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PubMed1987

Methionine residue accessibility in native subtilisin DY.

The three methionine residues of subtilisin DY were specifically modified into methionine sulfoxide using increasing amounts of chloramine T. By means of subsequent treatment with cyanogen bromide, gel chromatography, Edman degradation of the obtained peptides and the known structure of subtilisin DY it was established that Met222 is exposed to the surrounding solution, Met124 is partially exposed and Met199 is buried. The data obtained were confirmed on a computer graphics space movable model of subtilisin Carlsberg where Met222 was seen to be on the surface of the molecule and Met199 shielded by Tyr262, Ala179 and Leu196. Upon oxidation of Met222 of subtilisin DY by chloramine T, 25% of its caseinolytic activity was lost. This can be explained by the immediate adjacency to the active-site Ser221. An additional 5% loss of activity was observed at each subsequent methionine modification.

Amino Acid Sequence↗
PubMed2002

Myeloperoxidase-mediated protein oxidation: its possible biological functions.

Oxidation of proteins occurs both as a side-effect of aerobic energy metabolism and as an effect of specific metabolism of phagocytic polymorphonuclear granulocytes producing O2- and H2O2. In contrast to other cells, which control their H2O2 level by degrading it to O2 and H2O, polymorphonuclear neutrophilic leukocytes (PMN) use H2O2 as a substrate for oxidizing chloride ions to HOCl which rapidly react with all neighboring thiol, disulfide and amino residues. Chloramines, which are the most abundant HOCl reaction products, react with proteins, modifying only certain exposed methionine and cysteine residues. This may account for selective inactivation of a number of enzymes, carrier proteins and peptide mediators, including the alpha1-proteinase inhibitor, alpha2-macroglobulin and plasminogen activator inhibitor. Inactivaton of plasma proteinase inhibitors protects PMN elastase, collagenase, cathepsin G and other serine proteases in the inflammatory foci. This promotes proteolytic degradation of damaged tissue, removal of bacterial debris and wound healing, as well as tissue remodeling related to the inflammatory processes. Oxidative control of protease-anti-protease balance affects the development of the inflammatory processes. Moreover, inactivation of plasma proteinase inhibitors facilitates primary antigen processing, upregulates lymphocyte proliferative response and activates the local immune response. Oxidation produces a specific protein tagging which attracts and stimulates immune active cells. Therefore, humoral response against oxidatively modified proteins occurs more effectively than that of the native proteins. The effect is dose-dependent with respect to the amount of oxidant employed. Glycol aldehyde, which is the serine chloramine spontaneous decay product, in mice immunized with glycol aldehyde-modified egg-white albumin, yields specific IgG production manifold higher than that in mice immunized with native albumin. Immunopotentiation is produced by proliferation expansion of the same immunocompetent clones. Oxidative tagging of proteins may also affect the autoimmune-type reaction. Thus, a growing body of data suggest that the specific role of protein oxidation by activated PMN is oxidative protein tagging facilitating further development of the immune reaction.

Animals↗
PubMed1980

Iodination of thyroid-stimulating hormone for receptor-binding studies with human thyroid membranes: effects of specific activity and method of iodination.

Three different methods were compared for 125I-labelling of thyroid-stimulating hormone (TSH) for use in receptor-binding studies with human thyroid membranes: these were the chloramine-T, Bolton-Hunter and lactoperoxidase methods. Chloramine-T proved to be an inferior method to the other two. Iodinations to different specific activity (0.7--9.4 Bq/pg) were also compared: too high a specific activity led to reduced binding and a dramatic shift in the pH optimum for the TSH-receptor interaction. A specific acitivity of 2.5 Bq/pg should not be exceeded if binding of 125I-labelled TSH is to be representative of the binding of the natural hormone. Under these conditions, pH 7.5 was optimal for binding of TSH to its receptor. Repurification of the labelled TSH by receptor adsorption also proved to be essential.

Chloramines↗
PubMed1976

Some considerations in the preparation of raioiodoinsulin for radioimmunoassay and receptor assay.

125I-insulins, prepared by iodination with chloramine T in marked excess or by stepwise, stoichiometric addition of the oxidizing agent, were compared with respect both to their molecular distribution of iodine and to their suitability for use in a cultured lymphocyte receptor assay. Iodination of insulin in aqueous solution results in the same distribution of iodine atoms, independent of experimental method and dependent only on the average iodine number. This distribution can be calculated on the basis of a Monte Carlo simulation, For insulin iodinated at an average of 0.8 I atoms per molecule, approximately 50 per cent of the radioactivity is in other than monoiodoninsulin. Purification methods that separate on the basis of charge, such as starch-gel electrophoresis, are then required, to obtain monoiodoinsulin. More highly iodinated insulin do bind to the lymphocyte receptor, although, as in radioimmunoassay, the overiodinated species are less satisfactory for use as tracers. The shelf life of iodinated insulin appears to be related better to the average iodine content than to any other factor, presumably because of decay catastrophe. There is no evidence to suggest that exposure to chloramine T in marked excess for a few seconds is deleterious to insulin.

Chloramines↗
PubMed1976

Evaluation of chlorine compounds for surface disinfection by laboratory and in-use testing.

The disinfecting properties of chloramine and compounds containing chlorinated trisodium phosphate and potassium bromide or sodium dichlorisocyanurate and detergents were studied by laboratory and in-use tests and compared with a phenolic disinfectant containing detergents. In a modified Kelsey test all the preparations were effective in the recommended dilutions in clean conditions. The effectiveness of the chlorine-bromine disinfectant substantially decreased in the presence of organic material. The in-use testing was performed in infectious disease and intensive care wards. Chloramine was so disliked that the in-use test could not be carried out. In the infectious disease ward the total bacterial colony counts of the floors (disinfected once a day) were twice as high as those of the intensive care ward (disinfected 3 times a day) during the use of the phenolic and chlorine-bromine disinfectants. The frequent cleaning routine seemed thus to have an effect on the microbial contamination of the floors. The same difference was found in the contamination of the other non-vertical surfaces although they were disinfected only once a day in both wards. In both wards the total bacterial colony counts of all non-vertical surfaces were lower during the use of halogen compounds than during the use of the phenolic disinfectant. The staff, however, complained of the irritation of the skin and the mucous membranes when using chlorine disinfectants.

Chloramines↗
PubMed2001

Pathology associated with an aquareovirus in captive juvenile Atlantic halibut Hippoglossus hippoglossus and an experimental treatment strategy for a concurrent bacterial infection.

A large-scale mortality of larval and juvenile halibut Hippoglossus hippoglossus occurred at a semi-commercial halibut farm in Atlantic Canada. Investigation of the cause revealed aquareovirus particles in necrotic liver tissue of affected fish. Cytopathic effect on CHSE-214 cell lines occurred from all fish cultured for viruses, and the viral morphology of the particles in culture was consistent with that observed in necrotic host tissue. The virus was placed in the family of Reoviridae, genus Aquareovirus based on morphology and RT-PCR results. Multifocal hepatocellular necrosis was a consistent finding in all fish as well as acute necrosis of proximal renal tubules. Concurrent bacterial infections were present in some specimens. Fish experimentally treated with oxytetracycline or a combination of oxytetracycline and chloramine-T had a significantly lower mortality rate than untreated fish. Fish treated with chloramine-T alone had a significantly elevated mortality rate compared to controls. Despite supportive medical therapy, mortality levels in treated and untreated groups remained elevated, supporting the hypothesis that the primary pathogen was of viral origin. This is the first report of elevated mortalities in Atlantic halibut associated with an aquareovirus.

Animals↗
PubMed1992

High-performance liquid chromatography of 125I-labeled mouse epidermal growth factor radioiodinated by six different methods.

Six different procedures for radioiodination of mouse epidermal growth factor (EGF) all resulted in a heterogeneous 125I-labeled EGF preparation, as analyzed by reversed-phase HPLC. EGF preparations that had been iodinated with Chloramine T, lodogen, or lodo-beads were found mainly to consist of oxidized 125I-labeled EGF moieties. In contrast, the heterogeneous 125I-labeled EGF preparations obtained by using iodine monochloride, Protag-125, or lactoperoxidase-glucose oxidase-coupled beads (Enzymobeads) contained insignificant amounts of oxidized EGF entities. Ligand equivalence analysis (LEA) of distinct HPLC column fractions, obtained after preparative separation of Chloramine T-125I-labeled EGF, showed that the receptor-binding affinity of the tracer in all subfractions was less than the affinity of unlabeled EGF. This implies that HPLC purification of these 125I-labeled EGF preparations does not yield 125I-labeled EGF preparations with ligand equivalence. However, all but one HPLC column fraction of Enzymobeads-125I-labeled EGF showed ligand equivalence. Despite the small amount of the nonequivalent component in the Enzymobeads-labeled tracer, the nonchromatographed 125I-labeled EGF preparation showed ligand equivalence. No significant differences were observed in the maximal binding capacity of the different 125I-labeled EGF preparations.

Animals↗
PubMed2005

Design considerations for wastewater treatment by reverse osmosis.

Reverse Osmosis is finding increasing use for the treatment of municipal and industrial wastewaters due to the growing demand for high quality water in large urban areas. The growing success of membranes in this application is related to improved process designs and improved membrane products. Key factors which have been determined to result in successful operation of large-scale plants will be discussed. Factors which play a key role in the use of RO membranes include ultra or microfiltration pretreatment, low fouling membranes, flux rate, recovery and control of fouling and scaling. In particular, high flux rates can be used when UF or MF pretreatment is used. These technologies remove most of the suspended particles that would normally cause heavy fouling of lead elements. Typically, fluxes in the range of 17-21 lmh lead to cleaning frequencies in the range of 3-4 months. By combining the use of membrane pretreatment and chloramination of the feed water through chlorine addition, two of the primary sources of RO membrane fouling can be controlled. The use of chloramine has become a proven means to control biofouling in a membrane for wastewater applications. The other significant problems for RO membranes result from organics fouling by dissolved organics and scaling due to saturation of marginally soluble salts. The former can be a significant problem for membranes, due to the strong attraction forces. To some extent, these can be mitigated by making the membrane surface more hydrophilic or changing the charge of the membrane surface. To minimize fouling, many plants are turning to low fouling membranes. Extensive studies have demonstrated that the membrane surface is hydrophilic, neutrally charged over a broad pH range, and more resistant to organic adsorption. Also, an analysis of the potential scaling issues will be reviewed. In particular, calcium phosphate has been found to be one of the key scalants that will limit RO system recovery rate. Calcium phosphate concentrations can reach high values in many wastewaters, and scaling of this compound is not often modeled in most RO projection software. Various process options will be presented to evaluate the most economic means of avoiding phosphate scaling. Finally, data from major RO wastewater treatment plants will be presented to show how the RO membranes operate under actual conditions, utilizing many of these design features. Long-term data from the 2.6 mgd Bedok demonstration Plant demonstrate that the RO membranes operate consistently on wastewater. Experiences from the 8.5 mgd (32,000 m3/day) Bedok and 10.5 mgd (40,000 m3/day) Kranji plants will also be presented. These large plants started operation in the fall of 2002 and have demonstrated an effective means to reclaim high quality water from difficult source waters, such as municipal wastewaters.

Bioreactors↗
PubMed2006

Cellular processing in the SW1222 cell line of mAb A33 directly and indirectly radiohalogenated.

Investigations into the cellular processing of radiolabeled monoclonal antibodies (mAbs) for their further use in radioimmunodiagnosis and cancer therapy are needed in order to understand the fate of internalized and catabolized mAbs. The anti-colorectal cancer mAb, A33, was labelled with 76Br and 125I using the direct Chloramine-T method, or by labelling N-succinimidyl para-(tri-methylstannyl) benzoate and its further conjugation to the mAb. The cellular processing of the four conjugates was investigated in SW1222 cells in vitro. Uptake of mAb was rapid, peaking after 14-16 h. Intracellular degradation was slow and the early loss of radioactivity was due to dissociation of cell-surface bound mAb. The indirect labelling resulted in stronger binding of the mAb as well as prolonged intracellular retention of the radiolabel. Direct and indirect halogen radiolabelling results in different cell-processing patterns of radiolabels, and radioactive catabolic products follow different routes of cellular excretion. The results of this cellular study indicate that indirect labelling is preferable to the direct Chloramine-T method.

Antibodies, Monoclonal↗
PubMed1991

Six methods for direct radioiodination of mouse epidermal growth factor compared: effect of nonequivalence in binding behavior between labeled and unlabeled ligand.

Mouse epidermal growth factor (EGF) was radioiodinated by six different direct iodination methods. The 125I-labeled EGF preparations were distinguished by analyzing the binding of the radioligand to the EGF receptor (EGFR)-containing human placental membranes. The receptor-binding affinity of EGF labeled with Chloramine T was less than the affinity of unlabeled EGF, which precluded an accurate determination of the specific radioactivity of the 125I-labeled EGF preparation by "self-displacement analysis." Scatchard analysis of competitive binding data (increasing concentrations of unlabeled EGF) obtained with commercially prepared 125I-labeled EGF (Chloramine T method), according to the specific radioactivity stated by the manufacturer, resulted in a substantial underestimation of the apparent number of receptors. Iodination of EGF with Iodogen or Iodo-beads, reagents claimed to be more gentle because of their solid state, also yielded 125I-labeled EGF preparations that were not equivalent to the native EGF in receptor binding. In contrast, equivalence in the ligand-receptor interaction between labeled and unlabeled EGF could be achieved by iodinating EGF with iodine monochloride (ICl), Protag-125, or lactoperoxidase-glucose oxidase-coupled beads (Enzymobeads). Scatchard plots of saturation and competitive binding data obtained with these 125I-labeled EGF preparations produced identical results for apparent receptor number and apparent dissociation constants. Such radioiodinated EGF preparations yield relevant binding data in competition studies of labeled and unlabeled EGF.

Animals↗
PubMedDate not supplied

[Obtaining iodinated luteinizing hormone while preserving its biological properties].

The author describes a method of obtaining biologically-active 125I-labeled luteinzing hormone in iodation with chloramine T. The hormone and chloramine T concentration ratio of 1:2, and the reaction time of 20 sec was used. LH-125I with the specific activity of 27--30 microCi/microgram was bound with the receptor strictly specifically.

Animals↗
PubMed1986

Synergy between immunotoxins prepared with native ricin A chains and chemically-modified ricin B chains.

Ricin B chains treated with chloramine-T in the presence or absence of NaI show a 100-fold to 200-fold reduction in their ability to bind to the galactose-containing protein asialofetuin. Such treated B chains do not form covalently associated homodimers with treated B chains or heterodimers with native ricin A chains. Furthermore, they cannot enhance the toxicity of a ricin A chain-containing rabbit anti-human immunoglobulin (RAHIg-A) for Daudi cells. However, when such B chains are coupled to goat anti-rabbit Ig (GARIg), they potentiate the killing of RAHIg-A-treated Daudi cells only slightly less effectively than GARIg coupled to native B chains. Furthermore, if GARIg-B chain conjugates are treated with chloramine-T after coupling, they fail to bind to asialofetuin but enhance the killing of Daudi cells treated with RAHIg-A. These results demonstrate that the ability of ricin B chains to bind to galactose and to enhance the toxicity of ricin A chains (in the form of an antibody-A chain) can be operationally separated. Thus, the two functions of the B chain may reside on separate domains of the molecule.

Animals↗
PubMed1989

Stability of plasminogen activator inhibitor 1 (PAI-1).

The stability of PAI-activity has been studied at different conditions. The inactivation followed first order kinetics. Lowering the temperature and decreasing the pH both, increased the stability of PAI-1 dramatically. Addition of the PAI-1 binding protein, vitronectin, to reactivated PAI-1, about doubled the half-life of PAI-1 at all conditions studied. In the presence of chloramine T, the inactivation of reactivated PAI-1 was very rapid. In this case the protective effect of purified vitronectin, human plasma or fetal calf serum, but not of bovine serum albumin, was pronounced. The stability of the spontaneously active high Mr form of PAI-1 (partially purified or in plasma), constituting a complex between PAI-1 and vitronectin, was quite similar to reactivated PAI-1 in the presence of vitronectin. Addition of pure vitronectin, human plasma or fetal calf serum to such material had no further stabilizing effect. Reactivated PAI-1, which was inactivated by incubation at physiological conditions could again be fully reactivated, in contrast to chloramine T-oxidized PAI-1, which was irreversibly inactivated.

Antigens↗
PubMed1989

[Bactericidal effects of antiseptic gypsum].

In tests with plaster models we inspected the antibacterial effect of the dental plasters Dento-rock and Dento-dur. The results show, that models consisting of plaster that contains Chloramin T 80 were free of bacteria, even when produced from bacterial contaminated alginate impressions. After separation of the plaster models from the alginate impressions, a bactericide effect could be traced to a layer of 220 microns below the surface of the alginate. By using dental plaster of the tested types, it is therefore possible to eliminate the risk of a bacterial infection through plaster models, without any extra investing in time or labor. There is no sign of a negative influence of chloramin on physical parameters of the tested plasters.

Alginates↗
PubMed1989

[Toxicity of different endodontic antiseptics].

The median lethal dose (LD 50) of endodontic antiseptics sodium hypochlorite, chloramine and chlorhexidine digluconate was determined in experiment on albino mice using the method described by Litchfield and Wilcoxon. Sodium hypochlorite and chloramine have shown a twice stronger toxic effect than chlorhexidine digluconate. The difference was significant.

Animals↗
PubMed1979

[On the sporocidal action of dibromoisocyanuric acid (author's transl)].

Using the suspension test the action of dibromoisocyanuric acid (DBI) and--for the purpose of a comparison--elemental chlorine and Chloramine-T on three different spore strains at pH 7.0 and room temperature has been investigated. DBI proved to be a desinfecting agent by which at a concentration of 0.01 M positive halogene (= 1.45 g DBI/I) in 15--60 minutes (depending on the spore strain) a reliable kill can be achieved. Although using elemental chlorine shorter kill rates have been obtained (Chloramine-T was without any effect) DBI shows some advantages which also suggest its use in practice. As possible reasons for the differing efficiency of the three investigated substances the equilibrium concentrations of the hydrolysis products (HOCl resp. HOBr) and diffusion effects are discussed.

Bacillus↗
PubMed1979

Isolation and characterization of surface antigens from Schistosoma mansoni. I. Evaluation of techniques for radioisotope labeling of surface proteins from adult worms.

Radiolabeled surface proteins of adult Schistosoma mansoni were prepared by in vitro labeling of whole worms, and by labeling freeze-thaw surface membrane extracts. Incorporation of 125I into surface proteins was attempted using the lactoperoxidase, chloramine-T, iodosulfanilic acid, and Bolton-Hunter methods. Radiolabeling of whole worms with lactoperoxidase, chloramine-T and iodosulfanilic acid yielded a single protein peak (mol wt greater than 100,000) on SDS-PAGE, and showed considerable incorporation of label in the lipid fraction. Bolton-Hunter labeling of whole worms yielded four major peaks with molecular weights of 100,000, 60,000, 30,000 and 21,000, and minor peaks with molecular weights of 26,000, 36,000, 43,000, 68,000 and 78,000; three of the four major peaks corresponded to prominent bands in Coomassie blue-stained gels. Although carbohydrate-labeling techniques were not successful, a single carbohydrate band, molecular weight greater than 100,000, was detected was PAS staining. Radiolabeling of freeze-thaw extracts yielded results similar to those obtained with whole worms. Electron microscopy revealed the tegument to be left intact and undamaged after labeling with the Bolton-Hunter reagent.

Animals↗
PubMed1977

[Experiments on disinfection of vaccinia virus embedded in scabs and/or at the hand].

Vaccinia viruses embedded in rabbit dermal scabs were subjected to physical and chemical disinfection procedures. Scabs were suspended in vitro without saline or in physiological saline, and left for 1 hour at 70 to 90 degrees C. A complete inactivation was achived only in those scab samples which had been incubated at 90 degrees C for 1 hour and suspended in physiological saline. Scabs which had been placed in a disinfecting apparatus (Vacudes 4000) filled with mattrasses consistently proved to be free of infectious vaccinia viruses in each of the chosen programs. In addition scabs were subjected to disinfection by means of chemical disinfecting agents. The scabs had been placed in a chemical disinfecting suspension and left there for 90 minutes. Complete disinfection was obtained with glutaraldehyde 2%, formaldehyde 2%, Lysoformin 2% or 3%, phenol 5% and chloramine T 2%. Complete disinfection was likewise achieved after 3 hours treatment with some alchohols (ethylalcohol 80%, isopropylalcohol 7%, n-propylalcohol 60%), Amocid 5% and formaldehyde 1%.0.5% formaldehyde caused complete disinfection when applied for 6 hours. The only exception was a Quat which did not disinfect fully even after 18 hours application. Concerning the tests to disinfect the hands complete disinfection occurs when using chloramine T (1.5%) or isopropylalcohol (70%) in 2 to 5 minutes. Further tests were performed with scabs which were placed in sick rooms that were terminally disinfected with formaline vapor. It could be confirmed that the usual terminal disinfection with formaldehyde vapor was unable to completely disinfect the scabs. It is necessary to double the amount of formaldehyde (10 g formaldehyde per cubic metre of space) and prolong the period of treatment to 24 hours to achieve a greater degree of disinfection rate.

Animals↗