A multi-level analytical approach for detection and visualization of intracellular NO production and nitrosation events using diaminofluoresceins

Diaminofluoresceins are widely used probes for detection and intracellular localization of NO formation in cultured/isolated cells and intact tissues. The fluorinated derivative, 4-amino-5-methylamino-2′,7′-difluorofluorescein (DAF-FM), has gained increasing popularity in recent years due to its improved NO-sensitivity, pH-stability, and resistance to photo-bleaching compared to the first-generation compound, DAF-2. Detection of NO production by either reagent relies on conversion of the parent compound into a fluorescent triazole, DAF-FM-T and DAF-2-T, respectively. While this reaction is specific for NO and/or reactive nitrosating species, it is also affected by the presence of oxidants/antioxidants. Moreover, the reaction with other molecules can lead to the formation of fluorescent products other than the expected triazole. Thus additional controls and structural confirmation of the reaction products are essential. Using human red blood cells as an exemplary cellular system we here describe robust protocols for the analysis of intracellular DAF-FM-T formation using an array of fluorescence-based methods (laser-scanning fluorescence microscopy, flow cytometry and fluorimetry) and analytical separation techniques (reversed-phase HPLC and LC-MS/MS). When used in combination, these assays afford unequivocal identification of the fluorescent signal as being derived from NO and are applicable to most other cellular systems without or with only minor modifications.

Sensitivity analysis of an ocean carbon cycle model in the North Atlantic: an investigation of parameters affecting the air-sea CO2 flux, primary production and export of detritus

The sensitivity of the biological parameters in a nutrient-phytoplankton-zooplankton-detritus (NPZD) model in the calculation of the air-sea CO2 flux, primary production and detrital export is analysed. We explore the effect on these outputs of variation in the values of the twenty parameters that control ocean ecosystem growth in a 1-D formulation of the UK Met Office HadOCC NPZD model used in GCMs. We use and compare the results from one-at-a-time and all-at-a-time perturbations performed at three sites in the EuroSITES European Ocean Observatory Network: the Central Irminger Sea (60° N 40° W), the Porcupine Abyssal Plain (49° N 16° W) and the European Station for Time series in the Ocean Canary Islands (29° N 15° W). Reasonable changes to the values of key parameters are shown to have a large effect on the calculation of the air-sea CO2 flux, primary production, and export of biological detritus to the deep ocean. Changes in the values of key parameters have a greater effect in more productive regions than in less productive areas. The most sensitive parameters are generally found to be those controlling well-established ocean ecosystem parameterisations widely used in many NPZD-type models. The air-sea CO2 flux is most influenced by variation in the parameters that control phytoplankton growth, detrital sinking and carbonate production by phytoplankton (the rain ratio). Primary production is most sensitive to the parameters that define the shape of the photosynthesis-irradiance curve. Export production is most sensitive to the parameters that control the rate of detrital sinking and the remineralisation of detritus.

Creep and recovery of magnetorheological fluids: experiments and simulations

A direct comparative study on the creep-recovery behavior of conventional MR fluids is carried out using magnetorheometry and particle-level simulations. Two particle concentrations are investigated (ϕ=0.05 and 0.30) at two different magnetic field strengths (53 kA•m-1 and 173 kA•m-1) in order to match the yield stresses developed in both systems for easier comparison. Simulations are mostly started with random initial structures with some additional tests of using preassembled single chains in the low concentration case. Experimental and simulation data are in good qualitative agreement. The results demonstrate three regions in the creep curves: i) In the initial viscoelastic region, the chain-like (at ϕ=0.05) or percolated three-dimensional network (at ϕ=0.30) structures fill up the gap and the average cluster size remains constant; ii) Above a critical strain of 10 %, in the retardation region, these structures begin to break and rearrange under shear. At large enough imposed stress values, they transform into thin sheet-like or thick lamellar structures, depending on the particle concentration; iii) Finally in the case of larger strain values either the viscosity diverges (at low stress values) or reaches a constant low value (at high stress values), showing a clear bifurcation behavior. For stresses below the bifurcation point the MR fluid is capable to recover the strain by a certain fraction. However, no recovery is observed for large stress values.

The spaces of the Wednesday Play (BBC TV 1964–1970): production, technology and style

The BBC television drama anthology The Wednesday Play, broadcast from 1964-70 on the BBC1 channel, was high-profile and often controversial in its time and has since been central to accounts of British television’s ‘golden age’. This article demonstrates that production technologies and methods were more diverse at that time than is now acknowledged, and that The Wednesday Play dramas drew both approving but also very critical responses from contemporary viewers and professional reviewers. This article analyses the ways that the physical spaces of production for different dramas in the series, and the different technologies of shooting and recording that were adopted in these production spaces, are associated with but do not determine aesthetic style. The adoption of single-camera location filming rather than the established production method of multi-camera studio videotaping in some of the dramas in the series has been important to The Wednesday Play’s significance, but each production method was used in different ways. The dramas drew their dramatic forms and aesthetic emphases from both theatre and cinema, as well as connecting with debates about the nature of drama for television. Institutional and regulatory frameworks such as control over staff working away from base, budgetary considerations and union agreements also impacted on decisions about how programmes were made. The article makes use of records from the BBC Written Archives Centre, as well as published scholarship. By placing The Wednesday Play in a range of overlapping historical contexts, its identity can be understood as transitional, differentiated and contested.

Identification and quantification of glucosinolate and flavonol compounds in rocket salad (Eruca sativa, Eruca vesicaria and Diplotaxis tenuifolia) by LC–MS: Highlighting the potential for improving nutritional value of rocket crops

Liquid Chromatography Mass Spectrometry (LC-MS) was used to obtain glucosinolate and flavonol content for 35 rocket accessions and commercial varieties. 13 glucosinolates and 11 flavonol compounds were identified. Semi-quantitative methods were used to estimate concentrations of both groups of compounds. Minor glucosinolate composition was found to be different between accessions; concentrations varied significantly. Flavonols showed differentiation between genera, with Diplotaxis accumulating quercetin glucosides and Eruca accumulating kaempferol glucosides. Several compounds were detected in each genus that have only previously been reported in the other. We highlight how knowledge of phytochemical content and concentration can be used to breed new, nutritionally superior varieties. We also demonstrate the effects of controlled environment conditions on the accumulations of glucosinolates and flavonols and explore the reasons for differences with previous studies. We stress the importance of consistent experimental design between research groups to effectively compare and contrast results.

Production and purification of xylooligosaccharides from oil palm empty fruit bunch fibre by a non-isothermal process

Oil palm empty fruit bunches (OPEFB) fibre, a by-product generated from non-woody, tropical perennial oil palm crop was evaluated for xylooligosaccharides (XOS) production. Samples of OPEFB fibre were subjected to non-isothermal autohydrolysis treatment using a temperature range from 150 to 220 °C. The highest XOS concentration, 17.6 g/L which relayed from solubilisation of 63 g/100 g xylan was achieved at 210 °C and there was a minimum amount of xylose and furfural being produced. The chromatographic purification which was undertaken to purify the oligosaccharide-rich liquor resulted in a product with 74–78% purity, of which 83–85% was XOS with degree of polymerisation (DP) between 5 and 40.

Valorisation of side streams from wheat milling and confectionery industries for consolidated production and extraction of microbial lipids

Crude enzymes produced via solid state fermentation (SSF) using wheat milling by-products have been employed for both fermentation media production using flour-rich waste (FRW) streams and lysis of Rhodosporidium toruloides yeast cells. Filter sterilization of crude hydrolysates was more beneficial than heat sterilization regarding yeast growth and microbial oil production. The initial carbon to free amino nitrogen ratio of crude hydrolysates was optimized (80.2 g/g) in fed-batch cultures of R. toruloides leading to a total dry weight of 61.2 g/L with microbial oil content of 61.8 % (w/w). Employing a feeding strategy where the glucose concentration was maintained in the range of 12.2 – 17.6 g/L led to the highest productivity (0.32 g/L∙h). The crude enzymes produced by SSF were utilised for yeast cell treatment leading to simultaneous release of around 80% of total lipids in the broth and production of a hydrolysate suitable as yeast extract replacement.

Effect of impurities in biodiesel-derived waste glycerol on the performance and feasibility of biotechnological processes

The rapid development of biodiesel production technology has led to the generation of tremendous quantities of glycerol wastes, as the main by-product of the process. Stoichiometrically, it has been calculated that for every 100 kg of biodiesel, 10 kg of glycerol are produced. Based on the technology imposed by various biodiesel plants, glycerol wastes may contain numerous kinds of impurities such as methanol, salts, soaps, heavy metals and residual fatty acids. This fact often renders biodiesel-derived glycerol unprofitable for further purification. Therefore, the utilization of crude glycerol though biotechnological means represents a promising alternative for the effective management of this industrial waste. This review summarizes the effect of various impurities-contaminants that are found in biodiesel-derived crude glycerol upon its conversion by microbial strains in biotechnological processes. Insights are given concerning the technologies that are currently applied in biodiesel production, with emphasis to the impurities that are added in the composition of crude glycerol, through each step of the production process. Moreover, extensive discussion is made in relation with the impact of the nature of impurities upon the performances of prokaryotic and eukaryotic microorganisms, during crude glycerol bioconversions into a variety of high added-value metabolic products. Finally, aspects concerning ways of crude glycerol treatment for the removal of inhibitory contaminants as reported in the literature are given and comprehensively discussed

A new regime of carbon counting: the practices and politics of accounting for everyday carbon through C02e

Inspired by the commercial desires of global brands and retailers to access the lucrative green consumer market, carbon is increasingly being counted and made knowable at the mundane sites of everyday production and consumption, from the carbon footprint of a plastic kitchen fork to that of an online bank account. Despite the challenges of counting and making commensurable the global warming impact of a myriad of biophysical and societal activities, this desire to communicate a product or service's carbon footprint has sparked complicated carbon calculative practices and enrolled actors at literally every node of multi-scaled and vastly complex global supply chains. Against this landscape, this paper critically analyzes the counting practices that create the ‘e’ in ‘CO2e’. It is shown that, central to these practices are a series of tools, models and databases which, in building upon previous work (Eden, 2012 and Star and Griesemer, 1989) we conceptualize here as ‘boundary objects’. By enrolling everyday actors from farmers to consumers, these objects abstract and stabilize greenhouse gas emissions from their messy material and social contexts into units of CO2e which can then be translated along a product's supply chain, thereby establishing a new currency of ‘everyday supply chain carbon’. However, in making all greenhouse gas-related practices commensurable and in enrolling and stabilizing the transfer of information between multiple actors these objects oversee a process of simplification reliant upon, and subject to, a multiplicity of approximations, assumptions, errors, discrepancies and/or omissions. Further the outcomes of these tools are subject to the politicized and commercial agendas of the worlds they attempt to link, with each boundary actor inscribing different meanings to a product's carbon footprint in accordance with their specific subjectivities, commercial desires and epistemic framings. It is therefore shown that how a boundary object transforms greenhouse gas emissions into units of CO2e, is the outcome of distinct ideologies regarding ‘what’ a product's carbon footprint is and how it should be made legible. These politicized decisions, in turn, inform specific reduction activities and ultimately advance distinct, specific and increasingly durable transition pathways to a low carbon society.

Geography, skills and careers patterns at the boundary of creativity & innovation: digital technology and creative arts graduates in the UK

In the last decades, research on knowledge economies has taken central stage. Within this broader research field, research on the role of digital technologies and the creative industries has become increasingly important for researchers, academics and policy makers with particular focus on their development, supply-chains and models of production. Furthermore, many have recognised that, despite the important role played by digital technologies and innovation in the development of the creative industries, these dynamics are hard to capture and quantify. Digital technologies are embedded in the production and market structures of the creative industries and are also partially distinct and discernible from it. They also seem to play a key role in innovation of access and delivery of creative content. This chapter tries to assess the role played by digital technologies focusing on a key element of their implementation and application: human capital. Using student micro-data collected by the Higher Education Statistical Agency (HESA) in the United Kingdom, we explore the characteristics and location patterns of graduates who entered the creative industries, specifically comparing graduates in the creative arts and graduates from digital technology subjects. We highlight patterns of geographical specialisation but also how different context are able to better integrate creativity and innovation in their workforce. The chapter deals specifically with understanding whether these skills are uniformly embedded across the creative sector or are concentrated in specific sub-sectors of the creative industries. Furthermore, it explores the role that these graduates play in different sub-sector of the creative economy, their economic rewards and their geographical determinants.