Generation and Quantification of Ultrafine Particles through Terpene/Ozone Reaction in a Chamber Setting

Terpene/ozone reactions produce gas- and condensed-phase products and thus contribute to both indoor and outdoor aerosol. These reactions may be important in indoor settings, where terpenes are generated from indoor sources and ambient ozone can reach significant levels. Moreover, airway irritation has been observed in mice exposed to terpene oxidation products (OPs). The aim of this study was to characterize a system for generating and quantifying ultrafine particles formed through terpene/ozone reactions in preparation for inhalation toxicology experiments. Two common monoterpenes, f -pinene and d -limonene, and a hemiterpene, isoprene, were investigated. Ozone and gas-phase terpene were introduced continuously into a reaction flow tube, from which reaction products entered a plexiglass chamber. Particle number, mass, and size distribution (～15-750 nm) were monitored in the chamber for various reactant concentrations and air exchange rates (AERs). In all experiments, ozone was the limiting reagent and the reaction rate was much more rapid than the AER. Particles formed rapidly and in high concentrations in the pinene and limonene systems. Particle formation was slower in the isoprene system and fewer particles were formed; moreover, particle diameters were smaller. In all 3 systems, progressive growth of particles was observed due to condensation and coagulation processes. The isoprene system displayed instability with respect to aerosol characteristics and did not reach steady-state conditions. In the pinene system, ozone concentration was a strong predictor of steady-state particle number and mass concentration and particle diameter. The particle number was greater at higher AERs, but particles were smaller. This study is the first to incorporate measurement of ultrafine particles formed from terpene/ozone reactions into a controlled exposure chamber setting. Following system characterization, we will conduct mouse exposures to further investigate the respiratory effects of gas- and particle-phase terpene OPs.     

Minor Role of Mitochondrial Respiration for Fatty-Acid Induced Insulin Secretion

An appropriate insulin secretion by pancreatic beta-cells is necessary to maintain glucose homeostasis. A rise in plasma glucose leads to increased metabolism and an elevated cytoplasmic ATP/ADP ratio that finally triggers insulin granule exocytosis. In addition to this triggering pathway, one or more amplifying pathways—activated by amino acids or fatty acid—enhance secretion by promoting insulin granule recruitment to, and priming at, the plasma membrane. The aim of this study was to clarify the impact of the mitochondrial respiratory activity on fatty acid-induced insulin secretion that was assessed by an extracellular flux analyzer. Treatment of isolated mouse islets with glucose (20 mM) increased insulin secretion 18-fold and correlated with ATP-synthesizing respiration. Furthermore, oxygen consumption rate (OCR) significantly increased by 62% in response to glucose, whereas the addition of palmitate resulted only in a minor increase of OCR at both 2.8 mM (11%) and 20 mM glucose (21%). The addition of palmitate showed a pronounced increase of coupling efficiency (CE) at 2.8 mM glucose but no further insulin secretion. However, treatment with palmitate at 20 mM glucose increased insulin secretion about 32-fold accompanied by a small increase in CE. Thus, fatty acid induced respiration has a minor impact on insulin secretion. Our data clearly demonstrate that fatty acids in contrast to glucose play a minor role for respiration-mediated insulin secretion. In the presence of high glucose, fatty acids contribute partially to amplifying pathways of insulin secretion by further increasing mitochondrial activity in the islets of Langerhans.     

Polyethylene multiwalled carbon nanotube composites

Polyethylene (PE) multiwalled carbon nanotubes (MWCNTs) with weight fractions ranging from 0.1 to 10 wt% were prepared by melt blending using a mini-twin screw extruder. The morphology and degree of dispersion of the MWCNTs in the PE matrix at different length scales was investigated using scanning electron microscopy (SEM), transmission electron microscopy (TEM), atomic force microscopy (AFM) and wide-angle X-ray diffraction (WAXD). Both individual and agglomerations of MWCNTs were evident. An up-shift of 17 cm⁻¹ for the G band and the evolution of a shoulder to this peak were obtained in the Raman spectra of the nanocomposites, probably due to compressive forces exerted on the MWCNTs by PE chains and indicating intercalation of PE into the MWCNT bundles. The electrical conductivity and linear viscoelastic behaviour of these nanocomposites were investigated. A percolation threshold of about 7.5 wt% was obtained and the electrical conductivity of PE was increased significantly, by 16 orders of magnitude, from 10⁻²⁰ to 10⁻⁴ S/cm. The storage modulus (G′) versus frequency curves approached a plateau above the percolation threshold with the formation of an interconnected nanotube structure, indicative of ‘pseudo-solid-like’ behaviour. The ultimate tensile strength and elongation at break of the nanocomposites decreased with addition of MWCNTs. The diminution of mechanical properties of the nanocomposites, though concomitant with a significant increase in electrical conductivity, implies the mechanism for mechanical reinforcement for PE/MWCNT composites is filler-matrix interfacial interactions and not filler percolation. The temperature of crystallisation (T_c) and fraction of PE that was crystalline (F_c) were modified by incorporating MWCNTs. The thermal decomposition temperature of PE was enhanced by 20 K on addition of 10 wt% MWCNT.     

Transport of surface-modified nanoparticles through cell monolayers

We synthesized three peptides, a D-polyarginyl peptide (r8(FITC)), a Tat peptide (Tat(FITC)), and a control peptide (Cp(FITC)) and attached each to amino-CLIO, a nanoparticle 30 nm in diameter. We then examined the effective permeability, Peff, of all six materials through CaCo-2 monolayers. The transport of peptide-nanoparticles was characterized by a lag phase (0-8 h) and a steady-state phase (9-27 h). The steady-state Peff values for peptides were in the order r8(FITC)>Tat(FITC)=Cp(FITC). When r8(FITC) and Tat(FITC) peptides were attached to the nanoparticle, they conferred their propensity to traverse cell monolayers onto the nanoparticle, whereas Cp(FITC) did not. Thus, when the r8(FITC) peptide was attached to the amino-CLIO nanoparticle, the resulting peptide-nanoparticle had a Peff similar to that of this poly-D-arginyl peptide alone. The Peff of r8(FITC)-CLIO (MW approximately 1000 kDa) was similar to that of mannitol (MW=182 Da), a poorly transported reference substance, with a far lower molecular weight. These results are the first to indicate that the modification of nanoparticles by attachment of membrane-translocating sequence-based peptides can alter nanoparticle transport through monolayers. This suggests that the surface modification of nanoparticles might be a general strategy for enhancing the permeability of drugs and that high-permeability nanoparticle-based therapeutics can be useful in selected pharmaceutical applications.     

Synthesis and antiapoptotic activity of a novel analogue of the neutral sphingomyelinase inhibitor scyphostatin

The enantioselective synthesis of an analogue of scyphostatin, a potent inhibitor of the neutral sphingomyelinase, is described. The synthesis starts with cyclohexanone and a protected D-serine derivative. The key step is an asymmetric hydroxylation to access a hydroxycyclohexanone, which is transformed into a substituted hydroxycyclohexenone. This is converted into the scyphostatin analogue 14, a chemically and metabolically stabilised compound lacking the epoxy function of the natural congener and carrying a palmitic acid group instead of the native trienoyl residue. An evaluation of the biological activity of 14 revealed neutral sphingomyelinase inhibition in several in vivo test systems (monocytes, macrophages, hepatocytes) monitoring antiapoptotic effects and the inversion of phorbolester-induced translocation of green fluorescent protein labelled kinase (protein kinase C-alpha).     

Oxidative functionalization of carbon nanotubes in atmospheric pressure filamentary dielectric barrier discharge (APDBD)

The surface compositional and any structural changes that occur on carbon nanotubes using air-atmospheric pressure dielectric barrier discharge (APDBD) for functionalization are investigated employing Raman spectroscopy, X-ray photoelectron spectroscopy (XPS), thermal gravimetric analysis (TGA), X-ray diffraction (XRD), and neutron diffraction techniques. Atmospheric pressure plasmas (APP) are suggested to be particularly suitable for functionalization of aligned nanotubes, where wet chemical manipulation could damage or even destroy the highly desirable vertical alignment. In this work a detailed experimental study elucidating the effects of APDBD plasma treatment parameters (e.g. power density, discharge composition, inter-electrode gap and treatment time) on the electronic structure, physical, and chemical behaviour of carbon nanotubes has been conducted. In an atmospheric air we find an optimal oxidative functionalization of CNTs in our DBD system within few seconds (<5 s) at a discharge power of ∼0.5 kW. This investigation may find useful application as functionalization technique for CNT engineered devices and sensors.     

Orthogonal Peptide-Templated Labeling Elucidates Lateral ETAR/ETBR Proximity and Reveals Altered Downstream Signaling

Fine-tuning of G protein-coupled receptor (GPCR) signaling is important to maintain cellular homeostasis. Recent studies demonstrated that lateral GPCR interactions in the cell membrane can impact signaling profiles. Here, we report on a one-step labeling method of multiple membrane-embedded GPCRs. Based on short peptide tags, complementary probes transfer the cargo (e. g. a fluorescent dye) by an acyl transfer reaction with high spatial and temporal resolution within 5 min. We applied this approach to four receptors of the cardiovascular system: the endothelin receptor A and B (ET_AR and ET_BR), angiotensin II receptor type 1, and apelin. Wild type-like G protein activation after N-terminal modification was demonstrated for all receptor species. Using FRET-competent dyes, a constitutive proximity between hetero-receptors was limited to ET_AR/ET_BR. Further, we demonstrate, that ET_AR expression regulates the signaling of co-expressed ET_BR. Our orthogonal peptide-templated labeling of different GPCRs provides novel insight into the regulation of GPCR signaling.     

Structure-activity studies of RFamide peptides reveal subtype-selective activation of neuropeptide FF1 and FF2 receptors

Selectivity is a major issue in closely related multiligand/multireceptor systems. In this study we investigated the RFamide systems of hNPFF₁R and hNPFF₂R that bind the endogenous peptide hormones NPFF, NPAF, NPVF, and NPSF. By use of a systematic approach, we characterized the role of the C‐terminal dipeptide with respect to agonistic properties using synthesized [Xaa 7]NPFF and [Xaa 8]NPFF analogues. We were able to identify only slight differences in potency upon changing the position of Arg 7, as all modifications resulted in identical behavior at the NPFF₁R and NPFF₂R. However, the C‐terminal Phe 8 was able to be replaced by Trp or His with only a minor loss in potency at the NPFF₂R relative to the NPFF₁R. Analogues with shorter side chains, such as α‐amino‐4‐guanidino butyric acid ([Agb 7]NPFF) or phenylglycine ([Phg 8]NPFF), decreased efficacy for the NPFF₁R to 25–31 % of the maximal response, suggesting that these agonist–receptor complexes are more susceptible to structural modifications. In contrast, mutations to the conserved Asp 6.59 residue in the third extracellular loop of both receptors revealed a higher sensitivity toward the hNPFF₂R receptor than toward hNPFF₁R. These data provide new insight into the subtype‐specific agonistic activation of the NPFF₁ and NPFF₂ receptors that are necessary for the development of selective agonists.     

Assessment of German nitrous oxide emissions using empirical modelling approaches

Direct nitrous oxide (N₂O) emissions from agricultural soils contribute considerably to anthropogenic GHG emissions. Albeit a key source of emissions in many countries, direct N₂O emissions are still calculated and reported to the United Nations Convention on Climate Change using default emission factors defined in the IPCC guidelines (IPCC 1996, 2006). It is known that processes controlling production and transport of N₂O are highly sensitive to environmental conditions defined by weather, soil and management. The accuracy of N₂O emission budgets and the efficiency of mitigation can be improved if those dependencies are considered with regionalized emission factors. In this study an empirical method originating from soft computing techniques based on measured data is developed and applied to quantify direct N₂O emissions from agricultural soils at field and national level in Germany between 1990 and 2005. The method is used to derive maps of emission factor distribution of direct N₂O emissions of agricultural land in Germany. Model results are compared with alternative empirical approaches from literature. Results from developing empirical models show that grassland and cropland have to be differentiated according to the key controls driving N₂O emissions. N₂O emissions of German croplands are highly influenced by climatic conditions and soil properties. The variability of N₂O fluxes on grasslands is mainly driven by the fertilizer N applied. The model comparison using measured European N₂O emissions exhibits profound discrepancies between the models used on a regional scale. The nationwide budgets derived span a narrow range of −8 to 28% relative to direct N₂O emissions quantified by the German national inventory report. The emission factor of German agriculture estimated by the developed model is 0.91% of fertilizer N applied.