Structural characterisation of pre-processed thermoplastic protein derived from bloodmeal

Additives are required to convert bloodmeal powder into an extrudable thermoplastic protein-based bioplastic. These include a protein denaturant, a surfactant, a reducing agent and plasticisers. The objective of this work was to assess the structural changes induced in bloodmeal by these additives prior to extrusion. Structure was investigated using Fourier transform infrared (FT-IR) spectroscopy, wide angle X-ray scattering (WAXS) and synchrotron light based FT-IR microspectroscopy. FT-IR results suggested the additives reduced α-helical content. The shape of the amide I region (1600 – 1700 cm⁻¹, representing carbonyl group stretching in the protein backbone) is known to depend on protein secondary structures. Bloodmeal showed a broad, convoluted peak in this region, with a maximum in the range 1648 – 1658 cm⁻¹, associated with α-helices. With processing additives, a dip was seen in the α-helix region, with twin peaks emerging either side of it. Urea, one of the additives, also absorbs in the amide I region and may also contribute to a change in its shape. Analysis of the amide 3 region supported a reduction in the ratio of α helices to β sheets. Further support of structural changes was shown by WAXS. The additives decreased the sharpness of peaks corresponding to 4.8 Å and 10 Å, thought to represent intra-helix spacing and inter-helix packing respectively. FT-IR microspectroscopy at the Australian Synchrotron enabled spatial variations in secondary structure to be explored using peaks in the amide 3 region. Spatial distribution of secondary structure was detected in bloodmeal and thermoplastically modified bloodmeal prior to extrusion (PPM-TEG). Bloodmeal showed domain separation on the approximate order of 10 μm, whilst PPM-TEG appeared to have larger phases and overall reduced α-helical content, relative to beta sheets

Plasticization of Bloodmeal-based Thermoplastics

Water is the most common plasticizer for proteinbased thermoplastics, lowering the softening point to a allow processing without excessive degradation. The biggest drawback of using water a plasticizer is that water easily evaporates from the material during use or storage. This leads to embrittlement and loss of functionality over time. In this study a series of high molecular mass plasticizers were evaluated for their efficiency in plasticizing bloodmeal-based thermoplastics. It was found that propylene glycol, di and tri-ethylene glycol were most efficient in increasing the material’s ductility, as measured by elongation at break. Using 10 parts plasticizer per hundred bloodmeal (pphBM) in combination with 10 pphBM urea gave optimal results in terms of Young’s modulus, tensile strength and processability. The mechanical properties of plasticized samples showed a stronger dependency on moisture content, compared to unplasticized samples and reached higher equilibrium moisture content in a shorter time. Using 10 pphBM TEG as plasticizer in resulted in a plastic material with a Young’s modulus of 869 MPa, tensile strength of 14.7 MPa and an elongation at break of 46%

A Drosophila Model for Clostridium difficile Toxin CDT Reveals Interactions with Multiple Effector Pathways.

Clostridium difficile infections (CDIs) cause severe and occasionally life-threatening diarrhea. Hyper-virulent strains produce CDT, a toxin that ADP-ribosylates actin monomers and inhibits actin polymerization. We created transgenic Drosophila lines expressing the catalytic subunit CDTa to investigate its interaction with host signaling pathways in vivo. When expressed in the midgut, CDTa reduces body weight and fecal output and compromises survival, suggesting severe impairment of digestive functions. At the cellular level, CDTa induces F-actin network collapse, elimination of the intestinal brush border, and disruption of intercellular junctions. We confirm toxin-dependent re-distribution of Rab11 to enterocytes' apical surface and observe suppression of CDTa phenotypes by a Dominant-Negative form of Rab11 or RNAi of the dedicated Rab11GEF Crag (DENND4). We also report that Calmodulin (Cam) is required to mediate CDTa activity. In parallel, chemical inhibition of the Cam/Calcineurin pathway by Cyclosporin A or FK506 also reduces CDTa phenotypes, potentially opening new avenues for treating CDIs

Effect of oxidative treatment on the secondary structure of decoloured bloodmeal

Bloodmeal can be decoloured using peracetic acid resulting in a material with a pale-yellow colour which only needs sodium dodecyl sulphate, water and triethylene glycol to extrude into a semi-transparent bioplastic. Fourier-transform infrared (FTIR) spectroscopy using Synchrotron light was used to investigate the effect of peracetic acid treatment at various concentrations on the spatial distribution of secondary structures within particles of bloodmeal. Oxidation caused aggregation of helical structures into sheets and acetic acid suppressed sheet formation. Decolouring with peracetic acid led to particles with a higher degree of disorder at the outer edges and higher proportions of ordered structures at the core, consistent with the expected diffusion controlled heterogeneous phase decolouring reaction. The degradation of stabilizing intra- and intermolecular interactions and the presence of acetate ions results in increased chain mobility and greater amorphous content in the material, as evidenced by reduction in Tg and greater enthalpy of relaxation with increasing PAA concentration

Time dependent properties of thermoplastic protein produced from bloodmeal with sodium sulphite as an anti-crosslinking agent

The aim of this research was to investigate how the time dependent mechanical behaviour of bloodmeal-based thermoplastic was affected by varying sodium sulphite content at two injection moulding temperatures (120°C at exit or 140°C at exit). Thermoplastic protein was prepared by extrusion with 2, 3 or 4g sodium sulphite (SS), 3g sodium dodecyl sulphate, 10 g urea, 20 g triethylene glycol and 40 g water per 100 g bloodmeal, then injection moulded into test specimens. Pull to break, creep and stress relaxation tests were performed on conditioned samples and glass transition temperature (Tg) was determined by dynamic mechanical analysis. Ultimate tensile strength was 7.9, 7.6 and 5.6 MPa for samples moulded at 120°C and 7.6, 6.3 and 5.7 MPa when moulded at 140°C with 2, 3 and 4 g SS respectively. Experimental creep data was modelled with a 4 element model, consisting of a spring and dashpot in parallel, in series with an additional spring and dashpot. Plotting creep compliance versus time showed increasing chain mobility as SS content increased. Relaxation was modelled with the Struik equation for short-time experiments. Relaxation times were 530, 360 and 250 s with 2, 3 and 4 g SS respectively when moulded at the lower temperature. At 140°C, relaxation times were 440, 430 and 190 s for these SS contents. Tg was in the range 57-65°C (1 Hz peak in tanδ) for all samples, but was lowest for samples with 4 g SS. These results show that both increased sodium sulphite and the higher moulding temperature increased chain mobility in the processed plastic

Ion-channel-like behavior in lipid bilayer membranes at the melting transition

It is well known that at the gel-liquid phase transition temperature a lipid bilayer membrane exhibits an increased ion permeability. We analyze the quantized currents in which the increased permeability presents itself. The open time histogram shows a "-3/2" power law which implies an open-closed transition rate that decreases like $k(t) \propto t^{-1}$ as time evolves. We propose a "pore freezing" model to explain the observations. We discuss how this model also leads to the $1/f^{\alpha}$ noise that is commonly observed in currents across biological and artificial membranes.Comment: 5 pages, 4 figure

Three and four current reversals versus temperature in correlation ratchets with a simple sawtooh potential

Transport of Brownian particles on a simple sawtooth potential subjected to both unbiased thermal and nonequilibrium symmetric three-level Markovian noise is considered. The new effects of three and four current reversals as a function of temperature are established in such correlation ratchets. The parameter space coordinates of the fixed points associated with these current reversals and the necessary and sufficient conditions for the existence of the novel current reversals are found.Comment: 4 pages, 5 figures; some changes introduced; accepted for publication in Physical Review

The pattern and Loci of training-induced brain changes in healthy older adults are predicted by the nature of the intervention.

There is enormous interest in designing training methods for reducing cognitive decline in healthy older adults. Because it is impaired with aging, multitasking has often been targeted and has been shown to be malleable with appropriate training. Investigating the effects of cognitive training on functional brain activation might provide critical indication regarding the mechanisms that underlie those positive effects, as well as provide models for selecting appropriate training methods. The few studies that have looked at brain correlates of cognitive training indicate a variable pattern and location of brain changes - a result that might relate to differences in training formats. The goal of this study was to measure the neural substrates as a function of whether divided attentional training programs induced the use of alternative processes or whether it relied on repeated practice. Forty-eight older adults were randomly allocated to one of three training programs. In the SINGLE REPEATED training, participants practiced an alphanumeric equation and a visual detection task, each under focused attention. In the DIVIDED FIXED training, participants practiced combining verification and detection by divided attention, with equal attention allocated to both tasks. In the DIVIDED VARIABLE training, participants completed the task by divided attention, but were taught to vary the attentional priority allocated to each task. Brain activation was measured with fMRI pre- and post-training while completing each task individually and the two tasks combined. The three training programs resulted in markedly different brain changes. Practice on individual tasks in the SINGLE REPEATED training resulted in reduced brain activation whereas DIVIDED VARIABLE training resulted in a larger recruitment of the right superior and middle frontal gyrus, a region that has been involved in multitasking. The type of training is a critical factor in determining the pattern of brain activation

Separation quality of a geometric ratchet

We consider an experimentally relevant model of a geometric ratchet in which particles undergo drift and diffusive motion in a two-dimensional periodic array of obstacles, and which is used for the continuous separation of particles subject to different forces. The macroscopic drift velocity and diffusion tensor are calculated by a Monte-Carlo simulation and by a master-equation approach, using the correponding microscopic quantities and the shape of the obstacles as input. We define a measure of separation quality and investigate its dependence on the applied force and the shape of the obstacles