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- ItemSurface-Enhanced Raman Spectroscopy to Characterize Different Fractions of Extracellular Vesicles from Control and Prostate Cancer Patients(Basel : MDPI, 2021) Osei, Eric Boateng; Paniushkina, Liliia; Wilhelm, Konrad; Popp, Jürgen; Nazarenko, Irina; Krafft, ChristophExtracellular vesicles (EVs) are membrane-enclosed structures ranging in size from about 60 to 800 nm that are released by the cells into the extracellular space; they have attracted interest as easily available biomarkers for cancer diagnostics. In this study, EVs from plasma of control and prostate cancer patients were fractionated by differential centrifugation at 5000× g, 12,000× g and 120,000× g. The remaining supernatants were purified by ultrafiltration to produce EV-depleted free-circulating (fc) fractions. Spontaneous Raman and surface-enhanced Raman spectroscopy (SERS) at 785 nm excitation using silver nanoparticles (AgNPs) were employed as label-free techniques to collect fingerprint spectra and identify the fractions that best discriminate between control and cancer patients. SERS spectra from 10 µL droplets showed an enhanced Raman signature of EV-enriched fractions that were much more intense for cancer patients than controls. The Raman spectra of dehydrated pellets of EV-enriched fractions without AgNPs were dominated by spectral contributions of proteins and showed variations in S-S stretch, tryptophan and protein secondary structure bands between control and cancer fractions. We conclude that the AgNPs-mediated SERS effect strongly enhances Raman bands in EV-enriched fractions, and the fractions, EV12 and EV120 provide the best separation of cancer and control patients by Raman and SERS spectra.
- ItemFiber-based SORS-SERDS system and chemometrics for the diagnostics and therapy monitoring of psoriasis inflammatory disease in vivo(Washington, DC : Optica, 2021-1-28) Schleusener, Johannes; Guo, Shuxia; Darvin, Maxim E.; Thiede, Gisela; Chernavskaia, Olga; Knorr, Florian; Lademann, Jürgen; Popp, Jürgen; Bocklitz, Thomas W.Psoriasis is considered a widespread dermatological disease that can strongly affect the quality of life. Currently, the treatment is continued until the skin surface appears clinically healed. However, lesions appearing normal may contain modifications in deeper layers. To terminate the treatment too early can highly increase the risk of relapses. Therefore, techniques are needed for a better knowledge of the treatment process, especially to detect the lesion modifications in deeper layers. In this study, we developed a fiber-based SORS-SERDS system in combination with machine learning algorithms to non-invasively determine the treatment efficiency of psoriasis. The system was designed to acquire Raman spectra from three different depths into the skin, which provide rich information about the skin modifications in deeper layers. This way, it is expected to prevent the occurrence of relapses in case of a too short treatment. The method was verified with a study of 24 patients upon their two visits: the data is acquired at the beginning of a standard treatment (visit 1) and four months afterwards (visit 2). A mean sensitivity of ≥85% was achieved to distinguish psoriasis from normal skin at visit 1. At visit 2, where the patients were healed according to the clinical appearance, the mean sensitivity was ≈65%.
- ItemShape-Memory Metallopolymers Based on Two Orthogonal Metal–Ligand Interactions(Weinheim : Wiley-VCH, 2021) Meurer, Josefine; Hniopek, Julian; Bätz, Thomas; Zechel, Stefan; Enke, Marcel; Vitz, Jürgen; Schmitt, Michael; Popp, Jürgen; Hager, Martin D.; Schubert, Ulrich S.A new shape-memory polymer is presented, in which both the stable phase as well as the switching unit consist of two different metal complexes. Suitable metal ions, which simultaneously form labile complexes with histidine and stable ones with terpyridine ligands, are identified via isothermal titration calorimetry (ITC) measurements. Different copolymers are synthesized, which contain butyl methacrylate as the main monomer and the metal-binding ligands in the side chains. Zn(TFMS)2 and NiCl2 are utilized for the dual crosslinking, resulting in the formation of metallopolymer networks. The switching temperature can simply be tuned by changing the composition as well as by the choice of the metal ion. Strain fixity rates (about 99%) and very high strain recovery rates (up to 95%) are achieved and the mechanism is revealed using different techniques such as Raman spectroscopy. © 2021 The Authors. Advanced Materials published by Wiley-VCH GmbH
- ItemStealth Effect of Short Polyoxazolines in Graft Copolymers: Minor Changes of Backbone End Group Determine Liver Cell-Type Specificity(Washington, DC : ACS Publications, 2021) Muljajew, Irina; Huschke, Sophie; Ramoji, Anuradha; Cseresnyés, Zoltán; Hoeppener, Stephanie; Nischang, Ivo; Foo, Wanling; Popp, Jürgen; Figge, Marc Thilo; Weber, Christine; Bauer, Michael; Schubert, Ulrich S.; Press, Adrian T.Dye-loaded micelles of 10 nm diameter formed from amphiphilic graft copolymers composed of a hydrophobic poly(methyl methacrylate) backbone and hydrophilic poly(2-ethyl-2-oxazoline) side chains with a degree of polymerization of 15 were investigated concerning their cellular interaction and uptake in vitro as well as their interaction with local and circulating cells of the reticuloendothelial system in the liver by intravital microscopy. Despite the high molar mass of the individual macromolecules (Mn ≈ 20 kg mol-1), backbone end group modification by attachment of a hydrophilic anionic fluorescent probe strongly affected the in vivo performance. To understand these effects, the end group was additionally modified by the attachment of four methacrylic acid repeating units. Although various micelles appeared similar in dynamic light scattering and cryo-transmission electron microscopy, changes in the micelles were evident from principal component analysis of the Raman spectra. Whereas an efficient stealth effect was found for micelles formed from polymers with anionically charged or thiol end groups, a hydrophobic end group altered the micelles' structure sufficiently to adapt cell-type specificity and stealth properties in the liver. © 2021 The Authors. Published by American Chemical Society.
- ItemActivity and electron donor preference of two denitrifying bacterial strains identified by Raman gas spectroscopy(Berlin [u.a.] : Springer, 2022) Blohm, Annika; Kumar, Swatantar; Knebl, Andreas; Herrmann, Martina; Küsel, Kirsten; Popp, Jürgen; Frosch, TorstenHuman activities have greatly increased the input of reactive nitrogen species into the environment and disturbed the balance of the global N cycle. This imbalance may be offset by bacterial denitrification, an important process in maintaining the ecological balance of nitrogen. However, our understanding of the activity of mixotrophic denitrifying bacteria is not complete, as most research has focused on heterotrophic denitrification. The aim of this study was to investigate substrate preferences for two mixotrophic denitrifying bacterial strains, Acidovorax delafieldii and Hydrogenophaga taeniospiralis, under heterotrophic, autotrophic or mixotrophic conditions. This complex analysis was achieved by simultaneous identification and quantification of H2, O2, CO2, 14N2, 15N2 and 15N2O in course of the denitrification process with help of cavity-enhanced Raman spectroscopic (CERS) multi-gas analysis. To disentangle electron donor preferences for both bacterial strains, microcosm-based incubation experiments under varying substrate conditions were conducted. We found that Acidovorax delafieldii preferentially performed heterotrophic denitrification in the mixotrophic sub-experiments, while Hydrogenophaga taeniospiralis preferred autotrophic denitrification in the mixotrophic incubation. These observations were supported by stoichiometric calculations. The results demonstrate the prowess of advanced Raman multi-gas analysis to study substrate use and electron donor preferences in denitrification, based on the comprehensive quantification of complex microbial gas exchange processes. © 2021, The Author(s).
- ItemIsolation of bacteria from artificial bronchoalveolar lavage fluid using density gradient centrifugation and their accessibility by Raman spectroscopy(Berlin [u.a.] : Springer, 2021) Wichmann, Christina; Rösch, Petra; Popp, JürgenRaman spectroscopy is an analytical method to identify medical samples of bacteria. Because Raman spectroscopy detects the biochemical properties of a cell, there are many factors that can influence and modify the Raman spectra of bacteria. One possible influence is a proper method for isolation of the bacteria. Medical samples in particular never occur in purified form, so a Raman-compatible isolation method is needed which does not affect the bacteria and thus the resulting spectra. In this study, we present a Raman-compatible method for isolation of bacteria from bronchoalveolar lavage (BAL) fluid using density gradient centrifugation. In addition to measuring the bacteria from a patient sample, the yield and the spectral influence of the isolation on the bacteria were investigated. Bacteria isolated from BAL fluid show additional peaks in comparison to pure culture bacteria, which can be attributed to components in the BAL sample. The isolation gradient itself has no effect on the spectra, and with a yield of 63% and 78%, the method is suitable for isolation of low concentrations of bacteria from a complex matrix. Graphical abstract.
- ItemMultimodal Molecular Imaging and Identification of Bacterial Toxins Causing Mushroom Soft Rot and Cavity Disease(Weinheim : Wiley-VCH, 2021) Dose, Benjamin; Thongkongkaew, Tawatchai; Zopf, David; Kim, Hak Joong; Bratovanov, Evgeni V.; García-Altares, María; Scherlach, Kirstin; Kumpfmüller, Jana; Ross, Claudia; Hermenau, Ron; Niehs, Sarah; Silge, Anja; Hniopek, Julian; Schmitt, Michael; Popp, Jürgen; Hertweck, ChristianSoft rot disease of edible mushrooms leads to rapid degeneration of fungal tissue and thus severely affects farming productivity worldwide. The bacterial mushroom pathogen Burkholderia gladioli pv. agaricicola has been identified as the cause. Yet, little is known about the molecular basis of the infection, the spatial distribution and the biological role of antifungal agents and toxins involved in this infectious disease. We combine genome mining, metabolic profiling, MALDI-Imaging and UV Raman spectroscopy, to detect, identify and visualize a complex of chemical mediators and toxins produced by the pathogen during the infection process, including toxoflavin, caryoynencin, and sinapigladioside. Furthermore, targeted gene knockouts and in vitro assays link antifungal agents to prevalent symptoms of soft rot, mushroom browning, and impaired mycelium growth. Comparisons of related pathogenic, mutualistic and environmental Burkholderia spp. indicate that the arsenal of antifungal agents may have paved the way for ancestral bacteria to colonize niches where frequent, antagonistic interactions with fungi occur. Our findings not only demonstrate the power of label-free, in vivo detection of polyyne virulence factors by Raman imaging, but may also inspire new approaches to disease control. © 2021 The Authors. ChemBioChem published by Wiley-VCH GmbH
- ItemPhotophysics of Anionic Bis(4H-imidazolato)CuI Complexes(Weinheim : Wiley-VCH, 2022) Seidler, Bianca; Tran, Jens H.; Hniopek, Julian; Traber, Philipp; Görls, Helmar; Gräfe, Stefanie; Schmitt, Michael; Popp, Jürgen; Schulz, Martin; Dietzek‐Ivanšić, BenjaminIn this paper, the photophysical behavior of four panchromatically absorbing, homoleptic bis(4H-imidazolato)CuI complexes, with a systematic variation in the electron-withdrawing properties of the imidazolate ligand, were studied by wavelength-dependent time-resolved femtosecond transient absorption spectroscopy. Excitation at 400, 480, and 630 nm populates metal-to-ligand charge transfer, intraligand charge transfer, and mixed-character singlet states. The pump wavelength-dependent transient absorption data were analyzed by a recently established 2D correlation approach. Data analysis revealed that all excitation conditions yield similar excited-state dynamics. Key to the excited-state relaxation is fast, sub-picosecond pseudo-Jahn-Teller distortion, which is accompanied by the relocalization of electron density onto a single ligand from the initially delocalized state at Franck-Condon geometry. Subsequent intersystem crossing to the triplet manifold is followed by a sub-100 ps decay to the ground state. The fast, nonradiative decay is rationalized by the low triplet-state energy as found by DFT calculations, which suggest perspective treatment at the strong coupling limit of the energy gap law.
- ItemNovel Biobased Self-Healing Ionomers Derived from Itaconic Acid Derivates(Weinheim : Wiley-VCH, 2021) Meurer, Josefine; Hniopek, Julian; Dahlke, Jan; Schmitt, Michael; Popp, Jürgen; Zechel, Stefan; Hager, Martin D.This article presents novel biobased ionomers featuring self-healing abilities. These smart materials are synthesized from itaconic acid derivates. Large quantities of itaconic acid can be produced from diverse biomass like corn, rice, and others. This study presents a comprehensive investigation of their thermal and mechanical properties via differential scanning calorimetry (DSC), thermo gravimetric analysis (TGA), and FT-Raman and FT-IR measurements as well as dynamic mechanic analysis. Within all these measurements, different kinds of structure-property relationships could be derived from these measurements. For example, the proportion of ionic groups enormously influences the self-healing efficiency. The investigation of the self-healing abilities reveals healing efficiencies up to 99% in 2 h at 90 °C for the itaconic acid based ionomer with the lowest ionic content. © 2020 The Authors. Macromolecular Rapid Communications published by Wiley-VCH GmbH
- ItemComparison of Different Label-Free Raman Spectroscopy Approaches for the Discrimination of Clinical MRSA and MSSA Isolates(Birmingham, Ala. : ASM, 2022) Pistiki, Aikaterini; Monecke, Stefan; Shen, Haodong; Ryabchykov, Oleg; Bocklitz, Thomas W.; Rösch, Petra; Ehricht, Ralf; Popp, JürgenMethicillin-resistant Staphylococcus aureus (MRSA) is classified as one of the priority pathogens that threaten human health. Resistance detection with conventional microbiological methods takes several days, forcing physicians to administer empirical antimicrobial treatment that is not always appropriate. A need exists for a rapid, accurate, and cost-effective method that allows targeted antimicrobial therapy in limited time. In this pilot study, we investigate the efficacy of three different label-free Raman spectroscopic approaches to differentiate methicillin-resistant and -susceptible clinical isolates of S. aureus (MSSA). Single-cell analysis using 532 nm excitation was shown to be the most suitable approach since it captures information on the overall biochemical composition of the bacteria, predicting 87.5% of the strains correctly. UV resonance Raman microspectroscopy provided a balanced accuracy of 62.5% and was not sensitive enough in discriminating MRSA from MSSA. Excitation of 785 nm directly on the petri dish provided a balanced accuracy of 87.5%. However, the difference between the strains was derived from the dominant staphyloxanthin bands in the MRSA, a cell component not associated with the presence of methicillin resistance. This is the first step toward the development of label-free Raman spectroscopy for the discrimination of MRSA and MSSA using single-cell analysis with 532 nm excitation. IMPORTANCE Label-free Raman spectra capture the high chemical complexity of bacterial cells. Many different Raman approaches have been developed using different excitation wavelength and cell analysis methods. This study highlights the major importance of selecting the most suitable Raman approach, capable of providing spectral features that can be associated with the cell mechanism under investigation. It is shown that the approach of choice for differentiating MRSA from MSSA should be single-cell analysis with 532 nm excitation since it captures the difference in the overall biochemical composition. These results should be taken into consideration in future studies aiming for the development of label-free Raman spectroscopy as a clinical analytical tool for antimicrobial resistance determination.