Bactericidal properties of peracetic acid and hydrogen peroxide, alone and in combination, and chlorine and formaldehyde against bacterial water strains.

The bactericidal properties of peracetic acid, hydrogen peroxide, chlorine, and formaldehyde were compared in vitro using a rapid micromethod. A combination of peracetic acid and hydrogen peroxide was also tested to assess interactions. The activities of these agents, which are widely used as disinfectants, were evaluated against water isolates and culture collection strains. Peracetic acid and chlorine exhibited an excellent antimicrobial activity, with a relatively rapid destruction of 10(5) bacteria/mL. The time-dependent bactericidal activities of hydrogen peroxide and formaldehyde were the lowest. The combination of peracetic acid and hydrogen peroxide, tested by a checkerboard micromethod, was found to be synergistic. The minimal bactericidal concentration was established in terms of time for a given mixture of peracetic acid and hydrogen peroxide. Determination of bactericidal concentrations showed that synergy was maintained with increasing contact time. Concentrations for minimal times of treatment by chemicals that provided interesting activities in vitro were tested for disinfection of ultrafiltration membranes. The bactericidal activities of peroxygen compounds were confirmed and synergism was maintained in working conditions. Chlorine showed a loss of efficacy when used on membranes.     

The bactericidal, fungicidal and sporicidal properties of hydrogen peroxide and peracetic acid

The antimicrobial properties of aqueous solutions of peracetic acid and hydrogen peroxide have been compared. Peracetic acid exhibited excellent antimicrobial properties, especially under acidic conditions. Reductions by a factor of 10⁶ in the numbers of vegetative bacteria are obtained within 1 min at 25°C using a solution containing 1.3 mmol/l of peracetic acid. Rapid activity against bacterial spores and yeasts also occurs. Hydrogen peroxide is more effective as a sporicide than as a bactericide, with sporicidal action being obtained using a solution containing 0.88 mol/l. Bactericidal action is poor but hydrogen peroxide was bacteriostatic at concentrations above 0.15 mmol/l.     

Sporicidal activity of hydrogen peroxide and peracetic acid against Bacillus anthracis spores

The aim of the presented study was determined the effectiveness of sporicidal activity the peracetic acid and the hydrogen peroxide against B. anthracis spores. In the investigations was used B. anthracis stain "Sterne" 34F2. As inactivators were applied 0,5 % natriumthiosulphate and catalase. The obtained results show that the sporicidal effect of studied substances depends from their concentration and operates time. 5% water solution of peracetic acid shows the full sporicidal activity after outflow 120 minutes and the hydrogen peroxide about concentration 30% after outflow 180 minutes. However the hydrogen peroxide.     

Effects of different disinfection and sterilization methods on tensile strength of materials used for single-use devices

Driven by economic and time constraints, some medical centers and third parties are resterilizing single-use devices (SUDs) for reuse. The steam autoclave is quick, but most plastics used in SUDs cannot survive the temperature. Thus, a number of new methods of cleaning, disinfecting, and sterilizing these complex devices are being introduced on the market. The present study investigated the effects of a range of methods on the tensile strength of latex rubber, silicone elastomer, 2 different formulations of polyurethane, nylon, and high-density polyethylene (HDPE) specimens. The methods used were sodium hypochlorite bleach (Clorox), peracetic acid + hydrogen peroxide (Steris), formaldehyde gas (Chemiclave), low-temperature peracetic acid and gas plasma (Plazlyte), and low-temperature hydrogen peroxide gas plasma (Sterrad). The results showed that silicone elastomer was minimally affected, whereas the strengths of nylon, polyethylene, and latex were reduced by some of the methods. Depending on the formulation, the strength of polyurethane either increased or decreased. The data demonstrated that disinfection and sterilization can affect the tensile strength of certain materials used in medical devices.     

Role of Acinetobacter calcoaceticus 3,4-dihydrocoumarin hydrolase in oxidative stress defence against peroxoacids.

The physiological role of a bifunctional enzyme, 3,4-dihydrocoumarin hydrolase (DCH), which is capable of both hydrolysis of ester bonds and organic acid-assisted bromination of organic compounds, was investigated. Purified DCH from Acinetobacter calcoaceticus F46 catalysed dose- and time-dependent degradation of peracetic acid. The gene (dch) was cloned from the chromosomal DNA of the bacterium. The dch ORF was 831 bp long, corresponding to a protein of 272 amino acid residues, and the deduced amino acid sequence showed high similarity to those of bacterial serine esterases and perhydrolases. The dch gene was disrupted by homologous recombination on the A. calcoaceticus genome. The dch disruptant strain was more sensitive to growth inhibition by peracetic acid than the parent strain. On the other hand, the recombinant Escherichia coli cells expressing dch were more resistant to peracetic acid. A putative catalase gene was found immediately downstream of dch, and Northern blot hybridization analysis revealed that they are transcribed as part of a polycistronic mRNA. These results suggested that in vivo DCH detoxifies peroxoacids in conjunction with the catalase, i.e. peroxoacids are first hydrolysed to the corresponding acids and hydrogen peroxide by DCH, and then the resulting hydrogen peroxide is degraded by the catalase.     

A note on the effect of storage on the chemical resistance of spores of Bacillus subtilis SA22 and Bacillus subtilis globigii B17

Spores of Bacillus subtilis SA22, harvested from nutrient agar after 9 d at 30°C and stored in distilled water at 4°C, were unaltered in their resistance to 17.7% hydrogen peroxide or 0.04% peracetic acid after storage for up to 134 weeks. Three spore crops of B. subtilis globigii were unaffected by storage for up to 134 weeks with respect to 17.7% hydrogen peroxide resistance but were significantly more resistant to 0.04% peracetic acid following storage.     

Characterization of vanadium bromoperoxidase from Macrocystis and Fucus: reactivity of vanadium bromoperoxidase toward acyl and alkyl peroxides and bromination of amines

Vanadium bromoperoxidase (V-BrPO) has been isolated and purified from the marine brown algae Fucus distichus and Macrocystis pyrifera. V-BrPO catalyzes the oxidation of bromide by hydrogen peroxide, resulting in the bromination of certain organic acceptors or the formation of dioxygen. V-BrPO from F. distichus and M. pyrifera have subunit molecular weights of 65,000 and 74,000, respectively, and specific activities of 1580 units/mg (pH 6.5) and 1730 units/mg (pH 6) for the bromination of monochlorodimedone, respectively. As isolated, the enzymes contain a substoichiometric vanadium/subunit ratio; the vanadium content and specific activity are increased by addition of vanadate. V-BrPO (F. distichus, M. pyrifera, and Ascophyllum nodosum) also catalyzes the oxidation of bromide using peracetic acid. In the absence of an organic acceptor, a mixture of oxidized bromine species (e.g., hypobromous acid, bromine, and tribromide) is formed. Bromamine derivatives are formed from the corresponding amines, while 5-bromocytosine is formed from cytosine. In all cases, the rate of the V-BrPO-catalyzed reaction is much faster than that of the uncatalyzed oxidation of bromide by peracetic acid, at pH 8.5, 1 mM bromide, and 2 mM peracetic acid. In contrast to hydrogen peroxide, V-BrPO does not catalyze formation of dioxygen from peracetic acid in either the presence or absence of bromide. V-BrPO also uses phenylperacetic acid, m-chloroperoxybenzoic acid, and p-nitroperoxybenzoic acid to catalyze the oxidation of bromide; dioxygen is not formed with these peracids. V-BrPO does not catalyze bromide oxidation or dioxygen formation with the alkyl peroxides ethyl hydroperoxide, tert-butyl hydroperoxide, and cuminyl hydroperoxide.     

A colorimetric 96-well microtiter plate assay for the determination of urate oxidase activity and its kinetic parameters

Urate oxidase (E.C.1.7.3.3; uricase, urate oxygen oxidoreductase) is an enzyme of the purine breakdown pathway that catalyzes the oxidation of uric acid in the presence of oxygen to allantoin and hydrogen peroxide. A 96-well plate assay measurement of urate oxidase activity based on hydrogen peroxide quantitation was developed. The 96-well plate method included two steps: an incubation step for the urate oxidase reaction followed by a step in which the urate oxidase activity is stopped in the presence of 8-azaxanthine, a competitive inhibitor. Hydrogen peroxide is quantified during the second step by a horseradish peroxidase-dependent system. Under the defined conditions, uric acid, known as a radical scavenger, did not interfere with hydrogen peroxide quantification. The general advantages of such a colorimetric assay performed in microtiter plates, compared to other methods and in particular the classical UV method performed with cuvettes, are easy handling of large amounts of samples at the same time, the possibility of automation, and the need for less material. The method has been applied to the determination of the kinetic parameters of rasburicase, a recombinant therapeutic enzyme.     

Catalase activity and the survival of Pseudomonas putida, a root colonizer, upon treatment with peracetic acid

Peracetic acid is used as a sterilant in several industrial settings. Cells of a plant-colonizing bacterium, Pseudomonas putida in liquid suspension, were more sensitive to killing by peracetic acid when they lacked a major catalase activity, catalase A. Low doses of peracetic acid induced promoter activity of the gene encoding catalase A and increased total catalase specific activity in cell extracts. Microbes present in native agricultural soils rapidly degraded the active oxygen species present in peracetic acid. The simultaneous release of oxygen was consistent with a role for catalase in degrading the hydrogen peroxide that is part of the peracetic acid-equilibrium mixture. Amendment of sterilized soils with wild-type P. putida restored the rate of degradation of peracetic acid to a higher level than was observed in the soils amended with the catalase A-deficient mutant. The association of the bacteria with the plant roots resulted in protection of the wild-type as well as the catalase-deficient mutant from killing by peracetic acid. No differential recovery of the wild-type and catalase A mutant of P. putida was observed from roots after the growth matrix containing the plants was flushed with peracetic acid.     

Comparative sporicidal effects of liquid chemical agents.

We compared the effectiveness of glutaraldehyde, formaldehyde, hydrogen peroxide, peracetic acid, cupric ascorbate (plus a sublethal amount of hydrogen peroxide), sodium hypochlorite, and phenol to inactivate Bacillus subtilis spores under various conditions. Each chemical agent was distinctly affected by pH, storage time after activation, dilution, and temperature. Only three of the preparations (hypochlorite, peracetic acid, and cupric ascorbate) studied here inactivated more than 99.9% of the spore load after a 30-min incubation at 20 degrees C at concentrations generally used to decontaminate medical devices. Under similar conditions, glutaraldehyde inactivated approximately 90%, and hydrogen peroxide, formaldehyde, and phenol produced little killing of spores in suspension. By kinetic analysis at different temperatures, we calculated the rate of spore inactivation (k) and the activation energy of spore killing (delta E) for each chemical agent. Rates of spore inactivation had a similar delta E value of approximately 20 kcal/mol (ca.83.68 kJ/mol) for every substance tested. The variation among k values allowed a quantitative comparison of liquid germicidal agents.     

A near‐infrared fluorescent probe based on boric acid hydrolysis for hydrogen peroxide detection and imaging in HeLa cells

Using the characteristics of hydrogen peroxide that are able to cleave phenyl‐boric acid selectively and efficiently, we here report a dicyanoisophorone‐boric acid ( DCP‐BA )‐based near‐infrared (NIR) fluorescent probe for detection of hydrogen peroxide. This probe shows a rapid, highly selective, and sensitive detection process for hydrogen peroxide with a significant NIR fluorescent turn‐on response that has been successfully applied to detect exogenous hydrogen peroxide in HeLa cells.     

Oxidation of cytochrome c peroxidase to compound I by peroxyacids: evidence for rate-limiting diffusion through the protein matrix

The rate of the reaction between p-nitroperoxybenzoic acid and cytochrome c peroxidase (CcP) has been investigated as a function of pH and ionic strength. The pH dependence of the reaction between CcP and peracetic acid has also been determined. The rate of the reactions are influenced by two heme-linked ionizations in the protein. The enzyme is active when His-52 (pK(a) 3.8 +/- 0.1) is unprotonated and an unknown group with a pK(a) of 9.8 +/- 0.1 is protonated. The bimolecular rate constant for the reaction between peracetic acid and CcP and between p-nitroperoxybenzoic acid and CcP are (1.8 +/- 0.1) x 10(7) and (1.6 +/- 0.2) x 10(7) M(-)(1) s(-)(1), respectively. These rates are about 60% slower than the reaction between hydrogen peroxide and CcP. A critical comparison of the pH dependence of the reactions of hydrogen peroxide, peracetic acid, and p-nitroperoxybenzoic acid with CcP provides evidence that both the neutral and anionic forms of the two peroxyacids react directly with the enzyme. The peracetate and p-nitroperoxybenzoate anions react with CcP with rates of (1.5 +/- 0.1) x 10(6) and (1.6 +/- 0.2) x 10(6) M(-)(1) s(-)(1), respectively, about 10 times slower than the neutral peroxyacids. These data indicate that CcP discriminates between the neutral peroxyacids and their negatively charged ions. However, the apparent bimolecular rate constant for reaction between p-nitroperoxybenzoate and CcP is independent of ionic strength in the range of 0.01-1.0 M, suggesting that electrostatic repulsion between the anion and CcP is not the cause of the lower reactivity for the peroxybenzoate anion. The data are consistent with the hypothesis that the rate-limiting step for the oxidation of CcP to compound I by both neutral peroxyacid and the negatively charged peroxide ion is diffusion of the reactants through the protein matrix, from the surface of the protein to the distal heme pocket.     

Disinfectant effect of Persteril in combination with detergents

In laboratory conditions, the microbicidal effect, pH and changes in the content of peracetic acid and hydrogen peroxide were tested in Persteril at concentrations of 5 ml/l and 0.5 ml/l as well as in mixtures of these Persteril solutions with the detergents Jar, Pur, Hit, Corona, Sapon, Rekord and Universal. The efficiency and stability of Persteril solution in combination with the detergents were similar to those of Persteril aqueous solution. The tested mixtures ensured satisfactory bactericidal effect after 19-day storage. The sporicidal effect could be guaranteed during 5 days only at a concentration of 5 ml/l and provided disinfection was carried out by submerging. The above mixtures of Persteril and detergents have been recommended for one-stage disinfection in all types of medical facilities requiring simultaneous disinfection and washing.     

Inhibition of arachidonic acid incorporation into erythrocyte phospholipids by peracetic acid and other peroxides. Role of arachidonoyl-CoA: 1-palmitoyl-sn-glycero-3-phosphocholine acyl transferase

To explore possible mechanisms of the arachidonic acid deficiency of the red blood cell membrane in alcoholics, we compared the effect of ethanol and its oxidized products, acetaldehyde and peracetic acid, with other peroxides on the accumulation of [14C]arachidonate into RBC membrane lipids in vitro. Incubation of erythrocytes with 50 mM ethanol or 3 mM acetaldehyde had no effect on arachidonate incorporation. Pretreatment of erythrocytes with 10 mM hydrogen peroxide, 0.1 mM cumene hydroperoxide or 0.1 mM t-butyl hydroperoxide had little effect on [14C]arachidonate incorporation in the absence of azide. However, pretreatment of cells with N-ethylmaleimide, 0.1 mM peracetic acid or performic acid, with or without azide, inhibited arachidonate incorporation into phospholipids but not neutral lipids. In chase experiments, peracetate also inhibited transfer of arachidonate from neutral lipids to phospholipids. To investigate a possible site of this inhibition of arachidonate transfer into phospholipids by percarboxylic acids, we assayed a repair enzyme, arachidonoyl CoA: 1-palmitoyl-sn-glycero-3-phosphocholine acyl transferase (EC 2.3.1.23). As in intact cells, phospholipid biosynthesis was inhibited more by N-ethylmalemide and peracetic acid than by hydrogen peroxide, cumene hydroperoxide, and t-butyl hydroperoxide. Peracetic acid was the only active inhibitor among ethanol and its oxidized products studied and may deserve further examination in ethanol toxicity.     

Polyvinyl pyridine metal complex as permanent antimicrobial finishing for viscose fabric

Viscose fabrics were treated with polyvinyl pyridine (PVP) using padding technique, followed by oxidation with hydrogen peroxide or peracetic acid, which was prepared by the reaction of tetra acetyl ethylene diamine (TAED) with hydrogen peroxide. Peracetic acid gives higher oxidation of PVP than hydrogen peroxide. FTIR study proved the formation of N-oxide as a result of oxidation. Incorporation of copper and silver ion onto oxidized PVP was also proved by FTIR. The antimicrobial study emphasise that Cu/oxidized PVP and Ag/oxidized PVP have retarded the growth of bacteria significantly, and Ag/oxidized PVP has a far better biocidal activity. The antibacterial activity of both metal ions survived after washing 10 times.     

High-throughput Assays for Superoxide and Hydrogen Peroxide: DESIGN OF A SCREENING WORKFLOW TO IDENTIFY INHIBITORS OF NADPH OXIDASES*

Recent progress characterizing the reaction mechanism(s) of fluorescent probes with reactive oxygen species has made it possible to rigorously analyze these reactive species in biological systems. We have developed rapid high throughput-compatible assays for monitoring cellular production of superoxide radical anion and hydrogen peroxide using hydropropidine and coumarin boronic acid probes, respectively. Coupling plate reader-based fluorescence measurements with HPLC-based simultaneous monitoring of superoxide radical anion and hydrogen peroxide provides the basis for the screening protocol for NADPH oxidase (Nox) inhibitors. Using this newly developed approach along with the medium-throughput plate reader-based oximetry and EPR spin trapping as confirmatory assays, it is now eminently feasible to rapidly and reliably identify Nox enzyme inhibitors with a markedly lower rate of false positives. These methodological advances provide an opportunity to discover selective inhibitors of Nox isozymes, through enhanced conceptual understanding of their basic mechanisms of action.     

Clastogenic activity of caffeic acid and its relationship to hydrogen peroxide generated during autooxidation

Caffeic acid is a clastogenic cinnamic acid found in a conjugated form in a variety of foods. The possibility that the biological activity of caffeic acid is due to hydrogen peroxide generated during its autooxidation in solution was investigated using chromosome aberrations in Chinese hamster ovary cells as a test system. Freshly prepared caffeic acid at pH 7.00 contained only traces of hydrogen peroxide, as assayed by the molybdate-catalyzed release of I-3. Such solutions exhibited clastogenic activity which could not be accounted for by the level of hydrogen peroxide present, and which was not significantly diminished by the addition of catalase or horseradish peroxidase. 3-day-old solutions of caffeic acid exhibited higher levels of hydrogen peroxide, and increased biological activity. In such solutions, the clastogenic activity was catalase-sensitive and could be entirely accounted for by the level of hydrogen peroxide present.     

Removal of hydroperoxides by immobilized borohydride: a good method for purification of biochemical materials.

Borohydride was immobilized on a quaternary ammonium type anion exchange resin, Amberlite IRA-400, by an exchange reaction in N,N-dimethylformamide. The reducing ability of borohydride on the polymer beads was examined; 0.1 g resin was applied for about 30 min to 3 ml solutions of hydrogen peroxide, ethyl hydrogen peroxide, and peracetic acid, at a concentration of approximately 40 mM, m-chloroperbenzoic acid (3.13 mM), and 5-phenyl-4-pentenyl-1-hydroperoxide (1 mM), respectively. The solutions were then assayed for remaining hydroperoxide by use of horseradish peroxidase or prostaglandin H synthase. In addition, the effect of treatment on the ability of 5-phenyl-4-pentenyl-1-hydroperoxide to initiate the cyclooxygenase activity of prostaglandin H synthase was investigated. Results indicated that immobilized borohydride is very efficient in removing hydroperoxides. It can be used in either organic or aqueous media. It is convenient for both large and small scales, particularly important for purification of biochemical materials.