60 61 D I A M O N D L I G H T S O U R C E A N N U A L R E V I E W 2 0 1 9 / 2 0 D I A M O N D L I G H T S O U R C E A N N U A L R E V I E W 2 0 1 9 / 2 0 Imaging andMicroscopy Group Beamline I14 Evaluating the environmental impact of zinc oxide nanomaterials inmunicipal wastewater Related publication: Gomez-Gonzalez M. A., Koronfel M. A., Goode A. E., Al-Ejji M.,Voulvoulis N., Parker J. E., Quinn P. D., ScottT. B., Xie F.,YallopM. L., Porter A. E. & RyanM. P. Spatially resolved dissolution and speciation changes of ZnO nanorods during short-term in situ incubation in a simulated wastewater environment. ACS Nano 13 , 11049 (2019). DOI: 10.1021/acsnano.9b02866 Publication keywords: ZnO nanomaterials; X-ray fluorescencemicroscopy; Scanning electronmicroscopy; Spatially resolved in situ ZnO transformations Z inc oxide (ZnO) engineered nanomaterials are used in a wide range of consumer products, such as sunscreens, personal care products, and anti-bacterial agents. There are concerns about their potential impact as an environmental pollutant, with concentrations of ZnO nanomaterials in municipal water predicted to be of the order of milligrams per litre. ZnO nanomaterials may inhibit bacterial processes in urban wastewater treatment plants. They could also adversely affect the health of organisms in soils and waters once treated material is released back into the natural environment. What effect the ZnO nanomaterials will have on the environment depends on how they are transformed by chemical reactions with (e.g.) sulphur or organic carbon in environmental systems. Researchers need spatially-resolved in situ methods to track transformations in real- time. X-ray Fluorescence Microscopy (XFM) has emerged in recent years as a promising technique for imaging environmental samples due to its high spatial resolution and sensitivity to a wide range of elements. Researchers used time-resolved hard X-ray Fluorescence Microscopy on the Hard X-ray Nanoprobe beamline (I14) to monitor dynamic changes to the nanomaterials in situ. They were able to make predictions about the rates at which ZnO nanomaterials transform during the first stages of the wastewater treatment process. Their results can be used to generate fundamental knowledge about the fate of ZnO nanomaterials in conventional municipal sewage systems. Nanostructures of zinc oxide (ZnO) are at the forefront of application- driven nanotechnology because of their unique optical, piezoelectric, semiconductingandantibacterialproperties.Theirexceptionalproperties,such as nanometric size and high surface reactivity, which are utilised or engineered advantageously for many applications, may also increase their toxicity to the environment in the course of their synthesis, use, and disposal. Thus, there is an urgent need to understand their fate as transition from common usage to the environment via wastewater treatment and disposal plants, as well as any potential impact on the (micro)organisms. A methodology based on spatially resolved in situ X-ray Fluorescence Microscopy (XFM) was developed at I14, allowing observation of real-time dissolution and morphological and chemical evolution of synthetic template-grown ZnO nanorods (~ 725 nm length, ~ 140 nm diameter). An in-house liquid cell was designed by the I14 beamline staff, where the templated ZnO nanorods were aligned within the path of the X-ray beam, and subsequently incubated with ~ 200 µL of simulated sludge solution. Primary sludge, mainly formed by organic matter and humic substances, is one of the first physio-chemical barriers that influent or sewage waters encounter when entering the cleaning cycle of a regular wastewater treatment plants (WWTP) and, therefore, humic acid (10 mg L -1 ) was selected as simplified aquatic medium for this study. It is only by applying this simplified system – instead of a more complex real sludge – that accurate proof-of-concept of the feasibility of the technique can be generated. Consecutive XFM maps were acquired in situ during incubation of the ZnO nanorods for two short-time scales (1 hour and 3 hours), allowing evaluation of ZnO morphology changes within the initial stages of their release to sewage waters. A gradual decrease of the Zn fluorescence intensity was observed during the proposed in situ incubation periods. Over the 3-hours experiments, some of the Zn-hotspots dissolved extensively until they were no longer detectable. The expected ZnO nanorod dissolution favoured the release and transport of Zn 2+ ions within the simulated sludge solution, as confirmed by Inductively Coupled Plasma Mass Spectroscopy (ICP-MS). This partial nanorod dissolution was further established by ex situ scanning electron microscopy, exhibiting thinner rod structures smaller than ~200 nm length and ~20 nm width. To obtain chemical and speciation information along the Zn K-edge, further XFM image stacks were collected on the drained Zn-structures at different photon energies 1 . Nine energies (9-E) were selected as follows: i) three for background removal at Zn pre-edge; ii) five post-edge energies to differentiate between the distinctive features of the Zn-species; and iii) a spectrum image at the isosbestic point, where all the Zn-phases presented a similar fluorescence intensity irrespective of their speciation. Hence, speciation maps (SM) were calculated through fitting the absorption data from each pixel to the linear combination of the standard spectra after acquiring 9-E imaging scans (Fig. 1). Quantification maps (QM) were calculated by normalising the speciation maps to the isosbestic point, producing a visual estimation of the real extent of the Zn-species generation. A fluorescence map was also acquired at themaximumenergy of the Zn K-edge X-ray Absorption Near Edge Structure (XANES) spectrum (E max in Fig. 1) for total Zn intensity contribution. After 1 hour incubation of the ZnO nanorods in the simulated sludge (Fig. 1b), ZnS became the predominant species on the surface of the Zn-hotspot according to the SM, which was confirmed by the calculated QM. As expected, this differs from the spectra taken for the “as-synthesised” ZnO, which showed predominant ZnO contribution (Fig. 1a). After the 3 hours incubation, a different behaviour of the ZnO nanorods was found depending on the region analysed. In the first region (Fig. 1c), most of the Zn-hotspots remained as ZnO in the SM. The QM indicated a significant amount of ZnS species generated in areas depleted in ZnO. The second region analysed after 3 hours incubation (Fig. 1d) showed a more disperse variation in the Zn-species in the SM, with ZnS as the dominant species. Although ZnO was still a major component of the larger hotspots in the QM, trace amounts of diffuse Zn-phosphate (Zn 3 (PO 4 ) 2 ) and Zn adsorbed to iron-oxyhydroxides (Zn-Fe(ox)) were alsomapped after the 3 hours incubation, withdifferenttrends intheirspatialvariations.LeBars etal . 2 showedthatnano- ZnS is a major Zn species in raw liquid organic wastes, but it was a minority in solid and more processed organic-rich wastes. Furthermore, the formation of a ZnO-Zn 3 (PO 4 ) 2 core-shell structure has been reported in phosphate-rich environments 3 , where Zn 2+ can be complexed with aqueous PO 4 3- , resulting in the formation of an amorphous zinc phosphate hydrate precipitate 4 . Although Zn 3 (PO 4 ) 2 is the thermodynamically favoured species, the full transformation to these species is kinetically limited to longer reaction times (15 days) 3,4 , which were not achieved here. The higher solubility reported by Ma et al. 5 for the ZnS-shell/ZnO nanomaterials than that expected for bulk ZnS (Ksp = 2·10 -25 yields a solubility of 3.7·10 -10 mM) would kinetically favour the transformation to ZnS phase rather than to Zn 3 (PO 4 ) 2 under these conditions. To confirm the validity of the spectra extracted from the 9-energies speciation, a larger energy resolution image stack was acquired using 135-energies along the Zn K-edge. The generated XANES spectra were compared with the corresponding standards for the expected Zn-species, and subsequently analysed by linear combination fitting (LCF) analyses. An estimation of the ZnO composition of the samples was obtained by LCF, showing a decrease in the ZnO contribution after only 1-hour incubation in the simulated sludge to a ~27-41%, and a further significant decrease to an only ~12-17% of the total Zn content within 3-hours. In this work, spatially resolved maps of ZnO nanomaterial transformation within a simulated environmental medium have been acquired dynamically. These results demonstrated that: i) ZnO nanorods partially transformed to ZnS species predominantly within in situ short-time incubations (1 –3 hours) in a simulated primary sludge medium, ii) minor Zn 3 (PO 4 ) 2 and Zn adsorbed to Fe-oxyhydroxides were also measured in some non-sterically impeded regions, iii) the ongoing ZnO dissolution and the humic acid presence have an influence in the preferential transformation of the remaining ZnO nanorods. As nanomaterials formulations become more complex, the XFM technique would offer the possibility to characterise chemical transformations of individual nanoparticles within complex mixtures and coupled behaviour between the particles.When studying the structural and functional impacts of transformed nanomaterials on organisms, this information could, in future, be used to generate mechanistically underpinned insight to predict nanomaterial behaviour, refine guidelines for safe use and to inform remediation strategies of engineered nanomaterials. References: 1. Koronfel M. A. et al. Chemical evolution of CoCrMo wear particles: an in situ characterization study. J. Phys. Chem. C 123 , 9894 (2019). DOI: 10.1021/acs.jpcc.9b00745 2. Le Bars M. et al. Drastic change in zinc speciation during anaerobic digestion and composting: instability of nanosized zinc sulphide. Environ. Sci. Technol. 52 , 12987 (2018). DOI: 10.1021/acs.est.8b02697 3. Rathnayake S. et al. Multitechnique investigation of the pH dependence of phosphate induced transformations of ZnO nanoparticles. Environ. Sci. Technol. 48 , 4757 (2014). DOI: 10.1021/es404544w 4. Lv J., Dissolution and microstructural transformation of ZnO nanoparticles under the influence of phosphate. Environ. Sci. Technol. 46 , 7215 (2012). DOI: 10.1021/es301027a 5. Ma R., Sulfidation mechanism for zinc oxide nanoparticles and the effect of sulfidation on their solubility. Environ. Sci. Technol. 47 , 2527 (2013). DOI:10.1021/es3035347 Funding acknowledgement: This work and Miguel Gomez-Gonzalez contract were supported by the Natural Environment Research Council project: NE/N006402/1. Mary Ryan acknowledges support from the RAEng/Shell via a Research Chair in Interfacial Nanoscience. Reprinted (adapted) with permission from ACS Nano 2019, 13, 10, 11049-11061, Spatially Resolved Dissolution and Speciation Changes of ZnO Nanorods during Short-Term In Situ Incubation in a SimulatedWastewater Environment. Copyright 2019 American Chemical Society. Corresponding author: Miguel A. Gomez-Gonzalez, Diamond Light Source Ltd., miguel.gomez-
[email protected] Figure 1: 9-Energy X-ray Fluorescence Microscopy (XFM) of: (a) “as-synthesised” ZnO, (b) sample after the 1-hour incubation in a simulated sludge and (c-d) two different regions analysed in the sample after the 3-hours incubation. [Left] Fluorescence map acquired at E max =9669 eV. [Right] Speciation maps (SM) of the expected Zn-species, where the red colour equals a 100% compound contribution and the blue colour corresponds to 0%. Quantification maps (QM) for the same species were calculated after normalising the SM with the isosbestic point. The numbering on the scale bars on the QM provides an estimation of the Zn-species contribution. Scale bars = 750 nm.