{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,30]],"date-time":"2026-09-30T09:18:39Z","timestamp":1790759919131,"version":"4.1.0"},"reference-count":54,"publisher":"MDPI AG","issue":"13","license":[{"start":{"date-parts":[[2023,6,22]],"date-time":"2023-06-22T00:00:00Z","timestamp":1687392000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Nanomaterials"],"abstract":"<jats:p>In the present study, we addressed the knowledge gaps regarding the agglomeration behavior and fate of food-grade titanium dioxide (E 171) in human gastrointestinal digestion (GID). After thorough multi-technique physicochemical characterization including TEM, single-particle ICP-MS (spICP-MS), CLS, VSSA determination and ELS, the GI fate of E 171 was studied by applying the in vitro GID approach established for the regulatory risk assessment of nanomaterials in Europe, using a standardized international protocol. GI fate was investigated in fasted conditions, relevant to E 171 use in food supplements and medicines, and in fed conditions, with both a model food and E 171-containing food samples. TiO2 constituent particles were resistant to GI dissolution, and thus, their stability in lysosomal fluid was investigated. The biopersistence of the material in lysosomal fluid highlighted its potential for bioaccumulation. For characterizing the agglomeration degree in the small intestinal phase, spICP-MS represented an ideal analytical tool to overcome the limitations of earlier studies. We demonstrated that, after simulated GID, in the small intestine, E 171 (at concentrations reflecting human exposure) is present with a dispersion degree similar to that obtained when dispersing the material in water by means of high-energy sonication (i.e., \u226570% of particles &lt;250 nm).<\/jats:p>","DOI":"10.3390\/nano13131908","type":"journal-article","created":{"date-parts":[[2023,6,23]],"date-time":"2023-06-23T01:54:56Z","timestamp":1687485296000},"page":"1908","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":15,"title":["Agglomeration Behavior and Fate of Food-Grade Titanium Dioxide in Human Gastrointestinal Digestion and in the Lysosomal Environment"],"prefix":"10.3390","volume":"13","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-2739-740X","authenticated-orcid":false,"given":"Francesca","family":"Ferraris","sequence":"first","affiliation":[{"name":"Istituto Superiore di Sanit\u00e0\u2014National Institute of Health, 00161 Rome, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Andrea","family":"Raggi","sequence":"additional","affiliation":[{"name":"Istituto Superiore di Sanit\u00e0\u2014National Institute of Health, 00161 Rome, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4961-8405","authenticated-orcid":false,"given":"Jessica","family":"Ponti","sequence":"additional","affiliation":[{"name":"European Commission, Joint Research Centre (JRC), 21027 Ispra, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3482-2996","authenticated-orcid":false,"given":"Dora","family":"Mehn","sequence":"additional","affiliation":[{"name":"European Commission, Joint Research Centre (JRC), 21027 Ispra, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Douglas","family":"Gilliland","sequence":"additional","affiliation":[{"name":"European Commission, Joint Research Centre (JRC), 21027 Ispra, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sara","family":"Savini","sequence":"additional","affiliation":[{"name":"Istituto Superiore di Sanit\u00e0\u2014National Institute of Health, 00161 Rome, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6298-0700","authenticated-orcid":false,"given":"Francesca","family":"Iacoponi","sequence":"additional","affiliation":[{"name":"Istituto Superiore di Sanit\u00e0\u2014National Institute of Health, 00161 Rome, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Federica","family":"Aureli","sequence":"additional","affiliation":[{"name":"Istituto Superiore di Sanit\u00e0\u2014National Institute of Health, 00161 Rome, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Luigi","family":"Calzolai","sequence":"additional","affiliation":[{"name":"European Commission, Joint Research Centre (JRC), 21027 Ispra, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3454-7947","authenticated-orcid":false,"given":"Francesco","family":"Cubadda","sequence":"additional","affiliation":[{"name":"Istituto Superiore di Sanit\u00e0\u2014National Institute of Health, 00161 Rome, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2023,6,22]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"10726","DOI":"10.1021\/acs.langmuir.7b02729","article-title":"Impact of an Artificial Digestion Procedure on Aluminum-Containing Nanomaterials","volume":"33","author":"Sieg","year":"2017","journal-title":"Langmuir"},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"724","DOI":"10.1021\/acsanm.9b02236","article-title":"Environmental Impact of ZnO Nanoparticles Evaluated by in Vitro Simulated Digestion","volume":"3","author":"Voss","year":"2019","journal-title":"ACS Appl. Nano Mater."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"2441","DOI":"10.1021\/nn204728k","article-title":"Presence of Nano-Sized Silica during In Vitro Digestion of Foods Containing Silica as a Food Additive","volume":"6","author":"Peters","year":"2012","journal-title":"ACS Nano"},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"1198","DOI":"10.3109\/17435390.2012.726382","article-title":"Behaviour of silver nanoparticles and silver ions in an in vitro human gastrointestinal digestion model","volume":"7","author":"Walczak","year":"2012","journal-title":"Nanotoxicology"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"10","DOI":"10.1016\/j.tox.2011.11.004","article-title":"Models for oral uptake of nanoparticles in consumer products","volume":"291","author":"Roblegg","year":"2012","journal-title":"Toxicology"},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"609","DOI":"10.1002\/wnan.1333","article-title":"Mammalian gastrointestinal tract parameters modulating the integrity, surface properties, and absorption of food-relevant nanomaterials","volume":"7","author":"Bellmann","year":"2015","journal-title":"WIREs Nanomed. Nanobiotechnol."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"1255","DOI":"10.1515\/hsz-2015-0145","article-title":"Impact of food components during in vitro digestion of silver nanoparticles on cellular uptake and cytotoxicity in intestinal cells","volume":"396","author":"Lichtenstein","year":"2015","journal-title":"Biol. Chem."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"98","DOI":"10.1016\/j.tifs.2023.01.017","article-title":"Regulatory safety assessment of nanoparticles for the food chain in Europe","volume":"134","author":"Schoonjans","year":"2023","journal-title":"Trends Food Sci. Technol."},{"key":"ref_9","doi-asserted-by":"crossref","unstructured":"EFSA Scientific Committee, More, S., Bampidis, V., Benford, D., Bragard, C., Halldorsson, T., Hern\u00e1ndez-Jerez, A., Bennekou, S.H., Koutsoumanis, K., and Lambr\u00e9, C. (2021). Guidance on risk assessment of nanomaterials to be applied in the food and feed chain: Human and animal health. EFSA J., 19, e06768.","DOI":"10.2903\/j.efsa.2021.6768"},{"key":"ref_10","first-page":"59","article-title":"Titanium Dioxide: A Review","volume":"69","author":"Braun","year":"1997","journal-title":"J. Coat. Technol."},{"key":"ref_11","doi-asserted-by":"crossref","unstructured":"Verleysen, E., Waegeneers, N., Brassinne, F., De Vos, S., Jimenez, I.O., Mathioudaki, S., and Mast, J. (2020). Physicochemical Characterization of the Pristine E171 Food Additive by Standardized and Validated Methods. Nanomaterials, 10.","DOI":"10.3390\/nano10030592"},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"108492","DOI":"10.1016\/j.foodcont.2021.108492","article-title":"Towards a generic protocol for measuring the constituent particle size distribution of E171 in food by electron microscopy","volume":"132","author":"Verleysen","year":"2021","journal-title":"Food Control."},{"key":"ref_13","first-page":"e06585","article-title":"Safety assessment of titanium dioxide (E171) as a food additive","volume":"19","author":"Younes","year":"2021","journal-title":"EFSA J."},{"key":"ref_14","unstructured":"(2023, May 29). Food Directorate Health Canada State of the Science of Titanium Dioxide (TiO2) as a Food Additive. Available online: https:\/\/publications.gc.ca\/collections\/collection_2022\/sc-hc\/H164-341-2022-eng.pdf."},{"key":"ref_15","unstructured":"(2023, May 29). UK Committee on Toxicity of Chemicals in Food, C.P. and the E. Interim Position Paper on Titanium Dioxide, Available online: https:\/\/cot.food.gov.uk\/sites\/default\/files\/2022-01\/TiO2%20COT%20Interim%20position%20paper.pdf."},{"key":"ref_16","unstructured":"(2023, May 29). Food Standards Australia New Zealand (FSANZ) Titanium Dioxide as a Food Additive, Available online: https:\/\/www.foodstandards.gov.au\/consumer\/foodtech\/Documents\/FSANZ_TiO2_Assessment_report.pdf."},{"key":"ref_17","doi-asserted-by":"crossref","unstructured":"Bettencourt, A., Gon\u00e7alves, L.M., Gramacho, A.C., Vieira, A., Rolo, D., Martins, C., Assun\u00e7\u00e3o, R., Alvito, P., Silva, M.J., and Louro, H. (2020). Analysis of the Characteristics and Cytotoxicity of Titanium Dioxide Nanomaterials Following Simulated In Vitro Digestion. Nanomaterials, 10.","DOI":"10.3390\/nano10081516"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"37","DOI":"10.1016\/j.impact.2018.12.002","article-title":"Impact of protein-nanoparticle interactions on gastrointestinal fate of ingested nanoparticles: Not just simple protein corona effects","volume":"13","author":"Cao","year":"2018","journal-title":"Nanoimpact"},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"5975","DOI":"10.1039\/D1FO00499A","article-title":"In vitro digestion of food grade TiO2(E171) and TiO2 nanoparticles: Physicochemical characterization and impact on the activity of digestive enzymes","volume":"12","author":"Dudefoi","year":"2021","journal-title":"Food Funct."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"1087","DOI":"10.1080\/17435390.2017.1398356","article-title":"Potential impact of inorganic nanoparticles on macronutrient digestion: Titanium dioxide nanoparticles slightly reduce lipid digestion under simulated gastrointestinal conditions","volume":"11","author":"Li","year":"2017","journal-title":"Nanotoxicology"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"11568","DOI":"10.1039\/D0RA09706C","article-title":"Influences of a standardized food matrix and gastrointestinal fluids on the physicochemical properties of titanium dioxide nanoparticles","volume":"11","author":"Li","year":"2021","journal-title":"RSC Adv."},{"key":"ref_22","doi-asserted-by":"crossref","unstructured":"Marucco, A., Prono, M., Beal, D., Alasonati, E., Fisicaro, P., Bergamaschi, E., Carriere, M., and Fenoglio, I. (2020). Biotransformation of Food-Grade and Nanometric TiO2 in the Oral\u2013Gastro\u2013Intestinal Tract: Driving Forces and Effect on the Toxicity toward Intestinal Epithelial Cells. Nanomaterials, 10.","DOI":"10.3390\/nano10112132"},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"29","DOI":"10.1186\/s12989-018-0265-1","article-title":"Ingested engineered nanomaterials: State of science in nanotoxicity testing and future research needs","volume":"15","author":"Sohal","year":"2018","journal-title":"Part. Fibre Toxicol."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"1274","DOI":"10.1039\/C4JA00357H","article-title":"Single particle ICP-MS combined with a data evaluation tool as a routine technique for the analysis of nanoparticles in complex matrices","volume":"30","author":"Peters","year":"2015","journal-title":"J. Anal. At. Spectrom."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"2270","DOI":"10.1021\/ac402980q","article-title":"Single Particle Inductively Coupled Plasma Mass Spectrometry: A Powerful Tool for Nanoanalysis","volume":"86","author":"Laborda","year":"2013","journal-title":"Anal. Chem."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"15","DOI":"10.1016\/j.teac.2016.02.001","article-title":"Single particle inductively coupled plasma mass spectrometry for the analysis of inorganic engineered nanoparticles in environmental samples","volume":"9","author":"Laborda","year":"2016","journal-title":"Trends Environ. Anal. Chem."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"9979","DOI":"10.1021\/acs.jafc.0c07363","article-title":"Single-Particle ICP\u2013MS as a Screening Technique for the Presence of Potential Inorganic Nanoparticles in Food","volume":"69","author":"Vidmar","year":"2021","journal-title":"J. Agric. Food Chem."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"55","DOI":"10.1016\/j.jtemb.2019.04.003","article-title":"Possibilities of single particle\u2014ICP-MS for determining\/characterizing titanium dioxide and silver nanoparticles in human urine","volume":"54","author":"Badalova","year":"2019","journal-title":"J. Trace Elem. Med. Biol."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"1740","DOI":"10.1039\/C9JA00206E","article-title":"A critical review of single particle inductively coupled plasma mass spectrometry\u2014A step towards an ideal method for nanomaterial characterization","volume":"35","author":"Mozhayeva","year":"2019","journal-title":"J. Anal. At. Spectrom."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"1380","DOI":"10.1039\/C9JA00101H","article-title":"Combination of 47Ti and 48Ti for the determination of highly polydisperse TiO2 particle size distributions by spICP-MS","volume":"34","author":"Bucher","year":"2019","journal-title":"J. Anal. At. Spectrom."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"991","DOI":"10.1038\/s41596-018-0119-1","article-title":"INFOGEST static in vitro simulation of gastrointestinal food digestion","volume":"14","author":"Brodkorb","year":"2019","journal-title":"Nat. Protoc."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"170","DOI":"10.1016\/j.yrtph.2014.06.021","article-title":"Inter-laboratory validation of bioaccessibility testing for metals","volume":"70","author":"Henderson","year":"2014","journal-title":"Regul. Toxicol. Pharmacol."},{"key":"ref_33","doi-asserted-by":"crossref","unstructured":"Pelfr\u00eane, A., Cave, M.R., Wragg, J., and Douay, F. (2017). In Vitro Investigations of Human Bioaccessibility from Reference Materials Using Simulated Lung Fluids. Int. J. Environ. Res. Public Health, 14.","DOI":"10.3390\/ijerph14020112"},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"123","DOI":"10.1016\/j.tiv.2004.08.001","article-title":"Characterization of phagolysosomal simulant fluid for study of beryllium aerosol particle dissolution","volume":"19","author":"Stefaniak","year":"2005","journal-title":"Toxicol. Vitr."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"11","DOI":"10.1186\/s12989-015-0088-2","article-title":"Dissolution and biodurability: Important parameters needed for risk assessment of nanomaterials","volume":"12","author":"Utembe","year":"2015","journal-title":"Part. Fibre Toxicol."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"675","DOI":"10.1039\/b302257a","article-title":"Bioaccessibility testing of cobalt compounds","volume":"5","author":"Stopford","year":"2003","journal-title":"J. Environ. Monit."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"9361","DOI":"10.1021\/ac201952t","article-title":"Determining Transport Efficiency for the Purpose of Counting and Sizing Nanoparticles via Single Particle Inductively Coupled Plasma Mass Spectrometry","volume":"83","author":"Pace","year":"2011","journal-title":"Anal. Chem."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"2242","DOI":"10.1021\/es204168d","article-title":"Titanium Dioxide Nanoparticles in Food and Personal Care Products","volume":"46","author":"Weir","year":"2012","journal-title":"Environ. Sci. Technol."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"373","DOI":"10.3109\/17435390.2014.940404","article-title":"Using physiologically based pharmacokinetic (PBPK) modeling for dietary risk assessment of titanium dioxide (TiO2) nanoparticles","volume":"9","author":"Bachler","year":"2014","journal-title":"Nanotoxicology"},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"1404","DOI":"10.1080\/17435390.2016.1222457","article-title":"Oral intake of added titanium dioxide and its nanofraction from food products, food supplements and toothpaste by the Dutch population","volume":"10","author":"Rompelberg","year":"2016","journal-title":"Nanotoxicology"},{"key":"ref_41","doi-asserted-by":"crossref","unstructured":"EFSA Panel on Food Additives and Nutrient Sources added to Food (ANS) (2016). Re-evaluation of titanium dioxide (E 171) as a food additive. EFSA J., 14, 4545.","DOI":"10.2903\/j.efsa.2016.4545"},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"985","DOI":"10.1080\/17435390.2020.1778809","article-title":"Possible effects of titanium dioxide particles on human liver, intestinal tissue, spleen and kidney after oral exposure","volume":"14","author":"Brand","year":"2020","journal-title":"Nanotoxicology"},{"key":"ref_43","first-page":"1","article-title":"Review Possible Adverse Effects of Food Additive E171 (Titanium Dioxide) Related to Particle Specific Human Toxicity, Including the Immune System","volume":"22","author":"Bischoff","year":"2021","journal-title":"Int. J. Mol. Sci."},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"15","DOI":"10.1186\/s12989-018-0251-7","article-title":"Detection of titanium particles in human liver and spleen and possible health implications","volume":"15","author":"Heringa","year":"2018","journal-title":"Part. Fibre Toxicol."},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"107550","DOI":"10.1016\/j.foodcont.2020.107550","article-title":"Particle size analysis of pristine food-grade titanium dioxide and E 171 in confectionery products: Interlaboratory testing of a single-particle inductively coupled plasma mass spectrometry screening method and confirmation with transmission electron microscopy","volume":"120","author":"Geiss","year":"2020","journal-title":"Food Control."},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"1014","DOI":"10.1007\/s11051-012-1014-2","article-title":"Nanoparticle dispersion in environmentally relevant culture media: A TiO2 case study and considerations for a general approach","volume":"14","author":"Horst","year":"2012","journal-title":"J. Nanoparticle Res."},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"299","DOI":"10.1080\/10937404.2015.1074969","article-title":"Techniques and Protocols for Dispersing Nanoparticle Powders in Aqueous Media\u2014Is there a Rationale for Harmonization?","volume":"18","author":"Hartmann","year":"2015","journal-title":"J. Toxicol. Environ. Health Part B"},{"key":"ref_48","first-page":"6935","article-title":"Safety of Iron Hydroxide Adipate Tartrate as a Novel Food to Regulation (EU) 2015\/2283 and as a Source of iron in the Context of Directive 2002\/46\/EC","volume":"19","author":"Turck","year":"2021","journal-title":"EFSA J."},{"key":"ref_49","doi-asserted-by":"crossref","unstructured":"Pichot, R. (2015). Particle-Stabilized Emulsions and Colloids: Formation and Applications, RSC.","DOI":"10.1039\/9781782620143-00247"},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"2726","DOI":"10.3168\/jds.S0022-0302(97)76234-9","article-title":"The Influence of Fat Substitutes Based on Protein and Titanium Dioxide on the Sensory Properties of Lowfat Milks","volume":"80","author":"Phillips","year":"1997","journal-title":"J. Dairy Sci."},{"key":"ref_51","unstructured":"Kuznesof, P.M. (2006). Titanium Dioxide: Chemical and Technical Assessment, JECFA. Reviewed by M.V. Rao."},{"key":"ref_52","doi-asserted-by":"crossref","first-page":"221","DOI":"10.1002\/(SICI)1097-0010(199702)73:2<221::AID-JSFA708>3.0.CO;2-U","article-title":"The Improved Whitening of Minced Cod Flesh Using Dispersed Titanium Dioxide","volume":"73","author":"Meacock","year":"1997","journal-title":"J. Sci. Food Agric."},{"key":"ref_53","doi-asserted-by":"crossref","first-page":"152","DOI":"10.1039\/C8EN00760H","article-title":"Nanomaterial identification of powders: Comparing volume specific surface area, X-ray diffraction and scanning electron microscopy methods","volume":"6","author":"Dazon","year":"2018","journal-title":"Environ. Sci. Nano"},{"key":"ref_54","unstructured":"Mech, A., Rauscher, H., Rasmussen, K., Babick, F., Hodoroaba, V.-D., Ghanem, A., Wohlleben, W., Marvin, H., Br\u00fcngel, R., and Friedrich, C.M. (2020). The NanoDefine Methods Manual, European Commission, Joint Research Centre, Publications Office of the European Union."}],"container-title":["Nanomaterials"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2079-4991\/13\/13\/1908\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T19:58:33Z","timestamp":1760126313000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2079-4991\/13\/13\/1908"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2023,6,22]]},"references-count":54,"journal-issue":{"issue":"13","published-online":{"date-parts":[[2023,7]]}},"alternative-id":["nano13131908"],"URL":"https:\/\/doi.org\/10.3390\/nano13131908","relation":{},"ISSN":["2079-4991"],"issn-type":[{"value":"2079-4991","type":"electronic"}],"subject":[],"published":{"date-parts":[[2023,6,22]]}}}