Etablierung und Optimierung umweltfreundlicher Prozesse zur Modifikation steroidaler Verbindungen mittels Ganzzellkatalysen

Steroidale Verbindungen regulieren u.a. die Mineralstoff-Homöostaste, sowie Immun- und Entzündungsreaktionen und sind daher von hoher pharmazeutischer Relevanz. Die Herausforderung ihrer industriellen Synthese besteht neben hohen Ausbeuten in der Minimierung der einhergehenden Umweltbelastung. Zu diesem Zweck wurden in dieser Arbeit effiziente Ganzzellsysteme etabliert und optimiert, welche fähig sind ökologisch nachhaltige Teilsynthesen unterschiedlicher (Seco-) Steroide zu katalysieren. Zum einen konnte die effiziente Oxidation von Hydrocortison etabliert werden, welche, verglichen mit einem Referenzsystem, eine 1000-fach gesteigerte Cortisonausbeute von 14 g*L -1 aufwies. Eine SHE-Bewertung demonstrierte zudem eine erhöhte Ökoeffizienz im direkten Vergleich zu einem äquivalenten chemischen Verfahren. Ein weiteres Ganzzellsystem ermöglicht eine Hydroxylierung von Medran, welche als limitierender Teilschritt der Methylprednisolon-Synthese gilt. Hier konnte die Produktausbeute durch Optimierung verdoppelt werden. Die Etablierung einer Mischkultur zur Produktderivatisierung führte zu einer weiteren Erhöhung der Premedrolproduktion um 117% auf >1,5 g*L -1. Außerdem wurde die erste biotechnologische und daher hochselektive Hydroxylierung von Vitamin D2 zur Produktion von physiologisch aktiven Metaboliten entwickelt. All diese Ergebnisse unterstreichen das große Potenzial von Ganzzellsystemen, die ökologische Nachhaltigkeit industrieller Synthesen von Feinchemikalien zu erhöhen.Steroidal compounds regulate the mineral homeostasis as well as immune and inflammatory reactions and are, thus, highly relevant for pharma industry. Besides achieving high product yields, the industrial synthesis of these compounds is challenging in terms of minimizing accruing environmental pollution. For these reasons, this work focused on the establishment and optimization of diverse whole-cell systems to catalyze different sub-processes of the (seco-) steroid synthesis in an efficient and sustainable matter. In the first part, the efficient oxidation of hydrocortisone was enabled, which resulted in a 1000-fold increase of the product yield compared with a reference system. Furthermore, an SHE-assessment demonstrated improved ecological efficiency in comparison with an equivalent chemical process. A second whole-cell system catalyzes the hydroxylation of medrane, a limiting step of methylprednisolone synthesis. Here, the product yield was doubled by optimization. Moreover, the establishment of a mixed-culture fermentation for subsequent product derivatization led to a further enhancement of the premedrol production by 117%, up to 1.5 g L -1. Finally, the first biotechnological and thus, highly selective hydroxylation process of vitamin D2 was established for the production of physiologically active metabolites. All obtained results highlight the great potential of whole-cell systems to increase ecological sustainability for the industrial synthesis of fine chemicals.Deutsche Bundesstiftung Umwelt (DBU

Identification and circumvention of bottlenecks in CYP21A2-mediated premedrol production using recombinant Escherichia coli

Synthetic glucocorticoids such as methylprednisolone are compounds of fundamental interest to the pharmaceutical industry as their modifications within the sterane scaffold lead to higher inflammatory potency and reduced side effects compared with their parent compound cortisol. In methylprednisolone production, the complex chemical hydroxylation of its precursor medrane in position C21 exhibits poor stereo‐ and regioselectivity making the process unprofitable and unsustainable. By contrast, the use of a recombinant E. coli system has recently shown high suitability and efficiency. In this study, we aim to overcome limitations in this biotechnological medrane conversion yielding the essential methylprednisolone‐precursor premedrol by optimizing the CYP21A2‐based whole‐cell system on a laboratory scale. We successfully improved the whole‐cell process in terms of premedrol production by (a) improving the electron supply to CYP21A2; here we use the N‐terminally truncated version of the bovine NADPH‐dependent cytochrome P450 reductase (bCPR−27) and coexpression of microsomal cytochrome b5; (b) enhancing substrate access to the heme by modification of the CYP21A2 substrate access channel; and (c) circumventing substrate inhibition which is presumed to be the main limiting factor of the presented system by developing an improved fed‐batch protocol. By overcoming the presented limitations in whole‐cell biotransformation, we were able to achieve a more than 100% improvement over the next best system under equal conditions resulting in 691 mg·L−1·d−1 premedrol

Carbon Dynamics During the Formation of Sea Ice at Different Growth Rates

Controlled laboratory experiments have shed new light on the potential importance of brine rejection during sea-ice formation for carbon dioxide sequestration in the ocean. We grew ice in an experimental seawater tank (1 m3) under abiotic conditions at three different air temperatures (−40°C, −25°C, −15°C) to determine how different ice growth rates affect the allocation of carbon to ice, water, or air. Carbonate system parameters were determined by discrete sampling of ice cores and water, as well as continuous measurements by multiple sensors deployed mainly in the water phase. A budgetary approach revealed that of the initial total inorganic carbon (TIC) content of the water converted to ice, only 28–29% was located in the ice phase by the end of the experiments run at the warmest temperature, whereas for the coldest ambient temperature, 46–47% of the carbon remained in the ice. Exchange with air appeared to be negligible, with the majority of the TIC remaining in the under-ice water (53–72%). Along with a good correlation between salinity and TIC in the ice and water samples, these observations highlight the importance of brine drainage to TIC redistribution during ice formation. For experiments without mixing of the under-ice water, the sensor data further suggested stronger stratification, likely related to release of denser brine, and thus potentially larger carbon sequestration for ice grown at a colder temperature and faster growth rate

Politik, Journalismus, Medien. Kompetenzen von Kindern im Vor- und Grundschulalter (PoJoMeC)

Ab wann entwickeln Kinder konkrete Vorstellungen von Politik und welche Rolle spielen dabei Kinderinformations- und Unterhaltungsmedien? Wann werden z. B. Nachrichtensendungen von Kindern bewusst wahrgenommen? Ab wann wird Kindern der Zusammenhang von Politik und Massenmedien bewusst? Diese Fragen stehen im Zentrum des Forschungsprojekts PoJoMeC, das die Autor*innen in ihrem Beitrag vorstellen. Durch die Erhebung der kindlichen Konzept- und Einstellungsentwicklung wollen die Autor*innen dabei nicht nur Erkenntnisse über die politische und medienbezogene Partizipation von Kindern gewinnen. Die Ergebnisse sollen gleichzeitig auch einen Beitrag zur Qualitätssteigerung der Ausbildung künftiger Lehrkräfte und Journalist*innen leisten. (DIPF/Orig.)The article introduces the interdisciplinary research and development project PoJoMeC (= Politics, Journalism, Media – Competences of Preschool and Elementary School Children) funded by the Federal Agency for Civic Education (Germany). Among others, this longitudinal study seeks to explore the characteristics and development of ideas children at the age of 4 to 9 years have about politics, journalism and media (use). While research in the field of political didactic already focuses on young children, this is not the case in journalistic research yet. Furthermore, little knowledge exists on the influence of media and media use on the democratic socialization or the political understanding of preschool children. Based on our findings, we want to draw conclusions for pedagogical measures regarding political and civic education. (DIPF/Orig.

5-Keto-D-Fructose, a Natural Diketone and Potential Sugar Substitute, Significantly Reduces the Viability of Prokaryotic and Eukaryotic Cells

5-Keto-D-fructose (5-KF) is a natural diketone occurring in micromolar concentrations in honey, white wine, and vinegar. The oxidation of D-fructose to 5-KF is catalyzed by the membrane-bound fructose dehydrogenase complex found in several acetic acid bacteria. Since 5-KF has a sweetening power comparable to fructose and is presumably calorie-free, there is great interest in making the diketone commercially available as a new sugar substitute. Based on a genetically modified variant of the acetic acid bacterium Gluconobacter oxydans 621H, an efficient process for the microbial production of 5-KF was recently developed. However, data on the toxicology of the compound are completely lacking to date. Therefore, this study aimed to investigate the effect of 5-KF on the viability of prokaryotic and eukaryotic cells. It was found that the compound significantly inhibited the growth of the gram-positive and gram-negative model organisms Bacillus subtilis and Escherichia coli in a concentration-dependent manner. Furthermore, cell viability assays confirmed severe cytotoxicity of 5-KF toward the colon cancer cell line HT-29. Since these effects already occurred at concentrations of 5 mM, the use of 5-KF in the food sector should be avoided. The studies performed revealed that in the presence of amines, 5-KF promoted a strong Maillard reaction. The inherent reactivity of 5-KF as well as the Maillard products formed could be the trigger for the observed inhibition of prokaryotic and eukaryotic cells

Host shift induces changes in mate choice of the seed predator Acanthoscelides obtectus via altered chemical signalling

The mechanisms of host shift in phytophagous insects are poorly understood. Among the many proposed processes involved, sexual selection via semiochemicals has recently been suggested. This hypothesizes that sexual communication using pheromones is modified as a result of development on a new host, and such plant-induced phenotypic divergence in mate recognition cues can lead to reproductive isolation between host lines. We tested this hypothesis on Acanthoscelides obtectus, an oligophagous bruchid of Phaseolus vulgaris beans worldwide, which also develops in acceptable non-hosts, such as chickpea (Cicer arietinum L.). Male sex pheromone blends of the bean, chickpea and chickpea/bean host lines during artificially induced host shifts showed different composition. Bean-reared females did not distinguish between blends, whereas chickpea and chickpea/bean females preferred the chickpea male pheromone. However, electrophysiological (EAG) responses to male odour of antennae of the three female host lines were similar, all preferring bean-reared males. Egg-laying choice tests revealed a uniform preference for bean seeds across female host lines, even after multiple generations, whereas larvae did not distinguish between bean and chickpea seeds. We conclude that the development of divergent chemical signalling systems during host shifts does not facilitate the evolution of host races in A. obtectus, because oviposition preferences remain unaffected

Efficacy of Bifidobacterium animalis subsp. lactis (BB-12), B. infantis and Lactobacillus acidophilus (La-5) probiotics to prevent gut dysbiosis in preterm infants of 28+0–32+6 weeks of gestation: a randomised, placebocontrolled, double-blind, multicentre trial: the PRIMAL Clinical Study Protocol

Introduction The healthy ‘eubiosis’ microbiome in infancy is regarded as the microbiome derived from term, vaginally delivered, antibiotic free, breastfed infants at 4–6 months. Dysbiosis is regarded as a deviation from a healthy state with reduced microbial diversity and deficient capacity to control drug-resistant organisms. Preterm infants are highly sensitive to early gut dysbiosis. Latter has been associated with sepsis and necrotising enterocolitis, but may also contribute to long-term health problems. Probiotics hold promise to reduce the risk for adverse short-term outcomes but the evidence from clinical trials remains inconclusive and none has directly assessed the effects of probiotics on the microbiome at high resolution. Methods and analysis A randomised, double blind, placebo-controlled study has been designed to assess the safety and efficacy of the probiotic mix of Bifidobacterium animalis subsp. lactis, B. infantis and Lactobacillus acidophilus in the prevention of gut dysbiosis in preterm infants between 28+0 and 32+6 weeks of gestation. The study is conducted in 18 German neonatal intensive care units. Between April 2018 and March 2020, 654 preterm infants of 28+0–32+6 weeks of gestation will be randomised in the first 48 hours of life to 28 days of once daily treatment with either probiotics or placebo. The efficacy endpoint is the prevention of gut dysbiosis at day 30 of life. A compound definition of gut dysbosis is used: (1) colonisation with multidrug-resistant organisms or gram-negative bacteria with high epidemic potential or (2) a significant deviation of the gut microbiota composition as compared with healthy term infants. Dysbiosis is determined by (1) conventional microbiological culture and (2) phylogenetic microbiome analysis by high-throughput 16S rRNA and metagenome sequencing. Persistence of dysbiosis will be assessed at 12-month follow-up visits. Side effects and adverse events related to the intervention will be recorded. Key secondary endpoint(s) are putative consequences of dysbiosis. A subgroup of infants will be thoroughly phenotyped for immune parameters using chipcytometry. Ethics and dissemination Ethics approval was obtained in all participating sites. Results of the trial will be published in peer-review journals, at scientific meetings, on the website (www.primal-study.de) and via social media of parent organisations. Trial registration number DRKS00013197; Pre-results