This SOP describes the extraction of inorganic nitrogen (nitrate, NO3-, and ammonium, NH4+) from soil using 2 M potassium chloride (KCl). The KCl extraction is the standard method for determining plant-available inorganic nitrogen in soils (Keeney & Nelson, 1982; Mulvaney, 1996). The high ionic strength of the KCl solution displaces NH4+ from cation exchange sites and brings NO3- into solution, producing a filtered extract suitable for colorimetric, flow-injection, or ion chromatographic analysis (Maynard et al., 2008).
Does not apply to: Total nitrogen analysis (requires Kjeldahl or combustion digestion; see SOP-L407); dissolved nitrogen in water samples; semi-quantitative field screening (see SOP-L404)
Ammonium (NH4+): The protonated form of ammonia; held on negatively charged cation exchange sites in soil; released into solution by the high ionic strength of the 2 M KCl extractant; the primary plant-available nitrogen pool in many soils.
Cation exchange capacity (CEC): The total capacity of a soil to hold exchangeable positively charged ions; controls how much NH4+ can be stored and released; higher in clay-rich and organic-matter-rich soils.
KCl extraction: Shaking soil with 2 mol L-1 potassium chloride solution at a defined soil-to-solution ratio (typically 1:10 w/v); the standard procedure for quantifying plant-available inorganic nitrogen in soils (Keeney & Nelson, 1982).
Mineral nitrogen (mineral N): The sum of NH4+-N and NO3--N in the soil; the immediately plant-available nitrogen pool; expressed in mg N kg-1 dry soil.
Nitrate (NO3-): The oxidized form of inorganic nitrogen; anion, so not held on cation exchange sites; highly mobile in soil solution; measured colorimetrically or by flow injection analysis after KCl extraction.
Soil-to-solution ratio: The mass of soil (g) to volume of extractant (mL) used in the extraction; typically 1:10 for KCl extractions; must be consistent within and between studies for results to be comparable.
5 Safety and Hazards
WarningHazards
Chemical: 2 M KCl is a mild irritant; avoid contact with eyes and skin. Review the SDS before first use.
Physical: Reciprocating shaker can pinch fingers if containers are not secured properly.
The 2 M KCl concentration is the standard for inorganic N extraction (Keeney & Nelson, 1982; Mulvaney, 1996). Lower concentrations (e.g., 1 M) may underestimate exchangeable NH4+ in high-CEC soils (Maynard et al., 2008).
Weigh out 298.2 g KCl into a 2 L volumetric flask
Fill the flask approximately two-thirds full with distilled water
Place the flask on a magnetic stirrer and add a magnetic stir bar
Set the stirrer to low-medium speed and leave on for 4 hours until fully dissolved
Use a stir bar retriever to remove the magnetic stir bar
Fill the flask to the 2 L mark with distilled water and mix
TipTip
Prepare the KCl solution the day before extraction. It can be stored at room temperature in a capped container indefinitely. Verify the solution with a conductivity meter if stored for more than 6 months.
8.2 Moisture Subsample
A companion gravimetric moisture determination is required to express results on an oven-dry soil basis (Keeney & Nelson, 1982).
Set aside 10 g of soil from each sample for moisture determination
Place each 10 g subsample into a labeled aluminum weigh boat
Dry at 105 C for 24 hours and record the dry weight
The standard soil-to-solution ratio is approximately 1:5 to 1:10 (w/v) (Mulvaney, 1996). This SOP uses 7 g field-moist soil in 40 mL KCl, yielding an effective ratio of approximately 1:6 on an oven-dry basis for a typical soil at 15% gravimetric moisture.
Dispense 40 mL of 2 M KCl solution into labeled test tubes or extraction bottles
Add 7 g of field-moist soil to each tube (the total volume should reach approximately 50 mL). Record the exact mass of soil added to the nearest 0.01 g.
Cap the tubes securely
Place on the reciprocating shaker for 2 hours
Remove from the shaker and either:
Allow soil particles to settle for approximately 10 minutes (do not exceed 1 hour), or
Record the exact mass of field-moist soil added to each tube. This value, together with the gravimetric moisture content, is needed to calculate results on an oven-dry basis (see Appendix A).
8.4 Filtering
Set up filter funnels with Whatman #42 filter paper over labeled specimen containers or clean test tubes
Pour the settled or centrifuged solution into the funnels; do not fill above the filter paper line
Allow the solution to drain completely until only soil remains on the filter
Discard the used filter paper and soil
8.5 Extract Storage
Cap the filtered extract containers
If analysis will be completed within 48 hours, store at 4 C
If analysis will be delayed beyond 48 hours, freeze the extracts at -20 C until analysis (Keeney & Nelson, 1982)
Important
Extracts should be analyzed as soon as possible. Prolonged storage, even frozen, can alter NH4+ and NO3- concentrations due to microbial activity or chemical transformations (Maynard et al., 2008).
9 Calculations
9.1 Converting Extract Concentration to Soil Concentration
The colorimetric or flow-injection analyzer reports the concentration of NO3--N or NH4+-N in the filtered extract (mg N/L). To convert this to mg N per kg oven-dry soil (ppm):
\[
C_{\text{soil}} \;(\text{mg N kg}^{-1}) \;=\;
\frac{C_{\text{extract}} \;\times\; V_{\text{KCl}}}{m_{\text{dry}}}
\]
where:
\(C_{\text{extract}}\) = concentration in the filtered extract (mg N/L)
\(V_{\text{KCl}}\) = volume of KCl extractant added (L; typically 0.040 L)
\(m_{\text{dry}}\) = oven-dry equivalent mass of soil extracted (kg), calculated as:
where \(m_{\text{moist}}\) is the field-moist mass added (g, converted to kg) and \(\theta_g\) is the gravimetric moisture content expressed as a decimal (g water / g oven-dry soil).
See Appendix A for a worked R code example.
9.2 Soil Texture Correction Factor (Optional)
When using the semi-quantitative quick test (SOP-L404), a texture-based correction factor may be applied. For the quantitative KCl extraction described here, no texture correction is needed because the exact soil mass and moisture content are measured directly.
Texture
Moist Soil Factor
Dry Soil Factor
Sand
2.3
2.6
Loam
2.0
2.5
Clay
1.7
2.2
These factors are retained here for reference when comparing quick-test results to KCl extraction results.
10 Quality Control
Include a reagent blank (KCl solution only, no soil) in each extraction batch. The blank should read below the method detection limit for both NO3--N and NH4+-N.
Run extraction duplicates for at least 10% of samples. Duplicate results should agree within 15% relative percent difference (RPD).
Include a laboratory reference soil with known inorganic N content in each batch when available.
Record the KCl solution preparation date and batch number.
Verify that the analytical calibration curve for the colorimetric method has an \(r^2 \geq 0.995\) before reporting results.
Subtract the mean reagent blank concentration from all sample results if the blank reads above zero (Mulvaney, 1996).
11 Waste Disposal
Waste Type
Disposal Method
KCl extracts
Aqueous waste; drain with copious water if approved by EH&S, or collect for pickup
Used filter paper and soil
Standard trash
Unused KCl solution
Drain with water (non-hazardous at this concentration)
12 References
Hart, S. C., Stark, J. M., Davidson, E. A., & Firestone, M. K. (1994). Nitrogen mineralization, immobilization, and nitrification. In R. W. Weaver, J. S. Angle, & P. S. Bottomley (Eds.), Methods of soil analysis, part 2: Microbiological and biochemical properties (pp. 985–1018). Soil Science Society of America. https://doi.org/10.2136/sssabookser5.2.c42
Keeney, D. R., & Nelson, D. W. (1982). Nitrogen – inorganic forms. In A. L. Page, R. H. Miller, & D. R. Keeney (Eds.), Methods of soil analysis, part 2: Chemical and microbiological properties (2nd ed., pp. 643–698). American Society of Agronomy; Soil Science Society of America. https://doi.org/10.2134/agronmonogr9.2.2ed.c33
Maynard, D. G., Kalra, Y. P., & Crumbaugh, J. A. (2008). Nitrate and exchangeable ammonium nitrogen. In M. R. Carter & E. G. Gregorich (Eds.), Soil sampling and methods of analysis (2nd ed., pp. 71–80). CRC Press / Canadian Society of Soil Science. https://doi.org/10.1201/9781420005271.ch6
Mulvaney, R. L. (1996). Nitrogen: Inorganic forms. In D. L. Sparks, A. L. Page, P. A. Helmke, & R. H. Loeppert (Eds.), Methods of soil analysis, part 3: Chemical methods (pp. 1123–1184). Soil Science Society of America. https://doi.org/10.2136/sssabookser5.3.c38
13 Training Record
Before performing this procedure independently, personnel must:
Perform the extraction and filtering procedure under supervision
Calculate results for one batch using the R code in Appendix A and verify against the instructor’s values
Be signed off by the PI or lab manager
14 Appendix A: Calculation of Soil NO3--N and NH4+-N (R Code)
The following R code converts extract concentrations (mg N/L from the colorimetric analyzer) to soil concentrations (mg N/kg oven-dry soil, equivalent to ppm) for a batch of samples. It corrects for the exact mass of field-moist soil added and the gravimetric moisture content of each sample.
Code
# ===========================================================================# SOP-L403: KCl Extraction -- Convert extract concentration to soil ppm# ===========================================================================# ---- 1. Enter your data ---------------------------------------------------# Replace the example values below with your actual data.# Each vector must have the same length (one entry per sample).sample_id <-c("S01", "S02", "S03", "S04", "S05")# Mass of field-moist soil added to the extraction tube (g)soil_moist_g <-c(7.03, 7.10, 6.98, 7.05, 7.12)# Gravimetric moisture content (proportion, g water / g oven-dry soil)# From SOP-L400. Example: 0.15 means 15% moisture.theta_g <-c(0.152, 0.148, 0.161, 0.155, 0.143)# Volume of 2 M KCl added (mL) -- typically 40 mL for all samplesvol_kcl_mL <-rep(40, length(sample_id))# Extract concentrations from the colorimetric analyzer (mg N / L)no3_extract_mg_L <-c(3.42, 5.18, 2.76, 4.01, 6.33)nh4_extract_mg_L <-c(1.05, 0.87, 1.22, 0.93, 0.78)# Reagent blank concentrations (mg N / L) -- mean of your blanksblank_no3_mg_L <-0.02blank_nh4_mg_L <-0.01# ---- 2. Calculate oven-dry soil mass (kg) ----------------------------------soil_dry_g <- soil_moist_g / (1+ theta_g)soil_dry_kg <- soil_dry_g /1000# ---- 3. Blank-correct extract concentrations --------------------------------no3_corrected <- no3_extract_mg_L - blank_no3_mg_Lnh4_corrected <- nh4_extract_mg_L - blank_nh4_mg_L# Ensure no negative values after blank correctionno3_corrected[no3_corrected <0] <-0nh4_corrected[nh4_corrected <0] <-0# ---- 4. Convert to mg N / kg oven-dry soil (ppm) ----------------------------vol_kcl_L <- vol_kcl_mL /1000no3_ppm <- (no3_corrected * vol_kcl_L) / soil_dry_kgnh4_ppm <- (nh4_corrected * vol_kcl_L) / soil_dry_kg# ---- 5. Assemble results table -----------------------------------------------results <-data.frame(Sample = sample_id,Moist_g = soil_moist_g,Moisture_pct =round(theta_g *100, 1),Dry_g =round(soil_dry_g, 3),Vol_KCl_mL = vol_kcl_mL,NO3_extract = no3_extract_mg_L,NH4_extract = nh4_extract_mg_L,NO3_ppm =round(no3_ppm, 2),NH4_ppm =round(nh4_ppm, 2),Total_inorgN =round(no3_ppm + nh4_ppm, 2))# ---- 6. Print results --------------------------------------------------------cat("SOP-L403: KCl Extraction Results\n")cat("================================\n\n")print(results, row.names =FALSE)cat("\n")cat(sprintf("Reagent blank NO3-N: %.2f mg/L\n", blank_no3_mg_L))cat(sprintf("Reagent blank NH4-N: %.2f mg/L\n", blank_nh4_mg_L))cat("\nAll soil concentrations are mg N / kg oven-dry soil (ppm).\n")
14.1 Worked Example (Sample S01)
Using the example data above for sample S01:
Field-moist soil added: 7.03 g
Gravimetric moisture: 15.2% (0.152)
Oven-dry equivalent: \(7.03 / (1 + 0.152) = 6.102\) g = 0.006102 kg
KCl volume: 40 mL = 0.040 L
NO3--N in extract: 3.42 mg/L (blank-corrected: 3.40 mg/L)
General interpretation thresholds for extractable inorganic N in agricultural and wildland soils. These are approximate and vary by crop, climate, and soil type (Hart et al., 1994; Mulvaney, 1996).
Extractable Inorganic N (mg N/kg)
Interpretation
Notes
Less than 10
Low
Nitrogen likely limiting plant growth
10–25
Moderate
Adequate for many crops and native vegetation
25–50
High
Sufficient N; fertilization generally not needed
Greater than 50
Very high
Excess N; potential leaching or denitrification concern