Biomedical subjects

D G Mita

Publications and source records attributed to D G Mita.

12 recordsLinked to original sources

PubMed2005

Modulation of the catalytic activity of free and immobilized peroxidase by extremely low frequency electromagnetic fields: dependence on frequency.

A study of the influence of electromagnetic fields (EMF) of various frequencies, from 50 up to 400 Hz, on the catalytic activity of soluble and insoluble horseradish peroxidase (POD) was carried out. To simulate the conditions in which the enzyme operates in vivo, the POD was immobilized by entrapment on a gelatin membrane or by covalent attachment on a nylon graft membrane. The rate of inactivation of the soluble POD was found to exhibit positive and negative interactions with the 1 mT applied magnetic field, with an optimum positive effect at 130 Hz. The immobilized PODs, on the contrary, do not exhibit negative interactions, but show a maximum positive interaction at 150 Hz when entrapped and at 170 Hz when covalently attached. At 50 Hz and at frequencies higher than 250 Hz no effects were observed with insoluble POD. The optimum frequency of positive interaction between the EMF and the catalytic activity of the insoluble enzymes is shifted with respect to that of the soluble enzymes towards higher frequencies, the size of the shifts being dependent on the intensity of the physical forces involved in the immobilization process.

Catalysis↗

PubMed2003

In vitro studies of the influence of ELF electromagnetic fields on the activity of soluble and insoluble peroxidase.

The influence of an extremely low frequency (ELF) magnetic field (50 Hz and 1 mT, EMF) on the activity of a soluble and insoluble horseradish peroxidase (E.C. 1.11.17) has been studied as a function of time. Insoluble derivatives were obtained by enzyme entrapment into two different gelatin membranes or by covalent attachment of the enzyme on two nylon membranes, differently preactivated. Results have shown that the field affects the inactivation rate of the soluble enzyme, while no effects are observed with insoluble derivatives. Since in vivo enzymes are immobilised into the biomembrane bilayer or entrapped into the cytoplasmic mixture, one might speculate that our experimental conditions do not reflect the catalytic activity of the enzymes in vivo.

Benzothiazoles↗

PubMed2003

Protease removal by means of antiproteases immobilized on supports as a potential tool for hemodialysis or extracorporeal blood circulation.

This work studies protease concentration decrease in aqueous solutions in contact with a modified polyethersulphone graft membrane onto which antiproteases were immobilized. As a model of protease/antiprotease interaction, elastase and alpha1-antitrypsin were used. Experiments were carried out either under fixed amounts of immobilized antiproteases and variable protease concentration or under fixed protease concentration and variable amounts of immobilized antiproteases. In both cases, active protease concentrations decreased with increase in contact time with the membrane. Experimental conditions under which active elastase concentration becomes zero were also found. Occurrence of the same phenomenology has also been ascertained with protease solutions obtained from human blood neutrophils. The membrane activated with alpha1-antitrypsin showed differential inhibitory power on elastase and cathepsin G. This technology could open new perspectives in manufacturing new membranes to be used in hemodialysis and extracorporeal circulation when elastase is released.

Cardiopulmonary Bypass↗

PubMed2002

Advantages of using non-isothermal bioreactors for the enzymatic synthesis of antibiotics: the penicillin G acylase as enzyme model.

A new hydrophobic and catalytic membrane was prepared by immobilizing Penicillin G acylase (PGA, EC.3.5.1.11) from E. coli on a nylon membrane, chemically grafted with butylmethacrylate (BMA). Hexamethylenediamine (HMDA) and glutaraldehyde (Glu) were used as a spacer and coupling agent, respectively. PGA was used for the enzymatic synthesis of cephalexin, using D(-)-phenylglycine methyl ester (PGME) and 7-amino-3-deacetoxycephalosporanic acid (7-ADCA) as substrates. Several factors affecting this reaction, such as pH, temperature, and concentrations of substrates were investigated. The results indicated good enzyme-binding efficiency of the pre-treated membrane, and an increased stability of the immobilized PGA towards pH and temperature. Calculation of the activation energies showed that cephalexin production by the immobilized biocatalyst was limited by diffusion, resulting in a decrease of enzyme activity and substrate affinity. Temperature gradients were employed as a way to reduce the effects of diffusion limitation. Cephalexin was found to linearly increase with the applied temperature gradient. A temperature difference of about 3 degrees C across the catalytic membrane resulted into a cephalexin synthesis increase of 100% with a 50% reduction of the production times. The advantage of using non-isothermal bioreactors in biotechnological processes, including pharmaceutical applications, is also discussed.

Anti-Bacterial Agents↗

PubMed2002

The alpha1-antitrypsin/elastase complex as an experimental model for hemodialysis in acute catabolic renal failure, extracorporeal blood circulation and cardiocirculatory bypass.

A modified polyethersulphone graft membrane was loaded with antiproteases, with the aim of reducing the active protease blood concentration during hemodialysis in acute catabolic renal failure or cardiopulmonary bypass. As protease/antiprotease system, elastase and alpha1-antitrypsin were used. The concentration of active elastase in aqueous solutions decreased as function of contact time with the membrane, approaching saturation. A 40% loss of elastase activity was obtained at pH 7.4, which was not due to autolysis, which accounted for 5% of the loss. The highest reduction was achieved at pH 9.0 (25% higher than at pH 7.4). The saturation level of elastase decrease, calculated by means of the Einstein equation, was reached after more than 47 minutes. We speculate that a time reduction might be achieved either increasing the concentration of immobilized antiproteases, or increasing the rate of elastase movement across the membranes by hydraulic, osmotic, or temperature gradients. This technology can be applied to hemodialysis, and in extracorporeal blood circulation to promote elastase release.

Acute Kidney Injury↗

PubMed1999

A glucose biosensor operating under non-isothermal conditions: the dynamic response.

The results obtained with a glucose biosensor operating under non-isothermal conditions are presented and discussed. Glucose oxidase, immobilized onto Nylon membranes, was used as biological element. An amperometric two electrodes system was employed to measure the anodic current produced by oxidation of hydrogen peroxide. Non-isothermal conditions were characterised in terms of the temperature difference, delta T = Tw - Tc, and of the average temperature of the system, Tav = (Tw + Tc)/2, Tw and Tc being the temperature in the warm and cold half-cells constituting the biosensor. Comparison between the functioning of the biosensor under isothermal and non-isothermal conditions was performed. It was found that, under non-isothermal conditions, the dynamic response and sensitivity increased, while the response times and the detection limit decreased, if comparison was done with the same parameters measured under isothermal conditions. The increase of the dynamic response was found to be proportional to the applied temperature gradient.

Biosensing Techniques↗

PubMed1998

Advantages in using immobilized thermophilic beta-glycosidase in nonisothermal bioreactors.

Catalytic membranes, obtained by immobilizing thermophilic beta-glycosidase onto nylon supports, were used in a nonisothermal bioreactor to study the effect of temperature gradients on the rate of enzyme reaction. Two experimental approaches were carried out to explain the molecular mechanisms by which the temperature gradients affect enzyme activity. The results showed that the thermophilic enzyme behaved as the mesophilic beta-galactosidase, exhibiting an activity increase which was linearly proportional to the transmembrane temperature difference. The efficiency of the system proposed was determined by calculating two constants, alpha and beta, which represent respectively the percentage increase of enzyme activity when a temperature difference of 1 degrees C or a temperature gradient of 1 degrees C cm-1 were applied across the catalytic membrane. The increase of enzyme activity in nonisothermal bioreactors entailed a proportional reduction of production times. The advantages in using thermophilic enzymes immobilized in nonisothermal bioreactors are also discussed.

Bioreactors↗

PubMed1996

A non-isothermal bioreactor utilizing immobilized baker's-yeast cells: a study of the effect on invertase activity.

The behaviour of the enzyme invertase, located on the cell wall of baker's-yeast cells and entrapped in a gelatin membrane, was studied under isothermal and non-isothermal conditions. The reaction rate linearly increased with the applied transmembrane temperature gradient, with reference either to the average temperature or to the temperature on the warm side of the catalytic membrane. These results were obtained both when the bioreactor was operated under conditions of closed volumes and when the substrate-containing solutions are recirculated. The mathematical relationships have been elaborated between the temperatures read in the working solutions and those on the two faces of the catalytic membrane. Since the temperature difference across the membrane is smaller than that indicated by the thermocouples, the observed effects are greater than expected. The potential advantages of the use of a non-isothermal bioreactor in processes of industrial interest are discussed.

Biotechnology↗

PubMed1987

Temperature gradients and prebiological evolution.

Thermodiffusive transport of trace elements that play important roles in living organisms, such as molybdenum, nickel, copper, and vanadium, was studied in a nonisothermal biphasic system comprised of a liquid solution and jelly layers. Our intent was to mimic the effects of temperature gradients on prebiological evolution. Conditions were found, similar to those probably existing during development of early eobionts, under which all the elements tested were concentrated within the heated jelly. Nonisothermal matter transport through grossly porous artificial membranes--the process of thermodialysis--was next investigated to assess the behavior of compartmentalized, i.e., membrane bound, eobionts. Particular interest was dedicated to the continuity of nonisothermal transport phenomena in the homogeneous and heterogeneous (membrane) systems and to the ability of compartmentalized eobionts to withstand osmotic swelling by means of thermoosmotic transport. Interestingly enough, under the experimental conditions adopted, sodium/potassium countertransport is also found, suggesting a very early physicochemical origin of the sodium-potassium pump. Surprisingly enough, evidence of teleonomic behavior appears in those very simple analogs of prebiological systems.

Biological Evolution↗

PubMedDate not supplied

Modulation of membrane potential in algal cells by temperature gradients. A thermodynamic approach.

The aim of the present study is to ascertain whether transmembrane temperature gradients couple with transport of electric charge in living cells of Valonia utricularis and eventually measure the thermodynamic coupling coefficient (s). Simple experimental procedures are described that allow generation of temperature gradients of predetermined sense and intensity across the cell membrane. Simultaneous measurement of the potential difference is ensured by standard electrophysiological methods. The mathematical expressions that allow quantitative treatment of experimental results are indicated in the article and are based on standard nonequilibrium thermodynamic and electrophysiological formalism. The value of the coupling coefficient between temperature gradient and flow of electric charge is indicated and concisely discussed in terms of possible mechanisms of ionic membrane transport.

Biological Transport↗

PubMedDate not supplied

Increase in beta-galactosidase activity in a non-isothermal bioreactor utilizing immobilized cells of Kluyveromyces fragilis: fundamentals and applications.

The beta-galactosidase activity of Kluyveromyces fragilis cells immobilized in a kappa carrageenan gel was studied in a bioreactor functioning under isothermal and non-isothermal conditions. We observed an increase in enzyme activity which was found to be proportional to the intensity of the temperature gradient applied across the biocatalytic membrane, as well as to the average temperature of the bioreactor. The efficiency of such a non-isothermal bioreactor was calculated with respect to the yield of a bioreactor working under comparable isothermal conditions and was evaluated in terms of reduction of processing times in industrial applications. The possibility that enzyme activity in living cells is affected by non-isothermal conditions naturally existing owing to metabolic heat production is also discussed.

Bioreactors↗

PubMedDate not supplied

Enzyme reaction engineering: effect of methanol on the synthesis of antibiotics catalyzed by immobilized penicillin G acylase under isothermal and non-isothermal conditions.

The effect of methanol on the kinetically controlled synthesis of cephalexin by free and immobilized penicillin G acylase (PGA) was investigated. Catalytic and hydrophobic membranes were obtained by chemical grafting, activation, and PGA immobilization on hydrophobic nylon supports. Butyl methacrylate (BMA) was used as graft monomer. Increasing concentrations of methanol were found to cause a greater deleterious effect on the activity of free than on that of the immobilized enzyme. Methanol, however, improved the kinetic stability of cephalexin synthesized by free PGA, resulting in higher maximum yields. By contrast, immobilized PGA reached 100% yields even in the absence of the cosolvent. Cephalexin synthesis by the catalytic membrane was also performed in a non-isothermal bioreactor. Under these conditions, a 94% increase of the synthetic activity and complete conversion of the limiting substrate to cephalexin were obtained. The addition of methanol reduced the non-isothermal activity increase. The physical cause responsible for the non-isothermal behavior of the hydrophobic catalytic membrane was identified in the process of thermodialysis.

Anti-Bacterial Agents↗