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1A - Environmental analytical methods (MOP/19) In Vitro Biotransformation of Surfactants in Cellular and Subcellular Assays. Bernhard, Mary Jo1, Dyer, Scott1, Gimeno, Sylvia2, 1 The Procter & Gamble Company, Cincinnati, Ohio, USA2 The Procter & Gamble Company, Brussels, Belgium, Belgium ABSTRACT- We investigated the potential of using in vitro biotransformation to predict in vivo BCF. In vitro methods could yield several benefits: 1) refined BCFs from Kow-only approaches; 2) rapid screening capabilities; and 3) reduced animal use. Four in vitro systems were studied to identify a suitable method accounting for biotransformation that can be used to predict bioconcentration behavior of surfactants in whole fish. The in vitro systems investigated were an immortalized fish hepatic cell line (PHLC-1) and Cyprinus carpio primary hepatocytes, liver microsomes and homogenates. A nonionic and an anionic surfactant were used as test compounds to determine the feasibility of each in vitro system. Primary hepatocytes and PLHC-1 cells were dosed with nontoxic levels of radiolabeled test materials and timed samples obtained over 48h. Analyses included liquid scintillation counting (LSC), protein determination, and thin layer chromatography (TLC). LSC measured the bioconcentration by determining the partitioning of parent and metabolites associated between the cells and media. Protein measurement determines the cell mass. TLC establishes the distribution of parent and metabolites in each fraction. Biotransformation in sub-cellular systems was determined using RAD TLC at timed points over 4 hours. Parent compound loss, due to metabolism, in all four systems exceeded rates obtained from whole organism tests. Cellular BCFs were lower than whole organism BCFs. This work was supported by the European Risk Assessment of Surfactant Materials (ERASM). Key words: hepatocytes, Bioconcentration, microsomes, homogenates |
