High‐molecular‐weight poly(Gly‐Val‐Gly‐Val‐Pro) synthesis through microwave irradiation

In this study, we synthesized a polypeptide from its pentapeptide unit using microwave irradiation. Effective methods for polypeptide synthesis from unit peptides have not been reported. Here, we used a key elastin peptide, H‐GlyValGlyValPro‐OH (GVGVP), as the monomer peptide. It is difficult to obtain poly(Gly‐Val‐Gly‐Val‐Pro) (poly(GVGVP)) from the pentapeptide unit of elastin, GVGVP, via polycondensation. Poly(GVGVP) prepared from genetically recombinant Escherichia coli is a well‐known temperature‐sensitive polypeptide, and this temperature sensitivity is known as the lower critical solution temperature. When microwave irradiation was performed in the presence of various additives, the pentapeptide (GVGVP) polycondensation reaction proceeded smoothly, resulting in a product with a high molecular weight in a relatively good yield. The reaction conditions, like microwave irradiation, coupling agents, and solvents, were optimized to increase the reaction efficiency. The product exhibited a molecular weight greater than Mr 7000. Further, the product could be synthesized on a gram scale. The synthesized polypeptide exhibited a temperature sensitivity that was similar to that of poly(GVGVP) prepared from genetically recombinant E. coli. Therefore, this technique offers a facile and quick approach to prepare polypeptides in large amounts. Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd.     

Variability of n‐Alkanes and Nonacosan‐10‐ol in Natural Populations of Picea omorika

This is the first report of population variability of the contents of n‐alkanes and nonacosan‐10‐ol in the needle epicuticular waxes of Serbian spruce (Picea omorika). The hexane extracts of needle samples originated from three natural populations in Serbia (Vranjak, Zmajeva?ki potok, and Mile?evka Canyon) were investigated by GC and GC/MS analyses. The amount of nonacosan‐10‐ol varied individually from 50.05 to 74.42% (65.74% in average), but the differences between the three investigated populations were not statistically confirmed. The results exhibited variability of the composition of n‐alkanes in the epicuticular waxes with their size ranging from C₁₈ to C₃₅. The most abundant n‐alkanes were C₂₉, C₃₁, and C₂₇ (35.22, 13.77, and 12.28% in average, resp.). The carbon preference index of all the n‐alkanes (CPI_total) of the P. omorika populations (average of populations I–III) ranged from 3.3 to 11.5 (mean of 5.9), while the average chain length (ACL) ranged from 26.6 to 29.2. The principal component and cluster analyses of the contents of nine n‐alkanes showed the greatest difference for the population growing in the Mile?evka Canyon. The obtained results were compared with previous literature data given for other Picea species, and this comparison was briefly discussed.     

ALS‐associated fused in sarcoma (FUS) mutations disrupt Transportin‐mediated nuclear import

Mutations in fused in sarcoma (FUS) are a cause of familial amyotrophic lateral sclerosis (fALS). Patients carrying point mutations in the C‐terminus of FUS show neuronal cytoplasmic FUS‐positive inclusions, whereas in healthy controls, FUS is predominantly nuclear. Cytoplasmic FUS inclusions have also been identified in a subset of frontotemporal lobar degeneration (FTLD‐FUS). We show that a non‐classical PY nuclear localization signal (NLS) in the C‐terminus of FUS is necessary for nuclear import. The majority of fALS‐associated mutations occur within the NLS and impair nuclear import to a degree that correlates with the age of disease onset. This presents the first case of disease‐causing mutations within a PY‐NLS. Nuclear import of FUS is dependent on Transportin, and interference with this transport pathway leads to cytoplasmic redistribution and recruitment of FUS into stress granules. Moreover, proteins known to be stress granule markers co‐deposit with inclusions in fALS and FTLD‐FUS patients, implicating stress granule formation in the pathogenesis of these diseases. We propose that two pathological hits, namely nuclear import defects and cellular stress, are involved in the pathogenesis of FUS‐opathies.     

Sodium salicylate modulates inflammatory responses through AMP‐activated protein kinase activation in LPS‐stimulated THP‐1 cells

Sodium salicylate (NaSal) is a nonsteroidal anti‐inflammatory drug. The putative mechanisms for NaSal's pharmacologic actions include the inhibition of cyclooxygenases, platelet‐derived thromboxane A2, and NF‐κB signaling. Recent studies demonstrated that salicylate could activate AMP‐activated protein kinase (AMPK), an energy sensor that maintains the balance between ATP production and consumption. The anti‐inflammatory action of AMPK has been reported to be mediated by promoting mitochondrial biogenesis and fatty acid oxidation. However, the exact signals responsible for salicylate‐mediated inflammation through AMPK are not well‐understood. In the current study, we examined the potential effects of NaSal on inflammation‐like responses of THP‐1 monocytes to lipopolysaccharide (LPS) challenge. THP‐1 cells were stimulated with or without 10 ug/mL LPS for 24 h in the presence or absence of 5 mM NaSal. Apoptosis was measured by flow cytometry using Annexin V/PI staining and by Western blotting for the Bcl‐2 anti‐apoptotic protein. Cell proliferation was detected by EdU incorporation and by Western blot analysis for proliferating cell nuclear antigen (PCNA). Secretion of pro‐inflammatory cytokines (TNF‐α, IL‐1β, IL‐6) was determined by enzyme‐linked immunosorbent assay (ELISA). We observed that the activation of AMPK by NaSal was accompanied by induction of apoptosis, inhibition of cell proliferation, and increasing secretion of TNF‐α and IL‐1β. These effects were reversed by Compound C, an inhibitor of AMPK. In addition, NaSal/AMPK activation inhibited LPS‐induced STAT3 phosphorylation, which was reversed by Compound C treatment. We conclude that AMPK activation is important for NaSal‐mediated inflammation by inducing apoptosis, reducing cell proliferation, inhibiting STAT3 activity, and producing TNF‐α and IL‐1β.     

Large Power Amplification in Magneto‐Mechano‐Electric Harvesters through Distributed Forcing

Energy harvesting from extremely low frequency magnetic fields using magneto‐mechano‐electric (MME) harvesters enables wireless power transfer for operating Internet of Things (IoT) devices. The MME harvesters are designed to resonate at a fixed frequency by absorbing AC magnetic fields through a composite cantilever comprising of piezoelectric and magnetostrictive materials, and a permanent magnetic tip mass. However, this harvester architecture limits power generation because volume of the magnetic end mass is closely coupled with the resonance frequency of the device structure. Here, a method is demonstrated for maintaining the resonance frequency of the MME harvesters under all operating conditions (e.g., 60 Hz, standard frequency of electricity in many countries) while simultaneously enhancing the output power generation. By distributing the magnetic mass over the beam, the output power of the harvester is significantly enhanced at a constant resonance frequency. The MME harvester with distributed forcing shows 280% improvement in the power generation compared with a traditional architecture. The generated power is shown to be sufficient to power eight different onboard sensors with wireless data transmission integrated on a drone. These results demonstrate the promise of MME energy harvesters for powering wireless communication and IoT sensors.     

LC‐MS‐based metabolic characterization of high monoclonal antibody‐producing Chinese hamster ovary cells

The selection of suitable mammalian cell lines with high specific productivities is a crucial aspect of large‐scale recombinant protein production. This study utilizes a metabolomics approach to elucidate the key characteristics of Chinese hamster ovary (CHO) cells with high monoclonal antibody productivities (q_mAb). Liquid chromatography‐mass spectrometry (LC‐MS)‐based intracellular metabolite profiles of eight single cell clones with high and low q_mAb were obtained at the mid‐exponential phase during shake flask batch cultures. Orthogonal projection to latent structures discriminant analysis (OPLS‐DA) subsequently revealed key differences between the high and low q_mAb clones, as indicated by the variable importance for projection (VIP) scores. The mass peaks were further examined for their potential association with q_mAb across all clones using Pearson's correlation analysis. Lastly, the identities of metabolites with high VIP and correlation scores were confirmed by comparison with standards through LC‐MS‐MS. A total of seven metabolites were identified—NADH, FAD, reduced and oxidized glutathione, and three activated sugar precursors. These metabolites are involved in key cellular pathways of citric acid cycle, oxidative phosphorylation, glutathione metabolism, and protein glycosylation. To our knowledge, this is the first study to identify metabolites that are associated closely with q_mAb. The results suggest that the high producers had elevated levels of specific metabolites to better regulate their redox status. This is likely to facilitate the generation of energy and activated sugar precursors to meet the demands of producing more glycosylated recombinant monoclonal antibodies. Biotechnol. Bioeng. 2012; 109: 3103–3111. © 2012 Wiley Periodicals, Inc.     

Chemodiversity of Nonacosan‐10‐ol and n‐Alkanes in the Needle Wax of Pinus heldreichii

This is the first report of individual variability and population diversity of the contents of nonacosan‐10‐ol and n‐alkanes in the needle cuticular waxes of Bosnian pines originated from Montenegro, regarded as Pinus heldreichii var. leucodermis, and from Serbia, regarded as P. heldreichii var. pan?i?i. The amount of nonacosan‐10‐ol varied individually from 27.4 to 73.2% (55.5% in average), but differences between the four investigated populations were not statistically confirmed. The size of the n‐alkanes ranged from C₁₈ to C₃₃. The most abundant n‐alkanes were C₂₃, C₂₇, and C₂₅ (12.2, 11.2, and 10.8% in average, resp.). The carbon preference index (CPI) of the n‐alkanes ranged from 0.8 to 3.1 (1.6 in average), while the average chain length (ACL) ranged from 20.9 to 26.5 (24.4 in average). Long‐chain and mid‐chain n‐alkanes prevailed (49.6 and 37.9% in average, resp.). It was also found that the populations of P. heldreichii var. leucodermis had predominantly a narrower range of n‐alkanes (C₁₈? C₃₁) than the trees of the variety pan?i?i (C₁₈? C₃₃). Differences between the varieties were also significant for most of the other characteristics of the n‐alkane pattern (e.g., most abundant n‐alkanes, CPI, ACL, and relative proportion of short‐, mid‐, and long‐chain n‐alkanes). The principle component and cluster analyses of eleven n‐alkanes confirmed the significant diversity of these two varieties.     

Spiro‐Phenylpyrazole‐9,9′‐Thioxanthene Analogues as Hole‐Transporting Materials for Efficient Planar Perovskite Solar Cells

Perovskite solar cells have emerged as a promising technique for low‐cost, light weight, and highly efficient photovoltaics. However, they still largely rely on 2,2′,7,7′‐tetrakis(N,N‐di‐p‐methoxyphenylamine)‐9,9′‐spirobifluorene (Spiro‐OMeTAD) to serve as hole‐transporting materials (HTMs). Here, a series of HTMs with small molecular weight is designed, which are constructed on a spiro core involving phenylpyrazole and a second heteroaromatics, i.e., xanthene (O atom), thioxanthene (S atom), and acridine (N atom). Through varying from phenylpyrazole substituted xanthene ( PPyra‐XA ), thioxanthene ( PPyra‐TXA ), to acridine ( PPyra‐ACD ), their optical and electrochemical properties, hole mobilities, and the photovoltaic performance are optimized. As a consequence, PPyra‐TXA based device exhibits the highest power conversion efficiency (PCE) of 18.06%, outperforming that of Spiro‐OMeTAD (16.15%), which could be attributed to the enhancement of hole mobility exerted by the thioxanthene. In addition, the dopant‐free device shows PCE of 11.7%. These results open a new direction for designing spiro‐HTMs by simple modification of chemical structures.     

Ameliorative Effects of Curcumin on Fibrinogen‐Like Protein‐2 Gene Expression,Some Oxido‐Inflammatory and Apoptotic Markers in a Rat Model of l‐Arginine‐Induced Acute Pancreatitis

The aim of the study was to investigate the ameliorative effects of curcumin on fibrinogen like protein‐2 (fgl‐2), some oxido‐inflammatory and apoptotic markers in rat‐induced acute pancreatitis (AP). Seventy‐five albino rats were divided into control group, l ‐arginine (l ‐Arg)‐induced AP group, curcumin pre‐treated group before AP induction, curcumin post‐treated group after AP induction, and curcumin injected group only. AP group showed severe necrotizing pancreatitis confirmed by histopathological changes and elevations in serum amylase and lipase activities, levels of epithelial neutrophil‐activating peptide 78, tissue content of protein carbonyls, levels of tumor necrosis factor α, and caspase‐3 as well as myeloperoxidase activity. Significant elevation in pancreatic fgl‐2 mRNA expression was detected in AP group. Improvement of all parameters was detected with increase of caspase‐3 in both curcumin‐treated groups that confirmed curcumin ameliorative effects against AP through induction of apoptosis and inhibition of micro‐thrombosis, inflammation, and oxidative stress.     

Validated spectrofluorimetric method for the determination of carbamazepine in pharmaceutical dosage forms after reaction with 4‐chloro‐7‐‐nitrobenzo‐2‐oxa‐1,3‐diazole (NBD‐Cl)

A sensitive and simple spectrofluorimetric method has been developed and validated for the determination of the anti‐epileptic drug carbamazepine (CBZ) in its dosage forms. The method was based on a nucleophilic substitution reaction of CBZ with 4‐chloro‐7‐nitrobenzo‐2‐ oxa‐1,3‐diazole (NBD‐Cl) in borate buffer (pH 9) to form a highly fluorescent derivative that was measured at 530 nm after excitation at 460 nm. Factors affecting the formation of the reaction product were studied and optimized, and the reaction mechanism was postulated. The fluorescence–concentration plot is rectilinear over the range of 0.6–8 µg/mL with limit of detection of 0.06 µg/mL and limit of quantitation of 0.19 µg/mL. The method was applied to the analysis of commercial tablets and the results were in good agreement with those obtained using the reference method. Validation of the analytical procedures was evaluated according to ICH guidelines. Copyright © 2015 John Wiley & Sons, Ltd.     

Broad‐scale vegetation‐environment relationships in Eurasian high‐latitude areas

Question: How is tundra vegetation related to climatic, soil chemical, geological variables and grazing across a very large section of the Eurasian arctic area? We were particularly interested in broad‐scale vegetation‐environment relationships and how well do the patterns conform to climate‐vegetation schemes. Material and Methods: We sampled vegetation in 1132 plots from 16 sites from different parts of the Eurasian tundra. Clustering and ordination techniques were used for analysing compositional patterns. Vegetation‐environment relationships were analysed by fitting of environmental vectors and smooth surfaces onto non‐metric multidimensional scaling scattergrams. Results: Dominant vegetation differentiation was associated with a complex set of environmental variables. A general trend differentiated cold and continental areas from relatively warm and weakly continental areas, and several soil chemical and physical variables were associated with this broad‐scaled differentiation. Especially soil chemical variables related to soil acidity (pH, Ca) showed linear relationships with the dominant vegetation gradient. This was closely related to increasing cryoperturbation, decreasing precipitation and cooler conditions. Remarkable differences among relatively adjacent sites suggest that local factors such as geological properties and lemming grazing may strongly drive vegetation differentiation. Conclusions: Vegetation differentiation in tundra areas conforms to a major ecocline underlain by a complex set of environmental gradients, where precipitation, thermal conditions and soil chemical and physical processes are coupled. However, local factors such as bedrock conditions and lemming grazing may cause marked deviations from the general climate‐vegetation models. Overall, soil chemical factors (pH, Ca) turned out to have linear relationship with the broad‐scale differentiation of arctic vegetation.     

Global Rare Earth In‐Use Stocks in NdFeB Permanent Magnets

The rare earth elements are indispensible in modern technology, especially in the applications of permanent magnets. Very little quantitative information is available on rare earth elements used in permanent magnets, however. This study looks back to 1983, when neodymium‐iron‐boron (NdFeB) permanent magnets were first manufactured, and reaches to 2007, when the market of permanent magnets was well developed. We draw on the historical data on permanent magnets from China, Japan, the United States, and Europe to provide the first estimates of global in‐use stocks for four rare earth elements—praseodymium (Pr), neodymium (Nd), terbium (Tb), and dysprosium (Dy)—in NdFeB permanent magnets. In‐use stocks amount to 62.6 gigagrams (Gg) Nd, 15.7 Gg Pr, 15.7 Gg Dy, and 3.1 Gg Tb; these stocks, if efficiently recycled, could provide a valuable supplement to geological stocks as they are almost four times the 2007 annual extraction rate of the individual elements.