship between the particle concentration obtained from automated quantification and the mass concentration of MNPs. The inset graph shows the magnified results for mass concentration between 0.1... 50
Figure 3-8. Correlation analyses performed to assess the relationship between automated quantification and manual quantification results of MNPs. The inset graph shows the magnified results for manual quantification concentrations within the following ranges:... 52
Figure 3-9. Particle diameters of (a) 500-nm PS beads, (b) 1000-nm PS beads, and (c) fragmented MNPs dispersed in deionized (DI), lake, ground, river, and seawater are depicted in the box plot. The plot was created using a whisker model within the range of 1.5 times... 53
Figure 4-1. Electron microscopy images of PS MNPs of three different sizes. Scanning electron microscopy (SEM) images of (a) 0.05-, (b) 0.5-, and (c) 5-μm PS... 57
Figure 4-2. Microbial biomass (µg C g¯¹ soil) in soils treated with PS MNPs of three different sizes during the 40-d incubation period. Different lowercase letters indicate... 62
Figure 4-3. Inorganic N concentration (µg g¯¹ soil) in soils treated with the three sizes of PS MNPs at day 1. Different lowercase letter indicates significant differences... 63
Figure 4-4. Activities of β-1,4-glucosidase, xylosidase, β-1,4-N-acetyl-glucosaminidase, acid phosphatase, and cellobiohydrolase associated with C, N, and... 65
Figure 4-5. Differences in the relative abundance of bacterial groups in PS MNP- treated soils. The difference in the relative abundance between control soils and those... 67
Figure 4-6. Heatmap of the 25 most abundant genera of bacterial communities in soils treated with PS MNPs. Each row illustrates treatments of different sizes and... 69
Figure 4-7. Principal coordinate analysis (PCoA) graphs of microbial communities in soils based on the Bray-Curtis distance at the genus level. Soils treated with three sizes... 70
Figure 5-1. Natural water was collected in June 2022 from Ilgam Lake on the Konkuk University campus, in Seoul, Republic of Korea (a and b). For the exposure test, 400... 75
Figure 5-2. Characteristics of pristine and aged PS MNPs. TEM images of pristine (a), UV-aged (b), BSA-coated (c), and UV+BSA-aged and coated (d) PS MNPs. FT-... 80
Figure 5-3. Zeta potentials and hydrodynamic diameters of pristine and aged MNPs. Zeta potentials of MNPs dissolved in deionized (DI) water (a) and natural water (b).... 81
Figure 5-4. Microbial responses of pure culture upon exposure to pristine and aged PS MNPs shown as (a) oxidative stress expressed by the level of ROS generation, (b) membrane damage, (c) metabolic activity, (d) growth rate, and (e) DNA damage. Letters (a, b, c)... 84
Figure 5-5. Microbial responses of freshwater microbial communities upon exposure to pristine and aged PS MNPs shown as (a) oxidative stress expressed by the level of... 86
Figure 5-6. Extracellular enzyme activities of (a) β-1,4-N-acetylglucosaminidase (NAG), (b) β-1,4-glucosidase (BG), (c) cellobiohydrolase (BC), and (d)... 87
Figure 5-7. Cellular interactions between MNPs and E. coli analyzed by TEM (a - d), and fluorescence microscopy (e - h). The yellow arrow indicates PS MNPs stained... 88
Figure 5-8. Multidimensional scaling plots generated based on the microbial responses of pure culture (a) and freshwater microbial communities (b) upon exposure... 90
Figure 6-1. Regression curve for the concentration of 500-nm PS MNPs (mg L¯¹) and the absorbance at a wavelength of 270 nm. 96
Figure 6-2. Characteristics of pristine and aged PS MNPs. SEM images of pristine (a), UV-aged (b), BSA-coated (c), UV+BSA-aged and coated (d), and BSA+UV-aged... 99
Figure 6-3. Breakthrough curves of pristine and aged MNPs in porous media saturated with seawater (a-c), groundwater (d-f), and DI water (g-i) at 20, 25, and 30℃. 101
Figure 6-4. Retention profiles of pristine and aged MNPs after their transport in porous media saturated with seawater (a-c), groundwater (d-f), and DI water (g-i) at 20, 25, and 30℃. 102
Figure 6-5. Attachment efficiencies of pristine and aged MNPs in porous media saturated with seawater (a), groundwater (b), and DI water (c) at 20, 25, and 30℃. The bar indicates one standard error (n=3). Letters (a, b, c) indicate significant differences among... 103
Figure 6-6. Breakthrough curves of pristine (a, f, and k), UV-aged (b, g, and i), BSA-coated (c, h, and m), UV+BSA-aged and coated (d, i, and n), and BSA+UV-aged (e, j, and o) MNPs in porous media at 20, 25, and 30 ℃. 106
Figure 6-7. Effects of water temperature on the zeta potential of MNPs. Measurement was conducted after dispersing MNPs in 100 mM NaCl solution. The bar indicates... 107
Figure 7-1. Regression curve for the concentration of 500-nm PS MNPs (mg L¯¹) and the absorbance at a wavelength of 270 nm. 112
Figure 7-2. Characteristics of pristine and aged PS MNPs. FT-IR spectra of pristine and aged MNPs (a). Zeta potential (b) and hydrodynamic diameter (c) of pristine and... 114
Figure 7-3. Zeta potential of pristine and aged MNPs in NaCl (a) and CaCl₂ (b) solutions. Zeta potential (c) and hydrodynamic diameter (d) of pristine and aged... 115
Figure 7-4. Sedimentation profiles of pristine and aged MNPs in NaCl solutions with concentration of 100 to 900 mM at 20, 25, and 30℃. 118
Figure 7-5. Sedimentation profiles of pristine and aged MNPs in CaCl₂ solutions with concentration of 10 to 90 mM at 20, 25, and 30℃. 119
Figure 7-6. Aggregation kinetics of pristine and aged MNPs as a function of electrolyte concentrations. Attachment efficiency (α) of pristine and aged MNPs in NaCl solutions (a-c) and CaCl₂ solutions (d-f) at water temperatures of 20, 25, and 30 ℃. 120
Figure 7-7. DLVO energy profiles of pristine and aged MNPs in NaCl and CaCl₂ solutions at 25℃. 121
Figure 7-8. Short-term sedimentation profiles of pristine and aged MNPs in river water (a-c), lake water (d-f), groundwater (g-i), and seawater (j-l) at water temperatures of 20, 25, and 30 ℃. The bar indicates one standard error (n=3). 123
Figure 7-9. Long-term sedimentation profiles of pristine and aged MNPs in river water (a-c), lake water (d-f), groundwater (g-i), and seawater (j-l) at water temperatures of 20, 25, and 30 ℃. The bar indicates one standard error (n=3). 124
Figure 7-10. Attachment efficiencies (a-d) and DLVO energy profiles (e-h) of pristine and aged MNPs in natural waters at 20, 25, and 30℃. The bar indicates one standard error (n=3). Letters (a, b, c) indicate significant differences among MNP groups (p<0.05). 125
Figure 7-11. Attachment efficiencies of pristine and aged MNPs dispersed in 600 mM NaCl (a) and 30 mM CaCl₂ solutions at 20, 25, and 30℃. 128
Figure 7-12. Attachment efficiencies of pristine and aged MNPs dispersed in river water (a), lake water (b), groundwater (c), and seawater (d). 128
Figure 7-13. DLVO energy profiles of pristine and aged MNPs in 100 mM NaCl solution at water temperatures of 20, 25, and 30℃. The lower row graph illustrates the results obtained by magnifying the red box depicted in the upper row graph. 129