In vitro genotoxicity: a signal regulators could not dismiss
In laboratory tests on mammalian cells, both red iron oxide (Fe2O3) and black iron oxide (FeO.Fe2O3) produced signs of DNA damage, and this was seen for both nano-sized and micro-sized particles. The in vitro signal alone does not prove harm in people, but EFSA noted it could not be dismissed because no data from the site where exposure actually happens, the gut wall, were available. In vivo oral dosing studies in rats did not find genotoxic effects in blood cells, but EFSA said those tests did not cover the gastrointestinal lining, which is the relevant target.
Red (Fe2O3) and black (FeO.Fe2O3) iron oxide, both in nano and micro form, were positive in in vitro genotoxicity assays in mammalian cells; in vivo oral studies did not elicit genotoxic effects in rat blood cells, but no data for the gastrointestinal site-of-contact were available.
2015Regulatory reviewEFSA Panel on Food Additives and Nutrient Sources, Scientific Opinion on the re-evaluation of iron oxides and hydroxides (E 172) as food additives, EFSA Journal 2015;13(12):4317
EFSA concluded that 'an adequate assessment of the safety of E 172 could not be carried out because a sufficient biological and toxicological database was not available', and recommended that additional toxicological data be provided.
2015RegulatoryEFSA Journal 2015;13(12):4317
Carcinogenicity and reproductive toxicity: data were not available
When EFSA conducted its 2015 re-evaluation, studies covering long-term cancer risk and effects on reproduction or fetal development either did not exist or were submitted as unpublished company data that the Panel could not independently evaluate. This left two major safety questions formally unanswered. A follow-up EU data call was issued in 2016 and a new risk assessment (EFSA-Q-2021-00178) is ongoing as of 2025.
Carcinogenicity and reproductive and developmental toxicity data were not available and could not be evaluated by the EFSA Panel during the 2015 re-evaluation.
2015RegulatoryEFSA Journal 2015;13(12):4317
The European Commission issued a call for scientific and technical data on E172 in December 2016 to address the identified data gaps; a follow-up risk assessment (EFSA-Q-2021-00178) is listed as an ongoing risk assessment.
2016RegulatoryEuropean Commission call for data on E172, December 2016; EFSA ongoing work programme 2025
Nanoparticles in commercial E172 samples
Studies measuring real commercial E172 pigments found that nano-sized particles, those below 100 nanometres, were present in most samples tested. Particle size was not included in food-grade specifications at the time of the 2015 opinion, so manufacturers were not required to control or declare this fraction. Iron oxide nanoparticles have been shown to reach the intestinal wall in particle form after passing through artificial digestion. Cell studies showed minimal uptake across the intestinal barrier and no cytotoxicity at low concentrations, but liver cell studies at higher concentrations showed signs of stress. EFSA flagged the absence of particle-size specifications as a gap.
Nanoparticles were detected in all five commercial E172 pigments examined by transmission electron microscopy or small-angle X-ray scattering; four of five had a size median below 100 nm.
2020LabBauer et al., Food Chemistry, 2020 (presence of iron oxide nanoparticles in food pigment E172)
Iron oxide E172 particles passed through artificial gastrointestinal digestion without dissolving and reached intestinal Caco-2 cells in particle form; minor uptake into cells was detected but almost no transport across the barrier.
2024LabParticulate iron oxide food colorants (E 172) during artificial digestion and their uptake and impact on intestinal cells, Food and Chemical Toxicology, 2024
EFSA recommended that particle size and particle size distribution should be included in the specifications for E172 and that grouping of different E172 forms for risk assessment purposes is not advisable due to variation in physicochemical properties.
2015RegulatoryEFSA Journal 2015;13(12):4317; physicochemical characterisation study, 2025
Gut microbiota: high-dose animal findings
Rat studies using food-grade iron(III) oxide nanoparticles at high doses found changes in the gut microbiota, with increases in bacteria associated with inflammation and obesity and reductions in beneficial Bifidobacterium. These effects appeared at doses substantially higher than typical dietary exposure from food colouring use. Extrapolating animal high-dose findings directly to human consumers at normal intake levels is not straightforward, and no human studies have been conducted.
High-dose iron(III) oxide nanoparticle exposure in rats (200 mg/kg body weight over 28 days) changed the abundance and composition of gut bacteria, increasing inflammation-associated species and reducing Bifidobacterium; lower doses showed minimal effects on microbial diversity.
2024AnimalEffects of food-grade iron(III) oxide nanoparticles on cecal digesta- and mucosa-associated microbiota and short-chain fatty acids in rats, PMC10767317, MDPI Nutrients, 2024
Heavy metal impurities in the pigment
Iron oxide pigments can carry trace impurities of lead, arsenic, cadmium, and mercury from their production process. EU specifications set maximum limits for these contaminants in food-grade E172, and regulators have called for revision of those limits. The concern is about the impurities in the pigment, not the iron oxide itself.
Purity concerns for E172 include possible contamination with heavy metals including lead and arsenic; regulatory reviews have called for stricter impurity limits and revision of current EU specification maximum levels for cadmium, arsenic, lead, and mercury.
2015RegulatoryEFSA Journal 2015;13(12):4317; EU Commission Regulation (EU) No 231/2012 (specifications for E172)