2D-nanomaterials (2DNM) including graphene family nanomaterials (GFNs) and transition metal dichalcogenides (TMDs) are being researched and developed for commercial use, due to their wide spectrum of applications, ranging from electronic devices to sensors. GFNs include the Nobel Prize winning material, Graphene, and it is currently starting to be produced in mass scale.  Graphene Oxide (GO) is a functionalised form of graphene and is easily exfoliated in aqueous medium.  GOs suspend more regularly than graphene in water, and therefore offer more possibilities for up-scaling in certain applications of GFNs. However, the life cycle of these materials after their application and an understanding of their impact on the environment are still in their infancy.

Following an initial literature review of the toxicity of GFNs, it was found that:

• GOs may become the most prominent GFN material in the environment due to their dispersibility in aqueous media and their ease of production and scalability.
• There is currently very limited literature on the impact of GOs on terrestrial environments.

As mentioned above, there is very limited literature on the assessment of GOs on the environment especially against environmentally relevant bacteria. However, one of the characteristics frequently studied is the antimicrobial activity of these materials. A literature survey of GOs against common microbial highlighted the following:

• Physical characteristics such as the lateral dimension of GO affects the degree of interaction with bacteria 
• There is a lack of rigour in the characterisation of GOs exposed to bacteria and significant variability between the physical properties of samples tested making it hard to draw conclusions about effects of GO on  bacterial health and function.
• The mechanism of interaction between GO and soil bacteria is still unclear.

Here we exposed 2 different soil bacteria to different lateral sized GOs prepared by a Modified Hummer’s method, and employed established assays of bacterial activity together with spectromicroscopy to understand the interaction mechanism.

The hypotheses used to select the diameters were that large diameter GO could damage bacteria by wrapping around the bacteria, starving them of nutrients; smaller diameter GO could insert into the cell wall of the bacteria causing mechanical damage.

Toxicity assessments of nanomaterials against soil bacteria are still in their infancy and efforts to assess the toxicity of any 2DNM towards soil bacteria have not been performed yet. There are protocols published by the Centre for Ecology and Hydrology (CEH) who are leading experts in environmental science in the UK and globally to test nanomaterials against model soil bacteria. Therefore, the toxicity tests were carried out in collaboration with CEH.

To understand the interaction of GOs with bacteria, some samples were processed for TEM imaging. TEM imaging of carbon nanomaterials with cells can be powerful complementary technique to understand the mechanism of interaction. However, TEM imaging of soil bacteria as well as GOs with biological samples have not been widely used.  These images demonstrate the possibility of using imaging to understand mechanisms by which GO with different diameters interacts with soil bacteria.

The current dataset of toxicity assay suggests that GO ages over time which reduces its toxicity towards soil bacteria. It is also possible that GO acts as a food source for bacteria.  Next steps will be to characterise the GO using XPS and Raman spectroscopy to compare the functional chemistry of a fresh batch of GO and an aged batch of GO.  We will also compare the toxicity of a new batch of GO with the aged GO using the new modified ISO protocol.