Overview
ISO 14240-2:1997 - "Soil quality - Determination of soil microbial biomass - Part 2: Fumigation-extraction method" specifies a standardized laboratory procedure to estimate soil microbial biomass by measuring extractable organic carbon released from microbes after chloroform fumigation. The method is applicable across aerobic and anaerobic soils (including paddy soils) and over the full soil pH range. While the procedure can also estimate microbial nitrogen and ninhydrin‑reactive nitrogen, this part of the standard describes the carbon measurement.
Key topics and technical requirements
- Principle: Chloroform fumigation lyses intact microbial cells; released organic matter is extracted and the difference between fumigated and unfumigated extracts gives extractable microbial carbon (EC).
- Fumigation conditions: Ethanol‑free chloroform vapour, incubation in the dark at (25 ± 2) °C for 22–24 h, followed by removal of vapour by repeated evacuation.
- Extraction: Use 0.5 mol/L potassium sulfate (K2SO4) as extractant; common soil:extractant ratio is 1:4 (adjustable up to 1:30 for soils with very high organic matter).
- Analytical detection:
- Dichromate oxidation (chemical reflux and back‑titration) - conversion factor kEC = 0.38 for biomass carbon calculation.
- UV‑activated persulfate oxidation with IR/UV detection (automatic carbon analyzer) - kEC = 0.45 for biomass carbon calculation.
- Sampling and sample prep: Follow ISO 10381‑6 guidance; use moist samples (>30% water‑holding capacity) to ensure uniform chloroform distribution; typical sample mass equivalent to 25–50 g oven‑dry soil.
- Apparatus and reagents: Implosion‑protected desiccator, vacuum line, shaker, freezer, ethanol‑free chloroform, K2SO4, soda lime, etc.
- Quality/precision: Interlaboratory precision data are summarized in annex C; Annex A provides a normative method to measure water‑holding capacity (WHC).
Applications and who uses it
- Soil quality and fertility monitoring - assess microbial biomass as an indicator of soil health and nutrient cycling potential.
- Agricultural research and land management - evaluate effects of amendments, tillage, or crop rotations on microbial community size.
- Environmental assessment and remediation - monitor microbial activity in polluted or restored soils, including waterlogged/paddy systems.
- Academic and commercial soil laboratories - standardized protocol for comparable microbial biomass C data across studies and sites.
Typical users: soil scientists, agronomists, environmental consultants, laboratory analysts, and researchers studying carbon and nitrogen cycling.
Related standards
- ISO 14240‑1: Substrate‑induced respiration method (same series)
- ISO 10381‑6: Soil sampling - collection, handling and storage for aerobic microbial process assessment
- ISO 10694: Determination of organic and total carbon (dry combustion)
- ISO 11465: Determination of dry matter and water content (gravimetric method)
Keywords: ISO 14240-2:1997, soil microbial biomass, fumigation-extraction method, extractable organic carbon, chloroform fumigation, microbial biomass carbon, potassium sulfate extraction, soil quality.