Microscopy techniques are indispensable to the nanoanalytical toolbox and can provide accurate information on the number size distribution and number concentration of nanoparticles (NPs) at low concentrations (ca. ppt to ppb range) and small sizes (ca. <20 nm). However, the high capabilities of microscopy techniques are limited by the traditional sample preparation based on drying a small volume of suspension of NPs on a microscopy substrate. This method is limited by low recovery of NPs (ca. <10%), formation of aggregates during the drying process, and thus, the complete misrepresentation of the NP suspensions under consideration. This paper presents a validated quantitative sampling technique for atomic force microscopy (AFM) that overcomes the above-mentioned shortcomings and allows full recovery and representativeness of the NPs under consideration by forcing the NPs into the substrate via ultracentrifugation and strongly attaches the NPs to the substrate by surface functionalization of the substrate or by adding cations to the NP suspension. The high efficiency of the analysis is demonstrated by the uniformity of the NP distribution on the substrate (that is low variability between the number of NPs counted on different images on different areas of the substrate), the high recovery of the NPs up to 71%) and the good correlation (R > 0.95) between the mass and number concentrations. Therefore, for the first time, we developed a validated quantitative sampling technique that enables the use of the full capabilities of microscopy tools to quantitatively and accurately determine the number size distribution and number concentration of NPs at environmentally relevant low concentrations (i.e. 0.34-100 ppb). This approach is of high environmental relevance and can be applied widely in environmental nanoscience and nanotoxicology for (i) measuring the number concentration dose in nanotoxicological studies and (ii) accurately measuring the number size distribution of NPs; both are key requirements for the implementation of the European Commission recommendation for definition of nanomaterials.
Environmental context. Studies of manufactured nanoparticles (NPs) in the environment have been performed almost exclusively at high NP concentrations. These data lead to misunderstandings related to NP fate and effects at relevant environmental concentrations, which are expected to be low. A better understanding of the concentration-dependent behaviour of NPs will improve our understanding of their fate and effects under environmentally realistic conditions.Abstract. This rapid communication highlights the importance of nanoparticle concentration in determining their environmental fate and behaviour. Notably, two fate processes have been considered: dissolution and aggregation. The decrease in nanoparticle concentration results in increased dissolution and decreased aggregate sizes, inferring higher potential for environmental transport of nanoparticles. The behaviour (e.g, dissolution, aggregation, disaggregation) and fate (e.g. mobility, fugacity, non-transient (sink) or transient source) of nanoparticles (NPs) in environmental and toxicological media have been investigated for over a decade, typically at high NP concentrations (e.g. milligram per litre range) which are not relevant to the environment, [1] resulting in some potentially misleading assumptions that (i) NP behaviour is dominated by aggregation and thus their fate is dominated by sedimentation and removal from the water column, or, in porous media, deposition and removal from the aqueous phase [2] ; (ii) NP dissolution is limited for many NPs and rarely are all NPs dissolved fully in environmental and biological media over relevant timescales [1] and (iii) many NPs therefore impart little or no toxic risk to pelagic organisms as a result of limited NP dissolution and NP removal by aggregation and sedimentation. [3] Several NP groups (e.g. Ag NPs, Cu NPs, Cd NPs, ZnO) may undergo dissolution and release ions with well known toxic effects. [4] These various issues complicate NP risk characterisation and are exacerbated by the general use of high NP concentrations in NP fate, behaviour and ecotoxicological studies. [2,5] Use of high NP concentrations has been motivated by poor detection limits of available analytical techniques (e.g. dynamic light scattering, laser Doppler electrophoresis, UV-Vis spectroscopy) together with enhanced likelihood of observing more pronounced changes and effects at high NP concentrations. [6] Furthermore, most published nanoecotoxicological data are acute exposure studies, which also drive the high concentration selection bias in order to generate measurable biological responses. Many NPs tested for toxicity to aquatic organisms have been non-toxic on acute time scales until they reach unrealistically high exposure concentrations. Clear predictive linkages between unrealistic high acute exposures and more realistic low chronic exposures have not been established for aquatic systems, and are likely to be further complicated by differing concentration-dependent behaviours of NPs.Despite these concerns, little attention ha...
The present study involves the integrated network pharmacology and phytoinformatics-based investigation of phytocompounds from Ocimum tenuiflorum against diabetes mellitus-linked Alzheimer’s disease. It aims to investigate the mechanism of the Ocimum tenuiflorum phytocompounds in the amelioration of diabetes mellitus-linked Alzheimer’s disease through network pharmacology, druglikeness and pharmacokinetics, molecular docking simulations, GO analysis, molecular dynamics simulations, and binding free energy analyses. A total of 14 predicted genes of the 26 orally bioactive compounds were identified. Among these 14 genes, GAPDH and AKT1 were the most significant. The network analysis revealed the AGE-RAGE signaling pathway to be a prominent pathway linked to GAPDH with 50.53% probability. Upon the molecular docking simulation with GAPDH, isoeugenol was found to possess the most significant binding affinity (−6.0 kcal/mol). The molecular dynamics simulation and binding free energy calculation results also predicted that isoeugenol forms a stable protein–ligand complex with GAPDH, where the phytocompound is predicted to chiefly use van der Waal’s binding energy (−159.277 kj/mol). On the basis of these results, it can be concluded that isoeugenol from Ocimum tenuiflorum could be taken for further in vitro and in vivo analysis, targeting GAPDH inhibition for the amelioration of diabetes mellitus-linked Alzheimer’s disease.
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