Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Equilibrium optimization”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 1,405 records · Page 78Linked to original sources
PubMed2006

Triclosan release from coated polyglycolide threads.

Copolymerization of lactide, epsilon-caprolactone, and trimethylene carbonate is performed. The synthesis is focused on obtaining materials with adequate properties for application as a suture coating that could contain an antimicrobial agent like triclosan. An amorphous character, a low glass transition temperature, a moderate susceptibility to degradation, and a hydrophobic nature are the conditions required for the optimal behavior of this coating. These properties can be attained with a copolymer of composition 10:60:30. Triclosan is added to the surface of polyglycolide threads and its release is studied in different media with high-performance liquid chromatography. The influence of the temperature, the diameter of the thread, the initial concentration of the antibacterial agent, and the applied procedure on the incorporation of triclosan is also evaluated. A total release of triclosan is attained after a few days of exposure to a Dulbecco's based medium, whereas equilibrium concentrations are reached when a Sörensen hydrophilic medium is used. Partition and diffusion coefficients are also estimated.

Absorbable Implants↗
PubMed1998

In vitro study of alcohol dehydrogenase and acetaldehyde dehydrogenase encapsulated into human erythrocytes by an electroporation procedure.

The optimal conditions for electroporated/resealed loading of alcohol dehydrogenase (ADH) and/or acetaldehyde dehydrogenase (ALDH) into human erythrocytes were established prior to the study, with the following characteristics: 300 V, 1 ms pulse time, eight pulses every 15 min and 1 h resealing at 37 degreesC. High encapsulation yield and carrier cell recoveries were achieved. Cell volumes increase while hemoglobin contents decrease; in consequence a decrease in cell hemoglobin concentration was observed. A lower hypotonic resistance of loading erythrocytes (throughout osmotic fragility curves) and unaltered oxygen transport capability (as given by oxygen equilibrium curves) were observed. The stability against time (up to 168 h-7 days) of encapsulated individual enzymes, either ADH- or ALDH-red blood cells (RBCs), was studied at 4 degreesC and 37 degreesC, in comparison with that of free enzyme solutions. Both enzymes were released from carrier RBCs to the incubation medium. The stability of carrier RBCs was studied under similar conditions. Non-significant variations in hematological parameters were observed. However, the hemoglobin derivative forms showed modifications. The continuous degradation of ethanol by ADH-RBCs and coencapsulated ADH- and ALDH-RBCs, as a function of time (up to 70 h) suggests the use of these carrier RBCs as agents for complete metabolization of ethanol. The mentioned properties bare the possibility of using ADH and ALDH as carrier systems in in vivo situations.

Alcohol Dehydrogenase↗
PubMed2004

Simultaneous concentration and separation of enantiomers with chiral temperature gradient focusing.

A new technique is demonstrated for the simultaneous concentration and high-resolution separation of chiral compounds. With temperature gradient focusing, a combination of a temperature gradient, an applied electric field, and a buffer with a temperature-dependent ionic strength is used to cause analytes to move to equilibrium, zero-velocity points along a microchannel or capillary. Different analytes are thus separated spatially and concentrated in a manner that resembles isoelectric focusing but that is applicable to a greater variety of analytes including small chiral drug molecules. Chiral separations are accomplished by the addition of a chiral selector, which causes the different enantiomers of an analyte to focus at different positions along a microchannel or capillary. This new technique is demonstrated to provide high performance in a number of areas desirable for chiral separations including rapid separation optimization and method development, facile reversal of peak order (desirable for analysis of trace enantiomeric impurities), and high resolving power (comparable to capillary electrophoresis) in combination with greater than 1000-fold concentration enhancement enabling improved detection limits. In addition, chiral temperature gradient focusing allows for real-time monitoring of the interaction of chiral analyte molecules with chiral selectors that could potentially be applied to the study of other molecular interactions. Finally, unlike CE, which requires long channels or capillaries for high-resolution separations, separations of equivalent resolution can be performed with TGF in very short microchannels (mm); thus, TGF is inherently much more suited to miniaturization and integration into lab-on-a-chip-devices.

Buffers↗
PubMed2002

Mechanism of spontaneous DNA-DNA interaction of homologous linear duplexes.

Previously, we demonstrated the interaction of homologous linear duplexes with formation of four-way DNA structures on the model of five PCR products. We propose that homologous duplex interaction is initiated by the nucleation of several dissociated base pairs of the complementary ends of two fragments with Holliday junction formation, in which cross point migration occurs via spooling of DNA strands from one duplex to the other one, finally resulting in complete resolution into new or previously existing duplexes. To confirm that DNA-DNA interaction involves formation of four-way DNA structures with strand exchange at the cross point, we have demonstrated the strand exchange process between identical duplexes using homologous fragments, harboring either biotin label or (32)P-label. Incubation of the mixture resulted in the addition of (32)P-label to biotin-labeled fragments, and the intensity of (32)P-labeling of biotinylated fragments was dependent upon the incubation duration. DNA-DNA interaction is not based on surface-dependent denaturing, as Triton X-100 does not decrease the formation of complexes between DNA duplexes. The equilibrium concentration of Holliday junctions depends on the sequences of the fragment ends and the incubation temperature. The free energy of Holliday junction formation by the fragments with GC and AT ends differed by 0.6 kcal/mol. Electron microscopic analysis demonstrated that the majority of Holliday junctions harbor the cross point within a 300 base pair region of the fragment ends. This insight into the mechanism of homologous duplex interaction extends our understanding of different DNA rearrangements. Understanding of DNA-DNA interaction is of practical use for better interpretation and optimization of PCR-based analyses.

DNA↗
PubMed2005

Detailed ab initio studies of the conformers and conformational distributions of gaseous tryptophan.

A systematic and extensive conformational search has been performed to characterize the gas-phase tryptophan structures. A total of 648 unique trial structures were generated by allowing for all combinations of internal single-bond rotamers and were optimized at the DFT/B3LYP/6-311G* level of theory. A total of 45 local minima conformers were found. Further optimization of the 45 conformers with B3LYP and MP2/6-311++G** did not produce meaningful structural change, and accurate geometries, dipole moments, rotational constants, harmonic frequencies, and relative energies were then determined. Combined with statistical mechanics principles, the conformational distributions of gas-phase tryptophan at different temperatures are shown. The results clearly support the conclusion drawn by Compagnon et al. that only one dominant isomer existed in the molecular beam at 85 K and add further evidence that the supersonic jet expansion or embedding helium droplets did not produce an equilibrium distribution.

Computer Simulation↗
PubMed2006

Scaling down the Perdew-Zunger self-interaction correction in many-electron regions.

Semilocal density functional approximations (DFAs) for the exchange-correlation energy suffer from self-interaction error, which is believed to be the cause of many of the failures of common DFAs, such as poor description of charge transfer and transition states of chemical reactions. The standard self-interaction correction (SIC) of Perdew and Zunger mends some of these failures but spoils such essential properties as thermochemistry and equilibrium bond lengths. The Perdew-Zunger SIC seems to overcorrect many-electron systems. In this paper, we propose a modified SIC, which is scaled down in many-electron regions. The new SIC has an improved performance for many molecular properties, including total energies, atomization energies, barrier heights of chemical reactions, ionization potentials, electron affinities, and bond lengths. The local spin-density approximation (LSDA) benefits from SIC more than higher-level functionals do. The scaled-down SIC has only one adjustable parameter. Rationalization of the optimal value of this parameter enables us to construct an almost-nonempirical version of the scaled-down SIC-LSDA, which is significantly better than uncorrected LSDA and even better than the uncorrected generalized gradient approximation. We present an analysis of the formal properties of the scaled-down SIC and define possible directions for further improvements. In particular, we find that exactness for all one-electron densities does not guarantee correct asymptotics for the exchange-correlation potential of a many-electron system.

Journal Article↗
PubMed1994

A multi-ion permeation mechanism in neuronal background chloride channels.

Unitary current/voltage relationships of background Cl channels of rat hippocampal neurons were determined for varied gradients and absolute concentrations of NaCl. The channels revealed permeabilities for both Cl and Na ions. A hyperlinear increase of unitary conductance, observed for a symmetrical increase of salt concentration from 300 and 600 mM, indicated a multi-ion permeation mechanism. A variety of kinetic models of permeation were tested against the experimental current/voltage relationships. Models involving a pore occupied by mixed complexes of up to five ions were necessary to reproduce all measurements. A minimal model included four equilibrium states and four rate-limiting transitions, such that the empty pore accepts first an anion and then can acquire one or two cation/anion pairs. Three transport cycles are formed: a slow anion cycle (between the empty and single-anion states), a slow cation cycle (between the one- and three-ion states), and a fast anion cycle (between the three- and five-ion states). Thus, permeant anions are required for cation permeation, and several bound anions and cations promote a high rate of anion permeation. The optimized free-energy and electrical charge parameters yielded a self-consistent molecular interpretation, which can account for the particular order in which the pore accepts ions from the solutions. Although the model describes the mixed anion/cation permeability of the channel observed at elevated concentrations, it predicts a high selectivity for Cl anion at physiological ionic conditions.

Animals↗
PubMed2000

Toxicity and DNA binding of dextran-doxorubicin conjugates in multidrug-resistant KB-V1 cells: optimization of dextran size.

We previously showed that conjugating doxorubicin to very large 70-500 kDa dextran decreased its removal rate from P-glycoprotein (P-gp) over-expressing, multidrug-resistant KB-V1 cells. Furthermore these conjugates could act synergistically with other cancer drugs. In the drug-sensitive 3-1 clone, but not in the V1 subclone which was 300-fold more resistant to free doxorubicin, conjugation led to a size-related decrease in toxicity. Here we identified the optimal size of dextran for avoiding P-gp-mediated efflux and yet preserving as much as possible doxorubicin toxicity. Chemically reduced, intracellularly stable 3.4-10 kDa conjugates were prepared. Confocal microscopy and fluorescence quenching experiments showed that these conjugates entered nuclei and interacted with DNA. In 3-1 cells, but not in V1 cells, cytotoxicity of conjugates decreased 14- to 45-fold linearly related to log size of the carrier (r=0.95). In V1 cells toxicity of the 10 kDa conjugate exceeded that of free doxorubicin. After conjugation the equilibrium binding constant of the DNA-drug complex (KA) decreased only by up to 3-fold. In 3-1 cells, but not in VI cells, DNA binding kinetics was an important factor and toxicity could be linearly correlated to 1/KA of conjugate (r=0.94). Drug accumulation decreased with an increase in dextran size but drug removal was decreased only in V1 cells. It appeared that drug uptake was also sensitive to dextran conjugation. In Vl cells drug removal was sensitive to the P-gp inhibitor verapamil or energy starvation. Ratios of V1/3-1 toxicity, drug accumulation and drug removal correlated linearly with log dextran size. When these ratios equaled 1, dextran sizes were estimated to be 32, 103 and 21 kDa, respectively.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1976

Purification and characterization of phosphoglycerate mutase from methanol-grown Hyphomicrobium X and Pseudomonas AM1.

Phosphoglycerate mutase has been purified from methanol-grown Hyphomicrobium X and Pseudomonas AMI by acid precipitation, heat treatment, ammonium sulphate fractionation, Sephadex G-50 gel filtration and DEAE-cellulose column chromatography. The purification attained using the Hyphomicrobium X extract was 72-fold, and using the Pseudomonas AMI extract, 140-fold. The enzyme purity, as shown by analytical polyacrylamide gel electrophoresis, was 50% from Hyphomicrobium X and 40% from Pseudomonas AMI. The enzyme activity was associated with one band. The purified preparations did not contain detectable amounts of phosphoglycerate kinase, phosphopyruvate hydratase, phosphoglycerate dehydrogenase or glycerate kinase activity. The molecular weight of the enzymic preparation was 32000 +/- 3000. The enzyme from both organisms was stable at low temperatures and, in the presence of 2,3-diphosphoglyceric acid, could withstand exposure to high temperatures. The enzyme from Pseudomonas AMI has a broad pH optimum at 7-0 to 7-6 whilst the enzyme from Hyphomicrobium X has an optimal activity at pH 7-3. The cofactor 2,3-diphosphoglyceric acid was required for maximum enzyme activity and high concentrations of 2-phosphoglyceric acid were inhibitory. The Km values for the Hyphomicrobium X enzyme were: 3-phosphoglyceric acid, 6-0 X 10(-3) M: 2-phosphoglyceric acid, 6-9 X 10(-4) M; 2,3-diphosphoglyceric acid, 8-0 X 10(-6) M; and for the Pseudomonas AMI ENzyme: 3-4 X 10(-3) M, 3-7 X 10(-4) M and 10 X 10(-6) M respectively. The equilibrium constant for the reaction was 11-3 +/- 2-5 in the direction of 2-phosphoglyceric acid to 3-phosphoglyceric acid and 0-09 +/- 0-02 in the reverse direction. The standard free energy for the reaction proceeding from 2-phosphoglyceric acid to 3-phosphoglyceric acid was -5-84 kJ mol(-1) and in the reverse direction +5-81 kJ mol(-1).

Bacteria↗
PubMed1997

Measurement and simulation of water transport during freezing in mammalian liver tissue.

Optimization of cryosurgical procedures on deep tissues such as liver requires an increased understanding of the fundamental mechanisms of ice formation and water transport in tissues during freezing. In order to further investigate and quantify the amount of water transport that occurs during freezing in tissue, this study reports quantitative and dynamic experimental data and theoretical modeling of rat liver freezing under controlled conditions. The rat liver was frozen by one of four methods of cooling: Method 1-ultrarapid "slam cooling" (> or = 1000 degrees C/min) for control samples; Method 2-equilibrium freezing achieved by equilibrating tissue at different subzero temperatures (-4, -6, -8, -10 degrees C); Method 3-two-step freezing, which involves cooling at 5 degrees C/min. to -4, -6, -8, -10 or -20 degrees C followed immediately by slam cooling; or Method 4-constant and controlled freezing at rates from 5-400 degrees C/min. on a directional cooling stage. After freezing, the tissue was freeze substituted, embedded in resin, sectioned, stained, and imaged under a light microscope fitted with a digitizing system. Image analysis techniques were then used to determine the relative cellular to extracellular volumes of the tissue. The osmotically inactive cell volume was determined to be 0.35 by constructing a Boyle van't Hoff plot using cellular volumes from Method 2. The dynamic volume of the rat liver cells during cooling was obtained using cellular volumes from Method 3 (two-step freezing at 5 degrees C/min). A nonlinear regression fit of a Krogh cylinder model to the volumetric shrinkage data in Method 3 yielded the biophysical parameters of water transport in rat liver tissue of: Lpg = 3.1 x 10(-13) m3/Ns (1.86 microns/min-atm) and ELp = 290 kJ/mole (69.3 kcal/mole), with chi-squared variance of 0.00124. These parameters were then incorporated into the Krogh cylinder model and used to simulate water transport in rat liver tissue during constant cooling at rates between 5-100 degrees C/min. Reasonable agreement between these simulations and the constant cooling rate freezing experiments in Method 4 were obtained. The model predicts that the water transport ceases at a relatively high subzero temperature (-10 degrees C), such that the amount of intracellular ice forming in the tissue cells rises from almost none (= extensive dehydration and vascular expansion) at < or = 5 degrees C/min to over 88 percent of the original cellular water at > or = 50 degrees C/min. The theoretical simulations based on these experimental methods may be of use in visualizing and predicting freezing response, and thus can assist in the planning and implementing of cryosurgical protocols.

Animals↗
PubMedDate not supplied

Higher energy: is it necessary, is it worth the cost for radiation oncology?

The physical characteristics of the interactions of megavoltage photons and electrons with matter provide distinct advantages, relative to low-energy (orthovoltage) x rays, that lead to better radiation dose distributions in patients. Use of these high-energy radiations has resulted in better patient care, which has been reflected in improved radiation treatment outcome in recent years. But, as the desire for higher energy radiation beams increases, it becomes important to determine whether the physical characteristics that make megavoltage beams beneficial continue to provide a net advantage. It is demonstrated that, in fact, there is an energy range from 4 to 15 MV for photons and 4 to 20 MeV for electrons that is optimally suited for the treatment of cancer in humans. Radiation beams that exceed these maximum energies were found to add no advantage. This is because the costs (price of unit, installation, maintenance, shielding for neutron and photons) are not justified by either improved physical characteristics of the radiation (penetration, skin sparing, dose distribution) or treatment outcome. In fact, for photon beams some physical characteristics result in less desirable dose distributions, less accurate dosimetry, and increased safety problems as the energy increases for example, increasingly diffuse beam edges, loss of electron equilibrium, uncertainty in dose perturbations at interfaces, increased neutron contamination, and potential for higher personnel dose. The special features that make electron beams useful at lower energies, for example, skin sparing and small penetration, are lost at high energies. These physical factors are analyzed together with the economic factors related to radiation therapy patient care using megavoltage beams.

Cost-Benefit Analysis↗
PubMed2004

Mathematical modeling of salt-gradient ion-exchange simulated moving bed chromatography for protein separations.

The salt-gradient operation mode used in ion-exchange simulated moving bed chromatography (SMBC) can improve the efficiency of protein separations. A detailed model that takes into account any kind of adsorption/ion-exchange equilibrium, salt gradient, size exclusion, mass transfer resistance, and port periodic switching mechanism, was developed to simulate the complex dynamics. The model predictions were verified by the experimental data on upward and downward gradients for protein separations reported in the literature. All design and operating parameters (number, configuration, length and diameter of columns, particle size, switching period, flow rates of feed, raffinate, desorbent and extract, protein concentrations in feed, different salt concentrations in desorbent and feed) can be chosen correctly by numerical simulation. This model can facilitate the design, operation, optimization, control and scale-up of salt-gradient ion-exchange SMBC for protein separations.

Chromatography, Ion Exchange↗
PubMed1999

The effect of generic competition on prescription to over-the-counter switching.

OBJECTIVE: While it is generally accepted that the decision to switch a drug from the prescription market to the over-the-counter (OTC) market is based on an optimization problem that firms are solving, no attempts have been made to formalize the problem. The purpose of this article is to present a model of prescription to OTC switching that helps explain the role of potential generic competition in a firm's decision to switch. In particular, we examine what market conditions are necessary for the threat of generic competition to induce switching. DESIGN AND SETTING: The model is game-theoretic, played between an incumbent firm and a potential generic entrant, and is solved for its subgame perfect equilibrium. The incumbent first decides whether to apply to the FDA to switch to the OTC market. If the incumbent declines, then the potential generic entrant has the opportunity to apply for the switch. The FDA then accepts or rejects the application, and the generic chooses whether to enter the market. RESULTS: In equilibrium, when applying to switch is costless, switching occurs if the probability that the application will be approved by the FDA is strictly positive and the OTC market is characterized by first-mover advantages. Adding a cost to the application process places restrictions on the probability of FDA approval to offset the cost of applying. The probability of approval must be sufficiently high to offset the cost of the application. CONCLUSIONS: The model shows that switching from the prescription to OTC market may occur as a response to potential generic competition. Firms switch because they know that if they do not, a generic may initiate a switch and become the first mover in the OTC market.

Drug Costs↗
PubMed1996

Cerebrovascular reactivity: role of endothelium/platelet/leukocyte interactions.

In the last two decades, a tremendous amount of knowledge has been accumulated in various fields of biomedical research that discloses mechanisms of platelet/leukocyte/endothelium interactions. Occupying a strategically important location between circulating blood and underlying tissues, the endothelium effectively modulates both the functional state of the blood cells and the tone of vascular smooth muscle by generating or metabolizing a host of humoral substances. Under normal conditions, the endothelium releases agents with predominantly vasodilator and antiaggregant/anticoagulant activity that prevent thrombotic and angiospastic disorders. However, a variety of pathophysiological stimuli may trigger endothelial reorganization with the expression of different prothrombotic factors and activation of platelets and leukocytes that, combined, leads to blood cell adhesion to the endothelial monolayer, aggregation as thrombi, and the formation of numerous spasmogenic substances. Activation of the blood cells in the vicinity of the endothelium may induce endothelial dysfunction/injury, resulting in impairment of normal endothelial antispasmodic control. Within the microcirculatory bed, intravascular activation of the blood cells leads to scattered microvessel plugging, increased vascular permeability, edema formation, and cytotoxic actions of blood cell-released agents on the underlying tissue. A growing body of evidence suggests that these processes may be involved in pathophysiological cerebrovascular reactions including symptomatic angiospasm following subarachnoid hemorrhage, segmental occlusive constriction in atherosclerotic cerebral arteries, and constrictive vasomotion in microvessels. A perturbation in the delicate equilibrium between blood cells and endothelium in the microcirculation seems to be a factor aggravating ischemic brain damage or even primarily causing focal cerebral ischemia and scattered microinfarctions. Increased predisposition to these pathophysiologic events might influence unfavorably the effects of risk factors such as hypercholesterolemia, hypertension, and diabetes on cerebrovascular morbidity and mortality. Although the importance of blood cell/endothelium imbalance appears to be clear, its pharmacologic regulation is not sufficiently established. Some drugs have been demonstrated to limit platelet and/or leukocyte activity and protect the endothelial defense mechanisms, but the optimal therapeutic strategy has yet to be elaborated.

Animals↗
PubMed1998

Stochastic modeling and optimization of phage display.

Phage display, SELEX and other methods of combinatorial chemistry have become very popular means of finding ligands with high affinities to given targets. Despite their success, they suffer from numerous sources of error and bias, such as very low initial concentrations of species, non-specific binding, and the sampling of only a tiny fraction of the library at the end of an experiment. To understand the interaction of these errors and to better devise molecular search strategies that take the errors into account, I devise and analyze a highly detailed model of phage display. The model is specifically designed to study the influence of the stochastic nature of each laboratory step. The model includes phage multivalency, multiple classes of targets, and solid-phase equilibrium and washing, yet it is amenable to analytic results and rapid computer simulation. With both analytic and simulation approaches, I: (1) describe the effects of target concentration, phage valency, degree of background binding and other laboratory parameters on the probabilities of phage binding and of being selected; (2) show the effects of an increasing selection stringency strategy and how it results in a tradeoff between rapid library enrichment and high probability of sampling the best ligands; and (3) show how the number of phage sampled for detailed study at the end of a search alters search success. The work concludes with several practical suggestions for the control of selection stringency.

Bacteriophage M13↗
PubMed1999

Enzymatic synthesis of alpha-butylglucoside lactate: a new alpha-hydroxy acid derivative.

An alpha-hydroxy acid derivative, alpha-butylglucoside lactate, was successfully prepared by enzymatic transesterification of alpha-butylglucoside with a lactate alkyl ester in a non-aqueous medium using immobilized lipase as biocatalyst. Ester synthesis in organic solvent was optimized. Solvent choice was made on the basis of substrate solubility and enzyme stability in the medium. A solvent-free reaction using butyllactate as lactate donor led to the highest yields. In the presence of 0.5M alphabutylglucoside and 100 g/L Novozym(R), a 67 % yield could be obtained within 40 h at 50 degrees C. However, the presence of butanol by-product limited the reaction to a maximum that could not be exceeded in closed systems. The elimination of the alcohol under reduced pressure resulted in the complete equilibrium shift of the transesterification reaction in favor of synthesis; below 15 mbars, more than 95% of 0.5M alpha-butylglucoside could be converted within 30 h. Moreover, simultaneous evaporation of water allowed hydrolysis of butyllactate to be eliminated. Consequently, a very high alpha-butylglucoside lactate concentration (170 g/) could be obtained in a single batch reaction. A single purification procedure, consisting of butyllactate extraction with hexane, enabled the product to be obtained at a purity above 95% (w/w). 1H and 13C NMR analysis later demonstrated that lactic acid was exclusively grafted onto the primary hydroxyl group of alphabutylglucoside.

Bioreactors↗
PubMed2002

Selective recovery of lactate dehydrogenase using affinity foam.

Selective isolation of lactate dehydrogenase (LDH) from porcine muscle extract was studied using foam generated from the vigorous stirring of a non-ionic surfactant, Triton X-114 derivatized with Cibacron blue. The cloud point of the surfactant-dye conjugate was higher than that of the native Triton X-114, and also the foam prepared from the affinity surfactant was more rigid taking a longer time to collapse. The equilibrium dissociation constant between pure LDH and surfactant-dye conjugate was 5.0 microM as compared to the value of 2.2 microM for the enzyme and free dye as measured by differential spectroscopy. The isolation procedure involved mixing of the porcine muscle extract with the affinity foam, separating and collapsing the foam, and warming the solution formed to 37 degrees C to yield the surfactant-dye phase and an aqueous phase containing the enzyme. The effect of surfactant concentration and protein load on enzyme recovery and purification was investigated. Under optimal conditions, LDH was quantitatively recovered with high purification factor in a very short time. Both recovery and purification were higher when foam prepared from an equivalent mixture of surfactant-dye conjugate and unmodified surfactant was used. The selectivity of interaction between LDH and detergent-dye conjugate was confirmed by lowered recovery when NADH was included during the binding step.

Affinity Labels↗
PubMed1985

Quantitative 31P nuclear magnetic resonance analysis of metabolite concentrations in Langendorff-perfused rabbit hearts.

The quantitative analysis of the mobile high-energy phosphorus metabolites in isovolumic Langendorff-perfused rabbit hearts has been performed by 31P NMR utilizing rapid pulse repetition to optimize sensitivity. Absolute quantification required reference to an external standard, determination of differential magnetization saturation and resonance peak area integration by Lorentzian lineshape analysis. Traditionally accepted hemodynamic indices (LVDP, dp/dt) and biochemical indices (lactate, pyruvate) of myocardial function were measured concomitantly with all NMR determinations. Hemodynamically and biochemically competent Langendorff-perfused rabbit hearts were found to have intracellular PCr, ATP, GPC, and Pi concentrations of 14.95 +/- 0.25, 8.08 +/- 0.13, 5.20 +/- 0.58 and 2.61 +/- 0.47 mM respectively. Intracellular pH was 7.03 +/- 0.01. Cytosolic ADP concentration was derived from a creatine kinase equilibrium model and determined to be approximately 36 microM. Reduction of perfusate flow from 20 to 2.5 ml/min demonstrated statistically significant decreases in PCr, ATP, and pH as well as an increase in Pi that correlated closely with the independent hemodynamic and biochemical indices of myocardial function. The decrease in ATP and PCr concentrations precisely matched the increase in Pi during reduced flow. These results constitute the first quantitative determination of intracellular metabolite concentrations by 31P NMR in intact rabbit myocardium under physiologic and low flow conditions.

Adenosine Diphosphate↗