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Pyrolysis–Gas Chromatography
(2024)
The methodology of analytical pyrolysis-GC/MS has been known for several years, but is seldom used in research laboratories and process control in the chemical industry. This is due to the relative difficulty of interpreting the identified pyrolysis products as well as the variety of them. This book contains full identification of several classes of polymers/copolymers and biopolymers that can be very helpful to the user. In addition, the practical applications can encourage analytical chemists and engineers to use the techniques explored in this volume.
Process-induced changes in thermo-mechanical viscoelastic properties and the corresponding morphology of biodegradable polybutylene adipate terephthalate (PBAT) and polylactic acid (PLA) blown film blends modified with four multifunctional chain-extending cross-linkers (CECL) were investigated. The introduction of CECL modified the properties of the reference PBAT/PLA blend significantly. The thermal analysis showed that the chemical reactions were incomplete after compounding, and that film blowing extended them. SEM investigations of the fracture surfaces of blown extrusion films reveal the significant effect of CECL on the morphology formed during the processing. The anisotropic morphology introduced during film blowing proved to affect the degradation processes as well. Furthermore, the reactions of CECL with PBAT/PLA induced by the processing depend on the deformation directions. The blow-up ratio parameter was altered to investigate further process-induced changes proving synergy with mechanical and morphological features. Using blown film extrusion, the elongational behavior represents a very important characteristic. However, its evaluation may be quite often problematic, but with the SER Universal Testing Platform it was possible to determine changes in the duration of time intervals corresponding to the rupture of elongated samples.
Traditional and newly developed testing methods were used for extensive application-related characterization of transdermal therapeutic systems (TTS) and pressure sensitive adhesives (PSA). Large amplitude oscillatory shear tests of PSAs were correlated to the material behavior during the patient’s motion and showed that all PSAs were located close to the gel point. Furthermore, an increasing strain amplitude results in stretching and yielding of the PSA´s microstructure causing a consolidation of the network and a release with increasing strain amplitude. RheoTack approach was developed to allow for an advanced tack characterization of TTS with visual inspection. The results showed a clear resin content and rod geometry dependent behavior, and displays the PSA´s viscoelasticity resulting in either high tack and long stretched fibrils or non-adhesion and brittle behavior. Moreover, diffusion of water / sweat during TTS´s application might influence its performance. Therefore, a dielectric analysis based evaluation method displayed occurring water diffusion into the PSA from which the diffusion coefficient can be determined, and showed clear material and resin content dependent behavior. All methods allow for an advanced product-oriented material testing that can be utilized within further TTS development.
A firm link between endoplasmic reticulum (ER) stress and tumors has been wildly reported. Endoplasmic reticulum oxidoreductase 1 alpha (ERO1α), an ER-resident thiol oxidoreductase, is confirmed to be highly upregulated in various cancer types and associated with a significantly worse prognosis. Of importance, under ER stress, the functional interplay of ERO1α/PDI axis plays a pivotal role to orchestrate proper protein folding and other key processes. Multiple lines of evidence propose ERO1α as an attractive potential target for cancer treatment. However, the unavailability of specific inhibitor for ERO1α, its molecular inter-relatedness with closely related paralog ERO1β and the tightly regulated processes with other members of flavoenzyme family of enzymes, raises several concerns about its clinical translation. Herein, we have provided a detailed description of ERO1α in human cancers and its vulnerability towards the aforementioned concerns. Besides, we have discussed a few key considerations that may improve our understanding about ERO1α in tumors.
Introduction: A multitude of findings from cell cultures and animal studies are available to support the anti-cancer properties of cannabidiol (CBD). Since CBD acts on multiple molecular targets, its clinical adaptation, especially in combination with cancer immunotherapy regimen remains a serious concern.
Methods: Considering this, we extensively studied the effect of CBD on the cytokine-induced killer (CIK) cell immunotherapy approach using multiple non-small cell lung cancer (NSCLC) cells harboring diverse genotypes.
Results: Our analysis showed that, a) The Transient Receptor Potential Cation Channel Subfamily V Member 2 (TRPV2) channel was intracellularly expressed both in NSCLC cells and CIK cells. b) A synergistic effect of CIK combined with CBD, resulted in a significant increase in tumor lysis and Interferon gamma (IFN-g) production. c) CBD had a preference to elevate the CD25+CD69+ population and the CD62L_CD45RA+terminal effector memory (EMRA) population in NKT-CIK cells, suggesting early-stage activation and effector memory differentiation in CD3+CD56+ CIK cells. Of interest, we observed that CBD enhanced the calcium influx, which was mediated by the TRPV2 channel and elevated phosphor-Extracellular signal-Regulated Kinase (p-ERK) expression directly in CIK cells, whereas ERK selective inhibitor FR180204 inhibited the increasing cytotoxic CIK ability induced by CBD. Further examinations revealed that CBD induced DNA double-strand breaks via upregulation of histone H2AX phosphorylation in NSCLC cells and the migration and invasion ability of NSCLC cells suppressed by CBD were rescued using the TRPV2 antagonist (Tranilast) in the absence of CIK cells. We further investigated the epigenetic effects of this synergy and found that adding CBD to CIK cells decreased the Long Interspersed Nuclear Element-1 (LINE-1) mRNA expression and the global DNA methylation level in NSCLC cells carrying KRAS mutation. We further investigated the epigenetic effects of this synergy and found that adding CBD to CIK cells decreased the Long Interspersed Nuclear Element-1 (LINE-1) mRNA expression and the global DNA methylation level in NSCLC cells carrying KRAS mutation.
Conclusions: Taken together, CBD holds a great potential for treating NSCLC with CIK cell immunotherapy. In addition, we utilized NSCLC with different driver mutations to investigate the efficacy of CBD. Our findings might provide evidence for CBD-personized treatment with NSCLC patients.
Trade of wild-caught animals is illegal for many taxa and in many countries. Common regulatory procedures involve documentation and marking techniques. However, these procedures are subject to fraud and thus should be complemented by routine genetic testing in order to authenticate the captive-bred origin of animals intended for trade. A suitable class of genetic markers are SNPSTRs that combine a short tandem repeat (STR) and single nucleotide polymorphisms (SNPs) within one amplicon. This combined marker type can be used for genetic identification and for parentage analyses and in addition, provides insight into haplotype history. As a proof of principle, this study establishes a set of 20 SNPSTR markers for Athene noctua, one of the most trafficked owls in CITES Appendix II. These markers can be coamplified in a single multiplex reaction. Based on population data, the percentage of observed and expected heterozygosities of the markers ranged from 0.400 to 1.000 and 0.545 to 0.850, respectively. A combined probability of identity of 5.3*10-23 was achieved with the whole set, and combined parentage exclusion probabilities reached over 99.99%, even if the genotype of one parent was missing. A direct comparison of an owl family and an unrelated owl demonstrated the applicability of the SNPSTR set in parentage testing. The established SNPSTR set thus proved to be highly useful for identifying individuals and analysing parentage to determine wild or captive origin. We propose to implement SNPSTR-based routine certification in wildlife trade as a way to reveal animal laundering and misdeclaration of wild-caught animals.
In this work, the surface reactions of the homemade explosive triacetone triperoxide on tungsten oxide (WO3) sensor surfaces are studied to obtain detailed information about the chemical reactions taking place. Semiconductor gas sensors based on WO3 nanopowders are therefore produced and characterized by scanning electron microscopy, X-ray diffraction, and Raman spectroscopy. To analyze the reaction mechanisms at the sensor surface, the sensor is monitored online under operation conditions using Raman spectroscopy, which allows to identify the temperature-dependent sensor reactions. By combining information from the Raman spectra with data on the changing resistivity of the underlying semiconductor, it is possible to establish a correlation between the adsorbed gas species and the physical properties of the WO3 layer. In the results, it is indicated that a Lewis acid–base reaction is the most likely mechanism for the increase in resistance observed at temperatures below 150 °C. In the results, at higher temperatures, the assumption of a radical mechanism that causes a decrease in resistance is supported.
DT-13 attenuates inflammation by inhibiting NLRP3-inflammasome related genes in RAW264.7 macrophages
(2024)
Plant derived saponins or other glycosides are widely used for their anti-inflammatory, antioxidant, and anti-viral properties in therapeutic medicine. In this study, we focus on understanding the function of the less known steroidal saponin from the roots of Liriope muscari L. H. Bailey – saponin C (also known as DT-13) in lipopolysaccharide (LPS)-stimulated RAW264.7 macrophages in comparison to the well-known saponin ginsenoside Rk1 and anti-inflammatory drug dexamethasone. We proved that DT-13 reduces LPS-induced inflammation by inhibiting nitric oxide (NO) production, interleukin-6 (IL-6) release, cycloxygenase-2 (COX-2), tumour necrosis factor-alpha (TNF-α) gene expression, and nuclear factor kappa-B (NFκB) translocation into the nucleus. It also inhibits the inflammasome component NOD-like receptor family pyrin domain containing protein 3 (NLRP3) regulating the inflammasome activation. This was supported by the significant inhibition of caspase-1 and interleukin-1 beta (IL-1β) expression and release. This study demonstrates the anti-inflammatory effect of saponins on LPS-stimulated macrophages. For the first time, an in vitro study shows the attenuating effect of DT-13 on NLRP3-inflammasome activation. In comparison to the existing anti-inflammatory drug, dexamethasone, and triterpenoid saponin Rk1, DT-13 more efficiently inhibits inflammation in the applied cell culture model. Therefore, DT-13 may serve as a lead compound for the development of new more effective anti-inflammatory drugs with minimised side effects.
Pollution with anthropogenic waste, particularly persistent plastic, has now reached every remote corner of the world. The French Atlantic coast, given its extensive coastline, is particularly affected. To gain an overview of current plastic pollution, this study examined a stretch of 250 km along the Silver Coast of France. Sampling was conducted at a total of 14 beach sections, each with five sampling sites in a transect. At each collection site, a square of 0.25 m2 was marked. The top 5 cm of beach sediment was collected and sieved on-site using an analysis sieve (mesh size 1 mm), resulting in a total of approximately 0.8 m3 of sediment, corresponding to a total weight of 1300 kg of examined beach sediment. A total of 1972 plastic particles were extracted and analysed using infrared spectroscopy, corresponding to 1.5 particles kg−1 of beach sediment. Pellets (885 particles), polyethylene as the polymer type (1349 particles), and particles in the size range of microplastics (943 particles) were most frequently found. The significant pollution by pellets suggests that the spread of plastic waste is not primarily attributable to tourism (in February/March 2023). The substantial accumulation of meso- and macro-waste (with 863 and 166 particles) also indicates that research focusing on microplastics should be expanded to include these size categories, as microplastics can develop from them over time.
Design and characterization of geopolymer foams reinforced with Miscanthus x giganteus fibers
(2024)
This paper presents the effects of different amounts of fibers and foaming agent, as well as different fiber sizes, on the mechanical and thermal properties of fly ash-based geopolymer foams reinforced with Miscanthus x giganteus fibers. The mechanical properties of the geopolymer foams were measured through compressive strength, and their thermal properties were characterized by thermal conductivity and X-ray micro-computed tomography. Furthermore, design of experiment (DoE) were used to optimize the thermal conductivity and compressive strength of Miscanthus x giganteus reinforced geopolymer foams. In addition, the microstructure was studied using X-ray diffraction (XRD), Field emission scanning electron microscopy (SEM) and Fourier-Transform Infrared Spectroscopy (FTIR). Mixtures with a low thermal conductivity of 0.056 W (m K)−1 and a porosity of 79 vol% achieved a compressive strength of only 0.02 MPa. In comparison, mixtures with a thermal conductivity of 0.087 W (m K)−1 and a porosity of 58 vol% achieved a compressive strength of 0.45 MPa.
Spektroskopische Qualifizierung und Quantifizierung von Hyaluronsäure in Nahrungsergänzungsmitteln
(2023)
Background: the potency of drugs that interfere with glucose metabolism, i.e., glucose transporters (GLUT) and nicotinamide phosphoribosyltransferase (NAMPT) was analyzed in neuroendocrine tumor (NET, BON-1, and QPG-1 cells) and small cell lung cancer (SCLC, GLC-2, and GLC-36 cells) tumor cell lines. (2) Methods: the proliferation and survival rate of tumor cells was significantly affected by the GLUT-inhibitors fasentin and WZB1127, as well as by the NAMPT inhibitors GMX1778 and STF-31. (3) Results: none of the NET cell lines that were treated with NAMPT inhibitors could be rescued with nicotinic acid (usage of the Preiss–Handler salvage pathway), although NAPRT expression could be detected in two NET cell lines. We finally analyzed the specificity of GMX1778 and STF-31 in NET cells in glucose uptake experiments. As previously shown for STF-31 in a panel NET-excluding tumor cell lines, both drugs specifically inhibited glucose uptake at higher (50 μM), but not at lower (5 μM) concentrations. (4) Conclusions: our data suggest that GLUT and especially NAMPT inhibitors are potential candidates for the treatment of NET tumors.
A biodegradable blend of PBAT—poly(butylene adipate-co-terephthalate)—and PLA—poly(lactic acid)—for blown film extrusion was modified with four multi-functional chain extending cross-linkers (CECL). The anisotropic morphology introduced during film blowing affects the degradation processes. Given that two CECL increased the melt flow rate (MFR) of tris(2,4-di-tert-butylphenyl)phosphite (V1) and 1,3-phenylenebisoxazoline (V2) and the other two reduced it (aromatic polycarbodiimide (V3) and poly(4,4-dicyclohexylmethanecarbodiimide) (V4)), their compost (bio-)disintegration behavior was investigated. It was significantly altered with respect to the unmodified reference blend (REF). The disintegration behavior at 30 and 60 °C was investigated by determining changes in mass, Young’s moduli, tensile strengths, elongations at break and thermal properties. In order to quantify the disintegration behavior, the hole areas of blown films were evaluated after compost storage at 60 °C to calculate the kinetics of the time dependent degrees of disintegration. The kinetic model of disintegration provides two parameters: initiation time and disintegration time. They quantify the effects of the CECL on the disintegration behavior of the PBAT/PLA compound. Differential scanning calorimetry (DSC) revealed a pronounced annealing effect during storage in compost at 30 °C, as well as the occurrence of an additional step-like increase in the heat flow at 75 °C after storage at 60 °C. The disintegration consists of processes which affect amorphous and crystalline phase of PBAT in different manner that cannot be understood by a hydrolytic chain degradation only. Furthermore, gel permeation chromatography (GPC) revealed molecular degradation only at 60 °C for the REF and V1 after 7 days of compost storage. The observed losses of mass and cross-sectional area seem to be attributed more to mechanical decay than to molecular degradation for the given compost storage times.
ENaC channels
(2023)
The non-filarial and non-communicable disease podoconiosis affects around 4 million people and is characterized by severe leg lymphedema accompanied with painful intermittent acute inflammatory episodes, called acute dermatolymphangioadenitis (ADLA) attacks. Risk factors have been associated with the disease but the mechanisms of pathophysiology remain uncertain. Lymphedema can lead to skin lesions, which can serve as entry points for bacteria that may cause ADLA attacks leading to progression of the lymphedema. However, the microbiome of the skin of affected legs from podoconiosis individuals remains unclear. Thus, we analysed the skin microbiome of podoconiosis legs using next generation sequencing. We revealed a positive correlation between increasing lymphedema severity and non-commensal anaerobic bacteria, especially Anaerococcus provencensis, as well as a negative correlation with the presence of Corynebacterium, a constituent of normal skin flora. Disease symptoms were generally linked to higher microbial diversity and richness, which deviated from the normal composition of the skin. These findings show an association of distinct bacterial taxa with lymphedema stages, highlighting the important role of bacteria for the pathogenesis of podoconiosis and might enable a selection of better treatment regimens to manage ADLA attacks and disease progression.
The cystic fibrosis transmembrane conductance regulator (CFTR) anion channel and the epithelial Na+ channel (ENaC) play essential roles in transepithelial ion and fluid transport in numerous epithelial tissues. Inhibitors of both channels have been important tools for defining their physiological role in vitro. However, two commonly used CFTR inhibitors, CFTRinh-172 and GlyH-101, also inhibit non-CFTR anion channels, indicating they are not CFTR specific. However, the potential off-target effects of these inhibitors on epithelial cation channels has to date not been addressed. Here, we show that both CFTR blockers, at concentrations routinely employed by many researchers, caused a significant inhibition of store-operated calcium entry (SOCE) that was time-dependent, poorly reversible and independent of CFTR. Patch clamp experiments showed that both CFTRinh-172 and GlyH-101 caused a significant block of Orai1-mediated whole cell currents, establishing that they likely reduce SOCE via modulation of this Ca2+ release-activated Ca2+ (CRAC) channel. In addition to off-target effects on calcium channels, both inhibitors significantly reduced human αβγ-ENaC-mediated currents after heterologous expression in Xenopus oocytes, but had differential effects on δβγ-ENaC function. Molecular docking identified two putative binding sites in the extracellular domain of ENaC for both CFTR blockers. Together, our results indicate that caution is needed when using these two CFTR inhibitors to dissect the role of CFTR, and potentially ENaC, in physiological processes.
ESKAPEE Pathogen Biofilm Control on Surfaces with Probiotic Lactobacillaceae and Bacillus species
(2023)
Combatting the rapidly growing threat of antimicrobial resistance and reducing prevalence and transmission of ESKAPEE pathogens in healthcare settings requires innovative strategies, one of which is displacing these pathogens using beneficial microorganisms. Our review comprehensively examines the evidence of probiotic bacteria displacing ESKAPEE pathogens, with a focus on inanimate surfaces. A systematic search was conducted using the PubMed and Web of Science databases on 21 December 2021, and 143 studies were identified examining the effects of Lactobacillaceae and Bacillus spp. cells and products on the growth, colonization, and survival of ESKAPEE pathogens. While the diversity of study methods limits evidence analysis, results presented by narrative synthesis demonstrate that several species have the potential as cells or their products or supernatants to displace nosocomial infection-causing organisms in a variety of in vitro and in vivo settings. Our review aims to aid the development of new promising approaches to control pathogen biofilms in medical settings by informing researchers and policymakers about the potential of probiotics to combat nosocomial infections. More targeted studies are needed to assess safety and efficacy of different probiotic formulations, followed by large-scale studies to assess utility in infection control and medical practice.
Isovaleric acidemia (IVA), due to isovaleryl-CoA dehydrogenase (IVD) deficiency, results in the accumulation of isovaleryl-CoA, isovaleric acid and secondary metabolites. The increase in these metabolites decreases mitochondrial energy production and increases oxidative stress. This contributes to the neuropathological features of IVA. A general assumption in the literature exists that glycine N-acyltransferase (GLYAT) plays a role in alleviating the symptoms experienced by IVA patients through the formation of N-isovalerylglycine. GLYAT forms part of the phase II glycine conjugation pathway in the liver and detoxifies excess acyl-CoA’s namely benzoyl-CoA. However, very few studies support GLYAT as the enzyme that conjugates isovaleryl-CoA to glycine. Furthermore, GLYATL1, a paralogue of GLYAT, conjugates phenylacetyl-CoA to glutamine. Therefore, GLYATL1 might also be a candidate for the formation of N-isovalerylglycine. Based on the findings from the literature review, we proposed that GLYAT or GLYATL1 can form N-isovalerylglycine in IVA patients. To test this hypothesis, we performed an in-silico analysis to determine which enzyme is more likely to conjugate isovaleryl-CoA with glycine using AutoDock Vina. Thereafter, we performed in vitro validation using purified enzyme preparations. The in-silico and in vitro findings suggested that both enzymes could form N-isovaleryglycine albeit at lower affinities than their preferred substrates. Furthermore, an increase in glycine concentration does not result in an increase in N-isovalerylglycine formation. The results from the critical literature appraisal, in-silico, and in vitro validation, suggest the importance of further investigating the reaction kinetics and binding behaviors between these substrates and enzymes in understanding the pathophysiology of IVA.
Indoor spaces exhibit microbial compositions that are distinctly dissimilar from one another and from outdoor spaces. Unique in this regard, and a topic that has only recently come into focus, is the microbiome of hospitals. While the benefits of knowing exactly which microorganisms propagate how and where in hospitals are undoubtedly beneficial for preventing hospital-acquired infections, there are, to date, no standardized procedures on how to best study the hospital microbiome. Our study aimed to investigate the microbiome of hospital sanitary facilities, outlining the extent to which hospital microbiome analyses differ according to sample-preparation protocol. For this purpose, fifty samples were collected from two separate hospitals—from three wards and one hospital laboratory—using two different storage media from which DNA was extracted using two different extraction kits and sequenced with two different primer pairs (V1–V2 and V3–V4). There were no observable differences between the sample-preservation media, small differences in detected taxa between the DNA extraction kits (mainly concerning Propionibacteriaceae), and large differences in detected taxa between the two primer pairs V1–V2 and V3–V4. This analysis also showed that microbial occurrences and compositions can vary greatly from toilets to sinks to showers and across wards and hospitals. In surgical wards, patient toilets appeared to be characterized by lower species richness and diversity than staff toilets. Which sampling sites are the best for which assessments should be analyzed in more depth. The fact that the sample processing methods we investigated (apart from the choice of primers) seem to have changed the results only slightly suggests that comparing hospital microbiome studies is a realistic option. The observed differences in species richness and diversity between patient and staff toilets should be further investigated, as these, if confirmed, could be a result of excreted antimicrobials.
In memoriam Willy Lehnert
(2023)
Pitfalls of using sequence databases for heterologous expression studies - a technical review
(2023)
Synthesis of DNA fragments based on gene sequences available in public resources has become an efficient and affordable method that gradually replaced traditional cloning efforts such as PCR cloning from cDNA. However, database entries based on genome sequencing results are prone to errors which can lead to false sequence information and, ultimately, errors in functional characterization of proteins such as ion channels and transporters in heterologous expression systems. We have identified five common problems that repeatedly appear in public resources: 1) Not every gene has yet been annotated; 2) Not all gene annotations are necessarily correct; 3) Transcripts may contain automated corrections; 4) There are mismatches between gene, mRNA, and protein sequences; and 5) Splicing patterns often lack experimental validation. This technical review highlights and provides a strategy to bypass these issues in order to avoid critical mistakes that could impact future studies of any gene/protein of interest in heterologous expression systems. Abstract figure legend Projects involving heterologous gene expression are often characterised by similar steps. Initially, database research (A) is necessary to retrieve information of full of partial sequences of a gene of interest. A multitude of genome assemblies are annotated and deposited in public databases or that are available for refined search options using individual sequence information. The search results need to be scrutinised and compared to already available information (B). Once the sequence has been determined, DNA synthesis (C) by PCR or commercial synthesis are necessary for further cloning procedures (D). Eventually, the DNA needs to be transfected (E) and expressed in, e.g., eukaryotic cells (F). Finally, the expression of the gene of interest needs to be documented and its function analysed (G). This article is protected by copyright. All rights reserved.
Microbiome analyses are essential for understanding microorganism composition and diversity, but interpretation is often challenging due to biological and technical variables. DNA extraction is a critical step that can significantly bias results, particularly in samples containing a high abundance of challenging-to-lyse microorganisms. Taking into consideration the distinctive microenvironments observed in different bodily locations, our study sought to assess the extent of bias introduced by suboptimal bead-beating during DNA extraction across diverse clinical sample types. The question was whether complex targeted extraction methods are always necessary for reliable taxonomic abundance estimation through amplicon sequencing or if simpler alternatives are effective for some sample types. Hence, for four different clinical sample types (stool, cervical swab, skin swab, and hospital surface swab samples), we compared the results achieved from extracting targeted manual protocols routinely used in our research lab for each sample type with automated protocols specifically not designed for that purpose. Unsurprisingly, we found that for the stool samples, manual extraction protocols with vigorous bead-beating were necessary in order to avoid erroneous taxa proportions on all investigated taxonomic levels and, in particular, false under- or overrepresentation of important genera such as Blautia, Faecalibacterium, and Parabacteroides. However, interestingly, we found that the skin and cervical swab samples had similar results with all tested protocols. Our results suggest that the level of practical automation largely depends on the expected microenvironment, with skin and cervical swabs being much easier to process than stool samples. Prudent consideration is necessary when extending the conclusions of this study to applications beyond rough estimations of taxonomic abundance.
The epithelial sodium channel (ENaC) is a key regulator of sodium homeostasis that contributes to blood pressure control. ENaC open probability is adjusted by extracellular sodium ions, a mechanism referred to as sodium self-inhibition (SSI). With a growing number of identified ENaC gene variants associated with hypertension, there is an increasing demand for medium- to high-throughput assays allowing the detection of alterations in ENaC activity and SSI. We evaluated a commercially available automated two-electrode voltage-clamp (TEVC) system that records transmembrane currents of ENaC-expressing Xenopus oocytes in 96-well microtiter plates. We employed guinea pig, human and Xenopus laevis ENaC orthologs that display specific magnitudes of SSI. While demonstrating some limitations over traditional TEVC systems with customized perfusion chambers, the automated TEVC system was able to detect the established SSI characteristics of the employed ENaC orthologs. We were able to confirm a reduced SSI in a gene variant, leading to C479R substitution in the human α-ENaC subunit that has been reported in Liddle syndrome. In conclusion, automated TEVC in Xenopus oocytes can detect SSI of ENaC orthologs and variants associated with hypertension. For precise mechanistic and kinetic analyses of SSI, optimization for faster solution exchange rates is recommended.
Host-derived succinate accumulates in the airways during bacterial infection. Here, we show that luminal succinate activates murine tracheal brush (tuft) cells through a signaling cascade involving the succinate receptor 1 (SUCNR1), phospholipase Cβ2, and the cation channel transient receptor potential channel subfamily M member 5 (TRPM5). Stimulated brush cells then trigger a long-range Ca2+ wave spreading radially over the tracheal epithelium through a sequential signaling process. First, brush cells release acetylcholine, which excites nearby cells via muscarinic acetylcholine receptors. From there, the Ca2+ wave propagates through gap junction signaling, reaching also distant ciliated and secretory cells. These effector cells translate activation into enhanced ciliary activity and Cl- secretion, which are synergistic in boosting mucociliary clearance, the major innate defense mechanism of the airways. Our data establish tracheal brush cells as a central hub in triggering a global epithelial defense program in response to a danger-associated metabolite.
Microorganisms not only contribute to the spoilage of food but can also cause illnesses through consumption. Consumer concerns and doubts about the shelf life of the products and the resulting enormous amounts of food waste have led to a demand for a rapid, robust, and non-destructive method for the detection of microorganisms, especially in the food sector. Therefore, a rapid and simple sampling method for the Raman- and infrared (IR)-microspectroscopic study of microorganisms associated with spoilage processes was developed. For subsequent evaluation pre-processing routines, as well as chemometric models for classification of spoilage microorganisms were developed. The microbiological samples are taken using a disinfectable sampling stamp and measured by microspectroscopy without the usual pre-treatments such as purification separation, washing, and centrifugation. The resulting complex multivariate data sets were pre-processed, reduced by principal component analysis, and classified by discriminant analysis. Classification of independent unlabeled test data showed that microorganisms could be classified at genus, species, and strain levels with an accuracy of 96.5 % (Raman) and 94.5 % (IR), respectively, despite large biological differences and novel sampling strategies. As bacteria are exposed to constantly changing conditions and their adaptation mechanisms may make them inaccessible to conventional measurement methods, the methods and models developed were investigated for their suitability for microorganisms exposed to stress. Compared to normal growth conditions, spectral changes in lipids, polysaccharides, nucleic acids, and proteins were observed in microorganisms exposed to stress. Models were developed to discriminate microorganisms, independent of the involvement of various stress factors and storage times. Classification of the investigated bacteria yielded accuracies of 97.6 % (Raman) and 96.6 % (IR), respectively, and a robust and meaningful model was developed to discriminate different microorganisms at the genus, species, and strain levels. The obtained results are very promising and show that the methods and models developed for the discrimination of microorganisms as well as the investigation of stress factors on microorganisms by means of Raman- and IR-microspectroscopy have the potential to be used, for example, in the food sector for the rapid determination of surface contamination.
PURPOSE
Cervical cancer (CC) is caused by a persistent high-risk human papillomavirus (hrHPV) infection. The cervico-vaginal microbiome may influence the development of (pre)cancer lesions. Aim of the study was (i) to evaluate the new CC screening program in Germany for the detection of high-grade CC precursor lesions, and (ii) to elucidate the role of the cervico-vaginal microbiome and its potential impact on cervical dysplasia.
METHODS
The microbiome of 310 patients referred to colposcopy was determined by amplicon sequencing and correlated with clinicopathological parameters.
RESULTS
Most patients were referred for colposcopy due to a positive hrHPV result in two consecutive years combined with a normal PAP smear. In 2.1% of these cases, a CIN III lesion was detected. There was a significant positive association between the PAP stage and Lactobacillus vaginalis colonization and between the severity of CC precursor lesions and Ureaplasma parvum.
CONCLUSION
In our cohort, the new cervical cancer screening program resulted in a low rate of additional CIN III detected. It is questionable whether these cases were only identified earlier with additional HPV testing before the appearance of cytological abnormalities, or the new screening program will truly increase the detection rate of CIN III in the long run. Colonization with U. parvum was associated with histological dysplastic lesions. Whether targeted therapy of this pathogen or optimization of the microbiome prevents dysplasia remains speculative.
Several species of (poly)saccharides and organic acids can be found often simultaneously in various biological matrices, e.g., fruits, plant materials, and biological fluids. The analysis of such matrices sometimes represents a challenging task. Using Aloe vera (A. vera) plant materials as an example, the performance of several spectroscopic methods (80 MHz benchtop NMR, NIR, ATR-FTIR and UV-Vis) for the simultaneous analysis of quality parameters of this plant material was compared. The determined parameters include (poly)saccharides such as aloverose, fructose and glucose as well as organic acids (malic, lactic, citric, isocitric, acetic, fumaric, benzoic and sorbic acids). 500 MHz NMR and high-performance liquid chromatography (HPLC) were used as the reference methods.
UV-VIS data can be used only for identification of added preservatives (benzoic and sorbic acids) and drying agent (maltodextrin) and semiquantitative analysis of malic acid. NIR and MIR spectroscopies combined with multivariate regression can deliver more informative overview of A. vera extracts being able to additionally quantify glucose, aloverose, citric, isocitric, malic, lactic acids and fructose. Low-field NMR measurements can be used for the quantification of aloverose, glucose, malic, lactic, acetic, and benzoic acids. The benchtop NMR method was successfully validated in terms of robustness, stability, precision, reproducibility and limit of detection (LOD) and quantification (LOQ), respectively.
All spectroscopic techniques are useful for the screening of (poly)saccharides and organic acids in plant extracts and should be applied according to its availability as well as information and confidence required for the specific analytical goal. Benchtop NMR spectroscopy seems to be the most feasible solution for quality control of A. vera products.
Monitoring the content of dissolved ozone in purified water is often mandatory to ensure the appropriate levels of disinfection and sanitization. However, quantification bears challenges as colorimetric assays require laborious off-line analysis, while commercially available instruments for electrochemical process analysis are expensive and often lack the possibility for miniaturization and discretionary installation. In this study, potentiometric ionic polymer metal composite (IPMC) sensors for the determination of dissolved ozone in ultrapure water (UPW) systems are presented. Commercially available polymer electrolyte membranes are treated via an impregnation-reduction method to obtain nanostructured platinum layers. By applying 25 different synthesis conditions, layer thicknesses of 2.2 to 12.6 µm are obtained. Supporting radiographic analyses indicate that the platinum concentration of the impregnation solution has the highest influence on the obtained metal loading. The sensor response behavior is explained by a Langmuir pseudo-isotherm model and allows the quantification of dissolved ozone to trace levels of less than 10 µg L−1. Additional statistical evaluations show that the expected Pt loading and radiographic blackening levels can be predicted with high accuracy and significance (R2adj. > 0.90, p < 10−10) solely from given synthesis conditions.
This research studies in detail four different assays, namely DPPH (2,2-diphenyl-1-picrylhydrazyl), ABTS (2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)), FRAP (ferric ion reducing antioxidant potential) and FC (Folin-Ciocalteu), to determine the antioxidant capacity of standard substances as well as 50 organosolv lignins, and two kraft lignins. The coefficient of variation was determined for each method and was lowest for ABTS and highest for DPPH. The best correlation was found for FRAP and FC, which both rely on a single electron transfer mechanism. A good correlation between ABTS, FRAP and FC, respectively, could be observed, even though ABTS relies on a more complex reaction mechanism. The DPPH assay merely correlates with the others, implying that it reflects different antioxidative attributes due to a different reaction mechanism. Lignins obtained from paulownia and silphium have been investigated for the first time regarding their antioxidant capacity. Paulownia lignin is in the same range as beech wood lignin, while silphium lignin resembles wheat straw lignin. Miscanthus lignin is an exception from the grass lignins and possesses a significantly higher antioxidant capacity. All lignins possess a good antioxidant capacity and thus are promising candidates for various applications, e. g. as additives in food packaging or for biomedical purposes.
Im Rahmen dieser Arbeit wurden zunächst neuartige ionische Agarosederivate synthetisiert und anschließend umfassend charakterisiert. Anionische Agarosesulfate mit einer regioselektiven Derivatisierung in Position G6 wurden durch homogene Umsetzung in ionischer Flüssigkeit erhalten. Kationische Agarosecarbamate mit einstellbarem Funktionalisierungsgrad waren durch einen zweistufigen Syntheseansatz zugänglich. Hierzu wurden zunächst Agarosephenylcarbonate in einer homogenen Synthese hergestellt, im Anschluss folgte eine Aminolyse zu den gewünschten funktionalen Agarosederivaten. Die ionischen Agarosederivate waren bereits bei geringen Funktionalisierungsgraden vollständig löslich in Wasser. Damit war es möglich, Alginatmikrokapseln polyelektrolytisch zu beschichten und diese als Träger für eine kontrollierte Wirkstofffreisetzung zu verwenden. Ebenfalls konnten Kompositgele aus Agarose, Hydroxyapatit und Agarosederivaten hergestellt und charakterisiert werden. Im zweiten Teil wurden sowohl die Kompositträgermaterialien als auch die Alginatmikrokapseln mit vier verschiedenen Modellwirkstoffen (ATP, Suramin, Methylenblau und A740003) beladen und die Wirkstofffreisetzung über einen Zeitraum von zwei Wochen untersucht. Für die ionischen Modellwirkstoffe erwiesen sich Kompositträgermaterialien mit ionischem Agarosederivat, die beschichteten Mikrokapseln sowie die Kombination aus Komposit und Kapseln als effektiv, um die Freisetzung auf bis zu 40% zu verlangsamen. Für die schlecht wasserlösliche Substanz A740003, ein Rezeptorligand für die osteogene Differenzierung von Stammzellen, wurde eine stark verzögerte Freisetzung aus Polyelektrolytemikrokapseln festgestellt. Mithilfe von literaturbekannten und neu entwickelten Anpassungsmodellen gelang es, die Diffusion als Hauptmechanismus der Wirkstofffreisetzung zu identifizieren und die Freisetzungskurven mathematisch akkurat zu beschreiben und daraus Rückschlüsse über die einzelnen Phasen der Freisetzung zu ziehen.
Transdermal therapeutic systems (TTS) represent an up-to-day medication applied to human skin, which consists of a drug-containing pressure-sensitive adhesive (PSA) and a flexible backing layer. The development of a reliable TTS requires precise knowledge of the viscoelastic tack behavior of PSA in terms of adhesion and detaching. Tailoring of a PSA can be achieved by altering the resin content or modifying the chemical properties of the macromolecules. In this study, three different resin content of two silicone-based PSA – non-amine compatible, and less tack, amine-compatible – were investigated with the help of recently developed RheoTack method to characterize the retraction speed dependent tack behavior for various geometries of the testing rods. The obtained force-retraction displacement-curves clearly depict the effect of the chemical structure as well as the resin content. Decreasing the resin content shifts the start of fibril fracture to larger deformations states and significantly enhances the stretchability of the fibrils. To compare various rod geometries precisely, the force-retraction displacement curves were normalized to account for effective contact areas. The flat and spherical rods led to completely different failure and tack behaviors. Furthermore, the adhesion formation between TTS with flexible backing layers and rods during the dwell phase happens in a different manner compared to rigid plates, in particular for flat rods, where maximum compression stresses occur at the edges and not uniformly over the cross-section. Thus, the approach to follow ASTM D2949 has to be reconsidered for tests of these materials.
Lignin ist ein aromatisches Biopolymer, das in den Zellwänden von Pflanzen vorkommt. Es ist hauptsächlich aus drei sogenannten Monolignolen (p-Hydroxyphenyl (H), Guajakol (G) und Syringol (S)) aufgebaut, die über verschiedene Bindungen miteinander verknüpft sein können, und enthält eine Vielzahl an funktionellen Gruppen. Interessant für die Verwendung von Lignin sind dabei insbesondere die vielen phenolischen Hydroxygruppen, die als Ausgangsstoff bei der Synthese neuer Produkte dienen können, daneben aber auch für seine antioxidativen Eigenschaften verantwortlich sind. Da Struktur und Eigenschaften von vielen Faktoren wie Biomasse und Aufschlussprozess abhängen, ist eine detaillierte Charakterisierung der Lignine nötig, um Struktur-Eigenschafts-Beziehungen aufzuklären und so einen Schritt näher an eine mögliche stoffliche Nutzung zu kommen. Mit dieser Arbeit soll der Einfluss der Biomasse inklusive der verwendeten Partikelgröße sowie des Organosolv-Aufschlussprozesses auf die Monomerzusammensetzung, das Molekulargewicht und die Antioxidanz der isolierten Lignine untersucht werden.
Als Rohstoffe zur Ligningewinnung dienen die drei mehrjährigen lignocellulosereichen Low-Input-Pflanzen Miscanthus x giganteus, Silphium perfoliatum und Paulownia tomentosa, die momentan hauptsächlich zur Energiegewinnung genutzt werden. Im Rahmen der Bioökonomiestrategie der Europäischen Union soll der Schwerpunkt zukünftiger Bioraffinerien jedoch auf eine ganzheitliche Nutzung von Biomassen gelegt und so auch die stoffliche Nutzung fokussiert werden. Zusätzlich zu diesen drei Pflanzen werden auch Organosolv-Lignine aus den in der Literatur bereits gut beschriebenen Biomassen Weizenstroh und Buchenholz isoliert, und zwei Nadelholz-Kraft-Lignine als Vergleich herangezogen. Die Ergebnisse zeigen, dass die Art der Biomasse hauptsächlich die Monomerzusammensetzung beeinflusst: Gräser bestehen aus allen drei Monolignolen, Laubhölzer mehrheitlich aus S- und G-Einheiten, während Nadelhölzer nur aus G-Einheiten aufgebaut sind. Die Holzlignine besitzen zudem höhere Molekulargewichte sowie bessere antioxidative Eigenschaften als die Gras- und Krautlignine. Mit der feineren Vermahlung der Biomasse kann die Monomerzusammensetzung beeinflusst werden: der Einsatz kleinerer Partikelgrößen führt zu Ligninen mit einem höheren Gehalt an H-Einheiten, sowohl für Miscanthus als auch für Paulownia. Außerdem kann bei Paulownia die Ausbeute gesteigert und eine Zunahme des Molekulargewichtes beobachtet werden, wenn die kleinste Siebfraktion für den Organosolv-Aufschluss verwendet wird. Einen größeren Einfluss als der Mahlgrad der Biomasse haben die Autohydrolyse sowie der Organosolv-Aufschlussprozess selbst. Die Monomerzusammensetzung ändert sich aufgrund derselben Biomasse zwar kaum, die Bindungstypen zwischen den Monolignolen dagegen schon. Mit höherer Prozessstärke (Zeit, Temperatur, Ethanol-Konzentration) werden Etherbindungen gespalten, was den Anteil an phenolischen Hydroxygruppen und somit die Antioxidanz erhöht. Neben dieser Depolymerisation werden partiell auch Rekondensationsreaktionen beobachtet.
Die erzielten Ergebnisse liefern einen Beitrag zum Verständnis des Zusammenhangs zwischen Ligninquelle und -gewinnung mit der daraus resultierenden Ligninstruktur und Antioxidanz und bieten damit eine Grundlage für den Wandel von der energetischen hin zu einer nachhaltigen stofflichen Nutzung dieses nachwachsenden Biopolymers. Gerade über die Wahl der Aufschlussparameter können Struktur und Antioxidanz gezielt beeinflusst werden, was in zukünftigen Studien weiter fokussiert werden sollte.
Forensic DNA profiles are established by multiplex PCR amplification of a set of highly variable short tandem repeat (STR) loci followed by capillary electrophoresis (CE) as a means to assign alleles to PCR products of differential length. Recently, CE analysis of STR amplicons has been supplemented by high-throughput next generation sequencing (NGS) techniques that are able to detect isoalleles bearing sequence polymorphisms and allow for an improved analysis of degraded DNA. Several such assays have been commercialised and validated for forensic applications. However, these systems are cost-effective only when applied to high numbers of samples. We report here an alternative, cost-efficient shallow-sequence output NGS assay called maSTR assay that, in conjunction with a dedicated bioinformatics pipeline called SNiPSTR, can be implemented with standard NGS instrumentation. In a back-to-back comparison with a CE-based, commercial forensic STR kit, we find that for samples with low DNA content, with mixed DNA from different individuals, or containing PCR inhibitors, the maSTR assay performs equally well, and with degraded DNA is superior to CE-based analysis. Thus, the maSTR assay is a simple, robust and cost-efficient NGS-based STR typing method applicable for human identification in forensic and biomedical contexts.
The development of whole-genome amplification (WGA) techniques has opened up new avenues for genetic analysis and genome research, in particular by facilitating the genome-wide analysis of few or even single copies of genomic DNA, such as from single cells (prokaryotic or eukaryotic) or virions. Using WGA, the few copies of genomic DNA obtained from such entities are unspecifically amplified using PCR or PCR-related processes in order to obtain higher DNA quantities that can then be successfully analysed further.
This work presents an open source database with suitable retention parameters for prediction and simulation of GC separations and gives a short introduction to three common retention models. Useful computer simulations play an important role to save resources and time in method development in GC. Thermodynamic retention parameters for the ABC model and the K-centric model are determined by isothermal measurements. This standardized procedure of measurements and calculations, presented in this work, have a useful benefit for all chromatographers, analytical chemists, and method developers because it can be used in their own laboratories to simplify the method development. The main benefits as simulations of temperature-programed GC separations are demonstrated and compared to measurements. The observed deviations of predicted retention times are in most cases less than 1%. The database includes more than 900 entries with a large range of compounds such as VOCs, PAHs, FAMEs, PCBs, or allergenic fragrances over 20 different GC columns.
There & Back again: Developing a tool for testing of antimicrobial surfaces for space habitat design
(2023)
When optimizing the process parameters of the acidic ethanolic organosolv process, the aim is usually to maximize the delignification and/or lignin purity. However, process parameters such as temperature, time, ethanol and catalyst concentration, respectively, can also be used to vary the structural properties of the obtained organosolv lignin, including the molecular weight and the ratio of aliphatic versus phenolic hydroxyl groups, among others. This review particularly focuses on these influencing factors and establishes a trend analysis between the variation of the process parameters and the effect on lignin structure. Especially when larger data sets are available, as for process temperature and time, correlations between the distribution of depolymerization and condensation reactions are found, which allow direct conclusions on the proportion of lignin's structural features, independent of the diversity of the biomass used. The newfound insights gained from this review can be used to tailor organosolv lignins isolated for a specific application.
Analytical Chemistry I
(2023)
This workbook takes you through the successful work Harris, Textbook of Quantitative Analysis and is designed primarily for self-study. In five parts, the lecture content of analytical chemistry is summarized and explained using selected examples. Basic concepts of analytical chemistry are presented as well as the principle and various techniques of dimensional analysis and chromatography. UV/VIS, infrared and Raman spectroscopy are used to explain the investigation of molecularly present compounds, and selected techniques of atomic spectroscopy conclude the introduction to the fundamentals of analysis. The textbook's essential sections and illustrations are repeatedly referred to, which facilitates independent learning of the fundamentals of analytical chemistry.
The French–Italian Concordia Research Station, situated on the Antarctic Polar Plateau at an elevation of 3233 m above sea level, offers a unique opportunity to study the presence and variation of microbes introduced by abiotic or biotic vectors and, consequently, appraise the amplitude of human impact in such a pristine environment. This research built upon a previous work, which explored microbial diversity in the surface snow surrounding the Concordia Research Station. While that study successfully characterized the bacterial assemblage, detecting fungal diversity was hampered by the low DNA content. To address this knowledge gap, in the present study, we optimized the sampling by increasing ice/snow collected to leverage the final DNA yield. The V4 variable region of the 16S rDNA and Internal Transcribed Spacer (ITS1) rDNA was used to evaluate bacterial and fungal diversity. From the sequencing, we obtained 3,352,661 and 4,433,595 reads clustered in 930 and 3182 amplicon sequence variants (ASVs) for fungi and bacteria, respectively. Amplicon sequencing revealed a predominance of Basidiomycota (49%) and Ascomycota (42%) in the fungal component; Bacteroidota (65.8%) is the main representative among the bacterial phyla. Basidiomycetes are almost exclusively represented by yeast-like fungi. Our findings provide the first comprehensive overview of both fungal and bacterial diversity in the Antarctic Polar Plateau’s surface snow/ice near Concordia Station and to identify seasonality as the main driver of microbial diversity; we also detected the most sensitive microorganisms to these factors, which could serve as indicators of human impact in this pristine environment and aid in planetary protection for future exploration missions.
Electrical signal transmission in power electronic devices takes place through high-purity aluminum bonding wires. Cyclic mechanical and thermal stresses during operation lead to fatigue loads, resulting in premature failure of the wires, which cannot be reliably predicted. The following work presents two fatigue lifetime models calibrated and validated based on experimental fatigue results of an aluminum bonding wire and subsequently transferred and applied to other wire types. The lifetime modeling of Wöhler curves for different load ratios shows good but limited applicability for the linear model. The model can only be applied above 10,000 cycles and within the investigated load range of R = 0.1 to R = 0.7. The nonlinear model shows very good agreement between model prediction and experimental results over the entire investigated cycle range. Furthermore, the predicted Smith diagram is not only consistent in the investigated load range but also in the extrapolated load range from R = −1.0 to R = 0.8. A transfer of both model approaches to other wire types by using their tensile strengths can be implemented as well, although the nonlinear model is more suitable since it covers the entire load and cycle range.
Cyanobacteria are gaining considerable interest as a method of supporting the long-term presence of humans on the Moon and settlements on Mars due to their ability to produce oxygen and their potential as bio-factories for space biotechnology/synthetic biology and other applications. Since many unknowns remain in our knowledge to bridge the gap and move cyanobacterial bioprocesses from Earth to space, we investigated cell division resumption on the rehydration of dried Chroococcidiopsis sp. CCMEE 029 accumulated DNA damage while exposed to space vacuum, Mars-like conditions, and Fe-ion radiation. Upon rehydration, the monitoring of the ftsZ gene showed that cell division was arrested until DNA damage was repaired, which took 48 h under laboratory conditions. During the recovery, a progressive DNA repair lasting 48 h of rehydration was revealed by PCR-stop assay. This was followed by overexpression of the ftsZ gene, ranging from 7.5- to 9-fold compared to the non-hydrated samples. Knowing the time required for DNA repair and cell division resumption is mandatory for deep-space experiments that are designed to unravel the effects of reduced/microgravity on this process. It is also necessary to meet mission requirements for dried-sample implementation and real-time monitoring upon recovery. Future experiments as part of the lunar exploration mission Artemis and the lunar gateway station will undoubtedly help to move cyanobacterial bioprocesses beyond low Earth orbit. From an astrobiological perspective, these experiments will further our understanding of microbial responses to deep-space conditions.
The Concordia Research Station provides a unique location for preparatory activities for future human journey to Mars, to explore microbial diversity at subzero temperatures, and monitor the dissemination of human-associated microorganisms within the pristine surrounding environment. Amplicon sequencing was leveraged to investigate the microbial diversity of surface snow samples collected monthly over a two-year period, at three distances from the Station (10, 500, and 1000 m). Even when the extracted total DNA was below the detection limit, 16S rRNA gene sequencing was successfully performed on all samples, while 18S rRNA was amplified on 19 samples out of 51. No significant relationships were observed between microbial diversity and seasonality (summer or winter) or distance from the Concordia base. This suggested that if present, the anthropogenic impact should have been below the detectable limit. While harboring low microbial diversity, the surface snow samples were characterized by heterogeneous microbiomes. Ultimately, our study corroborated the use of DNA sequencing-based techniques for revealing microbial presence in remote and hostile environments, with implications for Planetary Protection during space missions and for life-detection in astrobiology relevant targets.
The white ground crater by the Phiale Painter (450–440 BC) exhibited in the “Pietro Griffo” Archaeological Museum in Agrigento (Italy) depicts two scenes from Perseus myth. The vase is of utmost importance to archaeologists because the figures are drawn on a white background with remarkable daintiness and attention to detail. Notwithstanding the white ground ceramics being well documented from an archaeological and historical point of view, doubts concerning the compositions of pigments and binders and the production technique are still unsolved. This kind of vase is a valuable rarity, the use of which is documented in elitist funeral rituals. The study aims to investigate the constituent materials and the execution technique of this magnificent crater. The investigation was carried out using non-destructive and non-invasive techniques in situ. Portable X-ray fluorescence and Fourier-transform total reflection infrared spectroscopy complemented the use of visible and ultraviolet light photography to get an overview and specific information on the vase. The XRF data were used to produce false colour maps showing the location of the various elements detected, using the program SmART_scan. The use of gypsum as the material for the white ground is an important result that deserves to be further investigated in similar vases.
Silicon carbide and graphene possess extraordinary chemical and physical properties. Here, these different systems are linked and the changes in structural and dynamic properties are investigated. For the simulations performed a classical molecular dynamic (MD) approach was used. In this approach, a graphene layer (N = 240 atoms) was grafted at different distances on top of a 6H-SiC structure (N = 2400 atoms) and onto a 3C-SiC structure (N = 1728 atoms). The distances between the graphene and the 6H are 1.0, 1.3 and 1.5 Å and the distances between the graphene layer and the 3C-SiC are 2.0, 2.3, and 2.5 Å. Each system has been equilibrated at room temperature until no further relaxation was observed. The 6H-SiC structure in combination with graphene proves to be more stable compared to the combination with 3C-SiC. This can be seen well in the determined energies. Pair distribution functions were influenced slightly by the graphene layer due to steric and energetic changes. This becomes clear from the small shifts of the C-C distances. Interactions as well as bonds between graphene and SiC lead to the fact that small shoulders of the high-frequency SiC-peaks are visible in the spectra and at the same time the high-frequency peaks of graphene are completely absent.
Modeling of Creep Behavior of Particulate Composites with Focus on Interfacial Adhesion Effect
(2022)
Evaluation of creep compliance of particulate composites using empirical models always provides parameters depending on initial stress and material composition. The effort spent to connect model parameters with physical properties has not resulted in success yet. Further, during the creep, delamination between matrix and filler may occur depending on time and initial stress, reducing an interface adhesion and load transfer to filler particles. In this paper, the creep compliance curves of glass beads reinforced poly(butylene terephthalate) composites were fitted with Burgers and Findley models providing different sets of time-dependent model parameters for each initial stress. Despite the finding that the Findley model performs well in a primary creep, the Burgers model is more suitable if secondary creep comes into play; they allow only for a qualitative prediction of creep behavior because the interface adhesion and its time dependency is an implicit, hidden parameter. As Young’s modulus is a parameter of these models (and the majority of other creep models), it was selected to be introduced as a filler content-dependent parameter with the help of the cube in cube elementary volume approach of Paul. The analysis led to the time-dependent creep compliance that depends only on the time-dependent creep of the matrix and the normalized particle distance (or the filler volume content), and it allowed accounting for the adhesion effect. Comparison with the experimental data confirmed that the elementary volume-based creep compliance function can be used to predict the realistic creep behavior of particulate composites.
Telogene Einzelhaare sind häufig vorkommende Spurentypen an Tatorten. Derzeit werden sie zumeist von der STR-Typisierung ausgeschlossen, weil ihre STR-Profile aufgrund geringer DNA-Mengen und starker DNA-Degradierung in vielen Fällen unvollständig und schwierig zu interpretieren sind. In der vorliegenden Arbeit wurde eine systematische Vorgehensweise angewandt, um Korrelationen zwischen der DNA-Menge und DNA-Degradierung zu dem Erfolg der STR-Typisierung aufzuweisen und darauf basierend den Typisierungs-Erfolg von DNA aus Haaren vorhersagen zu können.
Zu diesem Zweck wurde ein human- (RiboD) und ein canin-spezifischer (RiboDog) qPCR-basierter Assay zur Messung der DNA-Menge und Bewertung der DNA-Integrität mittels eines Degradierungswerts (D-Wert) entwickelt. Aufgrund der Lage der genutzten Primer, welche auf ubiquitär vorkommende ribosomale DNA-Sequenzen abzielen, ist das Funktionsprinzip schnell und kostengünstig auf unterschiedliche Spezies anzuwenden. Die Funktionsweise der Assays wurde mittels seriell degradierter DNA bestätigt und der humane Assay wurde im Vergleich zum kommerziellen Quantifiler? Trio DNA Quantification Kit validiert. Schließlich wurde mit den Assays an DNA aus telogenen und katagenen Einzelhaaren von Menschen und Hunden der Zusammenhang zwischen DNA-Menge und DNA-Integrität zu der Vollständigkeit der STR-Allele (Allel Recovery) von DNA-Profilen untersucht, die mittels kapillarelektrophoretischer (CE) STR-Kits erhaltenen wurde. Es zeigte sich, dass bei humanen Einzelhaaren die Allel-Recovery sowohl von der DNA-Menge als auch der DNA-Integrität abhängt. Dagegen war die DNA-Degradierung bei einzelnen Hundehaaren durchweg geringer und die Allel-Recovery hing allein von der extrahierten DNA-Menge ab.
Um die STR-Analytik degradierter humaner DNA-Proben weiter zu verbessern, wurde ein neuartiger NGS-basierter Assay (maSTR, Mini-Amplikon-STR) etabliert, der die 16 forensischen STR-Loci des European Standard Sets und Amelogenin als sehr kurze Amplikons (76-296 bp) parallel amplifiziert. Mit intakter DNA generierte der maSTR-Assay im Mengenbereich von 200 pg eingesetzter DNA reproduzierbare, vollständige Profile ohne Allelic Drop-ins. Bei niedrigeren DNA-Mengen traten vereinzelt Allelic Drop-ins auf, wobei unter Verwendung von mindestens 43 pg DNA vollständige Profile erhalten wurden.
Die kombinierte Strategie aus RiboD-Messungen der DNA-Menge und -Integrität und daraus resultierendem STR-Typisierungserfolg des maSTR-Assays wurde an degradierter DNA validiert. Anschließend wurde die Strategie auf DNA aus telogenen und katagenen Einzelhaaren angewandt und mit den Ergebnissen des CE-basierten PowerPlex? ESX 17-Kits verglichen, das dasselbe STR-Marker-Set analysiert. Dabei zeigte sich, dass der Erfolg der STR-Typisierung beider STR-Assays sowohl von der optimalen Menge der Template-DNA als auch von der DNA-Integrität abhängt. Mit dem maSTR-Assay wurden vollständige Profile mit ungefähr 50 pg Input-DNA für leicht degradierte DNA aus Einzelhaaren nachgewiesen, sowie mit ungefähr 500 pg stark degradierter DNA. Aufgrund der geringen DNA-Mengen von telogenen Einzelhaaren schwankte die Reproduzierbarkeit der maSTR-Ergebnisse, war jedoch stets dem PowerPlex? ESX 17-Kit in Bezug auf die Allel-Recovery überlegen.
Ein Vergleich mit zwei, hinsichtlich der Längenverteilung der Amplikons komplementären CE-basierten STR-Kits (PowerPlex? ESX 17 und ESI 17 Fast), sowie mit einem kommerziellen NGS-Kit (ForenSeq? DNA Signature Prep) ergab, dass nicht die Technik der NGS, sondern die Kürze der Amplikons der wichtigste Faktor zur Typisierung degradierter DNA ist. Der maSTR-Assay wies in allen Vergleichen mit den genutzten kommerziellen Kits jedoch eine höhere Anzahl an Allelic Drop-ins auf. Diese traten umso häufiger auf, je geringer die verwendete DNA-Menge und je stärker degradiert diese war.
Weil Profile mit Allelic Drop-ins Mischprofilen entsprechen, wurden die per maSTR-Assay generierten STR-Profile mit Verfahren zur Interpretation von Mischspuren untersucht. Bei der Composite-Interpretation werden alle vorkommenden Allele von Replikaten gezählt, bei der Consensus-Interpretation lediglich die reproduzierbaren Allele. Dabei stellte sich heraus, dass im Fall von wenigen Allelic Drop-ins (PowerPlex? ESX 17-generierte Profile) die Composite-Interpretation und bei Allelic Drop-in-haltigen Profilen (maSTR-generierte Profile) die Consensus-Interpretation am besten geeignet ist.
Schließlich wurde mittels der GenoProof Mixture 3-Software untersucht, inwieweit semi- und vollständig kontinuierliche probabilistische Verfahren bei der biostatistischen Bewertung der DNA-Profile aus Einzelhaaren geeignet sind. Dabei zeigte sich, dass der maSTR-Assay aufgrund der hohen Anzahl an Allelic Drop-ins den CE-basierten Methoden nur in Fällen von DNA leicht überlegen ist, die in ausreichender Menge und gering degradiert vorliegt. In diesem Bereich gelingt die Zuordnung des Profils aus Haaren zum Referenzprofil jedoch ebenfalls mittels CE-basierten Methoden.
Aus allen Ergebnissen wurde eine Empfehlung für die Handhabung von DNA aus ausgefallenen Einzelhaaren abgeleitet, die auf dem DNA-Degradierungsgrad in Kombination mit der DNA-Menge basiert. Die vorliegende Arbeit schafft somit eine Grundlage, um ausgefallene Einzelhaare in der Routine-Arbeit von kriminaltechnischen Ermittlungen nutzbar zu machen, sowie gegebenenfalls auf andere Spurentypen mit degradierter DNA geringer Menge anzuwenden. Dadurch könnte die Nutzbarkeit solcher Spurentypen für die forensische Kriminalistik erhöht werden, insbesondere wenn die standardmäßig verwendeten CE-basierten Methoden versagen. Perspektivisch ist die Technik der NGS aufgrund der großen Multiplexierbarkeit uniformer, kurzer Marker generell der CE-basierten Technik bei der Typisierung degradierter DNA überlegen.
The human enzymes GLYAT (glycine N-acyltransferase), GLYATL1 (glutamine N-phenylacetyltransferase) and GLYATL2 (glycine N-acyltransferase-like protein 2) are not only important in the detoxification of xenobiotics via the human liver, but are also involved in the elimination of acyl residues that accumulate in the form of their coenzyme A (coA) esters in some rare inborn errors of metabolism. This concerns, for example, disorders in the degradation of branched-chain amino acids, such as isovaleric acidemia or propionic acidemia. In addition, they also assist in the elimination of ammonium, which is produced during the transamination of amino acids and accumulates in urea cycle defects. Sequence variants of the enzymes have also been investigated, which may provide evidence of impaired enzyme activities, from which therapy adjustments can potentially be derived. A modified Escherichia coli strain was chosen for the overexpression and partial biochemical characterization of the enzymes, which may allow solubility and proper folding. Since post-translational protein modifications are very limited in bacteria, we also attempted to overexpress the enzymes in HEK293 cells (human-derived). In addition to characterization via immunoblots and activity assays, intracellular localization of the enzymes was also performed using GFP coupling and confocal laser scanning microscopy in transfected HEK293 cells. The GLYATL2 enzyme may have tasks beyond detoxification and metabolic defects and the preliminary molecular biology work has been performed as part of this project - the enzyme activity determinations were outsourced to a co-supervised bachelor thesis. The enzyme activity determinations with purified recombinant human enzyme from Escherichia coli provided a threefold higher activity of the sequence variant p.(Asn156Ser) for GLYAT, which should be considered as the probably authentic wild type of the enzyme. In addition, a reduced activity of the GLYAT variant p.(Gln61Leu), which is very common in South Africa, was shown, which could be of particular importance in the treatment of isovaleric acidemia, which is also common in South Africa. Intracellularly, GLYAT and GLYATL1 could be localized mitochondrially. As the analyses have shown, sequence variations of GLYAT and GLYATL1 influence their enzyme activity. As an example, the GLYAT variant p.(Gln61Leu) is frequently found in South Africa. In the case of reduced GLYAT activity, patients could be increasingly treated with L-carnitine in the sense of an individualized therapy, since the conjugation of the toxic isovaleryl-coA with glycine is restricted by the GLYAT sequence variation. Activity-reducing variants identified in this project are of particular interest, as they may influence the treatment of certain metabolic defects.
SLC6A14 (ATB0,+) is unique among SLC proteins in its ability to transport 18 of the 20 proteinogenic (dipolar and cationic) amino acids and naturally occurring and synthetic analogues (including anti-viral prodrugs and nitric oxide synthase (NOS) inhibitors). SLC6A14 mediates amino acid uptake in multiple cell types where increased expression is associated with pathophysiological conditions including some cancers. Here, we investigated how a key position within the core LeuT-fold structure of SLC6A14 influences substrate specificity. Homology modelling and sequence analysis identified the transmembrane domain 3 residue V128 as equivalent to a position known to influence substrate specificity in distantly related SLC36 and SLC38 amino acid transporters. SLC6A14, with and without V128 mutations, was heterologously expressed and function determined by radiotracer solute uptake and electrophysiological measurement of transporter-associated current. Substituting the amino acid residue occupying the SLC6A14 128 position modified the binding pocket environment and selectively disrupted transport of cationic (but not dipolar) amino acids and related NOS inhibitors. By understanding the molecular basis of amino acid transporter substrate specificity we can improve knowledge of how this multi-functional transporter can be targeted and how the LeuT-fold facilitates such diversity in function among the SLC6 family and other SLC amino acid transporters.
While many proteins are known clients of heat shock protein 90 (Hsp90), it is unclear whether the transcription factor, thyroid hormone receptor beta (TRb), interacts with Hsp90 to control hormonal perception and signaling. Higher Hsp90 expression in mouse fibroblasts was elicited by the addition of triiodothyronine (T3). T3 bound to Hsp90 and enhanced adenosine triphosphate (ATP) binding of Hsp90 due to a specific binding site for T3, as identified by molecular docking experiments. The binding of TRb to Hsp90 was prevented by T3 or by the thyroid mimetic sobetirome. Purified recombinant TRb trapped Hsp90 from cell lysate or purified Hsp90 in pull-down experiments. The affinity of Hsp90 for TRb was 124 nM. Furthermore, T3 induced the release of bound TRb from Hsp90, which was shown by streptavidin-conjugated quantum dot (SAv-QD) masking assay. The data indicate that the T3 interaction with TRb and Hsp90 may be an amplifier of the cellular stress response by blocking Hsp90 activity.
Microarray-based experiments revealed that thyroid hormone triiodothyronine (T3) enhanced the binding of Cy5-labeled ATP on heat shock protein 90 (Hsp90). By molecular docking experiments with T3 on Hsp90, we identified a T3 binding site (TBS) near the ATP binding site on Hsp90. A synthetic peptide encoding HHHHHHRIKEIVKKHSQFIGYPITLFVEKE derived from the TBS on Hsp90 showed, in MST experiments, the binding of T3 at an EC50 of 50 μM. The binding motif can influence the activity of Hsp90 by hindering ATP accessibility or the release of ADP.
Cathepsin K (CatK) is a target for the treatment of osteoporosis, arthritis, and bone metastasis. Peptidomimetics with a cyanohydrazide warhead represent a new class of highly potent CatK inhibitors; however, their binding mechanism is unknown. We investigated two model cyanohydrazide inhibitors with differently positioned warheads: an azadipeptide nitrile Gü1303 and a 3-cyano-3-aza-β-amino acid Gü2602. Crystal structures of their covalent complexes were determined with mature CatK as well as a zymogen-like activation intermediate of CatK. Binding mode analysis, together with quantum chemical calculations, revealed that the extraordinary picomolar potency of Gü2602 is entropically favoured by its conformational flexibility at the nonprimed-primed subsites boundary. Furthermore, we demonstrated by live cell imaging that cyanohydrazides effectively target mature CatK in osteosarcoma cells. Cyanohydrazides also suppressed the maturation of CatK by inhibiting the autoactivation of the CatK zymogen. Our results provide structural insights for the rational design of cyanohydrazide inhibitors of CatK as potential drugs.
There is an unmet need for the development and validation of biomarkers and surrogate endpoints for clinical trials in propionic acidemia (PA) and methylmalonic acidemia (MMA). This review examines the pathophysiology and clinical consequences of PA and MMA that could form the basis for potential biomarkers and surrogate endpoints. Changes in primary metabolites such as methylcitric acid (MCA), MCA:citric acid ratio, oxidation of 13C-propionate (exhaled 13CO2), and propionylcarnitine (C3) have demonstrated clinical relevance in patients with PA or MMA. Methylmalonic acid, another primary metabolite, is a potential biomarker, but only in patients with MMA. Other potential biomarkers in patients with either PA and MMA include secondary metabolites, such as ammonium, or the mitochondrial disease marker, fibroblast growth factor 21. Additional research is needed to validate these biomarkers as surrogate endpoints, and to determine whether other metabolites or markers of organ damage could also be useful biomarkers for clinical trials of investigational drug treatments in patients with PA or MMA. This review examines the evidence supporting a variety of possible biomarkers for drug development in propionic and methylmalonic acidemias.
The epithelial sodium channel (ENaC) is a heterotrimeric ion channel that plays a key role in sodium and water homeostasis in tetrapod vertebrates. In the aldosterone-sensitive distal nephron, hormonally controlled ENaC expression matches dietary sodium intake to its excretion. Furthermore, ENaC mediates sodium absorption across the epithelia of the colon, sweat ducts, reproductive tract, and lung. ENaC is a constitutively active ion channel and its expression, membrane abundance, and open probability (PO) are controlled by multiple intracellular and extracellular mediators and mechanisms [9]. Aberrant ENaC regulation is associated with severe human diseases, including hypertension, cystic fibrosis, pulmonary edema, pseudohypoaldosteronism type 1, and nephrotic syndrome [9].
Mobiles Laser-Schneidsystem zur Unterstützung der USBV-Entschärfung und Beweissicherung (mobiLaS)
(2022)
P30 - Das Elektrospinnen von halbleitenden Zinndioxidfasern für die Detektion von Wasserstoff
(2022)
Das Ziel dieser Arbeit ist die Entwicklung von dünnen keramischen Fasern als halbleitendes Sensormaterial zum Nachweis von Wasserstoff, möglichst bei Zimmertemperatur. Die elektrische Leitfähigkeit halbleitender Metalloxide ändert sich durch die Einwirkung von oxidierenden und reduzierenden Gasen auf die Oberfläche des Metalloxids. Dieser Effekt kann zur Messung der Gaskonzentration genutzt werden. Die Reaktion von Zinn(IV)-oxid mit Wasserstoff basiert auf der Reduktion des Zinn(IV)-oxids zum Zinn, wobei die Elektronen des Zinn(IV)-oxids im metallischen Zinn verbleiben und dort im nicht gebundenen Zustand zu einer Leitfähigkeitserhöhung beitragen. Die Reaktion des Wasserstoffes kann sowohl mit den Sauerstoffatomen des Oxids als auch mit adsorbierten Sauerstoffatomen an der Oxidoberfläche stattfinden.[ 6] Da die Reaktionen an der Oberfläche des Oxids stattfinden, sollten Sensoren mit einer großen Oberfläche im Vergleich zu metalloxidischen Bulkmaterialien eine höhere Empfindlichkeit aufweisen. [3] Die Verwendung von Fasern anstelle von Dünn- oder Dickschichten führt dabei zu einer besseren Sensitivität gegenüber Gasen.
Process-induced changes in the morphology of biodegradable polybutylene adipate terephthalate (PBAT) and polylactic acid (PLA) blends modified with various multifunctional chainextending cross-linkers (CECLs) are presented. The morphology of unmodified and modified films produced with blown film extrusion is examined in an extrusion direction (ED) and a transverse direction (TD). While FTIR analysis showed only small peak shifts indicating that the CECLs modify the molecular weight of the PBAT/PLA blend, SEM investigations of the fracture surfaces of blown extrusion films revealed their significant effect on the morphology formed during the processing. Due to the combined shear and elongation deformation during blown film extrusion, rather spherical PLA islands were partly transformed into long fibrils, which tended to decay to chains of elliptical islands if cooled slowly. The CECL introduction into the blend changed the thickness of the PLA fibrils, modified the interface adhesion, and altered the deformation behavior of the PBAT matrix from brittle to ductile. The results proved that CECLs react selectively with PBAT, PLA, and their interface. Furthermore, the reactions of CECLs with PBAT/PLA induced by the processing depended on the deformation directions (ED and TD), thus resulting in further non-uniformities of blown extrusion films.
Background: Cancer heterogeneity poses a serious challenge concerning the toxicity and adverse effects of therapeutic inhibitors, especially when it comes to combinatorial therapies that involve multiple targeted inhibitors. In particular, in non-small cell lung cancer (NSCLC), a number of studies have reported synergistic effects of drug combinations in the preclinical models, while they were only partially successful in the clinical setup, suggesting those alternative clinical strategies (with genetic background and immune response) should be considered. Herein, we investigated the antitumor effect
of cytokine-induced killer (CIK) cells in combination with ALK and PD-1 inhibitors in vitro on genetically variable NSCLC cell lines.
Methods: We co-cultured the three genetically different NSCLC cell lines NCI-H2228 (EML4-ALK), A549 (KRAS mutation), and HCC-78 (ROS1 rearrangement) with and without nivolumab (PD-1 inhibitor) and crizotinib (ALK inhibitor). Additionally, we profiled the variability of surface expression multiple immune checkpoints, the concentration of absolute dead cells, intracellular granzyme B on CIK cells using flow cytometry as well as RT-qPCR. ELISA and Western blot were performed to verify the activation of CIK cells.
Results: Our analysis showed that (a) nivolumab significantly weakened PD-1 surface expression on CIK cells without impacting other immune checkpoints or PD-1 mRNA expression, (b) this combination strategy showed an effective response on cell viability, IFN-g production, and intracellular release of granzyme B in CD3+ CD56+ CIK cells, but solely in NCI-H2228, (c) the intrinsic expression of Fas ligand (FasL) as a T-cell activation marker in CIK cells was upregulated by this additive effect, and (d) nivolumab induced Foxp3 expression in CD4+CD25+ subpopulation of CIK cells significantly increased. Taken together, we could show that CIK cells in combination with crizotinib and nivolumab can enhance the anti-tumor immune response through FasL activation, leading to increased IFN-g and granzyme B, but only in NCI-H2228 cells with EML4-ALK rearrangement. Therefore, we hypothesize that CIK therapy may be a potential alternative in NSCLC patients harboring EML4-ALK rearrangement, in addition, we support the idea that combination therapies offer significant potential when they are optimized on a patient-by-patient basis.
Cytokine-induced killer cells (CIK) in combination with dendritic cells (DCs) have shown favorable outcomes in renal cell carcinoma (RCC), yet some patients exhibit recurrence or no response to this therapy. In a broader perspective, enhancing the antitumor response of DC-CIK cells may help to address this issue. Considering this, herein, we investigated the effect of anti-CD40 and anti-CTLA-4 antibodies on the antitumor response of DC-CIK cells against RCC cell lines. Our analysis showed that, a) anti-CD40 antibody (G28.5) increased the CD3+CD56+ effector cells of CIK cells by promoting the maturation and activation of DCs, b) G28.5 also increased CTLA-4 expression in CIK cells via DCs, but the increase could be hindered by the CTLA-4 inhibitor (ipilimumab), c) adding ipilimumab was also able to significantly increase the proportion of CD3+CD56+ cells in DC-CIK cells, d) anti-CD40 antibodies predominated over anti-CTLA-4 antibodies for cytotoxicity, apoptotic effect and IFN-g secretion of DC-CIK cells against RCC cells, e) after ipilimumab treatment, the population of Tregs in CIK cells remained unaffected, but ipilimumab combined with G28.5 significantly reduced the expression of CD28 in CIK cells. Taken together, we suggest that the agonistic anti-CD40 antibody rather than CTLA-4 inhibitor may improve the antitumor response of DC-CIK cells, particularly in RCC. In addition, we pointed towards the yet to be known contribution of CD28 in the crosstalk between anti-CTLA-4 and CIK cells.
Introduction of Matrix-Filler Adhesion to Modelling of Elastic Moduli of Particulate Composites
(2022)
Cube in cube elementary volume (EV) concept serves to predict a filler-content dependent Young´s moduli of particle filled composites using moduli of a matrix EM and a filler EF. Paul and Ishai-Cohen derived formulas for composites moduli considering different load transfer boundaries in the EV assuming a complete filler-matrix adhesion. In this paper it is confirmed that their models represent the upper and lower bounds, respectively, with the respect to the experimental data. However, in vast majority of composites a filler-matrix adhesion is not complete. Therefore, an adhesion factor kadh gaining values between 0 and 1 was introduced into Paul´s model to consider the reduced adhesion as the reduction of the filler-matrix contact area for glass beads filled in polar and unpolar thermoplastic matrices as well as elastomer. The evaluation of these composite systems provides reasonable adhesion coefficients of PA66 > PBT > PP > PE-LD >> BR. It was also found that stiffening only occurs if kadh exceeds the minimum value adhesion of root square of E(M) divided by E(F). The determined kadh correspond to scanning electron microscopy observations of the composites fracture surfaces. Additionally, finite element analysis of the cubic and hexagonal arrangements of the EV show that the stress distributions are different, but they affect the calculated moduli only for the filler volume contents exceeding 20 %. The introduction of the filler-matrix adhesion provides more reliable predictions of Young´s moduli of particulate composites.
The utilization of simulation procedures is gaining increasing attention in the product development of extrusion blow molded parts. However, some simulation steps, like the simulation of shrinkage and warpage, are still associated with uncertainties. The reason for this is on the one hand a lack of standardized interfaces for the transfer of simulation data between different simulation tools, and on the other hand the complex time-, temperature- and process-dependent material behavior of the used semi crystalline polymers. Using a new vendor neutral interface standard for the data transfer, the shrinkage analysis of a simple blow molded part is investigated and compared to experimental data. A linear viscoelastic material model in combination with an orthotropic process- and temperature-dependent thermal expansion coefficient is used for the shrinkage prediction. A good agreement is observed. Finally, critical parameters in the simulation models that strongly influence the shrinkage analysis are identified by a sensitivity study.
Jet engines of airplanes are designed such that in some components damage occurs and accumulates in service without being critical up to a certain level of damage. Since maintenance, repair, and component exchange are very cost-intensive, it is necessary to predict efficiently the component lifetime with high accuracy. A former developed lifetime model, based on interpolated results of aerodynamic and structural mechanics simulations, uses material parameters estimated from literature values of standard creep experiments. For improved accuracy, an experimental procedure is developed for the characterization of the short-time creep behavior, which is relevant for the operation of turbine blades of jet engines. To consider microstructural influences resulting from the manufacturing of thin-walled single crystal turbine blades, small-scale specimens from used turbine blades are extracted and tested in short- and medium-time creep experiments. Based on experimental results and literature values, a creep model, which describes the fracture behavior for a wide range of creep loads, is calibrated and is now used for the lifetime prediction of turbine blades under real loading conditions.
Cathepsin K (CatK) is a target for the treatment of osteoporosis, arthritis, and bone metastasis. Peptidomimetics with a cyanohydrazide warhead represent a new class of highly potent CatK inhibitors; however, their binding mechanism is unknown. We investigated two model cyanohydrazide inhibitors with differently positioned warheads: an azadipeptide nitrile Gü1303 and a 3-cyano-3-aza-β-amino acid Gü2602. Crystal structures of their covalent complexes were determined with mature CatK as well as a zymogen-like activation intermediate of CatK. Binding mode analysis, together with quantum chemical calculations, revealed that the extraordinary picomolar potency of Gü2602 is entropically favoured by its conformational flexibility at the nonprimed-primed subsites boundary. Furthermore, we demonstrated by live cell imaging that cyanohydrazides effectively target mature CatK in osteosarcoma cells. Cyanohydrazides also suppressed the maturation of CatK by inhibiting the autoactivation of the CatK zymogen. Our results provide structural insights for the rational design of cyanohydrazide inhibitors of CatK as potential drugs.
Discrimination of Stressed and Non-Stressed Food-Related Bacteria Using Raman-Microspectroscopy
(2022)
As the identification of microorganisms becomes more significant in industry, so does the utilization of microspectroscopy and the development of effective chemometric models for data analysis and classification. Since only microorganisms cultivated under laboratory conditions can be identified, but they are exposed to a variety of stress factors, such as temperature differences, there is a demand for a method that can take these stress factors and the associated reactions of the bacteria into account. Therefore, bacterial stress reactions to lifetime conditions (regular treatment, 25 °C, HCl, 2-propanol, NaOH) and sampling conditions (cold sampling, desiccation, heat drying) were induced to explore the effects on Raman spectra in order to improve the chemometric models. As a result, in this study nine food-relevant bacteria were exposed to seven stress conditions in addition to routine cultivation as a control. Spectral alterations in lipids, polysaccharides, nucleic acids, and proteins were observed when compared to normal growth circumstances without stresses. Regardless of the involvement of several stress factors and storage times, a model for differentiating the analyzed microorganisms from genus down to strain level was developed. Classification of the independent training dataset at genus and species level for Escherichia coli and at strain level for the other food relevant microorganisms showed a classification rate of 97.6%.
The Chemotype of Chromanones as a Privileged Scaffold for Multineurotarget Anti-Alzheimer Agents
(2022)
Nanomedicine strategies were first adapted and successfully translated to clinical application for diseases, such as cancer and diabetes. These strategies would no doubt benefit unmet diseases needs as in the case of leishmaniasis. The latter causes skin sores in the cutaneous form and affects internal organs in the visceral form. Treatment of cutaneous leishmaniasis (CL) aims at accelerating wound healing, reducing scarring and cosmetic morbidity, preventing parasite transmission and relapse. Unfortunately, available treatments show only suboptimal effectiveness and none of them were designed specifically for this disease condition. Tissue regeneration using nano-based devices coupled with drug delivery are currently being used in clinic to address diabetic wounds. Thus, in this review, we analyse the current treatment options and attempt to critically analyse the use of nanomedicine-based strategies to address CL wounds in view of achieving scarless wound healing, targeting secondary bacterial infection and lowering drug toxicity.
Approximately 45% of global greenhouse gas emissions are caused by the construction and use of buildings. Thermal insulation of buildings in the current context of climate change is a well-known strategy to improve the energy efficiency of buildings. The development of renewable insulation material can overcome the drawbacks of widely used insulation systems based on polystyrene or mineral wool. This study analyzes the sustainability and thermal conductivity of new insulation materials made of Miscanthus x giganteus fibers, foaming agents, and alkali-activated fly ash binder. Life cycle assessments (LCA) are necessary to perform benchmarking of environmental impacts of new formulations of geopolymer-based insulation materials. The global warming potential (GWP) of the product is primarily determined by the main binder component sodium silicate. Sodium silicate's CO2 emissions depend on local production, transportation, and energy consumption. The results, which have been published during recent years, vary in a wide range from 0.3 kg to 3.3 kg CO2-eq. kg-1. The overall GWP of the insulation system based on Miscanthus fibers, with properties according to current thermal insulation regulations, reaches up to 95% savings of CO2 emissions compared to conventional systems. Carbon neutrality can be achieved through formulations containing raw materials with carbon dioxide emissions and renewable materials with negative GWP, thus balancing CO2 emissions.
Thermo-chemical conversion of cucumber peel waste for biobased energy and chemical production
(2022)
Therapeutic Treatments for Osteoporosis-Which Combination of Pills Is the Best among the Bad?
(2022)
Osteoporosis is a chronical, systemic skeletal disorder characterized by an increase in bone resorption, which leads to reduced bone density. The reduction in bone mineral density and therefore low bone mass results in an increased risk of fractures. Osteoporosis is caused by an imbalance in the normally strictly regulated bone homeostasis. This imbalance is caused by overactive bone-resorbing osteoclasts, while bone-synthesizing osteoblasts do not compensate for this. In this review, the mechanism is presented, underlined by in vitro and animal models to investigate this imbalance as well as the current status of clinical trials. Furthermore, new therapeutic strategies for osteoporosis are presented, such as anabolic treatments and catabolic treatments and treatments using biomaterials and biomolecules. Another focus is on new combination therapies with multiple drugs which are currently considered more beneficial for the treatment of osteoporosis than monotherapies. Taken together, this review starts with an overview and ends with the newest approaches for osteoporosis therapies and a future perspective not presented so far.
Characterization methods of pressure sensitive adhesives (PSA) originate from technical bonding and do not cover relevant data for the development and quality assurance of medical applications, where PSA with flexible backing layers are adopted to human skin. In this study, a new method called RheoTack is developed to determine (mechanically and optically) an adhesion and detaching behavior of flexible and transparent PSA based patches. Transdermal therapeutic systems (TTS) consisting of silicone-based PSAs on a flexible and transparent backing layer were tested on a rotational rheometer with an 8 mm plate as a probe rod at retraction speeds of 0.01, 0.1, and 1 mm/s with respect to their adhesion and detaching behavior in terms of force-retraction displacement curves. The curves consist of a compression phase to affirm wetting; a tensile deformation phase intercepting stretching, cavity, and fibril formation; and a failure phase with detaching. Their analysis provides values for stiffness, force, and displacement of the beginning of fibril formation, force and displacement of the beginning of a failure due to fibril breakage and detaching, as well as corresponding activation energies. All these parameters exhibit the pronounced dependency on the retraction speed. The force-retraction displacement curves together with the simultaneous video recordings of the TTS deformation from three different angles (three cameras) provide deeper insight into the deformation processes and allow for interpreting the properties’ characteristics for PSA applications.
Operating an ozone-evolving PEM electrolyser in tap water: A case study of water and ion transport
(2022)
While PEM water electrolysis could be a favourable technique for in situ sanitization with ozone, its application is mainly limited to the use of ultrapure water to achieve a sufficient long-time stability. As additional charge carriers influence the occurring transport phenomena, we investigated the impact of different feed water qualities on the performance of a PEM tap water electrolyser for ozone evolution. The permeation of water and the four most abundant cations (Na+, K+, Ca2+, Mg2+) is characterised during stand-by and powered operation at different charge densities to quantify underlying transport mechanisms. Water transport is shown to linearly increase with the applied current (95 ± 2 mmol A−1 h−1) and occurs decoupled from ion permeation. A limitation of ion permeation is given by the transfer of ions in water to the anode/PEM interface. The unstabilized operation of a PEM electrolyser in tap water leads to a pH gradient which promotes the formation of magnesium and calcium carbonates and hydroxides on the cathode surface. The introduction of a novel auxiliary cathode in the anolytic compartment has shown to suppress ion permeation by close to 20%.
Composite nanoparticles (NPs) consisting of lignin and different polysaccharide (PS) derivatives were prepared. In this synergistic approach, the PS derivative acts as biocompatible matrix that forms spherical NPs while lignin is a functional compound with therapeutic potential (e.g., antioxidative, antimicrobial, antiviral). Organosolv lignin and three different PS derivatives (cellulose acetate/CA, cellulose acetate phthalate/CAPh, xylan phenyl carbonate/XPC) were used in this study. Nanocomposites with particle sizes in the range of about 200–550 nm containing both types of biopolymers are accessible by dialysis of organic PS/lignin solutions against water. In particular, XPC and CAPh, which both contain aromatic substituents, were found to be suitable for incorporation of lignin within the PS nanomatrix. The present work paves the way for future studies in which the pharmaceutical potential and biocompatibility of composite NPs of lignin and PS derivatives with tailored properties are investigated.
Modern PCR-based analytical techniques have reached sensitivity levels that allow for obtaining complete forensic DNA profiles from even tiny traces containing genomic DNA amounts as small as 125 pg. Yet these techniques have reached their limits when it comes to the analysis of traces such as fingerprints or single cells. One suggestion to overcome these limits has been the usage of whole genome amplification (WGA) methods. These methods aim at increasing the copy number of genomic DNA and by this means generate more template DNA for subsequent analyses. Their application in forensic contexts has so far remained mostly an academic exercise, and results have not shown significant improvements and even have raised additional analytical problems. Until very recently, based on these disappointments, the forensic application of WGA seems to have largely been abandoned. In the meantime, however, novel improved methods are pointing towards a perspective for WGA in specific forensic applications. This review article tries to summarize current knowledge about WGA in forensics and suggests the forensic analysis of single-donor bioparticles and of single cells as promising applications.