2.1 General practices and analytical methods
2.1.1 Sampling and laboratory practices
During the collection of soil and pore water samples, attention was paid to minimize both the disturbance of the natural site and the influence of atmospheric oxygen on the samples. Soil samples were usually collected with an auger or, if larger amounts were required, with a spade, packed into bags, and either flash-frozen on dry ice or stored under cooled conditions, depending on the planned analyses or experiments. Collection of pore water samples for studies 1 and 3 was done with pore water peepers type Hesslein (1976). Oxygen influence on the water samples was minimized by conducting transport and storage of the peepers in a nitrogen-filled box. In studies 1 and 3, soil air composition (CO2 and oxygen) at the respective sampling sites was analyzed in approximately 10 to 20 cm depth using a portable gas analyzer (BIOGAS 5000, Geotech). For all experiments and sample preparations, ultrapure water and reagents of at least analytical degree were used. Oxygen-sensitive experiments (study 3) were conducted inside an anoxic chamber (Coy, 95%/5%
nitrogen/hydrogen).
For most analyses, the soil samples were freeze-dried, ground, and sieved (nylon, 1 mm mesh size). If the soil samples were used for analyses of potentially oxygen-sensitive parameters, i.e., for Mössbauer spectroscopy (study 2), X-ray absorption spectroscopy (XAS), 13C nuclear magnetic resonance spectroscopy (NMR), scanning electron microscopy energy-dispersive X-ray spectroscopy (SEM-EDS; all study 3) or for sequential extraction (study 1), preparation and storage was done inside an anoxic chamber. In addition, samples were stored under dark conditions to reduce photooxidation effects.
2.1.2 Analytical methods with respect to metal(loid)s
For the determination of total metal(loid) and sulfur contents, soil samples were digested in aqua regia using microwave-assisted digestion at 160 °C (MARS Xpress, CEM). Pore water and other liquid phase samples were stabilized in 0.45% H2O2 and 0.65% HNO3 directly after sampling. Analysis of total sulfur and metal(loid) concentrations (Al, As, Cd, Co, Cr, Cu, Fe, Mn, Ni, Zn) in soil digests and in pore water or any other liquid phase samples was conducted by inductively coupled plasma mass spectrometry (ICP-MS, X-Series 2, Thermo Scientific). All samples were filtered (0.2 µm, cellulose-acetate) and diluted at least 1:10 before ICP-MS analysis.
In study 1, a five-step sequential extraction procedure according to Fulda et al. (2013a) was conducted in order to gain information on operationally defined metal(loid) fractions in soil and thus on dominating binding mechanisms. This sequential extraction is a combination of the methods established by McGrath et al. (1988), Tessier et al. (1979), and Zeien and Brümmer (1989). The
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procedure included the following operationally defined fractions: (1) mobile (soluble and exchangeable, soluble metal(loid)-SOM complexes), (2) easily mobilizable (specifically adsorbed, bound to minerals labile at pH 5, and weak metal(loid)-SOM complexes), (3) organically bound (low affinity), (4) reducible (bound to amorphous and crystalline Fe and Mn (oxyhydr)oxides), (5) oxidizable (metal sulfides and bound to high affinity sites of SOM). To exclude any reactions induced by atmospheric oxygen, extraction steps 1 to 3 were conducted inside an anoxic chamber. Total metal(loid) contents in filtered (0.2 µm, cellulose-acetate) and diluted extracts were determined by ICP-MS.
In study 3, solid-phase speciation of Cu in natural and Cu-spiked soil samples was investigated using XAS at the Cu K-edge. Both X-ray absorption near edge structure (XANES) and extended X-ray absorption fine structure (EXAFS) spectra were collected at beamlines 4-1 and 4-3 of the Stanford Synchrotron Radiation Light Source (SSRL, Stanford, U.S.A.). Linear combination fitting was conducted both for normalized XANES and k3-weighted EXAFS sample spectra, using nine published spectra and one measured spectrum of Cu reference compounds. Details on spectra collection and data evaluation can be found in study 3. In addition, SEM-EDS data were collected in study 3 in order to detect any potential Cu-containing precipitates using a Leo Gemini 1530 (Carl Zeiss, Germany) with a Schottky emitter.
2.1.3 Methods used for soil and pore water characterization
Various other methods were used to characterize soil and pore water parameters, which are relevant for metal(loid) binding and mobility. The most important methods will briefly be mentioned in this chapter. Analytical details can be found in the individual studies.
Liquid samples were generally characterized by measuring pH and redox potential directly in solution or in soil suspensions. Methane and CO2 concentrations were determined by calculating dissolved concentrations from head space concentrations in septum vials using Henry’s law (Henry, 1803, Sander, 2015). Therefore, concentrations in the head space were measured with a gas chromatograph (SRI Instruments 8610C, U.S.) equipped with a methanizer and a flame ionization detector. In studies 1 and 3, additional photometrical analyses were conducted using a multiplate reader (Infinite 200 PRO, Tecan) and the ferrozine assay (Stookey, 1970) for determination of total Fe and ferrous Fe concentrations (absorbance measured at 570 nm), the methylene blue method (Cline, 1969) for determination of sulfide concentrations (absorbance measured at 650 nm), and the bathocuproine method (Maurer et al., 2013, Moffett et al., 1985) for determination of cuprous Cu (absorbance measured at 492 nm). Total dissolved organic carbon (DOC) and dissolved nitrogen concentrations were measured in filtered (0.45 µm, polyamide) liquid phase samples using thermo-catalytic oxidation with a TOC-VCPN Analyzer (Shimadzu). In study 3, DOM was characterized by Fourier-transform
Methods
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infrared spectroscopy (FTIR) with a Vector 22 spectrometer (Bruker Optik) in transmission mode using freeze-dried water extracts of soil samples.
Soil samples were generally characterized by measuring soil pH in 0.01 M CaCl2 solution at a soil-to-solution ratio (SSR) of 1:2.5. In addition, total soil organic carbon (SOC) and nitrogen contents were determined with a CHN elemental analyzer (Thermo Quest, Flash EA, 1112). In study 2, three soil samples were analyzed by Mössbauer spectroscopy in order to gain information on Fe mineralogy.
Spectra were collected at 77 K and 5 K using a closed-cycle exchange gas cryostat (Janis cryogenics) and a constant acceleration drive system (WissEL) in transmission mode with a 57Co/Rh source. In study 3, solid-phase SOM of soil samples was characterized by 13C NMR with an Avance III HD Spectrometer (Bruker) and soil mineralogy was characterized by X-ray powder diffraction (XRD) using a Philips X’Pert Pro diffractometer operating in reflection mode with Fe filtered CoKα1
radiation.
2.2 Spatial scale: determination of metal(loid) distribution patterns around a mofette (study 1)
In the first study of this thesis, the spatial distribution of the metal(loid)s Al, As, Cd, Co, Cr, Cu, Fe, Mn, Ni, and Zn in soil and pore water around the mofette Site A was determined. Soil samples were collected along a sampling grid with its center located in the degassing center and its margins in the non-CO2-influenced area. The samples derived from three depths representing the SOM-rich topsoil (0 to 5 cm depth), the alternating oxic/anoxic zone (25 to 35 cm), and the hypothetically permanently water-saturated zone (55 to 65 cm) of the Gleyic Fluvisol in the non-CO2-influenced area. To gain information on possible metal(loid) binding mechanisms, a five-step sequential extraction according to Fulda et al. (2013a) was conducted for the soil samples from the east transect. In addition, pore water samples from 10 to 60 cm depth were collected along the east transect of the sampling grid using pore water peepers type Hesslein (1976). Spatial distribution patterns of general soil and pore water parameters as well as total metal(loid) contents were plotted using universal Kriging interpolation.
Calculation of Euclidian distances between normalized distribution patterns allowed to cluster the investigated metal(loid)s into groups of high similarity. By comparing general soil parameters with metal(loid) contents and fractionation derived from sequential extraction, conclusions on CO2-induced long-term processes and on dominant binding mechanisms could be drawn for each individual metal(loid).
2.3 Temporal scale: time dependency of carbon dioxide induced metal(loid) mobilization in soil (study 2)
The second study of this thesis focused on the temporal scale of CO2-induced metal(loid) mobilization processes. Therefore, hitherto non-CO2-influenced soil from 20 to 30 cm depth of Site A (alternating oxic/anoxic zone) was incubated in triplicate with filtered (0.2 µm, cellulose-acetate) water from the
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river Plesná (used as pore water equivalent) in batch reactors and purged with a gas flow of 28 ± 5 mL CO2 min-1 for up to 42 days. For the first experiment, incubation temperature was varied (16, 22 and 35 °C) while in the second experiment there was a variation in initial soil conditions (fresh spring soil, stored spring soil, and fresh summer soil). Over the course of the experiment, the liquid phase was sampled and analyzed for general parameters (pH and redox potential) and for total concentrations of As, Cu, Fe, and Mn. Metal(loid)-specific mobilization rates could be calculated from concentration changes observed over time. The amounts of cultivatable heterotrophic aerobic, Fe(III)-reducing, and sulfate-reducing microorganisms were determined by most probable numbers (MPN).
Differences within the temperature treatments enabled the distinction between abiotic and biotic processes. Differences within the initial soil conditions helped to elucidate the influence of SOM content and Fe (oxyhydr)oxide crystallinity, determined by Mössbauer spectroscopy, on metal(loid) mobilization rates.
2.4 Case study on copper: sorption behavior and solid-phase speciation of copper in mofettes (study 3)
Study 3 of this thesis focused on Cu, since this element had shown some quite interesting behavior in Mehlhorn et al. (2014) and in study 2 of this thesis. Besides investigating Cu soil contents and Cu pore water concentrations with respect to dissolved CO2 along the east transect of Site A, a Cu spike experiment was conducted. Therefore, SOM-rich topsoil (5 to 15 cm depth) from 4 spots along the transect of Site A (‘Mofette A’, ‘Transition A1’, ‘Transition A2’, and ‘Reference A’) as well as from 2 spots of Site B (degassing center ‘Mofette B’ and non-CO2-influenced soil ‘Reference B’) were collected. The homogenized, fresh soil samples were incubated with 5 mM NaCl solution, pH 4.5, as background electrolyte at a SSR of 1:30 and spiked with up to 11.7 mmol kg-1 CuCl2 solution. Copper adsorption coefficients (Freundlich and linear coefficients) were calculated for the sorption isotherms, which could be obtained from this experiment. X-ray absorption spectra (XANES and EXAFS) were collected at the Cu K-edge for natural and Cu-spiked samples in order to elucidate the solid-phase speciation of Cu in these samples. In addition, a Cu-loaded soil sample was investigated by SEM-EDS in order to detect any potential Cu precipitates. The SOM and DOM composition of the respective soil samples were characterized by 13C NMR and FTIR, respectively.
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