Testing of Flavor and Fragrance Materials in Turkey Egg Genotoxicity Assay (TEGA) and Comparison of the Results in Ovo, in Vitro and in Vivo

Document Type


Publication Date

Spring 3-1-2017




The aim of the study was to investigate the use of the Turkey Egg Genotoxicity Assay (TEGA) as a non-animal alternative to in vivo follow-up studies. The genotoxic potential of 19 diverse flavor and fragrance (F&F) agents was assessed in the TEGA using 32P-nucleotide postlabeling (NPL) and comet assays to detect hepatic DNA adducts and strand breaks, respectively. The compounds were selected for testing based on their chemical structures and results in the GADD45a-Gluc ‘BlueScreen HC’ (BSHC) genotoxicity and the Ames mutagenicity assays. Two F&F materials (BDHCA and MEU) produced DNA adducts, and four materials (BDHCA, BMHCA, HEX and MAL) produced DNA strand breaks in the NPL and comet assays, respectively. Fourteen other tested compounds were negative in both NPL and comet assays. Of these14 materials, only 3 are negative in vitro, yet the majority of these materials are not shown to be genotoxic. Based on reports of oxidative DNA damage for two of the materials (MAL & HDMF), these compounds were tested in the enhanced comet assay using repair enzymes to identify oxidative DNA damage. In the enhanced comet assay positive comet findings for MAL were not confirmed, and only equivocal evidence of oxidative damage was found. Meanwhile, HDMF, produced positive results in the enhanced comet assay with formamidopyrimidine DNA glycosylase (FPG) enzyme digestion. In the test set, the TEGA had better specificity and sensitivity for in vivo data compared to in vitro test results. The observed non-concordance with in vitro data could be due to differences in the endpoints measured by the TEGA compared to those in vitro or due to the acknowledged higher rate of false positive results in in vitro systems. The TEGA showed high accuracy compared to standard in vivo genotoxicity assays for the prediction of genotoxic carcinogens as positive and the non-(genotoxic) carcinogens as negative. This findings are of high importance, since the end result of standard genotoxicity assays is typically the prediction of cancer risk.


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