B OZEL DUYGAN¹, S BEGGAH¹, S REY², M SEYREFIELD², J VAN DER MEER<sup>1

1</sup>Department of Fundamental Microbiology, University of Lausanne, Switzerland, ²Biotechnology Department, Firmenich SA, Switzerland

Existing biodegradation methods such as OECD tests are typically carried out at relatively high substrate concentrations such as 20 mgC/L that may result in under- or over- estimation of observed substrate utilization kinetics at environmentally relevant low concentrations. Furthermore, although well established, OECD guidelines are largely not suitable for testing hydrophobic and volatile compounds such as fragrances. Our main goal is to develop a standardized and validated growth-linked biodegradation test as an alternative method to existing biodegradation tests
 in the range of 0.1 - 10 mgC/L. Our methodological concept is based on comparative evaluation of cell density measurement by flow cytometer and substrate disappearance measurement by CO2 evolution and gas chromatography under assimilable organic carbon restricted conditions. We conduct our experiments with lake water microbial communities at starting cell density of 10E4 and 10E5 cells/mL in a defined mineral medium. Sodium benzoate, 1-octanol, anthraquinone and phenol are selected as primary positive controls for readily biodegradable compounds. We observed step-wise increase in the lake community size (at initial cell density of 10E4 cells/mL) at the expense of added positive control substance at concentrations of between 0.1 – 2 mgC/L and 1–2 mgC/L, respectively. Yield approximations from the observed community growth was in line with CO2 evolution test results for the target compounds. For accurate mass balance between compound and community size, we will further simultaneously measure lake water community growth and compound disappearance by gas chromatography. Lake water communities that enable biodegradation of the test compounds will be analyzed for diversity changes, and we will also further isolate and characterize degrader bacteria.