Electrocatalytic oxidation of ascorbic acid using a single layer of gold nanoparticles immobilized on 1,6-hexanedithiol modified gold electrode

This paper describes the electrocatalytic oxidation of ascorbic acid (AA) in phosphate buffer solution by the immobilized citrate capped gold nanoparticles (AuNPs) on 1,6-hexanedithiol (HDT) modified Au electrode. X-ray photoelectron spectrum (XPS) of HDT suggests that it forms a monolayer on Au surface through one of the two single bondSH groups and the other single bondSH group is pointing away from the electrode surface. The free single bondSH groups of HDT were used to covalently attach colloidal AuNPs. The covalent attachment of AuNPs on HDT monolayer was confirmed from the observed characteristic carboxylate ion stretching modes of citrate attached with AuNPs in the infra-red reflection absorption spectrum (IRRAS) in addition to a higher reductive desorption charges obtained for AuNPs immobilized on HDT modified Au (Au/HDT/AuNPs) electrode in 0.1 M KOH when compared to HDT modified Au (Au/HDT) electrode. The electron transfer reaction of [Fe(CN)6]4−/3− was markedly hindered at the HDT modified Au (Au/HDT) electrode while it was restored with a peak separation of 74 mV after the immobilization of AuNPs on Au/HDT (Au/HDT/AuNPs) electrode indicating a good electronic communication between the immobilized AuNPs and the underlying bulk Au electrode through a HDT monolayer. The Cottrell slope obtained from the potential-step chronoamperometric measurements for the reduction of ferricyanide at Au/HDT/AuNPs was higher than that of bare Au electrode indicating the increased effective surface area of AuNPs modified electrode. The Au/HDT/AuNPs electrode exhibits excellent electrocatalytic activity towards the oxidation of ascorbic acid (AA) by enhancing the oxidation peak current to more than two times with a 210 mV negative shift in the oxidation potential when compared to a bare Au electrode. The standard heterogeneous electron transfer rate constant (ks) calculated for AA oxidation at Au/HDT/AuNPs electrode was 5.4 × 10−3 cm s−1. The oxidation peak of AA at Au/HDT/AuNPs electrode was highly stable upon repeated potential cycling. Linear calibration plot was obtained for AA over the concentration range of 1–110 μM with a correlation coefficient of 0.9950. The detection limit of AA was found to be 1 μM. The common physiological interferents such as glucose, oxalate ions and urea do not show any interference within the detection limit of AA. The selectivity of the AuNPs modified electrode was illustrated by the determination of AA in the presence of uric acid.

GABAB1 and GABAB2 receptor subunits co-expressed in cultured human RPE cells regulate intracellular Ca2+ via Gi/o-protein and phospholipase C pathways

GABAB receptors associate with Gi/o-proteins that regulate voltage-gated Ca(2+) channels and thus the intracellular Ca(2+) concentration ([Ca(2+)]i), there is also reported cross-regulation of phospholipase C. These associations have been studied extensively in the brain and also shown to occur in non-neural cells (e.g. human airway smooth muscle). More recently GABAB receptors have been observed in chick retinal pigment epithelium (RPE). The aims were to investigate whether the GABAB receptor subunits, GABAB1 and GABAB2, are co-expressed in cultured human RPE cells, and then determine if the GABAB receptor similarly regulates the [Ca(2+)]i of RPE cells and if phospholipase C is involved. Human RPE cells were cultured from 5 donor eye cups. Evidence for GABAB1 and GABAB2 mRNAs and proteins in the RPE cell cultures were investigated using real time PCR, western blots and immunofluorescence. The effects of the GABAB receptor agonist baclofen, antagonist CGP46381, a Gi/o-protein inhibitor pertussis toxin, and the phospholipase C inhibitor U73122 on [Ca(2+)]i in cultured human RPE were demonstrated using Fluo-3. Both GABAB1 and GABAB2 mRNA and protein were identified in cell cultures of human RPE; antibody staining was co-localized to the cell membrane and cytoplasm. One-hundred μM baclofen caused a transient increase in the [Ca(2+)]i of RPE cells regardless of whether Ca(2+) was added to the buffer. Baclofen induced increases in the [Ca(2+)]i were attenuated by pre-treatment with CGP46381, pertussis toxin, and U73122. GABAB1 and GABAB2 are co-expressed in cell cultures of human RPE. GABAB receptors in RPE regulate the [Ca(2+)]i via a Gi/o-protein and phospholipase C pathway.

Emotional intelligence and job-demands resources model

Occupational stress research has consistently demonstrated negative effects for employees. Research also describes potential moderators of this relationship. While research has revealed some positive effects of emotional intelligence (EI) on employee adjustment, it has neglected investigation of their potential stress buffering effects. Based on the Job-Demand Resources model, it was predicted that higher trait emotional intelligence would act as a buffer to the potential negative effects of stressors on employee adjustment. Hierarchical multiple regression analyses with a sample of 306 nurses found no main effects of EI but revealed eight moderating effects. While some interactions support the buffering hypothesis, others revealed buffering for those with low EI. Findings are discussed in terms of theoretical and practical implications.

Competitive interactions of anti-carcinogens with serum albumin: A spectroscopic study of bendamustine and dexamethasone with the aid of chemometrics

Interactions between the anti-carcinogens, bendamustine (BDM) and dexamethasone (DXM), with bovine serum albumin (BSA) were investigated with the use of fluorescence and UV–vis spectroscopies under pseudo-physiological conditions (Tris–HCl buffer, pH 7.4). The static mechanism was responsible for the fluorescence quenching during the interactions; the binding formation constant of the BSA–BDM complex and the binding number were 5.14 × 105 L mol−1 and 1.0, respectively. Spectroscopic studies for the formation of BDM–BSA complex were interpreted with the use of multivariate curve resolution – alternating least squares (MCR–ALS), which supported the complex formation. The BSA samples treated with site markers (warfarin – site I and ibuprofen – site II) were reacted separately with BDM and DXM; while both anti-carcinogens bound to site I, the binding constants suggested that DXM formed a more stable complex. Relative concentration profiles and the fluorescence spectra associated with BDM, DXM and BSA, were recovered simultaneously from the full fluorescence excitation–emission data with the use of the parallel factor analysis (PARAFAC) method. The results confirmed that on addition of DXM to the BDM–BSA complex, the BDM was replaced and the DXM–BSA complex formed; free BDM was released. This finding may have consequences for the transport of these drugs during any anti-cancer treatment.

Protein loaded mesoporous silica spheres as a controlled delivery platform

Background The adsorption of bovine serum albumin (BSA) onto mesoporous silica spheres (MPS) synthesized from silica colloids was studied employing real time in situ measurements. The stabilities of the BSA at different pH values, their isoelectric points and zeta potentials were determined in order to probe the interactions between the protein and the mesoporous silica. Results The pore size of MPS was designed for protein, and this, coupled with an in depth understanding of the physico-chemical characteristics of the protein and MPS has yielded a better binding capacity and delivery profile. The adsorption isotherm at pH 4.2 fitted the Langmuir model and displayed the highest adsorption capacity (71.43 mg mL-1 MPS). Furthermore, the delivery rates of BSA from the MPS under physiological conditions were shown to be dependent on the ionic strength of the buffer and protein loading concentration. Conclusion Economics and scale-up considerations of mesoporous material synthesized via destabilization of colloids by electrolyte indicate the scaleability and commercial viability of this technology as a delivery platform for biopharmaceutical applications.

Affinity purification of plasmid DNA directly from crude bacterial cell lysates

We have shown previously that a sequence-specific DNA-binding protein based on the Lac repressor protein can isolate pre-purified DNA efficiently from simple buffer solution but our attempts to purify plasmids directly from crude starting materials were disappointing with unpractically low DNA yields. We have optimized tbe procedure and present a simple affinity methodology whereby plasmid DNA is purified directly by mixing two crude cell lysates, one cell lysate containing the plasmid and the other the protein affinity ligand, without the need for treatment by RNaseA. After IMAC chromatography, high purity supercoiled DNA is recovered in good yields of 100-150 μg plasmid per 200 mL shake flask culture. Moreover, the resulting DNA is free from linear or open-circular plasmid DNA, genomic DNA, RNA, and protein, to the limits of our detection. Furthermore, we show that lyophilized affinity ligand can be stored at room temperature and re-hydrated for use when required.

Enhancing methacrylate-monolith-based downstream processes to champion plasmid DNA production

Increasing numbers of preclinical and clinical studies are utilizing pDNA (plasmid DNA) as the vector. In addition, there has been a growing trend towards larger and larger doses of pDNA utilized in human trials. The growing demand on pDNA manufacture leads to pressure to make more in less time. A key intervention has been the use of monoliths as stationary phases in liquid chromatography. Monolithic stationary phases offer fast separation to pDNA owing to their large pore size, making pDNA in the size range from 100 nm to over 300 nm easily accessible. However, the convective transport mechanism of monoliths does not guarantee plasmid purity. The recovery of pure pDNA hinges on a proper balance in the properties of the adsorbent phase, the mobile phase and the feedstock. The effects of pH and ionic strength of binding buffer, temperature of feedstock, active group density and the pore size of the stationary phase were considered as avenues to improve the recovery and purity of pDNA using a methacrylate-based monolithic adsorbent and Escherichia coli DH5α-pUC19 clarified lysate as feedstock. pDNA recovery was found to be critically dependent on the pH and ionic strength of the mobile phase. Up to a maximum of approx. 92% recovery was obtained under optimum conditions of pH and ionic strength. Increasing the feedstock temperature to 80°C increased the purity of pDNA owing to the extra thermal stability associated with pDNA over contaminants such as proteins. Results from toxicological studies of the plasmid samples using endotoxin standard (E. coli 0.55:B5 lipopolysaccharide) show that endotoxin level decreases with increasing salt concentration. It was obvious that large quantities of pure pDNA can be obtained with minimal extra effort simply by optimizing process parameters and conditions for pDNA purification.

LacO-LacI interaction in affinity adsorption of plasmid DNA

Current approaches for purifying plasmids from bacterial production systems exploit the physiochemical properties of nucleic acids in non-specific capture systems. In this study, an affinity system for plasmid DNA (pDNA) purification has been developed utilizing the interaction between the lac operon (lacO) sequence contained in the pDNA and a 64mer synthetic peptide representing the DNA-binding domain of the lac repressor protein, LacI. Two plasmids were evaluated, the native pUC19 and pUC19 with dual lacO3/lacOs operators (pUC19lacO3/lacOs), where the lacOs operator is perfectly symmetrical. The DNA-protein affinity interaction was evaluated by surface plasmon resonance using a Biacore system. The affinity capture of DNA in a chromatography system was evaluated using LacI peptide that had been immobilized to Streamline™ adsorbent. The KD-values for double stranded DNA (dsDNA) fragments containing lacO1 and lacO3 and lacOs and lacO3 were 5.7 ± 0.3 × 10 -11 M and 4.1 ± 0.2 × 10-11 M respectively, which compare favorably with literature reports of 5 × 10-10 - 1 × 10-9 M for native laCO1 and 1-1.2 × 10-10 M for lacO1 in a saline buffer. Densitometric analysis of the gel bands from the affinity chromatography run clearly showed a significant preference for capture of the supercoiled fraction from the feed pDNA sample. The results indicate the feasibility of the affinity approach for pDNA capture and purification using native protein-DNA interaction.

A method to evaluate the response of the coronary circulation of perfused rat heart to adenosine

Exogenous adenosine causes a monophasic dilation of the coronary vessels in paced, perfused rat heart preparations. Because levels of endogenous adenosine in paced hearts may mask the presence of high potency adenosine receptors, we have developed a method to measure coronary vascular responses in a potassium-arrested heart. Hearts from adult male, Wistar rats were perfused at a constant flow rate of 10 mL/min in the nonrecirculating, Langendorff mode, using Krebs-Henseleit buffer. After 30 min, coronary perfusion pressure was 44 +/- 1 mmHg (mean +/- SEM). Hearts were then perfused with a modified Krebs-Henseleit buffer containing 35 mM potassium. Coronary perfusion pressure increased by 84 +/- 3 mmHg. Adenosine-induced reductions in coronary perfusion pressure were expressed as a percentage of the maximal increase in pressure produced by modified Krebs-Henseleit buffer from the equilibration level. A concentration-response curve for adenosine (n = 6) was biphasic and best described by the presence of two adenosine receptors, with negative log EC50 values of 8.8 +/- 0.3 and 4.3 +/- 0.1, representing 29 +/- 3 and 71 +/- 3%, respectively, of the observed response. Interstitial adenosine sampled by microdialysis during potassium arrest was 25% of the concentration found in paced hearts. Endogenous adenosine in nonarrested hearts may obscure the biphasic response of the coronary vessels to adenosine.

Bioaerosols associated with industrial green waste composting facilities and optimal microbiological indicators

This project developed and assessed a standard operating procedure for monitoring microbiological aerosol levels and dispersal from Australian industrial composting facilities. Development occurred via seasonal monitoring of such operations with evaluation of optimal microbial indicator organisms, sampling and analysis logistics. The resultant procedure allows practical end-user assessment of compost-associated bioaerosol levels, and potential health risks to proximal residential populations encroaching on such composting facilities and on-site industrial operations personnel.

The combination of plant-expressed cellobiohydrolase and low dosages of cellulases for the hydrolysis of sugar cane bagasse

Background The expression of biomass-degrading enzymes (such as cellobiohydrolases) in transgenic plants has the potential to reduce the costs of biomass saccharification by providing a source of enzymes to supplement commercial cellulase mixtures. Cellobiohydrolases are the main enzymes in commercial cellulase mixtures. In the present study, a cellobiohydrolase was expressed in transgenic corn stover leaf and assessed as an additive for two commercial cellulase mixtures for the saccharification of pretreated sugar cane bagasse obtained by different processes. Results Recombinant cellobiohydrolase in the senescent leaves of transgenic corn was extracted using a simple buffer with no concentration step. The extract significantly enhanced the performance of Celluclast 1.5 L (a commercial cellulase mixture) by up to fourfold on sugar cane bagasse pretreated at the pilot scale using a dilute sulfuric acid steam explosion process compared to the commercial cellulase mixture on its own. Also, the extracts were able to enhance the performance of Cellic CTec2 (a commercial cellulase mixture) up to fourfold on a range of residues from sugar cane bagasse pretreated at the laboratory (using acidified ethylene carbonate/ethylene glycol, 1-butyl-3-methylimidazolium chloride, and ball-milling) and pilot (dilute sodium hydroxide and glycerol/hydrochloric acid steam explosion) scales. We have demonstrated using tap water as a solvent (under conditions that mimic an industrial process) extraction of about 90% recombinant cellobiohydrolase from senescent, transgenic corn stover leaf that had minimal tissue disruption. Conclusions The accumulation of recombinant cellobiohydrolase in senescent, transgenic corn stover leaf is a viable strategy to reduce the saccharification cost associated with the production of fermentable sugars from pretreated biomass. We envisage an industrial-scale process in which transgenic plants provide both fibre and biomass-degrading enzymes for pretreatment and enzymatic hydrolysis, respectively.

A vibrational spectroscopic study of the silicate mineral kornerupine

We have studied the mineral kornerupine, a borosilicate mineral, by using a combination of scanning electron microscopy with energy-dispersive analysis and Raman and infrared spectroscopy. Qualitative chemical analysis of kornerupine shows a magnesium–aluminum silicate. Strong Raman bands at 925, 995, and 1051 cm−1 with bands of lesser intensity at 1035 and 1084 cm−1 are assigned to the silicon–oxygen stretching vibrations of the siloxane units. Raman bands at 923 and 947 cm−1 are attributed to the symmetrical stretching vibrations of trigonal boron. Infrared spectra show greater complexity and the infrared bands are more difficult to assign. Two intense Raman bands at 3547 and 3612 cm−1 are assigned to the stretching vibrations of hydroxyl units. The infrared bands are observed at 3544 and 3610 cm−1. Water is also identified in the spectra of kornerupine.

Independent distractor frequency and age-of-acquisition dffects in picture-word interference: FMRI evidence for post-lexical and lexical accounts according to distractor type

In two fMRI experiments, participants named pictures with superimposed distractors that were high or low in frequency or varied in terms of age of acquisition. Pictures superimposed with low-frequency words were named more slowly than those superimposed with high-frequency words, and late-acquired words interfered with picture naming to a greater extent than early-acquired words. The distractor frequency effect (Experiment 1) was associated with increased activity in left premotor and posterior superior temporal cortices, consistent with the operation of an articulatory response buffer and verbal selfmonitoring system. Conversely, the distractor age-of-acquisition effect (Experiment 2) was associated with increased activity in the left middle and posterior middle temporal cortex, consistent with the operation of lexical level processes such as lemma and phonological word form retrieval. The spatially dissociated patterns of activity across the two experiments indicate that distractor effects in picture-word interference may occur at lexical or postlexical levels of processing in speech production.

Progressive dysgraphia in a case of posterior cortical atrophy

Dysgraphia (agraphia) is a common feature of posterior cortical atrophy (PCA). However, detailed analyses of these spelling and writing impairments are infrequently conducted. LM is a 59-year-old woman with dysgraphia associated with PCA. She presented with a two-year history of decline in her writing and dressmaking skills. A 3D T1-weighted MRI scan confirmed selective bi-parietal atrophy, with relative sparing of the hippocampi and other cortical regions. Analyses of LM's preserved and impaired spelling abilities indicated mild physical letter distortions and a significant spelling deficit characterised by letter substitutions, insertions, omissions, and transpositions that was systematically sensitive to word length while insensitive to real word versus nonword category, word frequency, regularity, imagery, grammatical class and ambiguity. Our findings suggest a primary graphemic buffer disorder underlies LM's spelling errors, possibly originating from disruption to the operation of a fronto-parietal network implicated in verbal working memory.

An experimental study of the effects of work stress, work control, and task information on adjustment

The present study was designed to examine the main and interactive effects of task demands, work control, and task information on levels of adjustment. Task demands, work control, and task information were manipulated in an experimental setting where participants completed a letter-sorting activity (N= 128). Indicators of adjustment included measures of positive mood, participants' perceptions of task performance, and task satisfaction. Results of the present study provided some support for the main effects of objective task demands, work control, and task information on levels of adjustment. At the subjective level of analysis, there was some evidence to suggest that work control and task information interacted in their effects on levels of adjustment. There was minimal support for the proposal that work control and task information would buffer the negative effects of task demands on adjustment. There was, however, some evidence to suggest that the stress-buffering role of subjective work control was more marked at high, rather than low, levels of subjective task information.