nitrophenols and Hemolysis

Via the solvothermal reaction between Zn(II) or Mn(II) salts and 5-(3,4-dicarboxylphenoxy)nicotinic acid (H

Topics: Animals; Atherosclerosis; Coordination Complexes; Cytokines; Endothelial Cells; Enzyme-Linked Immunosorbent Assay; Hemolysis; Humans; Ligands; Manganese; NF-kappa B; Niacin; Nitrophenols; Polymers; Real-Time Polymerase Chain Reaction; Signal Transduction; Spectrometry, Fluorescence; Zinc

Adaptation of the colorimetric method for the determination of β-d-galactosidase, β-d-glucuronidase and α-l-fucosidase activities in serums from hemolyzed blood, the material currently being discarded.. The materials included serums from hemolyzed and non-hemolyzed blood, obtained from 26 healthy volunteers. The adaptation of the method involved precipitation of the proteins with trichloroacetic acid after incubating serums with substrates, but before determining the products of enzymatic reactions.. In serums from hemolyzed and non-hemolyzed blood of the same persons, we found high correlations among the results obtained using hemolyzed blood (with adapted) and non-hemolyzed blood (with non-adapted) methods.. We are able to determine the β-d-galactosidase, β-d-glucuronidase and α-l-fucosidase activities in serums from hemolyzed blood (with adapted) and non-hemolyzed blood (with non-adapted) methods, with the same accuracy and precision.

Topics: Adult; alpha-L-Fucosidase; beta-Galactosidase; Female; Glucuronidase; Hemoglobins; Hemolysis; Humans; Hydrogen-Ion Concentration; Kinetics; Male; Middle Aged; Nitrophenols; Young Adult

To prepare new copolymers, useful for gene delivery, based on alpha, beta-poly-(N-2-hydroxyethyl)-D, L-aspartamide (PHEA) as a polymeric backbone and bearing an oligoamine such as diethylenetriamine in the side chain. Moreover, in order to reduce solvent volume and make the reaction faster, microwave-assisted heating was used.. PHEA copolymers bearing different amounts of diethylenetriamine were prepared using bis(4-nitrophenyl) carbonate as a condensing agent with the use of microwaves. Chemical, physico-chemical and biological characterization of PHEA-diethylenetriamine copolymers and their complexes obtained with DNA were performed.. Copolymers showed good DNA complexing and condensing abilities depending on the oligoamine derivatization degree and good hemocompatibility. Moreover, plasmid DNA/copolymer polyplexes showed very good cytocompatibility on B16F10 and N2A cell lines.. Results support the use of these copolymers as gene delivery systems in the future. Finally, the use of microwaves makes the proposed synthetic method advantageous as time and solvents are saved.

Topics: Animals; Cell Line, Tumor; Cell Survival; DNA; Erythrocyte Aggregation; Gene Transfer Techniques; Hemolysis; Humans; Mice; Microwaves; Nitrophenols; Polyamines; Polyhydroxyethyl Methacrylate; Polymers

Two commercial amylase methods which employ p-nitrophenol derivatives of blocked maltoheptaosides suffer from negative interference due to fresh haemolysate. Specimen storage at room temperature, or pre-incubation at 37 degrees C or 57 degrees C removes the effect. Incubation of amylase reagent at 37 degrees C with fresh haemolysate where the final haemoglobin concentration was 0.011 g/L, showed a rapid fall in absorption around 414 nm which became stable after 150 min. Since p-nitrophenol, the product of the amylase reaction, is measured at 405 nm it is concluded that the negative interference from fresh haemolysis is due to the transitory fall in absorption around 405 nm. It is recommended that amylase measurements using this technique, particularly those performed as an emergency, should not be done on haemolysed specimens.

Topics: Amylases; Blood Chemical Analysis; Chromogenic Compounds; Drug Storage; Enzyme Stability; Glucosides; Hemoglobins; Hemolysis; Humans; Nitrophenols; Spectrophotometry, Ultraviolet; Temperature; Time Factors

Cell lines have been established that secrete a matched set of human chimeric IgM, IgG1, IgG2, IgG3, IgG4, IgE, and IgA2 antibodies that are directed against the hapten 4-hydroxy-3-nitrophenacetyl. These chimeric antibodies secreted from mouse plasmacytoma cells behave exactly like their authentic human counterparts in SDS-PAGE analysis, binding to protein A and in a wide range of serological assays. The antibodies have been compared in their ability to bind human C1q as well as in their efficacy in mediating lysis of human erythrocytes in the presence of human complement. A major conclusion to emerge is that whereas IgG3 bound C1q better than did IgG1, the chimeric IgG1 was much more effective than all the other IgG subclasses in complement-dependent hemolysis. The IgG1 antibody was also the most effective in mediating antibody-dependent cell-mediated cytotoxicity using both human effector and human target cells. These results suggest that IgG1 might be the favoured IgG subclass for therapeutic applications.

Topics: Animals; Antibodies, Monoclonal; Antibody-Dependent Cell Cytotoxicity; Cell Line; Complement Activating Enzymes; Complement C1; Complement C1q; Complement System Proteins; DNA, Recombinant; Electrophoresis, Polyacrylamide Gel; Genes, Immunoglobulin; Glycosylation; Haptens; Hemolysis; Humans; Immunoglobulins; Mice; Nitrophenols; Phenylacetates; Plasmacytoma; Plasmids; Transfection; Tumor Cells, Cultured

The non-dialysable fraction of haemolysate causes an apparent reduction of plasma alkaline phosphatase (ALP) activity using 4-nitrophenylphosphate as substrate. Analyses using four different buffers showed that the decrease in enzyme activity is affected by the buffer used. The percentage reduction in ALP activity is dependent on the initial ALP activity but not on the isoenzyme present. When diethanolamine was used as buffer, sample blanking almost completely compensated for the apparent reduction in enzyme activity. However, when aminomethylpropanol, aminomethylpropanediol and tris-carbonate buffers were used, it appeared that haemolysate reduced the catalytic activity of the enzyme, since sample blank correction had minimal effect on the results.

Topics: Alkaline Phosphatase; Buffers; Hemolysis; Humans; Nitrophenols; Organophosphorus Compounds; Spectrophotometry

Snake venom phospholipases A2 show a remarkable degree of amino acid sequence homology yet differ markedly in enzymatic and pharmacological activities. The basic phospholipase A2 from Naja nigricollis venom has much greater lethal potency, cardiotoxicity, hemolytic and anticoagulant activity than the acidic or neutral enzymes from Naja naja atra or Hemachatus haemachatus venoms, respectively, even though it has lower enzymatic activity than the latter two enzymes. Previous studies in which we selectively modified lysine and free carboxyl groups suggested that the pharmacological and enzymatic active sites are not identical. Tryptophan residues have been suggested as being involved in substrate binding although some phospholipases have no tryptophan. We investigated the effect of alkylating the tryptophans in N. nigricollis, N. n. atra, and H. haemachatus phospholipases A2 with 2-hydroxy-5-nitrobenzyl bromide. Chemical modification caused decreases in enzymatic activity, although the extent of inactivation varied with the enzyme and with the substrate (lecithin micelles, egg yolk, heart homogenates). The specificity of the enzymes for individual phospholipid substrates was not affected. Alkylation of the tryptophans also caused decreases in lethal, hemolytic, anticoagulant, and cardiotoxic potencies, which were similar to the extents of decrease in enzymatic activity. Our results suggest that tryptophans are not specifically associated with either the enzymatic or the pharmacological active site nor are essential for either activity.

Topics: 2-Hydroxy-5-nitrobenzyl Bromide; Alkylation; Animals; Elapid Venoms; Electric Stimulation; Heart Ventricles; Hemolysis; Kinetics; Male; Nitrophenols; Phospholipases; Phospholipases A; Phospholipases A2; Rats; Rats, Inbred Strains; Snakes; Species Specificity; Substrate Specificity; Tryptophan; Ventricular Function

The primary anti-(4-hydroxy-3-nitrophenyl)acetyl (anti-NP) antibody response of C57BL/6 mice was analyzed by several methods. Serum analyses by solid-phase radioimmunoassay (RIA) showed that stimulation with the thymus-independent (TI) type 1 antigen NP-LPS results in an anti-NP antibody response dominated by kappa (kappa) light chain bearing antibodies, whilst responses to NP-Ficoll (a TI-2 antigen) and NP-KLH (a thymus dependent antigen) are dominated by lambda (lambda) 1 light chain bearing antibodies. However, in all these responses NP-specific plaque-forming cells (PFCs) were predominantly heteroclitic, and inhibitable by anti-lambda antiserum. In addition, kappa-bearing IgM-producing hybridomas obtained by fusion of spleen cells from NP-LPS-immunized mice, although producing NP-specific antibodies detectable by RIA, were unable to produce NP-specific plaques. Direct determination of the affinity of 5 of those hybridomas by fluorometric titrations showed that their affinity is indeed lower than 10(-5) M. These results suggest that most NP-specific antibodies stimulated by NP-LPS are of too low affinity to be detected in a direct PFC assay, with the exception of a population of lambda-bearing antibodies. Therefore, the differential expression of kappa- or lambda-bearing antibodies in primary responses to the hapten NP presented on different carriers may be due to different affinity requirements for B-cell triggering via different activation pathways.

Topics: Animals; Antibody Affinity; Haptens; Hemolysis; Hemolytic Plaque Technique; Immunoglobulin kappa-Chains; Immunoglobulin lambda-Chains; Immunoglobulin Light Chains; Mice; Mice, Inbred C57BL; Nitrophenols; Phenylacetates; Radioimmunoassay

We have previously proposed the hypothesis that asymmetric membranes behave like bilayer couples: the two layers of the bilayer membrane can respond differently to a particular perturbation. Such a perturbation, for example, can result in the expansion of one layer relative to the other, thereby producing a curvature of that membrane. In experiments with erythrocytes and lymphocytes, we now demonstrate that different membrane perturbations which have opposite effects on membrane curvature can compensate and neutralize one another, as expected from the bilayer couple hypothesis. This provides a rational basis, for example, for understanding the effects of amphipathic drugs on a variety of cellular phenomena which involve shape changes of membranes.

Topics: Adenosine Triphosphate; Animals; Antigen-Antibody Reactions; Azides; Cell Membrane; Chlorpromazine; Dinitrophenols; Endocytosis; Erythrocytes; Hemolysis; Humans; Lymphocytes; Lysophosphatidylcholines; Mice; Nitrophenols; Oleic Acids

Topics: Animals; Antibodies; Cell Membrane; Chickens; Chromium Radioisotopes; Cytotoxicity Tests, Immunologic; Dinitrophenols; Erythrocytes; Female; Hemagglutination Tests; Hemolysis; Immune Sera; Immunity, Cellular; Lymphocytes; Lysine; Mice; Mice, Inbred BALB C; Nitrophenols; Rabbits; Spleen

Topics: Azides; Binding Sites; Cell Membrane; Chromatography, Ion Exchange; Densitometry; Electrophoresis, Polyacrylamide Gel; Erythrocytes; Hemolysis; Humans; Nitrophenols; Osmolar Concentration; Permeability; Sulfonic Acids; Sulfur Radioisotopes; Taurine; Tritium

Topics: Acid Phosphatase; Animals; Bacteriological Techniques; Coagulase; Deoxyribonucleases; Evaluation Studies as Topic; Fermentation; Hemolysis; Indoles; Mannitol; Microbial Sensitivity Tests; Nitrophenols; Organophosphorus Compounds; Penicillinase; Penicillins; Phenols; Pigments, Biological; Rabbits; Sheep; Spectrum Analysis; Staphylococcus

Topics: Animals; Catalysis; Chloromercuribenzoates; Esterases; Esters; Hemolysis; Hydrogen Peroxide; Hydrogen-Ion Concentration; Hydrolysis; Mercaptoethanol; N-Glycosyl Hydrolases; Nitrophenols; Organophosphorus Compounds; Phenylacetates; Sheep; Streptolysins; Sulfhydryl Compounds

Topics: Acrylates; Antibodies; Antigens; Complement Fixation Tests; Dinitrophenols; Electrophoresis; Erythrocytes; gamma-Globulins; Gels; Hemagglutination Tests; Hemolysis; Isoelectric Focusing; Methods; Nitrophenols; Proteins; Urea