OECD 208 Terrestrial Plant Test: Seedling Emergence and Seedling Growth Test

The “ OECD 208 Terrestrial plant testing: Seedling emergence and seedling growth testing” standard, published under the OECD guidelines on responsible business conduct for multinational companies (OECD guidelines), is designed to assess the effects on seedling emergence and early growth of higher plants after exposure to a test substance applied to the soil surface or soil.

Seeds are placed in contact with soil treated with the test substance and evaluated for effects, typically 14 to 21 days after 50 percent of the seedlings in the control group have emerged. The endpoints measured are visual assessment of seedling emergence, biomass measurements, shoot length, and visible detrimental effects on various plant parts. The test can be performed using a single concentration/rate to determine the dose-response curve or as a limit test depending on the purpose of the study. Appropriate statistical analysis is used to obtain the effective concentration or effective application rate for the most sensitive parameters of interest. Furthermore, the no-observed-effect concentration and the lowest observed-effect concentration can be calculated in this test.

The Organisation for Economic Co-operation and Development (OECD) develops standard test guidelines to assess the potential hazards of chemicals to human health and the environment. Among these, OECD Standard 208 is a critical tool for assessing the phytotoxicity of substances to higher plants.

The OECD 208 guideline is designed to evaluate the effects of a test substance, such as general chemicals, biocides, plant protection products, or composted materials, on seedling emergence and early growth of terrestrial plants. It is widely used to determine whether a substance is toxic to plants, including composted materials from biodegradable products such as plastics. This guideline is particularly important in regulatory frameworks such as ASTM D6400, which assess the environmental safety of composted materials.

This testing method focuses on two primary outcomes:

• Seedling emergence: The ability of seeds to germinate and emerge from soil treated with the test substance.
• Early growth: Growth and development of seedlings measured by parameters such as shoot length, biomass and apparent phytotoxic effects.

By evaluating these endpoints, the OECD 208 standard helps identify potential adverse effects of chemicals or materials on plant development and ensures that products entering the market are safe for ecosystems.

The OECD 208 test involves exposing seeds of selected plant species to soil treated with a test substance, either applied to the soil surface or mixed into the soil. The test typically takes 14 to 21 days after 50 percent of the control group seedlings have emerged.

This standard recommends testing a variety of plant species to account for taxonomic diversity and ensure reliable results. Typically, six to ten species from different plant families are selected. For example:

• Monocots: For example, oats, corn.
• Dicotyledons: For example, rapeseed, soybean, tomato, cucumber.
• Non-agricultural or wild species: For example, common yarrow or marigold.

The choice of species depends on the properties of the test substance and its intended use, such as herbicides or veterinary drugs.

The test method is performed under controlled conditions to ensure consistency and repeatability:

• Soil preparation: The soil is sieved to a depth of 2 mm and its properties (e.g., percentage organic carbon, pH, and electrical conductivity) are measured. For standard tests, artificial soil composed of quartz sand, sphagnum peat, kaolin clay, and calcium carbonate can be used.
• Pot size: Typically three pots per control and test substance group, 15 cm in diameter.
• Environmental conditions: Temperature 22 degrees plus/minus 1 degree, humidity 70 percent plus/minus 25 percent, photoperiod minimum 16 hours of light and light intensity 350 plus/minus 50 µE/m2/sec, wavelength 400-700 nm.
• Test duration: 14-21 days depending on species and study design.

The test can be administered in two formats:

• Dose-response testing: Multiple concentrations of the test substance are applied to determine a dose-response curve, allowing metrics such as EC50 (the concentration that causes 50 percent effect) to be calculated.
• Limit test: A single concentration is tested to confirm whether the substance has significant phytotoxic effects.

Seeds are planted in the treated soil, and test pots are sub-irrigated with nutrient solution as needed. Observations are made at regular intervals (e.g., on days 7, 14, and 21) to assess:

• Seedling emergence rate.
• Throughout the exile.
• Shoot fresh weight or biomass.
• Visible phytotoxic effects (e.g., chlorosis, necrosis, or abnormal growth).

For the test to be considered valid, the following criteria must be met:

• In the control group, seedling emergence should be at least 70 percent.
• Control seedlings should not show visible phytotoxic effects.
• At least 90 percent of the control seedlings that emerged must survive the duration of the study.

The OECD 208 standard is widely applied in a variety of regulatory and environmental contexts:

• Chemical safety assessment: Used to assess the phytotoxicity of industrial chemicals, pesticides and biocides under regulations such as the European Union Registration, Evaluation, Authorisation and Restriction of Chemicals (REACH).
• Biodegradation testing: As part of the ASTM D6400 standard, the OECD 208 standard evaluates the safety of composted materials derived from biodegradable plastics, ensuring that they do not release toxic components into the environment.
• Veterinary medicinal products: This standard has been adapted to test the effects of substances applied to manure and then mixed into the soil, simulating real-world agricultural scenarios.
• Herbicide development: This standard helps evaluate the impact of herbicides on non-target plants, ensuring they do not harm biodiversity or ecosystems.

The OECD 208 standard is a strong guideline, but some challenges remain:

• Species sensitivity: Plant species may not always reflect the sensitivity of wild or non-target plants, which has led to the inclusion of non-plant species in updated guidelines.
• Test duration: The 14-21 day period may not reflect long-term reproductive effects. This is addressed in complementary standards such as OECD 227 (for spray applications) or ISO 22030.
• Emergence variability: Some non-crop species fail to meet the 70 percent emergence criterion and require seed pretreatments such as cold stratification to enhance germination.

Recent updates to this standard have addressed these issues by increasing the number of recommended test types and clarifying regulatory requirements, providing broader applicability and improved sensitivity. The OECD 208 standard is considered adequate for long-term hazard assessment when testing a variety of species.

As a result, the OECD 208 standard is a cornerstone of environmental toxicology, providing a standardized method for assessing the phytotoxic effects of chemicals and composted materials on terrestrial plants. Its rigorous methodology, diverse species selection, and controlled conditions ensure reliable and reproducible results, making it a vital tool for regulators, industry professionals, and researchers. By identifying potential risks to plant life, this standard contributes to the safe development and use of chemicals, protecting ecosystems, and promoting sustainable practices.

Our organization, which has been operating for many years and meticulously follows global developments in science and technology, employs a team of experienced professionals and a broad infrastructure to conduct testing, measurement, and analysis in a wide range of areas for businesses across all sectors. In this context, we also provide testing services in accordance with the OECD 208 Terrestrial Plant Testing: Seedling Emergence and Seedling Growth Testing standard.