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Bisphenol A (BPA), a monomer commonly used to manufacture polycarbonate plastic products and the epoxy resins that line metal cans, has resulted in an increased human exposure because both products come in contact with food and beverages. Since BPA is an endocrine disruptor excessive exposure could affect human reproductive function. In response, various countries have adopted a range of measures, from voluntary reductions to outright bans of BPA in some products, mostly in baby bottles and containers for canned foods. Pregnancy-test like strips in combination with hand-held readers provide a simple and versatile tool to determine the presence of bisphenol A quantitatively, making it a very attractive analytical approach especially for on-site applications at critical points, such as environmental analysis, consumer protection, and materials testing.
Quaternary ammonium compounds (QACs) are widely used as active agents in disinfectants, antiseptics, and preservatives. Despite being in use since the 1940s, there remain multiple open questions regarding their detailed mode-of-action and the mechanisms, including phenotypic heterogeneity, that can make bacteria less susceptible to QACs. To facilitate studies on resistance mechanisms towards QACs, we synthesized a fluorescent quaternary ammonium compound, namely N-dodecyl-N,N-dimethyl-[2-[(4-nitro-2,1,3-benzoxadiazol-7-yl)amino]ethyl]azanium-iodide (NBD-DDA). NBD-DDA is readily detected by flow cytometry and fluorescence microscopy with standard GFP/FITC-settings, making it suitable for molecular and single-cell studies. As a proof-of-concept, NBD-DDA was then used to investigate resistance mechanisms which can be heterogeneous among individual bacterial cells. Our results reveal that the antimicrobial activity of NBD-DDA against Escherichia coli, Staphylococcus aureus and Pseudomonas aeruginosa is comparable to that of benzalkonium chloride (BAC), a widely used QAC, and benzyl-dimethyl-dodecylammonium chloride (BAC12), a mono-constituent BAC with alkyl-chain length of 12 and high structural similarity to NBD-DDA. Characteristic time-kill kinetics and increased tolerance of a BAC tolerant E. coli strain against NBD-DDA suggest that the mode of action of NBD-DDA is similar to that of BAC. As revealed by confocal laser scanning microscopy (CLSM), NBD-DDA is preferentially localized to the cell envelope of E. coli, which is a primary target of BAC and other QACs. Leveraging these findings and NBD-DDA‘s fluorescent properties, we show that reduced cellular accumulation is responsible for the evolved BAC tolerance in the BAC tolerant E. coli strain and that NBD-DDA is subject to efflux mediated by TolC. Overall, NBD-DDA’s antimicrobial activity, its fluorescent properties, and its ease of detection render it a powerful tool to study resistance mechanisms of QACs in bacteria and highlight its potential to gain detailed insights into its mode-of-action.
Engineered nanomaterials (NM) with their large surface-to-volume ratios and their for some materials observed size-dependent functional properties are of increasing relevance for current and future developments in various fields such as medical and pharmaceutical industry, computing and electronics or food and consumer products. The performance and safety of NM are determined by the sum of their intrinsic physicochemical properties. Especially, the particle surface chemistry, which is largely controlled by the chemical nature and density of functional groups (FG) and ligands, is an important key driver for NM performance, stability, and processibility as well as the interaction of NM with the environment. Thus, methods for FG quantification can foster the sustainable development of functional and safe(r) NM.
Aiming at the development of simple, versatile, and multimodal tools for the quantification of many bioanalytically relevant FG and ligands, we investigated and compared various analytical methods commonly used for FG quantification. This includes electrochemical titration methods, dye-based optical assays, and other instrumental analytical techniques such as nuclear magnetic resonance and thermal analysis methods.
The potential of our multimodal approach for FG quantification was demonstrated for commercial and custom-made polymeric and silica particles of varying FG, used as optical pH sensors. In the future, our strategy can contribute to establish multi-method characterization strategies to provide a more detailed picture of the structure-properties relationship.
Nowadays amorphous silica nanoparticles (SiO2-NP) are one of the most abundant engineered nanomaterials, that are highly stable and can be easily produced on a large scale at low cost. Surface functionalized SiO2-NP are of great interest in the life and material sciences, as they can be used e.g. as
drug carriers, fluorescent sensors, and multimodal labels in bioanalytical assays and imaging applications. Their performance in such applications depends not only on particle size, size distribution, and morphology, but also on surface chemistry, i.e. the total number of surface functional groups (FG)
and the number of FG accessible for subsequent functionalization with ligands or biomolecules, which in turn determines surface charge, colloidal stability, biocompatibility, and toxicity. Aiming at the development of simple, versatile, and multimodal tools for the quantification of many bioanalytically relevant FG and ligands, we investigated and compared various analytical methods commonly used for FG quantification. This includes electrochemical titration methods, dye-based optical assays, and other instrumental analytical techniques such as nuclear magnetic resonance and thermal analysis methods.
The potential of our multimodal approach for FG quantification was demonstrated for commercial and custom-made silica particles of varying FG, showing not only an influence of the synthesis methods on the number of FG but also on the performance. In the future, our strategy can contribute to establish multi-method characterization strategies to provide a more detailed picture of the structure-properties relationship.
A process spectroscopic approach for the monitoring of microplastics in environmental samples
(2018)
The potential of Raman and NIR process-spectroscopic as a rapid approach for the estimation of microplastics (MP) in soil matrix were tested. For this purpose, artificial MP/soil mixture samples consisting of standard soils or sand with defined ratios of MP (0,5 – 10 mass% polymer) from polyethylene (PE), polypropylene (PP), polystyrene (PS) and polyethylene terephthalate (PET) were prepared. MP particles with diameters < 2 mm and < 125 µm were obtained from industrial polymer pellets after cryo-milling. Spectra of these mixtures were collected by (i) a process FT-NIR spectrometer equipped with a reflection probe, (ii) by a cw-process Raman spectrometer and (iii) by a time-gated Raman spectrometer using fiber-optic probes. The evaluation of process-spectra was performed by chemometric methods. Whereas MP could be detected by Raman spectroscopy in coastal sand at 0.5 mass%, in standard soil detection of MP was limited to 10 – 5 mass% with the large fraction, but samples containing particles of the 125µm mass-fraction yielded no positive result at all. One reason for the lacking sensitivity could be fluorescence by soil organic matter and thus, in a next test time-gated Raman spectroscopy was applied. However, although being indeed more sensitive to the small particles this method failed at MP < 5 mass% indicating that fluorescence was not the major problem. Finally, FT-NIR was tested. Depending on the polymer, MP contents of 0,5 or 1 mass%, respectively, could be detected in standard soils and polymers identified. Furthermore, this approach could be used for the investigation of mixtures of up to four polymers and in real-world samples from bio-waste fermenter residues.
Regardless of its ban in agricultural use, the triazine derivative terbutryn is still used as a biocide against biofouling in building materials, such as façade colors and roof tiles. From there, it is washed off and may contaminate soil and surface waters, where it shows ecotoxicological behavior. For understanding of these leaching processes, analytical methods that can detect terbutryn in traces are necessary. In contrast to separation methods (Gas (GC) or liquid chromatography (LC)) coupled to mass spectrometry (MS), enzyme immunoassays (EIAs) have proven to be adequate tools for environmental monitoring. Compared to GC-or LC-MS, they are cheap, require only limited time and experimental expertise and enable high throughput. For quantification of terbutryn, EIAs have been established, with a limit of detection (LOD) of 50 ng L-1. For leaching experiments however, a lower LOD would be advantageous.
For detection of terbutryn we developed a direct competitive ELISA based on a monoclonal antibody: Terbutryn and an enzyme-terbutryn conjugate (“tracer”) compete for the binding sites of the immobilized antibody. The bound tracer converts a colourless substrate into a blue-coloured product, which can be detected via spectrophotometry. For quantification of terbutryn, a sigmoidal calibration curve is used. For establishing of the EIA, a new tracer based on a terbutryn surrogate (“hapten”) was synthesized. After optimization of the experimental conditions, the immunoassay exhibited a more than ten-fold increased sensitivity (LOD: 3 ng L-1), high stability against sample compounds (Ca2+, NaCl) and lower cross-reactivities against structurally related compounds, than reported before. Analysis of spiked samples was possible with high precision (inter-assay CV: < 10 %) and accuracy (recoveries from spiked samples: 89 % ± 10%). We therefore propose it as quick and reliable method for leaching studies of terbutryn out of building materials.
The rational synthesis and use of nanomaterials require the characterization of many different properties, ranging from particle size and size distribution over surface chemistry to more applicationrelevant features like optical, electrochemical, and magnetic properties. In the following, several methods for the characterization of functional groups on nanomaterials, like polymer and silica nanoparticles, semiconductor quantum dots, and lanthanide-based upconversion nanocrystals are presented. Additionally, procedures for the measurement of the key spectroscopic performance parameters of nanomaterials with linear and nonlinear photoluminescence, such as the photoluminescence quantum yield, are presented for the UV/vis/NIR/SWIR.
Advances in high-resolution imaging of cell cultures and tissues exposed to airborne particles
(2022)
High-resolution microscopy techniques are fundamental to our ability to investigate and quantify the toxicological effects of airborne particulate pollutants via imaging of in vitro respiratory models exposed to well-characterised particle suspensions and aerosols. However, all imaging methods are limited in their spatio-temporal resolution, information capacity and suitability for minimally invasive time lapse imaging. Many traditional methods are incompatible with the more complex three-dimensional model systems which have potential for greater correlation between in vitro studies and real-world health effects. New techniques can overcome these limits, increasing our ability to understand and measure biological responses to particulate exposure at subcellular, cellular and multicellular scales. We have investigated a variety of microscopy methods for imaging different respiratory models.