Protein encapsulation via porous CaCO3 microparticles templating

Porous microparticles of calcium carbonate with an average diameter of 4.75 microm were prepared and used for protein encapsulation in polymer-filled microcapsules by means of electrostatic layer-by-layer assembly (ELbL). Loading of macromolecules in porous CaCO3 particles is affected by their molecular weight due to diffusion-limited permeation inside the particles and also by the affinity to the carbonate surface. Adsorption of various proteins and dextran was examined as a function of pH and was found to be dependent both on the charge of the microparticles and macromolecules. The electrostatic effect was shown to govern this interaction. This paper discusses the factors which can influence the adsorption capacity of proteins. A new way of protein encapsulation in polyelectrolyte microcapsules is proposed exploiting the porous, biocompatible, and decomposable microparticles from CaCO3. It consists of protein adsorption in the pores of the microparticles followed by ELbL of oppositely charged polyelectrolytes and further core dissolution. This resulted in formation of polyelectrolyte-filled capsules with protein incorporated in interpenetrating polyelectrolyte network. The properties of CaCO3 microparticles and capsules prepared were characterized by scanning electron microscopy, microelectrophoresis, and confocal laser scanning microscopy. Lactalbumin was encapsulated by means of the proposed technique yielding a content of 0.6 pg protein per microcapsule. Horseradish peroxidase saves 37% of activity after encapsulation. However, the thermostability of the enzyme was improved by encapsulation. The results demonstrate that porous CaCO3 microparticles can be applied as microtemplates for encapsulation of proteins into polyelectrolyte capsules at neutral pH as an optimal medium for a variety of bioactive material, which can also be encapsulated by the proposed method. Microcapsules filled with encapsulated material may find applications in the field of biotechnology, biochemistry, and medicine.     

Computer simulation of a cerebellar cortex compartment

Here, Marr's theory of the cerebellum is elaborated at a detailed cellular level. The experimental grounds for the theory are briefly reviewed and problems related to (mossy-fibres) - (granule cell) coding are formulated. The properties of several particular types of (mossy-fibres) - (granule cell) connection matrices are illustrated through computer simulation.     

Fabrication and Characterization of Polyelectrolyte Microparticles with Protein

The incorporation of proteins into microparticles fabricated by layer-by-layer adsorption of oppositely charged polyelectrolytes (dextran sulfate and protamine) on protein microaggregates was studied. Microaggregates with insulin were prepared by two different techniques: 1) formation of insoluble polyelectrolyte complex consisting of insulin and dextran sulfate (aggregate size of 7-20 micro m), or 2) salting out of insulin from solution by sodium chloride (aggregate size of 5-13 micro m). Microparticles varying in the number of cycles (from 1 to 8) of polyelectrolyte adsorption on protein aggregates were examined and compared. Morphology of the microparticles was studied by scanning electron and optical microscopy. It was shown that polyelectrolyte microparticles retained the shape and dimensions of the initial protein aggregates used as a template. Ultrasonication of microparticles obtained using salted out protein aggregates resulted in the formation of stable nanoparticles (100-200 nm). Regulation of protein release from the microparticles of both types by varying the number of polyelectrolyte adsorption cycles and pH of the medium was demonstrated. Insulin not bound to polyelectrolytes was released from the microparticles at pH values between 6 and 8, which corresponds to the pH of the human small intestine and ileum.     

Interaction of Polymer Aggregates Based on Stearoyl-poly-N-vinylpyrrolidone with Blood Components

Stearoyl-poly-N-vinylpyrrolidone (PVP-stear) of various molecular weights (M(n) = 1500-5500) self-assemble in aqueous medium. Particles prepared from PVP-stear were characterized in terms of shape and size distribution, and the mechanical stability of the particles was studied. The interaction of PVP-stear and its aggregates with blood components was investigated. Aggregates formed by the polymers with M(n) = 1500-3500 in the presence of human serum are stable. The direct lytic action of PVP-stear preparations was studied using sheep and human erythrocytes. The influence of PVP-stear aggregates on the activation of complement system both on classical and alternative pathways was examined. The aggregates prepared from PVP-stear of various molecular weights had no effect on the activation of the complement system.     

Genetic diversity and differentiation of Gmelin larch Larix gmelinii populations from Evenkia (Central Siberia)

Within- and among-population diversity of Gmelin larch Larix gmelinii (Rupr.) Rupr. from Evenkia was inferred from data on 17 genes determining allozyme diversity of ten enzymes. More than 50% of the genes proved to be polymorphic. On average, each tree was heterozygous at 9.2% genes. Heterozygosity expected from the Hardy-Weinberg proportions was higher, 12.5%. A deficit of heterozygous genotypes was observed in all populations under study and attributed to inbreeding. With Wright's F statistics, average individual inbreeding was estimated at 26.6% relative to the population (F(IS)) and at 27.8% relative to the species (F(IT)). The greatest deficit of heterozygosity was observed for the youngest population. Within-population variation accounted for more than 98% of the total variation, while the contribution of among-population variation was 1.66%. Genetic distance between populations varied from 0.0025 to 0.0042, averaging 0.0035.     

An Electroacoustic Analysis for Determining the Effect of Amoxicillin on Microbial Cells

The probability of determining the effects of amoxicillin, which is one of β-lactam antibiotics, on microbial cells of Escherichia coli by the electroacoustic analysis method was shown for the first time. A piezoelectric resonator with a lateral electric field with a 1-mL liquid container was used as a biological sensor. It has been established that in the presence of amoxicillin the frequency dependence of the real and imaginary parts of the electrical impedance of a resonator loaded with a suspension of sensitive cells differs significantly from those of the resonator with a control of a microbial cell suspension without an antibiotic. When the resonator is loaded with the amoxicillin-resistant cell suspension, these dependencies are virtually the same. These results open prospects for the use of electroacoustic analysis methods to register the effect of β-lactam antibiotics on microbial cells and evaluate their antibacterial activity.     

Increase in the efficacy of synapses between parallel fibers and Purkinje cells after joint stimulation of climbing and parallel fibers in the frog

A study was made of the susceptibility of Purkinje cells to long-term plasticity changes produced by joint stimulation of two inputs: the parallel and the climbing fibers. Experiments were conducted on a preparation of isolated frog cerebellum, joined to the medulla by one peduncle. A total of 18 neurons were investigated which showed a monosynaptic response to stimulation of the parallel fibers and maintained stable background activity over a 2 h period. Curves were plotted throughout this time for the likelihood of a reaction occurring in Purkinje cells in response to stimulation of the parallel fibers. Level of current required to stimulate a Purkinje cell firing index of 0.5 (I_0.5) was calculated. Neurons in which compound response to the "climber" type had been produced by stimulating the medulla showed a I_0.5 of 0.7 (less than one unit) at the start and finish of experiments, which would suggest an increase in the efficacy of the synapses of parallel fibers in Purkinje cells when parallel and climbing fibers are stimulated simultaneously.Institute of Problems in Information Transmission, Academy of Sciences of the USSR, Moscow. Translated from Neirofiziologiya, Vol. 19, No. 2, pp. 156–164, March–April, 1987.     

Titration of active centers of enzymes by means of reversible inhibitors. Dipeptidyl-carboxypeptidase - inhibitor SQ 20881 from venom of the snake Bothrops jararca)

The essentials of estimation of the number of enzyme active sites by reversible inhibition are discussed. The necessity of evaluation of the substrate effect on the equilibrium of the systems with a rapidly dissociating enzyme -- inhibitor complex has been demonstrated. Some procedures for determination of the number of active sites of dipeptidyl-carboxypeptidase (EC 3.4.15.1) from bovine kidney cortex, using the competitive inhibitor SQ 20 881 (Glu-Trp-Pro-Arg-Pro-Gln-Ile-Pro-Pro) have been developed. The kinetic and equilibrium constants for the enzyme-inhibitor interaction (ki = 3.2 . 10(6) M-1s-1, k-i = 8 ms-1 and Ki = 2.5 +/- 0.5 nm) have been calculated.