Soil analyses

• Soil moisture
• Chlorophyll a
• Inorganic nitrogen
• Ammonium and nitrate
• Total N, total C, organic and inorganic C
• pH
• Electrical conductivity

Gravimetric Soil Moisture

Use analytical balance with precision to 0.1 mg.

1. Pre-weigh aluminum weighing dish.
2. Without taring dish, add field moist soil and weigh.
3. Place into drying oven at 105°C for 24 hours.
4. Remove oven-dry soil and place in desiccator to cool down. Weigh.
5. Calculate soil moisture:

Chlorophyll a

A. Sampling and extraction

1. Sample from top 5 mm of soil only.
2. All procedures are carried out in the dark to prevent degradation of chlorophyll.
3. Mix thoroughly and place 10 g soil in a 20 ml screw cap vial.
4. Add 10 ml of 90% acetone to each vial and swirl once gently.
5. Place vials in 5°C incubator overnight.
6. Adjust solution to pH 9.0. The solution must be basic because acidic chlorophyll contains pheophytin which will interfere with the spectrophotometric analysis of the chlorophyll.
7. Filter solution through a Whatman GF/C filter and place in a screwcap test tube. Keep refrigerated.

B. Determination of Chlorophyll a concentration

1. This procedure is carried out using a spectrophotometer.
2. Blank the spec with a 90% acetone solution.
3. Place 3 ml of extracted solution in a cuvette. Measure the absorbance at 664 nm and 750 nm. This must be done fairly quickly in order to prevent light from breaking down the chlorophyll.
4. After taking the initial measurements, add 0.1 ml of 1N HCl directly to the cuvette and agitate gently. Wait 90 seconds, then re-measure the absorbance at 665 nm and 750 nm. The acid will convert the chlorophyll to pheophytin by releasing a magnesium ion in an acidic environment.
5. Use the following equations to calculate the concentration of chlorophyll in your soil sample:

chlorophyll a (mg/m3) =

where

V1 = volume of extract in liters

V2 = volume of soil sample in m3

L = light path length of width of cuvette, in cm

664b= optical density of 90% acetone extract before acidification

= (absorbance at 664 nm) - (absorbance at 750 nm)

665a = optical density of 90% acetone extract after acidification

= (absorbance at 665 nm) - (absorbance at 750 nm)

C. References

1. Standard Methods for the examination of Water and Wastewater, Vol. 18, pages 10-18 to 10-19
2. The seasonal periodicity of algae on Antarctic fellfield soils. Holarctic Ecology 14: 112-120. 1991
3. Chlorophyll a determination of periphyton on sediments: identification of problems and recommendation of method. Freshwater Biology 20: 347-352. 1988.
4. A note on the extraction of chlorophyll from benthic algae using methanol. Aech. Hydrobiol. Beih. Ergebn. Limnol. 14: 88-90. 1980

KCl Extraction for Inorganic Nitrogen (Keeney and Nelson, 1982)

1. Weigh out 8 g air-dried, 2 mm sieved soil into a 125 ml Erlenmeyer flask.
2. Dispense 40 ml of 2M KCl into each Erlenmeyer flask including blanks.
3. Cover Erlenmeyer flasks with parafilm and place on a shaker. Shake for 1 hr on low speed setting.
4. Pour liquid through Whatman #40 filter paper.

NH4+ and NO3- determination

Ammonium and nitrate concentrations were determined using a Flow-Injection Autoanalyzer made by Lachat Instruments (Milwaukee, Wisconsin), model Quikchem AE System. [Lachat is now Zellweger Analytics, Inc., Lachat Instruments Division.] The instrument analyzes for NO3--N and NH4+-N colorimetrically as mg l-1 in KCl solution. The final results are presented as mg kg-1, where mg is NO3--N or NH4+-N and kg is soil mass.

Keeney, D.R. and D.W. Nelson. 1982. Nitrogen-Inorganic forms. In : A.L. Page (ed.). Methods of soil analysis. Chemical and microbiological properties. Agronomy 9 (2): 643-698. Am. Soc. of Agron., Madison, WI.

Total nitrogen, Total carbon, Organic carbon, Inorganic carbon

These parameters were analyzed using an elemental analyzer made by Carlo Erba Instruments (Milan, Italy). The instrument uses a combustion method converting all forms of N or C into N2 and CO2 gases. The N2 and CO2 gases are separated by gas chromatography and analyzed by a thermal conductivity detector. The final results were presented as unit mass of N or C in soil per unit mass of soil, expressed as g kg-1

Soil samples were dried and ground prior to analysis for total N and total C determination. For organic C determination, soil samples were pretreated with HCl to remove carbonate and bicarbonate, the major components of inorganic C. Inorganic C was determined by subtracting the organic C from the total C.

pH

These measurements were made in a 1:2 water suspension: one part of soil in two parts of deionized water without filtration. We used a Hach One Laboratory pH Meter and Model 44200 Combination Electrode (Hach Company, Loveland, Colorado). The measurements were taken at 22°C.

Electrical Conductivity

These measurements were made in a 1:5 water suspension: one part of soil in five parts of deionized water. The solution was centrifuged at 1000 g for 10 minutes to settle out particulates. We used an Orion Model 160 Conductivity Meter and Model 016010 4-Electrode Conductivity Cell (Orion Research Incorporated, Boston, Massachusetts). The measurements were recorded as dS m-1. All readings were adjusted to 25°C.

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