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OECD 301B
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Quick understanding of the test
OECD 301B - Method for the quantitative determination of the biodegradability of organic materials in an aqueous aerobic environment
Application
- The inoculum typically includes diverse microbial populations sourced from activated sludge, soil, or surface water.
- The test determines whether a chemical will degrade rapidly in the environment, reducing its potential for long-term ecological harm.
- Biodegradability data is mandatory under REACH and EPA regulations for chemical approval. Passing OECD 301B ensures regulatory compliance.
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Abstract
The OECD 301B (Modified Sturm Test), also known as the CO2 Evolution Test, is a widely recognized method for the quantitative determination of the biodegradability of organic materials in an aqueous aerobic environment. The biodegradability properties of several chemicals have a great impact on the environment, especially on aquatic systems. The 301 B test is one of six tests in the OECD series that have been widely adopted for assessing ready biodegradability. Its wide applicability is due to its virtue of simplicity and reliability.
The 301B test measures the release of carbon dioxide during the microbial degradation of a chemical. Conceptually, in the microbial degradation of an organic chemical, microorganisms respire, and the organism consumes O2 and produces CO2 as a byproduct. The production of CO2 is directly used as a measure to quantify biodegradation. The 301B test is specifically applicable for water-soluble, non-volatile chemicals.
Products to be tested with OECD 301B
This test applies to a wide variety of organic chemicals. This may include but is not limited to:
- Surfactants
- Detergents
- Pesticides
- Pharmaceuticals
- Industrial chemicals
- Personal care products
OECD-301B Test Conditions and Requirements
Testing requires specific conditions for an accurate measurement of biodegradation. The conditions appear as follows:
Test organisms:
Inoculum used in the test is usually obtained from activated sludge from sewage treatment plants. It could also be sourced from the natural environment, such as soil or surface water. This inoculum is a mixture of microbes such as bacteria, protozoa, and fungi that can break down organic matter.
Preconditioning of inoculum:
- Inoculum is pre-conditioned to experimental conditions. It consists of aerating activated sludge/secondary effluent for 5-7 days in a mineral medium at the test temperature.
- The concentration of the test substance should lie between 10 to 20 mg per liter of Dissolved Organic Carbon (DOC) or Total Organic Carbon (TOC).
- The test is carried out in aerobic conditions. The testing temperature is maintained between 22ยฐC ยฑ 2ยฐC to ensure sufficient microbial action. The test is conducted in diffuse light or in the dark to avoid photodegradation.
- The solution is passed through a continuous stream of air that is free from CO2 for the maintenance of aerobic conditions.
OECD 301B test methodology
- A measured volume of inoculated mineral medium is aerated with CO2 at a controlled rate in the dark or diffused light and incubated for 28 days.
- COโ produced during degradation is trapped using barium or sodium hydroxide.
- COโ is quantified by either titrating the residual hydroxide or measuring inorganic carbon.
- COโ from the test substance (minus blank control) is expressed as a percentage of theoretical COโ (ThCOโ).
- Additionally, the degree of biodegradation can also be assessed via DOC analysis at the start and end of incubation.
Importance of OECD 301B test
Biodegradability testing using OECD standards plays an important role in environmental risk assessment and ensuring compliance with regulatory bodies. The standardโs primary importance lies in its ability to:
- Assess biodegradability
It determines the rate and extent of the degradation of chemicals. Thus, this test helps determine if the chemical is biodegradable and non-toxic to the environment or if it is persistent.
- Regulatory compliance
Many regulatory bodies, such as the European Union’s REACH regulation, demand biodegradability data for chemicals registration and risk evaluation. The test is widely accepted for these aspects.
- Wastewater and environmental impact
The biodegradable nature of chemicals is important for the management of industrial wastewater and control of long-term environmental pollution.
Need help with Biodegradation testing!
At Microbe Investigations Switzerland, we provide reliable OECD 301B testing services to assist you in evaluating the biodegradability of your products, ensuring adherence to international regulations and lowering environmental impact. Our skilled team delivers precise and prompt results, facilitating your product development and compliance needs.
Reach out to us today to discover more about our testing services or to schedule your consultation.
Frequently Asked Questions
DR. Martinoz Scholtz
The test assesses the amount of CO2 emitted as a substance degrades biologically in an aerobic aquatic ecosystem.
The test duration is 28 days.
The primary components consist of the test substance, inoculum, mineral medium, and the CO2 measurement system.
OECD 301B (COโ Evolution Test) measures biodegradation by tracking the COโ produced during microbial respiration, making it suitable for a wide range of substances, especially those that may adsorb onto surfaces. OECD 301D (Closed Bottle Test) measures dissolved oxygen depletion in sealed bottles and is ideal for low-solubility, volatile substances.
This experiment is performed under aerobic conditions with a constant temperature at 22ยฐC ยฑ 2ยฐC.
This standard applies to water-soluble, non-volatile organic materials, such as detergents, surfactants, and other industrial chemicals.
The critical elements of the test include the preparation of the test solution, inoculation, aeration, CO2 trapping, and determination of the outcome.
Biodegradation is quantified by measuring the amount of CO2 produced throughout the degradation process using the titration method or by using an inorganic carbon analyzer.
For water-insoluble test substances, specific preparation techniques, such as using emulsifying agents, may be needed for testing.
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