Results of extended plant tests using more realistic exposure scenarios for improving environmental risk assessment of veterinary pharmaceuticals

Residues of veterinary medicinal products (VMPs) enter the environment via application of manure onto agricultural areas where in particular antibiotics can cause phytotoxicity. Terrestrial plant tests according to OECD guideline 208 are part of the environmental risk assessment of VMPs. However, this standard approach might not be appropriate for VMPs which form non-extractable residues or transformation products in manure and manure-amended soil. Therefore, a new test design with a more realistic exposure scenario via manure application is needed. This paper presents an extended plant test and its experimental verification with the veterinary antibiotics florfenicol and tylosin tartrate. With each substance, plant tests with four different types of application were conducted: standard tests according to OECD 208 and three tests with application of test substance via spiked manure either without storage, aerobically incubated, or anaerobically incubated for different time periods.

The verification of the extended plant test showed that seedling emergence and growth are comparable to a standard OECD 208 test and reliable effect concentrations could be established. As demonstrated in the present study, phytotoxicity of veterinary antibiotics can be significantly reduced by application via incubated manure compared to the standard plant test. Overall, the presented test design proved suitable for inclusion into the plant test strategy for VMPs.

In standard tests, the lowest NOEC was <0.06 mg/kg dry soil for florfenicol and 16.0 mg/kg dry soil for tylosin tartrate. Pre-tests showed that plant growth was not impaired at 22-g fresh manure/kg dry soil, which therefore was used for the final tests. The application of the test substances via freshly spiked as well as via aerobically incubated manure had no significant influence on the test results. Application of florfenicol via anaerobically incubated manure increased the EC10 by a factor up to 282 and 540 for half-maximum and for maximum incubation period, respectively. For tylosin tartrate, this factor amounted to 64 at half-maximum and 61 at maximum incubation period. The reduction of phytotoxicity was generally stronger when using cattle manure than pig manure and particularly in tests with cattle manure phytotoxicity decreased over the incubation period. Conclusions: The verification of the extended plant test showed that seedling emergence and growth are comparable to a standard OECD 208 test and reliable effect concentrations could be established. As demonstrated in the present study, phytotoxicity of veterinary antibiotics can be significantly reduced by application via incubated manure compared to the standard plant test. Overall, the presented test design proved suitable for inclusion into the plant test strategy for VMPs.