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Gold nanoparticle-catalyzed luminol chemiluminescence and its analytical applications.

Gold colloids with nanoparticles of different sizes were found to enhance the chemiluminescence (CL) of the luminol-H2O2 system, and the most intensive CL signals were obtained with 38-nm-diameter gold nanoparticles. UV-visible spectra, X-ray photoelectron spectra, and transmission electron microscopy studies were carried out before and after the CL reaction to investigate the CL enhancement mechanism. The CL enhancement by gold nanoparticles of the luminol-H2O2 system was supposed to originate from the catalysis of gold nanoparticles, which facilitated the radical generation and electron-transfer processes taking place on the surface of the gold nanoparticles. The effects of the reactant concentrations, the size of the gold nanoparticles. and some organic compounds were also investigated. Organic compounds containing OH, NH2, and SH groups were observed to inhibit the CL signal of the luminol-H2O2-gold colloids system, which made it applicable for the determination of such compounds.

Journal Article↗

Synthesis, characterization of dihydrolipoic acid capped gold nanoparticles, and functionalization by the electroluminescent luminol.

The use of gold nanoparticles as biological probes requires the improvement of colloidal stability. Dihydrolipoic acid (DHLA), a dithiol obtained by the reduction of thioctic acid, appears therefore very attractive for the stabilization and the further functionalization of gold nanoparticles because DHLA is characterized by a carboxylic acid group and two thiol functions. The ionizable carboxylic acid groups ensure, for pH > or = 8, the water solubility of DHLA-capped gold (Au@DHLA) nanoparticles, prepared by the Brust protocol, and the stability of the resulting colloid by electrostatic repulsions. Moreover almost all DHLA, adsorbed onto gold, adopts a conformation allowing their immobilization by both sulfur ends. It is proved by sulfur K-edge X-ray absorption near edge structure spectroscopy, which appears as an appropriate tool for determining the chemical form of sulfur atoms present in the organic monolayer. Such a grafting renders the DHLA monolayers more resistant to displacement by dithiothreitol than mercaptoundecanoic acid monolayers. The presence of DHLA on gold particles allows their functionalization by the electroluminescent luminol through amine coupling reactions assisted by 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and N-hydroxysuccinimide. As a luminol-functionalized particle is nine times as bright as a single luminol molecule, the use of the particles as a biological probe with a lower threshold of detection is envisaged.

Journal Article↗

Enzymatic activation and trapping of luminol-substituted peptides and proteins. A possible means of amplifying the cytotoxicity of anti-tumor antibodies.

Glutathione and glucose oxidase (EC 1.1.3.4) conjugates containing covalently bound luminol were prepared as prototypes for peptides and proteins with latent, enzyme-activatable chemical reactivity. In the presence of small quantities of activated horseradish peroxidase, conjugated luminol molecules were oxidized to unstable free radicals which reacted rapidly with soluble proteins and cells. These observations are of interest in regard to possible sequential localization reactions in which a few molecules of cell-bound antibody-horseradish peroxidase would be used to catalytically alter and trap many molecules of a second (luminol-substituted) enzyme, toxin, or hapten in the same area, as might be desirable in promoting selective cell destruction.

Antibodies, Anti-Idiotypic↗

Influence of polyclonal immunoglobulins on the polymorphonuclear leukocyte response to lipopolysaccharide of Salmonella enteritidis as measured with luminol-enhanced chemiluminescence.

In gram-negative sepsis, the activation of polymorphonuclear leukocytes (PMN) by lipopolysaccharide (LPS) and the resulting production of superoxide and other oxygen radicals may be an important cause of tissue damage. A suppression of the PMN response to LPS stimulation would be therapeutically beneficial. The aim of this study was to determine whether different polyclonal immunoglobulins (Igs; 5S-Ig, 7S-Ig, and 19S-Ig) influence the PMN response to LPS of Salmonella enteritidis in vitro. The respiratory burst activity of PMN was measured with luminol-enhanced chemiluminescence. After addition of a 5S-Ig solution containing F(ab')2 fragments of IgG and a 19S-Ig solution containing 12% polyclonal IgM, luminol-enhanced chemiluminescence was reduced by 27% (P < 0.05) and 46% (P < 0.005), respectively. However, after addition of a 7S-Ig solution containing polyclonal IgG, luminol-enhanced chemiluminescence was increased fourfold (P < 0.05). The results suggest that the influence of polyclonal Igs on PMN response to LPS stimulation is dependent on the Ig class, F(ab')2 fragments of IgG and IgM leading to LPS neutralization and IgG leading to the production of potentially toxic oxygen radicals.

Humans↗

Inhibition of luminol and lucigenin chemiluminescence by reducing organic compounds.

The effect of 22 reducing organic compounds on the chemiluminescent intensities and spectra of luminol-H(2)O(2)-Co(2+) and lucigenin-H(2)O(2)-Co(2+) reactions were examined. Many of the reductants inhibit the two chemiluminescent systems to different extents in the presence and absence of Co(2+). The magnitude of inhibition by reducing organic compounds is related to their molecular structure and reducing properties. The mechanism of chemiluminescent inhibition by reducing organic compounds is probably due to the reaction between reductants and radical intermediates of chemiluminescent reaction. Based on the inhibition of luminol and lucigenin chemiluminescence by reducing organic compounds, the possibility of analytical application was explored. Some reductants are detectable at picomolar levels.

Acridines↗

Inhibition of luminol-dependent luminescence and simultaneous generation of native luminescence of activated human polymorphonuclear leukocytes by addition of albumin.

Luminol-dependent luminescence (LDL) and luminol-independent, native luminescence (NL) of polymorphonuclear leukocytes were investigated with respect to the effects generated by the addition of albumin to the reaction medium. The cells were activated: (1) by simple surface attachment to a hydrophilic plastic, (2) by opsonized zymosan, (3) by phorbol myristate acetate, (4) by formylmethionyl-leucyl-phenylalaline. Both kinds of emissions were recorded simultaneously using a method of spectral discrimination. The addition of albumin resulted in an inhibition of LDL, which coincided with a generation of NL. The extent of the inhibition of LDL depended on the type of stimulus used. Maximum inhibition occurred with cells activated by attachment to plastic surfaces and minimum inhibition was observed with cells stimulated by opsonized zymosan. Different contributions of extracellularly released reactive oxygen-species may be responsible for this. It appears possible to discriminate between intra- and extracellular sites of oxygen-metabolites production using albumin simultaneously as extracellular quencher of LDL and as luminescent probe for NL.

Humans↗

Analysis of luminol-dependent chemiluminescence from granule depleted neutrophil cytoplasts reveals two different light-emitting mechanisms.

When neutrophil cytoplasts (granule-free vesicles of cytoplasm enclosed by plasmalemma) were exposed to the chemotactic peptide formylmethionyl-leucyl-phenylalanine, no luminol-dependent chemiluminescence was detected, despite a pronounced production of superoxide anions and hydrogen peroxide. Addition of purified myeloperoxidase (MPO) or human serum albumin (HSA) to the cytoplasts before the stimulus resulted in a chemiluminescence response. In contrast to the bimodal response obtained from normal PMNL, only a single peak of chemiluminescence was obtained from the cytoplasts responding to the peptide. The time-course of the response obtained in the presence of albumin was more prolonged than the response obtained in the presence of MPO. Furthermore, the involvement of different oxidative metabolites in the MPO and the HSA systems, respectively, was demonstrated by the accumulated chemiluminescence effect obtained when MPO, but not HSA, was introduced in the measuring system after FMLP addition. From these results it can be concluded that there are at least two different light-generating mechanisms in FMLP-induced luminol-dependent chemiluminescence of neutrophil cytoplasts. One of these is dependent on myeloperoxidase and possibly related to the myeloperoxidase-hydrogen peroxide reaction, whereas the other one is hydrogen peroxide independent.

Adult↗

Free fatty acid determination by peroxidase catalysed luminol chemiluminescence.

A sensitive, specific, and partly automatic method for the analysis of free fatty acids is described. The assay involves activation of free fatty acids by acyl-CoA synthetase (EC 6.2.1.3) followed by oxidation of the thioesters by acyl-CoA oxidase. The H2O2 formed is determined in a reaction catalysed by horseradish peroxidase (EC 1.11.1.7) using luminol as electron donor. The assay has a linear range of 0.05 to 5 nmol of different free fatty acids (C10-C18) in the original sample. The efficiency of the method toward capric, lauric, myristic, palmitic, palmitoleic, stearic, oleic, and linoleic acid measured as recovery of light emission compared to that of H2O2 standards, was over 90%. AffiGel 501 was used to covalently bind the free thiol group in CoASH eliminating interference of this substance in the peroxidase-luminol reaction.

Acyl-CoA Oxidase↗

Alkali-labile luminol derivatives as labels for quantitative binding studies with polyionic macromolecules.

Acylated derivatives of luminol (5-amino-2,3-dihydro-1,4-phthalazinedione) bearing a carboxyl or amino group can be linked by amide bonds to a macromolecule requiring labelling. Though themselves of low quantum yield these compounds are alkali-labile and can be detected at a similar level of sensitivity to the parent compound luminol. These cheap, readily accessible compounds are less hydrophobic than other currently employed chemiluminescent labels. They also lack a positively charged nitrogen atom which could complicate their covalent linkage to polyanionic compounds. They thus appear well suited for labelling heparin and other macromolecules which interact with the luminal surface of blood vessels.

Alkalies↗

Different influences of cytochalasin B on the activation of human neurophils settled onto petri dishes displayed by simultaneously detected native and luminol-dependent luminescence.

Cytochalasin B (CB) is known to interfere reversibly with the cytoplasmic contractile filamental network of mammalian cells. The role of the microfilament system in the mechanism of the reactive oxygen intermediates release of polymorphonuclear leukocytes (PMNL) was studied for different kinds of stimuli. PMNL from fresh human blood were treated with CB and stimulated by adherence on plastic surfaces, by opsonized zymosan, by phorbol myristate acetate and by N-formylmethionyl-phenylalaline. The production of reactive oxygen species were monitored by simultaneous detection of native, luminol-independent, luminescence (NL) and luminol-dependent luminescence (LDL) using a method of spectral discrimination. Different influences of CB on NL with respect to LDL as well stimuli-dependent influences of CB on the luminescence response of PMNL were observed. Especially phagocytosis-associated activation of PMNL was strongly inhibited by CB, whereas LDL was reduced to a much greater extent in comparison with NL. A firm involvement of the microfilament system is indicated, but it depends on the kind of stimulus engaged.

Actin Cytoskeleton↗

Luminol-dependent chemiluminescence in bovine eosinophils and neutrophils: differential increase of intracellular and extracellular chemiluminescence induced by soluble stimulants.

Luminol chemiluminescence was used to detect activation of the respiratory burst oxidase in bovine eosinophils and neutrophils. Extracellular and intracellular chemiluminescence were measured by supplementing the medium with horseradish peroxidase and catalase, respectively. Pure bovine eosinophils (greater than 90%), maximally stimulated with 1 nmol/l phorbol 12-myristate-13-acetate (PMA) showed ten times more extracellular luminol-dependent chemiluminescence (CL) than maximally stimulated pure bovine neutrophils (greater than 96%). Extracellular CL from eosinophils was preferably induced over intracellular CL by both PMA (27-fold difference) and platelet-activating factor (PAF) at 2 mumol/l (9-fold difference), but not by calcium ionophore A23187 (15 mumol/l). Time course information was used in the following experiments to distinguish between the mode of action of various stimulants. A progressively longer lag period was observed in eosinophil suspensions treated with decreasing doses of PMA, whereas platelet-activating factor induced a dose-dependent increase in the maximum response with no change in time to peak CL. The time course of extracellular CL was almost identical to intracellular CL for all stimulants tested, providing no evidence to suggest that extracellular CL stems from a different enzyme system than intracellular CL. Eosinophils generated most extracellular CL when stimulated with PMA, whereas neutrophils were most efficiently stimulated with A23187, which induced intracellular CL in eosinophils as well as in neutrophils. This accords with the greater tendency of neutrophils to ingest and kill microorganisms, whereas eosinophils are armed to destroy large extracellular targets.

Animals↗

Chemiluminescence intensities and spectra of luminol oxidation by sodium hypochlorite in the presence of hydrogen peroxide.

Hydrogen peroxide amplifies the chemiluminescence in the oxidation of luminol by sodium hypochlorite. A linear relationship between concentration of hydrogen peroxide and light intensity was found in the concentration range 5 x 10(-8)-7.5 x 10(-6) mol/l. At 7.5 x 10(-6) mol/l H2O2 the chemiluminescence is amplified 550-fold. The chemiluminescence spectra of these reactions have a wavelength maximum at 431 nm independent of the concentration of hydrogen peroxide. The results indicate that hydrogen peroxide is a necessary component in the chemiluminescent oxidation of the luminol by sodium hypochlorite.

Hydrogen Peroxide↗

Enhanced chemiluminescence in the oxidation of luminol and an isoluminol cortisol conjugate by hydrogen peroxide in reversed micelles.

The chemiluminescent oxidation of luminol and an isoluminol cortisol conjugate (ABI-COR) by hydrogen peroxide has been studied in cetyltrimethylammonium bromide (CTAB) reversed micelles in octane-chloroform (1:1). The maximum chemiluminescence intensity of both compounds is dependent on the initial concentrations of the H2O2 and substrates, the pH value of the micelle polar phase and the H2O/CTAB ratio. The optimum pH ranged from 8.5 to 9.5. Under comparable conditions, the chemiluminescence intensity for luminol was 15-fold higher than for the ABI-COR conjugate. A mechanism of oxidation of the substrates in reversed micelles is proposed and the possible mechanisms of inhibition by the substrate and oxidant is discussed.

Cetrimonium↗

The enhancement of iron-dependent luminol peroxidation by 2,2'-dipyridyl and nitrilotriacetate.

The generation of radicals from luminol and H2O2, in the presence of iron and iron chelates was monitored by measuring the chemiluminescence produced by further oxidation of these radicals. 2,2'-Dipyridyl enhanced the production of chemiluminescence in the presence of FeSO4, ferritin and haemosiderin but not FeCl3 or horseradish peroxidase. Nitrilotriacetic acid (NTA) enhanced chemiluminescence in the presence of both FeSO4 and FeCl3 but not ferritin or haemosiderin. The enhancement of chemiluminescence by iron chelation may have analytical applications and the process by which these iron chelates are able to generate radicals from the nitrogenous base luminol may be similar to that responsible for their toxic effects on DNA.

2,2'-Dipyridyl↗

Effect of oxygen abstraction on the peroxidase-luminol-perborate system: relevance to the HRP enhanced chemiluminescence mechanism.

Abstraction of oxygen from the HRP enhanced chemiluminescence system has no significant effect on the chemiluminescence generated. It is, therefore, proposed that in the peroxidase-luminol-perborate system at pH 7.3, chemiluminescence is generated by a direct reaction of diazaquinones with hydrogen peroxide and not, as generally assumed, from the reaction of luminol radicals with the molecular oxygen.

Borates↗

Studies of visible oscillating chemiluminescence with a luminol-H2O2-KSCN-CuSO4-NaOH system in batch reactor.

Oscillating chemical reactions are complex systems involving a large number of chemical species. In oscillating chemical reactions some species, usually reaction intermediates, exhibit fluctuation in concentration. Visible oscillating chemiluminescence, produced by the addition of luminol (3-aminophthalhydrazide) to the oscillating system H(2)O(2)-KSCN-CuSO(4)-NaOH, was investigated. In this study the effect of varying the concentration of H(2)O(2), KSCN, CuSO(4), NaOH and luminol was investigated in a batch reactor. We showed that the concentration of all components involved in the oscillating chemilumenscent reaction influenced the light intensity and the oscillation period.

Copper Sulfate↗

A comprehensive experimental study of industrial, domestic and environmental interferences with the forensic luminol test for blood.

This paper presents the fi rst comprehensive and quantitative study of substances that interfere with the forensic luminol test for blood. Two hundred and fifty substances have been selected on the basis of modern lifestyles and of contiguity with crime scenes. The intensity of the chemiluminescence produced by each substance has been measured relative to that of haemoglobin and the peak wavelength shift has also been determined. The following is a short list of nine substances that produce chemiluminescence intensities comparable with that of haemoglobin: turnips, parsnips, horseradishes, commercial bleach (NaClO), copper metal, some furniture polishes, some enamel paints, and some interior fabrics in motor vehicles. Care needs to be taken when the luminol test for blood is used in the presence of these substances.

Blood Chemical Analysis↗

Hydrazine-induced post-chemiluminescence phenomenon of permanganate-luminol reaction and its applications.

The post-chemiluminescence phenomenon arising from the permanganate-luminol reaction induced by hydrazine and isoniazid was investigated. When hydrazine or isoniazid was injected into the mixture after the end of the reaction of permanganate with alkaline luminol, a new chemiluminescence (CL) reaction was initiated and strong CL signal was detected. A possible CL mechanism is suggested, based upon the studies of the kinetic characteristics of the CL reaction, the UV-visible spectra, the CL spectra and some other experiments. The present reactions allow the determination of 0.1-10.0 mg/L hydrazine and 0.02-1.0 mg/L isoniazid, with detection limits of 0.03 mg/L and 0.006 mg/L, respectively. The method was applied to the determination of isoniazid in pharmaceutical preparations.

Antitubercular Agents↗