Fact sheet: Methanotrophic biodegradation—in situ

From: Public Services and Procurement Canada

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Biodegradation by methanotrophs is an in situ technology that allows transformation of chlorinated compounds by indigenous methanotrophic microorganisms for the treatment of contaminated groundwater and soil. Methanotrophs are aerobic bacteria that use methane (CH4) as a source of carbon and energy.

Methanotrophic bacteria produce the enzyme methane mono-oxygenase that oxidizes methane and transform various chlorinated compounds. Methanotrophs can therefore cometabolize chlorinated compounds in the presence of methane and oxygen (final electron acceptor). Cometabolism is the fortuitous transformation of a contaminant (such as TCE) by a microorganism without the organism directly benefiting from the transformation. For cometabolism to occur, an appropriate primary substrate (such as methane) must be present to serve as a source of carbon and energy for the microorganism that performs the contaminant transformation. Dissolved or gaseous nutrients, such as soluble fertilizers, may also be injected into the contaminated matrix.

The presence of methane in the environment is necessary for the development and growth of methanotrophs. If methane concentrations are too high, competition for the active site of the methane mono-oxygenase will favour the oxidation of methane. If not enough methane is present, the chlorinated compound will initially be degraded quickly, but will slow down as the contaminant is not a source of energy for the organism. Methane and oxygen injections are required to enhance the biodegradation of halogenated organic compounds. Dissolved or gaseous nutrients, such as soluble fertilizers, may also be injected into the contaminated matrix.

The oxidation of chlorinated compounds produces chlorinated epoxides that are unstable in water and quick to breakdown. Subsequent transformations are catalyzed by methanotrophic or other types of bacteria.

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Implementation of the technology

Methanotrophic biodegradation projects may include:

  • pre-treatment laboratory and/or pilot-scale trials
  • mobilization, site access and temporary facilities
  • reagent delivery, which could entail such measures as:
    • injection well/tip installation
    • infiltration trench/drain construction
    • injection or infiltration of aqueous treatment solutions
    • injection of slurries
    • injection of gases below the water table
    • deep soil mixing with solid or slurry reagents
    • groundwater extraction, amendment and re-injection
  • monitoring
  • decommissioning of injection equipment

Materials and storage

  • On-site storage is primarily a function of the compounds being applied to the groundwater systems and the manner of application.
  • Projects using periodic injections of material may bring materials to the site on an as-needed basis and avoid on-site storage. 
  • Injected materials vary widely according to contaminants, general groundwater composition and practitioners. A variety of proprietary mixtures are commonly used, including oxygen releasing compounds. Common generic compounds include urea (as a nitrogen source), ammonium nitrate (as a nitrogen source), dilute hydrogen peroxide, etc. Tanks of methane and oxygen gas or on-site oxygen generators may also be used.

Waste and Discharges

  • If treatment is successful, the primary residual is microbial biomass (which decays over time). Excess reagent typically cannot be recovered, and is generally consumed in situ.
  • System installation typically requires drilling or excavating in contaminated areas, resulting in the handling and disposal of contaminated soils, typically containerized and disposed of off-site.
  • Treated groundwater may transport bacteria, amendments, and degradation by-products out of the treatment zone. Hydraulic control may be required.

Recommended analyses for detailed characterization

Biological analysis

  • Total heterotrophic and specific bacterial counts (according to the contaminants of interest)

Chemical analysis

  • pH
  • Oxidation reduction potential (Eh)
  • Metals concentrations
  • Contaminant concentrations present in the following phases:
    • adsorbed
    • dissolved
    • free
  • Nutrient concentrations including:
    • ammonia nitrogen
    • total Kjeldahl nitrogen
    • nitrates
    • nitrites
    • total phosphorus

Physical analysis

  • Dissolved oxygen concentration
  • Dissolved methane concentration
  • Vadose zone oxygen, nitrogen dioxide, and methane concentrations
  • Temperature
  • Evaluation of biological conditions and ecological factors

Recommended trials for detailed characterization

Biological trials

  • Microcosm mineralization trial

Hydrogeological trials

  • Tracer tests


Tests examining the effect of temperature change on hydraulic conductivity and establishing the zone of freezing with a pilot scale tubing system are recommended to properly design the full-scale containment system.

Other information recommended for detailed characterization

Phase II

  • Contaminant delineation (area and depth)
  • Presence of receptors:
    • presence of potential environmental receptors
    • presence of above and below ground infrastructure
    • the risk of off-site migration

Phase III

  • Soil stratigraphy
  • Identification of preferential pathways for contaminant migration
  • Conceptual site model with hydrogeological and geochemical inputs
  • Characterization of the hydrogeological system including:
    • the direction and speed of the groundwater flow
    • the hydraulic conductivity
    • the seasonal fluctuations
    • the hydraulic gradient


  • Allows treatment of residual contamination within the vadose and saturated zones.
  • Allows treatment of dissolved contamination in groundwater.
  • Suitable for treatment of halogenated contaminants.

Applications to sites in northern regions

In situ methanotrophic biodegradation is potentially applicable to remote northern sites where impediments to material transport and injection equipment mobilization can be overcome. Cold temperatures can hamper biodegradation and microbial activity may only occur during the summer months, thus treatment time may take several years. Microbial activity may be possible in deep soil as temperatures (below permafrost) are relatively constant over the course of the year. 

Treatment type

Treatment type
Treatment typeApplies or Does not apply
In situ
Ex situ
Does not apply
Does not exist
Does not exist
Dissolved contamination
Free Phase
Does not exist
Does not exist
Residual contamination
Does not exist

State of technology

State of technology
State of technologyExist or Does not exist

Target contaminants

Target contaminantsApplies, Does not apply or With restrictions
Aliphatic chlorinated hydrocarbons
With restrictions
With restrictions
Does not apply
Does not apply
Monocyclic aromatic hydrocarbons
Does not apply
Non metalic inorganic compounds
Does not apply
With restrictions
Petroleum hydrocarbons
Does not apply
Phenolic compounds
Does not apply
Policyclic aromatic hydrocarbons
Does not apply
Polychlorinated biphenyls
Does not apply

Treatment time

Treatment time
Treatment timeApplies or Does not apply
Less than 1 year
Does not apply
1 to 3 years
3 to 5 years
More than 5 years

Long-term considerations (following remediation work)

Follow-up monitoring may be required to verify that the remediation objectives as well as applicable regulations are met once the groundwater system normalizes; after stimulation is withdrawn and excess biomass dies off. 

Secondary by-products and/or metabolites

Vinyl chloride, a toxic compound which can be produced during reductive dehalogenation of certain chlorinated compounds, is not produced by methanotrophic cometabolism. Laboratory or pilot scale studies, as well as strict control of injected materials are typically required.

Limitations and Undesirable Effects of the Technology

  • Fractured, compacted, hydrophobic, stratified, and/or a heterogeneous soil matrix could cause preferential pathways and impede the homogeneous distribution of injected methane and oxygen.
  • The competition between methane and halogenated organic compounds for the methane mono-oxygenase enzyme limit contaminant transformation.
  • The low solubility of oxygen and methane in water can limit contaminant biodegradation;
  • Low nutrients concentration may limit contaminant biodegradation.
  • Ideal when soil pH ranges from 5 to 8.
  • High contaminant concentrations can inhibit biodegradation of the halogenated organics compounds.
  • By design, methanogenic bioremediation will have large effects on parameters like oxidation-reduction potential, pH and total organic carbon. 
  • Injected fluids may displace (or “push”) contaminated pore water ahead of the injection front, leading to short-lived but dramatic changes in the distribution of groundwater contamination.
  • The use of oxygen and methane, two flammable gases,  poses health, safety and environmental risks (fire, suffocation in confined space and explosion).
  • If the treatment areas experience accident or upset conditions, contaminated groundwater may escape untreated.

Complementary technologies that improve treatment effectiveness

  • Methanotrophic biodegradation combined with a bioventing or biosparging system could be used under certain conditions.
  • The utilization of formate or methanol as a primary substrate is one solution to reduce the competition for mono-oxygenase, but has currently not shown acceptable results.
  • Bioaugmentation.

Required secondary treatments

Installation of a vapour extraction system to collect all gas emissions may be required where bioventilation or biosparging is used.

Application examples

Application examples are available at these addresses:


The use of methanotrophs in the biodegradation of chlorinated solvents has been demonstrated as an effective remediation strategy. Mineralization of 90% of TCE was observed at a contaminated site located, at Savannah River, USA, after the injection of gaseous methane, oxygen and nutrients. As with all remediation technologies, the efficacy and time required for remediation is dependent upon several factors, including the type and concentration of contaminants, the microbiological population and activity and the physical and chemical conditions of the contaminated site.

Measures to improve sustainability or promote ecological remediation

  • Schedule optimization for resource sharing and fewer days of mobilization.
  • Use of renewable energy and energy-efficient equipment (such as geothermal or solar energy for reagent delivery).
  • Select amendments with lower energy requirement for production
  • Encourage amendment supply by local producers.
  • Use of groundwater recirculation to maximize the use of amendments and lower number of injection wells.
  • Use of rainfall for amendments/nutrients mix for injection.
  • Use of recyclable bulk solution containers.

Potential impacts of the application of the technology on human health

Unavailable for this fact sheet


Author and update

Composed by : Karine Drouin, M.Sc., National Research Council

Updated by : Karine Drouin, M.Sc., National Research Council

Updated Date : April 1, 2008

Latest update provided by : Marianne Brien, P.Eng., Christian Gosselin, P.Eng., M.Eng., Golder Associés Ltée

Updated Date : March 22, 2019