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At least 883 records · Page 49Linked to original sources
PubMed1992

Purification and properties of urease from the cyanobacterium Anabaena cylindrica.

A purification procedure has been developed by which urease activity in extracts from the cyanobacterium Anabaena cylindrica was enriched 500-fold. The procedure involves MgSO4 precipitation at 55 degrees C and chromatography on hydroxylapatite and diethylaminoethyl sephadex. Its molecular weight was measured by sedimentation equilibrium in an airfuge to be 197,000 +/- 2000 with an estimated subunit molecular weight of 32,000 as determined by polyacrylamide gel electrophoresis. The pH- and temperature-dependence of the enzyme were determined and the activity found to be optimal at pH 8 and 30 degrees C, respectively. The concentration-dependence of the activation of the enzyme by Mg++ was measured, as were the effects on activity of a range of other metal ions.

Anabaena↗
PubMed1993

Development and optimization of reactivation techniques for carbamate-inhibited brain and plasma cholinesterases in birds and mammals.

Two biochemical assays were developed which promote and measure the induced reactivation of carbamate-inhibited cholinesterases in avian and mammalian brain and plasma samples. The effects of inhibitor concentration, temperature, and the extent of dilution on the achievement of a steady state equilibrium and the subsequent level and rate of recovery of brain cholinesterase activity were investigated. A similar procedure for reactivation of carbamate-inhibited plasma cholinesterase activity involved the removal of excess carbamate from a small sample volume (< 400 microliters). Both methods begin by measuring cholinesterase activity immediately following dilution and involve an incubation period during which conditions for spontaneous reactivation of the inhibited enzymes are maximized. Both assays are suitable for large-scale, rapid use and appear able to restore inhibited cholinesterase activity to levels closely approximating that of control values for each species tested. These methods will not only maximize the usefulness of cholinesterases in monitoring carbamate pesticide exposure but should prove to be extremely useful tools in the forensic assessment of carbamate exposure in human and wildlife pesticide incidents.

Aldicarb↗
PubMed1987

The effect of shape and position of the oxygen equilibrium curve on extraction and ventilation requirement in fishes.

The effects of haemoglobin-oxygen affinity and of cooperativity in oxygen binding on the oxygen extraction and ventilation requirement in fish are analysed using the mathematical model of Malte and Weber (1985). It is shown that a high affinity may allow a substantial reduction in ventilatory requirement and that optimal O2 uptake depends on a delicate balance between affinity and cooperativity. Using P50 and nH values from the literature, the oxygen extraction values predicted by the model for carp and rainbow trout coincide neatly with the values found for these species, indicating that the differences in oxygen extraction are a direct result of different blood oxygen binding properties.

Animals↗
PubMed2003

Optimization of experimental conditions for the preparative displacement chromatography of antitumor anthracycline antibiotics on carboxylic sorbents.

The physico-chemical conditions and the limits of the rates of mobile phases are determined when effective regimes of preparative chromatography in the conditions of sharpening the boundaries of chromatographic zones of anthracycline antibiotics are realized. The influence of pH on the equilibrium, kinetics and dynamics of sorption of anthracycline antibiotics (rubomycin, doxorubicin and carminomycin) on the carboxylic sorbents has been studied.

Adsorption↗
PubMed2004

Physiological levels of hydrocortisone maintain an optimal chondrocyte extracellular matrix metabolism.

OBJECTIVE: To investigate the effects of physiological doses of hydrocortisone on synthesis and turnover of cell associated matrix (CAM) by human chondrocytes obtained from normal articular cartilage. METHODS: Human articular cartilage cells were obtained from visually intact cartilage of the femoral condyles of five donors and maintained in culture for one week to reach equilibrium in accumulated CAM compounds. 0, 0.05, 0.20, and 1.0 micro g/ml hydrocortisone was added to the nutrient media during the entire culture period. Cells were liberated and levels of CAM aggrecan, type II collagen, and fibronectin, of intracellular IGF-1, IL1alpha and beta, and of their respective plasma membrane bound receptors IGFR1, IL1RI, and the decoy receptor IL1RII, were assayed by flow cytometry. RESULTS: In comparison with controls, hydrocortisone treated chondrocytes, at all concentrations, expressed significantly higher plasma membrane bound IGFR1. Intracellular IGF-1 levels remained unchanged. Together with these changes, reflecting an increased ability to synthesise extracellular matrix (ECM) macromolecules, hydrocortisone treated cells expressed significantly higher amounts of the plasma membrane bound decoy IL1RII. Concurrently, intracellular IL1alpha and beta levels and membrane bound IL1RI were down regulated. Levels of CAM aggrecan, type II collagen, and fibronectin were significantly up regulated in the chondrocytes treated with hydrocortisone. CONCLUSION: 0.05 micro g/ml hydrocortisone treated chondrocytes had decreased catabolic signalling pathways and showed an enhanced ability to synthesise ECM macromolecules. Because IL1 activity was decreased and the expression of IL1RII decoy receptor enhanced, more of the ECM macromolecules produced remained accumulated in the CAM of the chondrocytes. The effects were obtained at doses comparable with physiological plasma levels of hydrocortisone in humans.

Adolescent↗
PubMedDate not supplied

An equilibrium ELISA for the dosage of Androctonus australis garzonii (Aag) and Buthus occitanus tunetanus (Bot) scorpion venoms: set up and calibration.

Scorpion stings are very frequent in Tunisia; yet a method for evaluating envenoming severity and consequently victim treatment, has never been adequately established nor has its efficiency been properly evaluated. Indeed, a management of envenomed patients requires the optimization of envenoming antivenom immunotherapy. This task requires either, an accurate evaluation of toxicokinetic parameters of scorpion venoms in absence and in presence of antivenom, using animals as models, and the establishment of a quantitative relationship between human blood scorption venom levels, envenoming severities and clinical symptoms. A performant sandwich ELISA was set up and calibrated for measuring scorpion venom levels in human and rabbit sera. This assays was performed with polyclonal F(ab')2 specific to the two North African scorpion (Androctonus australis garzonii; Aag and Buthus occitanus tunetanus: Bot) venoms. It is simple, rapid, very sensitive (detection limit = 0.9 ng/ml) and shows a good linearity for venom concentrations in human sera comprised between 0.5 and 15 ng/ml. The ELISA is also reproducible: the coefficient of variation, determined at different venom concentrations (low: 4 ng/ml; medium: 8 ng/ml and high: 12 ng/ml) prepared in a pool of sera collected from several healthy donors, were lower than 10%. Such an ELISA has been successfully used, either in experimental toxinokinetic and immunotherapeutic studies carried out in rabbits or for the quantification of Aag and Bot venom levels in the serum of human victims stung by these scorpion.

Animals↗
PubMed1995

Mammalian protein geranylgeranyltransferase-I: substrate specificity, kinetic mechanism, metal requirements, and affinity labeling.

Protein geranylgeranyltransferase-I (PGGT-I) catalyzes the transfer of the 20-carbon prenyl group from geranylgeranyl pyrophosphate to the cysteine residue near the C-termini of a variety of eukaryotic proteins. Kinetic analysis of homogenous PGGT-I from bovine brain reveals that the reaction follows a sequential pathway in which either prenyl donor or acceptor can bind first to the enzyme and that the reaction operates at steady-state rather than at rapid equilibrium. Substrate inhibition by prenyl acceptor but not by prenyl donor suggests that geranylgeranyl pyrophosphate binding first to free enzyme is the kinetically preferred pathway. This is supported by isotope trapping experiments which show that the ternary complex goes on to products faster than the release of geranylgeranyl pyrophosphate from the complex. The KM for the interaction of geranylgeranyl pyrophosphate with PGGT-I is markedly affected by the structure of the prenyl acceptor bound to the enzyme. A detailed analysis of the substrate specificity of PGGT-I reveals that peptides which contain a C-terminal leucine are preferred (kcat/KM = 1-5 x 10(5) M-1 s-1) to those that end in serine (kcat/KM = 2-4 x 10(3) M-1 s-1) or phenylalanine (kcat/KM = 0.5 x 10(3) M-1 s-1). PGGT-I also catalyzes the farnesylation of peptides that have a C-terminal leucine; kcat for farnesylation and KM for farnesyl pyrophosphate are similar to those for geranylgeranylation, but the KM for the peptide is 30-fold higher. Geranyl pyrophosphate is utilized by PGGT-I but is a poor substrate. Optimal activity of PGGT-I is obtained in the presence of micromolar amounts of Zn2+ and mM amounts of Mg2+. Mn2+ or Cd2+ but not Co2+ can substitute for Zn2+ and for Mg2+. Metals are not required for tight-binding of geranylgeranyl pyrophosphate to PGGT-I, and the measured dissociation equilibrium constant for this binary complex is 16 nM. Photoaffinity analogues of geranylgeranyl pyrophosphate and farnesyl pyrophosphate were prepared and shown to exclusively label the beta-subunit. The implication of the results for the substrate specificity of protein prenylation in cells is briefly discussed.

Affinity Labels↗
PubMed2005

Single-step ion exchange purification of the coagulant protein from Moringa oleifera seed.

The coagulant protein from Moringa oleifera (MO) seed was purified using a single-step batch ion exchange (IEX) method. Adsorption and elution parameters were optimized. Impact of the purification on the reduction of organic and nutrient release to the water was studied. The matrix was equilibrated using ammonium acetate buffer, and the optimum ionic strength of NaCl for elution was 0.6 M. The time for adsorption equilibrium was between 90 and 120 min. Maximum adsorption capacity of the matrix, estimated with the Langmuir model, was 68 mg protein/g adsorbent. The purified protein does not release organic and nutrient loads to the water, which are the main concerns of the crude extract. This work suggests that a readily scalable single-step IEX purification method can be used to produce the coagulant protein and it can be carried out with locally available facilities. This will promote the use of MO in large water treatment plants and other industries.

Chromatography, Ion Exchange↗
PubMed1993

Random and site-specific immobilization of catalytic antibodies.

The effects of immobilization on the immunologic and catalytic activity of a catalytic antibody were compared for randomly immobilized (via glutaraldehyde) whole antibody and site-specifically immobilized (via the reactive sulfhydryl group at the base of the fragment) Fab' fragments. Upon immobilization, the specific binding capacity (n) and the catalytic activity decreased significantly for both systems. Increases in the Michaelis constant (KM) were accompanied by corresponding decreases in the equilibrium binding constant determined through immunoassays. For the immobilized Fab', n decreased dramatically with increased protein loading, suggesting that, despite the site-specific attachment and favorable orientation, molecular crowding denatured the Fab' fragments. These results also show that there is an optimal surface coverage, not necessarily at the maximum loading, for both immunologic and catalytic activity. Finally, the combining/active site conformation was probed using electron paramagnetic resonance (EPR) spectroscopy. In all antibody samples, there was no spectral evidence of conformational changes in the antibody active site.

Animals↗
PubMed2003

A new approach for immobilization of oligonucleotides onto piezoelectric quartz crystal for preparation of a nucleic acid sensor for following hybridization.

The aim of this study is to develop a nucleic acid sensor based on piezoelectric crystal microbalance system (QCM) for following hybridization. Piezoelectric quartz crystal surfaces were first treated in a glow-discharge apparatus with ethylene diamine (EDA) plasma at 15 W (discharge power), 2.5 min (incubation time) and 35 ml/min (monomer flow rate) to create amino groups on the crystal surfaces. The thickness of the EDA-plasma film formed was about 43+/-24 A. Then, the amino groups on the crystal surfaces were converted to aldehyde groups by reacting the amino groups with glutaraldehyde (GA) at different conditions. A GA concentration of 2.5% and an incubation time of 2 h were selected as optimal values at this step, corresponding to a GA surface concentration of about 270 ng/cm2. A double strand Oligonucleotides, having one extra base on 5'-end of one of the complementary strands, were immobilized through the amino groups of this base onto the GA-modified crystals. Optimal immobilization conditions were as follows: oligonucleotide concentration: 1 microg/ml; time: 3 h; pH: 9.2 carbonate buffer; ionic strength: 0.1; and temperature: 20 degrees C. The QCM sensor carrying the covalently bound strand was used in the hybridization experiments, which showed that equilibrium is achieved in about 5 min, and the frequency shift measured is related to the concentration of the target strand to be measured within the medium.

Adsorption↗
PubMed2005

Mathematical model describing gradient focusing methods for trace analytes.

The problem of gradient focusing for concentrating trace analytes is considered. Variation of buffer viscosity, conductivity, and possibly also the zeta-potential results in a focusing point where the electrophoretic velocity is balanced by the electroosmotic flow (EOF) and where the sample concentrates. The axial inhomogeneity also results in an induced pressure gradient that alters the EOF profile and therefore causes Taylor dispersion. The coupled hydrodynamics and transport problem leading to the achievement of a steady state is studied in the context of the lubrication approximation: all variations in the axial direction take place over a length scale very much larger than the characteristic channel width. A single length scale sigma(m) and a single time scale tau is found to completely determine the dynamics of the evolution close to the focusing point. Using appropriate scaled variables, the time evolution of the concentration profile near equilibrium can be described by an inhomogeneous advection diffusion equation that is free of all parameters. Explicit formulas are deduced for the location of the peak centroid and its width as a function of time. A simple graphical method is proposed for optimizing the performance of the system when some tunable external parameters are available.

Journal Article↗
PubMed1998

Determination of cannabinoids in water and human saliva by solid-phase microextraction and quadrupole ion trap gas chromatography/mass spectrometry.

Solid-phase microextraction (SPME) is applied to the determination of cannabidiol, delta 8-tetrahydrocannabinol (delta 8-THC), delta 9-tetrahydrocannabinol (delta 9-THC), and cannabinol in pure water and human saliva. The inherent extraction behavior of the cannabinoids in pure water is evaluated along with optimization of the method in human saliva. The commercially available poly(dimethylsiloxane) (PDMS) SPME fibers were found to be the best class for the cannabinoid analysis. Partition coefficients were found to be extremely large for all of the cannabinoids (log K > 4.0). Equilibrium times for the 7- and 30-micron PDMS fibers were 50 and 240 min, respectively. A shorter extraction time of 10 min with the 30-micron PDMS fiber may be used for multiple extractions from the same vial, thus conserving the sample necessary for analysis and speeding up the total analysis time. Recoveries for the cannabinoids in saliva, relative to pure water, were dramatically improved by a method developed in our laboratory involving addition of glacial acetic acid to the sample vial prior to performing SPME. Using this method, recoveries relative to SPME in pure water ranged from 21 to 47% depending on the cannabinoid. The linear range for spiked saliva samples was established at 5-500 ng/mL (r2 > 0.994) with precisions between 11 and 20% RSD. The ultimate level of detection by SPME for the cannabinoids in saliva was 1.0 ng/mL, with signal-to-noise values of > or = 12. A saliva sample collected 30 min after marijuana smoking was subject to SPME and traditional liquid-liquid extraction analysis. Internal standard quantitation results for delta 9-THC by both methods yielded comparable results, indicating that the SPME method of analysis is highly accurate and precise. The level of delta 9-THC by SPME was found to be 9.54 ng/mL for the saliva sample.

Cannabinoids↗
PubMed2005

Heats of formation of xenon fluorides and the fluxionality of XeF(6) from high level electronic structure calculations.

Atomization energies at 0 K and heats of formation at 0 and 298 K are predicted for XeF(+), XeF(-), XeF(2), XeF(4), XeF(5)(-), and XeF(6) from coupled cluster theory (CCSD(T)) calculations with new correlation-consistent basis sets for Xe. To achieve near chemical accuracy (+/-1 kcal/mol), up to four corrections were added to the complete basis set binding energies based on frozen core coupled cluster theory energies: a correction for core-valence effects, a correction for scalar relativistic effects, a correction for first-order atomic spin-orbit effects, and in some cases, a second-order spin-orbit correction. Vibrational zero-point energies were computed at the coupled cluster level of theory. The structure of XeF(6) is difficult to obtain with the C(3)(v)() and O(h)() structures having essentially the same energy. The O(h)() structure is only 0.19 kcal/mol below the C(3)(v)() one at the CCSD(T)/CBS level using an approximate geometry for the C(3)(v)() structure. With an optimized C(3)(v)() geometry, the C(3)(v)() structure would probably become slightly lower in energy than the O(h)() one. The calculated heats of formation for the neutral XeF(n)() fluorides are less negative than the experimental values from the equilibrium measurements by 2.0, 7.7, and 12.2 kcal/mol for n = 2, 4, and 6, respectively. For the experimental values, derived from the photoionization measurements, this discrepancy becomes even larger, suggesting a need for a redetermination of the experimental values. Evidence is presented for the fluxionality of XeF(6) caused by the presence of a sterically active, free valence electron pair on Xe.

Journal Article↗
PubMed2006

Underwater superhydrophobicity: theoretical feasibility.

The possibility of underwater superhydrophobicity is theoretically analyzed. Thermodynamic equilibrium and stability conditions are formulated, and the design goal is defined as minimizing the solid-liquid contact area. It is shown that for sufficiently high roughness ratios, underwater superhydrophobicity may be feasible and thermodynamically stable. In addition, some generic design optimization considerations are demonstrated.

Journal Article↗
PubMed1990

Separate [3H]-nitrendipine binding sites in mitochondria and plasma membranes of bovine adrenal medulla.

1. Two binding sites for the 1,4-dihydropyridine (DHP) derivative [3H]-nitrendipine have been found in the bovine adrenal medulla. The high-affinity site (Kd = 0.48 nM and Bmax = 128 fmol mg-1 protein) was specifically located in purified plasma membranes. The low-affinity site (Kd = 252 nM and Bmax = 169 pmol mg-1 protein) was located only in mitochondria. Chromaffin granule membranes lacked specific binding sites for [3H]-nitrendipine. 2. Kinetic analysis of the rates of association and dissociation of [3H]-nitrendipine, saturation isotherms and displacement experiments with unlabelled nitrendipine and PN200-110 revealed single, homogeneous populations of high- and low-affinity sites in plasma and mitochondrial membranes, respectively. 3. The high affinity site was sensitive to Ca2+ deprivation and heating; it was practically unaffected by changes in ionic strength of the medium and its optimal pH was slightly alkaline. This site exhibited a strong DHP stereoselectivity; diltiazem increased and verapamil decreased the affinity of [3H]-nitrendipine. 4. In contrast, binding of [3H]-nitrendipine to the low affinity site was more heat resistant and less affected by Ca2+ removal. Its optimal pH was slightly acid and the increase in ionic strength enhanced the number of available sites. The site had no DHP stereoselectivity. Verapamil decreased the dissociation constant of [3H]-nitrendipine acting in a non-competitive manner; diltiazem did not affect equilibrium binding parameters of [3H]-nitrendipine. 5. These results suggest that both biding sites reflect different receptor entities. The high-affinity binding site corresponds to the dihydropyridine receptor associated with the L-type calcium channel. The function of the mitochondrial, low-affinity binding site is, at present, unknown.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗
PubMed2004

Arc process parameters for single-walled carbon nanotube growth and production: experiments and modeling.

Collarets rich in single-walled carbon nanotubes (SWCNTs) have been grown using a direct current arc method. Arc process parameters such as current, pressure, and anode to cathode distance were varied experimentally and by modeling to provide an optimal working window. The best collaret yields were obtained when helium was used as a buffer gas. Mixing helium with argon in the buffer permits controlling nanotube diameters. In addition to an experimental study, a modeling approach was developed assuming local thermal equilibrium and homogenous and heterogeneous neutral chemistry. The gas-phase chemical model involves 81 neutral carbon species (C1, C2, . . ., C79, C60F, C70F) and 554 reactions with rates taken from data of Krestinin and Moravsky. Axial profiles of temperature, C atom, C2 radical, and fullerene distributions in the reactor are predicted as a function of process parameters. Carbon nanotube growth is considered by a set of surface reactions simulating open nanotube growth. Because nanotube surface chemistry is controlled by the local terminated bond and not by the bulk nanotube bond, a mechanistic approach based on the formal resemblance between the bonding and the structure of open nanotube and other carbon surfaces is proposed to explain nanotube growth. Predicted growth rates are in the range of 100 to 1000 microm/min.

Argon↗
PubMed2006

Double hepatic arterial phase MRI of the liver with switching of reversed centric and centric K-space reordering.

OBJECTIVE: The purpose of our study was to evaluate the clinical feasibility and usefulness of a 2D spoiled gradient-recalled echo MR sequence with serial switching of reversed centric and centric k-space reordering for high-spatial-resolution gadolinium-enhanced double hepatic arterial phase (HAP) MRI of the liver. SUBJECTS AND METHODS: MR images (frequency, 512; phase encoding without interpolation, 224; 6-mm thickness with 1-mm gap; 30 slices per 18 seconds) were obtained with multiphase imaging in which central k-space line data were filled 10, 21, 49, and 181 seconds after arrival of contrast medium in the abdominal aorta for the early HAP (reversed centric reordering, center of k-space lines acquired at end of acquisition), late HAP (centric reordering, center of k-space lines at beginning of acquisition), portal venous phase (centric reordering), and equilibrium phase (centric reordering), respectively, in 102 consecutive patients with suspected liver disease, including 48 untreated hepatocellular carcinomas (HCCs) in 35 patients. Images were quantitatively assessed for degree of contrast enhancement in the abdominal aorta, spleen, portal trunk, liver parenchyma, hepatic veins, and HCCs. Images were qualitatively assessed for the effectiveness of contrast enhancement in each phase and for degree of image degradation due to artifacts. RESULTS: Enhancement of the abdominal aorta peaked in the early HAP, of the portal trunk in the late HAP, and of the hepatic parenchyma and veins in the portal venous phase. Mean HCC-to-liver contrast peaked in the early HAP and turned to a negative value in the portal venous and equilibrium phases. Sufficient image quality was achieved in 99 (97%) of the patients. One of the other three patients had motion artifacts due to body motion, and the other two had unsatisfactory respiratory suspension. Scan timing for early and late HAP was optimal in 74 (73%) of the patients, for late HAP lagged in 20 (20%), for early HAP was premature in six (6%), and for early HAP lagged in five (5%) of the patients. CONCLUSION: We confirmed the feasibility and usefulness of a 2D gadolinium-enhanced double HAP spoiled gradient-recalled echo sequence incorporating serial switching of reversed centric and centric k-space reordering. This method has the potential for use in high-spatial-resolution double HAP MRI for the diagnosis of hypervascular HCC.

Adult↗
PubMed1987

[Parenteral use of a fat emulsion in the early postoperative period].

Parenteral application of fat emulsions has been established as useful in long term artificial nutrition especially during weaning off the ventilator. In order to ascertain whether these advantages can also be proved in nonventilated patients in the immediate postoperative phase, alternatively carbohydrates versus a new fat emulsion were investigated. 30 patients were randomly allocated to three different nutrition regimens. The caloric and nitrogen supplementation was identical for all groups, while the carbohydrate and fat content was different. Group I: only carbohydrates (as caloric source), group II: high fat load (3 g/kg/day), group III: medium fat dosage (1.5 g/kg/day). Routine laboratory parameters as well as acid-base-status and fat metabolism were evaluated. While most of the routine parameters were not significantly different between the groups, triglyceride levels in serum were highest in group II but never exceeding 280 mg/dl (-x). Nitrogen balance as well as protein metabolism were significantly improved using regimen III, thus documenting the beneficial effect of a medium fat dosage in the immediate postoperative phase. Parenteral nutrition combining amino acids, carbohydrates and a fat emulsion seems to be the optimal postoperative regimen even in patients without ventilatory support.

Abdomen↗