Photocurable Biopolymers for Coaxial Bioprinting.

Thanks to their unique advantages, additive manufacturing technologies are revolutionizing almost all sectors of the industrial and academic worlds, including tissue engineering and regenerative medicine. In particular, 3D bioprinting is rapidly emerging as a first-choice approach for the fabrication-in one step-of advanced cell-laden hydrogel constructs to be used for in vitro and in vivo studies. This technique consists in the precise deposition layer-by-layer of sub-millimetric hydrogel strands in which living cells are embedded. A key factor of this process consists in the proper formulation of the hydrogel precursor solution, the so-called bioink. Ideal bioinks should be able, on the one side, to support cell growth and differentiation and, on the other, to allow the high-resolution deposition of cell-laden hydrogel strands. The latter feature requires the extruded solution to instantaneously undergo a sol-gel transition to avoid its collapse after deposition.To address this challenge, researchers are recently focusing their attention on the synthesis of several derivatives of natural biopolymers to enhance their printability. Here, we present an approach for the synthesis of photocurable derivatives of natural biopolymers-namely, gelatin methacrylate, hyaluronic acid methacrylate, chondroitin sulfate methacrylate, and PEGylated fibrinogen-that can be used to formulate tailored innovative bioinks for coaxial-based 3D bioprinting applications.

Efficient recovery of thermostable polyhydroxybutyrate (PHB) by a rapid and solvent-free extraction protocol assisted by ultrasound.

Nowadays, there are great research interest in polyhydroxybutyrate (PHB) recovery protocols that reduce the use of organic solvents and efficiently recover this bacterial biopolymer. The present study reports an extraction protocol assisted by ultrasound, which is a rapid protocol that increases the amount of polymeric matter extracted, reduces the cellular digestion step with sodium hypochlorite and eliminates the use of organic solvents. Likewise, characterization studies by Fourier transform infrared spectroscopy (FTIR), proton nuclear magnetic resonance spectroscopy (1H NMR), differential scanning calorimetry (DSC), and X-ray diffraction (XRD) demonstrated that the biopolymer recovered by this protocol is PHB homopolymer with a major thermal-resistance and crystalline properties. Besides, in this study are reported the thermic and crystalline differences between the PHB obtained from the fermentation of complex carbohydrates (agavins) and simple sugars. The biopolymer obtained by this rapid extraction protocol would be suitable for ecological and biomedical applications, due to the low melting temperature, less than 50% crystallinity, and the lack of lipopolysaccharides. Therefore, this extraction protocol might represent an alternative to the traditional protocol based on NaOCl-chloroform and is part of the green trend to improve the PHB production.

A strongly adhesive hemostatic hydrogel for the repair of arterial and heart bleeds.

Uncontrollable bleeding is a major problem in surgical procedures and after major trauma. Existing hemostatic agents poorly control hemorrhaging from traumatic arterial and cardiac wounds because of their weak adhesion to wet and mobile tissues. Here we design a photo-reactive adhesive that mimics the extracellular matrix (ECM) composition. This biomacromolecule-based matrix hydrogel can undergo rapid gelling and fixation to adhere and seal bleeding arteries and cardiac walls after UV light irradiation. These repairs can withstand up to 290 mm Hg blood pressure, significantly higher than blood pressures in most clinical settings (systolic BP 60-160 mm Hg). Most importantly, the hydrogel can stop high-pressure bleeding from pig carotid arteries with 4~ 5 mm-long incision wounds and from pig hearts with 6 mm diameter cardiac penetration holes. Treated pigs survived after hemostatic treatments with this hydrogel, which is well-tolerated and appears to offer significant clinical advantage as a traumatic wound sealant.

Luminescent κ-Carrageenan-Based Electrolytes Containing Neodymium Triflate.

In recent years, the synthesis of polymer electrolyte systems derived from biopolymers for the development of sustainable green electrochemical devices has attracted great attention. Here electrolytes based on the red seaweeds-derived polysaccharide κ-carrageenan (κ-Cg) doped with neodymium triflate (NdTrif3) and glycerol (Gly) were obtained by means of a simple, clean, fast, and low-cost procedure. The aim was to produce near-infrared (NIR)-emitting materials with improved thermal and mechanical properties, and enhanced ionic conductivity. Cg has a particular interest, due to the fact that it is a renewable, cost-effective natural polymer and has the ability of gelling in the presence of certain alkali- and alkaline-earth metal cations, being good candidates as host matrices for accommodating guest cations. The as-synthesised κ-Cg-based membranes are semi-crystalline, reveal essentially a homogeneous texture, and exhibit ionic conductivity values 1⁻2 orders of magnitude higher than those of the κ-Cg matrix. A maximum ionic conductivity was achieved for 50 wt.% Gly/κ-Cg and 20 wt.% NdTrif3/κ-Cg (1.03 × 10-4, 3.03 × 10-4, and 1.69 × 10-4 S cm-1 at 30, 60, and 97 °C, respectively). The NdTrif-based κ-Cg membranes are multi-wavelength emitters from the ultraviolet (UV)/visible to the NIR regions, due to the κ-Cg intrinsic emission and to Nd3+, 4F3/2→4I11/2-9/2.

Effects of Gamma and Electron Radiation on the Structural Integrity of Organic Molecules and Macromolecular Biomarkers Measured by Microarray Immunoassays and Their Astrobiological Implications.

High-energy ionizing radiation in the form of solar energetic particles and galactic cosmic rays is pervasive on the surface of planetary bodies with thin atmospheres or in space facilities for humans, and it may seriously affect the chemistry and the structure of organic and biological material. We used fluorescent microarray immunoassays to assess how different doses of electron and gamma radiations affect the stability of target compounds such as biological polymers and small molecules (haptens) conjugated to large proteins. The radiation effect was monitored by measuring the loss in the immunoidentification of the target due to an impaired ability of the antibodies for binding their corresponding irradiated and damaged epitopes (the part of the target molecule to which antibodies bind). Exposure to electron radiation alone was more damaging at low doses (1 kGy) than exposure to gamma radiation alone, but this effect was reversed at the highest radiation dose (500 kGy). Differences in the dose-effect immunoidentification patterns suggested that the amount (dose) and not the type of radiation was the main factor for the cumulative damage on the majority of the assayed molecules. Molecules irradiated with both types of radiation showed a response similar to that of the individual treatments at increasing radiation doses, although the pattern obtained with electrons only was the most similar. The calculated radiolysis constant did not show a unique pattern; it rather suggested a different behavior perhaps associated with the unique structure of each molecule. Although not strictly comparable with extraterrestrial conditions because the irradiations were performed under air and at room temperature, our results may contribute to understanding the effects of ionizing radiation on complex molecules and the search for biomarkers through bioaffinity-based systems in planetary exploration.

Effect of ultrasound treatment during osmotic dehydration on bioactive compounds of cranberries.

The aim of the study was to investigate the influence of ultrasound treatment applied in osmotic solution on bioactive compounds, such as vitamin C, polyphenols, anthocyanins and flavonoids content as well as antioxidant activity in cranberries (Vaccinium oxycoccus). Ultrasound treatment was performed at the frequency of 21kHz for 30 and 60min in two osmotic solutions - 61.5% sucrose and 30% sucrose with an addition of 0.1% of steviol glycosides. Before the ultrasound treatment the material was subjected to cutting or blanching. The obtained results indicated that the influence of ultrasound waves on cranberries depends on a type of bioactive component. The ultrasound treated cranberries as well as the ones subjected to cutting or blanching enhanced by ultrasound were characterized mainly by a lower content of bioactive compounds.

Epitaxy of the bound water phase on hydrophilic surfaces of biopolymers as key mechanism of microwave radiation effects on living objects.

The research investigates the mechanism of microwave radiation effects on biological characteristics and structural-dynamic parameters of a sensor bioluminescence system. The research objects are a sterile growth medium (fish meal hydrolisate) and a bacterial culture. It has been established that irradiation causes changes of the growth medium spectral properties within the range of 200-350nm. Changes take place in the intensity and character of luminescence, as well as in relaxation parameters of nuclear magnetic resonance, growth characteristics of the bacterial culture, its cellular morphology and surface topology. The research results enabled us to establish the mechanisms of primary molecular processes that occur when the bacterial culture is exposed to microwave radiation. Transformation of the dynamic-structural state of adsorbed water phases on biopolymer surfaces has been found to be the key factor in the mechanism of microwave effects on living and water-containing objects.

Temporal control of xyloglucan self-assembly into layered structures by radiation-induced degradation.

Partially degalactosylated xyloglucan from tamarind seeds (Deg-XG) is a very appealing biopolymer for the production of in situ gelling systems at physiological temperature. In this work, we observe that the morphology of hydrogels evolves towards high degrees of structural organization with time, yielding to dense stacks of thin membranes within 24h of incubation at 37°C. We also explore the possibility offered by gamma irradiation of controlling the time scale of this phenomenon, the final morphology and mechanical properties of the system. Structural and molecular modifications of Deg-XG with dose are investigated by FTIR, dynamic light scattering (DLS) and rotational viscosimetry. The impact on gelation ability and gel strength is studied by rheological analysis. The morphology evolution is investigated by SEM analysis, and absence of cytotoxicity verified by MTS assay and optical microscopy of neuroblastoma cells.

High-molecular-weight poly(Gly-Val-Gly-Val-Pro) synthesis through microwave irradiation.

In this study, we synthesized a polypeptide from its pentapeptide unit using microwave irradiation. Effective methods for polypeptide synthesis from unit peptides have not been reported. Here, we used a key elastin peptide, H-GlyValGlyValPro-OH (GVGVP), as the monomer peptide. It is difficult to obtain poly(Gly-Val-Gly-Val-Pro) (poly(GVGVP)) from the pentapeptide unit of elastin, GVGVP, via polycondensation. Poly(GVGVP) prepared from genetically recombinant Escherichia coli is a well-known temperature-sensitive polypeptide, and this temperature sensitivity is known as the lower critical solution temperature. When microwave irradiation was performed in the presence of various additives, the pentapeptide (GVGVP) polycondensation reaction proceeded smoothly, resulting in a product with a high molecular weight in a relatively good yield. The reaction conditions, like microwave irradiation, coupling agents, and solvents, were optimized to increase the reaction efficiency. The product exhibited a molecular weight greater than Mr 7000. Further, the product could be synthesized on a gram scale. The synthesized polypeptide exhibited a temperature sensitivity that was similar to that of poly(GVGVP) prepared from genetically recombinant E. coli. Therefore, this technique offers a facile and quick approach to prepare polypeptides in large amounts. Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd.

Do twisted laser beams evoke nuclear hyperpolarization?

The hyperpolarization of nuclear spins promises great advances in chemical analysis and medical diagnosis by substantially increasing the sensitivity of nuclear magnetic resonance (NMR). Current methods to produce a hyperpolarized sample, however, are arduous, time-consuming or costly and require elaborate equipment. Recently, a much simpler approach was introduced that holds the potential, if harnessed appropriately, to revolutionize the production of hyperpolarized spins. It was reported that high levels of hyperpolarization in nuclear spins can be created by irradiation with a laser beam carrying orbital angular momentum (twisted light). Aside from these initial reports however, no further experimental verification has been presented. In addition, this effect has so far evaded a critical theoretical examination. In this contribution, we present the first independent attempt to reproduce the effect. We exposed a sample of immersion oil or a fluorocarbon liquid that was placed within a low-field NMR spectrometer to Laguerre-Gaussian and Bessel laser beams at a wavelength of 514.5nm and various topological charges. We acquired (1)H and (19)F NMR free induction decay data, either during or alternating with the irradiation that was parallel to B0. We observed an irregular increase in NMR signal in experiments where the sample was exposed to beams with higher values of the topological charge. However, at no time did the effect reach statistical significance of 95%. Given the measured sensitivity of our setup, we estimate that a possible effect did not exceed a hyperpolarization (at 5mT) of 0.14-6%, depending on the assumed hyperpolarized volume. It should be noted though, that there were some differences between our setup and the previous implementation of the experiment, which may have inhibited the full incidence of this effect. To approach a theoretical description of this effect, we considered the interaction of an electron with a plane wave, which is known to be able to induce electronic (e.g. in rubidium) and subsequent nuclear hyperpolarization. Compared to the plane wave, the additional transitions caused by a twisted wave are of the order of 10(-3) less. This suggests that the twist of the laser is unlikely to be responsible for the hyperpolarization of nuclear spins, unless a new mechanism of momentum transfer is identified.

Electron photodetachment dissociation for structural characterization of synthetic and bio-polymer anions.

Tandem mass spectrometry (MS-MS) is a generic term evoking techniques dedicated to structural analysis, detection or quantification of molecules based on dissociation of a precursor ion into fragments. Searching for the most informative fragmentation patterns has led to the development of a vast array of activation modes that offer complementary ion reactivity and dissociation pathways. Collisional activation of ions using atoms, molecules or surface resulting in unimolecular dissociation of activated ions still plays a key role in tandem mass spectrometry. The discovery of electron capture dissociation (ECD) and then the development of other electron-ion or ion/ion reaction methods, constituted a significant breakthrough, especially for structural analysis of large biomolecules. Similarly, photon activation opened promising new frontiers in ion fragmentation owing to the ability of tightly controlled internal energy deposition and easy implementation on commercial instruments. Ion activation by photons includes slow heating methods such as infrared multiple photon dissociation (IRMPD) and black-body infrared radiative dissociation (BIRD) and higher energy methods like ultra-violet photodissociation (UVPD) and electron photo detachment dissociation (EPD). EPD occurs after UV irradiation of multiply negatively charged ions resulting in the formation of oxidized radical anions. The present paper reviews the hypothesis regarding the mechanisms of electron photo-detachment, radical formation and direct or activated dissociation pathways that support the observation of odd and even electron product ions. Finally, the value of EPD as a complementary structural analysis tool is illustrated through selected examples of synthetic polymers, oligonucleotides, polypeptides, lipids, and polysaccharides.

Contribution of synchrotron radiation to photoactivation studies of biomolecular ions in the gas phase.

Photon activation of ions in the visible and ultraviolet range attracts a growing interest, partly for its promising applications in tandem mass spectrometry. However, this task is not trivial, as it requires notably high brilliance photon sources. Hence, most of the work in that field has been performed using lasers. Synchrotron radiation is a source continuously tunable over a wide photon energy range and which possesses the necessary characteristics for ion activation. This review focuses on the array of applications of synchrotron radiation in photon activation of ions ranging from near UV to soft X-rays.

Pressure-controlled motion of single polymers through solid-state nanopores.

Voltage-biased solid-state nanopores are well established in their ability to detect and characterize single polymers, such as DNA, in electrolytes. The addition of a pressure gradient across the nanopore yields a second molecular driving force that provides new freedom for studying molecules in nanopores. In this work, we show that opposing pressure and voltage bias enables nanopores to detect and resolve very short DNA molecules, as well as to detect near-neutral polymers.

Impact of ultrafast electronic damage in single-particle x-ray imaging experiments.

In single-particle coherent x-ray diffraction imaging experiments, performed at x-ray free-electron lasers (XFELs), samples are exposed to intense x-ray pulses to obtain single-shot diffraction patterns. The high intensity induces electronic dynamics on the femtosecond time scale in the system, which can reduce the contrast of the obtained diffraction patterns and adds an isotropic background. We quantify the degradation of the diffraction pattern from ultrafast electronic damage by performing simulations on a biological sample exposed to x-ray pulses with different parameters. We find that the contrast is substantially reduced and the background is considerably strong only if almost all electrons are removed from their parent atoms. This happens at fluences of at least one order of magnitude larger than provided at currently available XFEL sources.

Assembly of Ni(OH)2 nanoparticle films on aqueous surfaces induced by small amounts of toluene, and the study of their unmediated electrocatalytic oxidation toward some small biomolecules.

A simple strategy for the preparation of a Ni(OH)2 nanoparticle film is described. Ni(OH)2 nanoparticles were synthesized in an aqueous solution of Ni2+ and tert-butylamine in the presence of small amounts of toluene, which induced the nanoparticles to assemble a thin film on the aqueous surface. The obtained Ni(OH)2 nanoparticle film was easily transferred onto the electrode surfaces and exhibited stable electrochemical performance. The electrochemical behavior of various small biomolecules, including cysteine, homocysteine, glutathione, histidine, glycine, cystine, methionine, lysine, aspartic acid, glutamic acid, phenylalanine, ascorbic acid, uric acid and dopamine, were studied at the Ni(OH)2 nanoparticle-film-modified electrode. The Ni(OH)2 nanoparticle film exhibits excellent direct, unmediated electrocatalysis toward the oxidation of cysteine, homocysteine and ascorbic acid in a pH 7.4 buffer solution with a low onset potential and a high oxidation signal. This behavior differs from many reports in which small organic molecules are electrocatalyzed indirectly by the Ni(OH)2/NiOOH redox couple in a strongly alkaline solution.

Experimental assessment of the contribution of electrodynamic interactions to long-distance recruitment of biomolecular partners: Theoretical basis.

Highly specific spatiotemporal interactions between cognate molecular partners essentially sustain all biochemical transactions in living matter. That such an exquisite level of accuracy may result from encountering forces solely driven by thermal diffusive processes is unlikely. Here we propose a yet unexplored strategy to experimentally tackle the long-standing question of a possibly active recruitment at a distance of cognate partners of biomolecular reactions via the action of resonant electrodynamic interactions. We considered two simplified models for a preliminary feasibility investigation of the devised methodology. By taking advantage of advanced experimental techniques nowadays available, we propose to measure the characteristic encounter time scales of dually interacting biopartners and to compare them with theoretical predictions worked out in both the presence and absence of putative long-range electromagnetic forces.

Electrodiffusion of molecules in aqueous media: a robust, discretized description for electroporation and other transport phenomena.

Electrically driven transport of molecules and ions within aqueous electrolytes is of long-standing interest, with direct relevance to applications that include the delivery/release of biologically active solutes to/from cells and tissues. Examples include iontophoretic and electroporation-mediated drug delivery. Here, we describe a robust method for characterizing electrodiffusive transport in physiologic aqueous media. Specifically, we treat the case of solute present in sufficiently low concentration as to negligibly contribute to the total ionic current within the system. In this limiting case, which applies to many systems of interest, the predominant electrical behavior due to small ions is decoupled from solute transport. Thus, electrical behavior may be characterized using existing methods and treated as known in characterizing electrodiffusive molecular transport. First, we present traditional continuum equations governing electrodiffusion of charged solutes within aqueous electrolytes and then adapt them to discretized systems. Second, we examine the time-dependent and steady-state interfacial concentration gradients that result from the combination of diffusion and electrical drift. Third, we show how interfacial concentration gradients are related to electric field strength and duration. Finally, we examine how discretization size affects the accuracy of these methods. Overall these methods are motivated by and well suited to addressing an outstanding goal: estimation of the net ionic and molecular transport facilitated by electroporation in biological systems.

Efficacy of UV treatment in the management of bacterial adhesion on hard surfaces.

The efficacy of UV treatment to control bacterial adhesion onto hard surfaces was investigated in laboratory conditions. The major characteristics necessary for biofilm formation like extracellular polymeric substance (EPS) production, carbohydrate and protein concentration in EPS, and adhesion ability onto hard surface were studied using two bacterial strains isolated from marine biofilms. The results showed that there was a considerable difference between the control and UV treated bacterial cultures in their viability, production of EPS, and adhesion ability. The protein and carbohydrate concentration of the EPS and the adhesion of bacterial cells to surface were also considerably reduced due to UV treatment. This study indicates that treatment of water with UV light may be used to control biofilm development on hard surfaces.

Simulation of single-molecule trapping in a nanochannel.

The detection and trapping of single fluorescent molecules in solution within a nanochannel is studied using numerical simulations. As optical forces are insufficient for trapping molecules much smaller than the optical wavelength, a means for sensing a molecule's position along the nanochannel and adjusting electrokinetic motion to compensate diffusion is assessed. Fluorescence excitation is provided by two adjacently focused laser beams containing temporally interleaved laser pulses. Photon detection is time-gated, and the displacement of the molecule from the middle of the two foci alters the count rates collected in the two detection channels. An algorithm for feedback control of the electrokinetic motion in response to the timing of photons, to reposition the molecule back toward the middle for trapping and to rapidly reload the trap after a molecule photobleaches or escapes, is evaluated. While accommodating the limited electrokinetic speed and the finite latency of feedback imposed by experimental hardware, the algorithm is shown to be effective for trapping fast-diffusing single-chromophore molecules within a micron-sized confocal region. Studies show that there is an optimum laser power for which loss of molecules from the trap due to either photobleaching or shot-noise fluctuations is minimized.

Transport in disordered media with spatially nonuniform fields.

The theoretical treatment of transport in a disordered system in the presence of a system-wide force field F(x) or spatially varying macroscopic velocity field v(x) is developed in the framework of continuous time random walk (CTRW). The physical basis of CTRW and related fractional derivative equations relies on a mapping of the aggregate of transition rates w(s,s'), between sites s and s', in the Master equation describing the system kinetics, onto a joint probability distribution function psi(s,t). This distribution is calculated from the ensemble average of a position-dependent functional of w(s,s'); the procedure is effective when the scale of heterogeneities is much smaller than the system size. However, statistical homogeneity does not hold in the presence of large heterogeneities, which control the macroscopic v(x), or in the case of an interaction of F(x) with the transition rates. The transport equation, incorporating large-scale heterogeneity, involves the use of a local ensemble average to obtain a position-dependent psi(s,t;x); this determines a memory function, M(t;x), which is convoluted with the advection-dispersion operator. A prototype transport equation for a system with statistical inhomogeneity is developed as an integrodifferential equation. It is solved numerically for particles migrating with a steady-state Darcy velocity v(x , determined for different permeability fields and boundary conditions. The nature of the solutions as a function of key transport parameters (e.g., a characteristic time tc) is explored, and solutions are also compared to those of the advection-dispersion equation for v(x) and to a laboratory experiment. This transport equation is in contrast to the fractional Fokker-Planck equation, which is based on a decoupling of F(x) or v(x) with the transition rates w(s,s'). Further, an analytic expression for the effect of a variance of the ensemble average on the solution of the CTRW transport equation is derived.