In the present work, we prepared three different GO materials via a modified version of the Hummers' method under previously described endotoxin-free conditions . Our production process is characterised by the purification of graphite oxide that is synthesised after the conventional Hummers' method . Since this graphite oxide might still contain residual reagents such as manganese salts and the highly reactive dimanganese heptoxide , we performed multiple centrifugation rounds to progressively purify the mixture and remove these chemical impurities, alongside other metallic species and fulvic acids (Figure ). This purification step is not only essential to obtain the so called "GO layer" as previously described , but also to guarantee its high chemical purity. previously demonstrated that high concentrations of manganese and sulphur, resulting from the original Hummers' method, could have a highly detrimental impact on bacterial viability . On the other hand, neutralised and purified GO dispersions did not exert any significant effect compared to the deionised water control. The strict and controlled separation of the GO flakes from these toxic impurities was therefore implemented in our process in order to guarantee a highly pure GO product, which can be confidently assessed in terms of its biological effects following its exposure to biological systems. This purification step was also implemented to remove graphitic impurities from the final l-GO product (Figure ), due to their greater sedimentation rate compared to l-GO flakes.