Evaluation of the Suitability of Mammalian In Vitro Assays to Assess the Genotoxic Potential of Food Contact Materials

Abstract

Background: Non-targeted screening of food contact materials (FCM) for non-intentionally added substances (NIAS) reveals a great number of unknown and unidentified substances present at low concentrations. In the absence of toxicological data, the application of the threshold of toxicological concern (TTC) or of EU Regulation 10/2011 requires methods able to fulfill safety threshold criteria. In this review, mammalian in vitro genotoxicity assays are analyzed for their ability to detect DNA-damaging substances at limits of biological detection (LOBD) corresponding to the appropriate safety thresholds. Results: The ability of the assays to detect genotoxic effects varies greatly between substance classes. Especially for direct-acting mutagens, the assays lacked the ability to detect most DNA reactive substances below the threshold of 10 ppb, making them unsuitable to pick up potential genotoxicants present in FCM migrates. However, suitability for the detection of chromosomal damage or investigation of other modes of action makes them a complementary tool as part of a standard test battery aimed at giving additional information to ensure safety. Conclusion: improvements are necessary to comply with regulatory thresholds to consider mammalian genotoxicity in vitro assays to assess FCM safety.

HepGentox: a novel promising HepG2 reportergene-assay for the detection of genotoxic substances in complex mixtures

“…When comparing the HepGentox to the Ames test in Table 4 it can be seen that the mammalian assay only led to lower LEC values for the substances 7,12-DMBA and etoposide. For the other substances, the Ames test had superior LEC values, which was already observed in a literature survey by Pinter et al (2020). When looking at the reporter gene assays in Table S1, the BlueScreen TM HC and the p53 CALUX R , we found that the HepGentox had lower LEC values for 38% (5 out of 13) of the substances.…”

“…For this, the lowest effective concentration (LEC) value has to be taken into account, which is the lowest concentration of a genotoxin, where a positive result is obtained in a given in vitro bioassay. Recent publications cover the subject of analytical sensitivity of some genotoxicity assays (Rainer et al, 2018;Schilter et al, 2019;Pinter et al, 2020) and came to the conclusion that current methods are not sufficient for the analysis of complex mixtures. In this context, analytical sensitivity refers to an assay's ability to detect substances at low concentrations, meaning low LEC values respond to a high analytical sensitivity and will be referred to as such from now on.…”

Food contact materials: an effect-based evaluation of the presence of hazardous chemicals in paper and cardboard packaging

“…Compared to the classical chromosomal aberration test, where structural chromosomal damage is studied, the MN test allows detection of both structural and numerical alterations. The usage of TK6 cells has been highlighted to be both highly sensitive and specific (Pinter et al 2020). To our knowledge, this is the first study using a micronucleus test with TK6 cells to investigate paper and board FCMs.…”

Direct Comparison of the Lowest Effect Concentrations of Mutagenic Reference Substances in Two Ames Test Formats

“…Previous investigations [ 14 , 15 , 16 ] focused on the applicability of in vitro genotoxicity assays for packaging migrate safety assessment. Packaging migrates are typical complex mixtures which could contain low levels of genotoxic chemicals.…”

“…Its use has been recommended by an expert group of the International Life Science Institute [ 10 ] as part of a comprehensive safety assessment strategy. Compared to other in vitro tests based on mammalian cells, the Ames test exhibits several advantages, such as lower LEC values [ 14 , 16 ] for most substances, possibly resulting from tolerance to higher solvent concentrations [ 17 , 18 ].…”