Climate Change and Mitigation REU - Summer 2014
Methods and Materials
- Materials:
For this experiment, we gathered soil samples from plots under a continuous corn system at the North Agronomy Farm located at Kansas State University, Manhattan, Kansas. The plots contain soil known as a fine-silty, mixed, superactive mesic Cumulic Hapludoll, which is most commonly referred to as a Mollisol. They were divided into four replicate plots per treatment combination and have been part of an ongoing carbon sequestration study since 1990.
Main plots: no-till (NT) and conventional till (CT)
Sub plots: high manure (HM), high fertilizer (HF), and control (C)
High Manure: cow manure from 1990-2002 and compost from 2002-present (manually applied)
High Fertilizer: urea (manually applied)
Control: no fertilizer
- Soil sampling and treatment:
Each plot had soil samples taken with a 2 cm in diameter auger. These samples were then placed into bags that were marked with the test plot information, treatment information and their three depth categories: 0-5 cm, 5-15 cm, and 15-30 cm. Our study only focused on the 0-5 cm and 5-15 cm depth. The samples were air dried in a greenhouse environment, after which they were passed through an 8 mm sieve stacked on a 4 mm one. The 4-8 mm stieved aggregates were collected into sterile plastic containers and stored at 4°C.
Different sized water stable aggregates were procured by wet sieving the samples with a device similar to the Yoder wet-sieving apparatus (Mikha and Rice, 2004). This machine has stacked sieves that divide the water stable aggregates into three different sizes: > 2000 µm, 2000-250 µm, and < 250 µm. The larger sized aggregates were transferred into beakers and stored, while the < 250 µm aggregates were sorted again into 250-53 µm and < 53 µm by manually passing them through a sieve with smaller opening sizes. Once the smaller aggregates were sorted, they were also transferred into beakers after which all beakers were oven dried at 40°C. The dried water stable aggregates were ground with a mortar and pestle and had any roots or organic residue removed. Each sample was handled separately. No composite, homogenous samples were made.
- Soil analysis:
Soil Mn, Fe, and Al concentrations were determined by the acid hydroxylamine extracting method as presented by Ross et al. 1985; Wang et al. 1987. This method, initially created because it allows for amorphous Mn to be extracted with minimal dissolution of Fe oxides such as magnetite (Chao and Zhou, 1983), was later modified by Ross et al. (1985) and Wang et al. (1987). They compared it to the oxalate extraction method and found that their method allows for removing non-crystalline materials, such as hydrous Mn oxides, without dissolving any magnetite while also giving results for Al and Fe similar to the oxalate method.
The procedure required 0.100 g of < 2 mm air-dry soil to be passed through a 0.15 mm sieve for each sample. The exact weight was noted down, and the soil was placed into a 50 mL Eppendorf centrifuge tube with screw-cap.
Hydroxylamine hydrochloride-hyrochloric acid (0.25 M NH2OH·HCl, 0.25 M HCl) solution was prepared by adding 21.5 mL of concentrated hydrochloric acid (HCl) to 17.37 g of hydroxylamine hydrochloride (NH2OH·HCl) into a 1 L flask and adding deionized water until the flask met the 1 L volume requirement. The reagent was shaken gently, after which 25 mL of the hydroxylamine solution was pipetted into each centrifuge tube.
For dissolution to occur, the tubes were sealed tightly and were placed in a horizontal shaker for 16 hours. The tubes were then placed into an Eppendorf machine and centrifuged for 20 minutes at 510 rpm. Each centrifuged tube had 10 mL of the solution decanted into a small plastic vial. When all tubes were decanted, the vials were frozen to stop any possible microbial activity that might alter the organic content so that later DOC analysis could be performed.
The remaining supernatant from each sample was filtered through 42 Whatman filter paper into separately labeled plastic vials and the vials were stored at 4°C until all samples are ready for amorphous Mn, Fe and Al extraction.
To analyze the amorphous Mn as well as the acid hydroxylamine extractable Fe and Al concentrations, the Inductively Coupled Plasma-Optical Emission Spectrometer (ICP-OES) was used. This popular analytical machine allows for a wide range of trace metal concentrations to be measured.
- DOC analysis:
It was of interest to understand the extend of DOC adsorption by Mn, Fe, and Al. To do this, select samples were chosen from the NTHM, CTHM, CTC, and NTC plots for both 0-5 cm and 5-15 cm depths.
Prior to preparing the samples, glass vials were washed in a 10% HCl solution, rinsed five times with deionized water, and air dried in a suction hood to avoid C contamination. Samples were then passed through a 0.2 µm syringe filter into glass vials, after which the filtered samples were diluted by pipetting 2 mL of the filtered sample with a 5 mL graduated pipette into a clean glass vial and adding 10 mL of deionized water. After that, the TOC-L: Combustion catalytic oxidation method was used. this technique allows for efficient oxidation of easily decomposed, low molecular weight organic compounds, as well as not easily decomposed, insoluble, macromolecular organic compounds.
The ICP-OES works by injecting liquid samples into a radiofrequency-induced argon plasma with a nebulizer. The mist that reaches the plasma is then dried, vaporized and energized very rapidly because of high temperature collisional excitation. The plasma then gives off an atomic emission that can be "viewed by a radial or axial configuration, collected with a lens or a mirror, and then imaged onto the entrance slit of a wavelength selection device" (Xiandeng Hou et al. 2000).
- TOC analysis:
Data for TOC was obtained by measuring the CO2 released from a heated sample via dry combustion with a LECO C/N analyzer. The samples were pretreated with 1 N phosphoric acid to remove inorganic carbon.
- Statistical analysis:
All results were analyzed with the Statistical Analysis System (SAS) 9.3.