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J N Barbotin

Publications and source records attributed to J N Barbotin.

53 records · Page 3Linked to original sources
PubMed1986

Plasmid inheritability and biomass production: comparison between free and immobilized cell cultures of Escherichia coli BZ18(pTG201) without selection pressure.

Maintenance of the plasmid pTG201 in Escherichia coli BZ18 was studied for both free and immobilized cells during chemostat culture, in the absence of the antibiotic against which resistance was plasmid encoded. Electron microscopic observations of immobilized proliferant cells within carrageenan gel beads showed high cell concentrations and growth into distinct cavities. The plasmid which coded for the catechol 2,3-dioxygenase activity was stably maintained during 80 generations in the case of immobilized cells. A theoretical analysis founded on the compartmentalization resulting from the immobilized growth conditions was described. However, the model still showed a plasmid stability inferior to that determined experimentally. Hypotheses dealing with physiological changes of immobilized cells were presented. In addition, the high cell concentrations obtained in the outer 50 microns of the carrageenan gel beads gave a biomass productivity within this useful volume which was 20 times higher than in free-cell cultures.

Bacteriological Techniques↗
PubMed1984

The effects of oxygen solubility and high concentrations of salts on photosynthetic electron transport in chloroplast membranes.

Chloroplast membranes were isolated in different media containing Hepes [4-(2-hydroxyethyl)-1-piperazine-ethanesulphonic acid] and high concentrations of sorbitol (0.33 M), potassium citrate (0.75 M) or Na2SO4 (1.0 M). Due to the complexity of the media, the oxygen solubility is strongly modified by high concentrations of salts (oxygen solubility for 0.33 M-sorbitol, 0.21 mmol/litre; for 0.75 M-potassium citrate, 0.121 mmol/litre; and for 1.0 M-Na2SO4, 0.112 mmol/litre). The knowledge of these values is necessary to interpret the rate of O2 evolution. For thylakoids isolated in 'sorbitol buffer' and then tested in high concentrations of potassium citrate, a slight stimulation of O2 evolution is observed (143-173 mumol of O2/h per mg of chlorophyll a) with potassium ferricyanide as electron acceptor. When we monitor the potassium ferricyanide reduction, no stimulation of electron transport is obtained even if the observed phenomenon is identical with the Photosystem-II oxygen evolution. In the same experiments no stimulation of the photophosphorylation was recorded, but when thylakoids are directly isolated in 0.75 M-potassium citrate, O2 evolution, ferricyanide reduction and photophosphorylation are inhibited by high concentrations of salts. The behaviour of thylakoids seems to be influenced by their initial treatment.

Cell Membrane↗
PubMed1983

Growth and sporulation of entrapped Bacillus subtilis cells.

Growing Bacillus subtilis cells were immobilized in k-carrageenan gel beads. Growth and sporulation of entrapped cells were studied by two different methods: cell enumeration and transmission electron microscopy. Immobilized growing cells had a shorter generation time than free cells did, whereas sporulation timing was unchanged. When steady state was reached, cell density was very high in gel beads.

Bacillus subtilis↗
PubMed1982

Immobilized thylakoids in a cross-linked albumin matrix: effects of cations studied by electron microscopy, fluorescence emission, photoacoustic spectroscopy, and kinetic measurements.

Immobilization of lettuce (Lactuca sativa) thylakoids has been performed by using glutaraldehyde and bovine serum albumin. Confirming previous reports, a stabilization of the O(2) evolution activity of the photosystem II (PSII) under storage and functional conditions has been observed. The present work is devoted to the role played by mono-and divalent cations, during the immobilization process itself, on the O(2) production. Four types of measurements have been employed: kinetic measurements, low temperature (77 K) fluorescence emission, photoacoustic (PA) spectroscopy, and electron microscopy observations. We show that the effect of glutaraldehyde is complex because it acts as an inhibitor, a stabilizing agent, and a cross-linking reactive. In the present studies, the thylakoids are immobilized within a polymeric insoluble albumin matrix. The highest activity yield and the best storage conditions are obtained when 0.15 mm Na(+) (or K(+)), 1 mm Mg(2+), and 0.1 mm Mn(2+) are present in the resuspending media before the immobilization. Due to modifications of the ionic content during such a process, structural differences are observed on the stacking degree of thylakoids. No modification of the fluorescence and PA spectra after the immobilization are found. Furthermore, a correlation between activities and spectral changes have been shown: when the activities increase, the F(735) to F(695) ratio increases and the PA(676) to PA(440) ratio decreases.

Journal Article↗
PubMed1981

Kinetic, computer, and electron microscopic studies dealing with an artificial enzyme membrane: theoretical and experimental evaluation of the cytochemical demonstration of acetylcholinesterase.

The cytochemical demonstration of acetylcholinesterase is theoretically and experimentally evaluated with a model system. The model is an artificial proteic membrane in which acetylcholinesterase homogeneously is immobilized chemically by a bifunctional agent, glutaraldehyde. The copper-thiocholine histochemical method is studied kinetically and the system is simulated by computer calculations based on experimental kinetic parameters and numerical analysis methods. In addition, the corresponding electron micrographs are presented. These studies lead to the conclusions that the system is ruled by diffusional constraints and that enzyme distribution and repartition of the insoluble electron dense product are not circumscribed by any specific conditions.

Acetylcholinesterase↗
PubMed1980

Theoretical and experimental study of an immobilized bienzyme system. Computer calculations and electron microscopy visualization of local concentration profiles.

A cytochemical method was used in order to visualize the existence of local concentrations of metabolites within an immobilized bienzyme system. Glucose oxidase (EC 1.1.3.4) and peroxidase (EC 1.11.1.7) were immobilized together into an artificial proteic membrane obtained by a previously described co-cross-linking process using a bifunctional agent, glutaraldehyde. The cytochemical reagent 3,3'-diaminobenzidine was used as a hydrogen donor substrate and kinetic studies were performed with both free and immobilized forms of the enzymes; in the latter case, a new direct method was introduced which allowed kinetic studies on the visualization system itself. From experimental kinetic parameters, theoretical concentrations profiles were calculated by computer using numerical analysis methods. These results were discussed with special attention to the electron microscopic observations. Computer simulations were in good agreement with electron micrographs. No geometrical similarity was seen in our system between poly(3,3'-diaminobenzidine) precipitate distribution and peroxidase molecule homogeneous location.

Computers↗
PubMed1980

A histochemical model dealing with an immobilized glucose oxidase-peroxidase system. The influence of diffusion limitations on histochemical results.

A histochemical model dealing with an immobilized bienzyme system (glucose oxidase-peroxidase) is presented. The model is an artificial proteic membrane obtained by a previously described co-cross-linking process. The kinetic properties of free and immobilized horseradish peroxidase were studied when 3,3'-diaminobenzidine is used as a hydrogen donor substrate. A new direct method was developed for immobilized enzyme activity measurements. Computer simulation based on experimental kinetic parameters was performed in order to discuss electron microscopy results. By changing diffusion limitations, various profiles of insoluble product were visualized inside the proteic film and no geometrical similarity was seen between enzyme distributions and insoluble osmiophilic product patterns.

Computers↗
PubMed1979

Immobilization of L-glutamate dehydrogenase into soluble cross-linked polymers. ADP effect and electron microscopy studies.

Active soluble cross-linked L-glutamate dehydrogenase (L-glutamate: NAD(P)+ oxidoreductase (deaminating), EC 1.4.1.3) albumin polymers were produced. Electron microscopic studies and kinetic properties were studied with the polymer in solution and compared with previous published data about the enzyme immobilized inside proteic films (Barbotin, J.N. and Breuil, M. (1978) Biochim. Biophys. Acta 525, 18--27). The glutaraldehyde effect on activity yield, ADP and beta-NAD+ protection, stability and pH rate profile were studied and discussed. Apparent Michaelis constants were determined with soluble polymers produced with or without ADP during the grafting process. Experiments were performed on the regulatory properties of immobilized glutamate dehydrogeanse showing the decrease of ADP activation and GTP inhibition as compared to the free form. In other respects, electron microscopy observations showed morphological differences between the two populations of soluble polymers produced in presence of ADP, obtained after gel filtration on Sepharose 6B. Linear aggregates of high molecular weight and classical soluble polymers were obtained. Similar Km values and regulatory properties were exhibited by the two forms, demonstrating the absence of interdependence between the allosteric control and the polymerization of enzyme monomers.

Adenosine Diphosphate↗
PubMed1977

Experimental evidence for a kinetic and electrochemical memory in enzyme membranes.

The existence of hysteresis phenomena in artificial enzyme membranes due to the coupling of simple kinetic enzyme properties with diffusion transport processes is reported. The intramembrane pH of a urease coating on the surface of a glass pH electrode exhibits a hysteresis loop when the pH of the bulk solution varies cyclically. The steady-state potential of a urease membrane, as a function of the substrate concentration in the bulk solution, also exhibits a memory effect. The influence of the membrane's history on its overall behavior is visualized by electron microscopy. We interpret the results in terms of a coupling between the enzyme reactions and diffusion processes, without taking into account molecular effects.

Diffusion↗
PubMed1976

Blood compatibility of silicone rubber chemically coated with cross-linked albumin.

A new method is reported for chemically coating cross-linked albumin onto silicone rubber in an attempt to improve the blood compatibility of the material. Promising results were obtained in an evaluation using both vena cava and renal embolus antithrombogenicity tests ondogs. Coated silicone rubber rings induced much less renal emboli than uncoated ones after one week implantation.

Albumins↗