Scalable Production of Monodisperse Bioactive Spider Silk Nanowires

Elongated protein-based micro- and nanostructures are of great interest for a wide range of biomedical applications, where they can serve as a backbone for surface functionalization and as vehicles for drug delivery. Current production methods for protein constructs lack precise control of either shape and dimensions or render structures fixed to substrates. This work demonstrates production of recombinant spider silk nanowires suspended in solution, starting with liquid bridge induced assembly (LBIA) on a substrate, followed by release using ultrasonication, and concentration by centrifugation. The significance of this method lies in that it provides i) reproducability (standard deviation of length <13% and of diameter <38%), ii) scalability of fabrication, iii) compatibility with autoclavation with retained shape and function, iv) retention of bioactivity, and v) easy functionalization both pre- and post-formation. This work demonstrates how altering the function and nanotopography of a surface by nanowire coating supports the attachment and growth of human mesenchymal stem cells (hMSCs). Cell compatibility is further studied through integration of nanowires during aggregate formation of hMSCs and the breast cancer cell line MCF7. The herein-presented industrial-compatible process enables silk nanowires for use as functionalizing agents in a variety of cell culture applications and medical research.     

Alkynyl triazenes enable divergent syntheses of 2-pyrones

The 2-pyrone motif occurs frequently in bioactive natural products and is appreciated as synthetic intermediates. However, only few methods allow for diversifying functional group modifications on this relevant heterocycle. The distinct properties of 1-alkynyl triazenes promote a smooth addition of propiolic acids across the triple bond. Addition of catalytic amounts of silver salt induces cyclization to 2-pyrones. Depending on the reaction temperature, either 6-triazenyl or 5-triazenyl 2-pyrones are selectively formed. The triazenyl unit is subsequently replaced by a variety of valuable groups in a one-pot process yielding for instance 2-fluoro pyrones. The substitution occurs with an intriguing 1,5-carbonyl transposition. Moreover, the triazenyl group serves as traceless activating group for subsequent Diels–Alder cycloadditions and as a constituting unit for rare fused aminopyrazole pyrone heterocycles.

Substituted 2-pyrones are accessed in a silver-catalyzed one-pot process between propiolic acids and 1-alkynyl triazenes. Subsequent diversifying replacement of the triazenyl unit under 1,5-carbonyl transposition yields rare 2-fluoro pyrones.     

Particles trapped at the droplet interface in water-in-water emulsions

Water-in-water emulsions were formed by mixing incompatible aqueous solutions of dextran and poly(ethylene oxide) (PEO) in the presence of latex or protein particles. It was found that particles with a radius as small as 0.1 μm become trapped at the interface between the PEO- and dextran-rich phases with interfacial tensions down to 10(-6) N/m. The particles were visualized at the interface of the emulsion droplets using confocal laser scanning microscopy (CLSM) allowing determination of the contact angle. Various degrees of coverage with particles could be observed. On densely covered droplets, the particles had a hexagonal crystalline order. At intermediate coverage, transient clustering of the particles was observed. The diffusion coefficient of the particles at the interface was determined using multiparticle tracking. Fusion of droplets was observed in all cases leading eventually to macroscopic phase separation.     

Synthesis and Characterization of a Bio-Based Resin from Linseed Oil

Summary: The use of renewable raw materials in the polymer industries is becoming increasingly popular because of environmental concerns and the need to substitute fossil resources. Plant oils with triglyceride backbones can be chemically modified and used to synthesize polymers from renewable resources (biopolymers). In the present study, linseed oil was epoxidized using a chemo-enzymatic method based on Candida Antarctica lipase B (CALB) as a biocatalyst and the modified linseed oil was cured using maleinated linseed oil and a commercial polyamide resin. The amount of epoxidation achieved depended on the amount of lipase used and was determined by infrared (IR) and nuclear magnetic resonance (NMR) spectroscopies. With 20% (weight per weight) catalyst concentration based on the wt % of oil a degree of epoxidation of > 90% was achieved. The cross-linking reaction of epoxidized linseed oil with the maleinated linseed oil and the polyamide resin was studied using differential scanning calorimetry (DSC). DSC traces showed that an increase in epoxidation degree lead to larger values for the exothermic enthalpy integrals of the curing reactions and hence to a higher reactivity of the linseed oil towards the cross-linking agents.     

Lactosylated casein phosphopeptides as specific indicators of heated milks

Casein phosphopeptides (CPP) were identified in small amounts in milks heated at various intensities by using matrix-assisted laser desorption/ionization (MALDI) time-of-flight mass spectrometry. CPP selectively concentrated on hydroxyapatite (HA) were regenerated using phosphoric acid mixed in the matrix. Unphosphorylated peptides not retained by HA were removed by buffer washing. This procedure enhanced the MALDI signals of CPP that are ordinarily suppressed by the co-occurrence of unphosphorylated peptides. CPP, belonging to the β-casein (CN) family, i.e., (f1-29) 4P, (f1-28) 4P, and (f1-27) 4P, and the α_s2-CN family, i.e., (f1-21) 4P and (f1-24) 4P, were observed in liquid and powder milk. The lactosylated counterparts were specific to intensely heated milks, but absent in raw and thermized/pasteurized milk. Most CPP with C-terminal lysines probably arose from the activity of plasmin; an enzyme most active in casein hydrolysis. A CPP analogue was used as the internal standard. The raw milk signature peptide β-CN (f1-28) 4P constituted ~4.3% of the total β-CN. Small amounts of lactosylated peptides, which varied with heat treatment intensity, were detected in the milk samples. The limit of detection of ultra-high-temperature milk adjunction in raw or pasteurized milk was ~10%.     

Laser-induced microexplosion confined in the bulk of a sapphire crystal: evidence of multimegabar pressures

Extremely high pressures (approximately 10 TPa) and temperatures (5 x 10(5) K) have been produced using a single laser pulse (100 nJ, 800 nm, 200 fs) focused inside a sapphire crystal. The laser pulse creates an intensity over 10(14) W/cm2 converting material within the absorbing volume of approximately 0.2 microm3 into plasma in a few fs. A pressure of approximately 10 TPa, far exceeding the strength of any material, is created generating strong shock and rarefaction waves. This results in the formation of a nanovoid surrounded by a shell of shock-affected material inside undamaged crystal. Analysis of the size of the void and the shock-affected zone versus the deposited energy shows that the experimental results can be understood on the basis of conservation laws and be modeled by plasma hydrodynamics. Matter subjected to record heating and cooling rates of 10(18) K/s can, thus, be studied in a well-controlled laboratory environment.     

Harmonic entanglement with second-order nonlinearity

We investigate the second-order nonlinear interaction as a means to generate entanglement between fields of differing wavelengths and show that perfect entanglement can, in principle, be produced between the fundamental and second-harmonic fields in these processes. Neither pure second-harmonic generation nor parametric oscillation optimally produce entanglement; such optimal entanglement is rather produced by an intermediate process.     

Flocculation and percolation in reversible cluster-cluster aggregation

Off-lattice dynamic Monte-Carlo simulations were done of reversible cluster-cluster aggregation for spheres that form rigid bonds at contact. The equilibrium properties were found to be determined by the life time of encounters between two particles (t_e). t_e is a function not only of the probability to form or break a bond, but also of the elementary step size of the Brownian motion of the particles. In the flocculation regime the fractal dimension of the clusters is d_f=2.0 and the size distribution has a power law decay with exponent τ=1.5. At larger values of t_e transient gels are formed. Close to the percolation threshold the clusters have a fractal dimension d_f=2.7 and the power law exponent of the size distribution is τ=2.1. The transition between flocculation and percolation occurs at a characteristic weight average aggregation number that decreases with increasing volume fraction.