4D imaging reveals mechanisms of clay-carbon protection and release

doi:10.21203/rs.3.rs-54393/v1

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4D imaging reveals mechanisms of clay-carbon protection and release

https://doi.org/10.21203/rs.3.rs-54393/v1

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published 26 Jan, 2021

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posted 12 Aug, 2020

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Soil absorbs about 20% of anthropogenic CO2 emissions annually, and clay is the key carbon-capture material. Although sorption to clay is widely assumed to strongly retard the microbial decomposition of soil organic matter, enhanced degradation of clay-associated organic carbon has been observed under certain conditions. The conditions in which clay inhibits microbial decomposition remain uncertain because the mechanisms of clay-organic carbon interactions are not fully understood. Here we reveal the spatiotemporal dynamics of carbon sorption and release within clay aggregates and the role of enzymatic decomposition by directly imaging a transparent smectite clay on a microfluidic chip. We demonstrate that clay-carbon protection is due to the quasi-irreversible sorption of high molecular-weight sugars within clay aggregates and the exclusion of bacteria from these aggregates. We show that this physically-protected carbon can be enzymatically broken down into fragments that are released into solution. Further, we suggest improvements relevant to soil carbon models.

Geochemistry

Climatology

Climate Analysis and Modeling

clay aggregates

soil organic matter

carbon sorption

CO2

There is NO Competing Interest.

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Supplementary Material
FigureS1.jpg
Structures of 2-NBDG glucose (a) and FITC dextran (b). Images are from (a) Thermo Fisher and (b) Sigma Aldrich websites. Using Dynamic Light Scattering in the Princeton Biophysics Core Facility, we estimated that the hydrodynamic radii of 3-5 kDa dextran, 20 kDa dextran, and 70 kDa dextran are 2 nm, 5 nm, and 6 nm, respectively. The hydrodynamic radius of 2-NBDG glucose was too small to measure. The linkage that the enzyme dextranase breaks down is indicated by the black arrow. The hydrodynamic radius of the dextranase is 7 nm.
TableS1.jpg
Molecular weights of the organic substances used in this study
FigureS2.jpg
Calibration of the average fluorescence intensity of clay relative to the calculated concentration of organic matter sorbed to clay. (a) Cross-sectional images of clay aggregates after soaking for three days in solutions with fluorescent organic matter (3-5 kDa dextran) of different concentrations. The clay areas, shown by the red contours, were identified as patches of pixels with fluorescence intensity larger than the average intensity of the whole image. (b) The average fluorescence intensity within the clay increases with the organic matter concentration in clay Cs in an approximately linear manner at Cs < 1 g/g, with correlation coefficient R = 0.6 (in log scale), and then reaches a plateau (fluorescence intensity ≈ 0.7). The black line represents the linear regression between the average clay intensity and Cs in log scale at Cs < 1 g/g. The linear to plateau relationship is consistent with the fact that fluorescence intensity generally increases with increasing concentration of the fluorescent labels before the intensity of the fluorescent image saturates38. The laser intensity used to excite the fluorescence was the same for all measurements. The error bars are the standard errors of multiple imaging measurements from the same sample (same batch experiment). Cs was calculated from the fluorescence intensity of the supernatant solution, Caq, before and after adding clay (see Methods). (c) The concentration of organic matter in solution, Caq (open circles), was interpolated from calibration curves of average fluorescence intensity versus concentration (dashed lines), which were derived by measuring the fluorescence intensity for solutions with known dextran concentration (squares) in three different ranges. Three different laser intensities were used to resolve the fluorescence intensity in these three ranges. Note that the fluorescence intensity in the clay cannot be directly compared with the fluorescence intensity in the solution because clay intensifies the emitted fluorescence intensity.
FigureS3.jpg
intensities of green FITC 3-5 kDa dextran in buffer solution with and without 2 g/L dextranse, i.e., Idextranase and Ino-dextranas, shows less than 20% difference, which is insignificant considering that the fluorescence intensity varies by one order of magnitude during the sorption and desorption experiment.
FigureS4.jpg
Clay-carbon sorption isotherms of 2-NBDG glucose and 70 kDa dextran. (a) The concentration of glucose sorbed to clay, Cs, increased with increasing concentration of glucose in the solution, Ce. The error bars are standard errors of multiple measurements from one sample. The scatter of the data, especially at low carbon concentration, is likely because Ce and Cs were estimated from the fluorescence intensity of the supernatant (see Methods and Fig. S2), which was difficult to measure accurately at low concentration when the fluorescent signal was comparable to the background noise. Despite the scatter, the Cs versus Ce data are roughly consistent with the expectation of a linear adsorption isotherm (exemplified by the dashed line with a slope of 1). (b) The concentration of 70 kDa dextran sorbed to clay, Cs, was essentially invariant with Ce values ranging over two orders of magnitude. The plateau loading of 70 kDa dextran is essentially identical to that of 3-5 kDa dextran (Fig. 100 2(a)), Cs-plateau = 0.2 ± 0.1 g/g, as expected if this plateau corresponds to the formation of a monolayer of high molecular-weight organic matter on the clay surface. Note that the upper range of Ce in this study was limited by the solubility of glucose and dextran in water, which is on the order of 10 g/L.
FigureS5.jpg
Reversible sorption of glucose to clay was consistently observed in replicate sorption/desorption experiments. The symbol and the error bar represent the mean and the standard error of the average fluorescence intensity within 5 representative clay aggregates in one channel. The selected 5 clay aggregates were indicated by the red boxes in Fig. S7 (b). The experiment is similar to the one shown in Fig. 1(e), except that the sorption period (the gray region) is about seven times longer than in Fig. 1. The decrease in fluorescence intensity of 2-NBDG glucose during the sorption period likely reflects photobleaching of the fluorescent molecules.
FigureS6.jpg
Quasi-irreversible sorption of 3-5 kDa dextran to clay was consistently observed in replicate experiments. The symbol and the error bar represent the mean and the standard error of the average fluorescence intensity within 5 representative clay aggregates in one channel. The selected 5 clay aggregates (or groups of clay aggregates) were indicated by the red boxes in Fig. S7 (120 c). The experiment is similar to the one shown in Fig. 1(d), except that the fluorescence intensity was scanned every 10 minutes, less frequently than the 1-minute scanning interval used in Fig. 1(d). Because of the lower scanning frequency, the photobleaching of dextran was slower than in Fig. 1(d) during the desorption period. Note that compared with Fig. 1(d), the sorption of the dextran did not reach equilibrium at 16 hours, perhaps because the clay aggregates were more densely packed in this experiment (Fig. S7(c)), suggesting that the kinetics of organic matter uptake depend on the microstructure of the clay aggregates. The type of sorption, i.e., quasi-irreversible sorption, was consistent with Fig. 1(d).
FigureS7.jpg
Cross sectional images showing the arrangements of clay aggregates in microfluidic channels. The red boxes in (a), (b), and (c) show the 5 clay aggregates used to estimate the fluorescence intensity of carbon for the experiments shown in Fig. 1 (e), Fig. S5, and Fig. S6, respectively. The channel width is 300 μm for (a) and 400 μm for (b) and (c).
FigureS8.jpg
Replicate bacteria-clay culture experiments consistently show that bacteria are excluded 135 outside clay aggregates. The images were scanned with different resolutions, e.g. 0.48 μm, 0.48 μm, 0.24 136 μm, 0.03 μm for (a), (b), (c), and (d), respectively.
FigureS9.jpg
Cross-sectional profiles of dextran fluorescence intensity in clay and the diffusivity ofdextrans in clay. (a) Cross-sectional fluorescent image of a microfluidic channel containing clay micro aggregates, the same experiment as in Fig. 3. The green and red fluorescence intensities along a transect, the z-axis in (a), were plotted over time in (b) and (c), respectively; z = 0 μm denotes the edge of the clay aggregate. The color of the intensity profiles corresponds to the time indicated in the right color bar. (d) Diffusion coefficients were estimated from the fronts of the propagating fluorescence profiles. The green and red lines show the linear fits of the data and the corresponding legends show the diffusivities estimated from the linear fit.
FigureS10.jpg
No further sorption of high molecular-weight carbon to clay was observed after enzyme penetrated into clay. (a) Cross-sectional fluorescent image of a microfluidic channel containing clay micro-aggregates. (b) Fluorescence intensity profiles along the red transect, the z axis, shown in panel (a) at different times; z = 0 denotes the edge of the clay aggregate; tC, tnoC, tenzyme, and tC2 represent the start times of the injections of 3-5 kDa dextran, no organics, dextranase, and 3-5 kDa dextran (second injection), respectively. (c) The symbols and error bars represent the mean and the standard error of the average fluorescence intensity within the four immobile clay regions, outlined by the four black contours in panel (a).

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4D imaging reveals mechanisms of clay-carbon protection and release

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