Piericiden A Sensitivity, Site 1 Phosphorylation, and Reduced Nicotinamide Adenine Dinucleotide Dehydrogenase during Iron-limited Growth of Candida utilis

It has been reported that cells of Candida utilis, grown in continuous culture under iron-limited conditions, develop site 1 phosphorylation, without the appearance of piericidin sensitivity and without changes in the iron-sulfur centers of NADH dehydrogenase, on aeration in the presence of cycloheximide, as well as on increasing the supply of iron during growth. These findings were reinvestigated in the present study. The parameters and properties followed during these transitions were sensitivity of NADH oxidation to piericidin, presence or absence of coupling site 1, EPR signals appearing on reduction with NADH or dithionite, the specific activities of NADH oxidase, NADH-ferricyanide reductase, and NADH-5-hydroxy-1,4-naphthoquinone (juglone) reductase, and the kinetic behavior of NADH dehydrogenase in the ferricyanide assay. Monitoring the rates of oxidation of NADH in submitochondrial particles with artificial oxidants, observing the kinetics of the ferricyanide assay, and measuring the concentration of iron-sulfur centers elicited by EPR permitted ascertaining the type of NADH dehydrogenase present and its relative concentration in different experimental situations. It was found that on gradually increasing the concentration of iron during continuous culture (transition from ironlimited to iron- and substrate-limited growth), as well as on aeration of iron-limited cells, coupling site 1, piericidin sensitivity, NADH-ferricyanide activity, and iron-sulfur centers 1 and 2 increased concurrently, with concomitant decline of NADH-juglone reductase activity. Cycloheximide prevented all these changes. Iron-sulfur centers 3 plus 4 underwent relatively little increase during these transitions. It is concluded that in both of these experimental conditions a replacement of the type of NADH dehydrogenase present in exponential phase cells by that characteristic of stationary phase cells occurs and that the appearance of site 1 phosphorylation, piercidin sensitivity, and iron-sulfur centers 1 plus 2, all associated with the latter enzyme, is a consequence of this replacement. No evidence was found for the development of coupling site 1 without the appearance of piericidin sensir t

Reduced Nicotinamide Adenine Dinucleotide Dehydrogenase, Piericidin Sensitivity, and Site 1 Phosphorylation in Different Growth Phases of Candida utilis

Reports in the literature indicate that during the exponential phase of growth of Candida utilis NADH oxidation is insensitive to rotenone, that rotenone sensitivity is acquired during the transition to the late stationary phase and is again lost on catabolite repression. The acquisition and loss of rotenone sensitivity appears to be accompanied by similar changes in Site 1 phosphorylation but does not appear to be reflected in the rate of oxidation of NADH (by mitochondria) or of NAD-linked substrates (by mitochondria or whole cells). In the present paper evidence is presented that these fluctuations in sensitivity to inhibitors of NADH oxidation reflect the presence of different types of inner membrane-bound NADH dehydrogenases in different phases of growth. Thus inner membrane preparations from exponential phase cells contain an NADH dehydrogenase which reacts equally well with ferricyanide and juglone as electron acceptor, appears to be very labile, and lacks EPR signals corresponding to iron-sulfur Centers 1 and 2, whereas a new species, probably an iron-sulfur protein, with resonances at g|| = 2.01, and g⊥ = 1.92 in the reduced state, is present. This species is not significantly reduced by NADH. In corresponding preparations from late stationary phase cells NADH-ferricyanide activity is high, juglone reductase activity is low, and the enzyme is stable and exhibits the EPR signals of iron-sulfur Centers 1 and 2, whereas the EPR signals of iron-sulfur Centers 3 + 4 change very little on transition from exponential to stationary phase cells. There is also a decrease in cytochrome concentration. Most prominent among these is a b-type cytochrome (g = 2.54; 2.23; 1.87) which decreases 2- to 3-fold. The EPR detectable species with g|| = 2.01 and g⊥ = 1.92 in the reduced state is no longer detected. On catabolite repression of late stationary phase cells there is an 80 to 90% decline in NADH-ferricyanide activity, of iron-sulfur Centers 1 and 2, a 50 to 60% decrease of Centers 3 + 4, and an increase in a b cytochrome, but the specific activity in NADH-juglone reductase and NADH oxidase assays increases, the enzyme becomes once again labile, and the EPR detectable species with g|| = 2.01 and g⊥ = 1.92 appears on reduction with dithionite. All these changes are prevented by cycloheximide. The data suggest that sensitivity to piericidin A and coupling to energy conservation Site 1 are properties of the type of NADH dehydrogenase present in late stationary phase cells but not in exponential phase or of catabolite-repressed cells

Surface Defect Mitigation of Additively Manufactured Parts Using Surfactant-Mediated Electroless Nickel Coatings

The emergence of defects during the early production phases of ferrous-alloy additively manufactured (AM) parts poses a serious threat to their versatility and adversely impacts their overall mechanical performance in industries ranging from aerospace engineering to medicine. Lack of fusion and gas entrapment during the manufacturing stages leads to increased surface roughness and porosities in the finished part. In this study, the efficacy of employing electroless nickel–boron (Ni-B) deposition to fill and level simulated AM defects was evaluated. The approach to levelling was inspired by the electrochemical deposition techniques used to fill vias in the electronics industry that (to some extent) resemble the size and shape of AM-type defects. This work investigated the use of surfactants to attenuate surface roughness in electroless nickel coatings, thereby achieving the preferential inhibition of the coating thickness on the surface and promoting the filling of the simulated defects. A cationic surfactant molecule, CTAB (cetyltrimethyl ammonium bromide), and a nonpolar surfactant, PEG (polyethylene glycol), at different concentrations were tested using a Ni-B electrolyte for the levelling study. It was found that the use of electroless Ni-B to fill simulated defects on ferrous alloys was strongly influenced by the concentration and nature of the surfactant. The highest levelling percentages were obtained for the heavy-molecular-weight PEG-mediated coatings at 1.2 g/L. The results suggest that electroless Ni-B deposition could be a novel and facile approach to filling defects in ferrous-based AM parts

Asymmetric Organocatalysis and Continuous Chemistry for an Efficient and Cost-Competitive Process to Pregabalin

Herein, we present the scale up development of an innovative synthetic process to pregabalin. The process is underpinned by two enabling technologies critical to its success; continuous chemistry allowed a safe and clean production of nitroalkene, and asymmetric organocatalysis gave access to the chiral intermediate in an enantioenriched form. Crucial to the success of the process was the careful development of a continuous process to nitroalkene and optimization of the organocatalyst and of the reaction conditions to attain remarkably high turn-over frequency in the catalytic asymmetric reaction. Successful recycle of the organocatalysts was also developed in order to achieve a cost-competitive process

Movement and Physiological Demands of Australasian National Rugby League Referees.

Purpose: To evaluate the movement and physiological demands of the Australasian National Rugby League (NRL) referees, officiating with a ‘two referee’ (i.e., ‘lead’ and ‘pocket’) system and to compare the demands of the lead referee and pocket referees. Methods: 10 Hz global positioning system devices were used to obtain 86 data sets (‘lead’, n=41; ‘pocket’, n=45) on 19 NRL referees. Total distance, relative distance covered and heart rate per half and across match-play was examined within and between referees using t-tests. Distance, time and number of movement 'efforts' were examined in six velocity classifications (i.e., standing 7.0 m.s-1) using ANOVA. Cohen's d effect sizes were reported. Results: There were no significant differences between the ‘lead’ and ‘pocket’ referee for any movement or physiological variable. There was an overall significant (large; very large) effect for distance (% distance) and time (% time) (P 5.51 m.s-1. Conclusions: Findings highlight the intermittent nature of rugby league refereeing, but show that there were no differences in the movement and physiological demands of the two refereeing roles. Findings are valuable for those responsible for the preparation, training and conditioning of NRL referees, and to ensure training prepares for and simulates match demands

Biosemiotics, politics and Th.A. Sebeok’s move from linguistics to semiotics

This paper will focus on the political implications for the language sciences of Sebeok’s move from linguistics to a global semiotic perspective, a move that ultimately resulted in biosemiotics. The paper will seek to make more explicit the political bearing of a biosemiotic perspective in the language sciences and the human sciences in general. In particular, it will discuss the definition of language inherent in Sebeok’s project and the fundamental re-drawing of the grounds of linguistic debate heralded by Sebeok’s embrace of the concept of modelling. Thus far, the political co-ordinates of the biosemiotic project have not really been made explicit. This paper will therefore seek to outline 1. how biosemiotics enables us to reconfigure our understanding of the role of language in culture; 2. how exaptation is central to the evolution of language and communication, rather than adaptation; 3. how communication is the key issue in biosphere, rather than language, not just because communication includes language but because the language sciences often refer to language as if it were mere “chatter”, “tropes” and “figures of speech”; 4. how biosemiotics, despite its seeming “neutrality” arising from its transdisciplinarity, is thoroughly political; 5. how the failure to see the implications of the move from linguistics to semiotics arises from the fact that biosemiotics is devoid of old style politics, which is based on representation (devoid of experience) and “construction of [everything] in discourse” (which is grounded in linguistics, not communication study). In contrast to the post-“linguistic turn” idea that the world is “constructed in discourse”, we will argue that biosemiotics entails a reconfiguration of the polis and, in particular, offers the chance to completely reconceptualise ideology

A force profile analysis comparison between functional data analysis, statistical parametric mapping and statistical non-parametric mapping in on-water single sculling

Objectives: To examine whether the Functional Data Analysis (FDA), Statistical Parametric Mapping (SPM) and Statistical non-Parametric Mapping (SnPM) hypothesis testing techniques differ in their ability to draw inferences in the context of a single, simple experimental design. Design: The sample data used is cross-sectional (two-sample gender comparison) and evaluation of differences between statistical techniques used a combination of descriptive and qualitative assessments. Methods: FDA, SPM and SnPM t-tests were applied to sample data of twenty highly skilled male and female rowers, rowing at 32 strokes per minute in a single scull boat. Statistical differences for gender were assessed by applying two t-tests (one for each side of the boat). Results: The t-statistic values were identical for all three methods (with the FDA t-statistic presented as an absolute measure). The critical t-statistics (tcrit) were very similar between the techniques, with SPM tcrit providing a marginally higher tcrit than the FDA and SnPM tcrit values (which were identical). All techniques were successful in identifying consistent sections of the force waveform, where male and female rowers were shown to differ significantly (p < 0.05). Conclusions: This is the first study to show that FDA, SPM and SnPM t-tests provide consistent results when applied to sports biomechanics data. Though the results were similar, selection of one technique over another by applied researchers and practitioners should be based on the underlying parametric assumption of SPM, as well as contextual factors related to the type of waveform data to be analysed and the experimental research question of interest

MitoNeoD:a mitochondria-targeted superoxide probe

Mitochondrial superoxide (O2⋅−) underlies much oxidative damage and redox signaling. Fluorescent probes can detect O2⋅−, but are of limited applicability in vivo, while in cells their usefulness is constrained by side reactions and DNA intercalation. To overcome these limitations, we developed a dual-purpose mitochondrial O2⋅− probe, MitoNeoD, which can assess O2⋅− changes in vivo by mass spectrometry and in vitro by fluorescence. MitoNeoD comprises a O2⋅−-sensitive reduced phenanthridinium moiety modified to prevent DNA intercalation, as well as a carbon-deuterium bond to enhance its selectivity for O2⋅− over non-specific oxidation, and a triphenylphosphonium lipophilic cation moiety leading to the rapid accumulation within mitochondria. We demonstrated that MitoNeoD was a versatile and robust probe to assess changes in mitochondrial O2⋅− from isolated mitochondria to animal models, thus offering a way to examine the many roles of mitochondrial O2⋅−production in health and disease

Effect of additive concentration during copper deposition using EnFACE electrolyte

Copper deposition from solutions using high concentration of acid, metal ions and polyethylene glycol (PEG), and bis-(3-sulphopropyl) disulphide (SPS) and chloride ions (Cl-) is well known. A recent maskless micropatterning technology, which has the potential to replace the traditional photolithographic process, called EnFACE, proposed using an acid-free, low metal ion solution which is in direct contrast to those used in standard plating technology. In this work copper has been deposited using both standard electroplating solutions and those used in the EnFACE process. In the standard electrolyte 0.63 M CuSO4 and 2.04 M H2SO4 has been used, along with Gleam additives supplied by Dow Chemicals. For the Enface electrolyte, copper deposition has been carried out without any acid, and with different concentrations of additives between 17%-200% of those recommended by suppliers. 25 μm of metal has been plated on stainless steel coupons as suggested by ASTM, peeled off and subjected to ductility and resistance measurements. Scanning electron microscopy and electron back scatter diffraction have been carried out to determine the deposit morphology. It was found that copper deposits obtained from acid-free solutions containing low concentration of metal ion and additives produced copper deposits with properties which are comparable to those obtained from standard electrolytes. The optimum additive concentration for the EnFACE electrolyte was 50% of the supplier recommended value