A study on malt modification,used as a tool to reduce levels of beer hordeins

Storage proteins from barley, wheat and rye are toxic to gluten sensitive consumers. These consumers include those suffering from coeliac disease, which account for up to 1% of the global population, and non‐coeliac gluten sensitivity that may affect even greater numbers of the population. Codex Alimentarius has published guidelines and limits of gluten in gluten‐free foods, which are applied in Europe, and similar guidelines apply in the rest of the world. The storage proteins present in barley are hordeins. These proteins are broken down and used by the plant as a source of amino acids during germination and growth of the barley embryo. The objective of this study was to extend the germination stage of the malting process and look at the effect on beer hordeins. Standard MEBAK methods were used to develop an extended malting process and produce three different malts, germinated for 3, 5 or 7 days. The quality of malt was assessed and model beers were produced from each malt to test the effect of modification on levels of beer hordeins. Malt germinated for 7 days produced beer 18 mg/kg hordeins corresponding to a reduction of 44% compared with the beer made from malt germinated for 3 days characterized by a hordein content equal to 32 mg/kg. The malting loss was increased during the 7 days of germination but otherwise all malts were of high quality. The results showed that malting conditions have a significant impact on beer hordeins. Copyright © 2018 The Institute of Brewing & Distilling     

Prediction of Response to Cisplatin-Based Neoadjuvant Chemotherapy of Muscle-Invasive Bladder Cancer Patients by Molecular Subtyping including KRT and FGFR Target Gene Assessment

Patients with muscle-invasive urothelial carcinoma achieving pathological complete response (pCR) upon neoadjuvant chemotherapy (NAC) have improved prognosis. Molecular subtypes of bladder cancer differ markedly regarding sensitivity to cisplatin-based chemotherapy and harbor FGFR treatment targets to various content. The objective of the present study was to evaluate whether preoperative assessment of molecular subtype as well as FGFR target gene expression is predictive for therapeutic outcome—rate of ypT0 status—to justify subsequent prospective validation within the “BladderBRIDGister”. Formalin-fixed paraffin-embedded (FFPE) tissue specimens from transurethral bladder tumor resections (TUR) prior to neoadjuvant chemotherapy and corresponding radical cystectomy samples after chemotherapy of 36 patients were retrospectively collected. RNA from FFPE tissues were extracted by commercial kits, Relative gene expression of subtyping markers (e.g., KRT5, KRT20) and target genes (FGFR1, FGFR3) was analyzed by standardized RT-qPCR systems (STRATIFYER Molecular Pathology GmbH, Cologne). Spearman correlation, Kruskal–Wallis, Mann–Whitney and sensitivity/specificity tests were performed by JMP 9.0.0 (SAS software). The neoadjuvant cohort consisted of 36 patients (median age: 69, male 83% vs. female 17%) with 92% of patients being node-negative during radical cystectomy after 1 to 4 cycles of NAC. When comparing pretreatment with post-treatment samples, the median expression of KRT20 dropped most significantly from DCT 37.38 to 30.65, which compares with a 128-fold decrease. The reduction in gene expression was modest for other luminal marker genes (GATA3 6.8-fold, ERBB2 6.3-fold). In contrast, FGFR1 mRNA expression increased from 33.28 to 35.88 (~6.8-fold increase). Spearman correlation revealed positive association of pretreatment KRT20 mRNA levels with achieving pCR (r = 0.3072: p = 0.0684), whereas pretreatment FGFR1 mRNA was associated with resistance to chemotherapy (r = −0.6418: p < 0.0001). Hierarchical clustering identified luminal tumors of high KRT20 mRNA expression being associated with high pCR rate (10/16; 63%), while the double-negative subgroup with high FGFR1 expression did not respond with pCR (0/9; 0%). Molecular subtyping distinguishes patients with high probability of response from tumors as resistant to neoadjuvant chemotherapy. Targeting FGFR1 in less-differentiated bladder cancer subgroups may sensitize tumors for adopted treatments or subsequent chemotherapy.     

Microbeam Irradiation as a Simultaneously Integrated Boost in a Conventional Whole-Brain Radiotherapy Protocol

Microbeam radiotherapy (MRT), an experimental high-dose rate concept with spatial fractionation at the micrometre range, has shown a high therapeutic potential as well as good preservation of normal tissue function in pre-clinical studies. We investigated the suitability of MRT as a simultaneously integrated boost (SIB) in conventional whole-brain irradiation (WBRT). A 174 Gy MRT SIB was administered with an array of quasi-parallel, 50 µm wide microbeams spaced at a centre-to-centre distance of 400 µm either on the first or last day of a 5 × 4 Gy radiotherapy schedule in healthy adult C57 BL/6J mice and in F98 glioma cell cultures. The animals were observed for signs of intracranial pressure and focal neurologic signs. Colony counts were conducted in F98 glioma cell cultures. No signs of acute adverse effects were observed in any of the irradiated animals within 3 days after the last irradiation fraction. The tumoricidal effect on F98 cell in vitro was higher when the MRT boost was delivered on the first day of the irradiation course, as opposed to the last day. Therefore, the MRT SIB should be integrated into a clinical radiotherapy schedule as early as possible.     

Enhanced Magnetization in CoFe2O4 Through Hydrogen Doping

Magnetic spinel oxides have attracted extensive research interest due to their rich physics and wide range of applications. However, these materials invariably suffer suppressed magnetization, due to structural imperfections (e.g., disorder, anti-site defects, etc.). Herein, a dramatic enhanced magnetization is obtained with an increasement of 5 µ_B/u.c in CoFe₂O₄ (CFO) through ionic liquid gating induced hydrogen doping. The intercalated hydrogen ions lead to both distinct lattice expansion of ≈0.7% and notable Fe valence state reduction through electron doping, in which ≈17% Fe³⁺ is reduced into Fe²⁺. These facts collectively trigger a site-specific spin-flip on tetrahedrally coordinated Co²⁺ sites that enhances the net ferrimagnetic moment nearly to its theoretical maximum for perfect CFO.     

Tailoring Enzyme Stringency Masks the Multispecificity of a Lyngbyatoxin (Indolactam Alkaloid) Nonribosomal Peptide Synthetase

Indolactam alkaloids are activators of protein kinase C (PKC) and are of pharmacological interest for the treatment of pathologies involving PKC dysregulation. The marine cyanobacterial nonribosomal peptide synthetase (NRPS) pathway for lyngbyatoxin biosynthesis, which we previously expressed in E. coli, was studied for its amenability towards the biosynthesis of indolactam variants. Modification of culture conditions for our E. coli heterologous expression host and analysis of pathway products suggested the native lyngbyatoxin pathway NRPS does possess a degree of relaxed specificity. Site-directed mutagenesis of two positions within the adenylation domain (A-domain) substrate-binding pocket was performed, resulting in an alteration of substrate preference between valine, isoleucine, and leucine. We observed relative congruence of in vitro substrate activation by the LtxA NRPS to in vivo product formation. While there was a preference for isoleucine over leucine, the substitution of alternative tailoring domains may unveil the true in vivo effects of the mutations introduced herein.     

Investigation of molecular changes in wine during storage in refrigerated cabinets

Wine is a complex chemical mixture that is bound to change over time. Most wines are produced for consumption within months. Some premium wines are meant to be maturated for several years or even decades after bottling. The post-bottling evolution and the longevity of a wine depends on its initial chemical composition and the storage conditions. Temperature, exposure to light and the closure type are often mentioned as the most important storage influences. Especially elevated temperature is known to cause accelerated aging reactions in wine. Refrigerated wine storage cabinets promise to be the best storage option without the need of a wine cellar. They are available in different sizes and fit in every household. However, the influence of vibrations and low-interval temperature fluctuations caused by compressors are parameters that have been neglected in literature. The aim of this thesis was to investigate if vibrations and low-interval temperature fluctuations, which occur in refrigerated wine storage cabinets, have an influence on the post-bottling evolution of a wine. The influence of both parameters was studied separately from each other. The impact of vibration on oxidation and gas uptake from the headspace of a wine bottle into the wine was investigated using a model wine with saturated O2 and different headspace volumes. The study revealed that vibration promotes the dissolution of O2 from the headspace of bottle into the wine resulting in a faster SO2 consumption. Furthermore, it was shown that horizontal bottle position accelerated the O2 uptake significantly. It was concluded that the increased surface size between headspace and wine accelerates the O2 dissolution in wine. Also, bigger headspace volumes caused an accelerated O2 uptake into the wine. An experiment without any headspace volume revealed that the factors vibration and bottle position did not accelerate the O2 consumption in wine. This proves that vibration and bottle position accelerate only the dissolution of O2 in wine, but not the chemical reaction of O2 with wine constituents. The influence of vibration on the volatile profile of wine was investigated using Riesling sparkling and still wines sealed with different closures that were subjected to vibration for six months. Vibration caused no CO2 losses, SO2 and color changes in all wines indicating that vibration caused by compressors has no impact on the gas permeability of the used closures. However, vibration affected the volatile profile of sparkling wine and Riesling still wine sealed with a screw cap. Similar to the model wine study described earlier, it was shown that the equilibrium of volatile substances between the wine and the headspace in a bottle was influenced by vibration. The gas-liquid-equilibrium of some volatile compounds was shifted towards wine, while others were shifted towards headspace. As a result of this, the concentration of volatile compounds in wine is changed. Besides this indirect influence of vibration, the results of this study also suggested that specific degradation and formation reactions of volatile compounds are directly influenced by vibration. These multiple effects of vibration most likely explain why increasing vibration intensities could not be proportionally related to the observed volatile changes. The investigation of different wine styles revealed that the impact of vibration depends strongly on the initial composition of wine, age, and packaging conditions. Especially, headspace volume, closure type and CO2 pressure are likely to influence the equilibrium of volatile substances between the wine and the headspace in a bottle. Another study investigated the impact of low-interval temperature fluctuations on the volatile profile of wine. For this purpose, a Riesling wine was stored for two years under different temperature fluctuation patterns caused by compressors. Additionally, a model wine with nine volatile substances with known concentrations was stored for eight months under the same fluctuation patterns. The low- interval temperature fluctuations were compared to the mean value of the temperature fluctuations. Chemical and sensory analysis revealed that that low-interval temperature fluctuations accelerate wine aging reactions like ester hydrolysis and monoterpene degradation. Even small temperature amplitudes showed a significant impact on wine aging. The observed effect was explained by the Arrhenius equation which states that reaction rates exponentially increase with rising temperatures. A pump effect of air through the closure was initially assumed but not observed in this study. Small deviations in wine temperature, such as those caused by door openings of a refrigerator were found to be negligible. It was concluded that low-interval temperature fluctuations can accelerate wine aging reactions. The amplitude of the temperature fluctuations should be as small as possible during bottle storage of wine. This thesis showed that both parameters, vibration, and low-interval temperature fluctuations, have been proven to influence the evolution of wine during bottle storage. Regarding storage conditions in a refrigerated wine storage cabinet, those parameters should be monitored. Wine connoisseurs should therefore consider good wine cabinets, since some manufacturers emphasize on the importance to minimize vibrations and temperature fluctuations in their devices. The development of technology should be advanced to reduce both vibration and temperature fluctuations in refrigerated wine storage cabinets. Future research should focus on specific wine compounds in model systems and realistic vibration conditions to reveal the relationship between vibration intensities and reaction rates. The impact of low-interval temperature fluctuations on wine compositional changes should be investigated considering horizontal and vertical bottle positions. The calculated acceleration factors due to temperature fluctuations have to be verified by isotherm storage conditions at higher temperatures.