OECD 301B Biodegradability Testing – CO₂ Evolution
What is OECD 301B?
OECD 301B is a standardized test method used to evaluate the ready biodegradability of a substance by measuring carbon dioxide (CO₂) evolution under aerobic conditions. The method determines how effectively microorganisms convert organic carbon into CO₂ over a 28-day period.
Overview
OECD 301B, also known as the CO₂ Evolution (Modified Sturm) Test, is one of six OECD methods used to assess ready biodegradability in an aerobic aqueous environment.
The method is widely used for regulatory screening and environmental assessment of chemicals, particularly for determining whether a substance meets criteria for “readily biodegradable.”
At Situ Biosciences, OECD 301B testing is performed to evaluate biodegradability, support regulatory submissions, and guide product development decisions.
What OECD 301B Measures
OECD 301B measures the amount of carbon dioxide produced during microbial degradation of a test substance.
The test evaluates biodegradation by:
- Inoculating a mineral medium with microorganisms
- Adding the test substance as the primary carbon source
- Aerating the system with CO₂-free air
- Capturing evolved CO₂ in absorption solutions
- Quantifying CO₂ over time
Results are expressed as percent biodegradation based on theoretical CO₂ production (ThCO₂).
Because the method tracks mineralization directly, it provides a reliable measure of ultimate aerobic biodegradation.
How OECD 301B Testing Works
The method follows a standardized aerobic biodegradation procedure.
1. Sample Preparation
The test substance is added to a mineral medium at a known concentration, typically 10–20 mg carbon per liter.
2. Inoculum Addition
Microorganisms are introduced from sources such as activated sludge, sewage effluent, or surface water.
3. Aeration
CO₂-free air is continuously passed through the system to maintain aerobic conditions and prevent background CO₂ interference.
4. CO₂ Capture
Evolved CO₂ is trapped in alkaline absorption solutions such as sodium hydroxide or barium hydroxide.
5. CO₂ Measurement
The absorbed CO₂ is quantified at regular intervals, typically every few days throughout the 28-day test.
6. Calculation of Biodegradation
The amount of CO₂ produced is compared to the theoretical maximum (ThCO₂) to calculate percent biodegradation.
Common Test Conditions
| Parameter | Typical Conditions |
|---|---|
| Test Type | Aerobic ready biodegradability |
| Measurement | CO₂ evolution |
| Duration | 28 days |
| Environment | Aqueous aerobic system |
| Inoculum | Activated sludge or environmental microorganisms |
| Result Expression | Percent biodegradation (ThCO₂) |
| Standard Organization | OECD |
Benefits of OECD 301B
Regulatory acceptance
OECD 301B is widely accepted by regulatory agencies for assessing ready biodegradability.
Direct mineralization measurement
The method measures CO₂ production, providing direct evidence of complete biodegradation.
Standardized screening method
OECD 301B is part of a globally recognized framework for biodegradability testing.
Supports environmental claims
The test can support claims related to biodegradability when criteria are met.
Applicable to a wide range of chemicals
Suitable for many organic substances that are water-soluble and non-volatile.
Limitations of OECD 301B
Strict pass criteria
To be classified as readily biodegradable, substances must reach ≥60% biodegradation within a 10-day window during the 28-day test.
Controlled laboratory conditions
Results may not fully represent real-world environmental conditions.
Not suitable for all substances
Poorly soluble, volatile, or adsorbing substances may require alternative methods.
Potential variability
Biological systems can introduce variability depending on inoculum source and test conditions.
Why Choose OECD 301B Over Other Methods
Different OECD 301 methods evaluate biodegradability using different measurement approaches.
OECD 301B is commonly selected when direct measurement of CO₂ evolution and mineralization is preferred.
our future™ Program – Biodegradability Certification
Situ Biosciences offers the our future™ Program, a data-driven biodegradability certification designed to validate product performance through scientifically backed testing.
The our future™ Certified Biodegradable designation is rooted directly in biodegradation test results generated in our ISO 17025 accredited laboratory. Certification is based on rigorous evaluation using globally recognized test methods, ensuring your product meets defined biodegradability criteria.
By participating in the our future™ Program, companies gain:
- Verified biodegradability certification based on test data
- Transparent reporting, with supporting data available to customers
- Use of the Certified Biodegradable logo for marketing and product claims
- Alignment with global standards and regulatory expectations
This program provides a clear, data-backed way to substantiate biodegradability claims while building trust with customers and stakeholders.
Supported Test Methods
The our future™ Program supports a wide range of biodegradation pathways and materials, including both liquid and solid products.
Ready Biodegradability
OECD 301A, OECD 301B, OECD 301C, OECD 301D, OECD 301E, OECD 301F, OECD 310, ASTM D5864
Inherent Biodegradability
OECD 302B, OECD 302C, OECD 302D
Marine Biodegradability
Anaerobic Biodegradability
Ultimate Biodegradability
Why Enroll in the our future™ Program
Biodegradability claims are increasingly scrutinized across industries. The our future™ Program provides:
- Scientifically defensible claims backed by validated test data
- Improved regulatory alignment across global markets
- A competitive advantage through verified environmental performance
- Confidence for customers and partners through transparent data access
Get Started
To learn more about the our future™ Program or begin certification:
📞 847-483-9950
📧 info@situtest.com
Applications of OECD 301B Testing
OECD 301B is widely used across industries for evaluating biodegradability.
Common applications include:
- Chemical and material screening
- Regulatory submissions (e.g., REACH, TSCA)
- Product development and formulation optimization
- Environmental impact assessments
Frequently Asked Questions
What does OECD 301B measure?
OECD 301B measures the amount of CO₂ produced from microbial degradation of a substance under aerobic conditions.
How long does OECD 301B testing take?
The test typically runs for 28 days, although it may end earlier if biodegradation reaches a plateau.
What qualifies as “readily biodegradable”?
A substance must achieve at least 60% biodegradation within a 10-day window during the test period.
What is the difference between OECD 301B and ASTM D5511?
OECD 301B evaluates aerobic biodegradation, while ASTM D5511 evaluates anaerobic biodegradation under high-solids conditions.
Can OECD 301B be used for regulatory submissions?
Yes. OECD 301B is widely accepted for regulatory purposes and environmental classification.
Biodegradation Testing at Situ Biosciences
Situ Biosciences provides biodegradation testing for a wide range of materials including chemicals, polymers, and coatings.
Testing programs can be designed to support:
- biodegradability screening
- regulatory compliance
- product development
- environmental claim validation
For more information about OECD 301B testing, please contact Situ Biosciences.