A new pulsed electric field microreactor: comparison between the laboratory and microtechnology scale

This paper presents a new microreactor dedicated for pulsed electric field treatment (PEF), which is a pasteurization method that inactivates microorganisms with short electric pulses. The PEF microreactor consists of a flow-through channel with a constriction where the electric field is focussed. Compared to a laboratory-scale setup 25 times lower voltages were needed to obtain the same electric field strength due to the close electrode spacing. A finite element model showed that the electric field intensity is very homogeneous throughout the channel, which is crucial for the pasteurization processes. Experiments where artificial vesicles, loaded with carboxyfluorescein, were electroporated showed that the maximum transmembrane potential adequately described the processes both in the microreactor and the laboratory-scale setup, although the length scales are different. Electroporation started at a transmembrane potential of 0.5 V, reaching a maximum fraction of electroporated vesicles of 51% at a transmembrane potential of 1.5 V. The partial electroporation is not a result of the heterogenity of the vesicles or the electric field. With this new PEF microreactor it is possible to study the PEF process in more detail.     

"Clickphine": a novel and highly versatile P,N ligand class via click chemistry

[Structure: see text] A novel P,N-type ligand family (ClickPhine) is disclosed that is easily accessible using the Cu(I)-catalyzed azide-alkyne "click" cycloaddition. A diverse set of ligands was made in just three steps from readily available starting materials to give several homogeneous and a heterogeneous catalyst. Preliminary experiments show the efficacy of these ligands in the Pd-catalyzed allylic alkylation reaction.     

Conformational behavior of basic monomeric building units of glycosaminoglycans: isolated systems and solvent effect

Our work reports in detail the results of systematic large-scale theoretical investigations of the glycosaminoglycan building units 1-OMe DeltaIdoA-2SNa2 (1; 2H1 and 1H2 forms), 1-OMe GlcN-S6SNa2 (2), 1,4-DiOMe GlcNa (3), 1,4-DiOMe GlcN-S3S6SNa3 (4), 1,4-DiOMe IdoA-2SNa2 (5; 4C1, 1C4, and 2So conformations), and 1,4-DiOMe GlcN-S6SNa2 (6) at the BP86/TZ2P level of the density functional theory. The optimized geometries indicate that the most stable structure of these monomeric units in the neutral state is stabilized via "multifurcated" sodium bonds. The equilibrium structure of the biologically active anionic forms of the glycosaminoglycans studied changed considerably in a water solution. The computed interaction energies, DeltaE, of sodium coordinated systems 1-6 are negative and span a rather broad energy interval (from -130 to -590 kcal mol-1). Computations that include the effect of solvation indicated that in water the relative stability of Na+...ligand ionic bonds is considerably diminished. The computed interaction energy in water is small (from -20 to -53 kcal mol-1) and negative, that is, stabilizing.     

Catalyzed Peptide Bond Formation in the Gas Phase. Role of Bivalent Cations and Water in Formation of 2-Aminoacetamide from Ammonia and Glycine and in Dimerization of Glycine

The formation of 2-aminoacetamide from ammonia and glycine and N-glycylglycine from two glycine molecules with and without Mg²⁺, Cu²⁺, and Zn²⁺ cations as catalysts have been studied as model reactions for peptide bond formation using the B3LYP functional with 6–311+G(d,p) and 6–31G(d) basis sets. The B3LYP method was also used to characterize the nine gas–phase complexes of neutral glycine, its amide (2-aminoacetamide), and N-glycylglycine with Lewis acids Mg²⁺, Cu²⁺, and Zn²⁺, respectively. Further, the gas-phase hydration of metal-coordinated complexes of glycine, 2-aminoacetamide, and N-glycylglycine was also investigated. Finally, the effect of water on the structure and reactivity of the metal coordinated complexes was determined. Enthalpies and Gibbs energies for the stationary points of each reaction have been calculated to determine the thermodynamics of the reactions investigated. A substantial decrease in reaction enthalpies and Gibbs energies was found for glycine–ammonia and glycine–glycine reactions coordinated by Mg²⁺, Cu²⁺, and Zn²⁺ ions compared to those of the uncoordinated 2-aminoacetamide bond formation. The formation of a dipeptide is a more exothermic process than the creation of simple 2-aminoacetamide from glycine. The energetic effect of the transition metal ions Cu²⁺ and Zn²⁺ is of similar strength and more pronounced than that of the Mg²⁺ cation. The basicity order of the amides investigated shows the order: NH₂CH₂CO₂H < NH₂CH₂CONH₂ < NH₂CH₂CONHCH₂CO₂H. Interaction enthalpies and Gibbs energies of metal ion–amide complexes increase as Mg²⁺2+2+. In both reactant (glycine) and reaction products (2-aminoacetamide, N-glycylglycine) dihydration caused considerable reduction (about 200–500 kJ-mol^–1) of the strength of the bifurcated metal–amide bonds. Solvent effects also reduce the reaction enthalpy and Gibbs energy of reactions under study.     

Effect of metal ions (Li+, Na+, K+, Mg2+, Ca2+, Ni2+, Cu2+, and Zn2+) and water coordination on the structure of glycine and zwitterionic glycine

Interactions between metal ions and amino acids are common both in solution and in the gas phase. Here, the effect of metal ions and water on the structure of glycine is examined. The effect of metal ions (Li+, Na+, K+, Mg2+, Ca2+, Ni2+, Cu2+, and Zn2+) and water on structures of Gly.Mn+(H2O)m and GlyZwitt.Mn+(H2O)m (m = 0, 2, 5) complexes have been determined theoretically by employing the hybrid B3LYP exchange-correlation functional and using extended basis sets. Selected calculations were carried out also by means of CBS-QB3 model chemistry. The interaction enthalpies, entropies, and Gibbs energies of eight complexes Gly.Mn+ (Mn+ = Li+, Na+, K+, Mg2+, Ca2+, Ni2+, Cu2+, and Zn2+) were determined at the B3LYP density functional level of theory. The computed Gibbs energies DeltaG degrees are negative and span a rather broad energy interval (from -90 to -1100 kJ mol(-1)), meaning that the ions studied form strong complexes. The largest interaction Gibbs energy (-1076 kJ mol(-1)) was computed for the NiGly2+ complex. Calculations of the molecular structure and relative stability of the Gly.Mn+(H2O)m and GlyZwitt.Mn+(H2O)m (Mn+ = Li+, Na+, K+, Mg2+, Ca2+, Ni2+, Cu2+, and Zn2+; m = 0, 2, and 5) systems indicate that in the complexes with monovalent metal cations the most stable species are the NO coordinated metal cations in non-zwitterionic glycine. Divalent cations Mg2+, Ca2+, Ni2+, Cu2+, and Zn2+ prefer coordination via the OO bifurcated bonds of the zwitterionic glycine. Stepwise addition of two and five water molecules leads to considerable changes in the relative stability of the hydrated species. Addition of two water molecules at the metal ion in both Gly.Mn+ and GlyZwitt.Mn+ complexes reduces the relative stability of metallic complexes of glycine. For Mn+ = Li+ or Na+, the addition of five water molecules does not change the relative order of stability. In the Gly.K+ complex, the solvation shell of water molecules around K+ ion has, because of the larger size of the potassium cation, a different structure with a reduced number of hydrogen-bonded contacts. This results in a net preference (by 10.3 kJ mol(-1)) of the GlyZwitt.K+H2O5 system. Addition of five water molecules to the glycine complexes containing divalent cations Mg2+, Ca2+, Ni2+, Cu2+, and Zn2+ results in a net preference for non-zwitterionic glycine species. The computed relative Gibbs energies are quite high (-10 to -38 kJ mol(-1)), and the NO coordination is preferred in the Gly.Mn+(H2O)5 (Mn+ = Mg2+, Ca2+, Ni2+, Cu2+, and Zn2+) complexes over the OO coordination.     

Porous microcapsule formation with microsieve emulsification

A simple route is presented to prepare core-shell Eudragit microcapsules through a solvent extraction method with the use of microsieve emulsification. Droplets from a solution of Eudragit FS 30D (a commercial copolymer of poly(methyl acrylate-co-methyl methacrylate-co-methacrylic acid) 7:3:1) and hexadecane in dichloromethane are dispersed into water, using a micro-engineered membrane with well-defined pores, in a cross-flow setting. The dichloromethane is extracted from the droplets, which induces demixing in the droplets, leading to a hexadecane-rich core, and an Eudragit-rich shell. The obtained microcapsules have a narrow size distribution due to the microsieve emulsification process. The capsules have a porous shell as shown by SEM and AFM measurements. Their porosity and pore size is dependent on the ratios of Eudragit and hexadecane in the dispersed phase. At pH 7.1 and above Eudragit (FS 30D) dissolves in water; this pH change is used to release the contents of the microcapsule.     

Perindoprilat monohydrate

The title compound [systematic name: (1S)‐2‐((S)‐{1‐[(2S,3aS,7aS)‐2‐carboxyoctahydro‐1H‐indol‐1‐yl]‐1‐oxopropan‐2‐yl}azaniumyl)pentanoate monohydrate], C₁₇H₂₈N₂O₅·H₂O, (I)·H₂O, the active metabolite of the antihypertensive and cardiovascular drug perindopril, was obtained during polymorphism screening of perindoprilat. It crystallizes in the chiral orthorhombic space group P2₁2₁2₁, the same as the previously reported ethanol disolvate [Pascard, Guilhem, Vincent, Remond, Portevin & Laubie (1991). J. Med. Chem. 34 , 663–669] and dimethyl sulfoxide hemisolvate [Bojarska, Maniukiewicz, Sieroń, Fruziński, Kopczacki, Walczyński & Remko (2012). Acta Cryst. C 68 , o341–o343]. The asymmetric unit of (I)·H₂O contains one independent perindoprilat zwitterion and one water molecule. These interact via strong hydrogen bonds to give a cyclic R²₂(7) synthon, which provides a rigid molecular conformation. The geometric parameters of all three forms are similar. The conformations of the perhydroindole group are almost identical, but the n‐alkyl chain has conformational freedom. A three‐dimensional hydrogen‐bonding network of O—H...O and N—H...O interactions is observed in the crystal structure of (I)·H₂O, similar to the other two solvates, but because of the presence of different solvents the three crystal structures have diverse packing motifs. All three solvatomorphs are additionally stabilized by nonclassical weak C—H...O contacts.     

Characterization of β-Galactosidase Isoforms from Bacillus circulans and Their Contribution to GOS Production

A β-galactosidase preparation from Bacillus circulans consists of four isoforms called β-gal-A, β-gal-B, β-gal-C, and β-gal-D. These isoforms differ in lactose hydrolysis and galacto-oligosaccharide (GOS) synthesis at low substrate concentrations. For this reason, using a selection of the isoforms may be relevant for GOS production, which is typically done at high substrate concentrations. At initial lactose concentrations in between 0.44 % and 0.68 % (w/w), β-gal-A showed the least oligosaccharide formation, followed by β-gal-B and β-gal-C; most oligosaccharides were formed by β-gal-D. The differences in behavior were confirmed by studying the thermodynamics of lactose conversion with isothermal titration calorimetry since especially β-gal-A showed a different profile than the other isoforms. Also during the conversion of allolactose and 4-galactosyllactose at 0.44 % and 0.61 % (w/w), respectively, β-gal-A and β-gal-D showed clear differences. In contrast to above findings, the selectivity of the isoforms did hardly differ at an initial lactose concentration of 30 % (w/w), except for a slightly higher production of galactose with β-gal-A. These differences were hypothesized to be related to the different accessibility of the active sites of the isoforms for different-sized reactants. The initial GOS formation rates of the isoforms indicate that β-gal-A and β-gal-B are the best isoforms for GOS production at high lactose concentrations.     

Comparison of activated chromatography resins for protein immobilization

The adsorption of bovine serum albumin (BSA) to an immobilized camelid‐derived antibody fragment was investigated using six different activated resins, of which two are prototypes. The resins differed in base material, coupling chemistry and particle size. The adsorption, washing and desorption stage of the affinity chromatography process were taken into account. Dynamic binding capacities at 10% breakthrough ranged between 0.76 and 4.8 mg BSA/mL resin. The washing volume ranged between 2.9 and 10 column volumes. One of the resins did not concentrate BSA, while the highest concentration was 13‐fold. We present a method to rank and weigh the properties of the resins to find the optimal resin to meet specific requirements. For three of the resins the adsorption flow rate was varied, while the washing and desorption flow rate was kept the same. The dynamic binding capacity decreased with increasing flow rate, as expected. For one resin, the washing volume remained constant, but for the others it decreased with increasing adsorption flow rate. The number of column volumes required to purify a given amount of BSA increases with increasing flow rate, which indicates that higher flow rates do not necessarily speed up the process.