Biotransformation and biological impact of graphene and graphene oxide during simulated oral ingestion

Graphene is an innovative nanomaterial, made of two-dimensional honeycomb-like carbon lattice, with potential in many different applications. Studying the behaviour of graphene-related materials (GRMs) in biological systems is, therefore, crucial to assess possible side effects. In this work is studied the biotransformation and biological impact of few-layer pristine graphene (FLG) and graphene oxide (GOX), following ingestion as exposure route. To mimic FLG and GOX ingestion, an in vitro digestion assay based on a standardized operating procedure (SOP) is applied. The assay simulates the human ingestion of GRMs during their dynamic passage through the different environments of gastrointestinal (GI) tract (salivary, gastric, intestinal). Physical-chemical changes of GRMs during the digestion process are assessed through a detailed Raman spectroscopy characterization. Moreover, the effect of chronic exposure to digested GRMs on integrity and functionality of an in vitro model of intestinal barrier is also determined according to a second SOP. Our results showed that the FLG and GOX nanoflakes aggregates after experiencing the passage through the different environments of GI tract, with evident doping effects, due to the interaction of GRMs with digestive juice components and the strong pH variations. Interestingly, neither structural changes nor degradation of the nanomaterials is observed, suggesting that GRMs are bio-persistent nanomaterials when administered by oral route. Chronic exposure to digested GRMs does not affect intestinal barrier integrity and is not associated to inflammation and cytotoxicity, though possible long-term adverse effects cannot be ruled out, due to the observed bio-durability of the materials.