Glucose‐based microbial production of the hormone melatonin in yeast Saccharomyces cerevisiae

Melatonin is a natural mammalian hormone that plays an important role in regulating the circadian cycle in humans. It is a clinically effective drug exhibiting positive effects as a sleep aid and a powerful antioxidant used as a dietary supplement. Commercial melatonin production is predominantly performed by complex chemical synthesis. In this study, we demonstrate microbial production of melatonin and related compounds, such as serotonin and N‐acetylserotonin. We generated Saccharomyces cerevisiae strains that comprise heterologous genes encoding one or more variants of an L‐tryptophan hydroxylase, a 5‐hydroxy‐L‐tryptophan decarboxylase, a serotonin acetyltransferase, an acetylserotonin O‐methyltransferase, and means for providing the cofactor tetrahydrobiopterin via heterologous biosynthesis and recycling pathways. We thereby achieved de novo melatonin biosynthesis from glucose. We furthermore accomplished increased product titers by altering expression levels of selected pathway enzymes and boosting co‐factor supply. The final yeast strain produced melatonin at a titer of 14.50 ± 0.57 mg L^?1 in a 76h fermentation using simulated fed‐batch medium with glucose as sole carbon source. Our study lays the basis for further developing a yeast cell factory for biological production of melatonin.     

On the Effect of Prevalent Carbazole Homocoupling Defects on the Photovoltaic Performance of PCDTBT:PC71BM Solar Cells

The photophysical properties and solar cell performance of the classical donor–acceptor copolymer PCDTBT (poly(N‐9′‐heptadecanyl‐2,7‐carbazole‐alt ‐5,5‐(4′,7′‐di‐2‐thienyl‐2′,1′,3′‐benzothiadiazole))) in relation to unintentionally formed main chain defects are investigated. Carbazole–carbazole homocouplings (Cbz hc) are found to significant extent in PCDTBT made with a variety of Suzuki polycondensation conditions. Cbz hc vary between 0 and 8 mol% depending on the synthetic protocol used, and are quantified by detailed nuclear magnetic resonance spectroscopy including model compounds, which allows to establish a calibration curve from optical spectroscopy. The results are corroborated by extended time‐dependent density functional theory investigations on the structural, electronic, and optical properties of regularly alternating and homocoupled chains. The photovoltaic properties of PCDTBT:fullerene blend solar cells significantly depend on the Cbz hc content for constant molecular weight, whereby an increasing amount of Cbz hc leads to strongly decreased short circuit currents J_SC. With increasing Cbz hc content, J_SC decreases more strongly than the intensity of the low energy absorption band, suggesting that small losses in absorption cannot explain the decrease in J_SC alone, rather than combined effects of a more localized LUMO level on the TBT unit and lower hole mobilities found in highly defective samples. Homocoupling‐free PCDTBT with optimized molecular weight yields the highest efficiency up to 7.2% without extensive optimization.     

Osmotic Dehydration of Liposomal Dispersions: Influence of Particle Size and Electrostatic Deposition of Cold Water Fish Skin Gelatin

Liposomes are a promising delivery system for bioactives in food and nutraceuticals. Their practical application is limited by their physical and chemical instability caused by extrinsic factors. The physical stability of liposomes of three different sizes coated with cold water fish skin gelatin was assessed during osmotic dehydration at 2, 21 and 70 °C. Soy lecithin was used to prepare 1 % liposomal dispersions. The size distribution was controlled with high pressure homogenization (1500 bar) and extrusion through polycarbonate membrane (3 and 0.8 μm). Fish gelatin was adsorbed to the interface to make secondary liposomes. Liposomal dispersions were osmotically dehydrated while monitoring the relative weight, size and rheological properties. The primary liposomes had an initial mean volume diameter (d_4,3) of 0.09, 0.40 and 2.7 μm and a ζ-potential of ?55 mV. Secondary liposomes were 0.11, 0.45 and 3.4 μm with a ζ-potential of 25 mV. The size of liposomes influenced the stability of liposomes, with the smallest liposomes being stable for 30 min, corresponding to 80 % of the initial weight, while the larger liposomes were already aggregated. Secondary liposomes were stable to 120 min for the smaller liposomes and to 150 min for the largest liposomes corresponding to 40 % of the initial weight. Stability increased during dehydration at 2 °C. Coating the liposomes increased the physical stability of the liposomal dispersions at all temperatures. The results show that cold water fish skin gelatin is a viable option to coat liposomes of a wide size range.     

Objectively Measured Walking Duration and Sedentary Behaviour and Four-Year Mortality in Older People

BackgroundPhysical activity is an important component of health. Recommendations based on sensor measurements are sparse in older people. The aim of this study was to analyse the effect of objectively measured walking and sedentary duration on four-year mortality in community-dwelling older people.MethodsBetween March 2009 and April 2010, physical activity of 1271 participants (≥65 years, 56.4% men) from Southern Germany was measured over one week using a thigh-worn uni-axial accelerometer (activPAL; PAL Technologies, Glasgow, Scotland). Mortality was assessed during a four-year follow-up. Cox-proportional-hazards models were used to estimate the associations between walking (including low to high intensity) and sedentary duration with mortality. Models were adjusted for age and sex, additional epidemiological variables, and selected biomarkers.ResultsAn inverse relationship between walking duration and mortality with a minimum risk for the 3rd quartile (102.2 to128.4 minutes walking daily) was found even after multivariate adjustment with HRs for quartiles 2 to 4 compared to quartile 1 of 0.45 (95%-CI: 0.26; 0.76), 0.18 (95%-CI: 0.08; 0.41), 0.39 (95%-CI: 0.19; 0.78), respectively. For sedentary duration an age- and sex-adjusted increased mortality risk was observed for the 4th quartile (daily sedentary duration ≥1137.2 min.) (HR 2.05, 95%-CI: 1.13; 3.73), which diminished, however, after full adjustment (HR 1.63, 95%-CI: 0.88; 3.02). Furthermore, our results suggest effect modification between walking and sedentary duration, such that in people with low walking duration a high sedentary duration was noted as an independent factor for increased mortality.ConclusionsIn summary, walking duration was clearly associated with four-year overall mortality in community-dwelling older people.     

Development of a core outcome set for clinical trials in rosacea: study protocol for a systematic review of the literature and identification of a core outcome set using a Delphi survey

BackgroundRosacea is a chronic inflammatory disorder affecting millions of individuals worldwide. Diagnosis is based on signs and symptoms with management and treatment aimed to suppress inflammatory lesions, erythema, and telangiectasia. While many clinical trials of rosacea exist, the lack of consensus in outcome reporting across all trials poses a concern. Proper evaluation and comparison of treatment modalities is challenging. In order to address the inconsistencies present, this project aims to determine a core set of outcomes which should be evaluated in all clinical trials of rosacea.Methods/designThis project will utilize a methodology similar to previous core outcome set research. A long list of outcomes will be extracted over four phases: (1) systematic literature review, (2) patient interviews, (3) other published sources, and (4) stakeholder involvement. Potential outcomes will be examined by the Steering Committee to provide further insight. The Delphi process will then be performed to prioritize and condense the list of outcomes generated. Two homogenous groups of physicians and patients will participate in two consecutive rounds of Delphi surveys. A consensus meeting, composed of physicians, patients, and stakeholders, will be conducted after the Delphi exercise to further select outcomes, taking into account participant scores. By the end of the meeting, members will vote and decide on a final recommended set of core outcomes. For the duration of the study, we will be in collaboration with both the Core Outcome Measures in Effectiveness Trials (COMET) and Cochrane Skin Group - Core Outcome Set Initiative (CSG-COUSIN).DiscussionThis study aims to develop a core outcome set to guide assessment in clinical trials of rosacea. The end-goal is to improve the reliability and consistency of outcome reporting, thereby allowing sufficient evaluation of treatment effectiveness and patient satisfaction.

Electronic supplementary material

The online version of this article (doi:10.1186/s13063-016-1554-3) contains supplementary material, which is available to authorized users.     

Salt‐enhanced cultivation as a morphology engineering tool for filamentous actinomycetes: Increased production of labyrinthopeptin A1 in Actinomadura namibiensis

Salt‐enhanced cultivation as a morphology engineering tool for the filamentous actinomycete Actinomadura namibiensis was evaluated in 500‐mL shaking flasks (working volume 100 mL) with the aim of increasing the concentration of the pharmaceutically interesting peptide labyrinthopeptin A1. Among the inorganic salts added to a complex production medium, the addition of (NH₄)₂SO₄ led to the highest amount of labyrinthopeptin A1 production. By using 50 mM (NH₄)₂SO₄, the labyrinthopeptin A1 concentration increased up to sevenfold compared to the non‐supplemented control, resulting in 325 mg L^?1 labyrinthopeptin A1 after 10 days of cultivation. The performance of other ammonium‐ and sulfate‐containing salts (e.g., NH₄Cl, K₂SO₄) was much lower than the performance of (NH₄)₂SO₄. A positive correlation between the uptake of glycerol as one of the main carbon sources and nongrowth‐associated labyrinthopeptin productivity was found. The change in the cell morphology of A. namibiensis in conjunction with increased osmolality by the addition of 50 mM (NH₄)₂SO₄, was quantified by image analysis. A. namibiensis always developed a heterogeneous morphology with pellets and loose mycelia present simultaneously. In contrast to the non‐supplemented control, the morphology of (NH₄)₂SO₄‐supplemented cultures was characterized by smaller and circular pellets that were more stable against disintegration in the stationary production phase.     

Scale-up from shake flasks to fermenters in batch and continuous mode with Corynebacterium glutamicum on lactic acid based on oxygen transfer and pH

Scale-up from shake flasks to fermenters has been hampered by the lack of knowledge concerning the influence of operating conditions on mass transfer, hydromechanics, and power input. However, in recent years the properties of shake flasks have been described with empirical models. A practical scale-up strategy for everyday use is introduced for the scale-up of aerobic cultures from shake flasks to fermenters in batch and continuous mode. The strategy is based on empirical correlations of the volumetric mass transfer coefficient (k(L) a) and the pH. The accuracy of the empirical k(L) a correlations and the assumptions required to use these correlations for an arbitrary biological medium are discussed. To determine the optimal pH of the culture medium a simple laboratory method based on titration curves of the medium and a mechanistic pH model, which is solely based on the medium composition, is applied. The effectiveness of the scale-up strategy is demonstrated by comparing the behavior of Corynebacterium glutamicum on lactic acid in shake flasks and fermenters in batch and continuous mode. The maximum growth rate (micro(max) = 0.32 h(-1)) and the oxygen substrate coefficient (Y O2 /S= 0.0174 mol/l) of C. glutamicum on lactic acid were equal for shake flask, fermenter, batch, and continuous cultures. The biomass substrate yield was independent of the scale, but was lower in batch cultures (Y(X/S) = 0.36 g/g) than in continuous cultures (Y(X/S) = 0.45 g/g). The experimental data (biomass, respiration, pH) could be described with a simple biological model combined with a mechanistic pH model.