Approximately 1500g of soil or sediment were collected into Whirl-Pak(tm) bags with clean plastic scoops from 5 sections (0, 8, 16, 24, and 32 m away from the stream) along each transect. Five grams of sediment/soil were also collected from the top 1 cm for measurement of chlorophyll-a concentrations. Samples were mixed within the bags, placed into insulated coolers, and transported for analysis to the McMurdo laboratories. Until processing, the soils were stored in a 4_C refrigerator. Samples were analyzed by ASA Analytical Services on January 6th 1998.
Extraction of chlorophyll from the soil. All procedures were carried out in the dark or very low irradiance to avoid degradation of the chlorophyll. The soil samples were mixed thoroughly in the vials, and a sample of approximately 5 g was weighed out in to a 50 mL plastic centrifuge tube with a screw-top cap. 10 mL of a 50:50 DMSO/90% acetone solution was added to each sample and they were mixed thoroughly on a bench-top Vortex mixer for about 5 seconds. The vials were placed in a -4C constant temperature room, in the dark, and left for 12-18 hours.
Determination of chlorophyll a concentration. This was determined fluorometrically using a Turner model 111 fluorometer. A calibration using a known concentration of chlorophyll was carried out prior to sample analysis. The machine was blanked using a 50:50 DMSO/90% acetone solution. Each vial was mixed thoroughly, then centrifuged for 5 minutes at about 1800 RPM. A sample of approximately 4mL of the DMSO/acetone solution was taken from the top of the sample with a pipette, being careful not to get any soil particles in the solution. The sample was placed in a cuvette, in to the fluorometer and the fluorescence was recorded . This was done fairly quickly in order to prevent light from breaking down the chlorophyll. This measurement is called Fo, the initial fluorescence. After taking this reading, 0.1 mL of 1N HCl was added directly to the cuvette and the cuvette was gently agitated. After 20 seconds, the fluorescence was re-measured.
(During this step, the acid converts the chlorophyll to phaeophytin by releasing a magnesium ion in an acidic environment). This measurement is called Fa, the fluorescence after acidification. The solution was discarded in to a waste container, and the cuvette rinsed 3 times with DMSO/90% acetone solution before proceeding with the next sample. Data were received at NREL on disk and hard copy in the spring of 1998.
