Fact sheet: Anaerobic Bioremediation—in situ

From: Public Services and Procurement Canada

On this page


Anaerobic bioremediation is an in situ technology applicable for soil or groundwater contaminated with petroleum hydrocarbons or other organic contaminants in saturated conditions.

This technology consists of providing all the necessary compounds (electron acceptor, nutrients, substrate, etc.) to stimulate the indigenous anaerobic microorganisms (strict or facultative) to enhance contaminant biodegradation. Strict anaerobic microorganisms cannot use oxygen as the final electron acceptor in the respiratory chain and therefore, they use alternative electron acceptors such as nitrate (NO3-), dissolved manganese (Mn4+), ferric iron (Fe3+), sulfate (SO42-) and carbon dioxide (CO2). Alternative electron acceptors are more water-soluble than oxygen and can be more easily injected into the dissolved phase. Nutrients may be injected in a dissolved (such as commercial soluble fertilizer), or gaseous form. Addition of a substrate can help reduce the oxygen concentration and establishing anaerobic conditions.

Anaerobic bioremediation has seen extensive application for the remediation of chlorinated solvents contaminated sites. Anaerobic biodegradation of chlorinated organic compounds is discussed in the Reductive Dechlorination technology sheet.

In addition to the chlorinated compounds, in situ anaerobic bioremediation treatment has been demonstrated for multiple contaminants including petroleum hydrocarbons, ammonium, nitrate, sulfate, pesticides, explosives and dioxins. Depending on the contaminant and on the geochemistry of the contaminated media, practitioners may inject food-grade oils, hydrogen sources, nitrates, fertilizers, etc.

Internet links:

Implementation of the technology

  • Projects implementation may include:
    • Pre-treatment laboratory and/or pilot-scale trials;
    • Reagent delivery, which could entail such measures as:
    • Injection well/tip (drilling, hammering) installation.
    • Infiltration trenches/drain construction.
    • Injection or infiltration of aqueous treatment solutions.
    • Injection of slurries.
    • Injection of gasses below the water table (emitters, diffusers).
    • Deep soil mixing with solid or slurry reagents.
    • Groundwater extraction, addition of amendments and re-injection.
  • Monitoring.
  • Decommissioning of injection equipment.

Contaminants are degraded or transformed by the bacteria and the treatment is complete when the contaminant concentrations have reached the treatment objectives.

Issues with anaerobic bioremediation include the distribution of components required by the treatment in the subsurface and ensuring adequate environment for optimal microbial growth.

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 and nonproprietary mixtures are commonly used, such as lactate, molasses, acetate, methanol, ethanol, vegetable oil, slow-release hydrogen compounds, compost, etc.
  • Nitrate injection is regulated and may not be allowed under certain situations.

Waste and Discharges

  • If treatment is successful, the primary residual is microbial biomass (which decays over time).
  • Excess reagent may not be recovered, it is usually consumed on-site.
  • System installation typically requires drilling or excavating in contaminated areas, resulting in the handling and disposal of contaminated soils, typically disposed of off-site.
  • Treated groundwater may transport bacteria, amendments, and degradation by-products out of the treatment zone. Hydraulic controls 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)
  • Organic carbon content
  • Organic matter content
  • Metals concentrations
  • Contaminant concentrations present in the following phases:
    • adsorbed
    • dissolved
    • free
  • Nutrient concentrations including:
    • ammonia nitrogen
    • total Kjeldahl nitrogen
    • nitrates
    • nitrites
    • total phosphorus
  • Electron acceptor concentrations/reaction by-products including:
    • dissolved oxygen
    • nitrate
    • sulfate
    • ferric and ferrous iron
    • methane
    • dissolved manganese

Physical analysis

  • Dissolved oxygen concentration
  • Dissolved methane concentration
  • Temperature
  • Soil granulometry
  • 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 saturated zone, as well as dissolved-phase contamination in groundwater.
  • Suitable for contaminants that may be degraded or transformed under anaerobic (no oxygen) or anoxic (low oxygen concentration) conditions.
  • The soil permeability must be sufficiently high to allow the injection of electron acceptors and other substrates.

Applications to sites in northern regions

In situ anaerobic bioremediation 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
Does not apply
Does not apply
With restrictions
Does not apply
Monocyclic aromatic hydrocarbons
Non metalic inorganic compounds
With restrictions
With restrictions
Petroleum hydrocarbons
With restrictions
Phenolic compounds
Does not apply
Policyclic aromatic hydrocarbons
With restrictions
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 groundwater system normalizes and the applicable regulations are met after the stimulation is withdrawn and excess biomass dies off.

Secondary by-products and/or metabolites

  • By-product formation can be a significant concern. Toxic intermediates may occur in the degradation of some explosives and pesticides. Bench top and/or pilot testing, as well as strict quality control for injected materials, is typically required.

Limitations and Undesirable Effects of the Technology

  • This technology is not suitable when free phase is present;
  • Soil permeability limits the application of anaerobic bioremediation;
  • Fractured, compacted, hydrophobic, stratified, and/or heterogeneous soil matrices could cause preferential pathways making homogeneous injection of substrates hard to achieve;
  • High contaminant concentrations could inhibit biodegradation;
  • High metal concentrations can inhibit biodegradation;
  • Ideal pH conditions range from 6 to 8;
  • Biodegradation rates of organic compound under anaerobic conditions are slower than under aerobic conditions;
  • Understanding of anaerobic biodegradation mechanisms and system monitoring can be more complex than for aerobic biodegradation;
  • By design, bioremediation will have large effects on parameters like oxidation-reduction potential, pH and total organic carbon.
  • If the treatment areas experience accident or upset conditions, contaminated groundwater may escape untreated.

Complementary technologies that improve treatment effectiveness

  • Injection of nitrate with oxygen can induce biodegradation under microaerophilic conditions (about 2% to 10% of oxygen concentration), which may be an advantageous for certain contaminants;
  • Bioaugmentation.

Required secondary treatments

  • There is no required secondary treatment.

Application examples

Application examples are available at these links:


The treatment time for complete remediation using anaerobic bioremediation varies according to the type and concentration of contaminants, the indigenous microbial population 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 (for example, geothermal or solar energy for reagent delivery).
  • Use of amendments requiring less energy for their production.
  • Use of amendments manufactured locally.
  • Use of groundwater recirculation to maximize the use of amendments and lower number of injection wells
  • Use of alternative sources of water 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