Removing recently deposited and incompletely decomposed plant material from soil samples reduces the influence of temporary seasonal inputs on soil organic carbon measurements. Attraction to an electrostatically charged surface can be used to quickly remove a substantial amount of particulate organic matter.
Estimations of soil organic carbon are dependent on soil processing methods including removal of undecomposed plant material. Inadequate separation of roots and plant material from soil can result in highly variable carbon measurements. Methods to remove the plant material are often limited to the largest, most visible plant materials. In this manuscript we describe how electrostatic attraction can be used to remove plant material from a soil sample. An electrostatically charged surface passed close to dry soil naturally attracts both undecomposed and partially decomposed plant particles, along with a small quantity of mineral and aggregated soil. The soil sample is spread in a thin layer on a flat surface or a soil sieve. A plastic or glass Petri dish is electrostatically charged by rubbing with polystyrene foam or nylon or cotton cloth. The charged dish is passed repeatedly over the soil. The dish is then brushed clean and recharged. Re-spreading the soil and repeating the procedure eventually results in a diminishing yield of particulates. The process removes about 1 to 5% of the soil sample, and about 2 to 3 times that proportion in organic carbon. Like other particulate removal methods, the endpoint is arbitrary and not all free particulates are removed. The process takes approximately 5 min and does not require a chemical process as do density flotation methods. Electrostatic attraction consistently removes material with higher than average C concentration and C:N ratio, and much of the material can be visually identified as plant or faunal material under a microscope.
Accurate estimates of soil organic carbon (SOC) are important in evaluating changes resulting from agricultural management or the environment. Particulate organic matter (POM) has important functions in the ecology and physics of a soil but it is often short lived and varies based on several factors including season, moisture conditions, aeration, sample collection techniques, recent soil management, vegetation life cycle, and others1. These temporally unstable sources can confound estimates of long-term trends in stable and truly sequestered soil organic carbon2.
Despite being well-defined, common, and important, POM is not easily separated from soil nor is it easy to measure quantitatively. Particulate organic matter has been measured as that which floats in liquids (light fraction, typically 1.4-2.2 g cm-3), or as that which can be separated by size (e.g., > 53-250 µm or > 250 µm), or a combination of the two3,4,5. Both size-based and density-based techniques can influence the quantitative and chemical outcomes of POM measurement4. A careful visual inspection of soil that has been size-fractionated using routine methods often reveals long, narrow structures like roots and slivers of leaf or stem that have passed through the screen. Simply removing these structures by hand has been shown to substantially reduce measurements of total SOC2,6 but the method is notably subject to the diligence and visual acuity of the operator. POM separation from a soil sample as the light fraction during flotation in a dense liquid7 does not capture all POM, and excessive shaking during the flotation process can actually reduce the amount of light fraction recovered from a sample8. Flotation requires many steps and exposes the soil to chemical solutions which can change the chemical characteristics or dissolve and remove constituents that may be of interest4.
Alternative methods for removing POM have been used to avoid or augment the use of dense aqueous solutions. Kirkby, et al.6 compared light fraction removal using two flotation procedures to a dry sieving/winnowing method9. Winnowing was performed by passing a light current of air across a thin layer of soil to gently lift away the light from the heavy fraction. The dry sieving/winnowing performed similarly to the two flotation methods with regard to C, N, P, and S content; however, the authors suggest that dry sieving/winnowing produced "slightly cleaner" soils6. POM has also been separated from soil using electrostatic attraction10,11 in which organic particles are isolated by passing an electrostatically charged surface above the soil. The electrostatic attraction method successfully recovered POM, referred to as course organic particles, from dried, sieved (> 0.315 mm) soils with statistical repeatability comparable to other methods of size and density fractionation10.
Here we demonstrate how electrostatic attraction can be used to remove POM of sizes ranging from visible to microscopic. Unlike other reported methods, electrostatic attraction of fine soil also removes a small portion of mineral and aggregated soil which is visibly like the remaining soil. Given our results to date, it is reasonable to assume that the removal of a small portion of non-POM soil will have no substantial effect on the downstream analyses; however, this assumption should be verified for a specific soil if large proportions of the total soil sample are being removed electrostatically. The methods and examples provided here were performed on silt loam loess soils from a semi-arid environment.
This method may not be suitable for all soil types but has the advantages of being quick and efficient in removing particulate organic matter too small to remove manually or by an air current. Process speed is important in reducing fatigue, ensuring consistency, and encouraging greater replication for better accuracy of conclusions. Additionally, the ability to remove very small particulates is important in avoiding bias toward soils with larger rather than small particulate sizes.
1. Soil preparation
- Collect soil samples to the desired depth. Thoroughly dry the soil at 40 °C or following lab-specific standard protocols.
- Sieve the soil through appropriate-sized soil sieves to obtain approximately 10-25 g of sieved soil. Many studies use a 1- or 2-mm sieve. The amount of soil is based on the mass required for the downstream analyses and will impact the number of times the electrostatic removal step will need to be repeated.
- Place the soil in a clean, dry metal or glass flat-bottomed pan that is large enough for soil to be spread thin (at least 20 cm in diameter). Gently shake the pan horizontally to distribute the soil evenly in as thin a layer as possible.
2. Charge an electrostatic surface
- Hold a 100 mm diameter glass or polystyrene Petri dish top or bottom in one hand and vigorously rub the outer surface with a clean piece of nylon cloth, cotton cloth, or polystyrene foam several times. Perform the surface charging away from the sample to prevent fabric fragments from being introduced into the sample.
- Inspect the surface of the Petri dish to make sure it is clean.
3. Remove particulate organic matter
- Lower the charged surface to within 0.5 cm to 2 cm above the soil and move it horizontally to pick up as much particulate material as possible. Attraction to the surface can be noted visually and audibly.
- When the Petri dish no longer attracts additional particles, move the dish away from the sample.
4. Clean the electrostatic surface
- Hold the charged surface over a collection dish and use a fine brush to transfer the electrostatically attracted material from the Petri dish surface into the collection dish. A camel hairbrush works well.
5. Repeat until the yield of particulates decreases
- Repeat steps 2 through 4 until the number of organic matter particles being picked up decreases. Redistribute the soil sample by horizontal shaking of the soil pan to expose new material at the surface and continue electrostatic collection.
NOTE: The endpoint is arbitrary and depends on the judgement of the researcher. Inspection of the charged surface after exposure to the soil gives a visual indication of whether a significant amount of organic particulates are still being removed from the soil. The final products are soil with reduced particulate content, and concentrated POM containing a small amount of electrostatically removed soil.
The results presented here are based on the analysis of silt loam soils from agricultural sites in the Pacific Northwest (Table 1). Soils were collected to depths of 0-20 cm or 0-30 cm, dried at 40 °C, passed through a 2 mm sieve, and treated using a polystyrene surface charged with a nylon cloth.
The amount of soil electrostatically removed from a sample varied. About 1% to 6% of the total soil mass was removed (Table 2). In all cases the proportion of total sample C removed was greater than the soil mass removed. Also, the C concentration and C:N ratio of the electrostatically removed soil fraction was always greater than the remaining soil. These factors indicate that the method reduced the amount of incompletely decomposed organic substances.
The ambient conditions and the combination of materials used to produce the charged surface affected the results (Table 3). The electrostatic removal method is expected to be less effective in a more humid lab environment due to lower surface charges. All materials should be as dry as possible for the electrostatic process. Nylon is a good material for electrostatic charging because it is lint-free and, when used with polystyrene Petri dishes, should produce one of the greatest electrostatic charges12. Alternatively, some types of polystyrene foam work well in combination with glass. The combination of a glass dish and polystyrene foam removed a greater amount of soil and C than either glass(dish)/cotton or polystyrene(dish)/nylon combinations.
Regardless of the materials used for surface charging, electrostatic treatment removed a greater proportion of C from the soil and produced a sample with lower C:N ratio as compared to the forceps/winnowing method although differences were only significant with glass/foam. Comparatively, flotation was more effective than electrostatic treatment in removing concentrated particulate C from the sample as noted by the lowest C:N ratio of the remaining sample and greatest C:N of the removed fraction.
The electrostatic treatment can be repeated numerous times albeit the treatments will begin to remove greater proportions of soil due to diminishing amounts of particulates attracted to the dish surface. The effects of treatment endpoints were examined by collecting a series of three electrostatic samples one after the other from the same soil sample (Table 4). The first treatment collected the greatest amount of C and although the following two treatments collected less, both were still highly enriched in C compared to the remaining soil. The C:N ratio decreased in the removed fraction indicating greater proportions of soil to POM were removed with each successive step.
When performing the ES procedure using a polystyrene Petri dish, scratches on the polystyrene dish surface were visible, suggesting the possibility that C from the plastic dish could contaminate the soil samples. When the ES treatment was performed on washed, C-free sand using a polystyrene dish, there was no detectable C in ES fractions even after four repeated treatments on the same ES fraction (data not shown).
Finally, the amount of particulate material that could be electrostatically removed from the fine silt-size fraction that passed through a 53 µm screen was tested on five silt loam soils (Table 5). The electrostatically removed fractions demonstrated very little enrichment of particulate organic matter. Microscopic inspection reveals that POM does exist in the <53 µm fraction of these soils (Figure 1), but in very small quantities. If the fine soil fraction (i.e., <53 µm) contains very little POM, that fraction can be removed prior to electrostatic treatment to reduce the amount of soil being treated. Sieve the soil over a very fine sieve, such as 53 µm. Remove the soil from the top of the sieve and place in the tray for the electrostatic treatment, or simply use the sieve as the tray for spreading the sample. Return the fine fraction (soil passed through the sieve) to the electrostatically treated soil prior to chemical analysis.
|Soil||Soil Type||Management||Collection depth||Mean Annual Precipitation (mm)||Location|
|Thatuna||Thatuna silt loam (fine-silty, mixed, mesic Xeric Argialboll)||Wheat/fallow||0-30 cm||450||Pullman, WA|
|Ritzville-R||Ritzville silt loam (coarse-silty, mixed, superactive, mesic Calcidic Haploxeroll)||Wheat/fallow||0-30 cm||301||Ritzville, WA|
|Ritzville-E||Ritzville silt loam (coarse-silty, mixed, superactive, mesic Calcidic Haploxeroll)||Wheat/fallow||0-30 cm||290||Echo, OR|
|Walla Walla-M||Walla Walla silt loam (coarse-silty, mixed, superactive, mesic Typic Haploxeroll)||Wheat/fallow||0-30 cm||282||Moro, OR|
|NT-AW||Walla Walla silt loam (coarse-silty, mixed, superactive, mesic Typic Haploxeroll)||No-tillage annual winter wheat||0-20 cm||420||Pendleton, OR|
Table 1: Soils tested. List of samples used to compare the electrostatic process for particulate organic matter removal.
|Soils||Reps||Fraction||Proportion of total||C||N||C:N||Estimated POM C:N|
|Thatuna||10||Removed||0.01 (0.00)||0.05 (0.01)||54.02 (4.33)||2.85 (0.15)||18.68 (0.62)||24.39 (0.55)|
|Remainder||14.52 (0.15)||1.25 (0.01)||11.58 (0.11)|
|Ritzville-R||5||Removed||0.02 (0.01)||0.08 (0.03)||36.24 (3.29)||2.61 (0.21)||13.83 (0.16)||16.01 (0.15)|
|Remainder||9.61 (0.24)||0.95 (0.01)||10.10 (0.18)|
|Ritzville-E||8||Removed||0.02 (0.00)||0.07 (0.01)||36.73 (3.10)||2.65 (0.24)||13.89 (0.17)||15.94 (0.32)|
|Remainder||7.31 (0.10)||0.78 (0.01)||9.40 (0.07)|
|Walla Walla-M||5||Removed||0.02 (0.00)||0.04 (0.00)||15.88 (0.55)||1.17 (0.04)||13.54 (0.21)||17.37 (0.91)|
|Remainder||7.86 (0.05)||0.71 (0.01)||11.15 (0.20)|
|NT-AW||6||Removed||0.06 (0.01)||0.18 (0.02)||63.20 (9.25)||3.81 (0.47)||16.32 (0.50)||19.75 (0.49)|
|Remainder||15.7 (0.31)||1.40 (0.03)||11.21 (0.09)|
Table 2: Representative removal rates. The amount of soil in the electrostatically removed fraction (Removed) and the remaining soil fraction reduced in particulates (Remainder) as a proportion of the total sample mass and as the proportion of the total sample C. Also given are the concentrations of C, N and C:N. The estimated POM C:N gives the calculated C:N of the Removed fraction in excess of the concentrations in the Remainder, which is presumably the C:N of the POM removed. Numbers in parentheses are standard error of the mean. Analysis of variance indicated that Removed was greater than Remainder for both C and C:N (p > F of less than 0.0001). Replicates indicates the number of sample replicates per value. The electrostatic separation was performed with a polystyrene dish charged with nylon cloth after sieving out the fine fraction (<53 µm).
|Method†||Fraction||Proportion of total removed||C||N||C:N|
|ES polystyrene/nylon||Removed||0.03 (0.01)||0.08 (0.01)||31.34 (4.21)||1.95 (0.15)||15.99 (1.07)|
|Remainder||14.07 (0.35) ab||1.23 (0.02) ab||11.40 (0.18) ab|
|ES glass/cotton||Removed||0.04 (0.01)||0.10 (0.01)||28.20 (2.32)||1.87 (0.13)||15.08 (0.49)|
|Remainder||14.12 (0.32) ab||1.23 (0.02) ab||11.47 (0.12) ab|
|ES glass/foam||Removed||0.08 (0.02)||0.13 (0.03)||24.59 (2.85)||1.74 (0.11)||14.10 (1.11)|
|Remainder||13.95 (0.20) bc||1.20 (0.01) bc||11.60 (0.15) ab|
|ES glass/foam, humid||Removed||0.05 (0.01)||0.12 (0.02)||31.34 (4.58)||2.03 (0.2)||15.40 (0.75)|
|Remainder||13.96 (0.36) bc||1.23 (0.03) ab||11.30 (0.13) b|
|Forceps/Winnow||Removed||0.03 (0.01)||0.05 (0.01)||25.84 (2.61)||1.61 (0.09)||16.10 (1.40)|
|Remainder||14.86 (0.57) a||1.25 (0.04) a||11.90 (0.42) a|
|Flotation, 1.7 g cm3||Removed||0.01 (0.00)||0.10 (0.01)||141.28 (15.63)||7.63 (0.62)||18.50 (0.58)|
|Remainder||13.19 (0.58) c||1.18 (0.02) c||11.10 (0.50) b|
|Whole soil||14.50 (0.52) ab||1.25 (0.02) a||11.60 (0.44) ab|
|† ES combinations are noted as the composition of the dish followed by the charging surface. Foam is polystyrene.|
Table 3: Technique comparison. Removal of particulate organic matter from Thatuna soil using electrostatic attraction (ES), manual removal of visible particles with forceps and air (Forceps/winnow), and flotation on sodium iodide solution at 1.7 g cm-3. Electrostatic attraction was performed with a polystyrene dish charged with a nylon cloth, or a glass surface charged with a cotton cloth or polystyrene foam. Glass/foam was also tested under humidified conditions. Manual removal of particulates was performed by gently blowing air over the surface of a thinly spread soil to move it to the side and removing the visible residue with forceps. Data are the mean of six replicates. Means followed by a common letter are not significantly different according to the Tukey test at the 5% level of significance.
|Fraction||Proportion of total||C||N||S||C:N||Estimated POM C:N|
|1st treatment||0.01 (0.00)||0.04 (0.01)||48.70 (6.67)||2.93 (0.41)||0.27 (0.03)||16.6 (0.96) a||21.0 (1.88)|
|2nd treatment||0.01 (0.00)||0.03 (0.01)||32.07 (3.56)||2.30 (0.28)||0.23 (0.03)||14.1 (0.63) ab||18.4 (1.89)|
|3rd treatment||0.01 (0.00)||0.03 (0.01)||32.48 (4.68)||2.45 (0.40)||0.25 (0.04)||13.4 (0.46) bc||16.7 (1.29)|
|Remainder||0.60 (0.04)||0.60 (0.04)||12.02 (1.46)||1.11 (0.11)||0.14 (0.02)||10.8 (0.29)|
|< 53 µm fraction||0.37 (0.04)||0.03 (0.03)||9.51 (1.13)||0.96 (0.08)||0.11 (0.02)||9.7 (0.45)|
Table 4: Investigation of endpoints. Results of three successive electrostatic treatments to remove particulate organic matter. Average of three samples from the Thatuna soil and one each from the Ritzville-R, Ritzville-E, Walla Walla-M soils. The soil fraction passing through a 53 µm sieve was removed before electrostatic treatment and analyzed separately. Data are the mean of the six analyses with the standard error in parentheses. Analysis of variance produced p = 0.06 for both C and estimated POM C:N. Letters in the C:N column show significant differences between successive treatments at p < 0.05.
|Soils||Fraction||Proportion of mass||C||N||C:N||Difference in C:N|
Table 5: Particulate organic matter in the fine soil fraction. Test of electrostatic particulate removal on the fine fraction (<53 µm) of five soil samples from wheat cropping systems. An analysis of variance of Removed versus Remainder was not significant for C and C:N. The difference in C:N was not consistently greater in the removed fractions.
Figure 1: Visual identification of particulate organic matter. Microscopy images of the NT-AW soil as (A) whole soil, (B) removed fraction on the charged polystyrene surface, (C) <53 µm soil fraction, and (D) material that floated to the surface of a water slurry of the <53 µm fraction soil. Images were taken with 50x or 100x magnification. Images collected across several different focal points were combined in ImageJ software13 using the Stack Focuser plugin (https://imagej.nih.gov/ij/plugins/stack-focuser.html). Please click here to view a larger version of this figure.
The electrostatic attraction method was effective in removing POM from the silt loam soils. The method described here is slightly different from Kaiser, et al.10 which used a combination of glass/cotton. We treated all but the finest soil fraction and used polystyrene rather than glass due to the triboelectric difference, which for polystyrene/nylon is 100 nC/J compared to glass/ cotton at 20 nC/J12. Glass and polystyrene foam have proven effective and convenient in more recent experience. The relative humidity of the storage area and workspace could be an issue at some locations during certain seasons of the year. The methodology presented here was conducted in a workspace with consistently low (20% to 30%) relative humidity. Temperature would not be expected to change electrostatic attraction independent of humidity.
From our experience with the soils used for this research, the <53 µm soil can be sieved out of the sample before using the electrostatic process. Removal of the fine soil fraction prior to the electrostatic process seemed to improve attraction of particulates to the charged surface. Additionally, our soils did not appear to have significant quantities of particulates in the fine soil fraction, as indicated by its low C:N ratio. The electrostatic process was not effective at removing the organic particulates which were present in this soil fraction (Table 5). This might not be true of other soils.
Researchers need to consider whether they are willing to remove a small amount of mineral soil along with particulate organic matter. Theoretically, the non-organic particulate matter (mineral) soil and aggregates removed with the electrostatic fraction might be chemically different or be coated with organic matter of a different nature than the remaining soil sample that will be used for chemical analysis. If substantial amounts of mineral soil are being removed, a chemical comparison might be warranted.
Adequate removal of POM is an important process for soil C estimates. The electrostatic method has some advantages over other methods including dry removal and flotation. These advantages include the ability to remove very small particulates, reduce process time, and retain the POM fraction for additional analyses. This method may not be suitable for all soil types or ambient conditions thus researchers are encouraged to validate the method for their specific samples and conditions.
The authors have nothing to disclose.
This work was supported solely by USDA-ARS base funding. The authors greatly appreciate Mikayla Kelly, Caroline J. Melle, Alex Lasher, Emmi Klarer, and Katherine Son for their technical help.
|petri dish, glass or plastic|
|polystyrene foam, cotton or nylon cloth|
- Gosling, P., Parsons, N., Bending, G. D. What are the primary factors controlling the light fraction and particulate soil organic matter content of agricultural soils. Biology and Fertility of Soils. 49, (8), 1001-1014 (2013).
- Gollany, H. T., et al. Soil organic carbon accretion vs. sequestration using physicochemical fractionation and CQESTR simulation. Soil Science Society of America Journal. 77, (2), 618-629 (2013).
- Cambardella, C. A., Gajda, A. M., Doran, J. W., Wienhold, B. J., Kettler, T. A. Assessment methods for soil carbon. Kimble, J. M., Lal, R., Follett, R. F., Stewart, B. A. CRC Press. 349-359 (2001).
- Wander, M. Soil organic matter in sustainable agriculture. CRC Press. 67-102 (2004).
- Curtin, D., Beare, M. H., Qiu, W., Sharp, J. Does particulate organic matter fraction meet the criteria for a model soil organic matter pool. Pedosphere. 29, (2), 195-203 (2019).
- Kirkby, C. A., et al. Stable soil organic matter: A comparison of C:N:P:S ratios in Australian and other world soils. Geoderma. 163, (3-4), 197-208 (2011).
- Strickland, T. C., Sollins, P. Improved method for separating light- and heavy-fraction organic material from soil. Soil Science Society of America Journal. 51, (5), 1390-1393 (1987).
- Golchin, A., Oades, J. M., Skjemstad, J. O., Clarke, P. Study of free and occluded particulate organic matter in soils by solid state 13C Cp/MAS NMR spectroscopy and scanning electron microscopy. Soil Research. 32, (2), 285-309 (1994).
- Theodorou, C. Nitrogen transformations in particle size fractions from a second rotation pine forest soil. Communications in Soil Science and Plant Analysis. 21, (5-6), 407-413 (1990).
- Kaiser, M., Ellerbrock, R. H., Sommer, M. Separation of coarse organic particles from bulk surface soil samples by electrostatic attraction. Soil Science Society of America Journal. 73, (6), 2118-2130 (2009).
- Kuzyakov, Y., Biriukova, O., Turyabahika, F., Stahr, K. Electrostatic method to separate roots from soil. Journal of Plant Nutrition and Soil Science. 164, (5), 541 (2001).
- Lee, W. AlphaLab Inc., The Tribo-Electric Series. AlphaLab In, BC. (TriField.com). Available from: http://www.trifield.com/content/tribo-electric-series (2017).
- Schneider, C. A., Rasband, W. S., Eliceiri, K. W. NIH Image to ImageJ: 25 years of image analysis. Nature Methods. 9, (7), 671-675 (2012).