Recent advances and challenges in the recovery and purification of cellular exosomes

Exosomes are nanovesicles secreted by most cellular types that carry important biochemical compounds throughout the body with different purposes, playing a preponderant role in cellular communication. Because of their structure, physicochemical properties and stability, recent studies are focusing in their use as nanocarriers for different therapeutic compounds for the treatment of different diseases ranging from cancer to Parkinson's disease. However, current bioseparation protocols and methodologies are selected based on the final exosome application or intended use and present both advantages and disadvantages when compared among them. In this context, this review aims to present the most important technologies available for exosome isolation while discussing their advantages and disadvantages and the possibilities of being combined with other strategies. This is critical since the development of novel exosome‐based therapeutic strategies will be constrained to the effectiveness and yield of the selected downstream purification methodologies for which a thorough understanding of the available technological resources is needed.     

Non-isothermal short-range-order kinetics of binary alloys as influenced by solute-vacancy complexes

A model describing the roles of bound and unbound vacancies is proposed in order to predict defect decay and short-range-order kinetics of quenched binary alloys during linear heating experiments. This is an alternative treatment of a previous approach. The model has been applied to the differential scanning calorimetry (DSC) curves of Cu-5 at.% Zn quenched from different temperatures. An expression to calculate the activation energy for migration of solute-vacancy complexes was also developed which make use of DSC trace data. A value of 89.12±0.32 kJ mol^-1 was obtained for the above alloy. The relative contribution of bound and unbound vacancies to partition of effective activation energy corresponding to the ordering process as influenced by quenching temperature was also assessed. This revised version was published online in July 2006 with corrections to the Cover Date.     

Fleet configuration subject to stochastic demand: an application in the distribution of liquefied petroleum gas

We study the problem of configuring a fleet, in which vehicles receive information on-line about the demand that they should fulfil while they are on the road. In each district it must be decided the number of vehicles and their capacity. The objective function is to minimise the operational cost subject to constraints for the minimum delivery capacity, the maximum vehicle size and the average waiting time for customers. The last constraint is modelled as a queuing system that is adjusted according to the simulation of the delivery process of a Chilean company that distributes liquefied petroleum gas in portable cylinders. We provide the analytical form of all the components of the model, so it can be solved using a standard non-linear programming package. We show that the fleet may increase its sales by 3% and reduce the waiting time of customers 10% by allowing a set of vehicles to share the buffer of orders rather than having vehicles to exclusively serve smaller sectors.     

The oscillopolarographic determination of uranium and thorium in supporting electrolytes containing cupferron

Uranium(VI and IV) and thorium(IV) give cathodic indentations in supporting electrolytes prepared from 0.1M perchloric acid, 0.5 M ammonium thiocyanate and 5·10^-3M cupferron (solution A) or from 0.1 M succinic-succinate buffer pH 4, 0.1 M sodium chloride, 10^-3M cupferron and 0.05% gelatine (solution B). The uranium indentation on the dE/dt = f(E) curve (Q=0.75 and 0.73) permits its detection at the 3·10^-7M level. The thorium indentation (Q=0.78) permits its detection at the 4·10^-7M level in solution B. Methods for the elimination of interfering ions for the uranium determination are described. In the determination of thorium, Ga(III), Fe(III), Ti(IV) and U(VI) interfere.     

SOLVENT EFFECTS ON THE TAUTOMERIZATION OF 5′-DEOXYPYRIDOXAL. A PHOTOPHYSICAL STUDY

Abstract— Absorption and fluorescence spectra of 5′-deoxypyridoxal (DPL) in various pure solvents and mixtures were recorded both at room temperature and over the range10–65°C. The areas under the absorption bands were analyzed to obtain the mole fraction (f_N, f_z) of two tautomers (the zwitterionic, Z, and neutral, N, forms) in the ground state. The following spectral parameters were determined from the fluorescence spectra: Stokes shift (Δv), fluorescence quantum yield of the neutral form (Q_N), fluorescence ratio of the neutral to the zwitterionic form (ø_N/ø_Z) and the rate constant of tautomerization (k₁) from Z to N in the excited state. Some of these parameters (f_N, Δv, Q_N, k₁) were found to depend on the proton donor character of the solvent, whereas others (ø_N/ø_Z) depended on its dipole moment. Thus, the absorption and fluorescence spectra of DPL allow one to obtain information on the polarity and the concentration of –OH groups on its environment.     

Thermal behavior of short-range-order in quenched Cu-12 at% Mn assessed by DSC

Summary The ordering behavior of quenched Cu-12 at% Mn was investigated by differential scanning calorimetry (DSC) under increasing temperature conditions. The ordering process can be better explained in terms of a homogeneous short-range-order (SRO) rather than a disperse-short-range-order (DSRO) model as for concentrated Cu-γMn solid solutions. At the employed high quench rates ordering is established in one stage here termed stage 1, assisted by excess vacancies. This stage obeys a first order kinetics law, being the effective activation energy consistent with a weighed average sum of the activation energy for migration of free and bound vacancies. An estimated solute-vacancy binding energy of 15 kJ mol^-1 seems quite reasonable for this alloy together with the assessed activation energy for complex migration of 92.6 kJ mol^-1. By adopting a first SRO order parameter based in quasi-chemical theory in pair approximation and using boundary values calculated from displayed features of DSC traces, the retained degree of quenched-in order at room temperature was calculated. This procedure also enabled to estimate an ordering energy of -2.7 kJ mol^-1. The effect of quenching temperature demonstrate that for smaller vacancy sink densities, the retained degree of order at room temperature goes through a minimum.     

Degradation of Cephaloridine on Alkaline Hydrolysis

A kinetic study on the alkaline hydrolysis of cephaloridine ( 1 ) at pH 10.5 and 37° was carried out using ion-pair reversed-phase HPLC. The main resulting degradation products, the 7-epimer 2 of 1 , the Δ²-isomer 3 of 1 , and the 3-methylidene compound 4 were identified. The presence of a pyridinio group at C(3¹) results in a slightly increased formation constant for the 3-methylidene compound 4 and the 7-epimer 2, and introduces a new reaction: the isomerization of the double bond at C(3) in the dihydrothiazine ring to C(2).     

V2O5/TiO2 Catalytic Xerogels Raman and EPR Studies

Raman spectroscopy and Electron Paramagnetic Resonance (EPR) studies were performed on a series of V₂O₅/TiO₂ catalysts prepared by a modified sol-gel method in order to identify the vanadium species. Two species of surface vanadium were identified by Raman measurements, monomeric vanadyls and polymeric vanadates. Monomeric vanadyls are characterized by a narrow Raman band at 1030 cm^–1 and polymeric vanadates by two broad bands in the region from 900 to 960 cm^–1 and 770 to 850 cm^–1. The Raman spectra do not exhibit characteristic peaks of crystalline V₂O₅. These results are in agreement with those of X-ray Diffractometry (XRD) and Fourier Transform Infrared (FT-IR) previously reported (C.B. Rodella et al., J. Sol-Gel Sci. Techn., submitted). At least three families of V⁴⁺ ions were identified by EPR investigations. The analysis of the EPR spectra suggests that isolated V⁴⁺ ions are located in sites with octahedral symmetry substituting for Ti⁴⁺ ions in the rutile structure. Magnetically interacting V⁴⁺ ions are also present as pairs or clusters giving rise to a broad and structureless EPR line. At higher concentration of V₂O₅, a partial oxidation of V⁴⁺ to V⁵⁺ is apparent from the EPR results.     

Target-templated de novo design of macrocyclic d-/l-peptides: discovery of drug-like inhibitors of PD-1

Peptides are a rapidly growing class of therapeutics with various advantages over traditional small molecules, especially for targeting difficult protein–protein interactions. However, current structure-based methods are largely limited to natural peptides and are not suitable for designing bioactive cyclic topologies that go beyond natural l-amino acids. Here, we report a generalizable framework that exploits the computational power of Rosetta, in terms of large-scale backbone sampling, side-chain composition and energy scoring, to design heterochiral cyclic peptides that bind to a protein surface of interest. To showcase the applicability of our approach, we developed two new inhibitors (PD-i3 and PD-i6) of programmed cell death 1 (PD-1), a key immune checkpoint in oncology. A comprehensive biophysical evaluation was performed to assess their binding to PD-1 as well as their blocking effect on the endogenous PD-1/PD-L1 interaction. Finally, NMR elucidation of their in-solution structures confirmed our de novo design approach.

In silico design of heterochiral cyclic peptides that bind to a specific surface patch on the target protein (PD-1, in this case) and disrupt protein–protein interactions.     

4‐(2‐Hydroxyphenyl)‐2‐phenyl‐2,3‐dihydro‐1H‐1,5‐benzodiazepine and the 2‐(2,3‐dimethoxyphenyl)‐, 2‐(3,4‐dimethoxyphenyl)‐ and 2‐(2,5‐dimethoxyphenyl)‐substituted derivatives

The 1,5‐benzodiazepine ring system exhibits a puckered boat‐like conformation for all four title compounds [4‐(2‐hydroxyphenyl)‐2‐phenyl‐2,3‐dihydro‐1H‐1,5‐benzodiazepine, C₂₁H₁₈N₂O, (I), 2‐(2,3‐dimethoxyphenyl)‐4‐(2‐hydroxyphenyl)‐2,3‐dihydro‐1H‐1,5‐benzodiazepine, C₂₃H₂₂N₂O₃, (II), 2‐(3,4‐dimethoxyphenyl)‐4‐(2‐hydroxyphenyl)‐2,3‐dihydro‐1H‐1,5‐benzodiazepine, C₂₃H₂₂N₂O₃, (III), and 2‐(2,5‐dimethoxyphenyl)‐4‐(2‐hydroxyphenyl)‐2,3‐dihydro‐1H‐1,5‐benzodiazepine, C₂₃H₂₂N₂O₃, (IV)]. The stereochemical correlation of the two C₆ aromatic groups with respect to the benzodiazepine ring system is pseudo‐equatorial–equatorial for compounds (I) (the phenyl group), (II) (the 2,3‐dimethoxyphenyl group) and (III) (the 3,4‐dimethoxyphenyl group), while for (IV) (the 2,5‐dimethoxyphenyl group) the system is pseudo‐axial–equatorial. An intramolecular hydrogen bond between the hydroxyl OH group and a benzodiazepine N atom is present for all four compounds and defines a six‐membered ring, whose geometry is constant across the series. Although the molecular structures are similar, the supramolecular packing is different; compounds (I) and (IV) form chains, while (II) forms dimeric units and (III) displays a layered structure. The packing seems to depend on at least two factors: (i) the nature of the atoms defining the hydrogen bond and (ii) the number of intermolecular interactions of the types O—H...O, N—H...O, N—H...π(arene) or C—H...π(arene).