Browsing by Author "Tovar, Günter E. M."

Now showing 1 - 20 of 26

Open Access
Acid catalyzed cross‐linking of polyvinyl alcohol for humidifier membranes
(2021) Michele, Andre; Paschkowski, Patrick; Hänel, Christopher; Tovar, Günter E. M.; Schiestel, Thomas; Southan, Alexander
Polyvinyl alcohol (PVA) is a hydrophilic polymer well known for good film forming properties, high water vapor permeance JW, and low nitrogen permeance. However, depending on molar mass and temperature, PVA swells strongly in water until complete dissolution. This behavior affects the usability of PVA in aqueous environments and makes cross‐linking necessary if higher structural integrity is envisaged. In this work, PVA networks are formed by thermal cross‐linking in the presence of p‐toluenesulfonic acid (TSA) and investigated in a design of experiments approach. Experimental parameters are the cross‐linking period tc, temperature ϑ and the TSA mass fraction wTSA. Cross‐linking is found to proceed via ether bond formation at all reaction conditions. Degradation is promoted especially by a combination of high wTSA, tc and ϑ. Thermal stability of the networks after preparation is strongly improved by neutralizing residual TSA. Humidification membranes with a JW of 6423 ± 63.0 gas permeation units (GPU) are fabricated by coating PVA on polyvinyliden fluoride hollow fibers and cross‐linking with TSA. Summarizing, the present study contributes to a clearer insight into the cross‐linking of PVA in presence of TSA, the thermal stability of the resulting networks and the applicability as selective membrane layers for water vapor transfer.
Open Access
Active ester containing surfmer for one-stage polymer nanoparticle surface functionalization in mini-emulsion polymerization
(2018) Albernaz, Vanessa L.; Bach, Monika; Weber, Achim; Southan, Alexander; Tovar, Günter E. M.
Functional surface active monomers (surfmers) are molecules that combine the functionalities of surface activity, polymerizability and reactive groups. This study presents an improved pathway for the synthesis of the active ester containing surfmer p-(11-acrylamido)undecanoyloxyphenyl dimethylsulfonium methyl sulfate (AUPDS). Further, the preparation of poly(methyl methacrylate) and polystyrene nanoparticles (NPs) by miniemulsion polymerization using AUPDS is investigated, leading to NPs with active ester groups on their surface. By systematically varying reaction parameters and reagent concentrations, it was found that AUPDS feed concentrations between 2-4 mol-% yielded narrowly distributed and stable spherical particles with average sizes between 83 nm and 134 nm for non-cross-linked NPs, and up to 163 nm for cross-linked NPs. By basic hydrolysis of the active ester groups in aqueous dispersion, the positive ζ-potential (ZP) was converted into a negative ZP and charge quantities determined by polyelectrolyte titrations before and after hydrolysis were in the same range, indicating that the active ester groups were indeed accessible in aqueous suspension. Increasing cross-linker amounts over 10 mol-% also led to a decrease of ZP of NPs, probably due to internalization of the AUPDS during polymerization. In conclusion, by using optimized reaction conditions, it is possible to prepare active ester functionalized NPs in one stage using AUPDS as a surfmer in miniemulsion polymerization.
Open Access
Azido-functionalized gelatin via direct conversion of lysine amino groups by diazo transfer as a building block for biofunctional hydrogels
(2020) Keller, Silke; Bakker, Tomke; Kimmel, Benjamin; Rebers, Lisa; Götz, Tobias; Tovar, Günter E. M.; Kluger, Petra J.; Southan, Alexander
Gelatin is one of the most prominent biopolymers in biomedical material research and development. It is frequently used in hybrid hydrogels, which combine the advantageous properties of bio-based and synthetic polymers. To prevent the biological component from leaching out of the hydrogel, the biomolecules can be equipped with azides. Those groups can be used to immobilize gelatin covalently in hydrogels by the highly selective and specific azide-alkyne cycloaddition. In this contribution, we functionalized gelatin with azides at its lysine residues by diazo transfer, which offers the great advantage of only minimal side-chain extension. Approximately 84-90% of the amino groups are modified as shown by 1H-NMR spectroscopy, 2,4,6-trinitrobenzenesulfonic acid assay as well as Fourier-transform infrared spectroscopy, rheology, and the determination of the isoelectric point. Furthermore, the azido-functional gelatin is incorporated into hydrogels based on poly(ethylene glycol) diacrylate (PEG-DA) at different concentrations (0.6, 3.0, and 5.5%). All hydrogels were classified as noncyctotoxic with significantly enhanced cell adhesion of human fibroblasts on their surfaces compared to pure PEG-DA hydrogels. Thus, the new gelatin derivative is found to be a very promising building block for tailoring the bioactivity of materials.
Open Access
Beta-glucan production of Phaeodactylum tricornutum, Monodopsis subterranea and Cylindrotheca fusiformis during nitrogen depletion
(2023) Frick, Konstantin; Ebbing, Tobias; Yeh, Yen-Cheng; Schmid-Staiger, Ulrike; Tovar, Günter E. M.
AbstractBeta-glucans are polysaccharides that can be used for different applications, for example as an immunomodulator in food or feed or for managing high cholesterol levels. Certain microalgae species use beta-glucans as energy storage, accumulating them during nutrient depletion. In this study, we examined and compared beta-glucan production during nitrogen depletion in three different algae species, Phaeodactylum tricornutum, Monodopsis subterranea and Cylindrotheca fusiformis, grown in artificially illuminated flat panel airlift reactors, in order to determine the most promising microalgae species for beta-glucan production. Co-products such as fatty acids (especially eicosapentaenoic acid) and the carotenoid fucoxanthin (not produced by M. subterranea) were also considered. Biomass analysis showed that P. tricornutum cultures reached a maximal beta-glucan content of 317 ± 9 mg gDW-1, M. subterranea cultures reached 188 ± 6 mg gDW-1 and C. fusiformis cultures reached 129 ± 13 mg gDW-1. Furthermore, beta-glucan production was faster in P. tricornutum cultures. However, the maximum volumetric beta-glucan concentration reached was higher in M. subterranea cultures compared to P. tricornutum cultures as M. subterranea cultures produced more biomass during nitrogen depletion. In terms of possible co-products, P. tricornutum produced fucoxanthin and EPA, whereas M. subterranea did not produce fucoxanthin. However, M. subterranea exhibited a higher EPA content, which remained above 45 mg g-1 even after several days of nitrogen depletion. Overall, our results suggest that P. tricornutum and M. subterranea are both suitable species for beta-glucan production in flat panel airlift reactors.
Open Access
Charged triazole cross-linkers for hyaluronan-based hybrid hydrogels
(2016) Martini, Maike; Hegger, Patricia S.; Schädel, Nicole; Minsky, Burcu B.; Kirchhof, Manuel; Scholl, Sebastian; Southan, Alexander; Tovar, Günter E. M.; Boehm, Heike; Laschat, Sabine
Polyelectrolyte hydrogels play an important role in tissue engineering and can be produced from natural polymers, such as the glycosaminoglycan hyaluronan. In order to control charge density and mechanical properties of hyaluronan-based hydrogels, we developed cross-linkers with a neutral or positively charged triazole core with different lengths of spacer arms and two terminal maleimide groups. These cross-linkers react with thiolated hyaluronan in a fast, stoichiometric thio-Michael addition. Introducing a positive charge on the core of the cross-linker enabled us to compare hydrogels with the same interconnectivity, but a different charge density. Positively charged cross-linkers form stiffer hydrogels relatively independent of the size of the cross-linker, whereas neutral cross-linkers only form stable hydrogels at small spacer lengths. These novel cross-linkers provide a platform to tune the hydrogel network charge and thus the mechanical properties of the network. In addition, they might offer a wide range of applications especially in bioprinting for precise design of hydrogels.
Open Access
The choice of biopolymer is crucial to trigger angiogenesis with vascular endothelial growth factor releasing coatings
(2020) Claaßen, Christiane; Dannecker, Miriam; Grübel, Jana; Kotzampasi, Maria-Elli; Tovar, Günter E. M.; Stanzel, Boris V.; Borchers, Kirsten
Bio-based coatings and release systems for pro-angiogenic growth factors are of interest to overcome insufficient vascularization and bio-integration of implants. This study compares different biopolymer-based coatings on polyethylene terephthalate (PET) membranes in terms of coating homogeneity and stability, coating thickness in the swollen state, endothelial cell adhesion, vascular endothelial growth factor (VEGF) release and pro-angiogenic properties. Coatings consisted of carbodiimide cross-linked gelatin type A (GelA), type B (GelB) or albumin (Alb), and heparin (Hep), or they consisted of radically cross-linked gelatin methacryloyl-acetyl (GM5A5) and heparin methacrylate (HepM5). We prepared films with thicknesses of 8–10 µm and found that all coatings were homogeneous after washing. All gelatin-based coatings enhanced the adhesion of primary human endothelial cells compared to the uncoated membrane. The VEGF release was tunable with the loading concentration and dependent on the isoelectric points and hydrophilicities of the biopolymers used for coating: GelA-Hep showed the highest releases, while releases were indistinguishable for GelB-Hep and Alb-Hep, and lowest for GM5A5-HepM5. Interestingly, not only the amount of VEGF released from the coatings determined whether angiogenesis was induced, but a combination of VEGF release, metabolic activity and adhesion of endothelial cells. VEGF releasing GelA-Hep and GelB-Hep coatings induced angiogenesis in a chorioallantoic membrane assay, so that these coatings should be considered for further in vivo testing.
Open Access
Comparing three different Phaeodactylum tricornutum strains for the production of chrysolaminarin in flat panel airlift photobioreactors
(2022) Frick, Konstantin; Yeh, Yen-Cheng; Schmid-Staiger, Ulrike; Tovar, Günter E. M.
In recent years, various applications for algae-based ß-1,3-glucans have been postulated, including animal feed and human nutrition. Chrysolaminarin is a ß-1,3-1,6-glucan produced by diatoms such as Phaeodactylum tricornutum for energy storage. It is accumulated under nutrient-depleted cultivation conditions. In this study, the production of chrysolaminarin in artificially illuminated scalable flat panel airlift photobioreactors (FPA) was investigated by using P. tricornutum in a two-stage production process. In the growth stage primarily biomass is produced, and the subsequent nitrogen-depleted stage induces the accumulation of chrysolaminarin. Three P. tricornutum strains (SAG 1090-1a, SAG 1090-1b, SAG 1090-6) were cultured at laboratory scale in 6 L-FPA reactors under controlled light conditions to characterize the process and identify a production strain. The chrysolaminarin content of the algae strains was analysed and additionally their contents of eicosapentaenoic acid and fucoxanthin, both of which could be involved in a possible co-production. Strain SAG 1090-1b exhibited the highest biomass productivity and chrysolaminarin content (317 ± 9 mg gDW-1) after nitrogen depletion, and thus stood out as the most suitable for chrysolaminarin production in a two-stage process. A co-production of the three compounds is possible. However, during nitrogen depletion there occurred trade-offs between the compounds. As chrysolaminarin was produced, the amount of fucoxanthin in the culture stagnated or even decreased depending on the selected strain.
Open Access
Coumarin-4-ylmethyl- and p-hydroxyphenacyl-based photoacid generators with high solubility in aqueous media: synthesis, stability and photolysis
(2020) Adatia, Karishma K.; Halbritter, Thomas; Reinfelds, Matiss; Michele, Andre; Tran, Michael; Laschat, Sabine; Heckel, Alexander; Tovar, Günter E. M.; Southan, Alexander
(Coumarin‐4‐yl)methyl (c4m) and p‐hydroxyphenacyl (pHP)‐based compounds are well known for their highly efficient photoreactions, but often show limited solubility in aqueous media. To circumvent this, we synthesized and characterized the two new c4m and pHP‐based photoacid generators (PAGs), 7‐[bis(carboxymethyl)amino]‐4‐(acetoxymethyl)coumarin (c4m‐ac) and p‐hydroxyphenacyl‐2,5,8,11‐tetraoxatridecan‐13‐oate (pHP‐t), and determined their solubilities, stabilities and photolysis in aqueous media. The two compounds showed high solubilities in water of 2.77 mmol L−1±0.07 mmol L−1 (c4m‐ac) and 124.66 mmol L−1±2.1 mmol L−1 (pHP‐t). In basic conditions at pH 9, solubility increased for c4m‐ac to 646.46 mmol L−1±0.63 mmol L−1, for pHP‐t it decreased to 34.68 mmol L−1±0.62 mmol L−1. Photochemical properties of the two PAGs, such as the absorption maxima, the maximum molar absorption coefficients and the quantum yields, were found to be strongly pH‐dependent. Both PAGs showed high stabilities s24h ≥95 % in water for 24 h, but decreasing stability with increasing pH value due to hydrolysis. The present study contributes to a clearer insight into the synthesis, solubilities, stabilities, and photolysis of c4m and pHP‐based PAGs for further photochemical applications when high PAG concentrations are required, such as in polymeric foaming.
Open Access
Covalent incorporation of tobacco mosaic virus increases the stiffness of poly(ethylene glycol) diacrylate hydrogels
(2018) Southan, Alexander; Lang, Tina; Schweikert, Michael; Tovar, Günter E. M.; Wege, Christina; Eiben, Sabine
Hydrogels are versatile materials, finding applications as adsorbers, supports for biosensors and biocatalysts or as scaffolds for tissue engineering. A frequently used building block for chemically cross-linked hydrogels is poly(ethylene glycol) diacrylate (PEG-DA). However, after curing, PEG-DA hydrogels cannot be functionalized easily. In this contribution, the stiff, rod-like tobacco mosaic virus (TMV) is investigated as a functional additive to PEG-DA hydrogels. TMV consists of more than 2000 identical coat proteins and can therefore present more than 2000 functional sites per TMV available for coupling, and thus has been used as a template or building block for nano-scaled hybrid materials for many years. Here, PEG-DA (Mn = 700 g/mol) hydrogels are combined with a thiol-group presenting TMV mutant (TMVCys). By covalent coupling of TMVCys into the hydrogel matrix via the thiol-Michael reaction, the storage modulus of the hydrogels is increased compared to pure PEG-DA hydrogels and to hydrogels containing wildtype TMV (wt-TMV) which is not coupled covalently into the hydrogel matrix. In contrast, the swelling behaviour of the hydrogels is not altered by TMVCys or wt-TMV. Transmission electron microscopy reveals that the TMV particles are well dispersed in the hydrogels without any large aggregates. These findings give rise to the conclusion that well-defined hydrogels were obtained which offer the possibility to use the incorporated TMV as multivalent carrier templates e.g. for enzymes in future studies.
Open Access
Differentiation of physical and chemical cross-linking in gelatin methacryloyl hydrogels
(2021) Rebers, Lisa; Reichsöllner, Raffael; Regett, Sophia; Tovar, Günter E. M.; Borchers, Kirsten; Baudis, Stefan; Southan, Alexander
Gelatin methacryloyl (GM) hydrogels have been investigated for almost 20 years, especially for biomedical applications. Recently, strengthening effects of a sequential cross-linking procedure, whereby GM hydrogel precursor solutions are cooled before chemical cross-linking, were reported. It was hypothesized that physical and enhanced chemical cross-linking of the GM hydrogels contribute to the observed strengthening effects. However, a detailed investigation is missing so far. In this contribution, we aimed to reveal the impact of physical and chemical cross-linking on strengthening of sequentially cross-linked GM and gelatin methacryloyl acetyl (GMA) hydrogels. We investigated physical and chemical cross-linking of three different GM(A) derivatives (GM10, GM2A8 and GM2), which provided systematically varied ratios of side-group modifications. GM10 contained the highest methacryloylation degree (DM), reducing its ability to cross-link physically. GM2 had the lowest DM and showed physical cross-linking. The total modification degree, determining the physical cross-linking ability, of GM2A8 was comparable to that of GM10, but the chemical cross-linking ability was comparable to GM2. At first, we measured the double bond conversion (DBC) kinetics during chemical GM(A) cross-linking quantitatively in real-time via near infrared spectroscopy-photorheology and showed that the DBC decreased due to sequential cross-linking. Furthermore, results of circular dichroism spectroscopy and differential scanning calorimetry indicated gelation and conformation changes, which increased storage moduli of all GM(A) hydrogels due to sequential cross-linking. The data suggested that the total cross-link density determines hydrogel stiffness, regardless of the physical or chemical nature of the cross-links.
Open Access
Eclectic characterisation of chemically modified cell-derived matrices obtained by metabolic glycoengineering and re-assessment of commonly used methods
(2020) Keller, Silke; Liedek, Anke; Shendi, Dalia; Bach, Monika; Tovar, Günter E. M.; Kluger, Petra J.; Southan, Alexander
Azide-bearing cell-derived extracellular matrices (“clickECMs”) have emerged as a highly exciting new class of biomaterials. They conserve substantial characteristics of the natural extracellular matrix (ECM) and offer simultaneously small abiotic functional groups that enable bioorthogonal bioconjugation reactions. Despite their attractiveness, investigation of their biomolecular composition is very challenging due to the insoluble and highly complex nature of cell-derived matrices (CDMs). Yet, thorough qualitative and quantitative analysis of the overall material composition, organisation, localisation, and distribution of typical ECM-specific biomolecules is essential for consistent advancement of CDMs and the understanding of the prospective functions of the developed biomaterial. In this study, we evaluated frequently used methods for the analysis of complex CDMs. Sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) and (immune)histochemical staining methods in combination with several microscopic techniques were found to be highly eligible. Commercially available colorimetric protein assays turned out to deliver inaccurate information on CDMs. In contrast, we determined the nitrogen content of CDMs by elementary analysis and converted it into total protein content using conversion factors which were calculated from matching amino acid compositions. The amount of insoluble collagens was assessed based on the hydroxyproline content. The Sircol™ assay was identified as a suitable method to quantify soluble collagens while the Blyscan™ assay was found to be well-suited for the quantification of sulphated glycosaminoglycans (sGAGs). Eventually, we propose a series of suitable methods to reliably characterise the biomolecular composition of fibroblast-derived clickECM.
Open Access
Extrusion-based 3D printing of poly(ethylene glycol) diacrylate hydrogels containing positively and negatively charged groups
(2018) Joas, Sebastian; Tovar, Günter E. M.; Celik, Oguz; Bonten, Christian; Southan, Alexander
Hydrogels are an interesting class of materials used in extrusion-based 3D printing, e.g., for drug delivery or tissue engineering. However, new hydrogel formulations for 3D printing as well as a detailed understanding of crucial formulation properties for 3D printing are needed. In this contribution, hydrogels based on poly(ethylene glycol) diacrylate (PEG-DA) and the charged monomers 3-sulfopropyl acrylate and [2-(acryloyloxy)ethyl]trimethylammonium chloride are formulated for 3D printing, together with Poloxamer 407 (P407). Chemical curing of formulations with PEG-DA and up to 5% (w/w) of the charged monomers was possible without difficulty. Through careful examination of the rheological properties of the non-cured formulations, it was found that flow properties of formulations with a high P407 concentration of 22.5% (w/w) possessed yield stresses well above 100 Pa together with pronounced shear thinning behavior. Thus, those formulations could be processed by 3D printing, as demonstrated by the generation of pyramidal objects. Modelling of the flow profile during 3D printing suggests that a plug-like laminar flow is prevalent inside the printer capillary. Under such circumstances, fast recovery of a high vicosity after material deposition might not be necessary to guarantee shape fidelity because the majority of the 3D printed volume does not face any relevant shear stress during printing.
Open Access
Fermentation and recovery of cellobiose lipids using foam fractionation
(2023) Oraby, Amira; Hug, Daniel; Weickardt, Isabell; Maerz, Lea; Nebel, Sabrina; Kurmann, Jasper; Rupp, Steffen; Tovar, Günter E. M.; Zibek, Susanne
Cellobiose lipids (CL) are glycolipids secreted by many Ustilaginaceae species in aerobic fermentations characterised by excessive foaming. While increasing CL concentrations remains an aim for its industrial production, excessive foaming during fermentation presents a challenge even at laboratory scale. Foam fractionation (FF) provides a solution to the foaming problem and facilitates the proceeding purification of CL. Here, we present a first CL fermentation process applying FF. With our set-up, we manage to exploit the excessive foaming for continuous product separation. The set-up includes a foam collecting vessel (FCV) with inserts for CL accumulation and foamate recirculation to minimise biomass and nutrient loss. Integrating a foam column (FC) into the fermenter headspace enabled foam enrichment, resulting in the recovery of > 90% of the produced CL from the separated fractions consisting of foam depositions in the fermenter headspace and the FCV. We also increased the fermenter filling volume and thus achieved a higher fermentation capacity. The separated CL fraction was purified via ethanol extraction to obtain CL with purities > 90%. We further examined the effects of different culture media constituents, including biomass and CL, on foam generation and decay and assessed the effect of FC geometries on product enrichment and recovery. In this work, a FF set-up is presented that enables a stable CL fermentation without additional foam mitigation methods. At the same time, the application of FF separated a fraction that was highly enriched in CL during fermentation, resulting in highly pure CL after a simple ethanol extraction.
Open Access
Friction and wear behavior of deep drawing tools using volatile lubricants injected through laser-drilled micro-holes
(2021) Reichardt, Gerd; Henn, Manuel; Reichle, Paul; Umlauf, Georg; Riedmüller, Kim; Weber, Rudolf; Barz, Jakob; Liewald, Mathias; Graf, Thomas; Tovar, Günter E. M.
In deep drawing processes, the use of lubricants is mandatory in order to prevent wear on tools and surface damage to the formed sheet metal components. Here, frequently used lubricants are synthetic and mineral oils, emulsions, and waxes. However, these conventional lubricants have to be applied to the sheet material prior to the forming operation and removed afterwards by cleaning processes. Additionally, the lubricants often contain substances that are harmful to the environment and to human health. To counteract these economic and ecological disadvantages, research is currently being conducted on a novel tribological system. For this, volatile media such as liquid carbon dioxide and gaseous nitrogen are being used, and are introduced directly into the friction zones between the tool and the sheet metal material during deep drawing under high pressure through special laser-drilled micro-holes. This paper covers the latest investigations and findings regarding the design of flow-optimized micro-holes, the laser drilling process, the friction characterization on tool radii, and the tool wear to be expected when using the lubrication medium CO2.
Open Access
Improving determination of pigment contents in microalgae suspension with absorption spectroscopy : light scattering effect and Bouguer-Lambert-Beer law
(2023) Yeh, Yen-Cheng; Ebbing, Tobias; Frick, Konstantin; Schmid-Staiger, Ulrike; Haasdonk, Bernard; Tovar, Günter E. M.
The Bouguer-Lambert-Beer (BLB) law serves as the fundamental basis for the spectrophotometric determination of pigment content in microalgae. Although it has been observed that the applicability of the BLB law is compromised by the light scattering effect in microalgae suspensions, in-depth research concerning the relationship between the light scattering effect and the accuracy of spectrophotometric pigment determination remains scarce. We hypothesized that (1) the precision of spectrophotometric pigment content determination using the BLB law would diminish with increasing nonlinearity of absorbance, and (2) employing the modified version of the BLB (mBLB) law would yield superior performance. To assess our hypotheses, we cultivated Phaeodactylum tricornutum under varying illumination conditions and nitrogen supplies in controlled indoor experiments, resulting in suspensions with diverse pigment contents. Subsequently, P. tricornutum samples were diluted into subsamples, and spectral measurements were conducted using different combinations of biomass concentrations and path lengths. This was carried out to assess the applicability of the BLB law and the nonlinearity of absorbance. The chlorophyll a and fucoxanthin contents in the samples were analyzed via high-performance liquid chromatography (HPLC) and subsequently used in our modeling. Our findings confirm our hypotheses, showing that the modified BLB law outperforms the original BLB law in terms of the normalized root mean square error (NRMSE): 6.3% for chlorophyll a and 5.8% for fucoxanthin, compared to 8.5% and 7.9%, respectively.
Open Access
Influence of light conditions on the production of chrysolaminarin using Phaeodactylum tricornutum in artificially illuminated photobioreactors
(2023) Frick, Konstantin; Ebbing, Tobias; Yeh, Yen‐Cheng; Schmid‐Staiger, Ulrike; Tovar, Günter E. M.
The light conditions are of utmost importance in any microalgae production process especially involving artificial illumination. This also applies to a chrysolaminarin (soluble 1,3-β-glucan) production process using the diatom Phaeodactylum tricornutum. Here we examine the influence of the amount of light per gram biomass (specific light availability) and the influence of two different biomass densities (at the same amount of light per gram biomass) on the accumulation of the storage product chrysolaminarin during nitrogen depletion in artificially illuminated flat-panel airlift photobioreactors. Besides chrysolaminarin, other compounds (fucoxanthin, fatty acids used for energy storage [C16 fatty acids], and eicosapentaenoic acid) are regarded as well. Our results show that the time course of C-allocation between chrysolaminarin and fatty acids, serving as storage compounds, is influenced by specific light availability and cell concentration. Furthermore, our findings demonstrate that with increasing specific light availability, the maximal chrysolaminarin content increases. However, this effect is limited. Beyond a certain specific light availability (here: 5 µmolphotons gDW-1 s-1) the maximal chrysolaminarin content no longer increases, but the rate of increase becomes faster. Furthermore, the conversion of light to chrysolaminarin is best at the beginning of nitrogen depletion. Additionally, our results show that a high biomass concentration has a negative effect on the maximal chrysolaminarin content, most likely due to the occurring self-shading effects.
Open Access
Nano-in-micro-particles consisting of PLGA nanoparticles embedded in chitosan microparticles via spray-drying enhances their uptake in the olfactory mucosa
(2021) Spindler, Lena Marie; Feuerhake, Andreas; Ladel, Simone; Günday, Cemre; Flamm, Johannes; Günday-Türeli, Nazende; Türeli, Emre; Tovar, Günter E. M.; Schindowski, Katharina; Gruber-Traub, Carmen
Intranasal delivery has gained prominence since 1990, when the olfactory mucosa was recognized as the window to the brain and the central nervous system (CNS); this has enabled the direct site specific targeting of neurological diseases for the first time. Intranasal delivery is a promising route because general limitations, such as the blood-brain barrier (BBB) are circumvented. In the treatment of multiple sclerosis (MS) or Alzheimer’s disease, for example, future treatment prospects include specialized particles as delivery vehicles. Poly(lactic-co-glycolic acid) (PLGA) nanoparticles are well known as promising delivery systems, especially in the area of nose-to-brain (N2B) delivery. Chitosan is also broadly known as a functional additive due to its ability to open tight junctions. In this study, we produced PLGA nanoparticles of different sizes and revealed for the first time their size-time-dependent uptake mechanism into the lamina propria of porcine olfactory mucosa. The intracellular uptake was observed for 80 and 175 nm within only 5 min after application to the epithelium. After 15 min, even 520 nm particles were detected, associated with nuclei. Especially the presence of only 520 nm particles in neuronal fibers is remarkable, implying transcellular and intracellular transport via the olfactory or the trigeminal nerve to the brain and the CNS. Additionally, we developed successfully specialized Nano-in-Micro particles (NiMPs) for the first time via spray drying, consisting of PLGA nanoparticles embedded into chitosan microparticles, characterized by high encapsulation efficiencies up to 51%, reproducible and uniform size distribution, as well as smooth surface. Application of NiMPs accelerated the uptake compared to purely applied PLGA nanoparticles. NiMPs were spread over the whole transverse section of the olfactory mucosa within 15 min. Faster uptake is attributed to additional paracellular transport, which was examined via tight-junction-opening. Furthermore, a separate chitosan penetration gradient of ∼150 µm caused by dissociation from PLGA nanoparticles was observed within 15 min in the lamina propria, which was demonstrated to be proportional to an immunoreactivity gradient of CD14. Due to the beneficial properties of the utilized chitosan-derivative, regarding molecular weight (150-300 kDa), degree of deacetylation (80%), and particle size (0.1-10 µm) we concluded that M2-macrophages herein initiated an anti-inflammatory reaction, which seems to already take place within 15 min following chitosan particle application. In conclusion, we demonstrated the possibility for PLGA nanoparticles, as well as for chitosan NiMPs, to take all three prominent intranasal delivery pathways to the brain and the CNS; namely transcellular, intracellular via neuronal cells, and paracellular transport.
Open Access
A novel model extended from the Bouguer-Lambert-Beer law can describe the non-linear absorbance of potassium dichromate solutions and microalgae suspensions
(2023) Yeh, Yen-Cheng; Haasdonk, Bernard; Schmid-Staiger, Ulrike; Stier, Matthias; Tovar, Günter E. M.
The Bouguer-Lambert-Beer law is widely used as the fundamental equation for quantification in absorption spectroscopy. However, deviations from the Bouguer-Lambert-Beer law have also been observed, such as chemical deviation and light scattering effect. While it has been proven and shown that the Bouguer-Lambert-Beer law is valid only under very restricted limitations, there are only a few alternatives of analytical models to this law. Based on the observation in the experiments, we propose a novel model to solve the problem of chemical deviation and light scattering effect. To test the proposed model, a systematic verification was conducted using potassium dichromate solutions and two types of microalgae suspensions with varying concentrations and path lengths. Our proposed model demonstrated excellent performance, with a correlation coefficient (R2) exceeding 0.995 for all tested materials, significantly surpassing the Bouguer-Lambert-Beer law, which had an R2 as low as 0.94. Our results confirm that the absorbance of pure pigment solutions follows the Bouguer-Lambert-Beer law, while the microalgae suspensions do not due to the light scattering effect. We also show that this scattering effect leads to huge deviations for the commonly used linear scaling of the spectra, and we provide a better solution based on the proposed model. This work provides a powerful tool for chemical analysis and especially for the quantification of microorganisms, such as the concentration of biomass or intracellular biomolecules. Not only the high accuracy but also the simplicity of the model makes it a practical alternative to the existing Bouguer-Lambert-Beer law.
Open Access
Physical interactions strengthen chemical gelatin methacryloyl gels
(2019) Rebers, Lisa; Granse, Tobias; Tovar, Günter E. M.; Southan, Alexander; Borchers, Kirsten
Chemically cross-linkable gelatin methacryloyl (GM) derivatives are getting increasing attention regarding biomedical applications. Thus, thorough investigations are needed to achieve full understanding and control of the physico-chemical behavior of these promising biomaterials. We previously introduced gelatin methacryloyl acetyl (GMA) derivatives, which can be used to control physical network formation (solution viscosity, sol-gel transition) independently from chemical cross-linking by variation of the methacryloyl-to-acetyl ratio. It is known that temperature dependent physical network formation significantly influences the mechanical properties of chemically cross-linked GM hydrogels. We investigated the temperature sensitivity of GM derivatives with different degrees of modification (GM2, GM10), or similar degrees of modification but different methacryloyl contents (GM10, GM2A8). Rheological analysis showed that the low modified GM2 forms strong physical gels upon cooling while GM10 and GM2A8 form soft or no gels. Yet, compression testing revealed that all photo cross-linked GM(A) hydrogels were stronger if cooling was applied during hydrogel preparation. We suggest that the hydrophobic methacryloyl and acetyl residues disturb triple helix formation with increasing degree of modification, but additionally form hydrophobic structures, which facilitate chemical cross-linking.
Open Access
Precision 3D‐printed cell scaffolds mimicking native tissue composition and mechanics
(2020) Erben, Amelie; Hörning, Marcel; Hartmann, Bastian; Becke, Tanja; Eisler, Stephan A.; Southan, Alexander; Cranz, Séverine; Hayden, Oliver; Kneidinger, Nikolaus; Königshoff, Melanie; Lindner, Michael; Tovar, Günter E. M.; Burgstaller, Gerald; Clausen‐Schaumann, Hauke; Sudhop, Stefanie; Heymann, Michael
Cellular dynamics are modeled by the 3D architecture and mechanics of the extracellular matrix (ECM) and vice versa. These bidirectional cell‐ECM interactions are the basis for all vital tissues, many of which have been investigated in 2D environments over the last decades. Experimental approaches to mimic in vivo cell niches in 3D with the highest biological conformity and resolution can enable new insights into these cell‐ECM interactions including proliferation, differentiation, migration, and invasion assays. Here, two‐photon stereolithography is adopted to print up to mm‐sized high‐precision 3D cell scaffolds at micrometer resolution with defined mechanical properties from protein‐based resins, such as bovine serum albumin or gelatin methacryloyl. By modifying the manufacturing process including two‐pass printing or post‐print crosslinking, high precision scaffolds with varying Young's moduli ranging from 7‐300 kPa are printed and quantified through atomic force microscopy. The impact of varying scaffold topographies on the dynamics of colonizing cells is observed using mouse myoblast cells and a 3D‐lung microtissue replica colonized with primary human lung fibroblast. This approach will allow for a systematic investigation of single‐cell and tissue dynamics in response to defined mechanical and bio‐molecular cues and is ultimately scalable to full organs.